Handbook of Forensic Medicine [2 ed.] 9781119648550, 9781119648581, 9781119648611

Citation preview

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Handbook of Forensic Medicine

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Edited by

Burkhard Madea

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Institute of Forensic Medicine University of Bonn Bonn, Germany

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Volume 1

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Second Edition

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Handbook of Forensic Medicine

This second edition first published 2022 © 2022 John Wiley & Sons Ltd

All rights reserved. No part of this publication may be reproduced, stored in a retrieval system, or transmitted, in any form or by any means, electronic, mechanical, photocopying, recording or otherwise, except as permitted by law. Advice on how to obtain permission to reuse material from this title is available at http://www.wiley.com/go/permissions. The right of Burkhard Madea to be identified as the author of the editorial material in this work has been asserted in accordance with law.

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Registered Office John Wiley & Sons Ltd, The Atrium, Southern Gate, Chichester, West Sussex, PO19 8SQ, UK

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Editorial Office The Atrium, Southern Gate, Chichester PO19 8SQ, UK

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For details of our global editorial offices, customer services, and more information about Wiley products visit us at www.wiley.com.

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Wiley also publishes its books in a variety of electronic formats and by print-on-demand. Some content that appears in standard print versions of this book may not be available in other formats.

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Limit of Liability/Disclaimer of Warranty The contents of this work are intended to further general scientific research, understanding, and discussion only and are not intended and should not be relied upon as recommending or promoting scientific method, diagnosis, or treatment by physicians for any particular patient. In view of ongoing research, equipment modifications, changes in governmental regulations, and the constant flow of information relating to the use of medicines, equipment, and devices, the reader is urged to review and evaluate the information provided in the package insert or instructions for each medicine, equipment, or device for, among other things, any changes in the instructions or indication of usage and for added warnings and precautions. While the publisher and authors have used their best efforts in preparing this work, they make no representations or warranties with respect to the accuracy or completeness of the contents of this work and specifically disclaim all warranties, including without limitation any implied warranties of merchantability or fitness for a particular purpose. No warranty may be created or extended by sales representatives, written sales materials or promotional statements for this work. The fact that an organization, website, or product is referred to in this work as a citation and/or potential source of further information does not mean that the publisher and authors endorse the information or services the organization, website, or product may provide or recommendations it may make. This work is sold with the understanding that the publisher is not engaged in rendering professional services. The advice and strategies contained herein may not be suitable for your situation. You should consult with a specialist where appropriate. Further, readers should be aware that websites listed in this work may have changed or disappeared between when this work was written and when it is read. Neither the publisher nor authors shall be liable for any loss of profit or any other commercial damages, including but not limited to special, incidental, consequential, or other damages.

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Library of Congress Cataloging-in-Publication Data Names: Madea, B. (Burkhard), editor. Title: Handbook of forensic medicine / edited by Burkhard Madea. Description: Second edition. | Hoboken, NJ : Wiley, 2022. | Includes bibliographical references and index. Identifiers: LCCN 2021036998 (print) | LCCN 2021036999 (ebook) | ISBN 9781119648550 (hardback) | ISBN 9781119648581 (Adobe PDF) | ISBN 9781119648611 (epub) Subjects: MESH: Forensic Medicine Classification: LCC RA1063.4 (print) | LCC RA1063.4 (ebook) | NLM W 700 | DDC 614/.1–dc23 LC record available at https://lccn.loc.gov/2021036998 LC ebook record available at https://lccn.loc.gov/2021036999 Cover Design: Wiley Cover Image: © Comstock Images/Getty Set in 10/12pt MinionPro by Straive, Pondicherry, India

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Edition History John Wiley & Sons Ltd (1e, 2013)

Brief Contents

Preface to the First Edition

xxvii

Foreword by Denis A. Cusack

xxix

Foreword by Duarte Nuno Vieira

xxxi

PART I Duties of Forensic Medicine in Modern Societies

1

Burkhard Madea

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Duties of Forensic Medicine and the Forensic Medicine Practitioner19

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Burkhard Madea

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Nature and Definition of Death59 Certification of Death: External Postmortem Examination69

PART III Traumatology and Violent Death311 15

Postmortem Changes and Time since Death91 Burkhard Madea, Claus Henßge, Saskia Reibe, and Michael Tsokos

Legal Aspects of Traumatology and Violent Death313 Philip Beh

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Traumatology and Criminology317 Philip Beh

Burkhard Madea and Antonella Argo

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Mass Disaster Victim Identification291 Ruediger Lessig and Mechthild Prinz

Burkhard Madea

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Anthropology and Osteology267 Cristina Cattaneo and Daniele Gibelli

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Antonella Argo and Valeria Tullio Brigitte Tag

International Guidelines and Accreditation in Forensic Medicine43

PART II  Medical Aspects of Death

The Doctor, the Dead and the Relatives249

12 Transplantation257 13

Peter Wiegand, Burkhard Madea and Frank Mußhoff

5

Autopsy183 Peter J. T. Knudsen, Jørgen L. Thomsen, Wolfgang Grellner, Garyfalia Ampanozi, Jakob Heimer, Lars C. Ebert, Michael J. Thali, Burkhard Madea, Antti Sajantila

27

Forensic Medicine and Human Rights Hans Petter Hougen

4

Henrik Druid

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1 History3

9 Crime Scene and Crime Scene Investigations161

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Preface to the Second Edition 

Claas T. Buschmann, Marc Windgassen and Michael Tsokos

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Cremation151

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List of Contributors

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Contentsix

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Some Preliminary Remarks Concerning Biomechanical Principles321 Burkhard Madea and Heinz-­Dieter Wehner

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Brief Contents

Sequelae of Traumatic Injuries and Causes of Death327 Guy N. Rutty and Michael Tsokos

19

Vital Reactions349 Burkhard Madea, Elke Doberentz, Christian Jackowski, Wolfgang Grellner and Toshikazu Kondo

32

Burkhard Madea

33 Special Issues Regarding Expert Evidence in Violent Death763 Johanna Preuß-­Wössner, Rainer Amberg, Burkhard Madea, Jørgen L. Thomsen, Martin Worm-­Leonhard, and Paolo Procaccianti

20 Mechanical Trauma and Classification of Wounds375 Burkhard Madea, Stefan Pollak, Annette Thierauf-­Emberger, Veronique Henn, Christoph Meissner, Manfred Oehmichen, and Peter Mygind Leth

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PART IV Sudden and Unexpected Death from Natural Causes 801

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34 Natural Causes of Sudden Death803 Emanuela Turillazzi, Alessandro Santurro, Pietrantonio Ricci, Matteo Scopetti and Vittorio Fineschi

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Forensic Ballistics: Injuries from Gunshots, Explosives and Arrows459

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21

Bernd Karger

Wolfgang Keil, Philippe Lunetta, and Mattias Kettner, and Burkhard Madea

35 Postmortem Biochemistry as an Aid in Determining the Cause of Death845

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Asphyxiation505

Tomoya Ikeda, Naoto Tani, Takaki Ishikawa, Gerhard Kernbach-Wighton, Aurelio Luna

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23 Injuries Due to Heat607 Burkhard Madea and Elke Doberentz

36 Sudden and Unexpected Deaths in Infants and Sudden Infant Death Syndrome887

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24 Injuries Due to Cold627 Burkhard Madea and Elke Doberentz

Thomas Bajanowski, Mechtild M. Vennemann and Jan-­Peter Sperhake

25 Electrocution and Lightning639 Ryan Blumenthal

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26 Starvation and Neglect669

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Burkhard Madea and Elke Doberentz

Infanticide683

PART V  Clinical Forensic Medicine 37

38

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Thomas Bajanowski, Janine Helmus, Mechtild Vennemann and Sibylle Banaschak

28 Death During Pregnancy697

Marc Windgassen

30 Death in an Abnormal Position: Physical Restraint717 Sexual Homicide729 Philip Beh

Non-­Accidental Head Injury in Children939

39 Intimate Partner and Domestic Violence987 Saskia Etzold

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John C. Hunsaker III and Lisa B. E. Shields

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Sexual Abuse919

Robert A. Minns, Burkhard Madea, and Elke Doberentz

Eberhard Lignitz and Veronique Henn

29 Autoerotic Death707

917

Cristina Cattaneo, Daniele Gibelli and Marina Ruspa

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Medical Malpractice735

School Violence995 Gerhard Kernbach-­Wighton

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Violence Against Homosexuals1001 Gerhard Kernbach-­Wighton

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Brief Contents

42 Violence Against the Elderly1005 John C. Hunsaker III and Lisa B.E. Shields

Self-­harm1021 Saskia Etzold

44 Medical Age Assessment in  Living Individuals1027 Andreas Schmeling and Ernst Rudolf

Werner Jung

55 Effects of Vision and Visual Fields on Fitness to Drive1345 Patrick Vivell

56 Effects of Epilepsy on Fitness to Drive1355 Randi von Wrede

PART VI  Forensic Psychiatry

1055

57 Effects of Diabetes on Fitness to Drive1359 Wulf Quester

45 Forensic Psychiatry1057

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Alan R. Felthous and Henning Saß

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58 Epidemiology and Causal Factors in Fitness to Drive1375 Günter Berghaus

Ilkka Ojanperä

Olaf H. Drummer, Dimitri Gerostamoulos and Jennifer L. Schumann

Alan Wayne Jones

60 Effects of Illegal Drugs on Fitness to Drive1409

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47 Epidemiology and Adverse Drug Reactions1099

59 Driving Under the Influence of  Alcohol1387

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46 Legal Aspects of Toxicology1079

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1077

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PART VII  Toxicology

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48 Suspicion of Poisoning1107 Robert J. Flanagan, Graham R. Jones, Hans H. Maurer and Markus R. Meyer

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Olaf H. Drummer and Dimitri Gerostamoulos

Effects of Medicinal Drugs on  Fitness to Drive1423

62 Toxicological Markers of Chronic Alcohol Abuse1439 Fritz Pragst

63 Traffic Accidents1455

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50 Issues Affecting Interpretation: Stability and Artifacts1137

Frank Musshoff and Helena Fels

E. L. Theunissen, K. P. C. Kuypers, A. Vermeeren, Frederick Vinckenbosch and J. G. Ramaekers

49 Toxicological Analysis: Drug Screening and Confirmation1129

Gisela Skopp

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54 Effects of Cardiovascular Disease on Fitness to Drive1329

Heinz-­Dieter Wehner, Burkhard Madea and Guy Rutty ­

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51 Toxicokinetics/Toxicogenomics1155 Markus M. Meyer and Hans H. Maurer

52 Toxicology of Specific Substances1167 Alan Wayne Jones

PART IX  Identification

1495

64 Forensic DNA Analysis1497

PART VIII  Traffic Medicine

53 Driving Aptitude and Fitness to Drive1315 Frederick Vinckenbosch, Annemiek Vermeeren, Eef L. Theunissen and Johannes G. Ramaekers

Mechthild Prinz and Ruediger Lessig

1313 65

Forensic Anthropology1561 Cristina Cattaneo and Daniele Gibelli

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Forensic Odontostomatology1575 Rüdiger Lessig

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Brief Contents

PART X  The Doctor and the Law

1585

67 The Doctor and the Law1587

Further References

1637

Index1641

Brigitte Tag

PART XI  Insurance Medicine

1623

68 Personal Injury Assessment1625

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Teresa Magalhães and Duarte Nuno Vieira

Contents

Preface to the First Edition

xxvii

Foreword by Denis A. Cusack

xxix

Foreword by Duarte Nuno Vieira

xxxi

PART I Duties of Forensic Medicine in Modern Societies

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1 History3

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xxv

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Preface to the Second Edition

Peter Wiegand, Burkhard Madea and Frank Mußhoff

Burkhard Madea

PART II  Medical Aspects of Death 5

Duties of Forensic Medicine and the Forensic Medicine Practitioner19

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1.1 Definitions 3 1.2 Civilisations of the Near East and China4 1.3 Justinian enactments 6 1.4 Further developments and  Italian town charters 7 1.5 Forensic medicine as a book science 7 1.6 Forensic medicine as an experimental science 10 1.7 Current problems 15 References and further reading 16

Nature and Definition of Death59 5.1 5.2

Death and dying 59 Determination of death 60 References and further reading68

2.1 The Expert Witness Acknowledgements References and further reading

22 24 25

Forensic Medicine and  Human Rights

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Burkhard Madea and Antonella Argo

27 35 42

6.1 Introduction 6.2 Cause of death 6.3 Causes of death as shown by cause of death statistics

Hans Petter Hougen

3.1 Human rights issues 3.2 Torture Useful websites

57

Burkhard Madea

Burkhard Madea

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4.1 Introduction 43 4.2 Recommendations for forensic DNA laboratories based on  ISO/IEC 17025 47 4.3 Recommendations for forensic toxicological laboratories based on ISO/IEC 17025 49 4.4 Recommendations for forensic pathology based on ISO/IEC 1702553 4.5 Recommendations for clinical forensic medicine based on  ISO/IEC 17020 53 Appendix: Definitions (ENFSI Appendix 4) 54 Useful websites 54 References and further reading 54

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International Guidelines and Accreditation in Forensic Medicine43

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List of Contributors

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Brief Contents

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Certification of Death: External Postmortem Examination69 69 70 80

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Contents

10.1

10.2 Medicolegal Autopsy

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10.3 Clinical Autopsy

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198

Wolfgang Grellner

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References and further reading 205 10.6 Forensic Imaging and CT Angiography205 Garyfalia Ampanozi, Jakob Heimer, Lars C. Ebert and Michael J. Thali

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Peter J. T. Knudsen and Jørgen L. Thomsen

10.5 Exhumation

Claas T. Buschmann, Marc Windgassen and Michael Tsokos

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Peter J. T. Knudsen and Jørgen L. Thomsen

10.4 Autopsy Report

Cremation151

Short history of cremation Cremation in the modern day Process of cremation and modification of the corpse during cremation 8.4 Remnants of cremation 8.5 Conclusions 8.6 Embalming References and further reading

183

Peter J. T. Knudsen and Jørgen L. Thomsen

7.1 7.2

8.1 8.2 8.3

An Introduction to Autopsy Peter J. T. Knudsen and Jørgen L. Thomsen

Burkhard Madea, Claus Henßge, Saskia Reibe, and Michael Tsokos

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Peter J. T. Knudsen, Jørgen L. Thomsen, Wolfgang Grellner, Garyfalia Ampanozi, Jakob Heimer, Lars C. Ebert, Michael J. Thali, Burkhard Madea , and Antti Sajantila

Postmortem Changes and Time since Death91

Early Postmortem Changes 91 Later Postmortem Changes: Decomposition105 7.3 Timing of Death 111 7.4 Basic Forensic Entomology 125 References and further reading 132 7.5 Postmortem Injuries 133 References 142 References and further reading – Chapter 7.1–7.3 143

164 165 167 168 168 169 170 170 180

Autopsy183

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Consistency between cause of death diagnosis on the death certificate and following autopsy 80 6.5 Manner of death 81 6.6 Special constellations of circumstances in external postmortem examination 83 6.7 Checklist for the external postmortem examination 86 87 6.8 Identification of the corpse 6.9 Examination of the corpse 87 6.10 Completing the death certificate89 Useful website 89 References and further reading 89

9.4 Time of death 9.5 Examination of the body 9.6 Examination of the surroundings 9.7 Sample collection 9.8 Documentation 9.9 Cause and manner of death 9.10 Conclusions 9.11 Bloodstain pattern analysis References and further reading

6.4

References and further reading Bibliography 10.7 Molecular Pathology

151 151

217 218 222

Burkhard Madea

152 153 153 154 158

Crime Scene and Crime Scene Investigations161 Henrik Druid

9.1 General considerations 161 9.2 Equipment 162 9.3 Tasks for the forensic pathologist 163

References 10.8 Forensic Microbiology

232 235

Antti Sajantila

References 11

243

The Doctor, the Dead and the Relatives249 Antonella Argo and Valeria Tullio

11.1 Introduction 249 11.2 The work of mourning and its complexity249 11.3 Death, defence mechanisms, burnout, compassion fatigue and moral distress 250

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Contents

11.5



Death, communication and  reactions of family members 252 Assessment, recognition, cultural contexts, farewell to  the relative and death certificate253 References and further reading 253

Rüdiger Lessig with the assistance of Markus A. Rothschild and Klaus-Peter Benedix

14.9 Documentation and Quality Management297 Rüdiger Lessig and Mechthild Prinz



Useful websites 297 References and further reading297 14.10 Radiology Guidelines for Mass Fatalities299

12 Transplantation257 Brigitte Tag

12.1

Regulations and procedures of transplantation in selected countries257 12.2 Criteria for the diagnosis of  brain death 262 Acknowledgements 264 Useful websites 264 References and further reading 264

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References and further reading 14.11 Fire Fatalities

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References and further reading 14.12 Terrorist Attacks References

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PART III Traumatology and Violent Death311

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Mass Disaster Victim Identification291

15.1 Definitions 15.2 Basic legal principles 15.3 Conclusions 16

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294

Rüdiger Lessig and Mechthild Prinz

14.4 Standardisation

294 17

Rüdiger Lessig

14.5 Forensic Odontology Standards

295

Rüdiger Lessig

14.6 Forensic Molecular Biology Standards295 Rüdiger Lessig and Mechthild Prinz

14.7

Radiology Standards

296

Rüdiger Lessig

14.8 Chemical, Biological, Radiological, Nuclear Explosive Weapons 296

Traumatology and Criminology317 16.1 Introduction 16.2 Homicide 16.3 Non-­homicidal events 16.4 Suicide 16.5 Torture 16.6 Homicide, suicide or accident?

Rüdiger Lessig and Mechthild Prinz

14.3 DVI Team Structure

313 313 316

Philip Beh

An Introduction to Mass-Disaster Victim Identification 291 14.2 International Cooperation 292

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14.1

Legal Aspects of Traumatology and Violent Death313 Philip Beh

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Rüdiger Lessig and Mechthild Prinz

308

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Cristina Cattaneo and Daniele Gibelli

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303 304

Bertrand Ludes

Anthropology and Osteology267 13.1 Introduction 267 13.2 Preliminary steps 268 13.3 Diagnosis of species 272 13.4 Biological profile: the main role of anthropology273 13.5 Facial reconstruction 286 13.6 Geographical origin 287 Useful website 287 References and further reading 287

301 302

Guy Rutty

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Guy Rutty

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11.4

317 317 319 319 319 320

Some Preliminary Remarks Concerning Biomechanical Principles321 Burkhard Madea and Heinz-­Dieter Wehner

17.1 Introduction 321 17.2 Separation of coherence through the impact of tensile stress 321 17.3 Separation of coherence through the impact of shear stress 326 References and further reading 326

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Contents

Sequelae of Traumatic Injuries and Causes of Death327

21

Guy Rutty and Michael Tsokos

Bernd Karger

18.1 Introduction 18.2 Sepsis and multiple organ failure References and further reading 19

327 340 346

Vital Reactions349 Burkhard Madea, Elke Doberentz, Christian Jackowski, Wolfgang Grellner and Toshikazu Kondo

Vital reactions: An introduction

349

362

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References and further reading Wound-­Age Estimation: General Introduction and Methods

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Burkhard Madea, Elke Doberentz and Christian Jackowski

19.2

364

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Wolfgang Grellner

References and further readings 368 Wound-­Age Estimation: 368 Molecular Biology

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Toshikazu Kondo

References and further reading

374

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Veronique Henn

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References and further reading 371 19.4 Principles of Bone Fracture Healing 373

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20 Mechanical Trauma and Classification of Wounds375

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Burkhard Madea, Stefan Pollak, Annette Thierauf-­Emberger, Veronique Henn, Christoph Meissner, Manfred Oehmichen, and Peter Mygind Leth

20.1

Classification of Violence

375

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Burkhard Madea

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Stefan Pollak and Annette Thierauf-Emberger

Additional references 20.3 Homicides by Kicking

409 409

Veronique Henn

References and further reading 20.4 Forensic Neuropathology

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Asphyxiation505 Burkhard Madea, Wolfgang Keil, Philippe Lunetta, and Mattias Kettner

References 507 22.1 Injuries due to Asphyxiation and Drowning508 Wolfgang Keil, Philippe Lunetta, Richard Vann, and Burkhard Madea

380

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20.2 Blunt Force Injury

21.1 Introduction 459 21.2 Firearms 459 21.3 Ammunition 463 21.4 Shotgun ammunition 465 21.5 Interior ballistics 466 466 21.6 Exterior ballistics 21.7 Intermediate targets 467 21.8 Wound ballistics 467 21.9 Incapacitation 473 21.10 Forensic reconstruction 474 21.11 Differentiation of suicide and homicide487 21.12 The hand firing or steadying the gun488 21.13 Prior exposure of the entrance wound489 21.14 Defensive injuries 489 21.15 Accidents 489 21.16 Miscellaneous 490 21.17 Arrow wounds 491 492 21.18 Explosive injuries 21.19 Mechanisms of injury 492 References495

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19.1

Forensic Ballistics: Injuries from Gunshots, Explosives and Arrows459

411 412

References and further reading 22.2 Drowning Philippe Lunetta

References 573 22.3 Barotrauma and Diving-­Related Accidents576 Philippe Lunetta, Richard Vann, and Arthur Heickell

Christoph Meissner and Manfred Oehmichen

References and further reading 20.5 Sharp Force Injury

445 448

References 22.4 Immersion Time

588 589

Burkhard Madea

Peter Mygind Leth

References

549 556

457



References and further reading

596

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Contents

22.5 Death in High Altitude and  Barotrauma596

27

Infanticide683 Thomas Bajanowski, Janine Helmus, Mechtild Vennemann and Sibylle Banaschak

Mattias Kettner

References

603

27.1 Stillbirth 683 27.2 Neonaticide 685 27.3 Investigation of mothers and  neonates686 References 694

23 Injuries Due to Heat607 Burkhard Madea and Elke Doberentz

28 Death During Pregnancy697

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24 Injuries Due to Cold627

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Burkhard Madea and Elke Doberentz

24.1 Introduction 627 24.2 Pathophysiology 627 24.3 Clinical phases of hypothermia 629 24.4 Epidemiology 630 24.5 Morphological and biochemical changes630 24.6 Criminalistic aspects 636 References and further reading 637 25 Electrocution and Lightning639 Ryan Blumenthal

639 652 661 661 662

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25.1 Electrocution 25.2 Lightning 25.3 Conclusion Useful websites References and further reading

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26 Starvation and Neglect669 Burkhard Madea and Elke Doberentz

26.1 Introduction 669 26.2 Clinical starvation 669 26.3 Fatal starvation 671 26.4 Classification systems 674 26.5 Death from starvation 676 26.6 Physical neglect 677 26.7 Sarcopenia 678 26.8 Glossary 679 References and further reading680

28.1 Analysis of maternal mortality 697 28.2 Pregnancy as a physiological process with specific risks 697 28.3 Maternal death directly due to gestation698 28.4 Maternal death indirectly due to gestation703 703 28.5 Iatrogenic maternal death 28.6 Maternal death not due to gestation704 Useful websites 705 References and further reading 705

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23.1 Introduction 607 23.2 Burns from heat 607 617 23.3 Causes of death 23.4 Scalding 621 23.5 Generalised heat damage and hyperthermia622 23.6 Diagnosis 623 References and further reading 624

29 Autoerotic Death707 Marc Windgassen

29.1 29.2 29.3 29.4



General aspects 707 Categorisation of death cases 710 Possible mechanisms of death 710 Typical injuries to male genitalia and special cases, such as chemsex712 References and further reading 713

30 Death in an Abnormal Position: Physical Restraint717 John C. Hunsaker III and Lisa B. E. Shields

30.1 30.2 30.3 30.4 30.5

30.6

In-­custody and restraint death717 Extrinsic factors 719 Intrinsic factors 722 Controversies: excited delirium 723 Investigation of death in custody: restraint and  body position 724 Certification of death 725 References and further reading 726

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31

Contents

33.6

Sexual Homicide729

Mafia-­Related Deaths

Philip Beh



729 730 731 731 734

34 Natural Causes of Sudden Death803 Emanuela Turillazzi, Alessandro Santurro, Pietrantonio Ricci, Matteo Scopetti and Vittorio Fineschi

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Burkhard Madea

32.1 Introduction 735 735 32.2 Definitions 32.3 Epidemiology 736 751 32.4 Value of autopsy 32.5 New approaches 751 32.6 Conclusions 752 References and further reading752 32.7 Autopsy Findings and  Interpretation after Unsuccessful Resuscitation 755

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34.1 Introduction 34.2 Cardiac Causes of Sudden Death 34.3 Non-­Cardiac Causes of  Sudden Death References and further readings 34.4 COVID-­19-­associated Deaths

816 828 832

Klaus Püschel, Hanna Goebels and Fabian Heinrich

References and further reading

841

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761

35 Postmortem Biochemistry as an Aid in Determining the Cause of Death845

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Further reading

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33 Special Issues Regarding Expert Evidence in Violent Death763

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Johanna Preuß-­Wössner, Rainer Amberg, Burkhard Madea, Jørgen L. Thomsen, Martin Worm-­Leonhard, and Paolo Procaccianti

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Priority and Order of Injuries

763

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Johanna Preuß-­Wössner, Rainer Amberg, Burkhard Madea

765

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33.2 Physical Activity After Injuries and Survival Time Johanna Preuß-­Wössner, Rainer Amberg, Burkhard Madea

33.3

803 803

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Claas T. Buschmann

33.1

798

PART IV Sudden and Unexpected Death from Natural Causes 801

Medical Malpractice735



References and further reading

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31.1 Introduction 31.2 Profiling offenders in a sex-­related homicide 31.3 Crime assessment 31.4 Conclusions References and further reading 32

792

Paolo Procaccianti

Post-­Mortem Mutilation and Dismemberment767

Johanna Preuß-­Wössner, Rainer Amberg, Burkhard Madea



References and further reading771 33.4 Alcohol-­Related Deaths 772 Jørgen L. Thomsen

33.5 Death Due to Drug Addiction Jørgen L. Thomsen and Martin Worm-Leonhard

Tomoya Ikeda, Naoto Tani, Takaki Ishikawa, Gerhard Kernbach-Wighton, Aurelio Luna

35.1 Post-­mortem Biochemistry and Molecular Biology Tests

845

Tomoya Ikeda, Naoto Tani, Takaki Ishikawa

35.2 Indicators in each tissue 854 35.3 Conclusion 858 Acknowledgments 859 References 859 35.3 Integrating Data from Human Autopsy Investigations with Findings of Basic Medical Research864 Tomoya Ikeda, Naoto Tani, Takaki Ishikawa

Acknowledgments References 35.4 Post-­mortem Biochemistry as  an Aid in Determining the  Cause of Death

870 870

871

Gerhard Kernbach-­Wighton and Aurelio Luna

783

References and further reading

883

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Contents

39.3 Causes and reasons for  domestic violence 39.4 Effects and consequences of domestic abuse 39.5 The cycle of abuse 39.6 Diagnostic procedures References and further reading

36 Sudden and Unexpected Deaths in Infants and Sudden Infant Death Syndrome887 Thomas Bajanowski, Mechtild M. Vennemann and Jan-­Peter Sperhake

Gerhard Kernbach-­Wighton

N

LY

40.1 Introduction 995 40.2 Statistical data from various countries996 40.3 Motives and reasons for school violence and its identification 997 40.4 Prevention and interventional procedures997 References and further reading 998

917

Cristina Cattaneo, Daniele Gibelli and Marina Ruspa

41

Gerhard Kernbach-­Wighton

John C. Hunsaker III and Lisa B.E. Shields

42.1 Introduction 1005 42.2 Definition and scope of elder mistreatment1005 42.3 Types of abuse of the elderly 1007 42.4 Mimics of elder abuse 1010 42.5 Clinical evaluation in  1010 living patients 42.6 Medicolegal investigations 1012 42.7 Conclusions 1015 Useful websites 1015 References and further reading 1016

O

C

Non-­Accidental Head Injury in Children939

R

38

FO

Robert A. Minns

38.1

Non-­Accidental Head Injury in Children939

Robert A. Minns

References and further reading 38.2  Child Abuse

963 970

Burkhard Madea and Elke Doberentz



References and further reading

984

39 Intimate Partner and Domestic Violence987 Saskia Etzold

39.1 Introduction 39.2 Forms of domestic violence

987 987

41.1 Introduction 1001 41.2 Characteristics of gay violence 1001 41.3 Violence in straight versus homosexual relationships 1002 41.4 Problems with statistical data and underlying causes 1002 References and further reading 1003

42 Violence Against the Elderly1005

N

TR IB

U

TO

37.1 Introduction 919 37.2 Examination of the victim 920 37.3 Sexually transmitted infections 930 37.4 Toxicological analysis 932 37.5 Legal outcome 932 37.6 Juvenile pornography 933 Appendix 37.1 Approach to interpretation of medical findings in suspected sexual abuse 933 Appendix 37.2 Reminder checklist935 References and further reading 936

Violence Against Homosexuals1001

U

Sexual Abuse919

R

37

School Violence995

SE

PART V  Clinical Forensic Medicine

40

991 993 994 994

O

36.1 Introduction 887 36.2 Sudden infant death syndrome 888 36.3 Other main causes of death in SUDI cases and during childhood900 36.4 Practical investigation of  SUDI/SIDS905 36.5 Conclusions 906 References and further reading 906

991

43

Self-­harm1021 Saskia Etzold

43.1 Introduction and epidemiology 43.2 Classification and causes of ­ self-­harm groups 43.3 Characteristics of  self-­inflicted injuries

1021 1021 1023

xvi

Contents

43.4 Diagnostic procedures References and further reading

1024 1026

46.4 Drugs of abuse 46.5 Postmortem toxicology 46.6 Clinical forensic toxicology 46.7 Doping 46.8 Veterinary forensic toxicology 46.9 Sabotage, terrorism and chemical warfare Useful websites References and further reading

44 Medical Age Assessment in  Living Individuals1027 Andreas Schmeling and Ernst Rudolf

LY

47 Epidemiology and Adverse Drug Reactions1099

N

Olaf H. Drummer, Dimitri Gerostamoulos and Jennifer L. Schumann

O

SE

1055

TO

45 Forensic Psychiatry1057 Alan R. Felthous and Henning Saß

FO

R

C

O

N

TR IB

U

45.1 Definitions 1057 45.2 Inquisitorial versus adversarial legal systems 1058 45.3 Structure of forensic psychiatric examinations1059 45.4 Civil law, competencies and  general legal competency 1060 45.5 Competencies in criminal law 1061 45.6 Criminal responsibility 1063 45.7 Forensic security hospitals 1070 45.8 Formal education in the subspeciality of forensic psychiatry1071 45.9 Conclusions 1072 Legislation (codes and statutes) 1072 Landmark court cases (rulings and opinions) 1072 References and further reading 1072

PART VII  Toxicology

1077

46 Legal Aspects of Toxicology1079 Ilkka Ojanperä

46.1 Introduction 46.2 History of poisoning 46.3 Administrative issues

47.1 Introduction 1099 47.2 Investigating a Possible 1099 Adverse Drug Reaction 47.3 Epidemiology 1100 47.4 Evidence basis for toxicity 1100 47.5 Substandard and Falsified medical products 1102 47.6 Metabolic Aspects 1103 47.7 Transporters and Other Types of ADR1103 47.8 Conclusions 1104 Useful websites 1104 References and further reading 1104

R

PART VI  Forensic Psychiatry

1095 1096 1096

U

44.1 Introduction 1027 44.2 Medical age assessment in adolescents and young adults 1028 44.3 Age assessment in child pornographic image documents 1041 44.4 Age assessment in old-­age pension proceedings 1042 44.5 Age assessment in competitive sports1042 1043 44.6 Quality assurance 44.7 Ethical considerations in medical age assessment 1044 References and further reading 1046

1083 1086 1091 1095 1095

1079 1079 1080

48 Suspicion of Poisoning1107 Robert J. Flanagan, Graham R. Jones, Hans H. Maurer and Markus R. Meyer

48.1 Diagnosis of acute poisoning

1107

Robert J. Flanagan

48.2 Specimen collection for  forensic toxicology

1118

Graham R. Jones

48.3 Clinical signs and syndromes in intoxication1124 Hans H. Maurer and Markus R. Meyer



Useful website References and further reading

1127 1127

49 Toxicological Analysis: Drug Screening and Confirmation1129 Olaf H. Drummer and Dimitri Gerostamoulos

49.1 Introduction 49.2 Screening and confirmation tests 49.3 Isolation step 49.4 Instrumentation used in  forensic toxicology

1129 1130 1130 1131

xvii

Contents

49.5 49.6

Analytical methods 1133 Method performance 1134 Useful websites 1135 References and further reading1135

52 Toxicology of Specific Substances1167 A.W. Jones, F. Musshoff, T. Krämer, A. E. Steuer, D. Gerostamoulos, O.H. Drummer, G. Drasch, M. Balikova, J. Beyer, H. Teixeira, M. Thevis, W. Schänzer, H. Druid, and G. Skopp

52.1 50 Issues Affecting Interpretation: Stability and Artifacts1137

52.2 Chemistry of alcohols  52.3 Physicochemical properties  52.4 Determination in body fluids  52.5 Disposition and fate in the body 52.6 Mono-hydroxy alcohols  52.7 Aliphatic diols  52.8 Summary and conclusions  References 52.2 Illegal drugs

O

N

LY

Gisela Skopp

U

TR IB

N

O

C

51 Toxicokinetics/Toxicogenomics1155

R

Introduction and the LADME principle 51.2 Basic kinetic parameters 51.3 Toxicokinetics and toxicogenomics in forensic and clinical toxicology 51.4 Liberation/Release 51.5 Absorption 51.6 Distribution 51.7 Metabolism 51.8 Excretion 51.9 Conclusions References Further reading

FO

51.1

1155 1156

1168 1169 1169 1171 1175 1184 1188 1190 1196

SE

U

References and further reading 1209 52.3 New Psychoactive Drugs 1224 Olaf H. Drummer

R

References 52.4 Sedatives, Hypnotics

1234 1236

Thomas Kräemer and Andrea E. Steuer

References 1248 52.5 Narcotics and Other Analgesics 1252 Dimitri Gerostamoulos and Olaf H. Drummer

References 52.6 Toxic Elements

1257 1257

Gustav Drasch

References and further reading 52.7 Solvents and Gaseous Poisons

1265 1266

Marie A. Balíková

References and further reading 52.8 Natural Poisons

Markus M. Meyer and Hans H. Maurer

1167

Frank Musshoff

TO

50.1 Introduction 1137 50.2 Definition of stability and common causes of instability and artificial formation 1138 50.3 Stability of particular drugs and potential artifacts in blood collected from living individuals 1139 50.4 Stability of major drugs and  potential artifacts in urine collected from living individuals 1140 50.5 Stability of drugs and formation of artifacts in postmortem specimens1141 50.6 Sampling artifacts and measures to increase drug stability 1143 50.7 Influence of storage 1145 50.8 Instability of drugs and artificial formation during processing and analysis1146 50.9 Procedures for evaluating stability and artifacts 1148 50.10 Conclusions 1150 References and further reading 1150

Aliphatic Alcohols  Alan Wayne Jones

1273 1274

Jochen Beyer

References and further reading 52.9 Pesticides and Insecticides

1278 1278

Helena Teixeira

1158 1158 1158 1160 1161 1164 1165 1165 1165

52.10 Doping

1284

Mario Thevis and Wilhelm Schänzer

References 52.11 Cardiovascular drugs

1291 1292

Hiroshi Kinoshita

References 52.12 Paracetamol/Acetaminophen

1298 1299

Toshikazu Kondo

References

1301

xviii

Contents

52.13 Interpretation of Postmortem Forensic Toxicological Results

56.3 56.4 56.5 56.6

Legal aspects 1356 Individual risk increase 1356 Anti-seizure medication 1356 Epilepsy and general road safety1357 56.7 Outlook 1357 References 1357

1302

Henrik Druid and Gisela Skopp



References and further reading

PART VIII  Traffic Medicine

1311

1313

53 Driving Aptitude and Fitness to Drive1315 Frederick Vinckenbosch, Annemiek Vermeeren, Eef L. Theunissen and Johannes G. Ramaekers

57 Effects of Diabetes on Fitness to Drive1359 Wulf Quester

53.1 Introduction 1315 53.2 Empirical methods for identifying risk factors for road traffic accidents1316 53.3 Individual assessment of driving aptitude and fitness to drive 1322 53.4 Conclusion 1324 References and further reading 1324

U

SE

O

N

LY

57.1 Classification of diabetes 1359 57.2 Stages of diabetes 1361 57.3 Driving and treatment of diabetes1364 57.4 Driving performance and  diabetes1366 57.5 Therapeutic aspects for  1370 driving safely with diabetes 57.6 People with diabetes as victims of traffic injuries 1371 57.7 Conclusions 1372 References and further reading 1372

R

54 Effects of Cardiovascular Disease on Fitness to Drive1329

TO

Werner Jung

58 Epidemiology and Causal Factors in Fitness to Drive1375

N

TR IB

U

54.1 Introduction 1329 54.2 Derivation of the risk of harm formula1331 54.3 Coronary heart disease 1332 54.4 Arrhythmias 1333 54.5 Syncope 1338 54.6 Heart failure 1340 References and further reading 1341

O

55 Effects of Vision and Visual Fields on Fitness to Drive1345

C

Patrick Vivell

FO

R

55.1 Introduction 1345 55.2 Problems with vision 1345 55.3 Visual field disorders 1350 55.4 Condition after an eye operation1351 55.5 Problems of vision in older motorists1351 55.6 Effects of visual deficiencies 1352 References and further reading1353 56 Effects of Epilepsy on Fitness to Drive1355 Randi von Wrede

56.1 General rules 56.2 Background

1355 1356

Günter Berghaus

58.1 Introduction 1375 58.2 Epidemiological methods 1375 58.3 Examples of epidemiological studies1378 References and further reading 1385 59 Driving Under the Influence of  Alcohol1387 Alan Wayne Jones

59.1 Introduction 1387 59.2 Properties of alcohol 1388 59.3 Medicolegal alcohol determinations1389 59.4 Statutory alcohol limits for driving1394 59.5 Characteristics of traffic offenders1395 59.6 Alcohol ingestion and crash risk1398 59.7 Alcohol tolerance 1401 1402 59.8 Concluding remarks References 1404

xix

Contents

63.3 Motor-­Vehicle-­Assisted Suicide

60 Effects of Illegal Drugs on Fitness to Drive1409



References and further readings1472 1474 63.4 Railroad-­Related Deaths

Frank Musshoff and Helena Fels

60.1 Introduction 1409 60.2 Epidemiological data and its limitations1410 1411 60.3 Detection of impaired driving 60.4 Effects of drugs on driving 1411 60.5 Forensic toxicological analyses1416 References and further reading 1419

Guy Rutty

LY

References and further reading 1478 63.5 Aviation Fatalities 1479 Further reading 1489 References 1489 63.6 Transport Deaths Related to Animals, Wheelchairs, Mobility Scooters and Rickshaws 1490 References and further reading 1492

O

E. L. Theunissen, K. P. C. Kuypers, A. Vermeeren, Frederick Vinckenbosch and J. G. Ramaekers

N

Effects of Medicinal Drugs on  Fitness to Drive1423

Mechthild Prinz and Rüdiger Lessig

U

64.1 Stain Analysis 64.2 Paternity Testing

U 65

O

C

R

FO

63.1

Road Traffic Accidents

66 1455

Heinz-­Dieter Wehner and Burkhard Madea

Bibliography 1465 63.2 Sudden Natural Death while Driving1465 Guy Rutty



References and further reading

1467

64.3 Forensic DNA Phenotyping and Biogeographical Ancestry Determination1530 References 1543 Forensic Anthropology1561 65.1 Introduction 1561 65.2 Identification of human remains1562 65.3 Identification of the living 1567 65.4 The issue of quantification 1569 65.5 Not only identification: the  1570 analysis of trauma References and further reading 1572

63 Traffic Accidents1455 Heinz-­Dieter Wehner, Burkhard Madea and Guy Rutty ­

1525

Rüdiger Lessig and Mechthild Prinz

Cristina Cattaneo and Daniele Gibelli

N

TR IB

62.1 Introduction 1439 62.2 Trait markers and polygenic risk scores of alcohol use disorder1439 62.3 State biomarkers of alcohol consumption1440 62.4 Detection of chronic excessive alcohol consumption 1447 62.5 Control of abstinence 1447 References and further reading 1449

1497

Mechthild Prinz

62 Toxicological Markers of Chronic Alcohol Abuse1439 Fritz Pragst

1495

64 Forensic DNA Analysis1497

TO

61.1 Introduction 1423 61.2 Medicinal drugs that impair driving1424 61.3 Conclusions 1433 References and further reading 1434

SE

PART IX  Identification

R

61

1468

Guy Rutty

Forensic Odontostomatology1575 Rüdiger Lessig

66.1 Introduction 1575 66.2 Dental charting 1575 66.3 Age estimation 1577 66.4 Identification of an unknown person1579 66.5 Bite marks 1580 References and further reading 1583

xx

Contents

PART X  The Doctor and the Law

The Doctor and the Law1587

PART XI  Insurance Medicine

1623

68 Personal Injury Assessment1625 Teresa Magalhães and Duarte Nuno Vieira

67.1

68.1 Introduction 1625 68.2 Aim of personal injury 1626 assessment and reparation 68.3 General aspects for forensic medical assessment 1627 68.4 Specifics of forensic medical reports for personal injury assessment1630 68.5 Conclusions 1634 References and further reading 1634

N

O

SE

Further References

R TO U TR IB N O C R

1637

Index1641

U

Legal relevance of medical treatment1587 67.2 Legal aspects of personalised medicine1595 67.3 Assisted suicide, organised medically assisted suicide, and euthanasia1605 1610 67.4 Research on corpses 67.5 Legal and ethical aspects concerning the handling of  corpses and the display of  human remains 1615 Acknowledgements 1618 References and further reading 1618

LY

Brigitte Tag

FO

67

1585

List of Contributors

Budowle, Bruce, Center for Human Identification, University of North Texas Health Science Center, Texas, USA Bajanowski, Thomas, Institute of Legal Medicine, University of Duisburg-­Essen, Essen, Germany

LY

N

O

Drasch, Gustav, Institute of Forensic Medicine, University of Munich, Munich, Germany Druid, Henrik, Karolinska Institute Stockholm, Stockholm, Sweden Drummer, Olaf H., Victorian Institute of Forensic Medicine, Monash University, Southbank Melbourne, Victoria, Australia

U

TO

Balíková, Marie A., Institute of Forensic Medicine and Toxicology, First Faculty of Medicine and General Teaching Hospital, Charles University, Prague, Czech Republic

Doberentz, Elke, Institute of Forensic Medicine, University of Bonn, Bonn, Germany

SE

Argo, Antonella, Department of Biotechnology and Legal Medicine, Section of Legal Medicine, Palermo, Italy

Cattaneo, Cristina, Institute of Legal Medicine, Department of Biomedical Sciences for Health, University of Milan, Milan, Italy

U

Ampanozi, Garyfalia, Institute of Forensic Medicine, University of Zurich, Zurich, Switzerland

Budowle, Bruce, Center for Human Identification, University of North Texas Health Science Center, Texas, USA

R

Amberg, Rainer†, Wallstr. 20, Freiburg, Germany

TR IB

Banaschak, Sibylle, Institute of Forensic Medicine, University of Cologne, Cologne, Germany

N

Beh, Philip, Department of Pathology, University of Hong Kong, Hong Kong

O

Benedix, Klaus-­Peter, Bundeswehr Joint Medical Service, Regional Medical Support Command, Diez, Germany

C

Berghaus, Günter, Hohkeppeler Str. 76, Overath, Germany

FO

R

Beyer, Jochen, Department of Forensic Medicine, Victorian Institute of Forensic Medicine, Monash University, Southbank Melbourne, Victoria, Australia Biggs, Mike, Home Office Registered Forensic Pathologist, East Midlands Forensic Pathology Unit, Leicester Royal Infirmary, Leicester, UK Blumenthal, Ryan, Department of Forensic Medicine, University of Pretoria, Pretoria, South Africa Buschmann, Claas T., Institute of Legal Medicine, University Hospital Schleswig-­Holstein, Campus Lübeck: Kahlhorststr, 23562 Lübeck, Germany; Campus Kiel: Arnold-­Heller-­Str., 24105 Kiel, Germany

Ebert, Lars C., 3D Center Zurich, Institute of Forensic Medicine, University of Zurich, Zurich, Switzerland Fels, Helena, FTC München, Forensisch Toxikologisches Centrum GmbH, Munich, Germany Felthous, Alan R., Department of Neurology and Psychiatry, Saint Louis University, St Louis, MO, USA Fineschi, Vittorio, Department of Forensic Sciences, University Sapienza of Rome and Forensic Unit, Sant’Andrea Hospital, Rome, Italy Flanagan, Robert J., Toxicology Unit, Clinical Biochemistry, King’s College Hospital, London, UK Gerostamoulos, Dimitri, Victorian Institute of Forensic Medicine, Monash University, Southbank Melbourne, Victoria, Australia Gibelli, Daniele, Institute of Legal Medicine, Department of Biomedical Sciences for Health, University of Milan, Milan, Italy Goebels, Hanna, University Medical Center HamburgEppendorf, Institute of Legal Medicine, Hamburg, Germany Grellner, Wolfgang, Department of Legal Medicine, Medical University of Göttingen, Göttingen, Germany

xxii

List of Contributors

Heimer, Jakob, Institute of Forensic Medicine, University of Zurich, Zurich, Switzerland

Kernbach-­Wighton, Gerhard, Institute of Forensic Medicine, University of Bonn, Bonn, Germany

Heinrich, Fabian, University Medical Center HamburgEppendorf, Institute of Legal Medicine, Hamburg, Germany

Kettner, Mattias, University of Frankfurt, Institute of Forensic Medicine, Frankfurt, Germany

Helmus, Janine, Institute of Legal Medicine, University of Duisburg-Essen, Essen, Germany

Kinoshita, Hiroshi, Department of Forensic Medicine, Faculty of Medicine, Kagawa University, Kita, Kagawa 761-­0793

Henn, Veronique, Werdohler Str. 52, Lüdenscheid, Germany

Knudsen, Peter J. T., Institute of Forensic Medicine, University of Southern Denmark, Odense C, Denmark

Henßge, Claus†, Franzensbader Str. 24, Berlin, Germany

LY

Kondo, Toshikazu, Department of Forensic Medicine, Wakayama Medical University, Wakayama, Japan

Hougen, Hans Petter, Department of Forensic Medicine, Faculty of Health Sciences, University of Copenhagen, Copenhagen, Denmark

O

N

Kraemer, Thomas, Department of Forensic Pharmacology and Toxicology, Institute of Forensic Medicine, University of Zurich, Zurich, Switzerland

SE

U

Lessig, Rüdiger, Institute of Forensic Medicine, University of Halle (Saale), Halle (Saale), Germany

TO

Ikeda, Tomoya, Department of Legal Medicine, Osaka City University Medical School, Abeno, Osaka 545-­8585, Japan; Forensic Autopsy Section, Medico-­legal Consultation and Postmortem Investigation Support Center, Department of Legal Medicine, Osaka City University Medical School, Abeno, Osaka 545-­8585, Japan;

Kuypers, Kim P. C., Neuropsychology and Psychopharmacology, Faculty of Psychology and Neuroscience, University of Maastricht, Maastricht, the Netherlands

R

Hunsaker, John C., III, Justice and Public Safety Cabinet, Medical Examiner Division, Frankfort, KY, USA

U

Leth, Peter Mygind, Institute of Forensic Medicine, University of Southern Denmark, Odense C, Denmark

N

TR IB

Ishikawa, Takaki, Department of Legal Medicine, Osaka City University Medical School, Abeno, Osaka 545-­8585, Japan; Forensic Autopsy Section, Medico-­legal Consultation and Postmortem Investigation Support Center, Department of Legal Medicine, Osaka City University Medical School, Abeno, Osaka 545-­8585, Japan

C

O

Jackowski, Christian, Institute of Forensic Medicine, University of Bern, Bern, Switzerland

FO

R

Jones, Alan Wayne, Department of Forensic Toxicology, National Board of Forensic Medicine, Linköping, Sweden Jones, Graham R., Alberta Medical Examiners Office, Edmonton, Alberta, Canada Jung, Werner, Department of Cardiology, Academic Teaching Hospital Villingen of the University of Freiburg, ­Villingen-­Schwenningen, Germany

Lignitz, Eberhard, Werdohler Str. 52, Lüdenscheid, Germany Ludes, Bertrand, Université de Paris, BABEL, CNRS, F-­75012 Paris, France Luna, L. Aurelio, Department of Legal Medicine, Medical School, University of Murcia, Murcia, Spain Lunetta, Philippe, Department of Forensic Medicine, University of Helsinki, Helsinki, Finland Madea, Burkhard, Institute of Forensic Medicine, University of Bonn, Bonn, Germany Magalhães, Teresa, National Institute of Legal Medicine and Forensic Sciences (North Branch), University of Porto, Porto, Portugal Maurer, Hans H., Department of Experimental and Clinical Toxicology, University Hospital, Homburg/Saar, Germany

Karger, Bernd, Institute of Legal Medicine, University of Münster, Münster, Germany; bernd

Meissner, Christoph, Institute of Forensic Medicine, University of Lübeck, Lübeck, Germany

Keil, Wolfgang, Institute of Forensic Medicine, University of Munich, Munich, Germany

Meyer, Markus R., Department of Experimental and Clinical Toxicology, Saarland University, Homburg/Saar, Germany

xxiii

List of Contributors

Minns, Robert A., College of Medicine and Veterinary Medicine, University of Edinburgh, Edinburgh, UK; Robert

Rutty, Guy N., East Midlands Forensic Pathology Unit, University of Leicester, Leicester, UK

Mußhoff, Frank, Institute of Forensic Medicine, University of Bonn, Bonn, Germany

Sajantila, Antti, Department of Forensic Medicine, University of Helsinki, Finland; Forensic Medicine Unit, Finnish Institute of Health and Welfare, Finland

Oehmichen, Manfred, Institute of Forensic Medicine, University of Lübeck, Lübeck, Germany

Santurro, Alessandro, Department of Forensic Sciences, University Sapienza of Rome and Forensic Unit, Sant’Andrea Hospital, Rome, Italy

Ojanperä, Ilkka, Hjelt Institute, Department of Forensic Medicine, University of Helsinki, Helsinki, Finland

N

O

Scopetti, Matteo, Department of Forensic Sciences, University Sapienza of Rome and Forensic Unit, Sant’Andrea Hospital, Rome, Italy Shields, Lisa B. E., Norton Neuroscience Institute, Kentucky 40202, USA

TO

Procaccianti, Paolo, Department of Biotechnology and Legal Medicine, Section of Legal Medicine, Palermo, Italy

SE

Prinz, Mechthild, Formerly Department of Sciences, John Jay College of Criminal Justice, New York, NY, USA

Schumann, Jennifer L., Head Drug Harm Prevention Unit, Victorian Institute of Forensic Medicine, Monash University, Southbank, Australia

U

Preuß-­Wössner, Johanna, University of Frankfurt, Institute of Forensic Medicine, Frankfurt, Germany

Schmeling, Andreas, Institute of Legal Medicine, University of Münster, Münster, Germany

R

Pragst, Fritz, Forensic Toxicology, Institute of Legal Medicine, University Hospital Charité, Berlin, Germany

LY

Saß, Henning, University Hospital of Psychiatry, University of Aachen, Aachen, Germany

Pollak, Stefan, Institute of Legal Medicine, University of Freiburg, Freiburg, Germany

TR IB

U

Püschel, Klaus, University Medical Center HamburgEppendorf, Institute of Legal Medicine, Hamburg, Germany

Skopp, Gisela, Institute of Legal Medicine and Traffic Medicine, Heidelberg, Germany

Sperhake, Jan-­Peter, University Medical Center HamburgEppendorf, Institute of Legal Medicine, Hamburg, Germany

Ramaekers, Johannes G., Neuropsychology and Psychopharmacology, Faculty of Psychology and Neuroscience, University of Maastricht, Maastricht, the Netherlands

Steuer, Andrea E., Institute of Forensic Medicine, Department of Forensic Pharmacology and Toxicology, University of Zurich, Zurich, Switzerland

C

O

N

Quester, Wulf, Heart and Diabetes Center North ­Rhine-­Westphalia, Ruhr University Bochum, Bad Oeynhausen, Germany

Tag, Brigitte, Institute of Law, University of Zurich, Zurich, Switzerland

Etzold, Saskia, Institute of Legal Medicine and Forensic Science, University Medicine Berlin, Berlin, Germany

Tani, Naoto, Department of Legal Medicine, Osaka City University Medical School, Abeno, Osaka 545-­8585, Japan; Forensic Autopsy Section, Medico-­legal Consultation and Postmortem Investigation Support Center, Department of Legal Medicine, Osaka City University Medical School, Abeno, Osaka 545-­8585, Japan;

FO

R

Reibe, Saskia, Institute of Forensic Medicine, University of Bonn, Bonn, Germany

Ricci, Pietrantonio, Istituto di Medicina Legale, Ospandale “C. D’Avanzo” Rothschild, Markus A., Institute of Forensic Medicine, University of Cologne, Cologne, Germany Rudolf, Ernst, Attnang-­Puchheim, Austria

Teixeira, Helena, National Institute of Legal Medicine and Forensic Sciences (Center Branch), University of Coimbra, Largo de Sé Nova, Coimbra, Portugal

Ruspa, Marina, SVS (Sex Violence Aid Centre), Policlinic of Milan, ICCRS Cà Granda, Milan, Italy

Thali, Michael J., University of Zurich IRM/Forensic Institute, Zurich, Switzerland

xxiv

List of Contributors

Theunissen, Eef L., Neuropsychology and Psychopharmacology, Faculty of Psychology and Neuroscience, University of Maastricht, Maastricht, the Netherlands Thevis, Mario, Institute of Biochemistry, Center for Preventive Doping Research, German Sport University Cologne, Cologne, Germany

Vennemann, Mechtild M., Institute of Legal Medicine, University of Münster, Münster, Germany Vermeeren, Annemiek, Neuropsychology and Psychopharmacology, Faculty of Psychology and Neuroscience, University of Maastricht, Maastricht, the Netherlands

Thomas, Andreas, Institute of Biochemistry, Center for Preventive Doping Research, German Sport University Cologne, Cologne, Germany

Vinckenbosch, Frederick, Department of Neuropsychology and Psychopharmacology, Faculty of Psychology and Neuroscience, Maastricht University, Maastricht, The Netherlands

Tsokos, Michael, Institute of Legal Medicine and Forensic Sciences, Charité – University Medicine Berlin, Berlin, Germany

N

von Wrede, Randi, University Medicine Bonn, Bonn, Germany

SE

Wehner, Heinz-­Dieter (†), Society of Forensic Medical Examinations and Expert Services (GRUS), Tübingen, Germany

R

U

Wiegand, Peter, Institute of Forensic Medicine, University of Ulm, Ulm, Germany Windgassen, Marc, Institute of Legal Medicine and Forensic Science, Charité – University Medicine Berlin, Berlin, Germany

TO

Turillazzi, Emanuela, Institute of Forensic Pathology, University of Foggia, Foggia, Italy

Worm-­Leonhard, Martin, Institute of Forensic Medicine, University of Sourthern Denmark, Odense C, Denmark

FO

R

C

O

N

TR IB

U

Vann, Richard, Department of Anesthesiology, Center for Hyperbaric Medicine and Environmental Physiology, Duke University Medical Center, Divers Alert Network, Durham, NC, USA

Vivell, Patrick, Bruchsal Eye Hospital, Bruchsal, Germany

O

Thomsen, Jørgen L., Institute of Forensic Medicine, University of Southern Denmark, Odense C, Denmark

LY

Thierauf, Annette, Institute of Legal Medicine, University of Freiburg, Freiburg, Germany

Vieira, Duarte Nuno, National Institute of Legal Medicine and Forensic Sciences, University of Coimbra, Largo de Sé Nova, Coimbra, Portugal

Preface to the Second Edition

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During the preparation of the new edition, a chapter on the medicolegal aspects of COVID-­19 infections had of course to be included. Experts in forensic medicine could contribute to the understanding of the disease by performing systematic autopsies and doing research on the autopsy specimens (histology, immunohistochemistry, molecular pathology, etc.) together with clinical collaborations. The Corona pandemic and especially the financial consequences will accompany us probably for decades. I have to thank the publisher for making a second edition possible. Working together with Jenny Cossham, Emma Strickland and Katrina Maceda from the publisher was extremely encouraging. I have to thank all the contributors – the old ones and those who joined us for the second edition – for their support. Both the publisher and the editor tried everything to avoid a delay of the publishing process. Therefore, some small chapters of the previous edition had to be reprinted. Last but not the least, I have to thank my co-­workers at the Institute of Forensic Medicine at the University of Bonn, especially my secretary Stephanie Raabe, for their continuous support.

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Six years after the first edition, the preparation of a second edition was necessary. The book is now published as a three-­volume-­ set. Since the first edition, many scientific improvements have been made which had to be incorporated (for instance, new psychoactive substances (NPS), molecular photofitting and the determination of the geographic origin, progress in molecular pathology for the determination of the cause and manner of death, new vitality markers as heat shock proteins (HSPs) or aquaporins). The world was struck by terroristic attacks, for instance, in Paris, Nice and Berlin and other fatalities (Grenfell fire disaster), and of course recommendations on how to handle these cases are necessary. The importance of postmortem imaging for a fast identification of deceased was shown impressively. The beginning and the whole year 2020 was governed by the Corona pandemic with a lockdown in many countries for many weeks. This pandemic is also of significant importance in the field of forensic medicine: • Was the mortality rate increased? • What are the reasons for different mortality rates in different countries? • What are the causes of death in COVID-­ 19-­ positive patients?

Burkhard Madea Bonn, Autumn 2021

Preface to the First Edition

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evidence. One challenge of forensic medicine for the future will always be to keep up to date with other scientific improvements and to modernise itself. On the organisational level, in a number of countries, improvements of forensic medical services can be observed, in others unfortunately a decline, especially in countries where forensic medical services are mainly university-­based. In a time of an oversized competition about money, other medical disciplines are much more successful than the area of forensic medicine. The main tasks in forensic medicine are forensic investigation, reconstruction and expert witness reports. These missions are of course regulated according to national legislations, differing from country to country. Therefore, there are obvious geographical constraints in writing and editing a handbook on forensic medicine. On the other hand, not only countries in one continent but in all continents are growing together economically, and in the common future, a harmonisation of different scientific and legal systems is inevitable. Therefore, exchange of medico-­ information on an international level can do nothing, but good and legal medicine is no exception. Although medico-­ legal systems differ from country to country, in some countries a harmonisation of postgraduate education and also of daily routine procedures (standard operation procedures, for example, medico-­legal autopsy rules) can be observed. Therefore, quality aspects and quality control will be also addressed in this book. For example, in Europe, many institutes of forensic medicine are already accredited according to ISO 17025 for forensic purposes. International guidelines and international accepted quality standards will be of increasing importance in the future in a fast-­ growing together world. Authors from many countries contributed to this book. I am very much indebted for their input to this book and of course to the publisher for making the implementation of this book possible.

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The basic mission of forensic medicine has to be the proper application of justice. Not only justice itself but especially societies benefit from well-­functioning medico-­legal services concerning the: • detection and solving of crimes, • appearance of qualified expert witnesses at court, • the prevention of crimes (e.g., child abuse, torture, etc.) and also • the prevention of accidents (such as alcohol-­related accidents, drug-­related accidents, improved car engineering, etc.). Forensic medicine is an ancient medical discipline working at the borders or better cross sections of medicine and law. This is a book on forensic medicine and not only forensic pathology. It comprises all current aspects of forensic medicine commencing with the medical aspects of death, the certification of cause and manner of death, autopsy legislation, etc. The following chapters (as, for example, traumatology, sudden and unexpected deaths, clinical forensic medicine, etc.) deal with external violence, the physical signs of abuse and the differential diagnosis of natural versus unnatural causes of death. Of special importance is a chapter focusing on toxicology, since in the field of forensic medicine, further matrices are now available for toxicological analyses compared to clinical toxicology. Further questions dealt with are the tasks of general medico-­legal practice such as traffic medicine, identification, haemogenetics, medical law and forensic psychiatry. Knowledge of all sub-­ disciplines of forensic medicine is required in daily routine casework, since all results being obtained by different medical and scientific methods have to be integrated into one comprehensive reconstruction of events in the case in question. During the recent years, there have been a number of dramatic developments in forensic medicine – both on a technical and an organisational level. Great improvements have been made especially in the fields of forensic genetics and toxicology and also a new area – postmortem imaging – which was developed within a few years’ time. This implementation of new developments into daily routine casework improves considerably the overall validity of expert

Burkhard Madea Bonn, 2013

Foreword by Denis A. Cusack

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frameworks. These frameworks may have some significant ­differences as for example between the Civil Law System in Continental Europe and South America; the Common Law System in the United Kingdom, Ireland, Australia, New Zealand, Canada and parts of South East Asia; other mixed and singular Legal Systems in the United States, the Arab World, on the continent of Africa and in the Far East. But in Forensic Medicine there is a unity of purpose for the medical practitioner. It is a medical specialty in dynamic action for all citizens through combining and integrating professional standards in the specialty in accordance with best international shared practice and protocols without fear or favour and free of political influence or duress. It is through a harmonisation of forensic practice and understanding that this ancient yet modern branch of medicine can serve the objective demands of today’s global society in achieving both health and justice. It is in this context that this textbook edited by Professor Burkhard Madea constitutes an outstanding contribution to the modernised knowledge base and practice of Forensic & Legal Medicine in the 21st century. It comprises an impressive 68 chapters in 11 parts over 3 Volumes. It deals with all the core aspects of forensic medicine and includes a great array of areas from the duties of the forensic practitioner expanding through major areas of practice such as clinical diagnostics; post mortem issues at the gross anatomical level down to the molecular level in genetics; injuries; humanitarian medicine; medico-legal practice; death certification; psychiatry; toxi­ cology; criminology; road traffic medicine; and insurance medicine. The breadth and depth of the book and its comprehensive nature make it an extraordinarily valuable contribution which is all the more of great merit through the contributions of such an expanse of expert colleagues in Forensic and Legal Medicine. The textbook is a scholarly review of forensic skills and practice within our diverse legal systems with applicability to the necessary harmonisation of investigative procedures, standards and forensic accreditation. It is on such knowledge that we will base our training, peer review and quality assurance procedures which will be tested not least in the cauldron of court examination of evidence presented by forensic medical professional expert witnesses in the pursuit of societal justice. It is my honour to write this foreword to the 2nd Edition of this seminal textbook and to commend it to forensic specialists,

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The history of Forensic and Legal Medicine is an ancient one and well known to practitioners and students of the specialty. It traces its beginnings back to both ancient and more recent civilizations in Egypt, India, Babylon, Greece, Rome and China through such familiar names and texts as Imhotep, the Manusmitri, Hammurabi, Hsi Duan Yu, Hippocrates, Justinian and Charles V. In Europe the founding of Chairs in Legal Medicine in the late 19th century marked the start of the modern era of professional practice and education in the field. More recently a number of international organisations such as the International Academy of Legal Medicine; the European Council of Legal Medicine; the Academia Iberoamericana de Criminalística y Estudios Forenses; the Arab Union of Forensics and Toxicology; the African Society of Forensic Medicine; the Indo-Pacific Association of Law, Medicine and Science; and the International Association of Forensic Sciences have seen the development of a harmonised approach to practice and learning in the forensic specialty. It remains as one of the oldest medical specialties still practised in the 21st century. Its basic purpose is still close to its founding roots, to deliver a combination of health and ­justice to citizens by the application of medical principles to the justice system. It is both diverse in the areas it encompasses yet unified by its international applicability. It combines skills from clinical medicine, pathology and legal and ethical practices. Traditionally the four main subheadings of the specialty have been clinical forensic medicine; forensic pathology and thanatology; toxicology; and medical law and ethics including humanitarian forensic medicine and science. The modern concept of Translational Forensic Medicine may be defined as an interdisciplinary branch of the biomedical field supported by three main applicable pillars: scene, benchside and society. Forensic and Legal Medicine has retained a core and strategic place in both medical undergraduate and postgraduate education even though there has been a tendency to diminish the standing and place of the specialty in the core undergraduate medical curriculum in recent years. Indeed, this is one of the reasons this textbook is a timely reminder of the place of forensic medicine in medical training and practice for the greater good of society. Forensic & Legal Medicine is a discipline which requires specific skills to be applied in accordance with the various legal

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Foreword by Denis A. Cusack

Denis A. Cusack President of the European Council of Legal Medicine. Vice-President of the International Academy of Legal Medicine. Member of the Scientific Advisory Board of the Office of the Prosecutor of the International Criminal Court. Full Professor Emeritus of Forensic & Legal Medicine, University College Dublin, Ireland. Senior Coroner for the District of Kildare, Ireland.

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scholars and students and to all doctors whose professional practice involves forensic and medico-legal elements. I echo the eloquent words of Professor Duarte Nuno Vieira written in his Preface to the 1st Edition. I congratulate the Editor and Authors of this learned book which will be a substantial companion to the forensic community and full of practical and expert knowledge for the practitioners and students of Forensic & Legal Medicine for many years to come.

Foreword by Duarte Nuno Vieira

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the rhythm of life and are in tune with judicial requirements. Without good legal medicine there can never be good justice. This is the only field of knowledge that can truly shed light on situations as diverse and as complex as sexual abuse, physical and psychological trauma, the influence of toxic substances, identity and kinship, death by murder, suicide and accident, etc. The extraordinary progress which has been made over the last few years in many areas of legal medicine has yielded even more significant results, enabling the judicial system to become faster and more efficient, and allowing it to issue decisions that are more appropriate and scientifically grounded. As a science in constant evolution, legal medicine demands permanent contact with the latest legal, scientific, technological and judicial developments, as well as promoting discussion amongst its practitioners in order to solve common problems, share experiences and knowledge, harmonise methodology and compare results. Although legal medicine has achieved a great deal over the last few decades, there is still a long way to go. The road that lies ahead will never come to an end, because it will require constant adjustments and improvements, changing yet remaining always the same. Those who travel it will have to follow its course. Books are essential for this – they are the handrail on the flight of stairs that is the ongoing professional training that each of us undergoes throughout life – offering a means for the exchange of ideas, for the acquisition and consolidation of knowledge and skills, for making contact with different perspectives and recent doctrinal, technological and scientific developments and for understanding both the potential and the limitations of specialist activity, as well as the new challenges that are constantly appearing. This book, edited by Professor Burkhard Madea, offers an up-­ to-­date view of the various facets of legal medical intervention, particularly those specialist areas that have undergone the most pronounced development (such as personal injury appraisals under civil and labour laws, which still arouse considerable disagreement and controversy regarding doctrine and methodology). The fruit of incessant study, an exhaustive bibliographical review and, particularly, the practical experiences of a host of legal medical teachers and professionals (most of them household names in their respective fields), it is a repository of valuable knowledge and guidance, transmitting different interpretations and points of view which stimulate reflection. Hence, it is an essential companion for qualified specialist intervention, which always requires the careful weighing up of different interpretive possibilities and

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Born from the need for justice, legal medicine has had to adapt to a succession of scientific, technological, sociocultural and legal contexts throughout history. As a branch of science, it has been in constant evolution and, as such, has proved a source of inspiration and change for the law, stimulating improvements, amendments and legislative innovations. Specialist forensic work involves delving into mysteries of the unknown, solving the enigmas of existence and seeking answers for the large (and small) questions of life. It is also about adding to reality, about reinventing the world that we have inherited. Legal medicine has a mission that is far from easy, a mission that has always been (and always will be) fundamental to the correct application of justice. On it depend the honour and freedom of people. Legal medicine brings us into contact with people who carry with them the traumas of a life struck by misfortune, people that sometimes have the smell of death imbued in their skin. Hearing the voices of the victims and their families, their silent screams of anguish and despair, pain and anger, we have to make a daily effort to help them and to not give up on the world. Legal medicine is a field of study riddled with doubts, uncertainties, distress and nightmares. However, associated with this, there are also moments of great fulfilment, of intense professional and personal realisation  – as this is an activity that indisputably helps others. All who practise it have certainly had experiences with people and situations that have profoundly enriched their lives from a human and a spiritual point of view. Although there are differences between countries regarding the content and attributions of legal and forensic medicine, it is defined by the European Council of Legal Medicine as ‘the application of medical knowledge and methodology for the resolution of legal questions and problems for individuals and society. It involves the observation, documentation, collection, assessment and scientific interpretation of medical evidence deriving from clinical and postmortem investigations required for the different fields of law, including criminal, civil, work, family and administrative. Its core activities are clinical forensic medicine and forensic pathology, but other areas of science and expertise including forensic toxicology, forensic psychiatry, forensic genetics and forensic anthropology may be required depending on the nature of the case.’ As legal medicine is fundamental for the correct application of justice, it can only achieve its highest level with the collaboration of highly qualified legal medical departments that pulsate with

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FOREWORD BY DUARTE NUNO VIEIRA

stimulating food for thought, as well as guidance for professional practice. I congratulate the editor and the different authors on this contribution to legal medicine. It is indeed a great honour to have been invited to write this foreword.

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Duarte Nuno Vieira President of the European Council of Legal Medicine. Full Professor of Forensic Medicine and Forensic Sciences, University of Coimbra. President of the Portuguese National Institute of Legal Medicine and Forensic Sciences. Past President of the International Academy of Legal Medicine, of the International Association of Forensic Sciences, of the Mediterranean Academy of Forensic Sciences and of the World Police Medical Officers

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perspectives. This book will help legal medicine become more efficient and of better quality, bringing benefits to the field. Written in clear and simple language, yet rigorous and scientifically grounded, this is the work of competent and committed academics and professionals – people who are passionate about legal medical work, about teaching and the sharing and transmission of experiences and knowledge, who are deeply concerned with justice and the training of its practitioners. This book reveals the human, academic and professional merits of its editor, and attends to the differences in opinion that result from the wealth of human diversity and from the different perspectives that science can offer us. It is, in short, a book that deserves our attention. It is a pleasure for me to be able to recommend this book, not only to students and practitioners of legal medicine, but to anyone who, directly or indirectly, has an interest in legal medicine issues. I am sure it will give readers great satisfaction and provide

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PART I

Duties of Forensic Medicine in Modern Societies

1 | History 2 | Duties of Forensic Medicine and the Forensic Medicine Practitioner 3 | Forensic Medicine and Human Rights 4 | International Guidelines and Accreditation in Forensic Medicine Handbook of Forensic Medicine, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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1

History

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Burkhard Madea

Characteristic topics that are dealt within forensic medicine are summarised in Box 1.2. Forensic medicine as we experience it at the beginning of the 21st century has developed since the 19th century and from much older roots. The famous criminal code of Emperor Charles V, the Constitutio Criminalis Carolina, promulgated in 1532, is often called a landmark of the first importance in the history of legal medicine (Figure 1.1). R. P. Brittain (1965a) wrote:

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According to Sydney Smith (1951), forensic medicine may be defined briefly as consisting essentially of that body of medical and paramedical scientific knowledge which may be used for the purposes of administration of the law. Alfred Swaine Taylor has defined medical jurisprudence as ‘that science which teaches the application of every branch of medical knowledge to the purpose of the law’. According to a German definition by Schmidtmann (1905), the last editor of the famous Handbook of Forensic Medicine of Johann Ludwig Casper (1857), forensic medicine is a cross-­sectional discipline of medicine and natural sciences dealing with all medical evidence that is relevant for law. It deals with medical evidence not only in practice but also in research; furthermore, all legal essentials in health care are especially important for doctors as part of teaching, training and research. Apart from forensic pathology being the essential branch in the development of forensic medicine, in the last 20 years, clinical forensic medicine has developed as its own field of expertise. Clinical forensic medicine is that discipline of medicine which involves an interaction between law, judiciary and police dealing generally with living persons. There is of course no special date at which forensic medicine emerged as a recognisable separate scientific discipline. Several steps in the development of forensic medicine can be distinguished (Box 1.1): firstly the use of medical knowledge for legal and public purposes, secondly the compulsory medical testimony for the guidance of judges in special cases and thirdly the professionalisation as an own discipline.

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1.1 Definitions

Handbook of Forensic Medicine, Volume 1, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

‘It has commonly been considered as the true start of legal medicine, and hence Germany has been hailed as the country which gave birth to the discipline. It has been said that it caused medical men to be called in legal matters for the first time. This is not strictly true. They had been called on before as earlier enactments show. Without in any way minimising the advance the Carolina represented, it would be wrong to consider it as a phenomenon which occurred without logical antecedents, and as implying the legal medicine arose by a kind of spontaneous generation’. Indeed, the Bamberg Criminal Code was a model for the Constitutio Criminalis Carolina. However, there is a deep-­seated relationship between medicine and law dating back much earlier. These roots can be found in studies of nature, violation of law and its relation to medicine (injuries, violent death, pregnancy, stillbirth, rape, poisoning, etc.) and the need for experts to assist the law or a court – thus, defining the constitution of forensic medicine as a scientific discipline with the publication of monographs, subjects of special instructions and its own research (Table 1.1).

4

PART I  

DUTIES OF FORENSIC MEDICINE IN MODERN SOCIETIES

Box 1.1  Development of forensic medicine.

Box 1.2  History of characteristic topics in forensic medicine (according to Fischer-­Homberger, 1983).

Step 1 Medical knowledge is used for legal or public purposes • dependent on point of achievement both in law and medicine ○ Knowledge of medical plants, botany ○ Knowledge of injuries ○ Educational standards in medicine ○ Standards of competency ○ Legislation concerning disposal of the dead ○ Legislation concerning injuries ○ Compensation for injuries and deaths

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Step 2 Expert medical testimony must be obtained for the guidance of judges in cases of murder, wounding, poisoning, hanging, drowning, infanticide, abortion and malpractice

Responsibility • Age • Gender • Mental diseases, melancholia • Simulation • Disease or malice Sexuality and reproduction • Marriage, family • Impotence, infertility • Virginity • Conception and pregnancy • Duration of pregnancy • Superfoetation • Abortion • Infanticide ○ ‘Hydrostatic tests’ of lungs • Rape Injuries and violent deaths • Injuries ○ Prognosis of injuries and their locations ○ Lethality of wounds, grades of lethality ○ Relative fatality of wounds in different parts of the body • Suffocation • Poisoning Role of medicine for the public • Specialised medical profession • Educational standards • Standards of competency • Ethical standards • Malpractice

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Step 3 Further professionalisation: • Medicolegal examination • Giving evidence at court/medical expertise required at court • Publication of monographs • Teaching • Systematic research (decrease of the domain of magic and sorcery) • Knowledge gained by own practice replaces textbook knowledge (J. L. Casper) • Foundation of professorships • Foundation of own institutes • Foundation of societies

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The development and existence of a speciality of forensic medicine depend essentially on two factors: a sufficiently high development of the law and a sufficiently high development of medicine. As Ackerknecht has outlined, in very highly developed civilisations with sophisticated legal regulations, there is, on the one hand, no evidence that judges consult medical persons in assessing crime. On the other hand, despite high development of a rational medical art, no document exists that provides evidence for the use of medical experts in ancient Greece.

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1.2  Civilisations of the Near East and China Forensic medicine developed in relation to law, and it was often legal requirements which pushed forward improvements in forensic medicine. Forensic medicine as a scientific discipline developed when the domain of magic and sorcery was overcome. The early literate civilisations of the Near East and China had definite systems of law relating not only to crime but also to property, marriage and other civil matters. For instance, in Egypt, practice of medicine was subject to legal restrictions; the right to practice was restricted to members of a certain class with the intention that physicians had to study the precepts laid down by their predeces-

Source: Fischer-­Homberger (1983).

sors in certain ancient books (Smith  1951). Since physicians strictly had to adhere to the knowledge of ancient books, experiments and originality were not encouraged and, instead, witchcraft, magic and sorcery became dominant. As a result, good treatment was characterised by observing the authoritative ‘canon’, with the outcome that bad treatment or even malpractice originated from not properly observing the authoritative ‘canon’. In China at the beginning of the 14th century, a noteworthy volume entitled Hsi Yüan Lu (The Washing Away of Wrongs) was compiled on the procedure to be followed in investigating deaths, especially those under suspicious or obviously criminal circumstances. Sydney Smith, who has studied a comparatively modern edition of this book, describes his impression as follows: ‘I have not seen a translation of a really ancient copy of this book, but even from a comparatively modern edition (1843) one certainly gets the impression that there was a comprehensiveness in the scope of medicolegal procedure in ancient China that was not to be found in mediaeval Europe. The importance of a satisfactory examination of the wounds on a body is stressed, among other reasons, in order to check the validity of a confession or other statement. The sites where wounding is

5

HISTORY

likely to prove mortal are indicated. The preparations necessary for the examination of a body are described, and the examiner is warned not to be deterred by the unpleasant state of the corpse, but to make a systematic examination from the head downwards in every case. The difficulties caused by decomposition are clearly recognized, and the examiner is advised on the subject of counterfeited wounds. Sections are devoted to wounds caused by different agencies, such as blows from the fist or kicking, by various types of weapon, etc.; and to asphyxial deaths – f.i. by strangulation and drowning. The possibility of homicidal strangulation being passed off as suicide is discussed, also the means for distinguishing between the bodies of drowned persons and those thrown in after death. The possible confusion between ante-­mortem and post-­mortem burning is recognized, and poisoning is given considerable attention. The examiner is advised on the possible importance of examining the locus, and is warned that at an inquest nothing should be regarded as unimportant. . . . Altogether it is a remarkable document, and, although some of the methods and tests described are fantastic, there is no doubt that the real nature of the problems involved was clearly appreciated. As I have suggested, it is unfortunate that I cannot with certainty sort out the genuinely ancient from the more modern interpolations, but I am left with the conviction that in mediaeval times Chinese forensic medicine was far in advance of contemporary European practice’.

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(S. Smith 1951)

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Figure 1.1  Constitutio Criminalis Carolina (criminal code of Emperor Charles V). Source: Image from http://commons.wikimedia.org/wiki/ File:De_Constitutio_criminalis_Carolina_(1577)_13.jpg (image in public domain).

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CHAPTER 1  

220 BC

China

2200 BC

Babylon

10 BC

India Egypt

Bamboo with information on the rules and regulations for the examination of injuries Codex Hammurabi: rights and duties of physicians including medical malpractice

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Writings about pharmacology and pharmacognosy

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Table 1.1  Timetable of the history of forensic medicine (according to Payne-­James, Bonte and Smith).

449 BC

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Law of Manu: competence of witnesses at court Detailed laws concerning the medical profession, forensic medical investigation, stab wounds were categorised, closed head injuries with head fractures were described Official code for medical fees, penalties for medical malpractice, classification of injuries, abortion classified as crime Autopsies on human bodies were not permitted, use of physicians as expert witnesses was loose and ill-­defined

Rome

Lex duodecimo tabularum length of gestation (for the determination of legitimacy), disposal of the dead, poisoning, punishment depending on the degree of injury

460–355 BC

Hippocrates of Kos

Lethality of wounds, Hippocratic Oath as basis of medical ethics

384–322 BC

Aristotéle ¯s

Soul enters the body 40 days after conception

Ancient Rome

Numa Pompilius

Advocated cutting open the bodies of pregnant women after death to deliver the baby (caesarean section)

572 BC

Lex Aquilia

Lethality of wounds, ‘novus actus intervenium’, break in causation

138–78 BC

Lex Cornelia (Sulla)

If a physician had caused the death of his patient, he should be exiled or executed, prostitution and confinements supervised (Continued)

6

PART I  

DUTIES OF FORENSIC MEDICINE IN MODERN SOCIETIES

Table 1.1  (Continued) Gaius Julius Caesar

His body was examined after murder by the physician Antistius who discovered 23 stab wounds and declared only one to be fatal

23–79 BC

Plinius the Elder

Complaints that laws punishing incompetent or ignorant physicians were missing, drowning

131–200

Galen

Dealt with gladiators and their wounds, anatomic features, differences between neonates and adults lungs, stillbirths: Rubra, gravis, densa substantia carnis pulmonum, Alba, Levis, rara substantia carnis pulmonum in live births

483–565

Justinian

Digest ‘physicians are not ordinary witnesses, but give judgement rather testimony’ (Medici non sunt proprie testes, sed majus est judicium quam testimonium), proofs of pregnancy, sterility, hermaphrodism, simulation of diseases

5th–10th century

Leges Barbarorum Lex Euricianus Lex Visigothorum Lex Burgundionum Pactus Legis Salicae Pactus Legis Riduarie Edictus rothari Pactus Legis Alamannorum

Goths, Visigoths, Vandals, when medical experts have to be called, ‘Wergeld’ (blood money) had to be paid to the victim by the perpetrator or to relatives of the decedents, description of wounds

742–814

Charlemagne

Capitularies: expert medical testimony required in cases of wounding, abortion, rape, incest, infanticide and suicide

1140

Roger II of Sicily

Physicians required to have an examination prior to commencing their practice

Frederic II

Public examination of physicians, strict criteria for medical practice, versed in the teaching of Hippocrates and Galen, requirements for ordination as a doctor, at least 21 years of age, be legitimately born, have studied logic for three years, have studied medicine for five years, served a year of apprenticeship, on qualification had to swear an oath, had to treat the poor free, visit each of his patients twice by day and once by night as required A human body should be dissected in public once every five years

1100

Assizes of Jerusalem, Godfrey de Bouillon

In cases of alleged illness or murder, three experts were sent

1209

Pope Innocent III

1234

Pope Gregory IX

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100–44 BC

Appointment of physicians to the courts Compilatio decretalium: collection of all decisions and edicts related to medicolegal matters Bamberg Criminal Code, medical evidence necessary in certain cases

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1575

Ambroise Paré (1510-­1590)

1597

Giovanni Battista, Codronchi (1547–1628)

Methodus Testificandi

Fortunati Fidelis (1550-­1630)

De Relationibus Medicorum

Paolo Zacchia (1584–1659)

Quaestiones Medico-­Legales

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Constitutio Criminalis Carolina, legal medicine originated as an own speciality, requirement of a medical opinion in cases of murder, wounding, poisoning, hanging, drowning, infanticide and abortion

Book on medicolegal reports, death from lightning, antemortem vs. postmortem injuries, poisoning by carbon monoxide

Source: Adapted from Payne-­James (2005); Bonte (2000); Smith (1951).

1.3  Justinian enactments The Justinian enactments between AD 529 and 564 represent, according to Sydney Smith, the highest point of achievement in forensic medicine in the ancient world. Amongst many other

things, the Justinian Code provided guidelines for the regulation of the practice of medicine, surgery and midwifery; for the proof of competence by means of examinations; for the classes of ­physicians that were to be recognised; for the limitation of the number of physicians in each town and for the penalties to

CHAPTER 1  

Box 1.3  Dates of foundation of some European universities.

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be imposed for malpractice. The Justinian laws clearly recognised and defined an integrated medical profession, with required educational standards and standards of competency, in a way that had never previously been achieved. The medical expert, defined as ‘medici non sunt proprie testes, sed majus est judicium quam testimonium’, was not an ordinary witness, appearing for one side or for the other side, but assisted the judiciary by impartial interpretation. The Justinian Code enjoined the cooperation of medical experts in a broad field of legal problems, for instance, in the determination of the existence of pregnancy, in cases involving sterility, impotence or legitimacy, in cases of rape, in cases of poisoning, in cases involving the problem of survivorship, in cases which were complicated by the question of mental disease and in other comparable circumstances (Smith 1951).

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HISTORY

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In Bologna, according to the town charter, a medical expert must be at least 40 years of age and to have been a citizen of Bologna for at least 10 years. The first documented legal autopsy report of Bologna was signed by Bartolomeo da Varignana in 1302. The right of performing an autopsy was given to medical faculties, such as the Faculty of Medicine of the University of Montpellier in 1374. As in Italy, forensic medicine in France and Germany developed with the foundation of universities and medical faculties. Medical faculties even discussed and criticised court decisions. In 1478, the University of Cologne gave the following advice: ‘It is useful and necessary that those who die unexpectedly – god forbid this but unfortunately it happens so often – are opened and dissected immediately in order to examine the organs and find the cause of death or the lethal disease’. (Ackerknecht  1950/51). The physicians knew the limitations of only examining a body, they were aware of their responsibility. In the 17th century, it was recognised that autopsies were necessary to definitely clarify the cause of death, even if no signs of external violence were visible. Gottfried Welsch (1618–1690) in the first edition of his book Rationale Vulnerum Lethalium Judicium (1660) was already recommending forensic autopsies, especially in cases of intoxications, and that autopsies should be performed by doctors with experience in postmortem dissection.

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Already in the 12th century, physicians were being used as experts in cases of alleged illness or injuries. In the so-­called Assizes of Jerusalem from 1100, it was determined that if, because of alleged illness, a vassal could not appear before the lord’s court to plead his case, the lord must send to this man’s house three office men to decide on the issue – a physician, an apothecary and a surgeon. In cases of murder, these three experts were also sent, and they had to say what was the matter with him (the body) and where he has been injured and with what instrument it seems to them that the injuries had been inflicted. Similar regulations existed at the same time in Antioch: knights could only excuse their non-­ attendance before the court when medical experts confirmed the alleged illness (Bonte 2000). The Italian town charters played an important role from the 11th to the 13th century. The town charters were qualified with the help of the law faculties in the newly established universities (Box 1.3). For instance, in the town of Bologna, Hugo de Lucca was appointed expert of the magistrate of the city. It is likely that he was the first to perform legal autopsies between 1266 and 1275. Most of the Italian town charters determined that two experts, generally a physician and a surgeon, were responsible for postmortem examinations. An example for such an early report by medical experts is:

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in the occiput: one deadly wound in the upper jaw: one non-­fatal wound Sworn to be true on Saturday, February 12th (Ackerknecht 1950/51)

Bologna 1289 Master Albertus Maledova and Master Amoretus, physicians, who, on the injunction of Albertus of Gandino, judge, have seen and examined Jacobus Rustighelli in the Church of St. Catherine of Saracocia, wounded and dead, state in concordance, after having seen and examined, to have found the following: in the thorax: seven deadly wounds in the neck: one deadly wound in the middle of the forehead: two deadly wounds

1.5  Forensic medicine as a book science The period from the late 16th to the 18th century was characterised by books published on forensic medicine (Box 1.4). In 1575, Ambroise Paré published a book that dealt with medicolegal reports in ‘death from wounds or impotence or loss of any member’. He discussed abortion, infanticide, death by lightning,

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Box 1.4  Important books in the history of forensic medicine (17th–18th century).

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Alberti, Michael: Systema jurisprudentiae medicae . . . cum praefatione. christiani thomasii, Halae: Orphanotropheu 1725 (I. Band); Fulda: Tomus alter Schneebergae, 1729. Ammannus, Paulus: Medicina critica. Erfurti: Ohler; Hertz (Drucker) 1670. – Praxis vulnerum lethalium. Francofurti: Gleditsch 1690. Bohn, Johannes: De renunciatione vulnerum, seu vulnerum lethalium examen. Lipsiae: Gleditsch; Fleischer (Drucker) 1689. 2. Aufl. Amsterdam 1710. – De officio medici duplici, clinici nimirum ac forensis. Lipsiae: Gleditsch 1704. Cardanus, Hieronymus: De venensis libri tres. In: Opera omnia, 10 t. Lugduni: Huguetan & Ravaud 1663, t. 7, S. 275–355. Castro, Rodericus A.: Medicus-­politicus. Sive de officiis medico-­politicis tractatus. Hamburgi: Frobenius 1614. Codronchius, Baptista: De morbis veneficis ac veneficijs. Venetiis: De Franciscis 1595. – Methodus testificandi. In: De vitiis vocis, libri duo. Francofurti: Wechel 1597, S. 148–232. Fahner, Johann Christoph: Vollständiges System der gerichtlichen Arzneikunde. Ein Handbuch für Richter und gerichtliche Ärzte. 2 Bde., Stendal: Franzen und Grosse 1795; 1797. Fidelis, Fortunatus: De relationibus medicorum libri quatuor, In quibus ea omnia, quae in forensibus, ac publicis causis, medici referre solent, plenissime traduntur. Hrsg. v. Paul Amman, Lipsiae: Tarnov 1674. (1. Ausg. Palermo 1602). Frank, Johann Peter: System einer vollständigen medicinischen Polizey. 4 Bde.; Bd. I in 2. Aufl., Mannheim: Schwan 1784, Bde. 2–4 in 1. Aufl., Mannheim: Schwan 1780–1788; Bd. 5 Tübingen: Cotta 1813 (1. Auflage des I. Bandes 1779; ein Band 6 in 3 Abteilungen folgte Wien 1817–1819; Die Reihe der Supplementbände war erst 1827 abgeschlossen. Haller, Albrecht von: Vorlesungen über die gerichtliche Arzneiwissenschaft. Aus einer nachgelassenen lateinischen Handschrift übersetzt. 2 Bde., Bern: Neue typographische Gesellschaft 1782; 1784. (2. Band in 2 Teilen) Mende, Ludwig Julius Caspar: Kurze Geschichte der gerichtlichen Medizin. In: Ausführliches Handbuch der gerichtlichen Medizin; 1. Teil, Leipzig: Dyk 1819, S. 1–474. Metzger, Johann Daniel: Kurzgefasstes System der gerichtlichen Arzneiwissenschaft. Königsberg/Leipzig: Hartung 1793. Platner, Ernst: Untersuchungen über einige Hauptcapitel der gerichtlichen Arzei-­Wissenschaft durch beigefügt zahlreiche Gutachten der Leipziger medicinischen facultät erläutert. Aus dem Lateinischen übers. u. hrsg. v. C.E. Hedrich. Leipzig: Kupper 1820. Plenk, Josephus Jacobus: Anfangsgründe der gerichtlichen Arztneywissenschaft und Wundarztneykunst. Aus dem Lat. übers. v. F. August von Wasserberg. Wien: Gräffer 1782. Ploucquet, Wilhelm Gottfried: Abhandlung über die gewaltsame Todesarten, nebst einem Anhang von dem geflissentlichen Missgebähren. Als ein Beytrag zu der medicinischen Rechtsgelahrtheit. Tübingen: Berger o.J. Pyl, Johann Theodor (Hrsg.): Aufsätze und Beobachtungen aus der gerichtlichen Arzeneywissenschaft. Sammlungen 1–8, Berlin: Mylius 1783–1793. Suevus, Bernhardus: Tractaus de inspectione vulnerum lethalium et sanabilium praecipuarum partium corporis humani. Variis cum veterum, tum recentium medicorum observationibus, exemplis atque controversiis illustratus, non minus iurisconsultis quam medicis utilis atque necessarius. Marpurgi: Chemlin 1629. Teichmeyerus, Hermannus Fridericus: Institutiones medicinae legalis vel forensis. Ienae: Bielke 1723 (2. Auflage ebenda 1731). Welsch, Gottfried: Rationale vulnerum lethalium judiciium, in quo de vulnerum lethalium n atura, et causis; legitima item eorundem inspectione, ac aliis circa hanc materiam scitu dignis juxta, quam necessariis, agitur. Lipsiae: Ritzsche 1660. Zacchias, Paulus: Quaestiones medico-­legales. In quibus eae materiae medicae, quae ad legales facultates videntur pertinere, proponuntur, pertractantur, resolvuntur. Opus, iurispertis apprime necessarium, medicis perutile, caeteris non injucundum. Et. tertia, Amstelaedami: Blaeu, 1651. (1. Ausg. Rom Bd. 1: 1621; Bd. 2: 1625; Bd. 3-­4: 1628; Bd. 5: 1630; Bd. 6: 1634; Bd. 7: 1635.

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­ anging, drowning, feigned diseases and the differentiation of h antemortem and postmortem wounds  – all topics that still belong today to the field of forensic medicine. He also dealt with poisoning by carbon monoxide. An example of a report of Ambroise Paré on an abdominal wound resulting in abortion is as follows: I, Ambroise Paré, have come on the order of the great Provost to the Rue St. Houbré, to the house of Mr. M., where I have found a lady called Margaret in bed with a high fever, convulsions, and haemorrhage from her natural parts, as consequence of a wound that she has received in the lower abdomen situated three fingers below the umbilicus, in the right part, which has penetrated into the cavity, wounded and penetrated the uterus. She has therefore delivered before term a male infant, dead, well formed in all its limbs, which infant

has also received a wound in its head, penetrating into the substance of the brain. Therefore the above-­ mentioned lady will soon die. Certified this to be true in putting my signature, etc. In 1597, Codronchius, a physician of Imola, published his important Methodus Testificandi, in which he dealt with wounds, poisoning and sexual matters and gave models of reports. Another opus magnum was the work by Fortunatus Fidelis of Palermo entitled De Relationibus Medicorum, which was the first great general systematic treatise on legal medicine. In this, he deals firstly with matters of public health, secondly with wounds, simulated diseases and medical errors, thirdly with virginity, impotence, pregnancy and viability of the foetus, and finally with life and death, mortality of wounds, suffocation and death by ­lightning and poisoning.

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The greatest work in this early period was the Quaestiones Medico-­Legales of Paolo Zacchia, the principal physician to Pope Innocent X and Pope Alexander VII and an expert before the Rota Romana, the papal court of appeal (Figure  1.2). He published his monumental work between 1621 and 1635  in Rome (Figure 1.3), with two additional books published at Amsterdam in 1666. The books are divided into parts and these delve into specific questions dealing with: age, pregnancy, superfetation and moles, death during delivery, live birth and legitimacy, similarity and dissimilarity of children to their parents, dementia and insanity, poison and poisoning, impotence, feigned diseases, the plague and contagion, miracles, virginity and rape, fasting, wounds, mutilation and the salubrity of the air, water and places. Thus, by the middle of the 17th century, there was a well-­developed literature on this subject and the subject itself was recognised as an entity. In the 17th and early 18th centuries, it was mainly professors at universities in Central Germany (at Leipzig and Halle) who contributed to the development of forensic medicine. In 1660, Welsch (at Leipzig) wrote a book on wounds, dealing with their vitality. Ten years later, Ammann (also from Leipzig) produced a manuscript dealing with false believes in forensic medicine and in 1690 a more important contribution, his treatise on fatal wounding. Johannes Bohn of Leipzig in 1689 published his work De Renunciatione Vulnerum Lethalium Examen, which was of great importance at his time, and distinguished antemortem and postmortem wounds and wounds deadly per se (per se seu absolute lethalia) from accidental factors (ca accidenc lethalia). He was

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Figure 1.2  Paolo Zacchia (1584–1659), called the ‘father of forensic medicine’. Source: Image from the U.S. National Library of Medicine, History of Medicine Division.

Figure 1.3  Title cover of Paolo Zacchia’s book ‘Quaestiones Medico-­Legales’.

in favour of complete medicolegal autopsies instead of wound inspection and described already procedures to be followed. According to Bohn, in Germany, there was during the 18th century an almost uninterrupted production of treatises on legal medicine. However, in the 18th century, forensic medicine was a ‘book science’. It was the 19th century during which this book knowledge was replaced by personal experiences. Johann Ludwig Casper (1796– 1864) worked for nearly 40 years in the area of forensic medicine in Berlin. He transformed forensic medicine into a useful speciality based upon practical experiences and personal observations. His classic textbook Practisches Handbuch der Gerichtlichen Medicin (1856) was based on his extensive practical experience. Centres of legal medicine as it became a modern science in the 19th century were located in Berlin, Vienna, Glasgow, Edinburgh and London.

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Table 1.2  Historical development of forensic medicine in Germany.

Box 1.5  Types of autopsies.

1532

Constitutio Criminalis Carolina; expert evidence and expert witness mentioned

1835

Association of Badische Medical Officers for the Advancement of Staatsarzneikunde

1840

Association for Staatsarzneikunde in the Kingdom of Saxony

1868

Foundation of the section ‘Official Health Service and Staatsarzneikunde’ of the Society of German Natural Scientists and Doctors (Dresden)

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Foundation of an independent section ‘Forensic Medicine’ of the Society of German Natural Scientists and Doctors (Berlin)

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Decision to found the German Society of Forensic Medicine (Breslau)

1924

Forensic medicine belongs to the compulsory disciplines in examination

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Specialist in forensic medicine (GDR)

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Change of discipline’s name from Gerichtsmedizin to Rechtsmedizin

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Specialist in forensic medicine (FRG)

Anatomic autopsy Structure and function of human body Andreas Vesalius (1514–1564) Great progress in the 16th–18th century Clinical autopsy Cause and seat of diseases, aetiology, pathogenesis Giovanni Battista Morgagni (1682–1771) Marie François Xavier Bichat (1771–1802) Carl von Rokitansky (1804–1878) Rudolf Virchow (1821–1902)

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Forensic autopsy Cause and manner of death Causality of external violence for death Livebirth or stillbirth Medical malpractice Johannes Bohn (1640–1718) Johann Ludwig Casper (1796–1864) Eduard von Hofmann (1837–1897)

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The 19th century was the time of professionalisation of forensic medicine. Professionalisation is a term from sociology and means that those belonging to a profession can define their belongings independently. In many European countries, local communities of forensic medicine had been founded. Table 1.2 gives an example for Germany. Later national societies were founded, for instance, the German Society of Forensic Medicine in 1904. In 1924, forensic medicine became a compulsory discipline at medical exams in Germany, which was a real breakthrough. With forensic medicine becoming a compulsory discipline at medical examinations, teaching of forensic medicine of course became compulsory at medical faculties too and institutes and professorships were founded at most medical faculties. In the following, only a few countries can be mentioned. The history of forensic medicine in many European countries and worldwide can be found in the books Forensic Medicine in Europe and History of Forensic Medicine. For the professionalisation of forensic medicine, the forensic autopsy was essential. There are three types of autopsies: • the anatomic autopsy, • the clinical autopsy, and • the forensic autopsy (see Box 1.5). The anatomic autopsy was developed already in the 16th century, mainly in Italy, later in the Netherlands. The clinical autopsy was developed also in Italy (Padua), especially by Giovanni Battista Morgagni (1682–1771), where he wrote his famous book De Sedibus et Causis Morborum. There were already claims in the 17th century that in violent death an autopsy should be performed instead of just a wound inspection, but it took till the

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19th  century that forensic autopsies were regularly performed. Autopsies were not only in clinical medicine but also in forensic medicine one of the most important research tools, and in many countries, for instance, Austria and the German counties, autopsy regulations were released by the government which have to be strictly followed.

1.6  Forensic medicine as an experimental science

1.6.1 France According to Bertrand Ludes (2008), modern forensic medicine was born in France during the French Revolution with the closure of old universities and the creation of three new faculties of medicine in Paris, Strasbourg and Montpellier. Medical studies were reorganised in 1794 and professorships of forensic medicine were established in the new faculties. For instance, in 1789, François-­ Emmanuel Fodéré (1764–1835) published his legislation enlightened by physical sciences, and treatises of forensic medicine in public health, which represented the first French publication with forensic medicine in its title. Fodéré distinguished between civil forensic medicine, criminal forensic medicine, administrative forensic medicine or public health and health and medicine policing. He held the chair of forensic medicine and public health in Strasbourg where he published in 1830 a new edition of this treatise. He defined forensic medicine as follows: ‘By forensic medicine one means the application of physical, natural and medical knowledge to the legislation of the people, the administering of justice, local government, the maintenance of public health’. New horizons were opened for forensic medicine with the development of pathological anatomy and analytical

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1.6.2 Prussia

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Figure 1.4  Mathieu Joseph Bonaventure Orfila (1786–1853). Source: Image from https://en.wikipedia.org/wiki/Johann_Peter_Frank#/ media/File:Johann_Peter_Frank.jpg.

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One of the most remarkable experts in forensic medicine in the 19th century was Johann Ludwig Casper (1796–1864), the founder of modern forensic medicine in Prussia (Figure 1.5a,b). He was born in Berlin in 1796 and died there in 1864. Casper studied medicine in Berlin, Göttingen and Halle and became a medical doctor in 1819. Already at the age of 24, he received his postdoctoral lecture qualification for pathology and legal medicine. After his graduation, he studied in private and state institutions for public health in England and France for one year and in 1825 he was appointed a private counsel and member of the Royal Medical Council of Brandenburg. From 1834 on, he was senior private counsel of medicine and member of the scientific deputation for health care. In 1839, he was appointed professor and medicolegal officer for Berlin and in 1850 the director of the Institute of Forensic Medicine, at that time called ‘Unterrichtsanstalt für Staatsarzneikunde’ (School for State Medicine). Caspar published more than 170 papers, some at the beginning of his career also on medical statistics. He published on mortality and life expectancy with regard to different countries, gender and business (e.g., the mean life expectancy at this time was 38.5 years in England and only 21.3 years in Russia; the mean life expectancy for theologians was 65.1 years and 50.8 years for medical doctors). In 1852, Caspar founded the quarterly Journal of Forensic Medicine and Public Health and in 1857 the first edition of his practical handbook of forensic medicine was published. His practical handbook, which was also translated into English, was revolutionary, since its content was based on own observations. His motto was ‘non hypotheses condo, non optiones vendito, quod vidi scripsi’. In the preface of his handbook, he wrote:

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t­oxicology, both vigorously promoted by Mathieu Joseph Bonaventure Orfila (1786–1853) (Figure 1.4), one of the most influential men in the development of scientific forensic medicine in France. As a Spaniard, Orfila was the physician of Louis XVIII and the dean of the Paris faculty from 1830 to 1848. He published famous books such as Traité de Toxicologie (1813) and Leçons de Médecine Légale (1823) and did experimental work in both toxicology and classical forensic medicine, including on putrefaction and postmortem wounds. By 1840, Orfila was able to use a test that has coined almost a whole branch of crime: the arsenic test of J. Marsh (1795– 1846) of 1836. According to Brouardel, there was a dramatic decline of poisoning trials in the decade 1830–1840 due to the Marsh test. Orfila was not only a brilliant scientist and teacher but also a ‘courtroom star’. Other famous forensic scientists were Alphonse Devergie (1798–1879), author of a monumental treatise in 1853, P.C.H. Brouardel (1837–1902) and Auguste Ambroise Tardieu (1818–1879), who was a pupil of Orfila and like his master a courtroom star. They no longer produced ‘treatises’, but special monographs on particular issues such as hanging, abortion, poisoning, wounds, etc. Tardieu wrote the first book on sexual abuse on children and on battered children; subpleural haemorrhages are named after him. Brouardel held the chair of forensic medicine in Paris between 1879 and 1896 and also became dean of the faculty of medicine. In Lyon, forensic medicine was developed by Alexandre Lacassagne (1843–1924), who held the chair of forensic medicine in the faculty of medicine in Lyon for over 30 years (1880–1913).

‘In this book as in all my lectures in the last thirty-­six years I have striven especially against the prime failing of most authors on forensic medicine, viz., the separation of it from general medicine, and have endeavoured to purify it from all irrelevant rubbish, which has been so copiously accumulated in it by tradition, want of experience in forensic matters, and therefore ignorance of the proper relation which the medical jurist bears to the judge, as well as mistaken ideas as to the practical object of the science. . . . The correct appreciation of a simple dogma, which is unquestionably correct as it is to be unalterably maintained, leads of itself to the necessary reform in treating of juridical medicine. I mean the dogma that a medical jurist is – a physician – nothing more, nothing less, nothing else, and, as this simple dogma has been grossly misunderstood, to make it still more plain, I again repeat, he is a physician, and not a lawyer etc. Just as a technologist, artist, or any other craftsman must hold his knowledge or experience in his art or trade at the service of justice in the interest of the common need, so must the physician, and nothing else is required of him. . . .

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Figure 1.5  (a) Johann Ludwig Casper (1796–1864), founder of modern forensic medicine in Prussia. (b) Johann Ludwig Casper: Practisches Handbuch der Gerichtlichen Medicin (Handbook of Forensic Medicine) 1st edition published in 1857.

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This erroneous blending of medical and legal ideas and objects is also combined with another greater and more consequential error in the practice of forensic medicine. I mean the tendency to endeavour to obtain strict apodictical proof, such as was required by the practice of the older penal courts. . . . I demand in what other branch of general medical diagnosis, of which the forensic is but a part, is such indubitable certainty required, or where can it be attained?’ (Johann Ludwig Casper, preface to the third edition of his textbook, 1860)

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His practical handbook achieved eight editions, the last of them published by Schmidtmann in 1905. Caspar also published an atlas of forensic medicine. His son-­in-­law was his successor as head of the Unterrichtsanstalt für Staatsarzneikunde and built in Berlin the famous Institute of Forensic Medicine at the Charité from 1884 to 1886 (Figure  1.6). Liman had visited the Paris morgue and the plans for the institute in Berlin were based on the building in Paris.

1.6.3 Austria In Vienna, an institute of forensic medicine had been founded by 1804 as the Institute of Forensic Medicine and Medical Police.

The claim of a medical police goes back to Johann Peter Frank, who was appointed professor of surgery in 1794 at the University of Vienna. His most famous book was System einer Vollständigen Medicinischen Polizey (System of Complete Medical Police). However, from 1844 to 1875, forensic autopsies were performed by pathologists, mainly by the famous pathologist Carl von Rokitansky. In 1875, Eduard von Hofmann, who was the first professor of forensic medicine at the University of Innsbruck (since 1869), moved to Vienna (Figure 1.7). In 1878, he published his famous Lehrbuch der Gerichtlichen Medicin (Textbook of Forensic Medicine), which was translated into four languages (French, Russian, Italian and Spanish) (Figure 1.8). The 11th edition of this famous textbook was published by his pupil Albin Haberda in 1922. He also published a famous atlas of forensic medicine (1898) with numerous photographs and illustrations which can be still used today for teaching purposes (Figures 1.9–1.12). Around this time, Johann Peter Frank founded a museum of biological specimens in Vienna which can still be visited today in the so-­called ‘Narrenturm’. von Hofmann moved the forensic preparations out of the museum of pathology and founded his own collection of forensic preparations, which now comprises more than 2000 preparations; preparations are still exhibited that appeared in his own textbook and atlas of forensic medicine.

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Figure 1.6  Cut through the Institute of Forensic Medicine at the University of Berlin (built from 1884 to 1886). The institute served also as a morgue where unidentified deceased were exhibited for public viewing. At the first floor, a man is standing in front of a glass window, behind the window bodies are exhibited in cooling cells. The Paris morgue served as a model for the construction of the morgue in Berlin.

Figure 1.7  Eduard von Hofmann (1837–1897), professor of forensic medicine in Vienna from 1875 to 1887. His time in Vienna is called the golden age of forensic medicine. He published not only a famous textbook and an atlas of forensic medicine, but numerous articles throughout the whole discipline. He had many important pupils who performed outstanding experimental research.

Figure 1.8  Title cover of Eduard von Hofmann’s ‘Textbook of Forensic Medicine’.

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In the UK, the development of forensic medicine lagged behind Italy, France and Germany due to differences in legal systems and practices. In contrast to English common law, the approach of Roman canon law to legal decision-­making encouraged the development of forensic medicine because, according to Vanezis, technical evidence by experts could be more easily incorporated as sentences were made by judges. This was in contrast with common law trials where the use of juries tended to discourage testimony that could not easily be understood by lay people. However, by the end of the 18th century, chairs of forensic medicine were founded in Edinburgh and Glasgow. By 1834, 37 medical schools in Great Britain provided courses of instructions in forensic medicine. The subjects of courses had been made obligatory for the medical curriculum of every medical school the year before. The rise and the decline of forensic

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Figure 1.10  Stab wounds of the back to illustrate Langer’s lines, from Eduard von Hofmann, Atlas of Forensic Medicine.

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Figure 1.9  Atlas of Forensic Medicine (Atlas Der Gerichtlichen Medizin) by Eduard von Hofmann with numerous illustrations by the painter A. Schmitson.

medicine in the UK are entwined with the incorporation of forensic medicine into the medical curriculum. By 1944, the instruction in forensic medicine given to medical students was excessive. However, later, the Royal Commission on Medical Education in 1968 did not consider the subject at all. As a consequence, universities could claim with considerable justification that the provision of forensic medicine as a speciality of its own was not important, particularly when money was short and virtually no research came from those who specialised in the subject. A decline of academic forensic medicine cannot only be observed in the UK but also in Germany. Reasons for this are an inadequate financing of forensic medicine, competition on money between different medical disciplines and a scientific recognition that is based on impact factors and external funding.

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1.7  Current problems

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Figure 1.11  Suicide by hanging and strangulation mark, from Eduard von Hofmann, Atlas of Forensic Medicine.

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Forensic medicine developed as a recognisable separate scientific discipline in most European countries in the 19th century, but was not considered to be a separate academic discipline. More than 100 years ago, the famous German surgeon Theodor Billroth (1826–1894) wrote in a book on teaching and learning medicine at German-­speaking universities (1876) that there is no need to teach forensic medicine at universities, since it is not a science on its own, but rather a compilation of other independent sciences, and that the knowledge of these sciences is only used for practical purposes (e.g., judicial questions) (Figure 1.13). This approach is not only wrong but also proved hard to dispel and has accompanied our discipline for more than a century. Of course, forensic medicine has its own scientific profile and deals with questions which are not found in other disciplines. Already Billroth could have been aware of this, since the forensic physician Eduard von Hofmann (1837– 1897) was not only working at the medical faculty in Vienna at the same time as Billroth (see Figure 1.13) but also founded the famous Vienna School of Forensic Medicine with excellent experimental research.

Figure 1.12  Washerwoman’s skin, from Eduard von Hofmann, Atlas of Forensic Medicine.

In England and Wales, the number of professorships in forensic medicine has decreased dramatically, and in Germany, several institutes of forensic medicine were closed in the last 15 years. Forensic medicine, however, does have its own research profile and deals with questions and issues that are not found in other disciplines. These include: • Thanatology: postmortem changes, time of death, wound-­age estimation, distinction between antemortem and postmortem injuries, vitality of wounds, etc. • Traumatology as a basis of reconstruction. • Postmortem toxicology. • Toxicological analysis of various body fluids. • Hair analysis.

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Figure 1.13  (a) Cover page of the book of Theodor Billroth on learning and teaching medical sciences at universities in German-­speaking countries. (b) The figure on the left is Theodor Billroth (1826–1984), right beside him Eduard von Hofmann (1837–1897). This figure is taken from a picture of all members of the medical faculty of the University of Vienna. Obviously, Billroth did not recognise the importance of forensic medicine, importance of Eduard von Hofmann and his great experimental school and the specific profile of the discipline.

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driving ability. • Stain analysis. • Clinical forensic medicine.

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References and further reading

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Ackerknecht, E.H. (1950/51). Early History of Legal Medicine. Ciba Symposium 11, No. 7, pp.  1286–1304. Reprinted in: Burns, R.C. (1977). Legacies in Law and Medicine, pp. 247–271. New York: Science History Publications. Bonte, W. (2000). History/forensic medicine. In: J.A. Siegel, P. Saukko, G. Knupfer (eds.), Encyclopedia of Forensic Sciences, pp. 1064–1070. San Diego, San Francisco, New York: Academic Press. Brittain, R.P. (1965a). Origins of legal medicine: Constitutio Criminalis Carolina. Medico-­Legal Journal 33: 124–127. Brittain, R.P. (1965b). Origins of legal medicine: the origin of legal medicine in Italy. Medico-­Legal Journal 33: 168–173. Brittain, R.P. (1966a). Origins of legal medicine: Leges Barbarorum. Medico-­Legal Journal 34: 21–23. Brittain, R.P. (1966b). Origins of legal medicine: the Assizes of Jerusalem. Medico-­Legal Journal 34: 72–73. Brittain, R.P. (1966c). Origins of legal medicine: the origin of legal medicine in France. Medico-­Legal Journal 34: 168–174.

Brittain, R.P. (1967). Origins of legal medicine: Roman law: Lex Duodecim Tabularum. Medico-­Legal Journal 35: 71–72. Cattaneo, C. (2008). Legal Medicine in Italy. In: B. Madea and P. Saukko (eds.), Forensic Medicine in Europe, pp. 209–227. Lübeck: Schmidt-­Römhild. Camps, F.E. (1968). Gradwohl’s Legal Medicine, 2nd edn, pp. 1–14. Bristol: Wright. Clark, M. and Crawford, C. (eds.) (1994). Legal Medicine in History. Cambridge: Cambridge University Press. Corbella, J. (2004). Historia de Medicina Legal. In: G. Calabuig and E. Villanueva (eds.), Medicina Legal y Toxicologia, 6th edn, pp. 8–12. Barcelona: Masson. Dérobert, L. (1973). Histoire de la médicine légale. Zacchia 48: 1–37, 161–192, 341–382, 533–547. Feola, F. (2007). Profilo Storico della Medicina Legale. Dalle origini alle soglie del XX. secolo. Torino: Editioni Minerva Medico. Fischer-­Homberger, E. (1983). Medizin vor Gericht. Gerichtsmedizin von der Renaissance bis zur Aufklärung. Bern: Verlag Hans Huber. Geserick, G., Vendura, K., and Wirth, I. (2005). Das Institut für Rechtsmedizin der Charité in Berlin-­Mitte. Ansichten und Einblicke. Berlin, www.photobuch24.de Hausner, E. (2008). Historische Sammlung des Instituts für gerichtliche Medizin der Universität Wien. Wien: Edition Hausner, Desktop-­ Publishing, Oliver Hausner. Herber, F. (2002). Gerichtsmedizin unterm Hakenkreuz. Leipzig: Militzke.

National Research Council (2009). Strengthening Forensic Sciences in the United States. A Path Forward. Washington: National Academies Press. Payne-­James, J. (2005). History of forensic medicine. In: J. Payne-­James, R. Byard, T. Corey, and T. Henderson (eds.), Encyclopedia of Forensic and Legal Medicine, pp. 498–519. Amsterdam, Boston: Elsevier. Pounder, D.J. (2008). Scotland. In: B. Madea and P. Saukko (eds.), Forensic Medicine in Europe, pp. 343–356. Lübeck: Schmidt-­Römhild. Preuss, J. and Madea, B. (2009). Gerhard Panning (1900-­ 1944): A German forensic pathologist and his involvement in Nazi crimes during Second World War. American Journal of Forensic Medicine and Pathology 30: 14–17. Prokop, O. (1969). Karl Landsteiner zum Gedächtnis. Beitrage zur Gerichtlichen Medizin 26: 141. Rutty, G.N. (2005). The academic forensic pathologist. An endangered species? Forensic Science, Medicine and Pathology 1: 167–168. Schmidt, O. (1953). Gerichtliche Medizin in den ersten geschriebenen Rechten germanischer Staemme. Deutsche Gesellschaft für Gerichtlichen Medizin 42: 121–132. Smith, S. (1951). The history and development of forensic medicine. British Medical Journal 1: 599–607. Sung Tzu (1981). The Washing Away of Wrongs. Translated by Brian E. Mc Knight. Center of Chinese Studies. The University of Michigan. Thomas, F. (1974). Milestones in forensic science. Journal of Forensic Science 19: 241–254. Thorwald, J. (1965). Das Jahrhundert der Detektive: Weg und Abenteuer der Kriminalistik. Zürich: Droemer. Thorwald, J. (1966). Die Stunde der Detektive: Werden und Welten der Kriminalistik. Zürich: Droemer. Timmermanns, S. (2006). Postmortem: How Medical Examiners Explain Suspicious Deaths. Chicago: The University of Chicago Press. Vanezis, P. (2004). Forensic medicine: Past, present and future. Lancet 364: 8–9. von Neureiter, F. (1935). Anfaenge gerichtlicher Medizin nach den Stadtrechten des deutschen Mittelalters. Deutsche Gesellschaft für Gerichtliche Medizin 24: 1–7. Wirth, I., Geserick, G., and Vendura, K. (2008). Das Universitätsinstitut für Rechtsmedizin der Charité 1833–2008. Lübeck: Schmidt-­Römhild.

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Hunt, B. and Pounder, D. (2005). The forensic pathology initiative from Home Office pathologist to home-­ office pathologists? Journal of Clinical Forensic Medicine 12: 289–290. Jentzen, J.M. (2009). Death Investigation in America. Coroners, Medical Examiners and the Pursuit of Medical Certainty. Cambridge: Harvard University Press. Jones, A.W. (2007). The distribution of forensic journals, reflections on authorship practices, peer-­view and role of the impact factor. Forensic Science International 165: 115–128. Ludes, B. (2008). Forensic medicine in France. In: B. Madea and P. Saukko (eds.), Forensic Medicine in Europe, pp. 113–41. Lübeck: Schmidt-­Römhild. Luna Maldonado, A. and Pérez-­Cárceles, M.D. (2008). Forensic medicine in Spain. In: B. Madea and P. Saukko (eds.), Forensic Medicine in Europe, pp. 373–385. Lübeck: Schmidt-­Römhild. Madea, B. (2004). 100 Jahre Deutsche Gesellschaft für Gerichtliche Medizin/ Rechtsmedizin. Vom Gründungsbeschluss 1904 zur Rechtsmedizin des 21. Jahrhunderts. Deutsche Gesellschaft für Rechtsmedizin 2004. Madea, B. (2004). Special issue: 100th Anniversary of the German Society of Legal Medicine. Forensic Science International 144 (2–3): 83–302. Madea, B. (2008). Forensic Medicine in Germany. In: B. Madea and P. Saukko (eds.), Forensic Medicine in Europe, pp. 143–164. Lübeck: Schmidt-­Römhild. Madea, B. and Saukko, P. (2007). Future in forensic medicine as an academic discipline – focussing on research. Forensic Science International 165: 87–91. Madea, B. and Saukko, P. (2008). Forensic Medicine in Europe. Lübeck: Schmidt-­Römhild. Madea, B., Saukko, P., and Musshoff, F. (2007). Tasks of research in forensic medicine  – different study types in clinical research and forensic medicine. Forensic Science International 165: 92–97. Madea, B. (2017). History of Forensic Medicine. Berlin: Lehmann’s Media GmbH. Madea, B. (2017). History of the Autopsy. In: Madea B (ed.), History of Forensic Medicine, pp. 38–61. Berlin: Lehmann’s Media GmbH. Mallach, H.J. (1996). Geschichte der Gerichtlichen Medizin im deutschsprachigen Raum. Lübeck: Schmidt-­Roemhild. Mende, L.J.C. (1819). Ausfuehrliches Handbuch der gerichtlichen Medizin für Gesetzgeber, Rechtsgelehrte, Ärzte und Wundärzte, Vol. I. Leipzig: In-­der-­Dyk’sche Buchhandlung.

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achievement of elucidating and presenting evidence to demonstrate facts and truth to the required standard of proof within the justice system’.

According to D. N. Vieira, the basic mission of forensic medicine is the proper application of justice. The honour and freedom of individuals often depend on it. Much of what justice has been in the past few centuries has passed through forensic medicine, and much of what justice ought to be will pass through it in the future. Forensic medicine is always working at the frontier of new developments in the society and has to face these developments by research and expertise. For instance, new psychoactive substances (NPS) are a development of the last twenty years, but there were mainly experts in forensic medicine who developed analytical methods to detect NPS and founded the basis for new laws. Violation of human rights and torture are unfortunately still common today. Specialists in forensic medicine are the leading experts to protect human rights. Migration is a worldwide phenomenon of our time. Due to national legislation, age estimation in living is important and the appropriate methods were developed in forensic medicine. Like child abuse, the abuse of elderly was for a long time a neglected phenomenon. Again, there are experts in the field of forensic medicine who deal with these phenomena. Entry criteria to legal and forensic medicine may vary from country to country, but usually include a specific period of ­general medical training followed by postgraduate training in the field.

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The traditional and comprehensive definition of legal medicine as a special medical discipline that prepares medical knowledge for the purposes of law still remains a reliable description today. Thus, legal medicine is a typical interdisciplinary subject which includes the tasks outlined in Figure 2.1 and Box 2.1. According to the European Council of Legal Medicine (ECLM), the speciality profile was defined as follows:

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‘Legal and forensic medicine is the application of medical knowledge and methodology for the resolution of legal ­questions and problems for individuals and society. It involves the observation, documentation, collection, assessment, and scientific interpretation of medical evidence deriving from clinical and post-­ mortem investigations required for the ­different fields of law, including criminal, civil, work, family, and administrative. Its core activities are clinical forensic medicine and forensic pathology, but other areas of science and expertise including forensic toxicology, forensic psychiatry, forensic genetics, forensic anthropology may be required depending on the nature of the case.

Legal and forensic medicine requires specific skills to be applied in accordance with the various legal frameworks. It may need to determine not only the diagnosis (the what) but also to clarify and specify the circumstances which surround any event or incident (the who, why, when, where, and how). In order to achieve these goals, a unique combination of ­medical, forensic and legal knowledge is required which presupposes certain unique principles fundamental to the

Handbook of Forensic Medicine, Volume 1, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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Box 2.2  Typical and additional roles of a forensic physician.

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Typical roles • Determine fitness to be detained in custody • Determine fitness to be released • Determine fitness to be charged: competence to comprehend charge • Determine fitness to transfer • Determine fitness to be interviewed by the police or detaining body • Advise that an independent person is required to ensure rights for a vulnerable or mentally disordered individual • Assess alcohol and drug intoxication and withdrawal symptoms • Examine comprehensively a person’s ability to drive a motor vehicle • Undertake intimate body searches for drugs (e.g., anally/vaginally concealed drugs) • Carry out precise documentation and interpretation of injuries • Take forensic samples • Assess and treat personnel injured whilst on duty, including needle-­stick injuries • Pronounce life extinct at a scene of death and undertake a preliminary advisory role • Undertake mental-­state examinations • Examine adult complainants of serious sexual assault and the alleged perpetrators • Examine alleged child victims of neglect, physical or sexual abuse • Examine victims and assailants in alleged police assaults

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Figure 2.1  Tasks of forensic medicine. As a cross-­sectional discipline, it uses the knowledge of numerous different fields of expertise.

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Areas of competence for specialists in forensic medicine Investigations at the scene of death/crimea External examinations of bodies External examinations of bodies prior to cremationb Estimation of the time since deatha Identificationa Forensic anthropologyb Medicolegal autopsiesa Clinical forensic medicine, particularly focused on the examination of injuries in living individualsb Analyses for alcohol and drugsb Forensic toxicologya Clinical toxicologyb Expert opinions on the ability to drivea Expert opinions on the suitability for drivingb Expert evidence on legal responsibilitiesb Haemogeneticsb • Paternity diagnostics • Stain analyses Coroner’s autopsiesb Expert opinions given in courtb

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Box 2.1  Areas of competence for specialists in forensic medicine.

Additional roles • Expert opinion in courts and tribunals • Death in custody investigation • Provide opinion in fatal accident inquiries (FAI; Scotland) • Pressure group and independent investigators in ethical and moral issues Victims of torture War crimes Female genital mutilation (female circumcision) • Refugee medicine (medical and forensic issues) • Asylum-­seeker medicine (medical and forensic issues) Source: Expanded and modified from Howitt and Stark (1996) and Payne-­ James et al. (2011).

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 Services just offered by forensic medicine;  Services concentrated in forensic medicine.

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However, although there are some common rules, there are also considerable variations between countries. Different ­countries do not only have different laws but also different legal systems. For example, Europe represents a mosaic of sociocultural, economic and legal realities, and this is reflected in forensic medicine, it having always influenced a variety of operational modes, especially in terms of organisational systems, structures and functional competences (Vieira 2008). Whilst some countries still have a wealth of academic forensic units, in others, forensic medicine is mostly carried out by ­so-­called clinical forensic practitioners. By definition, clinical forensic medicine includes all medical (health care) fields related

to legal, judicial and police systems (Payne-­James and Busuttil 2003; Box 2.2). There is a wide range of practitioners who perform work in forensic medicine and forensic sciences: • Forensic pathologists. • Clinical forensic practitioners. • Forensic nurses (e.g., in the UK). • Forensic psychiatrists/psychologists. • Forensic odontologists. • Forensic scientists, especially in forensic toxicology and forensic genetics. • Forensic anthropologists. • Forensic archaeologists. In special cases, consultants from other clinical disciplines are asked to work together with the forensic pathologist or clinical

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inquiring authorities, and duties and rights of experts and witnesses. Knowledge in legal medicine is indispensable for every physician, since: • every doctor has to be able to carry out an external post-­ mortem examination; • most victims of violence are treated in hospitals and private practices and every physician has to be able to recognise and record injuries and might be summoned to court; • the physician has to know their own and the patient’s position in the respective legal system including all rights and obligations; • the doctor has to be informed about examinations that might be carried out in an institute of legal medicine in order to advise her or his patient adequately (e.g., on clinical toxicology). Additionally, the physician has to learn how medicolegal experts evaluate statements and findings, since she or he will also have to verify their validity and significance in practice (e.g., defensive manoeuvres in cases of child abuse, simulated illness, etc.). At conventional and traditional universities, forensic medicine is not only taught in the medical faculties but also in several other faculties that are closely linked to this subject; for example, faculties of law and of mathematics and natural sciences (food chemistry, pharmacy, biology, chemistry, etc.). It is the medicolegal commitment to research and teaching, in particular, that creates an indispensable basis for proper quality in services, thereby increasing the stability of law and justice in society. Legal medicine also provides advice to jurisdiction and politics regarding case law and required amendments. Medicolegal research deals with very specific subjects which should remain separate from other subject areas; this is essential for public and legal security. Its main tasks are: thanatology, traumatology as a basis for reconstruction of actions and movements, toxicological and molecular biological examinations of forensically relevant matrices, ballistics of wounds, epidemiology and causal research in alcohol-­or drug-­affiliated traffic accidents, establishment of limits regarding the ability to drive and those aspects of medical law and status that originate from forensic practice. Knowledge based on medicolegal routine and research has always been used in aspects of prevention. For example, an important increase in passive vehicle safety could be derived and developed from autopsy findings in road traffic accident victims. There can be no doubt that justice will only be able to reach its maximum effectiveness if it can rely on a wide collaboration of modern and dynamic medicolegal and forensic services, services which need to operate at the swift pace of life and legal needs (Vieira 2008). As there are various legal systems and diversity of different medicolegal systems worldwide, international cooperation and harmonisation are essential. As there is no international consensus on how forensic medicine should be delivered, there is a continuing need to learn about our similarities and differences, in order to further harmonise the system within medicolegal practice, especially medicolegal autopsy rules and laboratory procedures. The ECLM has meanwhile taken the initiative of European Union (EU) recognition of legal and forensic medicine as medical

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forensic practitioner (e.g., gynaecologists, paediatricians, emergency physicians, etc.). Separate disciplines could include forensic entomology and forensic botanic science. This system is characterised by a number of disadvantages; for instance, a number of different experts may be involved in the case. When they appear in court, the manners in which specific opinions are given by them vary, often against the interest of the victim. Secondary victimisation may occur, which could be avoided by changing the way that injuries, stains or traces are secured. Whilst in some countries, for instance Germany, both the clinical and the pathological aspects of forensic medicine are undertaken by the same individual, in other countries, this is strictly separated due to specialisation. Whereas for example a forensic pathologist in the UK or USA receives a training in pathology and then specialises in forensic pathology, a specialist in forensic medicine in Germany has to study both, forensic pathology and also clinical forensic medicine. Postgraduate training in forensic medicine differs from country to country; for example, in Germany, postgraduate specialisation in forensic medicine takes at least 5 years (60 months) according to the teaching regulations of the state medical chambers. One common problem in many countries is that forensic medicine is often extremely underfunded. Since forensic medicine carries out work for the police, justice and public health departments, and is also engaged in medical education and research, it should be financed by all four ministries in charge (ministries of justice, internal affairs, health care and science). However, often forensic subjects and forensic institutions are funded by justice ministries or ministries of science only and therefore appear to be relatively underfunded. Forensic medicine and forensic sciences therefore need to pay close attention to the great shifts of our day and age, and spare no effort to modernise themselves, to keep up-­ to-­date and to develop, so that they can play a fundamental role in the service of justice and community and remain at the centre of academically qualified research and teaching. In Scotland, funding of medicolegal services is provided by particular service contracts between the Crown Office (Ministry of Justice) and the appropriate university departments (of pathology) or the National Health Service (NHS). By this method, posts and administration within such departments are directly financed by the Crown Office based on contracts running for periods such as 5 years. After this period, contracts are reviewed and negotiated again. The system also applies to clinical medicolegal services. Although it can be assumed that this type of funding appears rather safe in terms of changing external factors and conditions, it might create problems regarding the independency of forensic medical experts originating from a conflict of interest. Furthermore, departments may be transformed to sole-­service providers, thus losing their appropriate academic input. University institutes of forensic medicine are responsible for the teaching of medical students within the faculty. Essential subjects of teaching are: post-­mortem examination, certificate of death, legal and ethical issues within the profession, signs of violence, securing and documentation of findings, contact with the

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2.1.2 Switzerland

In Switzerland, medical expert witnesses are needed for various purposes, and in particular to clarify questions regarding the following topics: the causal relationship between an accident and a damage to health, a patient’s concrete inability to work or earn money, a patient’s ability of judgement and the determination of various types of damages. Medical experts may provide for opinions in the context of judicial disputes, or outside such disputes (e.g., in the case of extrajudicial expert opinions by the Swiss Medical Association [FMH]).2 Moreover, there are numerous forms of opinions: (i) monodisciplinary expert opinions, (ii)  bidisciplinary expert opinions and (iii) multidisciplinary expert opinions. Even though there are only some legally binding guidelines, which apply to the preparation of expert opinions, the latter are usually structured as follows: After having referenced basic information on the patient and the expert, the opinion mentions the various sources used, as well as the examined person’s anamnesis and relevant medical records. The opinion then addresses the examined person’s details – that is, current condition, family and social anamnesis and education and work anamnesis  – before outlining the investigation findings. The opinion further outlines the diagnostics and assessments made, as well as all questions asked. A bibliography as well as an inventory of files must finally be included in the opinion (Siegel 2007).

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Fischer-­Homberger, E. (1983). Medizin vor Gericht. Gerichtsmedizin von der Renaissance bis zur Aufklärung. Bern: Verlag Hans Huber. Howitt, J. and Stark, M. (1996). The Role of the Independent Forensic Physician. Education and Research Committee of the Association of Police Surgeons. Association of Police Surgeons, Harrogate. Madea, B. (2008). Forensic Medicine in Germany. In: B. Madea and P. Saukko (eds.), Forensic Medicine in Europe, pp. 143–164. Lübeck: Schmidt-­Römhild. Madea, B. and Saukko, P. (2007). Future in forensic medicine as an academic discipline – focussing on research. Forensic Science International 165: 87–91. Madea, B. and Saukko, P. (eds.) (2008). Forensic Medicine in Europe. Lübeck: Schmidt-­Römhild. Madea, B., Saukko, P., and Musshoff, F. (2007). Tasks of research in forensic medicine  – different study types in clinical research and forensic medicine. Forensic Science International 165: 92–97. Madea, B. (2017). History of Forensic Medicine. Berlin: Lehmann’s Media GmbH. Payne-­James, J. and Busuttil, A. (2003). History and development of forensic medicine and pathology. In: J. Payne-­James, A. Busuttil, and W. Smock (eds.), Forensic Medicine: Clinical and Pathological Aspects, pp. 3–12. London: Greenwich Medical Media. Payne-­James, J., Jones, R., Karch, S.B., and Manlove, J. (2011). Simpson’s Forensic Medicine, 13th edn. London: Hodder Arnold. Vieira, D.N. (2008). Foreword. In: B. Madea and P. Saukko (eds.), Forensic Medicine in Europe, pp. 13–15. Lübeck: Schmidt-­Römhild. https:// www.ecml.org/ Wyatt, J., Squires, T., Norfolk, G., and Payne-­James, J. (2011). Oxford Handbook of Forensic Medicine. Oxford: Oxford University Press.

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References and further reading

Insurance companies, for example, health insurance, accident insurance and liability insurance, often depend on the knowledge of medical experts in order to clarify potential obligations to reimburse medication and medical treatment costs to insured persons. Moreover, different courts and the prosecution depend on the competences of expert witnesses in order to adequately assess medical facts underlying legal disputes. The following chapter aims to provide a rough outline of some important topics concerning the legal requirements and considerations applying to medical expert witnesses, with a particular focus on Switzerland and Germany.

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monospeciality, and, thus, as one of the guardians of the rights of the most vulnerable persons. Since forensic has its own specific scientific profile, especially university-­ based institutes of forensic medicine have to do research and improve the knowledge in the field.

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Judicial expert opinions

2.1.1  General considerations Under certain circumstances, doctors may be asked to act as expert witnesses and to provide corresponding expert opinions. In such cases, the medical expert has to answer the questions asked in an unbiased manner, to the best of his or her knowledge.1

As regards judicial expert opinions, a public prosecutor or a court may deem that precise current technical knowledge  – in casu medical knowledge – is necessary to determine or assess relevant facts of a case, and may thus appoint one or several experts.3 As the expert must be technically competent to assess the questions at stake, it may indeed be necessary to appoint a team of experts

 See the brochure of the Swiss Academy of Medical Sciences (SAMS) ‘Rechtliche Grundlagen im medizinischen Alltag’, available under: https://www.fmh.ch/files/ pdf7/rechtliche-­grundlagen-­2020-­de.pdf (last accessed 28 October 2020).

 See the webpage of the Swiss Medical Association (FMH) on extrajudicial expert opinions, available under: https://www.fmh.ch/ueber-­die-­fmh/organisation/fmh-­ gutachterstelle.cfm#i111678 (last accessed 28 October 2020). 3  Cf. Article 182 Swiss Criminal Procedural Code.

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Under certain circumstances, one may wish to resort to the knowledge of medical experts without necessarily involving the court. Then, the possibility of requesting an extrajudicial expert opinion makes sense. The Swiss Medical Association (FMH) established an office, which is specialised in the provision of extrajudicial expert opinions with regard to the complaint of patients concerning malpractice or an institutional fault which has possibly led to a health impairment. Using these services represents an opportunity to settle medical disputes whilst avoiding expensive legal proceedings.8 However, the expertise office of the FMH is not

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Expert opinions in the social insurance sector

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In Switzerland’s social insurance sector, the insurers are the ones who commission the expert opinions, as insurers are responsible for clarifying the facts in accordance with the principle of investigation (Swiss Insurance Medicine  2019). In case the insurance companies appoint an expert to provide for an expert opinion, Article 44 of the Swiss Federal Law on the General Part of Social Security Law9 is applicable. In most cases, the expert’s name must be disclosed to the parties and they may challenge the expert for ‘relevant reasons’ – that is, in case of a conflict of interests – and make counterproposals. Such a challenge may be justified by reasons of a formal nature, that is, personal conflict of interests or bias, but not by material reasons, such as the professional qualification of the expert witness.10 The latter concerns are however considered in the evaluation of the evidence. In the field of disability insurance, the legal regulations provide that multidisciplinary expert opinions – that is, expert opinions involving three or more medical disciplines  – addressing disability issues are to be attributed ‘randomly’ to experts, Article 72bis, paragraph 2 of the Regulation on Disability Insurance.11

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from different disciplines.4 The appointment of one or several experts may result from a request by a party or on the court’s own accord. Court-­appointed experts are subject to the same rules on conflicts of interests as judges.5 Moreover, such experts are subject to Article 307 of the Swiss Criminal Code (SCC) regarding perjury by an expert witness (Weiss 2018). However, the legal assessment of the medical facts may not be delegated to the expert witness and thus remains in the hands of legal experts (Aebi-­ Mueller et al. 2016). There are several requirements which apply to judicial expert opinions in order for their value to be recognised. In accordance with the Swiss Federal Supreme Court’s case law,6 the expert opinion must in particular be based on all-­round research and investigations. Moreover, all complaints which were received must be considered. The opinion must be provided with full knowledge and taking into account the patient’s previous records (i.e., anamnesis). Furthermore, the opinion must fit in the medical context and situation. Finally, the conclusions of the expert must be well-­founded. As regards the appreciation of judicial expert opinions, the principle of free sifting of evidence is applicable.7 The level of evidence required is that of ‘overwhelming likelihood’ (‘überwiegende Wahrscheinlichkeit’). This would be the case if there were no concrete objections to any of these elements. The judge must not deviate from the expert’s conclusions, except in case of justified reasons (Swiss Insurance Medicine 2019).

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4   Swiss Academy of Medical Sciences (SAMS), ‘Rechtliche Grundlagen im medizinischen Alltag’, available under: https://www.fmh.ch/files/pdf7/rechtliche-­ grundlagen-­2020-­de.pdf (last accessed 28 October 2020). 5  Cf. Article 183, paragraph 3 and Article 56 Swiss Criminal Procedural Code. 6  See Swiss Federal Supreme Court, decision 3 May 1996, BGE 122 V 157, 160. 7  See Swiss Federal Supreme Court, decision 14 June 1999, BGE 125 V 351; and decision 3  June 2015, BGE 141 V 281. The following provisions are moreover applicable in the social insurance sector: Article 55, paragraph 1 of the Swiss Federal Law on the General Part of Social Security Law (ATSG  – see footnote below) in connection with Article 19 of the Swiss Federal Act on Administrative Procedure of 20 December 1968 (SR 172.021) and Article 40 of the Swiss Federal Act on Federal Civil Procedure of 4 December 1947 (SR 273). 8  The cost for an expert opinion by the FMH amounts to max. 1,000 CHF (Hartmann 2019). For more information, see http://www.droitsdupatient.ch/ services-­befmh.cfm (last accessed 28 October 2020).

2.1.3 Germany General considerations Medical expert witnesses are also of great importance in Germany. The expert’s opinion is a possible means of evidence, and the involvement of an expert is always necessary when facts have to be established or questions have to be answered which the court cannot solve by its own expertise (Dettmeyer 2016). Although there is no legal definition of the term expert witness, a common definition provides that: ‘The expert witness is a natural person who possesses above-­average knowledge in a delimited field of the humanities and natural sciences, technology, economics, art or in any other field and makes this special expertise available to anyone on request in a personal, independent, impartial and objective manner’ (Schlund 2016). The expert witness is an assistant to the court and is not a decision maker.12

 Bundesgesetz über den Allgemeinen Teil des Sozialversicherungsrechts (ATSG) of 6 October 2000 (SR 830.1). 10  See Swiss Federal Supreme Court, decision 8 February 2006, BGE 132 V 93. 11  Swiss Federal Ordinance on Invalidity Insurance (IVV) of 17 January 1961 (SR 831.201). For further information, see https://www.bsv.admin.ch/bsv/de/home/ sozialversicherungen/iv/grundlagen-­gesetze/organisation-­iv/medizinische-­ gutachten-­iv.html (last accessed 20 October 2020); Swiss Federal Supreme Court, decision 28 June 2011, BGE 137 V 210. 12  Federal Court of Justice in criminal matters, 9,293; judicial power is exercised exclusively by the judge. 9

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Duties of the expert

The expert witness is selected by the public prosecutor, the police13 or the court.14 Legislation provides for the obligation to consult an expert, for example, on admission to a psychiatric hospital,15 on postmortem examination16 and on suspected poisoning.17 The law stipulates an obligation to act as an expert if he or she has been publicly appointed to render opinions of the required kind or if he or she publicly and commercially practises the science, art or trade the knowledge of which is a prerequisite for rendering an opinion or if he or she has been publicly appointed or authorised to exercise such profession.18 A doctor is publicly appointed with the licence to practise medical science and is obliged to act as an expert regardless of whether the activity is actually carried out (Meyer-­Grossner and Schmitt 2020). An autopsy is not to be performed by the physician who treated the deceased person during the illness that directly preceded the death, but might attend the autopsy to give information relating to the deceased’s medical history.19 The refusal of the expert opinion is possible, for example, because of heavy professional demands, the need for rest leave or professional unsuitability (Meyer-­Grossner and Schmitt  2020). The expert opinion may also be refused if this would violate the doctor’s duty of confidentiality (Dettmeyer 2016).

The rules for the expert witness are amongst others regulated in § 410, paragraph 1 Code of Civil Procedure, which states that the expert will provide the required expert opinion impartially and to the best of his knowledge and belief. Upon receipt of the order, the expert shall examine whether he or she accepts the expert opinion order or whether he or she may reject the appointment as expert. If the expert wants the revocation, he must explain why the expert opinion is unreasonable for him.23 The expert shall prepare the expert opinion himself and on his own responsibility.24 A delegation to assistants cannot take place without consulting the court or the client (Rosenberger 2020). The explanations must be as comprehensible as possible (Dettmeyer 2016). Because of the requirement of equality of arms, the explanations must be carried out in such a way that an informed opinion-­forming process is logically verifiable and comprehensible as far as possible for an open-­minded non-­physician.25 The expert shall be obliged to provide a defect-­free expert opinion, if he accepted the request (Dettmeyer, 2016). An expert is not entitled to compensation if the expert opinion does not meet the minimum requirements (Dettmeyer 2016). Important personal reasons which prevent the expert opinion from being issued or sudden circumstances which no longer allow the expert opinion to be issued as well as delays must be reported immediately (Dettmeyer, 2016).

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Liability of the expert According to § 839a German Civil Code, the expert has to make compensation for the damage incurred by a party to the proceedings as a result of a court decision based on an intentionally or by gross negligence submitted false expert opinion. By analogy, the liability of an expert who gives his opinion in a preliminary investigation by the public prosecutor is also applicable.26 An objectively incorrect opinion must have been given from the perspective of an expert.27 This may be due to insufficient establishment of facts, wrong conclusions or non-­consideration of prevailing doctrines (Schulze et al. 2017).

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The judge should reach an agreement with the expert by when the opinion can be issued.20 The client must carefully and precisely specify the report questions.21 The expert witness has the right, like stated in § 80 German Code of Criminal Procedure, to hear witnesses and defendants, inspect files and attend hearings for the preparation of his opinion. Furthermore, the client shall, for example, upon request, explain the assignment, arrange for the questioning of parties, communicate facts, allow the expert to participate in other investigations and issue other instructions (Dettmeyer 2016). After receiving the expert’s opinion, the court must review it for completeness and, if necessary, work towards its completion (Deutsch and Spickhoff 2008). The expert may be sworn in after the expert opinion has been issued by giving an oath that he has given his opinion impartially and to the best of his knowledge.22 If the court wants to deviate from the expert opinion, this must be justified by its own expertise or otherwise (Jessnitzer 1978).  Cf. § 161a, paragraph 1 German Code of Criminal Procedure.  Cf. § 73, paragraph 1 German Code of Criminal Procedure. 15  Cf. § 81 German Code of Criminal Procedure. 16  Cf. § 87 et seq. German Code of Criminal Procedure. 17  Cf. § 91 German Code of Criminal Procedure. 18  Cf. § 75 German Code of Criminal Procedure. 19  Cf. § 87, paragraph 2 German Code of Criminal Procedure. 20  Cf. § 73, paragraph 1 German Code of Criminal Procedure. 21  Higher Regional Court Hamm, decision 21 October 1968, VRS 36, 290. 22  Cf. § 79 German Code of Criminal Procedure.

Acknowledgements Thanks to Marie-­Hélène Spiess and Oliver Scharp for their assistance in drafting this chapter.

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23  Regional Court Bochum, decision 29 October 1985, 7 T 644/85, NJW 1986, 2890. 24  Court of Appeal Berlin, decision 26 April 2004, 20 U 57/03, Arztrecht in 2005, 273–274. 25  Federal Court, decision 24 June 1980, VI ZR 7/79 – Arztrecht 1981, 40–43. 26  Federal Court, MDR 2014, MDR 2014 p. 523 et seq. 27  Federal Court, decision 24 July 2014, III ZR 412/13.

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Aebi-­Mueller, R., Fellmann, W., Gaechter, T. et  al. (2016). Arztrecht. Bern: Staempfli Verlag. Canela, C., Buadze, A., Dube, A. et al. (2019) How do legal experts cope with medical reports and forensic evidence? The experiences, perceptions, and narratives of Swiss judges and other legal experts. Frontiers in Psychiatry 13:10:18. Christinat, R. (2019). Le procès en responsabilité civile médicale, Mise en œuvre en procédures civile et administrative. Basel/Neuchâtel: CN– Collection neuchâteloise. Dettmeyer, R. (2016). Die Rolle des medizinischen Sachverständigen – gesetzliche Vorgaben und Entscheidungen der Rechtsprechung. Rechtsmedizin 26, 316–324. Deutsch, E. and Spickhoff, A. (2008). Medizinrecht  – Arztrecht, Arzneimittelrecht, Medizinprodukterecht und Transfusionsrecht, 6th edn. Berlin/Heidelberg: Springer. Ebner G., Gächter, T., and Herzog-­Zwitter, I. (2020). Zur Rolle der medizinischen Begutachtung in der Missbrauchsbekämpfung. SZS 2020, pp. 113–121. Jessnitzer, K. (1978). Eigene Sachkunde des Richters bei der Rückrechnung. Blutalkohol 15, 315–322.

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Kuhn, M. and Poledna, T. (2007). Arztrecht in der Praxis, 2nd edn. Zurich: Schulthess Juristische Medien. Massimo, A. (2018). Zur geplanten Revision von Art. 44 ATSG. SZS 2018, pp. 144–160. Meyer-­Grossner, L. and Schmitt, B. (2020). Strafprozessordnung mit GVG und Nebengesetzen, 63rd edn. Munich: C.H. Beck. Rosenberger, R. (2020). In: F. Wenzel (2020). Handbuch des Fachanwalts. Medizinrecht, 4th edn, pp. 862–902. Köln: Luchterhand Verlag. Schlund, G. (2016). Kapitel 2. Das medizinische Gutachten im Zivilprozess. In: A. Ehlers (2016), Medizinische Gutachten im Prozess, Anwaltliche Strategie und Taktik beim Umgang mit Sachverständigen, 4th edn, pp. 7–49. Munich: C.H. Beck Medizinrecht. Siegel, A. (2007). 15. Kapitel: Empfehlungen zum Erstellen eines medizinischen Gutachtens – die Sicht des ärtzlichen Experten. In: M. Kuhn and T.  Poledna (2007), Arztrecht in der Praxis, 2nd edn, pp. 783–806. Zurich: Schulthess Juristische Medien. Swiss Academy of Medical Sciences SAMS (2020). Rechtliche Grundlagen im medizinischen Alltag, 3rd edn. Bern: SAMS. Swiss Insurance Medicine (2019). Les expertises médicales en Suisse, 3rd edn. Steinhausen: Swiss Insurance Medicine. Weiss, M. (2016). Beweiswürdigung medizinischer Gutachten im Sozialversicherungsrecht  – kritische Anmerkungen. HAVE 2016, pp. 417–424.

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Forensic Medicine and Human Rights

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grown. Thirty years ago, virtually no forensic pathologist dealt with the subject, but today there is a wide acceptance of the importance of forensic documentation of human rights ­violations. For instance, the subject forms a natural part of international forensic conferences. The UN High Commissioner for Human Rights has relied on forensic documentation for many years, and other international organisations (such as the Organization for Security and Co-­operation in Europe (OSCE) and Organization of American States (OAS)) frequently make use of forensic expertise in human rights violation cases. During the last 10 years, the International Committee of the Red Cross (ICRC) has also made use of forensic expertise in their work. The forensic human rights work today is based on different activities: exhumation and examinations of bodies in singular grave or mass graves (forensic archaeologists, forensic anthropologists, forensic pathologists, forensic odontologists, crime scene technicians and morgue technicians); autopsies on fresh bodies (mainly forensic pathologists and morgue technicians); examination of living torture victims (forensic pathologists and psychiatrists); studies of documents such as medical records, police reports and autopsy reports (forensic pathologists); court appearances, mainly international courts such as the regional human rights courts (the European, the African and the Inter-­ American International Crime Courts, the International Criminal Tribunal for the former Yugoslavia (ICTY), etc.) and last but not the least publication of reports, scientific articles, text books and conducting training courses.

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Forensic pathologists are trained in trauma description, interpretation and report writing. They are used to cases including all kinds of violence, and, therefore, it is natural that forensic expertise is used in the documentation of human rights violations. In post-­World War II society, human rights are based on the United Nations (UN) Universal Declaration of Human Rights from 1948. This declaration was not ratified as a convention until 1984. The majority of UN member countries have ratified the convention, which has been integrated into their legislation, and from 1994, the UN has had a high commissioner for human rights based in the Geneva office. Other conventions have also been passed during the years and are integrated in national legislations. Article 3 of the Declaration of Human Rights states that ‘Everyone has the right to life, liberty and security of person’. Article 5 states that ‘No one shall be subjected to torture or to cruel, inhuman or degrading treatment or punishment’. These are the articles that are relevant for forensic medicine. Forensic documentation of human rights violations started in the 1970s, primarily with the work of Amnesty International’s Danish medical group. Their results were reported, and soon other organisations such as Physicians for Human Rights took up this important work. During the last four decades, hundreds of publications have appeared on documentation of human rights violations, mainly torture. The interest in and importance of the forensic documentation of human rights violations have steadily

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Handbook of Forensic Medicine, Volume 1, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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Mass killings have unfortunately been – and still are – the reality in several parts of the world. Often, the bodies are dumped in graves, single graves or mass graves, and often the graves are disguised. It is important to locate the graves, open them, exhume and examine the bodies (Figure 3.1). There are several reasons why this should be done. First of all, the relatives need to know what has happened to their next of kin. The identification and verification of the cause of death are also important as evidence in criminal and human rights courts, and, finally, documentation of the atrocities is important for the prevention of new human rights abuses.

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The African Court of Justice and Human Rights, previously named the African Court on Human and Peoples’ Rights, was founded by the African Union in 2004. The court is located in Arusha in Tanzania. The jurisdiction of the court is 15 of the African states  – those countries that are member states of the Protocol to the African Charter on Human and Peoples’ Rights. The European Court of Human Rights was established in 1959 by the Council of Europe. The court is located in Strasbourg, France, and its jurisdiction is the 47 member states of the Council of Europe. The Inter-­American Court of Human Rights was founded by the OAS in 1979. The court is based in San José, Costa Rica, and together with the Inter-­ American Commission on Human Rights, it makes up the human rights protection system of the OAS. Its jurisdiction is 21 of the South and Central American states. The United States and Canada have not ratified the convention. There is no human rights court for the Asian or Pacific region, although the question has been raised several times over the years, most recently at an Asia-­Pacific conference of lawyers.

The International Criminal Court was founded by UN in 2002 and is situated in The Hague. One hundred and twenty of the UN members have ratified the statutes of the court. It is a permanent tribunal court to prosecute individuals for genocide, crimes against humanity, war crimes and crimes of aggression. The court opens investigations when it finds it necessary; for instance, in the Democratic Republic of Congo, Uganda, Darfur (Sudan), etc. The court frequently publishes a list of people who have been indicted and among them are both previous and present heads of states. The first verdict was in 2012 when Thomas Lubanga from Congo was sentenced to 14 years imprisonment for forcing children to fight as soldiers.

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Ad hoc tribunals are a known system for dealing with the international prosecution of war crimes, among them genocide. The first tribunal was set up in Nuremberg in 1945 after the Holocaust during the Nazi regime. Tokyo War Crimes Tribunal lasted from 1946 to 1948, where Japanese military and political leaders were charged with war crimes committed in Japanese-­occupied countries during World War II. The ICTY was founded in 1993 in The Hague by the UN after the beginning of the Balkan Wars. The tribunal is scheduled to end in 2014. Several Balkan leaders have been sentenced to long imprisonment, while the cases against the two main characters, Radovan Karadžic and Ratko Mladić, are only in their initial phase. The International Criminal Tribunal for Rwanda is, like the ICTY, under the auspices of the UN and is situated in Arusha, Tanzania. Several of the main perpetrators in the genocide in Rwanda have been sentenced, while others still are awaiting trial in detention. Extraordinary Chambers in the Courts of Cambodia, also known as the Khmer Rouge Tribunal, is a national court established pursuant to an agreement between the Royal Government of Cambodia and the UN to charge senior members of the Khmer Rouge for serious violations of the Cambodian Penal Code and international humanitarian law between 1975 and 1979. The court was founded in 2003 and is situated in Phnom Penh. The end date is still not scheduled.

Figure 3.1  Exhumation of a single grave, the Philippines.

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belongings and document with photographs at all stages of the process, and the morgue technician assists the pathologist. This teamwork has proven very efficient on many occasions in all continents of the world. Fingerprint experts may also play an important role in the identification process if the bodies are sufficiently preserved, and if reliable comparison fingerprints are available. The assistance of a forensic geneticist may also be required to establish the DNA type of the deceased. The forensic anthropologist can reveal gender, age, height, ethnicity and bone trauma. The pathologist can document and interpret bone and soft tissue trauma and other pathology, if soft tissue is present, and establish the cause of death. The forensic odontologist can examine the denture and make a comparison with antemortem data, if they exist, and in that way identify the deceased. The forensic geneticist can compare the DNA of the deceased with possible live family members or with DNA from the deceased’s toothbrush, shaving gear, etc.

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The first challenge is often to locate a grave. Unmarked graves have been found inside cities, in open land and in forests, even in remote mountain areas. Finding the location of a grave often needs assistance from botanists, geologists, archaeologists and advanced technology such as satellite photos and earth radar, which have been helpful in several cases. Once the grave is identified, it must be opened carefully, so that evidence is not destroyed. Many mass graves have been opened with shovels and even excavators, with the inevitable result of broken and co-­mingled bones. Forensic archaeologists and forensic anthropologists play the key role in this part of the work, ensuring that the excavation is done thoroughly and is ­documented in the right way. Thereafter, the bodies are transported to a suitable place where they can be examined thoroughly by the pathologist to establish the cause of death, and by the anthropologist and forensic odontologist to establish the identity (if possible). Crime scene technicians register all the personal

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In 1999, North Atlantic Treaty Organization (NATO) forces threw the Serbian Army out of Kosovo. The UN initiated forensic work immediately after NATO had taken over, and several international teams worked in the country doing exhumations and autopsies, primarily to document the causes of deaths of thousands of people who had been killed by the Serbs, and also to identify the deceased bodies – lying on the ground, in wells and in rivers, buried in the countryside or in cemeteries or lying more or less incinerated in burned houses. Forensic teams were either working as stationary teams in tents converted into primitive autopsy facilities or were working as mobile teams doing open air autopsies (Figure 3.2). The work continued in 2000, but then all the autopsies were performed at a central facility, a previous factory, that was transformed into an autopsy facility during the winter 1999–2000 (Figure 3.3). International teams consisted of experts from more than 30 ­different countries, and in the year 2000 and thereafter, the work was carried out according to the guidelines of the Minnesota Protocol.

Figure 3.2  Open air autopsy on a hot summer day, Kosovo.

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Figure 3.3  Autopsy teams, Kosovo.

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In several cases, forensic genetics also plays an important role in the documentation of human rights abuses. Examples are in Argentina and Chile where many people disappeared during the military regimes in the 1970s and early 1980s. On many occasions, newborn children were taken from detained women and adopted by families of the regimes. The true identities of these children have later been revealed through DNA analyses. The identification of victims from mass graves in Latin America, the Balkans and other regions has later been made using DNA techniques. The International Commission on Missing Persons (ICMP) maintains a DNA laboratory in Sarajevo, Bosnia that processes samples of mortal remains on a large scale. In addition, the Sarajevo laboratory maintains an active programme of technical development and new methods of validation. Interpol has developed an international person identification form, the disaster victim identification (DVI) form, where both antemortem and postmortem information can be registered. The antemortem forms are yellow and the postmortem forms are pink; apart from that the forms are the same. They are divided into a general part (clothing, personal documents, jewellery, etc.), a pathology part (scars, tattoos, etc., plus external and internal traumatology and pathology) and an odontology part. All information can be processed electronically with software especially

developed for the purpose, if needed in incidents with several unidentified bodies.

3.1.4  Forensic observers and inspectors International forensic observers may also be required for autopsies in high-­profile cases, where the death is, or is suspected to be, due to human rights violations. Forensic observers may be present at an autopsy to ensure impartiality and to check the correct procedures are carried out; they are often present as representatives at the request of international organisations such as OAS, OSCE, etc. The situation of the local forensic pathologist can be difficult, as most forensic pathologists are government employees, and in cases of human rights violations, the suspected perpetrators may be representatives of the authorities. The presence of an international observer can therefore be of great help for the local pathologist. Unfortunately, local pathologists can be under severe pressure and therefore reluctant to sign reports opposing the authorities’ view on a case. A sad example is what happened in an African country some years ago, where a well-­known pathologist, who on several occasions witnessed against the authorities, died in a car accident that was never investigated thoroughly. A suspicion of foul play was evident.

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The Colombian Revolutionary Armed Forces, FARC, had kidnapped 12 local politicians from the city of Cali and kept them hostage for 5 years. Suddenly, it was announced that they were dead and buried by the guerrillas in FARC-­occupied territory. FARC said that they were killed by the military, and the army announced that they were killed by FARC. The public demanded that the bodies should be handed over to the families for burial, but FARC would not let the authorities enter their occupied area to exhume the bodies. After long negotiations, the ICRC, together with the Swiss, Spanish and French embassies, managed to get permission from FARC to go in and exhume the bodies. The bodies were buried in a remote jungle area, and the graves were difficult to locate. After several days of intense search and digging, the bodies were finally found and transported to the city of Cali for forensic examination (Figure 3.4). All bodies were autopsied under the strict supervision of an impartial, international expert committee consisting of forensic pathologists, odontologists and anthropologists from Argentina, Canada, Denmark and Portugal. The committee was formed by OAS. All the bodies were identified. They had all died of multiple gunshot wounds, but it was impossible to determine whether they had been shot by FARC or the army. No irregularities were detected in the way the bodies were autopsied or identified.

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Figure 3.4  Body bags with killed hostages, Colombia.

3.1.5  Autopsy protocols Forensic autopsies and preparations of autopsy reports should preferably be done according to the Minnesota Protocol. This

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On some occasions, international experts on inspection missions have been harassed by the local authorities, especially when they have represented non-­governmental organisations (NGOs); for instance, Amnesty International. Case example

A human rights activist was killed in 1996 in his house in Nairobi, Kenya by unknown intruders. Amnesty International was contacted by the NGO for whom the deceased had been working. The autopsy was scheduled for the next day, and Amnesty International was able to find a forensic pathologist who left for Nairobi a few hours later. The next morning the forensic pathologist was taken to the city morgue, where the autopsy was to be performed. The morgue supervisor had been informed by the police that activists would come with a foreign forensic pathologist, and he tried to prevent his presence at the autopsy. The doctor doing the autopsy arrived and was informed about the foreign expert and had no objections to his presence; but midway through the autopsy, the morgue supervisor came with the police and together they virtually threw the foreign export out of the morgue. After some arguing, however, he managed to get in again. After the autopsy, which showed that the cause of death was blunt force to the head caused by a heavy blunt object, the foreign pathologist was interviewed by a newspaper and the next morning there was a front-­page article with a photo, where the pathologist told that he had been harassed. The host organisation did all they could to hide the pathologist, and his embassy had to take several precautions to get him safely out of the country. A few years prior to this incident, a doctor representing Physicians for Human Rights was detained for several days after he had been present at an autopsy where foul play was suspected.

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As forensic pathologists in many countries of the world are also trained in clinical forensic medicine, the examination of living victims of human rights abuses, that is, torture victims, should whenever possible be performed by them. In countries where clinical examinations are not part of forensic pathology work, specially trained clinicians should perform the examination of these persons. However, forensic pathologists and most clinicians are not specialists in psychiatry, and since torture victims almost invariably have mental sequelae, a psychiatric evaluation should be included if possible as a natural part of the examination. Torture is described in more detail later in this chapter. Torture victim examinations can either be done during fact-­ finding missions performed by NGOs, such as Amnesty International and Physicians for Human Rights, or by ­international organisations such as the UN (the High Commissioner of Human Rights Special Rapporteur on Torture), or can be performed by local forensic doctors. Local forensic doctors in countries where torture takes place may be at risk if they examine torture victims, and therefore often need protection or assistance from, for instance, the UN or other international organisations. However, there are unfortunately several accounts of doctors who have done this important work at ­personal risk and have suffered the ultimate consequences. Situations like this can be avoided if the examination is performed by, for example, the Special Rapporteur on Torture, as he or she and their team always visit a country with an official invitation from the government. Luckily, however, several doctors have done – and still do – important torture documentation in less-­democratic countries without suffering major harassment from the authorities.

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Fact-­finding missions may also be used to document large-­scale forced population displacement and forced labour. An example is a series of investigations performed in the border area of Thailand and Burma. The dictatorship in Burma often forced locals to work as slaves on road building and other public projects and as carriers for the army. At the same time, they often force entire village populations to abandon their land and move to more remote areas. As soon as the inhabitants are forced out of their houses, the village is mined to hinder them from returning home. Such mining unfortunately results in many land mine victims (Figure 3.5), and the result of the slavery is severe suffering and often death (Figure 3.6). Hundreds of thousands of people live as refugees in the Western world and many of these people are torture victims. Groups of doctors, such as Physicians for Human Rights in the USA and the Amnesty International medical group in Denmark, have for many years examined torture victims who have fled from their country of origin. This work is important not only for the torture victim (documented torture may improve the possibilities of obtaining asylum) but also for documentation of the atrocities, using publication to induce pressure on both local and ­international authorities. Examination of victims immediately after the torture has taken place is usually difficult, as many authorities keep the victims until bruises, excoriations, etc., have disappeared. However, local trained doctors have over the years documented a substantial amount of information on recent torture victims. Recent torture cases have also been examined during different international fact-­finding missions. Medical groups in the Western countries mostly examine victims with sequelae to the torture. Torture victims should always be examined according to the Istanbul Protocol. This document is the result of work done by an international expert group and was adopted by the UN in 1999. The manual is widely available and is used all over the world, thanks to years of promotion work, including training courses, performed by several NGOs.

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protocol forms part of the ‘Manual on the Effective Prevention and Investigation of Extra-­ legal, Arbitrary and Summary Executions’. The manual was adopted by the UN in 1990 and contains legal, forensic pathology and forensic anthropology chapters. Unfortunately, this manual is not as well known among forensic pathologists as it is in the international courts.

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The political climate in the West African country Togo became unstable early in the 21st century. Human rights had deteriorated considerably, and due to the poor human rights conditions, the European Union (EU) put a large economic support to the country on hold. The government promised to improve the human rights conditions, and the Special Rapporteur was invited to visit the country. In 2008, the rapporteur at that time, Professor Manfred Nowak, and his team, also including a forensic pathologist, visited the country and paid many non-­ announced inspection visits to police stations, detention centres, prisons and military camps. Several violations of human rights conditions, both judicial and medical, were detected, but no case of torture was found. It was obvious that the human rights situation had improved, even though it was far from perfect. Later on, the EU reinstalled economic support to the country.

3.1.7 Forensic evaluation based on documents The need for forensic evaluation in human rights violation cases may arise several years after the person or persons in question are dead, and exhumation and further investigation of the body may be impossible for various reasons (e.g., cremation). However, as forensic pathologists are used to preparing reports based not only on autopsy findings but also including information from medical records and police reports, evaluations can be performed based on documents. This evaluation may of course not be of the same quality as if one had been present at the autopsy, but it can be valuable in court cases, particularly if photographs of the crime scene, exhumation, autopsy, etc., exist. Experienced human rights forensic pathologists are frequently approached by different human rights organisations,

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Figure 3.5  Land mine victim, Burma. He was smuggled out of the country and the photo was taken at a hospital in Thailand.

Figure 3.6  Forced labour, Burma. Villagers are forced to carry heavy weights of cargo over long distances. The painting was made by a survivor.

families of victims or their lawyers and sometimes also by authorities, to evaluate a case. In these cases, as in all other forensic work, impartiality is of the utmost importance, and one should always be careful not to overinterpret findings and statements. It is also important to be aware of the possibility of inaccuracy in the original autopsy report due to

circumstances under which the local pathologists work. Again, most forensic pathologists are government employees, and, in many countries, this is unfortunately often incompatible with total impartiality. Case reviews have on several occasions played an important role in the outcome of human rights ­violation cases.

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Case example

A Danish pathologist and a Portuguese forensic pathologist reviewed a case from Egypt, where a young man was beaten to death by police officers. The officers were finally sentenced to 4 years’ imprisonment. The report from the foreign forensic pathologists played an important role in the outcome of the case. The death of the young man due to police brutality played an important role in the start of the Egyptian Revolution in the beginning of 2011. In a case at the Inter-­American Court of Human Rights in the second half of 2011, the state of Ecuador was accused of negligence and violation of human rights. In 1992, a man was caught by the police during a robbery. He fired at the police, they answered the fire and the man was wounded. The police took him to a hospital where he was observed for some hours and then discharged. The police brought him back to the detention centre, but his situation deteriorated rapidly. He was re-­hospitalised and was kept for several days only treated with intravenous glucose water. He became febrile and unconscious, and was then transferred to a hospital in a larger city, where he was operated on and the surgeons found a perforated intestine and severe peritonitis. The man died a few days after the surgical intervention. The family complained of neglect and human rights abuse from the doctors and the hospital management and filed a case against them. The case went through the national court system and finally ended in the Inter-­American Court of Human Rights in 2011. The court asked two internationally known forensic experts to review the case. At this time there was no body to examine, so the experts were left with documents only. Based on the hospital records, police reports and some witness information, the experts made a report that clearly stated that it was a case of severe medical neglect. The court found the state of Ecuador guilty and sentenced the state to pay a considerable economic compensation to the family, together with an obligation to publish the verdict throughout the country, and to change and strengthen the rules regarding treatment in public hospitals.

In 1991, two young brothers, 14 and 17 years old, were approached by the police in the city of Callao, Peru. The police suspected that they were planning a theft. The boys were taken outside the city in a police car, where they were beaten up and shot through the hands, and finally shot in the head. The family was worried when the boys did not come home for supper and they searched for the boys everywhere, but with no result. Finally, the family received information that the boys could be dead, lying in the morgue. The family went to the morgue and identified the boys. The boys were autopsied, and the cause of death was stated as gunshot wounds to the head. The family took a few photos of the dead boys lying in their coffins prior to the funeral. The family sued the authorities and finally, after several years, the responsible policemen were sentenced to several years in prison. However, the family wanted compensation, and the case went all the way to the Inter-­American Court of Human Rights. Their lawyer, an international human rights defence lawyer, who was working primarily pro bono, asked two Spanish forensic pathologists to review the case. They made a report that was included in the case file. When the case came up in the Inter-­American Court in 2003, the lawyer asked two other doctors, one with many years’ experience in anti-­torture work, and the other, an experienced forensic pathologist, to act as expert witnesses in the court. They travelled to San José and witnessed for the court on the torture aspects and forensic pathology interpretations of the injuries based on the documents of the case and the amateur photos. The state of Peru was found guilty and was sentenced to pay a considerable economic compensation, to publish the verdict and to name a public school after the boys.

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Case examples

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3.1.8  Expert witnesses in international courts

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In contrast to other medical specialists, forensic pathologists are used to appearing in court as expert witnesses. This forms an important part of the forensic pathologist’s work, as he or she here has the possibility of further explaining and underlining the facts of the report in a language understood by non-­ medical professionals. The different international courts vary in their use of forensic expertise in human rights violation cases; for instance, the ICTY only occasionally has used forensic expertise, while forensic experts have often acted as expert witnesses in cases brought before the Inter-­American Court of Human Rights. The special crime court in Cambodia dealing with cases against former Khmer Rouge leaders has only recently considered the idea of consulting forensic expertise. It is also thought-­provoking that many human rights lawyers are unfamiliar with the capabilities and usefulness of forensic expertise in the court room.

3.1.9  Training and research Training is a cornerstone in forensic human rights. Physicians for Human Rights, the International Rehabilitation Council for Torture Victims (IRCT), the Norwegian Medical Association and others have web-­based training programmes. Summer schools, PhD courses, seminars and conferences are arranged by several universities and NGOs. Tuition is on several levels  – from ­grassroots to the highest postgraduate academic level. One training programme that has recently finished in Turkey comprised the teaching of more than 4000 doctors, practising lawyers and judges in the understanding and application of the Istanbul Protocol. This vast training programme was planned and carried out by the IRCT and the Human Rights Foundation of Turkey. Every year, the IRCT arranges Istanbul Protocol training courses in collaboration with their local representatives in different parts of the world, and the Argentine Forensic Anthropology Team (Equipo Argentino de Antropología Forense; EAAF) arranges combined forensic anthropology/forensic pathology/forensic genetics courses in collaboration with local human rights organisations in both Africa and Asia. Written manuals and other teaching materials are produced by several different organisations and are easily accessible via the Internet. The IRCT publishes a free Medline indexed scientific journal, Torture, which publishes a

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Relatively few forensic specialists work with human rights violation cases, and the majority do it part time, as their main occupation is elsewhere; for instance, in universities or state forensic institutions. The work is international, and the experts collaborate or represent different organisations; the need for an international expert network has been felt by many of us for years. In 2009, such a network was formed by IRCT and the University of Copenhagen. Since then, the network has been activated on several occasions and has shown to be effective and rapid. The network has, for instance, already published an operational manual for forensic examination methods by medical teams, has made a statement against hooding that led to the practice now being condemned in Britain and has also intervened effectively in specific human rights abuse cases.

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3.1.10  International expert network

CAT consists of 10 independent experts elected by the member states. All member states are obliged to submit regular reports to CAT on how the rights are being implemented. States must report initially 1 year after acceding to the convention and then every 4 years. CAT examines each report and addresses its concerns and recommendations to the member state(s) in the form of ‘concluding observations’. In addition to the reporting procedure, the convention establishes three other mechanisms through which CAT performs its monitoring functions: (i) CAT may also, under certain circumstances, consider individual complaints or communications from individuals claiming that their rights under the convention have been violated; (ii) undertake inquiries and (iii) consider inter-­ state complaints. According to the convention, torture is absolutely prohibited under all circumstances, both in war and in peacetime. Unfortunately, many of the countries who already have integrated the convention in their national legislation fail to meet these requirements. Even though physical punishment (e.g., whipping) is not included in the torture definition, this act is inconsistent with Article 5 of the UN Declaration of Human Rights. However, several of the UN member countries still practise physical punishment and the death penalty. The Geneva Conventions and their additional protocols are at the core of international humanitarian law, the body of international law that regulates the conduct of armed conflict and seeks to limit its effects. They specifically protect people who are not taking part in the hostilities (civilians, health workers, aid workers) and those who are no longer participating in the hostilities, such as the wounded, the sick, shipwrecked soldiers and prisoners of war. The conventions date back to 1864 when the first was adopted by 12  nations based on an initiative by Henri Dunant, the founder of the ICRC. New protocols were added after international diplomacy conferences during the years on the initiatives of the ICRC and the Swiss government. In 1949, the modern version was signed after yet another international conference in Geneva and the conventions were adopted by the UN. Later, additional protocols have been included which deal with: • The protection of persons deprived of liberty, especially in non-­international armed conflicts. • Mechanisms of control for the respect of international humanitarian law and reparations to victims of violations. • The protection of internally displaced persons. • The protection of the natural environment. The Council of Europe is an international organisation promoting cooperation between all countries in Europe in the areas of legal standards in human rights, democratic development, the rule of law and cultural corporation. It was founded in 1949, and has 47 member states with some 800 million citizens. In 1987, the European Convention for the Prevention of Torture and Inhuman or Degrading Treatment or Punishment was adopted by member states of the Council of Europe. It was subsequently amended by two protocols that entered into force in 2002.

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variety of human-­rights-­related (mainly forensic) articles. There are many activities arranged, but the need for knowledge is immense. Unfortunately, the documentation of human rights violations is still far from complete.

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3.2 Torture

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The UN definition of torture is stated in Article 1 of the Convention Against Torture and Other Cruel, Inhuman or Degrading Treatment or Punishment from 1948. Torture was already mentioned in the Human Rights Declaration from 1948 (Article 5), but with no clear definition. Under the UN Convention Against Torture of 1984, torture involves intentional infliction of pain, by a public official, to obtain information. The full definition of torture is:

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Any act by which severe pain or suffering, whether physical or mental, is intentionally inflicted on a person for such purposes as obtaining from him or a third person information or a confession, punishing him for an act he or a third person has committed or is suspected of having committed, or intimidating or coercing him or a third person, or for any reason based on discrimination of any kind, when such pain or suffering is inflicted by or at the instigation of or with the consent or acquiescence of a public official or other person acting in an official capacity. This definition excludes ‘pain or suffering arising only from, inherent in or incidental to lawful sanctions’, which seems designed to permit the death penalty. The Convention Against Torture has been ratified by most of the UN member countries. To monitor the prevention of torture and other cruel, inhuman or degrading treatment or punishment, the UN has established the Committee Against Torture (CAT).

DUTIES OF FORENSIC MEDICINE IN MODERN SOCIETIES

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The deliberate, systematic or wanton infliction of physical or mental suffering by one or more persons acting alone or on the orders of any authority to force another person to yield information, to make a confession or for any other reason.

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The eight paragraphs of the Tokyo Declaration are listed in the following text, and six of the paragraphs deal directly or ­indirectly with torture. 1. The physician shall not countenance, condone or participate in the practice of torture or other forms of cruel, inhuman or degrading procedures, whatever the offense of which the victim of such procedures is suspected, accused or guilty, and whatever the victim’s beliefs or motives, and in all situations, including armed conflict and civil strife. 2. The physician shall not provide any premises, instruments, substances or knowledge to facilitate the practice of torture or other forms of cruel, inhuman or degrading treatment or to diminish the ability of the victim to resist such treatment. 3. When providing medical assistance to detainees or prisoners who are, or who could later be, under interrogation, physicians should be particularly careful to ensure the confidentiality of all personal medical information. A breach of the Geneva Conventions shall in any case be reported by the physician to relevant authorities. 4. The physician shall not use nor allow to be used, as far as he or she can, medical knowledge or skills, or health information

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specific to individuals, to facilitate or otherwise aid any interrogation, legal or illegal, of those individuals. 5. The physician shall not be present during any procedure during which torture or any other forms of cruel, inhuman or degrading treatment are used or threatened. 6. A physician must have complete clinical independence in deciding upon the care of a person for whom he or she is medically responsible. The physician’s fundamental role is to alleviate the distress of his or her fellow human beings, and no motive, whether personal, collective or political, shall prevail against this higher purpose. 7. Where a prisoner refuses nourishment and is considered by the physician as capable of forming an unimpaired and rational judgement concerning the consequences of such a voluntary refusal of nourishment, he or she shall not be fed artificially. The decision as to the capacity of the prisoner to form such a judgement should be confirmed by at least one other independent physician. The consequences of the refusal of nourishment shall be explained by the physician to the prisoner. 8. The WMA will support, and should encourage the international community, the National Medical Association and fellow physicians to support, the physician and his or her family in the face of threats or reprisals resulting from a refusal to condone the use of torture or other forms of cruel, inhuman or degrading treatment. The definition of torture in the Tokyo Declaration is much wider than the UN definition. It is not restricted to government officials or people acting on behalf of the authorities, and in contrast to the UN definition it does not exclude physical punishment. One has to take into account that the Tokyo Declaration is written by doctors, for doctors, and only the highest ethical standards can be accepted for doctors. The UN Declaration is a legal document binding states and is incorporated into the legislation. Prior to the UN Declaration of Human Rights in 1948, there were no international rules in this field, but prohibition of torture was incorporated into national legislation in many Western countries at the end of the 18th and the beginning of the ­ 19th ­centuries. Torture is well known from history and has been used openly by authorities to provoke confessions and information, and/or as part of punishment. The Christian Church played an active role in torture for about 600 years from the 13th to the 19th century. The most widely known are the Spanish and Portuguese Inquisitions. Not only the Catholic Church but also the different Protestant churches frequently applied torture, usually in collaboration with the secular authorities. Witch-­hunting followed by summary legal proceedings – including torture-­provoked confessions and ending with burning in public of the ‘guilty’ while still alive – was not uncommon in Europe for several centuries. Torture disappeared at least officially in the Western world during the 18th and 19th centuries, but reappeared during the totalitarian regimes in Russia and Germany in the 20th century. Today, torture is prohibited in the Western world and is rare, although not completely absent. In many other parts of the world, torture is unfortunately part of the interrogation procedures used

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The European Convention for the Prevention of Torture and Inhuman or Degrading Treatment or Punishment or, more shortly, the Committee for the Prevention of Torture (CPT) is the anti-­torture committee of the Council of Europe. The CPT visits places of detention in the member states. The visits are unannounced and are carried out by small teams of CPT members, who usually call in additional experts, such as forensic doctors. After each visit a report about the findings and recommendations is drawn up and sent to the respective governments. After 20 years of experience, this European model was adapted and generalised by the UN through the Optional Protocol to the Convention Against Torture (OPCAT) in 2006. So far, 60 UN member countries have ratified the protocol, and a further 22 countries have signed but not yet ratified it. OPCAT, which entered into force in June 2006, has created the Subcommittee on Prevention of Torture (SPT). The SPT has a mandate to visit places where persons are deprived of their liberty in the membership countries. Under the OPCAT, membership countries should establish independent national preventive mechanisms for the prevention of torture at the domestic level. The World Medical Association (WMA) has a set of international guidelines for physicians concerning torture and other cruel, inhuman or degrading treatment or punishment in relation to detention and imprisonment – the Tokyo Declaration. It was adopted in 1975. According to the declaration, torture is defined as:

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Blunt force injuries Many reports show that blunt force injury, such as beating and kicking, is a part of almost all torture sessions (Table 3.1). Many torture victims have reported that blunt force injury has been

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Table 3.1  Torture methods reported by 235 torture victims from 35 different countries. Percentages exceed 100, as almost all victims had been tortured in many ways.

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Many different torture methods have been reported, both physical and psychological. Psychological torture methods will just be briefly discussed, as forensic pathology is focused on signs of physical torture.

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3.2.1 Torture methods

used as the ‘introduction’ to the real torture sessions with more sophisticated methods, while other torture sessions only include blunt violence. Blunt violence is mostly applied on the body, but also often against the arms, legs, head and face. The violence is often kicking with boots or hitting with fists, especially in the face. Different forms of instruments are also often used. A police truncheon is an effective weapon, which can cause severe damage. Instead of truncheons, which can be of different lengths and can be handled with one or two hands, the police in some countries also use long sticks such as lathis in Nepal and India. The effects of this torture include bruising and swelling, and fractures of the arms, fingers, ribs and skull. Fractures of the facial bones and teeth are also not infrequent. Blunt violence to the head can cause intracranial haemorrhages. Kicking and stamping on the body can lead to lacerations of internal organs, pneumothorax and fatal haemorrhage. A special kind of blunt force torture is falanga, also called falaka or bastinado, which is beating of the foot soles with truncheons, sticks, metal wires or other elongated instruments. The victim is usually lying down, either prone or supine, during the torture session. Falanga is not only painful to the feet, but the pain often spreads to the rest of the body and the head. The feet swell and if the beating has been intense and prolonged, the skin of the foot sole may burst. Not infrequently the torture victim is forced to jump up and down on bare feet or walk on uneven surfaces, stones, etc., after the falanga torture, which is extremely painful. This torture method has been used in the Middle East for centuries, but has also spread to both Europe (Greece, in the junta regime in the 1960s and Spain in the Franco period) and Asia (India, Nepal) and even to Central and South America, although reports of falanga are scarce from that part of the world. Whipping has been an integrated part of torture and punishment for thousands of years. It is well described from most of the civilisations in Asia, Africa and Europe. In Europe, it was used by both the Greeks and the Romans and later by both the Inquisition and secular authorities. Whipping was also a frequently used punishment for slaves in America. Today, whipping is still accepted as punishment in several Middle East countries, and also in some Asian and African countries (Figure 3.7). Either a whip or a cane is used for the whipping sessions. The victim is usually tied to a pole and is then whipped on the naked back. Whipping is a torture method that may be combined with the application of salt, pepper and chilli in the wounds afterwards. Whipping causes intense pain, and prolonged and fierce whipping may cause death. The wounds often become infected and may lead to death.

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by the police and other authorities. The yearly reports by Amnesty International clearly document this sad fact. In almost all countries where torture takes place, it is not officially admitted by the authorities, as they have incorporated both the UN Declaration of Human Rights and conventions against torture in their legislation. It seems as though the most important ‘reason’ for torture today is not to obtain information or to punish, but to break down the mind and personality of persons who represent opposition and therefore are a threat to non-­democratic regimes.

Reported use

Blunt trauma (beatings, including falanga, kicking, whipping)

99%

Positional torture (suspension, streaking of limbs, forced positioning)

53%

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Torture method

34%

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Burns (with cigarettes, heated instruments, hot or caustic liquids) Electrical torture

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24% 21%

Asphyxiation

18%

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Penetrating injuries (stab and gunshot wounds)

Sexual violence

18%

Crush injuries (crushing fingers or heavy rollers to injure thighs or back)

17%

Chemical exposure (salt, chilli pepper, gasoline in wounds or body orifices)

11%

Poor detention conditions (cold, hot, solitary confinement, forced nudity)

9%

Sensory deprivation (sound, light, sense of time, sleep restriction)

8%

Near-­drowning A torture method that was frequently used in Latin America under the military regimes in the 1960s, 1970s and 1980s was the submarino (submarine). There were two versions of this torture, the wet and the dry. The wet submarino, also called ‘la bañera’, is where water is used. The water is often polluted with urine, faeces or vomit and the victim’s head held under water

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Electrical torture

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This torture method has been, and unfortunately still is, widespread. Electrodes are usually placed on sensitive parts of the body, such as the genitalia, anus, nipples, tongue and lips. Several torture victims have reported that equipment with variable amperes has been used. Electric cattle prods have also been used on many occasions as torture instruments. In a more recent and infamous case, the crown prince of Abu Dhabi tortured a previous business associate by inserting an electric cattle prod in the victim’s anus. A variety of electric shock devices or weapons are available on the market, such as taser guns, electric shock batons or stun belts. Several police forces use these devices, and they are unfortunately also frequently used as torture instruments. CAT reports that taser use can be a form of torture due to the acute pain caused and warns about the possibility of death in some cases. The use of stun belts has been condemned by Amnesty International as torture, not only for the physical pain the device causes, but also for their heightened abuse potential. Electric shock causes immediate pain, but can also cause cardiac arrest due to universal depolarisation.

‘Telefono’

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‘Telefono’ is the Spanish word for telephone and the technique consists of simultaneous slapping of both ears. This results in acute, severe earache and headache and usually tympanic rupture. The method was frequently used in Latin America, Greece and Spain during the military regimes.

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until nearly reaching the point of drowning. This is usually repeated several times. The other form is the dry submarino, where a plastic bag is placed over the head and held firmly to the neck, so no air can enter. The plastic bag is then kept over the victim’s head until the point of choking, then torn off and often replaced again several times. Both wet and dry submarino techniques were also commonly used under the Franco regime in Spain and the methods have also been reported from other parts of the world. Waterboarding is a form of torture in which water is poured over the face of an immobilised captive, thus causing the individual to experience the sensation of drowning. Waterboarding, like submarino, can cause extreme pain, damage to the lungs, brain damage from oxygen deprivation, other physical injuries including broken bones due to struggling against restraints, ­lasting psychological damage and death. The term waterboard torture appeared in press reports as early as 1976 from Cambodia. Although a variety of specific techniques are used in waterboarding, the victim’s face is usually covered with cloth or some other thin material, and the subject is immobilised on his/her back. Water is then poured onto the face over the breathing passages, causing an almost immediate gag reflex and creating the sensation of drowning. In 2007, it was reported that the Central Intelligence Agency (CIA) was using waterboarding on extrajudicial prisoners and that the Department of Justice had authorised the procedure, even though the United States government hanged Japanese soldiers for waterboarding US prisoners of war in World War II. The CIA confirms using waterboarding on three Al-­Qaeda suspects in 2002 and 2003. During the presidency of George W. Bush, US government officials at various times said they did not believe waterboarding to be a form of torture.

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Figure 3.7  Whipping, Togo. The victim was accused of the theft of two chickens and was whipped to provoke a confession.

Burning Burning, especially with cigarettes, is a widespread torture method usually applied in combination with other torture methods. Apart from the pain, the burns can get infected, and large burns may be lethal.

Finger torture Fingers are often subjected to torture. One method is to place, for instance, a pencil between the fingers and press them together. This is very painful, as the sensitive periost of the phalanges is affected. The nails can also be the target of torture by pressing a sharp object up under the nail or pulling the nail out. Fingers can also be broken or crushed – the Chilean singer and guitar player Víctor Jara had his fingers and wrists broken so that he was no longer able to play the guitar; soon after he was killed by the military.

Suspension torture Different forms of suspension as part of the torture have been – and still are –widely used. The victims are suspended by the hands or head down by the feet. A variant of suspension is the ‘Palestinian hanging’ or ‘strappado’, where the victim’s hands are

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One of the witnesses in the trail against Saddam Hussein told that he was arrested in Dujail following an assassination attempt against Saddam and later taken to the Baghdad headquarters of the Mukhabarat intelligence agency. Interrogators forced him to strip and then hung him from his feet before beating him with hoses and applying electric shocks to him, including ‘sensitive parts’ of his body, the man testified. At one point, Saddam’s half-­ brother and former intelligence chief Barzan Ibrahim joined the interrogation along with two men in civilian clothes. Ibrahim asked one of the men to light a cigarette for him, and he then put it out on the witness’s head, the court heard.

There are many forms of positional torture, all of which tie or restrain the victim in contorted, hyperextended or other unnatural positions, which cause severe pain and may produce injuries to ligaments, ­tendons, nerves and blood vessels. Characteristically, these forms of torture leave few, if any, external marks or radiological findings, despite subsequent frequently severe chronic disability. Positional torture is directed towards the tendons, joints and muscles. The ‘five-­point tie’ is a technique of trussing up a captive used in several African countries. A rope is tied round the wrists, ankles and neck or mouth, holding the trunk tightly in extreme extension. An attempt to relieve the pain by moving one limb tightens it more around the others. In China, many forms of shackling are used as punishment and are given nicknames to disguise the appallingly painful methods used. For instance, Su Qin bei jian (literally ‘Su Qin carries a sword on his back’) describes the shackling of one arm pulled over the shoulder to the other which is twisted behind the back. Another is liankao, describing various methods of shackling the hands and feet behind the back. Cramped or distorted postures and prolonged standing are used routinely in many countries. An example is in Israel where ‘moderate physical pressure’ is permitted by law. Several techniques have been devised by the General Security Service (Shin Bet) and are routinely used to put detainees under undue stress. In shabeh, the victim is shackled for hours to a low chair whose front legs have been shortened so that the detainee must constantly struggle to avoid sliding off. Shabeh is usually combined with methods of abuse, such as placing an often filthy sack over the victim’s head, exposing him or her to loud music and sometimes to extreme temperatures and sleep deprivation. In gambaz, the detainee is forced to crouch on his toes in the ‘frog’ position for long periods. In kas’at tawila, the subject is made to kneel with his back up against a table and his cuffed arms resting on the table behind him while the interrogator’s legs push against his shoulders. A small chamber nicknamed the ‘refrigerator’ is used to keep the victim immobile for hours or days. The use of coffin-­like boxes, where victims have been forced to stand up for several days, has been reported from the Turkish Kurdistan. CAT has determined that restraining detainees in painful positions is by itself an act of both torture and cruel, inhuman or degrading treatment. It recently determined that the use of ‘short shackling’ by US personnel constitutes either torture or cruel, inhuman or degrading treatment and has recommended that the method be prohibited.

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Case example

Positional torture

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tied behind the back before the victim is suspended by the hands. Weights may be added to the body to intensify the effect and increase the pain. The name ‘Palestinian hanging’ is most probably a misnomer, since it is neither a Palestinian nor an Israeli invention, nor used more frequently in that region than in other parts of the world. The suspension method seems to have been widely used in Europe up to the beginning of the 19th century; it was also one of many methods of punishment used in German concentration camps during the Nazi regime. The Khmer Rouge regime of Cambodia in the 1970s also used this variant of suspension as part of the torture. Suspension is usually combined with blunt force violence, burning, chemical torture (acid or caustic) or electric torture.

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Another variation of suspension is ‘crucifixion’, where the victim is tied with spread arms to a horizontal pole or a cross. The parrot’s perch, also called ‘Pau de arara’, refers to a physical torture technique designed to cause severe joint and muscle pain (to the wrists, ankles, legs and knees), as well as headaches. The position also causes congestion of the head with the risk of brain damage and may also impede respiration and circulation, causing death. The technique consists of a tube, bar or pole placed over the victim’s biceps and behind the knees while tying the victim’s ankles and wrists together. The entire assembly is suspended between two metal platforms forming what looks like a parrot’s perch. This technique is believed to originate from Portuguese slave traders, who used it as a form of punishment for disobedient slaves. Its use was widespread by agents of the political police of the Brazilian military dictatorship against political dissidents in the 1960s and 1970s, and it is still believed to be in use by Brazilian police forces, although outlawed. The device was often used as a restraint for other torture techniques, such as waterboarding, nail pulling, branding, electric shocks and sexual torture. Suspension causes severe pain to the region that has been tied, but depending on the method of suspension, it may also cause severe pain of, for instance, the shoulder joints as well as causing ruptures of tendons and luxation of shoulder joints. ‘Palestinian hanging’ may also cause fractures of the scapular part of the shoulder joint and may impede respiration. Suspension with the head down may cause brain damage due to congestion and may lead to death due to circulatory collapse.

3.2.2 Sequelae to torture Most examinations of torture victims are performed months to years after the torture has taken place. All wounds are healed, and the physical signs that can be detected are scars, neurological symptoms, broken or missing teeth, musculoskeletal and visual or hearing impairment, etc. Many torture methods leave no scars or other physical sequelae and are therefore not possible to document

DUTIES OF FORENSIC MEDICINE IN MODERN SOCIETIES

Burning

Case example

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Scars after burning may show keloid formation and contractions may be present. The scars may be either hyper-­or hypo-­ pigmented, and if on the head, alopecia is often prominent. Pattern scars may be present if cigarettes, irons or other hot objects with a typical appearance have been used in the torture. Usually, the scars have a non-­specific appearance.

A 23-­year-­old Turkish Kurd was arrested for deserting from military service. He was repeatedly tortured for about 4 weeks. He was tied to a cross that was raised so that his feet could not touch the ground. In this position, he was repeatedly beaten with truncheons, and electrodes were placed on his penis and his chest, and he received several electric shocks. Every time he fainted, cold water was poured over him. His testicles were squeezed several times, and he was forced to eat faeces and drink urine. Several times he suffered near-­drowning: his head was pressed down into the toilet until the point of drowning. He was also forced to stand on one leg, and at other times, he was forced to be on his knees with his hands tied on his back to his wrists. He was forced to remain in this position for many hours. He was also threatened several times with sexual torture, and he witnessed family members being tortured in front of him. He was sentenced to 17 years’ imprisonment. In the prison, he was often beaten with sticks until he fainted, and was burned with cigarettes on his arms and body when he woke up again. After some months, he was transferred to another prison, where he was not tortured physically but was placed in solitary confinement for 2 years. After that, he was released and went underground until he was able to leave the country. At the examination 3 years later, he had several non-­specific scars on the body and also circular scars with the diameter of cigarettes on both his chest and back (Figure 3.8). He suffered from severe PTSD.

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Blunt force, especially against the head, may lead to more or less permanent headache and visual and hearing impairment. Peripheral nerve damage may cause permanent neurological symptoms, and incorrectly healed fractures may cause chronic pain and impaired movement of the limbs. X-­ray may be valuable to verify healed fractures. Due to the proximity between the plantar aponeurosis and the bones of the foot, the compartment with nerves and vessels is small and the aponeurosis is so inflexible and tightly attached that there is very little space for expansion of the compartment. If a person is subjected to falanga torture, the result is swelling due to oedema and haemorrhages. If the falanga torture is severe and prolonged, both nerves and vessels in the area may be injured and become necrotic due to the increased pressure caused by the swelling. The fatty tissue in the foot sole plays an important role as shock absorber and this tissue may necrotise as well. The result is a foot sole with nerve damage and necrosis. In severe cases, the impaired circulation can lead to gangrene of the toes and even to the entire forefoot. Pain in the foot soles propagating to the legs is not uncommon among falanga victims, and in severe cases, the torture victims can never walk again. Whipping leaves striped scars. They may in some cases have a typical hyper-­pigmented tramline appearance that can be seen in fresh lesions. This appearance is due to the fact that the whipping caused the rupture of small vessels at the edge of the lesions and pigment remains in the skin afterwards. The distance between the parallel tramlines determines the diameter of the instrument used for whipping. The scars are often without the tramlines and are either hyper-­or hypo-­pigmented. Africans not infrequently show keloid formation of the scars.

Electrical torture usually leaves no or very faint marks, and sequelae are therefore impossible or difficult to detect. If any scars are detected, they are small and discreet, and usually found on the genitalia or in the mouth.

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somatically. The scars are often non-­specific and are therefore not sufficient per se to document torture. The torture history is therefore often more important than the physical findings.

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A 42-­year-­old Sikh from India was detained by the police after a religious ceremony. He was detained for 2 weeks, where he was tortured several times. He was hit all over the body with lathis and ropes. He was also forced to lie down and was tied with a belt and submitted to falanga with lathis. He was tied around the ankles and hung with the head down for a long time, while he received kicks and beatings with lathis. He was released, but arrested again half a year later and detained for 10 days. During this detention, he was again beaten and submitted to falanga, suspended with the head down, and burned on his back with a hot iron bar. On one occasion, chilli powder was inserted into his anus. After the 10 days, the family managed to bribe a warden and he was released and smuggled out of the country. At the examination 1 year after the last torture session, he had several irregular scars scattered around the body and limbs and several 10–15-­cm-­long, 1–2-­cm-­wide hyper-­pigmented scars on his back. He also had discreet linear scars around the ankles. Both foot soles were flattened and he had distinct pressure-­ provoked pains. He had clear symptoms consistent with posttraumatic stress disorder (PTSD).

Figure 3.8  Close-­up of scars after cigarette burns.

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FORENSIC MEDICINE AND HUMAN RIGHTS

3.2.4  Examination of torture victims

Psychological sequelae to torture are usually not difficult to detect by experienced psychiatrists. The symptoms may be so prominent that they are also obvious to other doctors. A vast majority of torture victims suffer from PTSD. Torture victims re-­experience the torture sessions through flashbacks and nightmares. They avoid stimuli associated with torture. They easily become angry and often suffer from sexual dysfunctions. They find it difficult to concentrate and their memory is bad. The PTSD usually develops a long time after the torture, and the symptoms are permanent without proper treatment. PTSD is not torture-­specific, but is a psychological reaction to severe stress. This is important to know when a doctor examines torture victims who also are refugees or asylum seekers in other country. These people have been exposed to extreme stress, and many of them suffer from PTSD due to their situation. However, studies have shown that refugees who have been tortured have more severe mental symptoms than non-­tortured refugees from the same countries.

The examination should follow the guidelines of the Istanbul Protocol. The protocol is an official UN document and is therefore accepted by the majority of UN member states. Medical evaluations of torture may be useful evidence in legal contexts for various reasons: (i) it may play a role in the identification of a perpetrator responsible for torture; (ii) it may support a political asylum application; (iii) it may establish conditions under which false confessions have been obtained by state officials and (iv) it may establish regional practices of torture. Medical evaluations may also be used to identify the therapeutic needs of the torture survivors, and serve as a testimony in human rights investigations. One should always be conscience that torture victims most probably suffer from PTSD, and it may be very difficult for him or her to talk about the torture. It is therefore very important for the doctor to obtain the torture victim’s confidence. Often an interpreter is needed and here again confidence is important. It is not enough that the torture victim and the interpreter understand each other. Several interview situations have been made impossible due to the fact that the interpreter and the torture victim have represented warring factions in internal or regional conflicts. It is also advisable for the doctor not to wear a white coat, as some torture victims have experienced doctors or other health personnel monitoring the torture. Invasive examination procedures should be kept to an absolute minimum, as many torture victims are extremely sensitive to pain. The anamnesis has to be thorough, as physical sequelae are often absent or very discreet. An examination of a torture victim is a time-­consuming procedure, and is usually a very stressful situation for the victim. It is important to have access to psychological or psychiatric assistance, as there is a considerable risk of re-­traumatisation of the torture victim. A psychiatric evaluation of the torture victim is a necessary part of the examination, and the forensic doctor should incorporate the results in the final report. A trained forensic doctor following the guidelines of the Istanbul Protocol will usually know if there is consistency between the reported torture and the findings at the examination. It is also important for the examining doctor to have as much knowledge as possible of the frequently used torture methods in the region. As in many other forensic reports, firm conclusions may not be reached, but it is important to state the degree of consistency between the anamnesis and the objective physical and psychological findings. It is also important to recognise that no findings are not necessarily the same as no torture.

Case example

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A 23-­year-­old Syrian Kurd was arrested by the police when he participated in a demonstration against the authorities. He was detained for 7 months without being brought before a judge. During the detention, he was tortured on several occasions. He was kicked and punched all over the body, and he was hit on the legs with truncheons. On several occasions, he was undressed and hit all over with belts. On other occasions, he was forced into a tyre that was rolled across a courtyard, while he was being kicked and hit with truncheons. He was blindfolded during most of the torture sessions and was kept in solitary confinement for most of the detention period. At the examination 4 years later, no scars were detected apart from one on his knee, which was not related to the torture. However, he had severe symptoms consistent with PTSD.

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3.2.3  Psychological torture

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Psychological torture is often combined with physical torture or may be the only torture form. A common psychological torture form is blindfolding; other forms are threats, mock executions, forcing the victim to watch the torture of family members or friends, forcing torture victims to torture others and humiliation (e.g., forced nudity), which are common in several parts of the world. Exposure to light (e.g., flickering lights or intense light 24 hours a day) and sleep deprivation are well-­documented torture methods. Solitary confinement is also considered by many as torture. In 2008, the Special Rapporteur on Torture concluded that ­‘prolonged isolation of detainees may amount to cruel, inhumane or degrading treatment or punishment and, in certain instances, may amount to torture’. Solitary confinement may be necessary in short pre-­trial periods to avoid influence on the police investigation. However, several countries, including some EU countries, seem to underestimate the psychological consequences of solitary confinement.

3.2.5  Treatment of torture victims Treatment of illnesses is not the work of forensic doctors. It is, however, important for forensic doctors to know that there are treatment possibilities, and it is the obligation of a forensic ­doctor – as it is for other medical personnel – to refer a patient to other specialists when necessary. The treatment of torture victims

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has been developed during the last 30 years. The first rehabilitation centre for torture victims was founded in Copenhagen in 1992. Since then, many rehabilitation centres have been founded throughout the world and an extensive international collaborative network enhances further evolution of the field. Unfortunately, the need for treatment by far exceeds the capacity of the centres.

Useful websites

International Rehabilitation Council for Toture Victims, http://www.irct. org. Istanbul Protocol, http://www2.ohchr.org/english/about/publications/ docs/8istprot.pdf. Office of the High Commissioner for Human Rights, http://www.ohchr. org. Physicians for Human Rights, http://physiciansforhumanrights.org/. The Advocates for Human Rights, http://www.theadvocatesforhuman rights.org/4Jun20046.html#III. (All last accessed 18 December 2012.)

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Council of Europe, http://www.coe.int. International Committee of the Red Cross, http://www.icrc.org/.

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International Guidelines and Accreditation in Forensic Medicine

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Peter Wiegand, Burkhard Madea and Frank Mußhoff

required for different fields of law, including criminal, civil, work, family and administrative. Its core activities are clinical forensic medicine and forensic pathology, but other areas of science and expertise including forensic toxicology, forensic psychiatry, forensic genetics and forensic anthropology may be required depending on the nature of the case.

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In general, forensic science refers to the examination of scenes of crime, recovery of evidence, laboratory examinations, interpretation of findings and presentation of the conclusions for intelligence purposes or for use in court (ILAC 2002). Activities range from instrumental analysis such as of blood alcohol and drugs, DNA typing of items at scenes of crime, identification and paternity testing to the forensic investigation of the (injured) human body (dead or alive) based on clinical examination or different pathological methods (autopsy, histology, etc.). Laboratory investigations in forensic science involve the examination of a wide range of items and substances using the highest analytical standards in compliance with the requirements of ISO/IEC 17025, as interpreted for forensic science laboratories (ISO/IEC 1998, 2005). One aim of the European Council of Legal Medicine (ECLM), founded in Cologne in 1993, is the recognition of legal and forensic medicine as a medical mono-­speciality, and, thus, as one of the guardians of the rights of the most vulnerable persons. The European Council of Legal Medicine shall be the official body dealing with matters relating to the discipline of legal medicine in Europe. The ECLM shall deal with all scientific, educational and professional matters pertaining to this discipline on a European level. According to documents published by the ECLM, legal and forensic medicine is the application of medical knowledge and methodology for the resolution of legal questions and problems for individuals in society. It involves the observation, documentation, collection, assessment and scientific interpretation of medical evidence deriving from clinical and postmortem investigations

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Handbook of Forensic Medicine, Volume 1, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

4.1.1  Autopsy standards in forensic medicine The creation of autopsy standards as a measure of quality control of medicolegal autopsies dates back to the 19th century. For example, in 1855, a decree was released in Austria on postmortem medicolegal external examinations which is still in force today. This very detailed decree comprises 134 paragraphs and consists of four major parts: 1. General remarks on the postmortem medicolegal examination (paragraphs 1–46). 2. Description of the medicolegal examination (paragraphs 47–97). 3. Postmortem examination in cases of suspected intoxication (paragraphs 98–111). 4. Postmortem examination in newborns (paragraphs 112–134). The second main part contains detailed descriptions on the postmortem dissection of the body and all organs (paragraphs 58–97). Similar decrees were also released in other German states, such as Prussia; the Prussian autopsy regulation is quite famous and is edited by the pathologist Rudolf Virchow, giving standard autopsy protocols (Figure 4.1). All these national autopsy regulations contained detailed descriptions of how to carry out an

DUTIES OF FORENSIC MEDICINE IN MODERN SOCIETIES

6. Autopsy procedures. 7. Autopsy report. 8. Specific procedures.

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Although the document is a ‘recommendation’ by nature, it has legal implications because the Council of Europe member countries have to implement these principles in their national legislation (Saukko and Knight 2004). However, in many countries, the legal regulations have not yet been changed in line with these recommendations. One of the suggested rules is that an autopsy should be carried out in all obvious or suspected unnatural deaths, even when there is a delay between causative events and death, in particular: • Homicide or suspected homicide. • Sudden unexpected death including sudden infant death. • Violence of human rights such as suspicion of torture or any other form of ill treatment. • Suicide or suspected suicide. • Suspected medical malpractice. • Accidents, whether transportational, occupational or domestic. • Occupational diseases and hazards. • Technological or environmental disasters.

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autopsy, which instruments should be used, what had to be retained for further analysis and which terms should be used in the autopsy protocol. In order to maintain quality control and quality assurance, theses autopsy protocols were also reviewed by juries. Today we have national guidelines for autopsy standards in many countries, and to improve international cooperation, medicolegal autopsy rules have been standardised; for example, in 1991, the General Assembly of the United Nations endorsed the model autopsy protocol of the United Nations. In Europe, Recommendation No. R(99)3 on the harmonisation of medicolegal autopsy rules was adopted by the Committee of Ministers on 2 February 1999 (Council of Europe  1999, cited in Byard and Winskog  2012). This recommendation has meanwhile been updated by the ECLM 2014 in Dubai. Similar to the old autopsy regulations, this European recommendation consists of several parts: 1. Principles and rules relating to medicolegal autopsy procedures. 2. Scene investigation. 3. Autopsy physicians. 4. Identification. 5. General considerations.

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Figure 4.1  Rudolf Virchow’s (1821–1902) famous autopsy technique in the morgue of the Charité hospital with special regard of forensic medical practice.

International Guidelines and Accreditation in Forensic Medicine

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tion is carried out at the local medical council by two experts in forensic medicine and one physician of the medical chambers. The oral exam lasts at least half an hour, after which the physician and the board of examiners inspect the written reports the candidate presented at the examination. The guidelines for anatomical pathology residency training in forensic pathology, endorsed by the National Association of Medical Examiners (NAME) in the USA, recommend that during training each anatomical pathology resident should receive training in forensic pathology. During this training, it is recommended that each resident will: • assist in, or perform under direct forensic pathologist supervision, medicolegal autopsies; • learn the general principles of autopsy and biosafety; • understand the statutory basis for the medicolegal death investigation systems, and the requirements to serve as medical examiner or coroner or forensic pathologist; • learn how the medicolegal death investigation system interacts with the criminal and civil legal systems and public health and safety agencies; • learn the three core elements of a medicolegal autopsy: scene/ death investigation, autopsy and toxicology; • learn to recognise the common injury patterns seen in blunt force trauma, sharp force injury, firearms injury, motor vehicle fatalities, asphyxial injuries, temperature and electrical injuries and suspected child and elder abuse; • learn to recognise common postmortem changes, including decomposition patterns; • understand the causes and autopsy findings in cases of drug-­ related and toxin-­related fatalities; • understand the causes and autopsy findings in cases of sudden, unexpected and natural death; • understand the importance of proper documentation in medicolegal autopsies; • understand the concept of evidence recognition, collection, preservation, transport, storage, analysis and chain of custody; • understand the basic disciplines of forensic science and their relevance to death investigation systems; • understand the concept and application of clinical forensic medicine; • learn the importance of professional interaction with families and the public; • understand the proper method of death certification; • understand and correctly apply the terms ‘cause’, ‘manner’ and ‘mechanism’ of death. These recommendations of course only provide a minimum working knowledge of forensic pathology. It is particularly important that the training takes place in an accredited anatomical pathology residency training programme. Training programmes, whole institutes of forensic medicine or laboratory branches within one institute may be accredited. Accreditation is based on the recommendations of the national societies on various topics.

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activity. • Unidentified or skeletonised bodies. Of course, medicolegal experts must exercise their functions with total independence and impartiality. They should not be subject to any form of pressure and should be objective, in particular, in the presentation of their results and conclusions. Whenever possible, medicolegal autopsies should be carried out by two physicians of whom at least one should be qualified in forensic pathology. The different stages in the determination of cause and manner of death can be summarised as follows: 1. External examination. 2. Autopsy. 3. Supplementary investigations such as histology, immunohistochemistry, toxicology, postmortem biochemistry and molecular pathology. The external examination has the lowest evidential value, with much more detail emerging from the autopsy. Nevertheless, also concerning the external examination and filling out the death certificate, very important recommendations have been published for instance by the College of American Pathologists. Histology is a routine part of postmortem examinations not only where the cause of death remains unclear based on macroscopic findings but also in cases where the causes of death are known. Recommendation No. R(99)3 therefore includes regulations covering which parts of organs and body fluids should be retained for histology and subsequent investigations. The quality of an autopsy and its protocol depend not only on international recommendations but also on the qualifications of the doctors who perform the autopsy. The postgraduate education and qualifications in forensic medicine or forensic pathology vary from country to country. The concept of forensic medicine is common in the European continent, where physicians are trained not only in forensic pathology but also in ­clinical examinations, toxicology, etc., whilst in the UK, the United States and elsewhere, the concept of forensic pathology is predominant. For example, in Germany, the postgraduate specialisation in forensic medicine takes at least 5 years according to the teaching regulations of the medical chambers. Of these 60 months, six are spent in clinical pathology, six in psychiatry or forensic psychiatry and six further months can be spent either in pathology or public health, pharmacology, toxicology or p ­ sychiatry. The remaining three and a half years have to be spent in forensic medicine; at least 400 complete external examinations of bodies with detailed descriptions have to be carried out, 25 crime scene investigations have to be done and a minimum of 300 forensic autopsies have to be performed with special emphasis on the relation between morphological findings and ­traumatic mechanisms. Furthermore, 2000 histological investigations are mandatory, 200 cases of oral or written reports for court have to be prepared, at least 10 cases of stains have to be analysed and 25 forensic osteological and odontological investigations are required. Once a postgraduate trainee has completed his or her further education, an examina-

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• Identification of representative samples and the selection of minimum amounts of material required to obtain meaningful results for interpretative purposes. • Positive and negative controls. • Guidance on methods for sampling that aid/assure the prevention of cross-­contamination. • Preservation/storage of relevant material for subsequent analysis.

Labelling and documentation

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To fulfil the chain of custody record, it is essential to know who has handled which item and what he/she did with it. The laboratory should describe all items and evidential material. Labels should be attached to each item at the time of packing. Whilst the legal status and use of labels can vary, the minimum details should be recorded and directly and unequivocally attributed to each package (ENFSI 2008).

4.1.3 Laboratory standards in forensic medicine: quality management in forensic DNA analysis and forensic toxicology

Personnel

The laboratory should define the standards of competence required for individuals to undertake particular responsibilities, the training required and the assessments that are needed. For each area of responsibility, therefore, the laboratory should specify requirements under the headings of qualifications and experience, competencies, training and assessment and maintenance of competency, documented on the individual’s training records (e.g., practical tests, oral examinations, casework supervision) (ENFSI 2008, 2016).

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Each laboratory needs to write a handbook on forensic quality management that describes the basic laboratory procedures in accordance with ISO/IEC 17025 (ISO/IEC 1998, 2005, 2017). Minimum requirements are: the aims and scope, organisation and management, personnel qualification and training, facilities and equipment, validation and controls, analytical procedures, calibration and maintenance, proficiency testing, corrective action, documentation and reports, internal and external audits and safety (Butler 2001; GTFCh 2004, 2006a, 2006b, 2009a, 2009 b; European Network of Forensic Science Institutes (ENFSI) 2008, 2016). The different analytical laboratory processes are documented in standard operation procedures (SOPs), including all the necessary information to achieve correct and reproducible results (e.g., DNA extraction, polymerase chain reaction (PCR) analysis based on different short-­tandem-­repeat (STR) kits).

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In order to achieve the recognition of legal and forensic medicine as a medical mono-­speciality in Europe, the European Council of Legal Medicine has meanwhile published further recommendations. For instance, concerning: • accreditation of forensic pathology services in Europe; • principles for on-­ site forensic and medicolegal scene and corpse investigation; • guidelines for the examination of victims of sexual assault; • harmonisation of forensic and medicolegal examinations of persons; • guidelines for the examination of suspected elder abuse. Altogether, the guidelines of the European Council of Legal Medicine cover not only forensic pathology but the whole field of forensic medicine including clinical forensic medicine.

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Assessment at the laboratory

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Before starting work on any case, the expert should always carry out an assessment of the information available, of the risk of contamination or any other issue that could affect the integrity of the items. This should be done before the items provided for examination are submitted to the laboratory.

Sampling The actual sampling plan (ENFSI 2008) should consider the circumstances of the specific case and the amount of material available for examination. A chain of custody record should be maintained from the receipt of items/samples, which details each person who was involved in the process (ILAC 2002). The following points should be considered:

Validation All technical procedures must be fully validated before they are used on casework items. Validation is the confirmation by examination and the provision of effective evidence that the particular requirements for a specific intended use are fulfilled. The laboratory should use only validated techniques and procedures for the examination and interpretation of results in casework whenever possible (ENFSI 2008). Minimum requirements are: • Documented, scientifically reliable procedures and quality control concepts. • Definition of critical aspects and limitations of the techniques/ methods. • Effective and high-­quality methods, materials and equipment to enable reliable and robust results. • Appropriately trained and competent laboratory staff.

Equipment and measurement traceability The laboratory needs a procedure for the maintenance and calibration of the equipment used in the examination of evidential material. Calibrations and performance checks are necessary where equipment settings can significantly affect the test or analytical result (e.g., temperature, volumetric equipment, microscopes).

International Guidelines and Accreditation in Forensic Medicine

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Laboratory examinations and anticontamination precautions

The laboratory design should be based on written procedures for taking and maintaining case notes to support the conclusions drawn in reports (Butler 2001). All records must be produced in accordance with the requirements of the particular legal system in force in the member country and sufficiently detailed to allow another forensic expert to evaluate the quality and reliability of the work. Where evaluation and interpretation are required, the following information should be provided in the expert report (ENSFI 2008): • Background case information available. • Description of evidence and methodology. • Discriminating power of the analytical methods used. • Degree of certainty that can be assigned to a result or identity. • Information available in relevant databases. The laboratory should list published validated data that are available and recommend approaches for the evaluation and interpretation of examination results.

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Proficiency testing assesses the laboratory systems and the competence of individuals participating in the tests. The laboratory should specify the number of proficiency tests taken each year. Participants in the tests should follow the standard laboratory procedures for casework. Tests have to be completed and returned to the proficiency test organisation for evaluation. The design and implementation of proficiency tests should be carried out in accordance with the recommendations of the international forensic guidelines on the conduct of proficiency tests and collaborative exercises (ENFSI 2008, 2015).

Reporting the results: evaluation and interpretation

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Calibration is carried out for many types of analyses using synthetic standards containing the analytes being tested, prepared from chemicals of known composition (ILAC 2002). Reference collections of data/items/materials representative for casework (e.g., drug samples, frequency databases) should be implemented, documented and controlled. Analytical performance should be monitored by operating quality-­control schemes including, for example, reference collections, positive and negative controls and repeated testing (for further details, see ILAC 2002, Chapter 5.9).

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To avoid contamination, written guidelines and training need to be available to assist individuals in managing the specific risks associated with individual analyses (ILAC 2002; ENFSI 2008; Wilson-­Wilde 2018). One important new aspect in the ISO/IEC 17025:2017 is the extended implementation of a risk-­based concept focusing on the avoidance of contamination and wrong sample codification (e.g., four-­eyes principle, barcodes, reanalysis of samples; Figure  4.2). Anticontamination precautions should be implemented for all examinations; if these include materials that may be required for subsequent analysis, extreme caution should be taken because of the sensitivity of current techniques, including the wearing of appropriate barrier clothing (e.g., gloves, face masks). The minimum anticontamination precautions are: • Condition of packaging on arrival at the laboratory (e.g., any damage, leaks). • Storage of materials (e.g., separation of controls from material originating from suspects). • Use of separate work areas if there are cross-­contamination risks (e.g., suspect to victim, suspect to other suspect).

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4.2  Recommendations for forensic DNA laboratories based on ISO/IEC 17025

Basic laboratory procedures are provided in ISO/IEC 17025 (2017). In addition, recommendations for forensic DNA analyses are available which have been specified and adopted during method development. There is an increasing importance of international DNA databases (see the recommendations on DNA typing (Editorial 1991, 1992a, b) and ENFSI recommendations (ENFSI 2008, 2011, 2016)).

Personnel qualifications All staff working in the laboratory need to be specifically qualified for the work required. The technical manager (head of the laboratory) must have an academic degree (minimum bachelor, usually masters and PhD) that involved basic knowledge in the

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DUTIES OF FORENSIC MEDICINE IN MODERN SOCIETIES

Accommodation and environmental conditions

Depending on the equipment used by the forensic DNA laboratory, the relevance of measurement-­specific requirements for testing and calibration should be defined. The volumetric equipment needs a minimum of one calibration check per year, but should be continually visually examined. Temperature profiles of PCR cyclers should be calibrated once a year and can be additionally monitored by control samples on documented block positions, which should be systematically varied.

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Investigation on test items and those from a scene of crime should be followed by defined criteria on the efficiency background. Prioritisation depending on the relevance of a selected item is usually necessary for complex casework analysis, including multiple items. If possible, only a minimum amount of material should be selected for further sampling and DNA typing. Photographic documentation of all items and selected traces represents a basic step of the sampling procedure. Positive and negative controls are relevant to check equipment, reagents and materials.

Handling of items/samples The laboratory must be able to demonstrate that the items/samples examined are those submitted to the laboratory. Appropriate packaging and labelling should be checked. For all samples, the chain of custody must be fulfilled (sample documentation, labelling of selected traces, persons involved in the procedure, dates and locations of the sample and traces). Furthermore, the quality or condition (e.g., degradation) of the item/biological material submitted should be documented. Extracted DNA not used from traces should be stored under adequate conditions (refrigerator) to enable later investigations.

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The forensic laboratory area needs defined areas of security to minimise the risk of contamination. Access must be controlled and limited, especially to evidence storage areas and the operational area of the laboratory. Working areas must be separated in pre-­and post-­PCR rooms to avoid cross-­contamination; standard separation is into: (i) examination and preparation of samples and crime scene items, (ii) DNA extraction and PCR set-­up and (iii) PCR amplification and electrophoretical separation of amplified DNA. Written procedures have to be available for environmental monitoring, cleaning procedures and decontamination.

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field of genetics/molecular genetics. Furthermore, she/he must have detailed knowledge and experience in molecular biology and forensic genetics (including good working practice, casework examination and supervision of experts). Continuous training is necessary to improve knowledge in new DNA-­typing methods, biostatistic approaches and laboratory strategies. Technical staff should be qualified for laboratory work in molecular biology/genetics. The technical manager is responsible for permanent training and optimising the qualifications of the technical staff based on written standards of competence. Scientific and technical training/education has to be documented for each person (maintenance of competence; e.g., workshops, scientific meetings, training courses) and competence and qualifications for different analytical techniques for evidence samples and DNA methods should be personally defined (authorised). The preparation of a final report lies within the responsibility of the scientifically qualified staff (technical manager and/or deputies).

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Analytical testing, calibration and method validation

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The technical procedures used must be forensically validated. Usually, new commercial kits/products are validated by the companies based on defined standard protocols. If no variation of the validated standard protocol was carried out, such products need only documented confirmation to demonstrate the reliability of the procedure and the results obtained. The development of new (in-­house) methods requires complete validation, usually based on comparison with established methods (e.g., body fluid identification based on mRNA assays, DNA phenotyping; Haas et al. 2011; Kayser  2015). Furthermore, variations on established standard protocols must be validated to confirm the results including experiments to investigate limits and specificity of the system (e.g., control samples and dilutions, mixture experiments, real casework samples). Only high-­quality standard materials and reagents should be used and pre-­tested for their reliability. Lot/batch numbers must be recorded. Instruments and equipment should be maintained properly and calibrated with documented logs for calibration records. If available, national or international standards should always be used for calibration.

Assuring the quality of test results The laboratory has to monitor the analytical performance and quality of the procedures in place. Internal quality controls are necessary to demonstrate reliability and correctness of the results including reference samples, positive and negative controls, replicate testing, internal standards, defined allelic ladders and alternative methods, if necessary. If possible, the extracted DNA should be typed in two or more independent analyses. For the interpretation of results, written criteria must be available (profile acceptance criteria: percentage of stutter, artefact peaks, minimal and maximal peak highs, allelic imbalance (heterozygotes)). Analyses of mixed stain patterns and/or low-­template DNA in particular need cautious interpretation of DNA profiles based on the context of published guidelines (ENFSI 2011). Optimised analysis of mixed profiles may be possible using massive parallel sequencing methods (De Knijff 2019). Proficiency testing: for external controls, two tests per year are necessary including analysis conditions, which have to be comparable to routine casework.

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Accommodation and environmental conditions It must be guaranteed that unauthorised persons do not have access to any of the laboratory rooms. Unauthorised persons may only visit the laboratory if accompanied by authorised personnel. The laboratory space must be large enough to accommodate adequate laboratory equipment for a clear identification and quantitative determination of individual substances. Substance samples and biological material must be processed in separate laboratory rooms; contamination has to be excluded. Sufficient cooling and deep freeze units should be available for the proper storage of analytical standards and samples and their protection from unauthorised access. Apart from the basic equipment, the appliances currently needed in an analytical laboratory generally consist of: • Gas chromatography with special detectors such as a nitrogen-­ specific detector, electron capture and flame ionisation detectors or a mass spectrometer. • High-­performance liquid chromatography with special detectors such as a diode array detector, UV and fluorescence detectors or a mass spectrometer. • Immunochemical and photometric analyses. Other procedures or equipment that deliver equivalent results may also be used.

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Interpreting results in the field of forensic toxicology requires knowledge of multiple areas, especially the natural sciences, medicine and jurisprudence. Forensic toxicologists have to bear in mind, once they commence an analysis, that their report might be introduced as evidence in a criminal or civil court. Fundamental knowledge based on education and training in relevant disciplines is therefore the basis for analysing samples, interpreting results and providing reports. The provision of reports in particular always has to be checked against the background of forensic aspects and possible consequences for a suspect or victim (AAFS 2009).

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4.3  Recommendations for forensic toxicological laboratories based on ISO/IEC 17025

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The test report has to include all the information required by ISO/IEC 17025. All received items and selected traces must be listed and – depending on the individual case – described further. The basic investigation methods must be documented in writing. Criteria for interpretation of the results have also to be documented by the laboratory and need to be explained in the report if necessary (e.g., guidelines for mixed stain analysis, biostatistical calculations, population data).

the human body, especially in anatomy, physiology, pathology and biochemistry, are essential. The differentiation between intoxication and forensic psychopathology is of importance as well as back-­calculations (especially regarding blood alcohol concentrations), legal matters and general pathology. One important and challenging aspect of interpretation in forensic toxicology is postmortem toxicology. Postmortem diagnostic findings in poisoning, thanatochemistry (especially changes in drug distributions) and aspects of exhumation also have to be considered. Technical staff should be qualified for all the laboratory work required in forensic toxicology. The technical manager is responsible for permanent training and optimising the qualifications of the technical staff. Scientific and technical training/education must be personally documented (maintenance of competence; e.g., workshops, scientific meetings, training courses). The preparation of final reports is within the responsibility of the scientifically qualified personnel and staff (technical manager and/or deputies).

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Reporting the results

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Personnel qualifications

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The supervisor of the laboratory where the designated tests are carried out should have a university degree in life sciences or medicine (preferably a PhD), relevant further education and forensic toxicological experience. For example, the German Society of Toxicological and Forensic Chemistry (Gesellschaft für Toxikologische und Forensische Chemie, GTFCh) awards titles in the designated field of work (‘Forensic Toxicologist GTFCh’ or ‘Forensic Chemist GTFCh’). The postgraduate modules taken as part of the forensic toxicologist GTFCh course are summarised in Table 4.1 (GTFCh 2006a). Furthermore, continuous professional training is necessary to improve knowledge of analytical techniques, new drugs on the market, regulations, biostatistic approaches and laboratory strategies (GTFCh  2006b). Other societies have similar education requirements. A knowledge of pharmacology and toxicology is important for the interpretation of obtained data. This includes structures of molecules and structure–response relationships, physical ­mechanisms, spectrums of efficacy, adverse reactions, interactions, modes of application, xenobiotic metabolisms and organ toxicology of drugs. Toxicity tests and other aspects of pharmacodynamics and pharmacokinetics and toxicodynamics and toxicokinetics, respectively, also represent important knowledge for the interpretation of toxicological and clinical cases. Furthermore, basics of human biology, including knowledge of the functions of

Analytical testing, calibration and method validation New analytical methods to be used in forensic toxicology require careful method development and thorough validation of the final procedures. Analytical methods in forensic toxicology may either be used for screening and identification of drugs, poisons and/or their metabolites in biological fluids or tissues, for their quantification in these matrices, or for both. For quantitative bioanalytical procedures, there is a general agreement that, at the least, the following validation parameters should be evaluated: selectivity, calibration model (linearity), stability, accuracy (bias), precision (repeatability, intermediate precision) and the lower limit of

Table 4.1  Postgraduate professional education modules for the German ‘Forensic Toxicologist GTFCh’ qualification. Subject

Domain

Details

Basics of human biology Toxicology/ pharmacology

Basic knowledge of the function of the human body (anatomy, physiology, biochemistry) Structure, physical mechanism, spectrum of efficacy, adverse reaction, interaction, mode of application, toxicogenetics/ pharmacogenetics

Toxicodynamics and toxicokinetics/ pharmacodynamics and pharmacokinetics

Xenobiotic metabolism, organ toxicology, toxicity tests, structure– response relationship Definition of terms, conditions of effect of toxins, influence of endogenic and exogenic factors, postmortem diagnostic findings in poisoning, principles of inquest, sampling in suspicion of poisoning, exhumation, thanatochemistry, metals, non-­metal inorganic poisons, organic poisons, legal aspects

Alcohol

Retrograde calculation, law, alcohol consumption after the critical incident and prior to blood sampling

Other substances affecting the central nervous system

Pharmacokinetics of the most important intoxicating substances, intoxicating substances in road traffic incidents, aspects of medicine in road traffic incidents, sampling, determination of chronic abuse, retrograde calculations, legal matters, pathology, intoxication versus forensic psychopathology

Substances not affecting the central nervous system

Analyses and appraisal of samples taken from living humans

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Postmortem toxicology

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Forensic toxicology

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Basics of forensic genetics

Analysis of illicit drugs

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Basics of forensic chemistry

Analysis of other non-­biological seizures

Epidemiology of acute poisoning

Certification

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Clinical toxicology

Accreditation

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Quality management

Legal aspects in diagnostics and treatment of poisoning Measures taken in acute poisoning

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Differential diagnosis and cardinal symptoms in poisoning

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Therapy of acute poisoning

Toxicologically relevant parameters

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Clinical chemistry

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Symptoms of the most frequent types of poisoning, therapeutic methods, antidotes

Basics of pharmacy Legislation

Relevant laws and regulations

Narcotics law, road traffic law and criminal law (e.g., intoxication effects and criminal responsibility)

Rights and duties of experts Current court judgements passed Analytical toxicology

Material of investigation

Standard matrices Alternative matrices (e.g., hair, saliva, tissue samples)

Pre-­analytical aspects

Sampling and storage

Analytics

Sample preparation Qualitative and quantitative analysis procedures including validation

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Table 4.1  (Continued) Subject

Domain

Details Quality assurance

Method development Post-­analytical aspects

Sample storage

Biostatistics Plausibility of forensic toxicological results Interpretation of forensic toxicological results

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tigation. Within Germany, forensic toxicological testing is not being performed on whole blood, but on serum or plasma (if these can be obtained from the samples). For toxicological testing with or without determination of blood alcohol concentrations, a blood sample preserved without additives as well as a blood sample with fluoride (especially for the determination of cocaine) should be used to obtain serum or plasma. It should be stated in the expert report whether whole blood or serum/plasma was used. Regarding the collection of urine samples, special measures must be observed depending on the individual request, such as collection under visual control in drug abstinence checks (compare also evaluation criteria on fitness to drive diagnostics). For transportation, the sample material should be packed in shockproof and tightly sealed containers with no exposure to heat or light. The analytical questions and possible corresponding analyses determine how quickly a sample should be transported and whether special transport conditions (e.g., deep freezing) apply. In special cases, regulations on the transport of hazardous goods on the roads must be observed by the sender. Special recommendations exist for the sampling of materials for toxicology investigations during autopsy, depending on the individual case (Table 4.2). Sometimes photographic documentation of the samples is necessary. Positive and negative controls are relevant to check equipment, reagents and materials.

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­ uantification (LLOQ). Additional parameters that may be releq vant include the limit of detection (LOD), recovery, reproducibility and ruggedness (robustness). For qualitative procedures, a general validation guideline is currently not available, but there seems to be agreement that at least selectivity and the LOD should be evaluated and that additional parameters such as precision, recovery and ruggedness (robustness) might also be important. For methods using liquid chromatography/mass spectrometry (LC–MS), experiments for the assessment of possible matrix effects (i.e., ion suppression or ion enhancement) should always be part of the validation process, particularly if they utilise electrospray ionisation (ESI). The guidance documents issued by the GTFCh for quality control in forensic toxicological analyses comprise numerous details concerning sample preparation, requirements for immunoassays, performance control of chromatographic procedures or identification criteria in mass spectrometric detection. Special recommendations are given for the validation of new methods in forensic toxicology and for forensic hair analysis as well as for postmortem toxicology. A well-­known guideline for the validation of new methods has been published by Peters et al. (2007). The measurement of uncertainty is an important parameter for each analytical method and the general basic principles make up the Guide to the Expression of Uncertainty in Measurement (2008), on which the EURACHEM/CITAC Guide (2000) for chemical analytical methods and the Nordtest Technical Report (2003) for environmental analytical methods are based. The GTFCh (2009b) prefers an estimation of measurement uncertainty via collaborative testing and intermediate precision determined by control samples. Only high-­quality standard materials and reagents should be used and pre-­tested for reliability. Lot/batch numbers must be recorded. Instruments and equipment should be properly maintained and also calibrated with documented logs for calibration records.

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Expert opinion on records

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Forensic expertise

For example, consideration of problem, limit values

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Forensic expertise

Sampling If not regulated by adequate guidelines or recommended otherwise, the testing laboratory should inform the applicant about the type, quantity, storage conditions and transport conditions of the required sample materials, in order to guarantee an adequate inves-

Handling of items/samples The laboratory must be able to demonstrate that the items/samples examined were those submitted to the laboratory. Appropriate packaging and labelling should be checked. For all samples, the chain of custody must be fulfilled (sample documentation, labelling of the selected trace, persons involved in the procedure, dates and locations of samples and traces). Furthermore, the quality or condition (e.g., degradation) of the item/biological material submitted should be documented. The laboratory has to immediately inform the applicant in cases where the sample is damaged, the sample is unsuitable for testing, the sample amount is too small for the research task or where the required analysis cannot be carried out by the laboratory. After receipt, samples should be stored appropriately so that analytes do not decompose and samples are not contaminated.

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Table 4.2  Recommendations for sampling prior to, or during, all autopsies or autopsies where the cause of death remains uncertain or special problems have to be addressed. All autopsies

Additional specimens in cases in which the cause of death remains uncertain

Special problems

Scalp hair; alternatively body hair

Vitreous humour

Specimens that can be collected prior to autopsy Blood from the vena femoralis; alternatively a sample from the vena subclavia Vomitus from the scene Urine

Liquor cerebrospinals Finger and toe nails Skin and subcutaneous tissue Smear tests from the skin and mucosa

Muscle tissue Fatty tissue Contents of the small and large intestines Pericardial fluid Fluid from the thoracic cavity Bones, bone marrow Entomological species

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Bile Liver Lungs Brain Kidneys

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Heart blood Gastric contents

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Specimens that should be collected immediately after opening the thoracic and abdominal cavities or after organ removal

Source: GTFCh (2004).

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known, the time of the incident must be included. The name of the person responsible for the analysis and external representation must be indicated. If statements are made, all relevant connecting facts should be included in the expert report if no other arrangement was made with the client. When analysing biological material, the results must be explicitly assignable to the person from whom the sample was taken, as well as to the blood withdrawal system. The methods used and the nature of the test material analysed (e.g., whole blood, serum, plasma, remaining blood) should be stated. The use of unsuitable withdrawal or sampling systems should be indicated. The possible loss of analyte due to delayed or inadequate storage of, for example, serum or plasma samples (freezing conditions of a minimum of −15 °C are required) or to unsuitable analysis should be mentioned in the report, as well as the basic investigation methods. Criteria for the interpretation of the results have to be documented by the laboratory and need to be explained in the report if necessary.

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After completion of the analysis and the final report, remnants of sample material and original containers (blood withdrawal systems, tubes, containers, etc.) should be stored according to the applicable administrative regulations  – at least 6  months, and blood samples for a minimum of 2 years. In general, a storage period of 2 years is advised. If other laws or regulations apply, these should be followed. The applicant should be informed of the storage period. Original containers including all sample remains must be presented on demand.

Assuring the quality of test results

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The laboratory has to monitor analytical performance and quality of the procedures. Internal quality controls are necessary to demonstrate reliability and correctness of the results, including reference samples, positive and negative controls, replicate testing, internal standards and alternative methods if necessary. For the interpretation of results, written criteria must be available (acceptance criteria). Aspects of quality assurance and quality control in forensic toxicological analyses are described in detail in the GTFCh (2009b) guidelines including recommendations for acceptance criteria. External quality control is done by proficiency testing that includes analysis conditions and is comparable to that of routine casework. Collaborative testing complements the internal accuracy monitoring of laboratories and simultaneously guarantees an objective supervision of accuracy or bias of the results of qualitative and quantitative forensic toxicological analyses.

Reporting the results The test report has to include all the information required by ISO/IEC 17025. At the least, dates and times of sample collections and sample receipt as well as the period of analysis (beginning and end of analysis) must be given as header data in the report. If

Documentation The head of the laboratory is responsible for the written documentation of all methodological instructions used by the laboratory – such as the SOPs and procedures for all important operations in the laboratory, included as part of the quality management handbook. The instructions should comply with approved quality ­criteria and must be audited (e.g., within the framework of an accreditation). Application forms, status protocols and all ­documents, such as evaluations of measurement results and analyses, measurement protocols, calibrations, chromatograms, ­spectra, analysis reports or expert reports as well as the assay procedure, must be filed and stored in a way that they may be presented to an expert authorised by court at any time. On the basis of the ­documents, the correct analytical procedure and the expert opinion about the results must be deducible. It should be clear which

International Guidelines and Accreditation in Forensic Medicine

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Even low-­technology autopsies have to provide the following information: • Establish the cause of death. • Assist in determining the manner of death (i.e., homicide, suicide, etc.). • Compare the premortem and postmortem findings. • Produce accurate vital statistics. • Monitor public health. • Assess the quality of medical practice. • Instruct and educate medical students and physicians. • Identify new and changing diseases. • Evaluate the effectiveness of therapies such as drug administration, surgical techniques and prostheses. • Reassure family members. • Protect against false liability claims and settle valid claims quickly and fairly (Lundberg 1998). The main tasks of the forensic autopsy are, of course, establishing the cause and manner of death. Another important task is the reconstruction of the course of events or clarification of a causal connection with preceding bodily harm. A further important task may be identification. The autopsy itself, especially the clinical autopsy, represents a measure of quality control on clinical diagnostics and therapy. However, the autopsy has to follow quality criteria as well. Quality criteria and quality control for the forensic autopsy were already established in the 19th century. Of special importance today is Recommendation No. R(99)3 of the Committee of Ministers to EU member states on the harmonisation of medicolegal autopsy rules in Europe (Council of Europe 1999, cited in Byard and Winskog 2012) (meanwhile updated by the ECLM in Dubai in 2014). Medicolegal autopsies should be performed whenever possible by two physicians, of whom at least one should be qualified in forensic pathology or forensic medicine. Qualification in forensic pathology or forensic medicine takes about 5 years and follows individual national regulations. The autopsy has to be carried out step by step beginning with identification of the body, an external and internal examination and then completing the autopsy report in compliance with manuals of autopsy practice. For measuring and weighing (e.g., body weight, body height, organ weights), calibrated instruments have to be used. Minimal rules have to be applied in the sampling of tissues and body fluids: in all autopsies, the basic sampling scheme includes specimens from the main organs for histology

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and femoral blood (for alcohol and toxicology), urine and gastric contents. If death is related to physical violence, sampling also includes the injuries (e.g., to determine wound age) and any foreign materials seen in wounds. As for the laboratory sectors of forensic science, contamination has to be avoided and proper identification of samples has to be ensured. SOPs must be available for every step of the autopsy and the chain of custody of autopsy samples from the autopsy room to the laboratory must be guaranteed. Autopsy technicians must have a special qualification. As in clinical pathology, histology is an integral part of the autopsy. For postmortem biochemical analyses, the vitreous humour should be sampled. After autopsy, the body has to be returned in a dignified condition and it is the task of the forensic pathologist and the autopsy technician to ensure this. In the United States, the standard for quality in death investigation for medical examiner offices is accreditation by NAME. Accreditation attests that an office has a functional governing code, adequate staff, equipment, training and a suitable physical facility, and can produce a forensically documented, accurate, credible death investigation product. According to the US National Research Council (2009), most coroner systems cannot qualify for accreditation because of problems related to size, insufficient staff and equipment and insufficiently trained personnel, which inhibits their ability to perform a competent physical examination, make and/or exclude medical diagnoses on dead bodies and make determinations of the cause and manner of death. Also, many medical examiner systems are constrained by budget, lack of staff, lack of equipment and insufficient facilities and cannot meet NAME standards. In other countries, for example, in Germany, most institutes of forensic medicine were accredited during the last 10 years according to ISO 17025 for forensic purposes. As already mentioned, the NAME has elaborated an accreditation/certification checklist which should be served as decision-­ making support to assist inspectors appointed to evaluate applicants. In the same spirit of the NAME accreditation standards, the European Council of Legal Medicine board decided to set up an ad hoc working group with a mission to elaborate an accreditation/certification procedure similar to the NAME, taking into account the realities of forensic medicine practices in Europe and restricted postmortem investigation (see Mangin et al. 2015).

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person(s) performed the analysis and which expert is responsible for the final result. The person responsible for the analysis has to ensure that the research was performed according to the documents in force. Documentation may also be electronic if accessibility is guaranteed over the relevant period of storage. Documents must be retained for at least 6 years unless the relevant administrative regulations stipulate longer retention periods.

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4.5  Recommendations for clinical forensic medicine based on ISO/IEC 17020 International Laboratory Accreditation Cooperation (ILAC) has extended their guide to encompass ISO/IEC 17020, as it applies to crime scene investigations (ISO/IEC 2004). In additional to crime scene investigations (e.g., photographs of the body as it is found, notification of all relevant artefacts, protection of the deceased’s hands and head with paper bags, investigations for determination of time since death), second postmortem examinations before cremation and physical investigations of living

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individuals after bodily harm should follow this recommendation. Crime scene investigations, external examinations before cremation and bodily investigations should be carried out whenever possible by board-­certified experts in forensic medicine or forensic pathology, at the least under the supervision of a qualified expert. The ECLM has published recommendations also in the field of clinical forensic medicine.

• Trace evidence/material: refers to forensic examinations in

which the following types of material, amongst others, are involved: fibres, hairs, glass, paint, soil, etc. • Validation: confirmation by examination and provision of objective evidence that the particular requirement for a specific intended use is fulfilled. [ISO 8402:1994 – 2.18]. Other relevant definitions can be added or superfluous ones removed as required by the particular evidence type, but those given should cover most aspects.

Appendix: Definitions (ENFSI Appendix 4)

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References and further reading AAFS (American Academy of Forensic Sciences) (2009). So You Want to Be a Forensic Scientist! http://www.aafs.org/choosing-­ career (last accessed 28 December 2011). Anon. (2007). Vorschrift für die Vornahme der gerichtlichen Totenbeschau. Reichsgesetzblatt für das Kaisertum Österreich, Verordnung der Ministerien des Inneren und der Justiz vom 28. Jänner 1855. In: M. Hochmeister, M. Grassberger and T. Stimpfl (eds.), Forensische Medizin für Studium und Praxis, Vol. 2. Wien: Maudrich. AWMF (Arbeitsgemeinschaft der Wissenschaftlichen Medizinischen Fachgesellschaften) (2007). Leitlinien der Deutschen Gesellschaft für Rechtsmedizin: Die rechtsmedizinische Leichenöffnung. Leitlinien-­Register No. 054/001, Entwicklungsstufe 3. http://www.awmf.org/uploads/tx_ szleitlinien/054-­001_S3_Die_rechtsmedizinische_Leichenoeffnung_12-­ 2007_12-­2012.pdf (last accessed 28 December 2011). Bär, W., Brinkmann, B., Budowle, B. et al. (1997). DNA recommendations. Further report of the DNA Commission of the ISFH regarding the use of short tandem repeat systems. International Journal of Legal Medicine 110, 175–176. Bär, W., Brinkmann, B., Budowle, B. et al. (2000). DNA Commission of the International Society for Forensic Genetics: Guidelines for mitochondrial DNA typing. International Journal of Legal Medicine 113, 193–196. Burton, J.L. and Rutty, G.N. (2010). The Hospital Autopsy. A Manual of Fundamental Autopsy Practice. London: Hodder Arnold. Butler, J.M. (2001). Forensic DNA Typing. San Diego: Academic Press. Byard, R.W. and Winskog, C. (2012). Histology in forensic practice  – required or redundant? Forensic Science, Medicine and Pathology, 8 (1): 56–57 discussion 58–72. College of American Pathologists (2006a). Basic Competencies in Forensic Pathology. A Forensic Pathology Primer. Northfield, IL: College of American Pathologists.

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American Board of Forensic Toxicology, http://www.abft.org. National Association of Medical Examiners, Autopsy Standards and Inspections Checklist, https://www.thename.org/inspection-­ accreditation. European Council of Legal Medicine, http://eclm.eu/. (All last accessed 20 March 2020.)

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mine whether quality activities and related results comply with planned arrangements and whether these arrangements are implemented effectively and are suitable to achieve objectives. [ISO 8402:1994 – 4.9] Calibration: a set of operations that establish, under specified conditions, the relationship between values and quantities indicated by a measuring instrument or measuring system, or values represented by a material measure or a reference material, and the corresponding values realised by standards. [International Vocabulary of Basic and General Terms in Metrology 1993  – 6–11 and ISO/IEC Guide 25:1990 – 3.6] Competence: a person’s qualification for the job by virtue of their training and/or experience and demonstrated knowledge, skills and abilities. Competence assessment: a formal assessment to check whether or not an individual meets the standards of performance. Management/administrative review: a review of the case file and report, in each case, to ensure that the customer’s needs have been properly addressed, and there has been compliance with laboratory policy and, for the report, editorial correctness. Proficiency test: the use of inter-­laboratory comparisons to determine the performance of individual laboratories for specific tests or measurements and to monitor laboratories’ continuing performance. [ISO/IEC Guide 43-­1 (1997). Proficiency Test by Interlaboratory Comparison – Part 1: Development and Operation of Proficiency Testing Schemes] Quality assurance: all the planned and systematic activities implemented within the quality system, and demonstrated as needed, to provide adequate confidence that an entity will fulfil the requirements for quality. [ISO 8402: 1994 – 3.5] Quality control: operational techniques and activities that are used to fulfil the requirements for quality. [ISO 8402:1994 – 3.4]. Raw data: record of results of analyses and examinations in the form in which those results were interpreted by the original analyst. Scientific/technical review: review of a case file and report for the reliability and interpretation of the scientific findings. Systematic error: any discrepancy due to improper instrument function or setting.

Useful websites

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• Audit: a systematic and independent examination to deter-

International Guidelines and Accreditation in Forensic Medicine

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Gjertson, D.W., Brenner, C.H., Baur, M.P. et  al. (2007). ISFG: Recommendations on biostatistics in paternity testing. Forensic Science International: Genetics 1: 223–231. GTFCh (Gesellschaft für Toxikologische und Forensische Chemie (2004). Recommendations for sampling postmortem specimens for forensic toxicological analyses and special aspects of a postmortem toxicology investigation. Toxichem Krimtech 71 (2), 101–107. (English translation, https://www.gtfch.org/cms/images/stories/files/Recommendations%20 for%20sampling%20postmortem%20specimens%20%28Appendix%20 D%29.pdf (last accessed 28 December 2011.) GTFCh (Gesellschaft für Toxikologische und Forensische Chemie (2006a). Professional title ‘Forensic toxicologist GTFCh’. Toxichem Krimtech 73 (3), 118–125. (English translation, http://www.gtfch. org/cms/index.php/forensictoxicologist (last accessed 28 December 2011.) GTFCh (Gesellschaft für Toxikologische und Forensische Chemie (2006b). Continuing education regulations for ‘Forensic toxicologist GTFCh’. Toxichem Krimtech 73 (3), 126–127. (English translation, http://www.gtfch.org/cms/index.php/ftcontinuingeducationregulatio ns (last accessed 28 December 2011.) GTFCh (Gesellschaft für Toxikologische und Forensische Chemie (2009a). Requirements for the validation of analytical methods. Toxichem Krimtech 73 (3), 185–208. https://www.gtfch.org/cms/files/ GTFCh_Richtlinie_Anhang%20B_Validierung_Version%201.pdf (last accessed 28 December 2011). GTFCh (Gesellschaft für Toxikologische und Forensische Chemie (2009b). Guideline for quality control in forensic-­toxicological analyses. Toxichem Krimtech 76 (3): 142–176. (English translation, https:// www.gtfch.org/cms/images/stories/files/Guidelines%20for%20 quality%20control%20in%20forensic-­toxicological%20analyses%20 %28GTFCh%2020090601%29.pdf (last accessed 28 December 2011.) Haas, C., Hanson, E., Kratzer, A. et al. (2011). Selection of highly specific and sensitive mRNA biomarkers for the identification of blood. Forensic Science International: Genetics 5, 449–458. ILAC (International Laboratory Accreditation Cooperation) (2002). Guidelines for Forensic Science Laboratories. ILAC-­G19:2002, ILAC Secretariat, Silverwater, Australia. ISO/IEC (International Organization for Standardization/International Electrotechnical Commission) (1998). General Criteria for the Operation of Various Types of Bodies Performing Inspection. ISO/IEC 17025:1998. ISO/IEC (International Organization for Standardization/International Electrotechnical Commission) (2004). General Criteria for the Operation of Various Types of Bodies Performing Inspection. ISO/IEC 17020:2004. ISO/IEC (International Organization for Standardization/International Electrotechnical Commission) (2005). General Requirements for the Competence of Testing and Calibration Laboratories. ISO/IEC 17025:2005. JCGM (Joint Committee for Guides in Metrology) (2008). Guide to the Expression of Uncertainty in Measurement (GUM), http://www.bipm. org/utils/common/documents/jcgm/JCGM_100_2008_E.pdf (last accessed 28 December 2011). Kayser, M. (2015). Forensic DNA phenotyping: predicting human appearance from crime scene material for investigative purposes. Forensic Science International: Genetics 18: 33–48.

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College of American Pathologists (2006b). Cause of Death and Death Certificate. Important Information of Physicians, Coroners, Medical Examiners and the Public. Northfield, IL: College of American Pathologists. Council of Europe (1999). Committee of Ministers. Recommendation No. R(99)3 of the Committee of Ministers to Member States on the Legal Autopsy Rules. Adopted by the Harmonisation of Medico-­ Committee of Ministers on 2 Feb 1999. Cusack, D., Ferrara, S.D., Keller, E. et  al. (2017). European Council of Legal Medicine (ECLM) principles for on-­site forensic and medico-­ legal scene and corpse investigation. International Journal of Legal Medicine 131: 1119–1122. De Knijff (2019). From next generation sequencing to now generation sequencing in forensics. Forensic Science International: Genetics 38, 175–180. Editorial (1992a). DNA recommendations – 1992 report concerning recommendations of the DNA Commission of the International Society for Forensic Haemogenetics relating to the use of PCR-­ based polymorphisms. International Journal of Legal Medicine 105, 63–64. Editorial (1992b). Second DNA recommendations  – 1991 report concerning recommendations of the DNA commission of the International Society for Forensic Haemogenetics relating to the use of DNA polymorphisms. International Journal of Legal Medicine 104, 361–364. Editorial (1993). Statement by DNA Commission of the International Society for Forensic Haemogenetics concerning the National Academy of Sciences report on DNA Technology in Forensic Science in the USA. Forensic Science International 59, 1–2. ENFSI (European Network of Forensic Science Institutes) (2008). Guidance on the Production of Best Practice Manuals within ENFSI. Issue No. 008, issue date 1 May 2008. ENFSI (European Network of Forensic Science Institutes) (2016). ENFSI Guideline for Evaluative Reporting in Forensic Science. Approved version 3.0, issue date 8 May 2015. ENFSI (European Network of Forensic Science Institutes) (2011). DNA-­ database Management – Review and Recommendations – ENFSI DNA Working Group. EURACHEM/CITAC (2000). Quantifying Uncertainty in Analytical Measurement, 2nd edn. http://www.measurementuncertainty.org/pdf/ QUAM2000-­1.pdf (last accessed 28 December 2011). European Council of Legal Medicine (2014). Harmonization of Medico-­ Legal Autopsy Rules. ECLM update of the principles and rules related to medico-­legal autopsy procedures. Updated Dubai Jan 2014. Finkbeiner, W., Ursell, P. and Davies, R. (2004). Autopsy Pathology. A Manual and Atlas. Philadelphia: Elsevier. Forensic Toxicologist Certification Board (2003). Certification Requirements. http://home.usit.net/~robsears/ftcb/index.htm (last accessed 28 December 2011). FTC (Forensic Toxicology Council) (2010). Briefing: What is Forensic Toxicology? http://www.abft.org/files/WHAT%20IS%20FORENSIC% 20TOXICOLOGY.pdf (last accessed 28 December 2011). Gill, P., Brenner, C., Brinkmann, B. et al. (2001). DNA Commission of the International Society of Forensic Genetics: recommendations on forensic analysis using Y-­chromosome STRs. International Journal of Legal Medicine 114, 305–309.

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National Research Council (2009). Strengthening Forensic Science in the United States. Washington D.C.: National Academies Press. Nordtest (2003). Handbook for Calculation of Measurement Uncertainty in Environmental Laboratories. Technical Report 537, Nordic Council of Ministers, Espoo, Finland. Peters, F.T., Drummer, O.H. and Musshoff, F. (2007). Validation of new methods. Forensic Science International 165, 216–224. Pomara, C. and Fineschi, V. (2007). Manuale – atlante die technica autoptica forense. Padua: Piccin. Saternus, K. and Madea, B. (2007). Forensic Autopsy. Handling of the Human Corpse. Research in Legal Medicine, Vol. 36. Lübeck: Schmidt-­Römhild. Saukko, P. and Knight, B. (2004). Knight’s Forensic Pathology, 3rd edn. London: Edward Arnold. Skopp, G. (2004). Preanalytical aspects in post-­ mortem toxicology. Forensic Science International 142 (2–3), 75–100. Skopp, G. (2010). Postmortem toxicology. Forensic Science, Medicine and Pathology 6, 314–325. Thali, M. and Bolliger, A. (2011). Quality aspects in autopsy versus virtopsy. In: M.J. Bogusch (ed.), Quality Assurance in the Pathology Laboratory, pp. 112–135. Boca Raton, FL: CRC Press. US Department of Justice, Office of Justice Programs, National Institute of Justice (2013). Death Investigation: A Guideline for the Scene Investigator. www.ojp.usdoj.gov (last accessed 21  January 2013). Virchow, R. (1876). Die Sektionstechnik im Leichenhause des Charité-­ Krankenhauses mit besonderer Rücksicht auf gerichtsärztliche Praxis. Berlin: August Hirschwald. Waters, B.L. (2009). Handbook of Autopsy Practice, 4th edn. x, Totowa: Humana Press. Wilson-­Wilde, L. (2018). The international development of forensic science standards – a review. Forensic Science International 288, 1–9. Wirth, I. (2005). Zur Sektionstätigkeit im Pathologischen Institut der Friedrich-­Wilhelms-­Universität zu Berlin von 1856 bis 1902. Ein Beitrag zur Virchow Forschung. Berlin: Logos Verlag.

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Keller, E., Santos, C., Cusack, D. et al. (2019). European council of legal medicine (ECLM) guidelines for the examination of suspected elder abuse. International Journal of Legal Medicine 133: 317–322. Kirschbaum, K.M. and Musshoff, F. (2011). Quality assurance aspects of interpretation of results in clinical and forensic toxicology. In: M.J. Bogusz (ed.), Quality Assurance in the Pathology Laboratory: Forensic, Technical, and Ethical Aspects, pp. 345–362. Boca Raton, FL: CRC Press. Ludes, B., Geraut, A., Väli, M. et al. (2018). Guidelines examination of victims of sexual assault harmonization of forensic and medico-­legal examination of persons. International Journal of Legal Medicine 132: 1671–1674. Ludwig, J. (2002). Handbook of Autopsy Practice, 3rd edn. Totowa: Humana Press. Lundberg, E.D. (1998). Low-­tech autopsies in the era of high-­tech medicine. Continued value for quality assurance and patient safety. JAMA 208: 1273–1274. Madea, B. (2012). Histology in forensic practice – required or redundant? A commentary. Forensic Science, Medicine and Pathology 8, 64–65. Madea, B. and Saukko, P. (eds.) (2008). Forensic Medicine in Europe. Lübeck: Schmidt-­Römhild. Madea, B., Saukko, P., Oliva, A. and Musshoff, F. (2010). Molecular pathology in forensic medicine. Introduction. Forensic Science International 203, 3–14. Madea, B. (2015) Rech.tsmedizin. Befunderhebung-­Rekonstruktion  – Begutachtung, 3rd edn. Berlin Heidelberg New York: Springer. Madea, B. (2016). Estimation of the Time since Death, 3rd edn. Boca Raton FL: CRC Press Taylor & Francis Group. Madea, B. (2017). History of Forensic Medicine. Berlin: Lehmann’s Media GmbH. Mangin, P., Bonbled, F., Väli, M. et al. (2015). European Council of Legal Medicine (ECLM) accreditation of forensic pathology services in Europe. International Journal of Legal Medicine 129: 395–403. Matuszewski, B.K., Constanzer, M.L. and Chavez-­Eng, C.M. (2003). Strategies for the assessment of matrix effect in quantitative bioanalytical methods based on HPLC-­MS/MS. Analytical Chemistry 75 (13), 3019–3030.

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5 | Nature and Definition of Death 6 | Certification of Death: External Postmortem Examination 7 | Postmortem Changes and Time since Death 8 | Cremation 9 | Crime Scene and Crime Scene Investigations 10 | Autopsy 11 | The Doctor, the Dead and the Relatives 12 | Transplantation 13 | Anthropology and Osteology 14 | Mass Disaster Victim Identification Handbook of Forensic Medicine, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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Burkhard Madea

Short agony (within minutes)

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5.1  Death and dying

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This occurs in many forensically relevant cases (e.g., strangulation, drowning, bleeding to death by stab wounds with involvement of large vessels), but also in cases of death with natural causes (acute myocardial infarction, fulminant pulmonary embolism). Short forms of agony in violent death cases are often characterised by strong reactions in respiration (e.g., asphyxial suffocation with dyspnoea in contrast to hypoxic suffocation), in circulation (e.g., tachycardia, rise in blood pressure) and in the central nervous system (CNS) (e.g., hanging with convulsions due to hypoxia).

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5.1.1  Agonal period

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Death and dying are processes which are characterised by loss of the functions of the great organ systems (cardiovascular system, respiratory system, nervous system) and their coordination. Loss of the coordination of the great organ systems reveals a dissociation of the function of the different organs. The agonal period may be initiated either by disease or trauma. The final crisis leads to a state of vita minima in which no vital signs may be apparent and to a state of death characterised by irreversible cessation of circulatory or respiratory arrest (Figure 5.1). Under special clinical conditions, brain death may replace the classical signs of death (i.e., irreversible circulatory and respiratory arrests and their consequences postmortem lividity and rigor mortis).

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The period of dying is called ‘agony’ and its duration can vary depending on the damaging agent and the remaining reaction facilities of the patient.

Ultrashort or ‘non-­existing’ agony (fractions of a second) This occurs if the body is completely damaged (e.g., by explosion).

Handbook of Forensic Medicine, Volume 1, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

Long agony (within hours) This occurs as the final phase of chronic illness (e.g., tumours); in this case, death is obvious to outsiders. Manner and duration of agony are relevant for the progress of postmortem changes. The term ‘agony’ (death struggle) characterises many violent deaths because sensitively experienced attacks upon a person’s life (during fear, defence, resistance) can lead to intense agonal reactions, partly due to a special pathophysiology of the death process (e.g., asphyxial suffocation). By contrast, many death processes due to pathological internal causes are accompanied by hypoxia of the brain with subconsciousness and unconsciousness occurring so that death is not experienced by the individual anymore.

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acute crisis incomplete dysregulation of vital functions vita reducta

disease

agonal period

trauma

final crisis complete dysregulation of vital functions

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vita minima

individual death

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autolysis

postmortal phase

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supravital reactions

N

irreversible cessation of circulation, respiratory arrest brain death with cessation of life support

putrefaction

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biologic death

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decomposition

TO

Figure 5.1  Scheme and terms of the agonal period. The agonal period is initiated either by disease or trauma.

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Depending on the functional system that is significantly damaged and on the intensity of the damage, three types of death processes can be distinguished: 1. So-­called ‘central death’. 2. Protracted heart and circulatory failure. 3. Acute cardiac arrest. During the final crisis of an agonal period, vital signs may not be apparent and people who are still alive may falsely be pronounced dead (Figure 5.2). In most cases in which death was certified for people still being alive, intoxication with central nervous system depressants in combination with hypothermia is the reason for the lifeless condition. Therefore, especially in cases of hypothermia, the old proverb of emergency physicians that ‘no one is dead until he is warm and dead’ should be kept in mind. The death of an individual is determined either by irreversible loss of circulatory and respiratory functions or brain death. In the agonal period, due to loss of coordination of the great organ systems, vital reactions may steadily decrease (Figure 5.3). Brain death or irreversible circulatory or respiratory arrest represents the death of an individual. However, different tissues survive the death of the individual for a considerable period of time dependent on their metabolic activities. The death of the last cell of the body is called ‘biological’ or ‘cellular’ death, a moment which is of no practical or legal relevance. Of medical and legal importance is only the time of individual death, namely, the brain death.

Figure 5.2  Older lady lying dressed at the bank of a river. Vital signs were missing. Although postmortem changes (lividity, rigor mortis) were missing, she was pronounced dead by an emergency physician. Probably cold stiffening was mixed up with rigor mortis. Cause of death was an intoxication with central nervous system depressants.

5.2  Determination of death Establishing definitely that death has occurred is quite straightforward. Cessation of vital functions can be diagnosed with certainty by the following criteria:

CHAPTER 5  

5.2.1  Apparent death

Dying

Intermediate life

Biological death

Agonal reactions Supravital reactions

1. Presence of definite signs of death (livor mortis, rigor mortis, advanced postmortem changes).

2. Failure of attempted resuscitation for around 30 minutes, con-

O

Do not fill out any death certifications if there are no certain signs of death.

If there is doubt, the physician has to wait for help (emergency doctor) or admission to hospital has to be arranged. In any case, the behaviour of the physician depends substantially on information about anamnesis and the patient’s prognosis. The so-­called ‘uncertain signs of death’, such as fixed pupils, wide pupils, areflexia, loss of cardiac activity, loss of respiration and dropping core temperature, are meaningless if examination is improperly done, especially if reversibility/irreversibility of the condition is not questioned (Box 5.2). For example, absence of peripheral pulse within centralisation of circulation as occurs in hypothermia does not disclose anything about a lack of cardiac activity. Minimum breathing excursions from abdominal respiration will not be seen in fully clothed people. A body surface that feels cold when the ambient ­temperature is low and wet clothing is worn does not give any information about the core temperature. When pupil width is estimated, a possible intoxication always has to be kept in mind (the so-­called ABC toxins: alcohol, amphetamine, belladonna, cannabis).

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firmed by about 30  minutes of a flatline electrocardiogram (ECG) despite carrying out appropriate measures and after ruling out general hypothermia or intoxication by central nervous system depressant drugs. 3. Brain death (can be determined under clinical conditions only during assisted ventilation). 4. Physical trauma that is incompatible with life. As the physician has to make the differential diagnosis of whether a patient has died, or if there is a case requiring reanimation, only those regulations make sense that oblige the physician to carry out external postmortem examination ‘immediately’ upon receiving the report about the case of death. Determination of death is uncomplicated if there are obvious postmortem changes (rigor mortis, livor mortis, putrefaction, injuries that are incompatible with life). Again, we can observe that even if obvious postmortem changes are present (livor mortis, rigor mortis), resuscitation is only being carried out because of a lack of knowledge about these signs. When it comes to determination of death, uncertainty is seen especially within the time interval between collapsing apparently lifelessness and the appearance of early postmortem changes (livor mortis about 20–30 minutes postmortem as the first sign), or in the phase of vita minima and vita reducta with signs of life that are not necessarily ‘catching one’s eye’ if examination is not done thoroughly. Dying and death are processes being characterised by a loss of functions of the body systems (circulatory system, respiratory system and central nervous system) and loss of their coordination. The loss of coordination leads to an increasing dissociation of organ functions with the final point ‘individual death’.

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Figure 5.3  Diagram of agonal period, individual death, supravital period and biological death. Of medical and legal importance is the moment of the individual death.

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Local vital reactions

Systemic reactions

N

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Vital reactions

Determination of death might be difficult within the period of vita minima and vita reducta with increasing devitalisation before early postmortem changes appear due to heart and circulatory arrest. Within the states of vita minima and vita reducta with dysregulation of functional systems  – their reducing coordination and increasing devitalisation – looking for signs of life (respiration, circulation) may be abandoned by the attendee and therefore signs might not be seen if the body is only examined superficially. Complex causes and conditions leading to vita minima and vita reducta are summarised by Prokop and Göhler (1976), forensic medical examiners based in Berlin, and called ‘AEIOU rule’ (Box 5.1). If the conditions of the AEIOU rule are suspected – intoxication with hypnotics, carbon monoxide and alcohol; hypothermia; electrical accidents; apoplexy; brain pressure; metabolic coma; epileptic seizures; hypoxic brain damage; missing signs of life and missing certain signs of death occurring at the same time  – great care is required. In these cases, the following is essential:

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Cardiac and respiratory arrest

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Nature and Definition of Death

Box 5.1  Complexes of causes for determining vita minima/vita reducta. A alcohol, anaemia, anoxemia E electricity, stroke of lightning I injury (head injury) O opium, anaesthetics, neuropharmacological drugs U uraemia (and other metabolic coma), hypothermia Source: From Prokop and Göhler (1976).

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5.2.2  Brain death

• • •

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From a medical and legal point of view, failure of heart resuscitation is a reliable criterion to stop proper but unsuccessful reanimation. Normally, cardiopulmonary resuscitation will be stopped if there is no success after 30–40 minutes or if heart reactivation seems unlikely. This includes the following criteria: 1. Absence of spontaneous electrical activity (zero voltage line in the ECG). 2. Signs of electromechanical decoupling (deformed QRS complexes in the ECG, but absence of a palpable pulse on large vessels). 3. Continuous ventricular fibrillation with frequency deceleration and progressive decrease in amplitudes. These signs show definite and irreversible cardiac death if the patient is normothermic and there are no other specific conditions. If the patient is hypothermic, there is suspected ‘near-­ drowning’ or intoxication; resuscitation has to be performed longer than 30–40 minutes until detoxification occurs or the body warms up. It is only then that it makes sense to stop reanimation. Centrally acting medication and general hypothermia have to be excluded from the diagnosis. Emergency doctors have formulated the principle: ‘No one is dead until he is warm and dead’. Determination of death may follow if resuscitation is unsuccessful and the core temperature is at least 32 °C or above. Irreversibility of loss of function in clinical death can be assured even before postmortem changes occur if the abovementioned regulations are respected and necessary additional instrumental examinations (e.g., ECG) are carried out. This is in accordance with regulations issued by German Medical Association concerning reanimation and emergency care. Livor mortis should appear soon after unsuccessful resuscitation lasting 30–40  minutes. In these cases, the presence of postmortem changes may also be proven without delay. If there is no possibility to make additional instrumental examinations (e.g., ECG), the physician has to wait for postmortem changes (rigor mortis, livor mortis) to appear.

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•

N

• •

Unlike the beginning of life, for a long period, lawyers did not define the end of life. According to a famous lawyer of the beginning of the 19th century, Friedrich Karl von Savigny (1779–1861), death is such a simple natural event that, unlike birth, it does not need exact definitions of its elements. With advancement of reanimation and intensive care, especially the possibility to replace heart and circulatory and respiratory functions by machines, further criteria for the definition of death became necessary, namely, in cases of severe brain damage and/or irreversible loss of integrative function due to primary or secondary damage. The forensic pathologists of the 19th century (e.g., Eduard von Hofmann (1837–1897)) knew that the destruction or significant damage to one or several organs that are essential for life represents a primary cause of death. Bichat (1771–1802) wrote already at the beginning of the 19th century: ‘Each way of sudden death indeed starts with interrupted circulation of blood, respiration or brain activity. One of these functions suspends first  – the other stop gradually’. The irreversible loss of brain function was therefore long known and accepted as cause of death before the introduction of determined criteria of brain death. However, a clearer definition of brain death became necessary in the mid-­20th century as respiratory and circulatory functions could be replaced by machines. According to Moskopp (2017), there are assumptions that brain death was ‘invented’ at the Harvard Medical School in 1968 for the single purpose to harvest organs. In his review on the history of the concept of brain deaths, he outlines that the concept of brain death has been evolved in Europe between 27 August 1952 and 11 March 1960 entirely unrelated to the harvesting of organs for transplantation. Interestingly, in the report of the Ad Hoc Committee of the Harvard Medical School to examine the definition of brain death, Pope Pius XII is cited as follows:

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• •

fixed pupils, dilated pupils areflexia missing cardiac activity dropping core temperature are no ‘certain signs of death’, because absence of peripheral pulse ≠ absence of heart activity minimum breathing excursions within abdominal respiration are not ‘catching one’s eye’ fully clothed persons body surface may feel cold when the ambient temperature is low and wet clothing is worn was it tested, maybe on hands? ≠ core temperature (warmth regulation in coldness – minimum peripheral blood) flow pupil’s width? Reaction to light stimulus? ABC toxins proprioceptive and polysynaptic reflexes, other reflexes than corneal reflex?

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• • • •

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Apparent death – vita minima–vita reducta

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Box 5.2  So-­called uncertain signs of death.

‘In an address, “The Prolongation of Life” (1957), Pope Pius XII raised many questions; some conclusions stand out: (1) In a deeply unconscious individual vita functions may be maintained over a prolonged period only by extraordinary means. Verification of the moment of death can be determined, if at all, only by a physician. Some have suggested that the moment of death is the moment when irreparable and overwhelming brain damage occurs. Pius XII acknowledged that it is not “within the competence of the Church” to determine this. (2) It is incumbent on the physician to take all reasonable, ordinary means of restoring the spontaneous vital functions and consciousness, and to employ such extraordinary means as are available to him to this end. It is not obligatory, however, to continue to use extraordinary means indefinitely in hopeless cases’ (Pius XII, 1958). The authors of the report of the Ad Hoc Committee of the Harvard Medical School conclude: ‘It is the church’s view that a

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Nature and Definition of Death

Box 5.4  The legal framework in Germany.

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Section 16 of the German Transplantation Law (Transplantationsgesetz, TPG) entrusts the German Medical Association with the task of establishing written guidelines, in accordance with the current state of medical scientific knowledge, for the rules for the determination of death according to Section 3 Abs. 1 S. 1 Nr. 2 TPG, and for the procedural rules for the determination of the final, irreversible loss of all functions of the cerebrum, cerebellum and brainstem (ILBF) according to Section 3 Abs. 2 Nr. 2 TPG. This guideline, according to Section 16 Abs. 1 S. 1 Nr. 1 TPG, was last revised by the German Medical Association in 2015, as recommended by the Scientific Advisory Board and with the approval of the Federal Ministry of Health (1). The special legally conferred role of the German Medical Association extends beyond the creation of guidelines and includes a role as guarantor.

O

N

Source: From Brandt and Angstwurm (2018).

Intoxication Hypothermia Circulatory shock Endocrine or metabolic coma Neuromuscular block Therapeutic modifications of the CNS ○○ Barbiturates ○○ Other hypnotics ○○ Sedatives ○○ Analgetics

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Box 5.5  Conditions preventing the clinical diagnosis of brain death.

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time comes when resuscitative efforts should stop and death be unopposed’. Brain death is defined as the irreversible loss of all functions of the brain, including those of the brainstem. The essential findings in brain death are coma, absence of brainstem reflexes and apnoea. An evaluation of brain death should be considered in patients who have undergone massive irreversible brain injury from an identifiable cause. A patient determined as brain dead is both legally and clinically dead. Accordingly, in other legislations for example in Germany, brain death is defined as the irreversible loss of the general function of brain, cerebellum and brainstem, whereas heart and circulatory functions are maintained by controlled ventilation. Brain death is the death of a human being. Today instead of brain death also the term ‘irreversible loss of brain function’ (ILBF) is used. The diagnosis of ILBF means that the patient’s brain has irretrievably ceased to function despite the ongoing maintenance of cardiovascular function by the methods of intensive care medicine (Box 5.3). In Section  3 of the German Transplantation Law (Transplantationsgesetz, TPG), ILBF is described as ‘the final, irreversible loss of all functions of the cerebrum, cerebellum and brainstem’. The significance of ILBF as a reliable sign of death has been definitely established in many countries in position statements of multiple involved organisations, in Germany; for instance, the German Medical Association (Box 5.4), medical scientific speciality societies and religious communities. The diagnosis of brain death is primarily clinical. No other tests are required if a full clinical examination, including each of two assessments of brainstem reflexes and a single apnoea test, is conclusively performed. According to the guidelines for the determination of brain death issued by the New  York State Department of Health, brain death cannot be diagnosed in the absence of either complete clinical findings consistent with brain

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Box 5.3  The aetiology, pathogenesis and relevance of irreversible loss of brain function (ILBF).

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• The aetiology and pathogenesis of ILBF ILBF can be caused by any type of brain disease or injury that raises the pressure within the bony cranial vault above the arterial blood pressure and thereby leads to cessation of the cerebral circulation. For example, ischaemia and/or hypoxia due to a transient cardiac arrest can cause ILBF. • The relevance of ILBF in intensive care medicine The determination of ILBF is an indispensable diagnostic instrument in intensive care medicine, independently of the matter of tissue or organ donation. The question whether ILBF is present arises when the brain functions that are regularly checked in the intensive care are absent, whilst gas exchange and circulatory function continue to be sustained by means of artificial ventilation or extracorporeal oxygenation. In this situation, it must be decided whether: ○○ intensive care measures are to be terminated, or else ○○ tissue and/or organ transplantation are to be initiated, in accordance with the wishes of the deceased. Source: From Brandt and Angstwurm (2018).

death or confirmatory tests demonstrating brain death. The diagnosis of brain death is based on a clinical evaluation of brain function including the following: 1.  Coma (of known aetiology). 2.  Reaction to pain (trigeminus). 3.  Pupillary reaction to light. 4.  Pupillary size. 5.  Corneal reflex. 6.  Eyelid reflex. 7.  Oculocephalic reflex. 8.  Oculovestibular reflex. 9.  Gag reflex. 10.  Cough reflex. 11.  Masseter reflex. 12.  Oculocardiac reflex. 13.  Carotid sinus reflex. 14.  Atropine test. Causes of irreversible coma include severe head injury, hypertensive intracerebral haemorrhage, ruptured aneurysma, subarachnoid haemorrhage, hypoxic–ischaemic brain insults and fulminant hepatic failure. Confounding factors such as drug intoxication, neuromuscular blockade, hypothermia or other metabolic abnormalities must be ruled out (Box 5.5). If toxicants such as barbiturates are present, levels need not be zero, but should be in a range that would normally not be expected

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to interfere significantly with consciousness. Lack of significant hypothermia or hypotension is based on a core temperature over 32 °C at an age ≥18 years and systolic blood pressure over 90 mmHg at age >80 years. For other ages, different age-­specific norms have to be considered. The three essential findings in brain death are coma or unresponsiveness, absence of brainstem reflexes and apnoea. The determination of brain death verifies these findings by the clinical indications listed in Box 5.6 and shown in Figure 5.4.

Box 5.6  Guidelines for determining brain death. Coma or unresponsiveness: no cerebral motor response to pain in all extremities (nail-­bed pressure) and supraorbital pressure Absence of brainstem reflexes • Pupils ○○ No response to bright light ○○ Size: midposition (4 mm) to dilated (9 mm) • Ocular movement ○○ No oculocephalic reflex (testing only when no fracture or instability of the cervical spine or skull base is apparent) ○○ No deviations of the eyes to irrigation in each ear with 50 ml of cold water (tympanic membranes intact; allow 1 minute after injection and at least 5 minutes between testing on each side) • Facial sensation and facial motor response ○○ No corneal reflex ○○ No jaw reflex (optional) ○○ No grimacing to deep pressure on nail bed, supraorbital ridge or temporomandibular joint • Pharyngeal and tracheal reflexes ○○ No response after stimulation of the posterior pharynx ○○ No cough response to tracheobronchial suctioning • Apnoea test

Confirmatory tests

Coma, absence of brainstem reflexes, apnoea

No other appropriate causes

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Full clinical examination

Confirmatory tests

Acute primary or secondary brain damage

EEG: absence of electrical activity during at least 30 minutes in cases of infratentorial brain damage and children below 2 years obligatory

O

Primary

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Proof of irreversibility

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Clinical symptoms

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Acute primary or secondary brain damage

Source: New York State Department of Health (2005).

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In some circumstances, a confirmatory test verifying brain death may be necessary; sometimes confirmatory testing may precede other aspects of determination of brain death. Confirmatory tests are shown in Box  5.7. All phases of the determination of brain dead should be documented in patient’s medical records (Figure 5.5). If the diagnosis of brain death is carried out under intensive care unit conditions, it is neither difficult nor likely to be wrong if the established procedures are followed. However, in the majority of all cases, death is established by the classical signs of death following irreversible circulatory and respiratory arrests.

Secondary

Supratentorial Alternative:

Adults

Children above 2 years

After 12 hours

Children below 2 years

Absence of evoked potentials in cases of supratentorial and secondary brain damage Newborns Alternative:

Complementary examinations

Absence of intracerebral circulation

Diagnosis

Diagnosis

After 24 hours After 72 hours

Figure 5.4  Flow chart for the diagnosis of brain death. Source: German Medical Association (1998).

After 72 hours

Immediate

Box 5.7  Confirmatory tests for determining brain death. • Angiography (conventional, computerised tomographic, magnetic resonance and radionuclide): Brain death confirmed by demonstrating the absence of intracerebral filling at the level of the carotid bifurcation or Circle of Willis. • Electroencephalography: Brain death confirmed by documenting the absence of electrical activity during at least 30 minutes of recording that adheres to the minimal technical criteria for EEG recording in suspected brain death. • Nuclear brain scanning: Brain death confirmed by absence of uptake of isotope in brain parenchyma and/or vasculature, depending on isotope and technique. • Somatosensory evoked potentials: Brain death confirmed by bilateral absence of N20–P22 response with median nerve stimulation. • Transcranial Doppler ultrasonography: Brain death confirmed by small systolic peaks in early systole without diastolic flow, or reverberating flow, indicating very high vascular resistance associated with greatly increased intracranial pressure.

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Source: New York State Department of Health (2005).

Figure 5.5  Brain death checklist. Source: New York State Department of Health (2005).

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Figure 5.5  (Continued)

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According to a recent review on the relevance of ILBF as a reliable sign of death, there is an ongoing need for clear explanation of the diagnostic entity called ‘irreversible loss of brain function’, as the absolute reliability of this diagnosis and its significance continue to be widely misunderstood. According to the authors, the determination of death as an objective medical scientific matter is often not clearly distinguished from various other aspects of death, thus as its metaphysical and cultural aspects and the ways in which the living deal with the dead. The current principles underlying the German guidelines for the diagnosis of ILBF are represented

in Figure 5.6. According to the review by Brandt and Angstwurm, the ILBF involves the loss of all regulatory circuits in which the brain participates and of the physical basis of consciousness and personality. The spontaneity of function of other organs besides the brain and their integration into the individual as a unitary living being are now impossible. ILBF is a reliable sign of human death, and the diagnosis of ILBF, as determined by the procedure required in the guidelines, is reliable. There has not been any known case of incorrect determination of ILBF after proper application of the required standardised diagnostic procedures.

CHAPTER 5  

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Nature and Definition of Death

FIGURE Diagnostic evaluation for irreversible loss of brain function in the first 2 years of life

Diagnostic evaluation for irreversible loss of brain function from 2nd birthday onward

II. Clinical manifestations Coma + absent brainstem reflexes + apnea*2

O

after ≥ 72 h

Supplementary testing

Clinical manifestations

Supplementary testing

(EEG, SEP, or EAEP, Doppler/ duplex ultrasonography, perfusion scintigraphy, CT angiography*4)

Coma +

(EEG, Doppler/ duplex ultrasonography, perfusion scintigraphy, CT angiography*4)

U

after ≥ 72 h

after ≥12 h

TR IB

Clinical manifestations Coma + absent brainstem reflexes + apnea*2

Supplementary testing and

(EEG, EAEP, Doppler/ duplex ultrasonography, perfusion scintigraphy*5)

C

O

N

absent brainstem reflexes + apnea*2

Child from 29 days to 2nd birthday

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at once

obligatory, no waiting period required

Neonate Up to age 28 days

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Primary infratentorial brain damage

at once

(EEG, EAEP, Doppler/ duplex ultrasonography, perfusion scintigraphy*5)

III. Demonstration of irreversibility (clinical & ancillary tests)

TO

after ≥12 h

and

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III. Demonstration of irreversibility*3 Secondary brain damage

II. Supplementary testing

N

II. Clinical manifestations Coma + absent brainstem reflexes*1 + apnea*2

Primary supratentorial brain damage

I. Prerequisites Age ≥ 37 weeks of gestation (since last menstrual period) Acute, severe brain damage No other cause of the manifestations of loss of brain function

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I. Prerequisites Acute, severe brain damage No other cause of the manifestations of loss of brain function

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Three-step algorithm for the determination of irreversible loss of brain function (1) *1 If not all of the required clinical deficits are testable, supplementary ancillary testing is mandatory

FO

*2 If an apnea test cannot be performed, or if the initial paCO2 is above 45 mmHg, the loss of brainstem function must be additionally documented by the demonstration of cerebral circulatory arrest *3 For the procedure to be followed in case of combined brain damage, see Section 3 of the guideline *4 See Comment 9 in the guideline (CT angiography is validated for use only in patients aged 18 years or older) *5 See comments 6 and 9 in the guideline (perfusion scintigraphy is required after the 2nd clinical examination after the stated waiting period) CT computed tomography FAFP early auditory evoked potentials FFG electroencephalography SFP somatosensory evoked potentials Figure 5.6  Three-­step algorithm for the determination of irreversible loss of brain function. Source: Brandt and Angstwurm (2018).

MEDICAL ASPECTS OF DEATH

FO

R

C

O

N

TR IB

U

LY

TO

R

U

Ad Hoc Committee of the Harvard Medical School (1968). A definition of irreversible coma. Report of the ad hoc committee of the Harvard Medical School to examine the definition of brain death. JAMA 205: 337–340. Bichat, M.X. (1796). Recherches physiologiques sur la vie et la mort. Reproduction en Facsimile de l’edition de 1796. GauthierVillars: Paris. Brandt, S.A. and Angstwurm, H. (2018). The relevance of irreversible loss of brain function as a reliable sign of death. Deutsches Ärzteblatt International 115: 675–681. Bundesärztekammer (German Medical Association) (1998). Richtlinien zur Feststellung des Hirntodes. Deutsches Ärzteblatt 95: A1861–A1868. Department of Health (1998). A Code of Practice for the Diagnosis of Brain Stem Death. March. London: Department of Health. Hofmann, von E (1878). Lehrbuch der Gerichtlichen Medizin. Wien: Urban und Schwarzenberg. Link, J., Schaefer, M., and Lang, M. (1994). Concepts and diagnosis of brain death. Forensic Science International 69, 195–203.

Madea, B. (2015). Rechtsmedizin: Befunderhebung  – Rekonstruktion  – Begutachtung. 3rd edn. Berlin, Heidelberg, New York: Springer Verlag. Mollaret, P., Bertrand, I., and Mollaret, H. (1959). Previous coma and necrosis of the central nervous system. Revue Neurologique 101: 116–139. Moskopp, D. (2017). Das Konzept des Hirntodes wurde in Europa zwischen 1952 und 1960 entwickelt. Eine Übersicht zur Historie. Nervenheilkunde 30: 423–432. New York State Department of Health (2005). Guidelines for Determining Brain Death. New York. Oehmichen, M. (1994). Brain death: Neuropathological findings and forensic implications. Forensic Science International 69, 205–219. Pius XII (1958). The prolongation of life. Pope Speaks 4 (4): 393–398. Prokop, O. and Göhler, W. (1976). Forensische Medizin, 3rd edn. Stuttgart: Fischer. Wijdicks, E.FM., Varelas. P.N., and Gronset, G.S. (2010). Evidence based guidelines update: Determining brain death in adults: Report of the quality standard subcommittee of the American Academy of Neurology. Neurology 74, 1911–1918. Wuermeling, H.B. (1994). Brain death as an anthropological or as a ­biological concept. Forensic Science International 69, 247–249.

N

References and further reading

O

PART II  

SE

68

LY

SE

O

Burkhard Madea and Antonella Argo

N

6

Certification of Death: External Postmortem Examination

diseases and cause of death. An exact definition of the manner of death is essential to guarantee legal certainty, but may also have consequences for other legal areas (e.g., civil law, insurance law, compensation law). The legal bases for external postmortem examinations vary from country to country, and even between different counties of the same country. In Germany, for instance, each of the 16 counties has its own funeral law. An external postmortem examination has to be carried out once a dead body is found. A human corpse is defined as follows: 1. The body of a dead person, as long as tissue continuity has not been destroyed by putrefaction. 2. The body of a dead neonate (irrespective of the body weight) if it has completely left the womb, and if after leaving the womb it showed one of the three signs of life (heartbeat, umbilical pulsation and breathing). 3. A stillbirth (stillborn baby weighing ≥500 g). 4. The head or torso separated from a body that cannot be reassembled. The following are not corpses: 1. Skeletons, partial or complete. 2. Miscarriages (stillborn fetuses with a birth weight 75%   Anomalous LCA origin for the right sinus with interarterial course

Anomalous RCA origin from the left sinus with interarterial course Wrong aortic sinus coronary artery anomalies without interarterial course High take-­off from the tubular portion   Anomalous LCx origin from the right sinus or coronary artery Anomalous LAD origin with course anterior to the pulmonary artery Coronary ostial plication Intramyocardial course of LAD (myocardial bridge) Small vessel disease (Continued)

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Table 6.5  (Continued) Highly probable

Uncertain

Myocardial diseases

Acute diffuse myocarditis (any morphological type)

Hypertrophic CM Arrhythmogenic CM Dilated CM Idiopathic fibrosis (non-­ischaemic LV scar) Multifocal myocarditis Sarcoidosis Storage diseases Amyloidosis

Focal myocarditis Idiopathic LV hypertrophy   Hypertensive heart disease Hypertrabeculation (non-­compacted) myocardium

Native/prosthetic valve diseases

Mitral valve papillary muscle or chordae tendineae rupture with mitral valve incompetence and pulmonary oedema Thrombotic block or endocarditis vegetations on valve prosthesis Laceration/dehiscence/leaflet escape of valve prosthesis with acute valve incompetence

Calcific aortic valve stenosis with LV hypertrophy and fibrosis   Myxoid degeneration of the mitral valve (prolapse) with atrial dilatation or LV myocardial fibrosis and intact chordae

Moderate aortic valve sclerosis without LV hypertrophy/mitral annular calcification   Dystrophic calcification of the membranous septum (± mitral annulus/aortic valve) Aortic insufficiency (dilated aortic annulus) Myxoid degeneration of the mitral valve (prolapse) without atrial dilatation or LV fibrosis and intact chordae

U

TO

O C R FO Others

Massive pulmonary embolism Haemopericardium (aortic rupture/ dissection or cardiac rupture) Myxoma or other tumour/ thrombus obstructing a valve orifice

Haemorrhage of the sub-­aortic septum Fibrosis of RBB and LBB (Lenègre disease)

Tetralogy of Fallot, surgical repair ± pulmonary valve incompetence and RV dilatation Congenital aortic stenosis (supra, sub or valvular) or isthmic coarctation with LV hypertrophy and fibrosis CHD with septal defects, repaired or unrepaired, and obstructive pulmonary vascular disease (Eisenmenger syndrome) CHD with perimembranous VSD, postero-­inferior rim stich Corrected GA (unrepaired) Ebstein’s anomaly Atrioventricular anomalous pathway (Kent fascicle) TGA, atrial or arterial switch operation CHD with RV–pulmonary artery conduit repair CHD with univentricular or one and a half repair Ross operation

CHD with septal defect, no obstructive pulmonary vascular disease, repaired or unrepaired Any other unrepaired CHD

Intramural ventricular/septal tumour

Atrial septum lipoma   Congenital partial absence of pericardium

TR IB

BAV and/or isthmic coarctation with aortic dissection

N

Congenital heart diseases

N

O

SE U AV node cystic tumour Purkinje cell hamartoma Sarcoidosis of the AV conduction system Surgical stiches, perimembranous

R

Conduction system diseases

LY

Certain

Note – AV: atrioventricular, BAV: bicuspid aortic valve, CHD: congenital heart disease, CM: cardiomyopathy, LAD: left anterior descending, LBB: left bundle branch, LCA: left coronary artery, LCx: left circumflex, LV: left ventricle, RBB: right bundle branch, RCA: right coronary artery, RV: right ventricle, TGA: transposition of the great arteries, VSD: ventricular septal defect. Source: From Basso et al. (2017).

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Case report 1

SE

O

N

LY

A 37-­year-­old doctor complained for several days of pain in the stomach region and took the medicine Nexium®. In the morning, he came to hospital and was found lifeless in his room. The cause of death remained unclear. The autopsy revealed as cause of death: internal haemorrhage into the abdominal cavity due to ruptured aneurysm of the arteria lienalis. Underlying disease was a fibromuscular dysplasia (Figure 6.2a,b).

U

(a)

(b)

TR IB

U

TO

R

Figure 6.2  Ruptured dissection aneurysm of the arteria lienalis, fibromuscular dysplasia. (a) Elastica van Gieson staining (20:1). (b) Alcian blue staining (200:1).

Case report 2

FO

R

C

O

N

An 18-­year-­old student was found dead in the bathroom of his flat. Neither pre-­existing diseases nor drug or alcohol problems were known. The cause of death remained unclear. The autopsy revealed a thickened right ventricular chamber with a 5 cm in diameter area of subepicardial fatty tissue with severe haemorrhage (Figure 6.3a,b). Cause of death was obviously an arrhythmogenic right ventricular dysplasia.

(a)

(b)

Figure 6.3  (a) and (b) Fatty infiltration of the right ventricular wall with haemorrhage into the subepicardial fatty tissue.

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Case report 3

SE

O

N

LY

A 12-­year-­old boy was playing with his cousin at the street. He suddenly complained of shortness of breath and fell unconscious. Reanimation was unsuccessful. Cause of death remained unclear. The autopsy revealed great scars especially of the ventricular septum (Figure 6.4 a,b). Cause of death was severe myocardial fibrosis.

U

(a)

(b)

TO

R

Figure 6.4  (a) and (b) Great scar of the ventricular septum, severe myocardial fibrosis.

over 85 years and over 100 years of age have shown that, in each case, morphologically ascertainable underlying diseases and immediate causes of death were present. If appropriate, the diagnosed diseases that contributed to the occurrence of death may be descriptively listed as a multifactorial converging type of death, in order to avoid ‘makeshift’ diagnoses. Regarding deaths attributable to medical procedures, the first notable point is a considerable discrepancy between the deaths recorded in the German federal statistics as due to complications of medical and surgical treatment and the data derived from epidemiological research on death cases due to maltreatment. Epidemiological research for Germany revealed that 17,500 deaths per year are suspected to be results of medical malpractice – these figures are consistent with international data – whereas the German Federal Statistical Office mentioned in the year 2007 only 399 deaths as complications of medical and surgical ­treatment; in the year 2017, there were 2120 reported deaths. It  appears clear that there exist a considerable number of ­unreported cases raising the question of whether in relevant ­circumstances the attending physician should issue the death certificate, or whether, irrespectively of the existence or otherwise of suspicion, such cases should always be subject to an official investigation.

FO

R

C

O

N

TR IB

U

death, both the diagnosis of the cause of death and the underlying disease may be difficult. In addition, one may be guided by ‘death types’ that have been described as a ‘thanatological bridge’ between the underlying disease and the immediate cause of death (Figure 6.5): 1. Linear type of death: Underlying disease and immediate cause are within one organ system. 2. Diverging type of death: Organ-­specific underlying cause, but non-­organ-­specific immediate cause. 3. Converging type of death: Underlying diseases in various organ systems lead to death via a final pathogenetic phase common to all of them. 4. Complex type of death: Underlying diseases in various organ systems with more than one non-­organ-­specific immediate cause of death. If the cause of death remains unclear in a case of unexpected death of a healthy person, this should be noted on the death certificate. The Federal Statistical Office recommendations on entering the cause of death and important terms are shown in Table 6.6. When only insufficient information is available to describe the immediate cause of death (e.g., because the patient did not seek medical attendance), descriptive statements should be used such as ‘prostatic carcinoma with lung and bone metastases’ or ‘found dead with a history of severe coronary atherosclerosis and recent myocardial infarction’. Typical errors in death certificates are summarised in Table 6.7, a case illustration with various errors is given in Table 6.8. Finally, particular problems arise with deaths in old age or in connection with medical procedures. ‘Senility’ or ‘old age’ are not causes of death. Retrospective examinations of deaths of those

COVID-­19-­associated deaths Concerning the Corona pandemic in 2019–2020, the mortality rate in different countries differed widely. The mortality rate was

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Linear dying type Severe coronary atherosclerosis and thrombosis

Myocardial infarction

Death

Cardiac tamponade

Linear dying type with a vascular disease and cardiac cause of death, 75-year-old male Suicidal ingestion of amitryptiline

Fatal concentration in femoral blood and several organs

No other pathological autopsy findings

Linear dying type with a drug overdose as cause of death, 43-year-old female Pulmonary oedema, brain oedema, fluid cadaveric blood

Fatal morphine concentration

N

Linear dying type with a drug overdose as cause of death in a 25-year-old male

O

Diverging dying type

Tumour anaemia

Example: 45year-old woman with metastatic cancer and unspecific cause of death

SE

Severe metastasis Carcinoma of gall bladder

Death

Tumour cachexia

U

Tumour intoxikation

LY

i.v. ingestion of heroin

R

Converging dying type

TO

Severe coronary atherosclerosis

Acute coronary insufficiency

Stomach ulcer with recurrent bleeding

Death

Example: 63-yearold man with vascular and pulmonary disease – cerebral as well as pulmonary cause of death

TR IB

U

Chronic emphysema and bronchitis

Death

Example: 79-yearold man with vascular, gastric and pulmonary disease

Complex dying type

N

Arterial hypertension Severe atherosclerosis of the basal cerebral arteries

O

Chronic emphysema bronchitis

Encephalomalacia

Bronchopneumonia

R

C

Figure 6.5  Types of death. Source: According to Leis (1982) and Thieke and Nizze (1988); case examples from Thieke and Nizze (1988) and Madea (2019).

FO

Table 6.6  Causes of death: examples and important aspects (according to the German Federal Statistical Office recommendations). Cause of death

Comments

Pneumonia

Primary, hypostatic, aspirational, other underlying causes Pathogen If seen as a consequence of immobility or debility: the cause of immobility or debility

Infection

Primary or secondary, pathogen If primary: bacterial or viral If secondary: more information about the primary infection required

Urinary infection

Localisation in the urinary tract, pathogen, other underlying causes If a consequence of immobility or debility: the cause of the immobility or debility

Renal failure

Acute, chronic or terminal; underlying cause, for example, hypertension, atherosclerosis, cardiovascular disease If a consequence of immobility or debility: the cause of immobility or debility (Continued)

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Table 6.6  (Continued) Comments

Hepatitis

Acute or chronic, alcohol-­related If viral: type A, B, C, D or E

Infarction

Atherosclerotic, due to thrombosis or embolism

Thrombosis

Arterial or venous, including vascular designation Intracranial sinus, purulent, non-­purulent, venous (state which vein) If postoperative or in an immobile patient: disease that was the reason for surgery or immobilisation

Pulmonary embolism

If younger than 75 years: cause If postoperative: disease that was the reason for surgery or immobilisation

Leukaemia

Acute/subacute/chronic Lymphatic/myelogenous/monocytic

Alcohol/medical drugs/ narcotics

Long-­term abuse or drinking Dependency

Complication of surgery

Disease that was the reason for surgery

Dementia

Cause (e.g., senility, Alzheimer’s disease, multiple infarctions)

Accidental death

Details of circumstances (e.g., cyclist hit by a car) Accident, suicidal, violent assault, circumstances unknown Accident site (e.g., on street, at home); activity at the time of death (playing golf, going to the movies, working)

Tumour

Benign, malignant, location, metastases

U

Table 6.7  Definition of major and minor errors in death certificates.

TO

R

U

SE

O

N

LY

Cause of death

Definition

TR IB

Type of error

Major

Mechanisms or non-­specific conditions listed as the underlying cause of death

Improper sequencing

Sequence of events does not make sense; underlying cause of death not listed on the lowest completed line in part I

O

N

Mechanisms of death listed without an underlying cause

Two or more causally unrelated, aetiologically specific diseases listed in part I

C

Competing causes

Abbreviations

R

Minor

Abbreviations used to describe diseases No time intervals listed in Part I or Part II

Mechanism of death followed by a legitimate underlying cause of death

Use of a mechanism, but qualified by an aetiologically specific cause of death

FO

Absence of time intervals

Source: From Myers and Farquhar (1998).

high in Italy and Spain, but comparatively low in Germany. For Heinsberg, a small city in North Rhine-­ Westphalia and one ­hotspot of Corona virus infection, a mortality rate of 0.37% was calculated by virologists investigating this hotspot which was much below the mortality rate of 1.98% calculated by the Johns Hopkins University for Germany.

Concerning the dying type, most people had severe pre-­ existing diseases (converging or complex dying type). Deaths of healthy people without pre-­existing diseases were obviously not observed or very rare. The case fatality ratio (CFR) (ratio between number of infections to fatalities) was with 12.8% very high for Italy, comparatively low with 2.2% for Germany.

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Table 6.8  Case illustration showing types of mistakes in stating the cause of death. A 75-­year-­old male smoker with a 5-­year history of pulmonary emphysema is admitted to hospital because of an exacerbation of his lung disease. In hospital, Haemophilus influenzae pneumonia is diagnosed. Independently of this, he has a 10-­year history of coronary heart ­disease. While in hospital his condition worsens, but he does not wish to be intubated and artificially ventilated. One week after admission, he is found dead in his bed. Four alternative ways of staging the cause of death on the death certificate are shown (A–D). In this case, only ­alternative D is correct.

Finding(s)

Approximate time interval from onset of disease to death

(a)

Respiratory arrest

–

Functional end status given, time interval from onset of disease to death omitted

(b)

–

–

–

(c)

–

–

–

Part II

Coronary heart disease

–

–

(a)

Emphysema

–

(b)

Pneumonia

–

(c)

–

–

Part II

Coronary heart disease

–

Disease leading to death

Mistake type

U Emphysema

(b) Part II

R TO

N

–

5 years

Competing causes of death that have no causal connection

Coronary heart disease

10 years

–

–

–

–

Haemophilus influenzae pneumonia

1 week

–

Emphysema

5 years

–

–

–

–

Coronary heart disease

10 years

–

O

N

D Part I

C

(a)

R

(b)

FO

(c) Part II

–

TR IB

(a)

Incorrect sequencing of the underlying cause and the immediate cause of death, time intervals absent –

U

C Part I

O

SE

B Part I

LY

A Part I

Source: Case scenario adapted from Myers and Farquhar (1998).

Typical causes of death in COVID-­19 infection cases are, for instance: 75-­year-­old male

86-­year-­old male Ia

Acute respiratory acidosis

1 day

Ic

COVID-­19 infection

10 days

II

Diabetes mellitus Adipositas permagna

10 years

Ia

Acute respiratory distress syndrome (ARDS)

2 days

Ib

Ib

Pneumonia

10 days

Ic

COVID-­19 infection

14 days

Id

Chronic obstructive pulmonary disease (COPD) grade 4

years

II

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PART II  

MEDICAL ASPECTS OF DEATH

the underlying disease and code this underlying disease in accordance with the International Classification of Diseases regulations. Against the background of increasingly multifactorial death processes, however, monocausal representations of death can only partially fulfil the requirements of cause of death statistics and the data derived from them about indicators for health.

51-­year-­old female Ia

Brain haemorrhage

2 days

Ib

Cerebral metastasis

4 weeks

Ic

Mammary carcinoma

3 years

II

Diabetes mellitus, COVID-­19 infection

  (14 days)

6.4  Consistency between cause of death diagnosis on the death certificate and following autopsy

Cerebral haemorrhage, pneumonia

Ib

Brain metastasis, COVID-­19 infection

(4 weeks)

Ic

Mammary carcinoma

3 years

II

Diabetes mellitus

Numerous studies have been published on the validity of clinically determined causes of death as entered on the death certificate in comparison to pathoanatomical findings. The Görlitz study (1986–1987), with a nearly 100% autopsy rate (1060 deaths, in 1023 of which a postmortem was carried out), showed incongruity between certificate and autopsy findings in totals of 45% of male deaths and 48.8% of female deaths. Among hospital deaths, there were incongruities of the underlying disease in 42.9% of men and 44% of women; among deaths in care homes, the corresponding figures were 63.2% of men and 57.8% of women; and for deaths occurring elsewhere (at home, in public, etc.), the rates were 41.3% for men and 50.7% for women. Among iatrogenic deaths, the rate of incongruities between underlying diseases as determined clinically and found at autopsy was as high as 72%, and inconsistencies for immediate causes of death were 45.8%. Numerous studies have differentiated and operationalised the discrepancies between clinically determined causes of death and those determined at autopsy (major mistake, class 1; major mistake, class 2; minor mistake; Box 6.1). According to various statistics, class 1  major mistakes having consequences for treatment and survival of the patient occur in 11–25% of all deaths, while class 2 major mistakes, with no consequences for treatment and survival, are found in 17–40% of all death cases.

FO

R

C

U

O

N

TR IB

U

TO

In 2017, 932,272 deaths were reported in Germany; according to the Federal Statistical Office, in about 96% of these, the causes of death were natural. Even just for the location where death occurred, there are no uniform data for the whole of Germany; but more than 50% of deaths today occur in hospital (according to own data), about 25% at home and around 15% in care homes. The remaining 10% are divided among transport accidents, work accidents and so on. In 2017, of 19,952,735 inpatient admissions, 7,606,379 of these were cases belonging to the field of internal medicine. A large number of admissions (the second largest) – 4,369,706 patients – were done to the departments of surgery. Within internal medicine departments, most deaths were seen in cardiology, followed in descending order by gastroenterology, haematology and geriatrics. Out of a total of 932,272 deaths in 2017, 344,524 were due to cardiovascular conditions, most common of which was ‘ischaemic heart disease’. The second most common group of causes of death was ‘malignant neoplasms’, with 235,681 deaths. It should be borne in mind that deaths for the various disease groups vary considerably among different age groups. Up to an age of 40 years, unnatural death is more frequent than death from natural diseases (internal cause of death), but beyond this age, deaths from malignancies and cardiovascular diseases become more frequent than unnatural deaths. These data from the German Federal Statistical Office are taken from the coding of the entries in the death certificate on the underlying disease(s) and immediate causes of death, and it is only the underlying disease which is taken into account in the current cause of death statistics. On the contrary, in the state (land) statistical offices, the statement on the underlying disease is not automatically used for the cause of death statistics, but the coders examine the entries on each death certificate, determine

R

6.3  Causes of death as shown by cause of death statistics

SE

O

N

Ia

LY

51-­year-­old female

Box 6.1  Discrepancies between causes of death determined clinically and at autopsy. • Major mistake, class 1 ○○ Clinically missed diagnosis that proves at autopsy to have been the underlying cause of death and/or the main immediate cause of the patient’s death. If the diagnosis had been made on time, the patient’s life could have been prolonged, at least for a time. • Major mistake, class 2 ○○ Clinically missed diagnosis that would not have affected the management and course of the disease had it been made before death. • Minor mistake ○○ Diseases or medical facts discovered during autopsy that have no direct causal connection with the underlying or immediate cause of death. Source: From Shojana et al. (2002).

Certification of Death: External Postmortem Examination

No. (%)

Infection • Fungal pneumonia (1 case with coexisting fungal myocarditis) • Viral pneumonia • Bacterial pneumonia • Miliary tuberculosis and bacterial meningitis • Meningitis of unknown aetiology • Toxoplasmosis involving lung, liver and brain • Candidemia confirmed by postmortem blood culture • Renal abscess

15 (45.5) 6

C

O

N

TR IB

U

Item and findings

Pulmonary embolism

2 2 1 1 1 1 1

  8 (24.2)   6 (18.2)  3        2      1

Cardiovascular • Retroperitoneal and/or intraabdominal haemorrhage • Cartilaginous emboli

  3 (9.1)  2  1

Immunologic • Anaphylactic laryngeal oedema

  1 (3.0)  1

Total

33 (100)

FO

R

Malignancy • Lymphoma involving multiple organs ○○ Diffuse large B-­cell lymphoma ○○ Follicular lymphoma ○○ Peripheral T-­cell lymphoma • Pulmonary carcinoma ○○ Small cell carcinoma ○○ Squamous cell carcinoma • Gastric adenocarcinoma

Source: From Marshall and Milikowski (2017).

LY

6.5 Manner of death

SE

O

N

According to cause of death statistics, around 3.8% of deaths in Germany result from unnatural causes (Figure  6.6). More than 9200 cases per year are due to suicides, 11,000 due to accidents at home, just over 3300 result from traffic accidents and 352 deaths from physical assaults. Retrospective analyses of death certificates for which the manners and causes of death have been checked at autopsies suggest that unnatural deaths are around 33–50% more frequent than it is reflected by the federal statistics, and that it could be assumed that there are around 81,000 unnatural deaths every year. From the judicial point of view, a particular concern has to be the number of homicides that remain undiscovered by means of medical death certification. A multicentre study suggests that every year around 1200 homicides remain unidentified on death certificates in Germany (Brinkmann 1997). This large number of unrecorded cases was repeatedly confirmed by incidental findings of homicide or even serial murders (including those in care homes and hospitals). Six percent of hospital physicians regularly attest exclusively a natural death; 30% tick the box for natural death even in cases of violence, poisoning, suicide or following medical intervention. In assessing the manner of death, the certifying physician decides whether a death will ever come to the attention of the investigating authorities. Assessing the manner of death is thus an extremely important task not only from the judicial point of view (detection of homicides), but also in terms of the interests of the bereaved (for example, compensation claims after a fatal accident). ‘Natural’ is a death from an internal cause (disease), where the deceased person has suffered from a disease that can precisely be identified and from which death was anticipated; the death occurred entirely independent from any external factors of legal significance. The prerequisite for attesting a natural death is thus the existence of an underlying disease of death known from the patient’s medical history with a poor prognosis as to survival. ‘Unnatural’, by contrast, is a death attributable to an event caused, triggered or influenced from outside, irrespective of whether due to the fault of the patient himself or herself or of a third party. Unnatural deaths, therefore, are those due to: • Physical assault. • Accident (irrespective of whose fault).

TO

Table 6.9  Findings with potential impact on survival or treatment (class 1 discrepancy).

81

The concept of diagnostic error would correspond to the class 1 major mistake. A diagnostic error is assumed to have occurred when, at the end of the diagnostic decision process, a disease is definitely presumed to be present in a patient when in fact it later proves not to be so. Furthermore, when a treatment is initiated that is not appropriate for the disease identified at the later date, and the failure to recognise the disease that is actually present has led to a worsening of the patient’s prognosis, diagnostic error is assumed.

U

According to a meta-­analysis by Shojania et  al. (2002), class 1 major mistakes have decreased in the past 40 years, but do still occur in about 8–10% of deaths. Even recent evaluation of autopsy-­detected errors (class 1 discrepancy) revealed 10% class 1 errors (Table 6.9). However, the rate of agreement or disagreement between clinically and autopsy-­determined causes of death depends on many variables, such as: 1. The definition of the cause of death. 2. The evaluated disease class. 3. Age. 4. The patient group under investigation (outpatient, inpatient, specialised hospital). 5. The duration of the hospital stay. 6. The predictability of the death (expected vs. unexpected). 7. The autopsy rate. No comparison between clinically and autopsy-­determined causes of death taking into account these variables in a differentiated way has yet been carried out, and none is to be expected under the current regulatory framework (at least in Germany) on performing clinical autopsies. This is particularly true for ­outpatient deaths, which are almost never subjected to autopsy except when ordered by the courts.

R

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Trauma, poisoning, and certain other consequences of external causes/unnatural causes of death (30 571)

Accident at home (6361) Intentional self-harm (9402) Violent assault (451) Accident at work or at school (471)

827 155

30 571

Traffic accident (5011) Sports accident, accident at play Natural causes of death (796 505) (145)

N

LY

Other accidents and all other entries (8730)

O

Figure 6.6  Causes of death, showing the proportion of unnatural causes of death among deaths in Germany 2017. Source: Courtesy of the Federal Statistical Office, Germany (www.destatis.de; last accessed 25 March 2020).

U

SE

assumes that a clear cause of death can be given. In this regard, it is worrying that about 50–70% of physicians certify a ‘natural’ death for death following femoral neck fractures, 20% for deaths during injections and 30–40% for intraoperative deaths. If on the one hand unnatural deaths are considerably underrepresented in official statistics, and, on the other hand, both physicians in private practice and those working in emergency departments report attempts by the police to influence them to certify a death as natural although no cause of death is apparent, the death ought to be certified at least as unexplained. In an anonymous survey of randomly selected physicians, 41% of physicians in private practice and 47% of emergency room physicians reported such attempts to influence death certification. The background of these attempts is that investigating authorities have a teleologically narrowed understanding of the term ‘unnatural death’ as meaning ‘death in which there is a possibility of third-­ party guilt’. If a natural death is certified, no further investigations are necessary. Indicators that a death may be unnatural may arise from the case history and findings: for example, sudden death without known previous illness, prima facie accidents and suicides, presence of farewell letters and so on. Findings that tend to indicate unnatural death include conjunctival haemorrhages, unusual colour of livor mortis, remains of tablets in the oral area and signs of injury. Unsuitable criteria for indications of natural death are age, especially when no pre-­ existing life-­ threatening diseases are known, and the absence of visible trauma. Regarding deaths in hospital, especially when the patient was under medical treatment for a period long enough, the error rate should also be relatively low; problem areas include failure to identify causal connections to trauma at the beginning of the fatal causal chain, and deaths related to medical procedures. In the inpatient setting, there are occasional reports of initially unrecognised series of killings by physicians or nursing personnel. The danger of errors and scope for deception are without doubt greatest when death is certified by private practice

FO

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N

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Homicide. Poisoning. Suicide. Treatment errors/medical malpractice. Fatal consequences of any of the first six points. The interval between an external event at the beginning of the causal chain that leads to death and the occurrence of that death can thus be indefinitely long (it may be years). If the cause of death cannot be ascertained when the death certificate is issued, the manner of death will therefore also remain unclear. Various regulations relating to the certification of death in different states (länder in Germany) and a draft outline for a federal-­ wide death certification process from the German Medical Association envisage explicitly that attestation of a natural death requires examination of the unclothed body. Section  3 of the Bavarian Interment Regulations, for example, says: ‘. . . determination of a natural death requires in every case that the medical external examination on which the death certificate is based be carried out with the body of the deceased completely undressed. This examination of the completely undressed body shall include all regions of the body including all body orifices, the back, and the scalp’. Sensible though this requirement is, there is no doubt that it is regularly disregarded. If the physician fails to meet the required standards of thoroughness, however, he or she has committed a regulatory offence. Alternatively, it must be recognised that completely undressing a dead body in cases of expected death in hospital will not lead to any gain of further information and can face the certifying physician with objective problems (e.g., when complete rigor mortis is established, and no support personnel are available to help). Furthermore, this requirement fails to take into account the difference between expected and unexpected deaths. The manner of death remains undetermined if the cause of death cannot be identified on examination even with the help of the medical history. The attestation of natural death always

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• • • • •

Certification of Death: External Postmortem Examination

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N

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Mistakes: • Giving the functional status as cause of death without relating it to an underlying cause. • Incorrect sequence from the final cause of death to the underlying cause. • Time intervals not stated. • Overlooking a causal connection with an external event (e.g., trauma) at the beginning of the causal chain that led to death. • Inadequate or misinterpreted clinical information. • Efforts to conceal diagnoses that might cause uneasiness or ­distress to family members.

deaths (compared to autopsy rates of 20–30% in England, Scotland and Wales, Sweden and Finland). These autopsies, which are required for valid cause of death statistics and for the planned National Mortality Register, would, however, have to be remunerated adequately, which unfortunately they are not at present.

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Problems that may repeatedly arise within the hospital context are: 1. Deaths in connection with medical interventions. 2. Deaths following injury from a fall or other violent events, in which the causal connection to violence from another person or another external event is not identified and therefore death is wrongly certified as natural. For deaths occurring unexpectedly linked to medical interventions, the manner of death should always be certified as ‘unexplained’, so that an official autopsy can be carried out to investigate objectively the underlying and immediate cause(s) of death. This is the only basis an opinion can be expressed on any question of  maltreatment. Certifying the manner of death as unexplained or unnatural does indeed not signify an admission of maltreatment. For physicians in private practice, the main problems may arise when bodies are found at home, patients die unexpectedly and with deaths occurring at old age. If the cause of death cannot be established from external examination of the body or from interviewing any doctors previously involved in treatment, this should be recorded and the manner of death certified as unexplained. With old people, there is always the question whether or not case history and severity of the diagnosed disease(s) explain the occurrence of death at this particular moment. Mistakes and risks of the medical examination of a body are summarised in Box 6.2. Whenever the cause of death cannot be established by external examination, an autopsy should be carried out, as this is still usual in many countries in Europe. In Germany, the present autopsy rate is less than 5% of all deaths; the rate of hospital autopsies in particular has been dropping sharply in recent years, while judicial autopsies have remained relatively stable at 2% of

Beware: • Never certify a death in the absence of definite sign(s) of death. • Examine the unclothed body carefully. • Review the patient’s medical history: ○○ What was the clinical diagnosis? ○○ What was the sequence of events leading to death? ○○ Can manner and circumstances in which death occurred be explained by the confirmed clinical diagnosis? ○○ How certain are the diagnoses of the underlying and immediate causes of death? • Was there any external event at the beginning of the causal chain that led to death? If yes, the death should be certified as unnatural. • If death occurred in the context of medical interventions, the manner of death is unexplained or unnatural. • If the cause of death cannot be established even after interviewing doctors who previously treated the patient, it remains unclear, and so does the manner of death. • Resist attempts to be influenced how you complete the death certificate.

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6.5.1  Problem areas

Box 6.2  Errors and risks of the medical examination for death certification.

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­ hysicians at home; typical mistakes and sources of error, in p order of frequency, are: • Inexperience. • General carelessness. • Careless examination of the body. • Consideration of the emotional state of relatives. Furthermore, there are sometimes also unfavourable external conditions, such as poor lighting and simply not being adequately trained for the job, or that there was no possibility of calling an appropriately qualified certifying physician. Particularly physicians in private practice can find themselves being confronted with a collision of interests, since they are also the physician of the relatives of the deceased. Attesting an unexplained death puts them at risk of triggering investigations that could lose the relatives as patients. Compared to a physician working in private practice, the hospital physician is in a more protected position (death in the medically dominated environment of a hospital rather than in the private area).

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6.6 Special constellations of circumstances in external postmortem examination 6.6.1  Unexpected death during hospitalisation About 2–2.5% of all hospitalised patients die at hospital. In Germany, of 17.5  million hospitalised patients, around 400,000 die each year. Death cases are mainly due to severe underlying diseases or accidents that have led to hospitalisation. However, unexpected deaths occur at hospital too, due

MEDICAL ASPECTS OF DEATH

cases are expected to be homicides or around the disposal of a homicide victim in the bathtub. As causes of unexpected deaths, carbon monoxide poisoning, drug intoxications and electric devices being used in the bathtub play important roles. Electric burns may be absent in bathtub deaths. However, porcelain white stripes and a demarcation of postmortem lividity according to the water level may be seen. Even violent deaths in bathtubs may show only few if any external injuries. In a well-­known case of crime history, the offender drowned his victims (his wives who trusted him) when they took a bath by taking their legs and pulling them out of the bathtub until their heads were under water. In cases of deaths in the bathroom, therefore, the following checklist may be helpful.

1. Drug, sources of carbon monoxide, water in the bathtub

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(water level, water running, water temperature, bath additives). 2. If the bathtub is empty, lines of marks in the bathtub ­corresponding to the previous water level. 3. Electric devices near to or in the bathtub. 4. Switch position of hairdryers or room heaters. 5. Farewell letters in the premises.

TO

Death in police custody occurs mainly in males within the fourth to fifth decade of life. Causes of death range from sudden natural death (especially due to cardiovascular diseases) to suicides (mostly by hanging), undiagnosed head injuries and intoxications (alcohol intoxication or mixed alcohol–drug intoxication) or combinations of both. Special risk factors for deaths in police custody are higher age, pre-­existing natural diseases, injuries, blood alcohol concentrations higher than 300  mg/100  ml and combined alcohol–drug intoxication. Therefore, prior to imprisonment, a thorough investigation to determine the fitness of the individual to be detained in custody has to be carried out. People under the influence of alcohol may show impaired consciousness. In cases with additional head trauma, a differential diagnosis between impaired consciousness due to alcoholisation or head trauma is important.

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Findings at the scene

6.6.2  Deaths in police custody

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to as yet diagnosed diseases, as for example in acute coronary insufficiency or pulmonary embolism. Special problems include diagnostic or therapeutic misadventures. When unexpected deaths occur during hospitalisation, especially in connection with medical procedures, the manner of death should be classified as unnatural or unascertained and the police should be notified. Other types of unnatural deaths during hospitalisation are accidental falls, deaths during fixation, scaldings when persons are being left unattended in bathrooms and also suicides (in particular, in psychiatric hospitals). However, homicides, even serial homicides, occur in hospital, committed by doctors and nurses. Therefore, each case of an unexpected death during hospitalisation requires a thorough investigation.

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Examination of the corpse

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1. Basic examination: clothing, postmortem lividity, rigor ­mortis,

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6.6.3  Deaths in prison

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Of deaths in prison, 30–40% are due to natural diseases, mostly cardiovascular diseases. Comparatively rare are deaths due to accidents by intoxication. Homicides occur only very rarely. Of greater importance are suicides (mainly by hanging) that are particularly committed by male prisoners in the third decade of life and at the beginning of imprisonment in single cells. Clothing, belts, pieces of bedclothes and electric cables are often used for strangulation and windows and parts of sanitary installations or heating pipes for suspension. Even in prison cells occupied by more than one prisoner, unnoticed suicides can be observed.

6.6.4  Death in the bathroom Unexpected deaths in the bathroom are a particular challenge, since only 10–30% of these cases are due to natural mechanisms. Of particular importance are accidents and suicides, but 5% of

body core temperature (rectal temperature), water temperature. 2. Signs of water immersion such as washerwomen’s skin. 3. Signs of drowning (froth extruding from mouth and nose). 4. Findings that are related to possible intoxication. 5. Electrical burns: blisters, ‘linear electric burns’, linear demarcation of postmortem lividity according to the water level. 6. Death due to pre-­existing diseases such as epilepsy with seizures (tongue bites). 7. Injuries which are not compatible with the findings at the scene.

6.6.5  Deaths in psychiatric hospitals There is a higher suicide rate of mentally ill patients, especially due to depressive and schizophrenic disorders, in psychiatric hospitals. They are mainly committed by younger patients in the third to fourth decade of life. Hanging is one of the most frequently used methods followed by intoxications and falling from height. If suicide by jumping from height is suspected, the horizontal distance between the final position of the body and the point where the person jumped off has to be measured and checked for plausibility.

Certification of Death: External Postmortem Examination

Table 6.10  Findings at the external postmortem examination in cases of intoxication. Possible toxic substances

Skin bleedings

Each poison that may lead to damage of liver parenchyma; in particular, phosphorus, Amanita phalloides

N

Hydrogen sulphide, cyanide, ammonia

‘Holzer’ blisters Miosis*

Mydriasis*

Hypnotics (barbiturates)

So-­called MnOP toxins: morphine, opiates, nicotine, phosphoric acid esters, physostigmine, pilocarpine, prostigmine, barbiturates So-­called ABC toxins: ethanol, Amanita muscaria, Amanita pantherina, atropine, cannabinoids, quinine, cocaine, colchicine, cyanides, methanol, scopolamine

In particular lyes, acids but also halogens, phenol and its derivates, paraquat, trichloroethylene

Salivation

Phosphoric acid esters, Amanita muscaria

Light red nails

Carbon monoxide

U

Acid traces

Livor mortis:

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N

The suspicion of poisoning depends on three circumstances: case history, conditions under which death occurred and findings at the external postmortem examination. Intoxication as a cause of death has to be taken into account under the following circumstances: 1. Young healthy people without any medical history. 2. Simultaneous ‘sickness’ of several people. 3. Mentally ill people. 4. Drug addicts. 5. Third parties that might be interested in the death of an individual (e.g., elimination of a relative who is perceived as a burden for carers, rich testators, high value of life insurance, enemies, rivals). 6. People who have good access to poisons (e.g., chemists, biologists, physicians, nurses, photographers, goldsmiths, etc.). Symptoms that may be a hint for intoxications are: 1. Sudden and unexpected illness, sudden collapse and immediate death. 2. Seizures, agitation, loss of consciousness, coma, sudden unresponsiveness. 3. Respiratory depression, dyspnoea, apnoea, snorkelling. 4. Changes of the diameter of the pupil and salivation. 5. Froth in mouth and nostrils. 6. Abdominal pain, emesis or diarrhoea.

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Odour

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6.6.7  Death by poisoning

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Finding

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Death at the steering wheel may be due to accidents, suicides or pre-­existing natural diseases. Suspicion for suicidal death is raised if the cause for an accident is not evident; for example, driving on a straight street with high speed against the pillar of a bridge with the safety belt unused. In case of sudden death behind the steering wheel, there are often only minor injuries seen on the deceased, and there may be a typical driving pattern, such as striking parked cars, collision with oncoming vehicles, drifting to the opposite lane or attempts to stop the car. Sudden natural death at the steering wheel mostly affects men within the sixth to seventh decade of life dying due to ischaemic heart disease. Electrophysiologically, fatal cardiac arrhythmia develops over a period of about 2 minutes. Warning symptoms due to damage of the cerebral function are recognised and the car driver tries to stop the car to avoid severe accidents. If serious injuries are found following unexplained accidents at external postmortem examination, an autopsy should be carried out to clarify two substantial questions: 1. Is there any internal natural disease that possibly caused the accident? 2. Is there an internal disease that may have contributed to death compared to the accident-­related injuries?

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6.6.6  Deaths at the steering wheel

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External findings of the external postmortem examination are mostly non-­specific, but there may be certain changes that might indicate intoxication (Table  6.10). Furthermore, great attention should be paid to findings at the scene of death (e.g., glasses or bottles sometimes with remnants of substances, drug containers, warning colours of poisons, vomit, etc.) (Figure 6.7a,b). Besides poisoning by oral ingestion, poisoning via the vaginal or anal pathway occurs, both in cases of suicides and homicides.

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When thick clothing was worn and the ground was soft, there might be a significant discrepancy between the absence of externally visible injuries due to the blunt force impact and extensive injuries revealed at autopsy.

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• ash grey

Methanol

• brownish

Methemoglobin-­former (e.g., nitrites, nitrobenzene, chlorates, soaps, aromatic amino compounds)

• light red

Carbon monoxide, cyanides

Several injection sites

Opiates

General icterus

‘Liver toxins’, phosphorus intoxication, mushroom poisoning

Mees’ lines

Arsenic, thallium

Dark gingival seam, stomatitis

Lead, mercury, bismuth

‘Paw’ position of hands

Hydrocyanic acid, strychnine, phosphoric acid esters

Hair pulled out easily

Thallium

 Because of agonal (hypoxia of the brainstem) and postmortem changes only of

*

limited value.

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MEDICAL ASPECTS OF DEATH

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(a)

(b)

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Figure 6.7  Warning colours in suicidal intoxication. (a) Combined suicide by oxydemeton-­methyl and hanging, as evidenced by the blue stain flow from the left ankle of the mouth and strangulation mark. (b) Intensive blue colour of mouth and lips in a case of suicidal intoxication with solvents.

6.7.2  Date and time of the external postmortem examination

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6.6.8  Discovery of several corpses

These should be noted on the patient’s medical notes.

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If more than one body is found at the scene of death or when a body and a dead animal or a body and another person in poor health are found together, an unnatural death should be assumed unless the contrary is proven. Coincidental deaths of two people at the same time represent an absolutely rare event. When several people are found dead at one scene, the following constellations are possible: • Homicide. • Homicide with survivor. • Homicide with suicide of the offender. • Homicide with attempted suicide. • Suicide, extended suicide, suicide with survivor. • Accident. • Death due to underlying natural diseases.

6.7  Checklist for the external postmortem examination 6.7.1  Who initiated external postmortem examination/who notified doctor 1. The bereaved (in cases of death at home). 2. Head of an institution. 3. Medical staff. 4. Failed emergency aid. 5. Police (somebody found dead).

6.7.3  Place of the external postmortem examination 1. In hospital:

a. Death was expected; b. Death was unexpected. 2. At home: a. Death was expected; b. Death was unexpected; c. Discovery of corpses. 3. In public: a. Accident; b. Suddenly collapsed; c. Emergency aid; d. Discovery of corpses.

6.7.4  Description of the surroundings where the body was found 1. Outdoors or indoors (doors and windows opened or closed, describe type and way of locking).

2. Inside and outside ambient temperatures, weather conditions; is the heating system running or not?

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Certification of Death: External Postmortem Examination

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3. Finding of corpses indoors: what kind of room is it?

5. External cardiac massage. 6. Complications (rib fractures, misintubation, pneumothorax). 7. Defibrillation.

6.7.5  Description of the immediate surroundings next to the corpse

6.8.3  Position of the corpse

1. Condition of the premises (e.g., tidy, neglected, searched, etc.). 2. Information about use of alcohol, illicit drugs, pharmaceuti-

tilt.

2. Arms/legs stretched, bent, splayed out. 3. Gender, age (estimate if necessary). 4. Body length, body weight, nutritional status. 5. Body adhesions: blood, faeces, sperm, dirt (locations). 6. Trace flow of blood and secretions (course, dried).

N

cals (e.g., bottles, cans, drugs, prescriptions, utensils of drug abusers). 3. Weapons, tools for strangulation lying next to the corpse, pools of blood or traces of blood.

1. Supine position, prone position, lateral position, head-­down

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What is the position of the corpse? Describe clothing present.

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6.7.6  Information about diseases

6.9  Examination of the corpse

Health insurance certificates, drugs, prescriptions.

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6.9.1  Postmortem changes 1. Lividity (livor mortis): position, colour (pink, cherry-­red: car-

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6.8  Identification of the corpse

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1. The individual is known to the doctor. 2. Identity documents of the deceased (passport, driving licence)

bon monoxide; cold environment, brownish-­red: methaemoglobin; normal colour: bluish-­purplish), intensity, expansion (little expansion: internal/external blood loss; anaemia). 2. Rigidity (rigor mortis): determine the degree of rigor mortis in finger joints and larger joints (present/not present, doughy/ smooth, hard, breaking the rigidity manually not possible, reappears after breaking). 3. Drying: of lips, genitals, conjunctivae, extremities. 4. Supravital reactions: determine idiomuscular pad, electric excitation of skeletal muscle, measure deep rectal temperature. 5. Putrefaction: greenish discolouration of abdominal skin, epidermal detachment, skin blisters, facial/abdominal/scrotal swelling with gas; superficial veins of the skin are visible, putrefactive fluids in mouth and nostrils, hair pulls out easily, nails fall off. 6. Corpse fauna: fly eggs in nostrils, palpebral fissures, corners of the eyes, corners of the mouth or penetrating skin lesions; fly maggots (lengths); fly pupation, pupae; empty pupal shells. 7. Is the degree of postmortem changes compatible with indicated time of death?

present.

3. Family members/police/others can give information about

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identity.

6.8.1  Condition of clothing

TR IB

4. Identification is impossible.

N

1. Arranged or disarranged. 2. Buttons are in buttonholes, buttons are torn out or button tape

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is damaged.

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3. Zippers are unzipped or closed. 4. Kind of outerwear/underwear and footwear. 5. Damage and dirt of clothing and footwear. 6. Stains of blood, secretions on fabric of clothing or tissue. 7. Scratches on footwear/soles. 8. Watches, jewellery, contents of pockets. 9. Modification of clothing during external postmortem examination (clothes are cut, torn).

6.8.2 Resuscitation 1. Situation when emergency doctor arrived. 2. Medical actions during resuscitation (see records of rescue services).

3. Injections (note injection sites to differentiate between earlier and recent ones).

4. Intubation (difficulties, complications).

6.9.2 Systematic examination of the corpse 1.  Odour: press on costal arch, take a smell at mouth and nose (alcoholisation: aromatic odour; hydrogen cyanide: bitter almond odour; E 605: garlic-­like odour; uraemia, acetone). 2.  Oedema of the lower legs. 3.  Pressure marks on knees/ankles: ‘Holzer’ blisters in cases of hypnotic intoxication with sedatives and barbiturates. 4.  Scars: scars on the inner wrist if attempted suicide in the past, surgical scars, scars after cut/stab wounds.

MEDICAL ASPECTS OF DEATH

TR IB

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N

11. 

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9.  10. 

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8. 

N

7. 

b. Perioral/perinasal dryings/reddening of the skin; c. Occlusion of the mouth and the nose. 14.  Neck: injuries (dryings, skin bleedings, subhaemorrhages, scratches of the epidermis, tools for strangling found around the neck, ligature furrow, ligature mark): horizontal mark ascending to one side or to the nape of the neck, similar or different ligature impressions, double ligature marks, subcutaneous bleeding in the middle of the ligature mark. 15.  Trunk/thorax/extremities: a. Injury signs: dryings, abrasions, haemorrhages, subcutaneous haematomas penetrating skin lesions, abnormal mobility (cervical spine: pull head gently and turn it to each side); b. Pelvic ring (press/palpate spina iliaca anterior superior and symphysis); c. Arms: fingertip bruising on the inner upper arms, defensive wounds on the extensor side, on forearms (ulnar/little finger side), on the backs of the hands; abrasions on the backs of the hands due to atonic falls; gunshot residues, blood splatters, injuries caused by the slide of a weapon in cases of suicidal shot injury. 16.  Electric burns: on hands, flexor aspects of fingers, soles, toes. 17.  Anus, genitals: anal bleeding, genital bleeding (signs of injury, foreign objects, adhesion of secretion, sperm, faeces). 18.  General status of care and nutrition (important in cases of neglect: infants, toddlers, disabled, elderly, invalids). 19.  Decubital ulcers, pressure sores: localisation, size, status of care.

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6. 

forearm, back of the hand, and also interdigital skin between fingers and toes, mucosa of the oral vestibule, tongue, groin, penis; older injection sites and injection sites of different ages arranged like Baily’s beads (‘shoot bands’), skin abscesses or scars thereof. Signs of pregnancy: darker aureolae mammae, yellowish fluid running out of mamilla when squeezed, striae on lower abdomen and thighs; palpatory findings over abdomen, fundal height. Hairy scalp palpation: swelling, haematoma, injuries of the scalp, palpable crepitation. Facial: lesions of prominent parts (eyebrows, zygomatic arches, nose, chin – injuries due to falls when sudden death occurred), bleedings, eyelid swelling (monocular haematoma, lips and mucosa of the oral vestibule with lacerations when blunt force impacts/beating occurred), bleeding from the external auditory meatus (and/or from the mouth and nose); basilar skull fracture  – punctate petechial haemorrhages of the facial skin (on eyelids, in conjunctivae of the eyelids, mucosa of the oral vestibule) during compression of the throat (manual strangulation/ligature), chest compression; detailed examination of the neck in every case. Facial swelling and cyanosis. Eyes: a. Opened, closed, dryings of the sclerae; b. Pupil width: of equal or different size, small/middle/large width; c. Haemorrhages of the conjunctivae of the eyeballs. Mouth and nose: a. Froth: pulmonary oedema (congestive heart failure), opiate intoxication, drowning; b. Death due to compression of the neck, vomitus in the oral cavity; c. Foreign material in the oral cavity; d. Suicidal or homicidal intoxication: particles of tablets; e. Trace flow from a corner of the mouth, trace flow of saliva in cases of hanging (salivation due to pressure to pterygopalatine ganglion); f. Blood in the oral cavity and in the oral vestibule: blunt force (lacerations of the lips and the mucosa of the oral vestibule); g. Shot wound in the mouth; h. Teeth: fixed, condition of denture; acid burn of lips, trace flows; i. Teeth-­framed prints on lips and the mucosa of the oral vestibule: manual occlusion of the mouth; j. Upper gastrointestinal bleeding: in case of gastric bleeding, possibly haematinised blood; k. Acid or alkaline intoxication. Tongue: behind, between the rows of teeth, injuries due to bitemarks. Nose: a. Nasal skeleton with abnormal mobility, contents of nostrils, trace flows;

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5.  Injections sites: drug addiction, then check crook of the arm,

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12.  13. 

Necessary instruments for the external postmortem examination 1. Single-­use gloves. 2. Two forceps (to ectropionise the conjunctivae of the eyelids). 3. Torch light if illumination is low.

6.9.3  Anamnesis/conditions of death and time of death 1. Death was expected, defined underlying disease with a poor prognosis was known; time and conditions of death are compatible with diagnosis and prognosis. 2. Sudden and unexpected death; no anamnestic information regarding underlying diseases that could directly lead to death.

6.9.4  Who performed the external postmortem examination 1. Treating physician. 2. Doctor with information given by treating physicians. 3. Neutral doctor when death occurred during surgery (exitus in tabula).

4. Doctor with no information given by treating physicians.

Certification of Death: External Postmortem Examination

1. Functional final states (such as respiratory, circulatory or car-

N

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TO

diac arrest, cerebral death, old age) are not ‘causes of death’ but constituent parts of the dying process. 2. Cachexia and bleeding to death are causes of death, but the underlying diseases have to be named, such as: a. Cachexia caused by anorexia nervosa; b. Bleeding to death caused by an aneurysm that ruptured into the abdominal cavity; c. Bleeding to death caused by stab wounds of the chest with injuries of heart and lungs. 3. Note the obligation to inform the a. Police: if death is unnatural, the manner of death is unclear or the identity of the deceased is not known; b. Health authority: if there is suspicion of communicable diseases and/or transmitted diseases according to the infection protection act.

N

6.10  Completing the death certificate

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death certificate has to be filled out (ask treating physician if additional information is available on the case medical history). 2. If it is impossible to undertake the external postmortem examination at the place where the corpse was found, inform the police. 3. If death is unnatural or manner of death is unclear, inform police. 4. If there is suspicion of carbon monoxide intoxication, inform residents and police in order to determine the source.

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1. When external postmortem examination is completed, the

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Useful website

www.destatis.de

(last

accessed

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Statistisches Bundesamt, 25 March 2020).

References and further reading Basso, C., Corrado, D., and Thiene, G. (1999). Cardiovascular causes of sudden death in young individuals including athletes. Cardiology in Review 7: 127–135. Basso, C., Calabrese, F., Corrado, D., and Thiene, G. (2001). Postmortem diagnosis in sudden cardiac death victims: Macroscopic, microscopic and molecular findings. Cardiovascular Research 50: 290–300. Basso, C., Aguilera, B., Banner, J. et  al. (2017). Guidelines for autopsy investigation of sudden cardiac death: 2017 update from the Association for European Cardiovascular Pathology. Virchows Archiv 471: 691–705.

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Berzlanovic, A., Keil, W., Waldhoer, T. et al. (2005). Do centenarians die healthy? An autopsy study. Journals of Gerontology 60: 862–865. Brinkmann, B. (1997). Fehlleistungen bei der Leichenschau in der Bundesrepublik Deutschland. Ergebnisse einer multizentrischenStudie (I) und (II). Archiv für Kriminologie 199: 2–12, 65–74. Burton, J.L. (2007). The external examination: an often neglected autopsy component. Current Diagnostic Pathology 13: 357–365. Burton, J.L. and Rutty, G.N. (2010). The Hospital Autopsy, 3rd edn. London: Hodder Arnold. Ferris, J.A.J. (2005). Autopsy. Adult. In J. Payne-­James, R.W. Byard, T.C. Corey, and C. Henderson (eds.), Encyclopedia of Forensic and Legal Medicine, Vol. 1, pp. 183–192. Goldman, L., Sayson, R., Robbins, S. et  al. (1983). The value of the autopsy in three medical eras. New England Journal of Medicine 308 (17): 1000–1005. Hanzlick, R. (1993). Death Certificate. The need of further guidance. American Journal of Forensic Medicine and Pathology 14, 249–252. Hanzlick, R. (2006). Cause of Death and the Death Certificate: Important Information for Physicians, Coroners, Medical Examiners and the Public. Northfield IL: College of American Pathologists. Hanzlick, R. (2007). Death Investigation – Systems and Procedures. Boca Raton: CRC Press. Interpol (2002). Disaster Victim Identification Forms (DVI). https://www. interpol.int/How-­we-­work/Forensics/Disaster-­Victim-­Identification-­ DVI (last accessed 25 March 2020). Junbelic, N.I. (2005). Mass disasters. Role of forensic pathologists. In J. Payne-­James, R.W. Byard, T.S. Corey, and C. Henderson (eds.), Encyclopedia of Forensic and Legal Medicine, Vol. 1, pp. 197–207. Amsterdam: Elsevier. Leis, J. (1982). Die Todesursache unter indivudal-­ pathologischen Gesichtspunkten. Deutsche Medizinische Wochenschrift 107,  1069–1072. Lundberg, G. (1998). Low-­ tech autopsies in the era of high-­ tech ­medicine. Continued value for quality assurance and patient safety. JAMA 280 (14):1273–1274. Madea, B. (2015). Rechtsmedizin. Befunderhebung, Rekonstruktion, Begutachtung, 3rd edn. Berlin, Heidelberg, New York: Springer. Madea, B. (2019). Die ärztliche Leichenschau: Rechtsgrundlagen  – Praktische Durchführung – Problemlösungen, 4 edn. Berlin, Heidelberg, New York: Springer. Madea, B. and Rothschild, M. (2010). The postmortem external examination. Deutsches Ärzteblatt International 107 (33): 575–588. Madea, B. and Kernbach-­Wighton, G. (2012). External post mortem examination. In: P. Saukko and J. Siegl (eds.), Elsevier Encyclopedia of Forensic Sciences, 2nd edn. Amsterdam: Elsevier. Madea, B., Doberentz, E., and Radbruch, L. (2019). Leichenschau bei unerwarteten Todesfällen im Krankenhaus. Kriminalistik 10: 570–578. Madea, B., Doberentz, E., and Kristiansen, G. (2020). Rückläufige Obduktionszahlen in Deutschland: Konsequenzen für medizinische Versorgung, Wissenschaft und Rechtspflege. LUHRI Verlag, Bonn. Madea, B. and Doberentz, E. (2020). Die Ärztliche Leichenschau. Teil I. CME Zertifizierte Fortbildung, Springer Medizin, 16 (1–2): 9–19. Madea, B. and Doberentz, E. (2002). Die Ärztliche Leichenschau. Teil II. CME Zertifizierte Fortbildung, Springer Medizin, 16 (3): 9–23.

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6.9.5 Measures/consequences

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Richter, M. (1905). Gerichtsärztliche Diagnostik und Technik. Leipzig: Hirztel. Rutty, G.N. (2010). The external examination. In J.L. Burton and G.N. Rutty (eds.), The Hospital Autopsy, pp. 90–103. London: Hodder Arnold. Shojania, K., Burton, E., McDonald, K. et al. (2002). The autopsy as an outcome and performance measure. Evidence report/technology assessment number 58 (prepared by the University of California at San Francisco–Stanford Evidence-­based Practice Centre under contract no. 290-­97-­0013) AHRQ Publication for health care research and quality, October 2002. Thieke, Ch. and Nizze, H. (1988). Sterbenstypen: Thanatologische Brücke zwischen Grundleiden und Todesursache. Pathologe 9, 240–244. Wagner, S. (2009). Death Scene Investigation: A Field Guide. Boca Raton, FL: CRC Press.

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Madea, B. and Weckbecker, K. (2020). Todesfeststellung und Leichenschau für Hausärzte. Berlin, Heidelberg, New York: Springer. Magrane, B.P., Gilliland, G.F., and King, D.A. (1997). Certification of death by family physicians. American Family Physician 56: 1433–1438. Marshall, H.S. and Milikowski, C. (2017). Comparison of clinical diagnosis and autopsy findings: Six-­year retrospective study. Archives of Pathology & Laboratory Medicine 141 (9): 1262–1266. Maudsley, G. and Williams, E.N. (1996). Inaccuracy in death certification – where are we now? Journal of Public Health Medicine 18: 59–66. Myers, K. and Farquhar, D.R.E. (1998). Improving the accuracy of death certification. Canadian Medical Association Journal 158: 1317–1323. Orth, J. (1908). Was ist Todesursache? Berliner Klinische Wochenschrift 45: 485–490. Pounder, D., Jones, M., and Peschel, H. (2011). How can we reduce the number of coroner autopsies? Lessons from Scotland and the Dundee initiative. Journal of the Royal Society of Medicine 104: 19–24.

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7

Postmortem Changes and Time since Death

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Burkhard Madea, Claus Henßge, Saskia Reibe, and Michael Tsokos

muscles may be preserved up to 20 hours postmortem. Some of the supravital reactions are of great practical importance in forensic medicine, since they can easily be examined at the scene of death and may provide immediate results regarding the time elapsed since death: these are the mechanical and electrical excitability of skeletal muscle and the pharmacological excitability of the iris.

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With irreversible cardiac or respiratory arrest, immediate muscular flaccidity occurs and all muscle tone is lost. Due to the loss of cardiac activity, early signs of death develop as postmortem lividity and rigor mortis. Depending mainly on ambient temperature, cause of death (infectious diseases, sepsis) and medication (antibiotics), the body will decompose sooner or later.

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7.1 Early Postmortem Changes

7.1.1 Supravitality

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R

C

O

N

In most cases, the irreversible circulatory or respiratory arrest is the main criterion for death. However, metabolism of tissues does not cease immediately after death, but continues for a number of hours. The main energy-­producing metabolic processes in the early postmortem period are the creatine-­kinase reaction followed by anaerobic glycolysis. During this period of intermediary life, supravital reactions can be examined and are defined as reactions of tissue on postmortem excitation. The duration of the supravital period is much longer than that of the resuscitation period, as known from physiology and experimental surgery (Figure  7.1.1). The resuscitation period is defined as the time of global ischaemia following the time when the ability to recover expires; the supravital period covers also the period hereafter, characterised by increasing irreversible damage of structure and function. The resuscitation period of skeletal muscles under normothermia lasts for perhaps 2–3 hours, whereas the supravital electrical excitability of skeletal

Handbook of Forensic Medicine, Volume 1, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

Mechanical excitability of the muscle The mechanical excitability of skeletal muscles is examined by rigorously hitting a muscle with the back of a heavy knife or a chisel; for example, the biceps brachii muscle at a perpendicular direction to the longitudinal arm axis. Other muscles can be examined as well, but reference values for estimating the time since death are available only for the biceps brachii muscles. There are three reaction patterns of the muscles depending on the duration of the postmortem interval (Figure 7.1.2): 1. In the first phase, mechanical excitation of the muscle reveals a contraction of the whole muscle (propagated excitation). This first phase of idiomuscular contraction is identical to Zsako’s muscle phenomenon. The first degree of idiomuscular contraction, a propagated excitation, can be observed up to 2 hpm (hours postmortem). 2. In the second phase, a strong and typically reversible idiomuscular pad develops. This phase may be seen as long as 5 hpm (Figure 7.1.3). 3. In the last phase, a weak idiomuscular pad develops which may persist over a longer period of time (up to 24 hours). This

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Transient Circulation ischaemia Spontaneous function

MEDICAL ASPECTS OF DEATH

Ischaemia

Structure

Time

Permanent ischaemia Circulation

Ischaemia

LY

Spontaneous function

Figure 7.1.3  Typical idiomuscular pad of the biceps brachii muscle. Source: Courtesy of Professor G. Dotzauer.

O

N

Reagibility

Structure Morphological changes

Recovery Latency period Survival period time Resuscitation period Latency of recovery Supravital period

Initial investigations on the electrical excitability of skeletal muscles were already carried out at the end of the 18th and the beginning of the 19th century, and electrical excitability was recommended as one method for estimating the time since death. Meanwhile, extensive investigations have been carried out using techniques to objectify muscular contraction. However, for practical purposes, most investigations on postmortem electrical excitability of skeletal muscle are based on verbal descriptions and subjective grading of the muscular response to excitation – the muscular contraction – according to the following criteria: 1. The strength of contraction. 2. The spread of movement to areas distant from the electrodes. In the early postmortem interval, excitation leads to strong contractions of muscles and the excitation will spread to muscles distant from the electrodes, while with an increasing postmortem interval the contraction will become weaker and the muscular response will be confined to the place of excitation. This reaction pattern can be seen in most muscles. In the final phase, only a fascicular twitching or movement of the electrodes will be visible. The most extensive investigations have been carried out for the orbicularis oculi muscles, since movements of these muscles are easily visible. For the orbicularis oculi muscles, puncture electrodes are inserted at a distance of 15–20 mm into the nasal part of the upper eyelid and 5–7 mm deep; for the mouth, puncture electrodes are placed on both sides 10 mm away from the angles of the mouth (Figure 7.1.4). The muscular response is graded into six degrees (I–VI). In the very early postmortem interval (degree VI), the whole ipsilateral muscle will contract; in degree V, only upper and lower eyelids and forehead will show a reaction, and with an increasing postmortem interval the reaction will be confined to the place of excitation (Figure 7.1.5, Table 7.1.1). For stimulation, a small square-­wave generator producing constant current rectangular impulses of 30 mA and 10 ms duration with a repetition rate of 50 ms is used (Figure 7.1.6). Together

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Figure 7.1.1  Diagram of the duration of the supravital period (bottom) compared to the resuscitation period (top). The supravital period after irreversible circulatory arrest exceeds the duration of the resuscitation period after transient ischaemia by far. Source: After Madea (2007).

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Time

Electrical excitability of skeletal muscle

SE

Irreversible Reversible

% 40 Contraction of the whole muscle

N

30

C

O

Strong and reversible

20

Weak and persistent 24 h

0

FO

R

10

2

4

6

8

10

12 hpm

Idiomuscular contraction

Figure 7.1.2  Three phases of idiomuscular contraction after mechanical excitation of the muscle; frequency of a positive reaction (y axis) over the postmortem interval (x axis); hpm, hours postmortem. Source: After Dotzauer (1958).

weak idiomuscular pad can be seen in a time interval up to 12 hpm. If the idiomuscular pad is not visible, it should be palpated for, otherwise skin incisions may be necessary to demonstrate the presence of an idiomuscular pad.

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CHAPTER 7   Postmortem Changes and Time since Death

Table 7.1.1  Upper and lower confidence limits for the six degrees of electrical excitability in hours in different random samples: forensic, clinical pathology, emphysema and haematoma of the eyelid. Degree

Forensic Clinical Haematoma/emphysema pathology pathology of the eyelid

I.

5–22

3–16

5–16

0–16

3.5–13

1.5–9

3–8

1–7

LY 1–7

1–6

1–6

O

N

TR IB

U

TO

R

U

SE

Figure 7.1.4  Position of electrodes for examining electrical excitability of facial muscles.

2–7

N

II. 1/3–2/3 upper eyelid III. Whole upper eyelid IV. Upper and lower eyelids V. Upper and lower eyelids + forehead VI. Upper and lower eyelids + forehead + cheek

29: traumatic emphysema 27.3–52: postmortem emphysema 32: traumatic haematoma

O

Local upper eyelid

8.5 h

I

10.5

5.5 h

II

8.25

4.75 h

III

5.5

2.5 h

IV

4.5

2.5 h

V

3.5

2.5 h

VI

FO

R

C

13.5

Figure 7.1.5  Six degrees of positive reaction after stimulation of the orbicularis oculi muscle (see also Table 7.1.1).

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Table 7.1.2  Postmortem excitability of the iris after injection of different drugs. Postmortem Concentration (%) excitability (h)

Drug

Mydriatics Norepinephrine/epinephrine Tropicamide Atropine/cyclopentolate Miotics Acetylcholine

1% 0.25% 1%/0.5%

14–46 5–30 3–10

5%

14–46

LY

reaction lasts at least for 1 hour. If no change is visible after this time, the reaction is negative (Table 7.1.2). It is sufficient to examine the pharmacological excitability of the iris just with one drug, since examining a double reaction after injection of a mydriaticum at first instance followed by a mioticum does not provide further information on the time elapsed since death. In contrast to younger investigations, who want to use these data for forensic death time estimation, these data were checked in routine casework in numerous cases, furthermore in field studies and proved to be reliable. Further supravital reactions are the reagibility of the sweat glands on the injection of drugs, sperm motility, vitality of leucocytes, DNA incorporation and postmortem blood clotting. However, these supravital reactions have not gained any practical relevance in forensic medicine.

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with the grading mean values, the 95% limits of confidence for the different degrees of reaction are presented in Figure 7.1.5. The 95% limits of confidence for the six degrees of electrical excitability of the orbicularis oculi muscle have been proven to be reliable on randomly selected case material and also in field studies. However, in cases showing haematoma or emphysema of the eyelid, electrical excitability may last much longer than corresponding to the upper 95% limits of confidence for the particular degree (see Table  7.1.1, last column). The same can be seen in cases of hypothermia. Alternatively, in cases of long-­lasting diseases with long terminal episodes, the duration of electrical excitability is shorter than in cases of sudden death due to the glycogen responsible for the reaction being already depleted during the lifetime. Other muscles can be examined as well, and reliable reference data are available, for example, for the orbicularis oris muscle. The muscles of the thenar or hypothenar may react on electrical stimulation up to about 11 hours postmortem.

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O

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Figure 7.1.6  Square-­wave generator for defined stimulation of facial muscles enabling the procurement of the data in Table 7.1.1 and Figure 7.1.7. Producer and supplier: http://j-­peschke. homepage.t-­online.de (last accessed 1 April 2020).

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O

N

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Postmortem lividity

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Pharmacological excitability of the iris

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Compared to skeletal muscles, the smooth iris muscle is irritable to electrical and pharmacological stimulation for a much longer period of time. In some cases, excitability on subconjunctival injection of noradrenaline or acetylcholine may be observed up to 46 hpm (Table 7.1.2). For practical purposes, the pharmacological excitability of the iris following subconjunctival injection of drugs can be recommended, but the drugs should not be injected into the anterior eye chamber. The starting diameter of the pupil has to be recorded prior to injection using a transparent multidiameter scale. About 0.5 ml solution of either noradrenaline, tropicamide, atropine or acetylcholine should be injected. A positive reaction can be seen within 5–30 minutes with the diameter of the pupil becoming greater (atropine, tropicamide, noradrenaline) or smaller (acetylcholine). The duration of the

After irreversible circulatory arrest, postmortem lividity develops at the earliest and therefore initial postmortem change. Following circulatory arrest, the hydrostatic pressure becomes the leading force within the parallelogram of forces consisting of blood pressure, structural barriers, tissue turgor and pressure of underlying surfaces. Hypostasis means the movement of bodily fluids according to gravity. All fluid compartments are involved in hypostasis, not only the intravascular but transcellular fluids as well. Influenced by gravity, blood sinks into the lowest parts within the vascular system of the body, in a supine position towards the back, the buttocks, thighs, calves and back of the neck. Irregular pink patches in the face, especially over the cheeks in the agonal period, are caused by local stasis and are called ‘Kirchhofrosen’ (Table 7.1.3). Postmortem lividity visible on the skin is a result of the sinking of blood into the capillaries of the corium. Postmortem lividity may be visible within 20–30  minutes after death, in the early stages still as pink patches which are regularly and gradually confluent with an increasing postmortem interval. Due to consumption of oxygen, the pink colour of lividity (Figure 7.1.7a) changes to dark pink or bluish. In the areas of deep hypostasis, cutaneous petechial haemorrhages due to capillary ruptures may develop, called vibices (Figure 7.1.7b). Of diagnostic and criminalistic relevance is not only the development but also the colour of lividity, its distribution over the body and the phenomena fixation (disappearance

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CHAPTER 7   Postmortem Changes and Time since Death

Table 7.1.3  Lividity: causes, consequences and phenomena checked on the body. Cause

Consequences

Phenomenon

Decrease of intensity of myocardial contraction Cardiac arrest, hydrostatic pressure

Stasis

‘Kirchhofrosen’, local stasis with patchy discolouration during the agonal period due to centralisation of the circulation Livores with a ‘shifting’ quality and ‘disappearance on pressure’

Vascular permeability Autolysis, putrefaction

Haemoconcentration Diffusion of haemoglobin

Hypostasis

O

N

LY

Decrease of shifting and disappearance on pressure No shifting, no disappearance on pressure

(e)

N

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TO

(c)

(b)

R

U

SE

(a)

(f)

O

(d)

FO

R

C

Figure 7.1.7  Postmortem lividity. (a) Postmortem lividity in a supine position, pink colour due to storage in a refrigerator. (b) Petechial-­like haemorrhages due to capillary ruptures in areas of deep hypostasis (vibices). (c) Zonal segmentation of hypostasis: partly dark blue, partly red. (d) Brownish colour of hypostasis in case of methaemoglobin intoxication. (e) Patterned hypostasis of the back due to the underground. (f) Complete disappearance of lividity on light blunt pressure.

after turning the body) and disappearance (blanching) on blunt thumb pressure. In case of intoxication by carbon monoxide and cyanides, the colour of hypostasis usually appears cherry pinkish, with methaemoglobin intoxication brownish (Table 7.1.4, Figure 7.1.7d). Due to the lack of dissociation of oxyhaemoglobin, a bright pink colour of lividity may be seen in hypothermia as well. If a body is transferred from a cold environment into normal room temperature, a typical zonal segmentation of hypostasis may be seen with the colour being dark blue in the rewarmed areas (Figure 7.1.7c). Of predominant criminalistic significance regarding lividity are the phenomena of ‘disappearance on pressure and disappearance after turning the body’. In the early stage, lividity will com-

pletely disappear on soft thumb pressure. With an increasing postmortem interval, this pressure must increase as well. Later, the lividity will disappear only incompletely on blunt pressure and, finally, it will not disappear at all. If the body is turned in the early postmortem interval, some or all of the hypostasis may move down to the most dependent areas according to gravity. In a comparatively later postmortem interval, only some of the hypostasis will slip down to the new dependent area and only a faint blanching of the lividity will be noted in the former dependent area (Figure 7.1.8). With increasing postmortem interval, disappearance on thumb pressure and relocation after shifting decreases and then ceases completely. This effect is caused by an increasing

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Table 7.1.4  Postmortem lividity discolouration. Aetiology

Colour

Mechanism

Normal Carbon monoxide Cyanide Fluoroacetate Refrigeration/hypothermia

Blue-­purplish Pink, cherry-­red Pink, cherry-­red Pink, cherry-­red Pink, cherry-­red

Sodium chloride/nitrite, nitrate Hydrogen sulphide

Brown Green

Venous blood Carboxyhaemoglobin Excessively oxygenated blood due to inhibition of cytochrome oxidase Same as mentioned in the earlier text Oxygen retention in cutaneous blood by cold air Left shifting of the Hb O2 dissociation curve Methaemoglobin Sulfhaemoglobin

O

N

haemoconcentration of intravascular erythrocytes due to transcapillary extravasation of plasma. The intravascular haemoconcentration is the main reason for the disappearance gradually decreasing on thumb pressure and after shifting. In a comparatively later postmortem interval, haemolysis and haemoglobin diffusion into the perivasal tissue start, but this only becomes a secondary mechanism contributing to the fixation of hypostasis. Elements of hypostasis that change with an increasing postmortem time interval are: 1. Confluence, maximum, disappearance on thumb pressure. 2. Complete or incomplete disappearance after shifting. Longitudinal studies on these criteria in large random samples are lacking and cross-­sectional studies are of limited value only. The best statistical data available were summarised by Mallach (1964), who calculated mean values, standard deviations and 95% limits of confidence based on a number of textbook reports (Table  7.1.5). Since better data are lacking, these data are still undisputed. However, these data do not represent absolute threshold values. Investigations using a quantitative measurement of livor mortis have not yet gained practical importance.

SE

Up to 6 hpm complete shifting

LY

Source: Modified according to Spitz et al. (2006).

TO

R

U

6–12 hpm incomplete shifting

TR IB

U

Over 12 hpm no shifting

N

Figure 7.1.8  Shifting of lividity after turning the body. Source: After Patscheider and Hartmann (1993).

O

Table 7.1.5  Time courses of different criteria of lividity according to calculations by Mallach (1964) based on textbook reports. Range of scatter (h)

Standard deviation (h)

Lower limit

Upper limit

Lower limit

Number of quotations Upper limit used for calculations

0.75 2.5 9.5

0.5 1 4.5

– 0.75 0.5

2 4.25 18.25

0.25 1 3

3 4 16

17 5 7

5.5

6

–

17.5

1

20

5

17

10.5

–

37.5

10

36

4

3.75 11 18.5

1 4.5 8

2 2.25 2.5

5.5 20 34.5

2 4 10

6 24 30

11 11 7

C

Second standard deviation (h)

R

Mean value x (h)

FO

Criteria of lividity

Development Confluence Greatest distension and intensity Displacement: 1. Complete on thumb pressure 2. Incomplete on sharp pressure (forceps) Displacement after turning the body: 1. Complete 2. Incomplete 3. Only little pallor

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CHAPTER 7   Postmortem Changes and Time since Death

(for instance, over the neck in a supine position) are observed.

2. Confluence is stated if separate areas of discolouration of

Each part, deprived of supple government, shall, stiff and stark and cold, appear like death. With irreversible circulatory arrest, all muscles of the body will become completely flaccid due to a loss of tone. In the early postmortem interval, adenosine triphosphate (ATP) can be resynthesised due to the creatin-­kinase reaction and anaerobic glycolysis. Once the ATP level has fallen below 85% of the initial value, actin and myosin filaments will contract and subsequently the subjective impression of stiffening of the muscle will set on (Table 7.1.6). In practice, development and state of rigor mortis are proven subjectively by flexing a joint: either the muscles are flaccid or during development of rigor mortis a certain degree of resistance may be felt (Figure 7.1.9). Once rigor has fully developed, even a strong investigator cannot flex or stretch a joint. Biochemical, physiological and mechanical properties of the rigor (prerigor and postrigor) phases are summarised in Table 7.1.7.

7.1.3  Rigor mortis

U

The second postmortem change and sign of death is rigor mortis, developing in normal ambient temperatures about 3–4 hpm

SE

O

moderate intensity are noted. 3. Maximum can be stated if during the death scene investigation and autopsy hypostasis did not increase. 4. The criterion thumb pressure is positive if lividity disappears completely on soft thumb pressure. 5. Complete relocation is given when all hypostasis shifts down to the new dependent areas. This may be observed at the scene when a body is turned from a face-­down into a supine position. 6. In incomplete shifting, hypostasis remains also in the former dependent areas, but shifts as well to a greater or lesser extent to the new dependent areas.

LY

1. Beginning is stated if mottled patches at lower parts of the body

following primary flaccidity. Rigor mortis was misjudged as a sign of death up to the 19th century, although Shakespeare (Romeo and Juliet, Act. IV, Scene 1) described already all the elements of rigor mortis very well as follows:

N

7.1.2  Stages of postmortem lividity

TO

R

Table 7.1.6  Overview of the biochemical, mechanical, morphological and physiological bases of rigor mortis. Establishment of rigor

Biochemistry

ATP level: 0.435 ± 0.555 mg/g muscle

Mechanics

–

Morphology

–

Physiology

Exponential decrease of membrane potential: above −55 mV propagated excitation possible; below this level up to −30 mV only local concentration on excitation

– NH3↑ ATP level decreased to less than 85% of the original value – Spontaneous elongation of Stiffness ↑, plasticity firstly the loaded muscle; increased, then reduced plasticity ↑ Contraction of loaded muscle Swelling and destruction Irreversible elongation of Appearance of fine transverse muscle; decoupling of of mitochondria and striations (bridging between myofilaments, the sarcoplasmic A and I filaments) with a disintegration of reticulum periodicity of 400 Å structure – – –

FO

R

C

O

N

TR IB

U

Rigor phase

Post rigor phase, secondary flaccidity

Delay period

(a)

(b)

Figure 7.1.9  (a) Due to rigor mortis, the lower leg is fixed against gravity. (b) Objects in the hand such as branches must not be mixed up with instantaneous rigor mortis or cadaveric spasm.

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Table 7.1.7  Time course of different criteria of rigor mortis.

Rigor state

Average postmortem ± standard deviation (h)

Beginning Maximum Duration Complete resolution

 3 ± 2  8 ± 1 57 ± 14 76 ± 32

Range of scatter (second standard deviation) (h) Lower limit

Upper limit

Number of quotations used for calculation

–  6 29 12

 7  10  85 140

26 28 27 27

N O SE U R

Figure 7.1.10  Re-­establishment of rigor mortis after breaking. If rigor is broken during development of rigor mortis (at D or E), it will re-­establish at a lower level (at D, or E). If rigor is broken (e.g., at F) after it has fully developed (B), it will not re-­establish at all. Source: After Madea (2007).

FO

R

C

O

N

TR IB

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Rigor must not be confused with cold stiffening. When rigor is present, hypostasis must be present as well, but in cases of cold stiffening (body core temperature 30–33 °C), hypostasis is usually absent. Development and state of rigor mortis should be examined not only in one but in several joints (mandibular joints, fingers, knees, elbows) to get an impression if it is still in progression or has already reached its maximum. Rigor mortis does not start simultaneously in all muscles. Nysten’s rule (1881) that rigor starts in the mandibular joints and then affects muscles of the trunk, next the lower and finally the upper extremities applies to most cases of death in a supine position. However, in cases with glycogen depletion during protracted agony in the lower extremities, rigor will start there. Resolution of rigor mortis is due to protein degradation (increase of NH3). Rigor mortis does not start simultaneously in different fibres of a muscle but successively. This phenomenon can be used for a rough estimation of the time since death as well. If some fibres have already become stiff, this stiffness can be broken by flexing a joint; rigor may now develop in other, not yet stiffened fibres. Dependent on the time when stiffness has been broken, rigor may develop again on a higher or lower level unless it was already fully developed (Figure 7.1.10). This phenomenon of re-­establishment of rigor mortis may be observed up to 8 hpm, and in rare cases up to 12 hpm. Younger investigations on 314 joints of 79 bodies revealed a re-­ establishment of rigor mortis in 38.5% of joints in the time interval from 7.5 until 19 hpm. These ‘observations’ are in contrast to experimental findings (objective measurement of rigor mortis, duration of supravital period of skeletal muscle, more than 20,000 objective measurements of muscular contraction) and are lacking physiological explanation and are obviously not valid. Secondary flaccidity may become apparent in normal ambient temperature after 2 days. In lower ambient temperatures, a fully developed rigor mortis may be preserved for 2 weeks and even longer. Cadaveric rigidity, cataleptic spasm or instantaneous rigor stand for phenomena always being mentioned in textbooks, but which are non-­existent in practice. None of the cases reported in the literature stands up to criticism. Rigor mortis is not only established in skeletal muscles but in smooth muscles; for example, of the skin. Rigidity of the smooth musculi arrectores pilorum can be seen as gooseflesh (cutis anserina; Figure 7.1.11).

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Source: According to calculations by Mallach (1964) based on textbook reports.

Figure 7.1.11  Gooseflesh of the right forearm due to rigidity of the smooth musculi arrectores pilorum.

Development, duration and resolution of rigor mortis are dependent on the amount of glycogen in the muscles at the moment of death, ambient temperatures and so on. Therefore, rigor may develop very fast in people who die during or soon after physical exertion or exhaustion or from electrocution. All of the criteria mentioned in the earlier text concerning rigor mortis

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CHAPTER 7   Postmortem Changes and Time since Death

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too. The cooling of a body is decelerated by postmortem heat production only within the first hours of postmortem and only to a minor degree; interindividual differences due to varying glycogen levels have to be taken into account. The liver must be seen as an exception, since liver temperatures are usually higher than other central core temperatures. Lundquist calculated roughly: heat release is approximately 0.4 kcal/g glycogen. Glycogen of the body 350 g is thus approximately 140/587 kcal/kJ heat. Thus, this amount of heat currently released would lead to a current increase in temperature of approximately 2.5  °C based on a mean specific heat of 0.8  kcal/kg. Roughly estimated, this amount of heat could explain about one in six of the postmortem temperature plateau. 2. In cases of non-­stationary heat conduction, the cooling rate is proportional to the temperature guide number of the body tissue. The temperature guide number is the quotient of heat conductivity and specific heat capacity. The temperature guide numbers of biological tissues are three decimal powers lower compared to good heat conductors. Although the heat conductivity of fatty tissue is significantly lower than that of muscle tissue, it also has a low heat capacity. Temperature guide numbers of fat and muscle tissues have a ratio of 1:1.3. Therefore, the assumption that bodies with much adipose tissue probably cool much slower down than those with smaller fat depots is not true. 3. Convective heat transport within the body stops with irreversible circulatory arrest. Heat exchange is then mainly based on conduction. The conductive heat transport within the body is mainly due to temperature differences of neighbouring tissue layers. Since the heat conductivity of body tissues is rather low, the conductive heat transport within the body proceeds slowly. From these physical facts, the following conclusions can be derived: 4. In case of temperature differences between core and surface of the body, or body surface and surrounding temperature, heat transport basically proceeds radially, from the body core to the surface (cooling) or the other way round (warm-­up). 5. The cooling (or warm-­up) process begins with a significant increase of temperature differences within the body in a radial direction (core-­surface). This increase is caused by temperature differences between surrounding and surface temperatures. These differences cause either heat dissipation from the warmer surface to the cooler surrounding or vice versa, and heat absorption by the cooler surfaces from the warmer surroundings. This heat dissipation or absorption results in a reduction or an increase of the temperatures of the most superficial tissue layers. Thus, a temperature difference to the adjacent and deeper tissue layers develops and causes heat transport in the direction of the temperature gradient. This process proceeds slowly until the deep tissue layers of the axial centre are affected by temperature reduction or increase. A visible sign of this process is the postmortem temperature plateau of cooling curves that are located far away from surfaces. Since the local temperature within the body or parts of a body

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(development, re-­establishment, full development, duration and resolution) are time-­dependent. This is evident from one of the rare studies on rigor mortis of the 19th century. Niderkorn (1872), who determined the times necessary for the completion of rigor mortis in 113 bodies, found it fully established after 4–7 hours in 76 corpses (67%). In two cases, rigidity was complete within 2 hpm and in two others within 13 hpm. However, there is a wide range of the interindividual variability which is due to endogenous and exogenous factors. Longitudinal studies on large random samples are lacking; however, results of animal experiments taking into account various factors influencing the time course of rigor mortis have been published. Devices for an objective measurement of rigor mortis have also been developed, but have not yet gained practical importance. The best available data despite all the justified criticism originate again from Mallach (1964), based on his literature compilation (1811–1969) with statistical analyses (Table 7.1.7). These values again cannot claim to be absolute limit values. As lividity, rigor mortis can only provide a rough estimate and no greater accuracy can be expected. It should never be examined solely. Rigor is examined by flexing or stretching joints. Beginning can be stated if a slight rigor can be observed in some joints. Complete development or maximum means that it has strongly developed in all joints. Re-­establishment can be stated if rigor is found again in a joint (mostly elbow) some time (hours) after breaking it. Mostly rigor is broken during death scene investigation and examined again at autopsy.

Temperatures of corpses

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Body temperatures are of practical medicolegal interest for the following tasks: 1. Determination of the time since death. 2. Diagnosis of hyperthermia/hypothermia as cause of/contributing to death. Both these tasks depend on each other. Hyperthermia or hypothermia derived from body temperature can only be diagnosed in very rare cases. Preconditions are a known time period since death or reliable information concerning ambient temperature after death. Alternatively, estimations of the time since death derived from body temperatures have to take into account the possibility of hyperthermia/hypothermia at the time of death.

Basic principles for body temperature after death 1. Heat production after death is low and is mainly due to anaer-

obic glycolysis. Anaerobic glycolysis stops within a few hours after death with the degradation of the body’s glycogen/glucose to lactic acid. Subsequently, the heat production is low

MEDICAL ASPECTS OF DEATH



Tr Ta , To Ta

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Q

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ing. Already early authors in the 19th century were using rectal temperatures and described a lag time; the postmortem temperature plateau before an exponential body cooling according to Newton’s law commences (Figure 7.1.12). The temperature plateau is because central axial temperatures cannot begin to decrease until a heat gradient is set up between the core of the body and its surface. This delay is variable and may last for some hours (point 6). A mathematical expression of the rectal cooling after death was published by Marshall and Hoare (1962), taking into account the whole sigmoidal shape of the cooling curve. Marshall and Hoare performed body cooling experiments under ‘standard conditions of cooling’ which are defined as a ‘naked body with dry surfaces, lying extended on a thermically indifferent base, in still air’. Their mathematical model of body cooling is expressed in a two-­exponential term:

A x exp B x t

1 A x exp

AxB A 1

x t,



where Q = standardised temperature; Tr = rectal temperature at any time t; To = rectal temperature at death (t = 0); Ta = ambient temperature; A = constant; B = constant; t = time of death.

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varies widely at the moment of death, an exact local definition of the site of measurement is necessary. The temperature which is taken at a certain postmortem time at one site under defined cooling conditions depends on the radius of the part of the body and the radial depth of the measuring site. 6. The postmortem temperature plateau is mainly determined physically (point 5) and is only slightly intensified by postmortem heat production (point 1). The duration of the temperature plateau depends on the same influencing factors as the body cooling in the pure exponential cooling phase following the plateau. These influencing factors are: radius of the body part, radial position of the measuring site and cooling condition. The shorter the temperature plateau is, the steeper the curve drops away. Central cooling curves of cooling dummies which are used to simulate rare cooling conditions and warmed up to 37 °C also show a postmortem temperature plateau. The postmortem temperature plateau causes the sigmoidal shape of the cooling curve of any central axial site of measurement. This sigmoidal shape is a combination of the postmortem temperature plateau and the following single exponential cooling curve according to Newton’s law of cooling. 7. The mechanisms of heat dissipation of the living must not be transferred to those of bodies. This is due to points 1 and 3 as well as the missing insensible evaporation of the body. 8. Postmortem body cooling is due to the following factors: conduction, convection, radiation and water evaporation. Heat dissipation by water evaporation has the lowest significance. Conduction may play a major role in those parts of the body lying on a surface; convection at those parts of the body which are not lying on a surface. Only a very small part of body cooling is caused by radiation, since the temperature of the body surface decreases rapidly towards the environmental temperature. For practical purposes, it is very important to examine the body at the scene, and especially to evaluate if there are any thermal interactions between temperature measuring site and surroundings regarding conduction, convection and radiation. Sources of radiation in the environment of a body – sun, radiator, oven, lamps with intensive heat radiation, walls with significant lower or higher temperature than the environmental media (air, water)  – as well as forced convection (wind, water drift) or special supporting bases (metal plates, concrete in groundwater level) have an important impact on the cooling velocity, especially of unclothed bodies. The rate of cooling depends on various conditions and can be altered by several factors: a. ambient conditions (temperature, wind, rain, humidity); b. weight of the body, mass/surface area ratio; c. posture of the body (extended or thighs flexed on the abdomen); d. clothing/covering. Different temperature probe sites were used (surface skin temperature, axilla, liver, rectum, brain temperature), but for practical purposes only central core temperatures (rectum, brain) are of value today. Body cooling does not follow Newton’s law of cool-

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T0

Temperature (°C)

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Postmortal temperature plateau

T0–Ta Tr–Ta

Ta Time (hpm) Figure 7.1.12  Sigmoidal shape of the cooling curve which is best described by the two-­exponential term of Marshall and Hoare (1962). The quotient

Tr Ta To Ta

is a good measure of the progress of

cooling. If this quotient Q is 100 mg/dl and, in a second step, cases with a terminal episode >6 hours, the 95% limits of confidence could be reduced to ·22 and ·20 hours. Recently, several statistical approaches for a more accurate estimation of the time since death have been recommended:

< <


<


>

> >

1. In most investigations, the postmortem interval has been used

> > > > > > > > > >

>

MEDICAL ASPECTS OF DEATH

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31 30 29 28 27 26 25 24 23 22 21 20 19 18 17 16 15 14 13 12 11 10 9 8 7 6 5 4 3 2 1

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Case numbers

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Gastric contents and time since death

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Table 7.3.2  Precision of time-­of-­death estimation using vitreous potassium in different random samples. Urea was used as internal standard; statistical parameters of the entire sample and subgroups. Urea as internal standard

n Intercept Slope Correlation coefficient Variance (S²) Standard deviation (Syx) 95% limits of confidence (h)

Entire sample

Urea < 100 mg/dl

Change (%)

270 (170) 6.10 (5.99) 0.20 (0.2033) 0.89 (0.86)

288 (138) 6.02 (5.88) 0.18 (0.1877) 0.91 (0.89)

-15.5 -1.3 -10 -2.2

8.57 2.93 (3.42)

5.09 2.25 (2.62)

-40.6 -23.2

±25.51 (±34)

±21.78 (±22)

-14.6

Source: The data in brackets are from Madea et al. (1989).

Gastric content alone allows only a rough estimation of the interval between last meal and death. State of digestion and the distribution of the last meal in stomach and upper intestine have for a long time been proposed as a method to estimate the time since death. For the estimation of the time since death, the volume of the stomach content compared to the volume of the last meal and transportation distance into the small intestine must be known. Even if the volume of the last meal is not known, the type of the meal (breakfast, lunch, dinner) may allow rough estimations of the daytime when death occurred. Gastric emptying has been studied and quantified in the last decades using different methods (e.g., radiological, intubation-­aspiration, radioisotopes, ultrasound, absorption kinetics of orally administered solutes, ferromagnetic traces). Liquids leave the stomach much faster than solids. While gastric emptying for liquids follows an obviously exponential function, solids show a linear emptying pattern.

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the following conclusions can be derived: if at autopsy 50% of the volume of the last meal is found, the last food intake was about 3–4 hours prior to death with 98% confidence limits not shorter than 1 hour and not longer than 10 hours. If 90% of the last meal is found in the stomach, the last ingestion took place probably within the last hour prior to death with 98% limits of confidence of not more than 3–4 hours. However, conditions for a delayed and accelerated gastric emptying have to be kept in mind (Box 7.3.1). A different reference curve exists for mere carbohydrate nourishment, since gastric emptying obviously seems to proceed much faster in such cases. A reference curve for baby food was also compiled. It can be used in the same way as the curve for mixed food. Special instructions for microscopic and immunologic identification of stomach content are available. The macroscopic identification of stomach content should be carried out according to the suggestions of Holczabek (1961): in order to get a first impression of the type of food, we use plastic sieves (plastic is used to avoid any kind of contamination). Primarily, the stomach content is put into a plastic sieve with holes of 2 mm in diameter placed above a bowl with ice. The stomach content is then purified by using water. In all cases without advanced digestion, the composition of food can immediately be identified to a great extent. The volume might be estimated by putting it on a dish. By this, it might also be estimated how much of the stomach content has already left the stomach. If the type of food cannot be identified, a small particle can be used for microscopic examination, identified and stored in 10% formalin, and the rest of

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The following gastric emptying times are given in the literature: 1–3 hours for a light small-­volume meal; 3–5 hours for a medium-­ sized meal; 5–8 hours for a large meal. However, different anatomical and functional disorders may cause delayed or accelerated gastric emptying (Box 7.3.1). According to Horowitz and Pounder (1985), only the solid compartments of a mixed solid and liquid meal should be considered and the weight of the stomach content should be compared with an estimated weight of the last meal and reference made to the known 50% emptying times for the solid components of meals of various sizes. Tröger et  al. (1987) compared the gastric content (volume) found at autopsy to time and volume of last meals on an autopsy collective of 47 cases (sudden and unexpected death, exclusion of brain tumours, operations of gastrointestinal tract, intoxication, blood alcohol level). Gastric contents in per cent of the volume of the last meal were plotted against the survival time. Only a gastric volume over 10 g was considered. Regression line and 90 and 98% confidence limits were calculated (Figure 7.3.7). From this graph,

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• Transient delayed gastric emptying • Postoperative illness • Acute viral gastroenteritis • Hyperglycaemia • Drugs: morphine, anticholinergics, levodopa, β-­adrenergic agonists, nicotine • Stress: labyrinthine stimulation, cold, pain, pectin supplementation • Chronic gastric stasis • Diabetes mellitus • Postsurgical (truncal vagotomy with pyloroplasty and antrectomy) • Gastroesophageal reflux • Anorexia nervosa • Progressive systemic sclerosis • Chronic idiopathic intestinal pseudo-­obstruction • Amyloidosis • Myotonia dystrophica • Familial dysautonomia • Dermatomyositis • Tachygastria • Paraplegia • Idiopathic myocardial infarction • Acute abdomen • Laparotomia • Physiological (liquids, acid, lipids, left-­side position) • Accelerated gastric emptying • After gastric surgery: • vagotomy • antrectomy/subtotal gastrectomy • Zollinger–Ellison syndrome • Duodenal ulcer disease • Reserpine • Physiological: liquids, hunger

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Box 7.3.1  Aetiology of delayed and accelerated gastric emptying.

Source: After Horowitz and Pounder (1985) and Tröger et al. (1987).

Figure 7.3.7  Relation between gastric volume and a mixed meal in per cent of ingested volume and time after ingestion with regression line, 90% and 98% limits of confidence. Source: From Tröger et al. (1987).

MEDICAL ASPECTS OF DEATH

relatively constant over a longer period of time and during day and night, while air temperatures differ not only from day to night but from one day to the next. Morphological findings which have to be taken into consideration for an estimation of the duration of immersion in immersed bodies are listed in the following text.

External findings

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a. washerwomen’s skin; b. loosening of nails; c. peeling of skin; d. loss of nails. 9. Feet: a. washerwomen’s skin; b. loosening of nails; c. peeling of skin; d. loss of nails.

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1.  Rigor mortis. 2.  Lividity. 3.  Marbling. 4.  Bloating of face, scrotum and subcutaneous tissue. 5.  Discolouration of skin (green, black, reddish). 6.  Loss of epidermis. 7.  Loss of hairs. 8.  Hands:

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the remaining stomach content is kept – necessarily with the rinsing liquid – for chemical examinations. The rinsing liquid in the ice bowl is then filtered through a fine mesh with special attention given to the leading elements. Leading elements are those particles that are only slowly digested and thus can indicate the path of the food through the intestinal tract. These leading elements are mainly fruits (e.g., poppy seeds, caraway, fruit stones, cloves and peppercorns). Microscopic examination of the intestine content often shows an affiliation of the leading elements with special types of food. The leading elements can also provide quite detailed information about the progression of digestion before death, since they can be found in large sections of the intestine. For examination of the small intestine, it is placed between two marks on the table (1.5 metres distance between the marks) so that its approximate length can be determined. The intestine is being ligated at three points (in all cases in which an examination might be of high importance at five points) before removing it from the body. Thus, shifting of the content can be mostly avoided. In the next step, the intestine is opened and the content put in a sieve with 1 mm holes. Prior to the examination with a sieve, the content of the intestine is checked regarding amount, colour, consistency and odour. Notice possible filling of the lymphatic vessels of the intestinal wall and within the mesentery. During the examination, it is often possible to identify food particles with the naked eye and test specimens retained for microscopic examination. The large intestine also has to be examined in three portions. The content of the large intestine is also put into a sieve with 2  mm holes, and it is washed with water using a shower head. With this method, it is quite easy to separate solid parts from the content of the intestine.

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Internal findings

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1. Volume of transudate in pleural cavity. 2. Heart without blood. 3. Liquefaction of brain.

Putrefaction

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Putrefaction is a bacterial process predominantly influenced by environmental factors, mainly the ambient temperature, and by underlying diseases and body proportions as well. Advanced stages of putrefaction may be seen within a few hours after death; those in moderate or cold climates not seen after weeks. Even in relatively constant ambient temperatures, the progression of putrefaction varies considerably. Recent H-­magnetic resonance spectroscopic investigations on metabolites emerging during decomposition of brain tissue (Scheurer et al. 2003) offer promising results that even decomposition may be used as a reliable method to estimate the time since death. However, these longitudinal studies on postmortem changes are still on an experimental level and decomposition under different ambient temperatures has to be studied. One weak point may be that decomposition has been studied in isolated brains and skulls; thus, an invasion of bacteria from the gastrointestinal tract is not possible. There is no sound approach to the use of stages of putrefaction for estimating the time since death while the body is lying in air. For bodies recovered from water, a quite good and reliable method for estimating the minimum and maximum water time has been developed based on putrefactive changes which are visible at external examination or at the dissection of the body. This is mainly due to the fact that water temperatures are

The warmer the water, the sooner a definite stage of putrefaction is achieved. From the mean water temperature for each month and the stages of decomposition, the German forensic pathologist Reh (1969) developed a chart with minimum time intervals of immersion. For all 16 parameters for estimating the minimum time of immersion, see Table 7.3.3. On the left-­hand side of the table, a list of useful criteria can be found; in the first line the months, in the second line the mean water temperature and in the following line the minimum time interval in days are shown. As many criteria as possible should be used for estimating the minimum time interval since death. With more than only one or two criteria, the result will become more reliable. For estimating the minimum time interval, the mean water temperature which is nearest to the actual water temperature at the time of recovery should be used. With this chart, not only the minimum time interval of immersion can be estimated but also the maximum interval by considering those criteria which have not yet developed. If in June marbling, bloating and discolouration of the body have developed, and the nails are loose but not lost, it may be concluded that the interval of immersion is over 3 days but below 8 days. The author’s personal experience with this chart is quite good, since it is much better than the old rules of thumb because it takes the actual temperature for the progression of putrefaction into consideration.

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Table 7.3.3  The German forensic pathologist Reh (1969) developed this chart showing minimum time intervals* of immersion (days). Chart to estimate the minimum time interval of immersion. First line: month; second line: median water temperature for the month; left column: signs of putrefaction and malzeration; following columns: minimal time interval in days. If e.g., in July marbling, distension of tissues by gas, discolouration of the body, peeling of the epidermis, loosening of finger and toe nails, peeling of the skin of hands and feet and a liquefied brain are observed, the minimum time interval of immersion would be about 2–3 days. Jan.

Feb.

Mar.

April

May

June

July

Aug.

Sept.

Oct.

Nov.

Dec.

3,5°

3,9°

5,8°

9,9°

13,0°

17,4°

18,6°

18,6°

17,3°

13,2°

8,8°

4,7°

32 35

25 25

16 (23) 16 (23)

9–10 10

4–5 4–5

2 2–3

1–2 2

2 3

3 3–4

4–5 7

10 10

17 17

35

25

16 (23)

(14)

4–5

2

2

3

3–4

7

10

17

35 35 (1)

25 25 (1) 28–30

16 (23) 16 (23) (12 h)

4–5 4–5

3 2–3 (6 h)

2 2–3

3 3

3–4 3–4 2h

7 7

10 10 2h

17 17 (1)

Over 35 35

(40) 30–32 (45)

23 23

16 16

5 10

2–3 3

3 3

3 3–4

3–4 4

11 7

17 20

28 28

Over 53 (1)

45 (1)

30 (40) (12 h)

21 (1)

14

8 (6 h)

3 1/2 h

4

10 2h

Over 11

20 2h

Over 35 (1)

11. 12.

Median water temperature (°C) Marbling Distension of tissues by gas Discolouration of the body Peeling of the epidermis Hair lost Hands: beginning of wrinkling Nails become loose Peeling of skin in glove form Nails lost Feet: beginning of wrinkling Nails become loose Peeling of skin

Over 53 Over 53

40 60

26 (35) 35

17 16

10 10

5 5

3 3

4 5–6

8 8–9

17 20

28 28

13.

Nails lost

Over 53

Over 60

53

35

25 (40)

18 (35)

Over 10 3–4

3

Over 10 3

Over 10 11

5

Over 39 35

32–34 (40) 30 (40)

23 (23)

14–15 14–16

9 5

4 3–4

3 3

3 3

5 6

11 10

Over 20 Over 20 20 17

Over 35

Transudate in pleural cavities* Heart without blood Brain liquefied

Over 28 5

3

14.

Over 35 10

12 Over 11 (14) Over 11

9. 10.

15. 16.

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(16) 10–12

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 > 500 ml in adults. Note: All times are minimum time of duration of immersion except values in brackets which are maximum times.

*

28 28

MEDICAL ASPECTS OF DEATH

However, compared to the average monthly water temperatures listed in the table, the actual water temperatures have since risen during the last 40 years. Especially in summer, reliable results can only be expected when the actual water temperature is similar to the temperatures in the table. Especially for higher water temperatures, the time interval of immersion may be underestimated when using the table, since systematic observations on the progression of putrefaction in correlation to the higher water temperatures are missing. Therefore, as task for the future, the table should be adapted to the risen water temperatures.

Immunohistochemical methods as an aid in estimating the time since death

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A few years ago, Wehner et al. (1999–2002) developed a method to estimate the time since death using immunohistochemical staining.

Proteins undergo degradation after death and this reduces their stainability. Previous studies have explored the development of a method to estimate the time since death using immunohistochemical staining. A systematic analysis demonstrated that calcitonin can still be stained 4 days, thyroglobulin 5 days, glucagon 6 days and insulin 12 days after death. After 12 days, calcitonin and thyroglobulin can no longer be stained, glucagon after 14 days and insulin after 29 days. A control study with 105 cases with known time since death generally confirmed the original data; however, for calcitonin and thyroglobulin, we observed earlier negative stainings. A positive staining is shown in Figure 7.3.8. When using the method in casework, the wider limits according to the results of both studies have to be taken into account to avoid false estimation of the time since death. Although the immunohistochemical detection of antigens allows only a rough estimation of the time since death, it may be helpful in individual cases.

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(b)

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(a)

(c)

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Figure 7.3.8  (a) Pancreas tissue anti-­human insulin 200×, autopsy 8 days after death. (b) Pancreas tissue anti-­human glucagon 200×, autopsy 8 days after death. (c) Thyroid gland tissue anti-­human thyroglobulin 400×, autopsy 8 days after death. (d) Thyroid gland tissue anti-­human calcitonin 400×, autopsy 11 days after death. Source: From Ortmann et al. (2017).

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Case example

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A woman was found dead murdered 8 days after have been seen last alive (Figure 7.3.9). Cause of death was a ligature strangulation; furthermore, there was a postmortem incised wound of the anterior neck. Due to a lack of putrefactive change, postmortem interval was thought to be not more than 2 days. The question of the police was of course: Is an earlier time of death possible? There would be completely different criminalistic approaches: either the murderer killed the woman immediately and hit her in the field, or the murderer had the woman in his power for some days, killed her and then brought the body back to the place of discovery. In this case, even methods with a low precision may be helpful in forensic casework. Immunohistochemical stainings were carried out

on thyroid gland and pancreas and these stainings were positive for insulin, glucagon and thyroglobulin, and negative for calcitonin. Based on the immunohistochemical staining, time since death was not more than 5  days and less than 12  days. Two days after the autopsy, vitreous humour was withdrawn. The potassium value was 33.8 mmol/l. Using different formulas, time since death was over 6 to 7  days. Using the formula of Zilg, which takes into account also the ambient temperature, a postmortem interval between time of missing and time of discovery of 8 days is possible. A shorter time since death (for instance, 2 days) can even be ruled out. Therefore, in comparable cases, even methods with a low precision of death-­ time estimation should be applied and the relevant material taken at autopsy.

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Figure 7.3.9  (a) Place of discovery. (b) Ligature strangulation mark and postmortem incised wound. Source: From Madea et al. (2019).

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7.4.1 Succession

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Any decaying organic material, including carcass, is a natural habitat for a diverse range of arthropods. They use it for nourishment, breeding site, mating or hiding place. As the material decomposes, it undergoes a series of changes offering different species exactly what they are specialised on. Several succession studies were carried out in different countries (Anderson and VanLaerhoven 1996; Arnaldos et  al. 2001; Archer 2003; Bharti and Singh 2003; Grassberger and Frank 2004; Watson and Carlton 2005; Eberhardt and Elliot 2008) to understand the order in which necrophagous species respond to different stages of decomposition and to correlate further species and decomposition stage to estimate a postmortem interval in real cases (Goff 1993). For the studies, animal carcasses were exposed in the open to monitor the different stages of decay and to sample corresponding arthropods. Nevertheless, each trial has to be regarded as an individual experiment, since the habitat and the climate have serious effects on the insect fauna. Figure  7.4.1 shows exemplarily different stages of decay of human body (1–5). All of the corpses were autopsied in the

Institute of Forensic Medicine, Bonn, Germany: each cadaver has been found outdoors under different climatic scenarios in Central Germany; however, the general progress of decay can be followed. In Figure 7.4.1, specimens are grouped above the stage of decomposition in which they were observed or collected. This picture is not complete, it is rather a snapshot view and also the following list of species and their descriptions are an introduction only. Among the earliest visitors of a cadaver are blowflies (Diptera: Calliphoridae) (A in Figure  7.4.1). They deposit their eggs in areas where the hatching first instar larvae can feed on soft tissue (Norris 1965). Another early visitor can be a wasp (Alysia manducator, Hymenoptera: Braconidae) (B in Figure 7.4.1). It is attracted to the odour of decaying meat and acts as parasite to blowfly larvae (Matthews 1974). The latter feeds on the decomposing tissue which is softened and cracked down by the activity of bacteria. A predator of the feeding larvae is the beetle Creophilus maxillosus (Coleoptera: Staphylinidae) (D in Figure  7.4.1) (Greene 1996). After cycling through several larval stages, the next developmental step for blowfly larvae is pupariation; in the pupal stage (E in Figure  7.4.1), the metamorphosis from larvae to adult blowfly takes place (Denlinger 1994). A common parasitoid of the puparium is the wasp Nasonia vitripennis (Hymenoptera: Pteromalidae) (F in Figure 7.4.1) (Fabritius and Klunker 1991). After parasitisation,

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Figure 7.4.1  Visitation of several interacting insect groups in different stages of decomposition. 1, fresh (a few hours after death); 2, active decay (2 weeks after death); 3, advanced decay (3 weeks after death); 4, advanced decay (2 months after death); 5, dry or skeletonised (2 years after death). See text for more explanation.

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one puparium can become the nutrition site for hundreds of new specimens of N. vitripennis, as the wasps are very small (see size comparison in F of Figure 7.4.1). In an advanced stage of decay, specimens of Piophilidae (Diptera) are attracted to the corpse (G in Figure 7.4.1). Their larvae (H in Figure 7.4.1) have the ability to jump as a defensive mechanism against their predators: the beetle Necrobia rufipes (Coleoptera: Cleridae) (I in Figure 7.4.1). In general, the longer the decomposition proceeds, a higher diversity of arthropod families arrive to benefit from the ever-­ changing habitat. This later stage (stage 4  in the figure) also attracts several species from the beetle family of Staphylinidae (K in Figure 7.4.1). They all prey on fly larvae. Another beetle that is frequently found in later stages of decay is Nicrodes littoralis (Coleoptera: Silphidae) (L in Figure 7.4.1). Its larvae can be found on cadavers a few weeks after death (Matuszewski et al. 2008). Additionally, other dipteran families can be frequently observed; for example, Muscidae. In stage 4, pupae of Fannia manicata (Diptera: Fanniidae) were found (J in Figure  7.4.1). These larvae also feed on decaying material.

The last species shown is the beetle Necrophorus vespilloides (Coleoptera: Silphidae) (M in Figure  7.4.1). This picture was taken during an experiment exposing small-­piglet carcasses. The beetle is associated with carcass, but is known for burying small-­ vertebrate carcasses and preparing them as food for their young (Scott 1998; Kalinová et  al. 2009). Therefore, they are mostly observed in succession experiments with small-­animal carcasses. Human cadavers are too big, so that N. vespilloides has, so far, not been reported in real forensic cases. This is one of the problems when succession studies are carried out with small-­animal carcasses; it is only partially representative for human-­sized cadavers. Furthermore, succession studies are conducted in several different places worldwide, which makes it difficult to compare and use them in other places. To understand the local fauna, it is strongly recommended to conduct such experiments and to compare the results to the insects actually found on corpses investigated in the local institute for forensic medicine. There are also species that occur predominantly in indoor locations. Specimens of the scuttle fly Megaselia scalaris (Diptera:

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Blowflies (Calliphoridae) are among the first species attracted towards carcass and are therefore very important for postmortem interval determination. Decomposing material is a protein source for themselves as well as breeding site for their progeny (Lane 1975; Putmann 1977; Smith and Wall 1997). If a freshly dead piglet is exposed in the open field, the first blowflies arrive within minutes. Two different behaviours of blowflies can be observed: extending their proboscis to absorb liquid or extending their ovipositor at distinct regions to find a proper place to oviposit (Figure 7.4.2). If a pregnant blowfly finds a suitable place, it will deposit its eggs in a clutch. After a few hours, it can be observed that blowflies prefer spots where eggs have already been deposited to oviposit as well, so that big egg aggregations are formed (Browne et al. 1969). Also, it seems that blowflies accumulate at convenient spots on the carcass (Figure 7.4.3); accumulated flies act as an attractant for other flies (Norris 1965). In general, blowflies try to find oviposition sites that offer the best conditions for their progeny.

The development of blowflies includes four stages: egg stage, larval stage, pupal stage and imago stage (Tao 1927). During the larval stage, three instars can be defined: first, second and third instar, where the latter is divided due to behavioural changes in feeding and postfeeding larvae. As the hatching first instar larvae can only feed on moist tissue, blowflies choose a body’s natural orifices or wounds to oviposit. This ensures a food supply for the hatching first instar larvae. The first three instars each undergo a moult in order to reach the next developmental stage; the stages can be distinguished by the number of respiratory slits at the posterior end of the larvae. The third instar stage lasts for longer than the first two. The larvae feed on the substrate as third instars, then leave the food source to find a suitable place for pupation, entering the postfeeding stage (Arnott and Turner 2008). About one-­third of the pre-­adult development time is spent in the postfeeding larval stage (Greenberg 1990).

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The general life cycle of blowflies

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Phoridae) are most often found on corpses discovered indoors, as they are very small and can enter even enclosed environments (Manlove and Disney 2008). Also, Fannia canicularis is often found on carcass indoors, its common name is little housefly. Fannia scalaris occurs indoors whenever the circumstances are primitive and neglected and is associated with lavatories and cesspits, which lead to the common name latrine fly (Benecke and Lessig 2001). To sum up, succession studies can be a useful tool to classify species in their relation to certain decomposition stages, although no clear cuts can be made from one stage to another. Moreover, every succession study is highly dependent on the habitat and climate. Nevertheless, it is important to know the local fauna to be able to judge the entomological findings in each individual case.

Figure 7.4.2  Blow flies on carcass: (a) Extended proboscis of blow fly. (b) Extended ovipositor of blow fly.

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Then, pupation sets in and the imago develops within the pupal case until eclosion (Lowne 1890). This last stage persists for about half of the time of the total development. Hatching of the adult flies completes the developmental cycle.

alcohol. If neither boiling water nor alcohol is available, storing the insects as cool as possible is advisable until ethanol can be added.

Identification of insects

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It is not complicated to collect the desired insect specimens from a corpse. Any liquid safe container can be used; additionally, a pair of tweezers and some ethanol (80%) or any other alcohol (60% and above), a piece of paper and a pencil. For collecting the insects from a body, one should decide for at least three areas to collect from; for example, head area, genitals and extremities. Take three containers, three small pieces of paper and label them properly with a pencil (only with a pencil, everything else will wash off): date, time, collection area, initials and location of the corpse during collection. Put the piece of paper inside the container and collect only from the area corresponding to the label. Use the pair of tweezers to collect larvae or crawling insects and put them in the container. You should not fill the container more than half; instead of overloading a container, prepare another one for the same collection area. Try to collect insects of different sizes and stages, and, if possible, take rather more specimens than less. However, more than approximately 100 specimens from one collection area are not necessary. Before closing the container, fill the container with ethanol or other alcohol (60% upwards). A more advanced way to collect specimens (in terms of equipment needed) is to pour boiling water over the collected insects (Adams and Hall 2003), wait a few minutes, dispose of the water and then add ethanol. This ensures that the larvae are dead immediately after boiling; otherwise they will survive a few hours within the alcohol and might blacken over time due to internal decomposition processes. However, if it is not possible to boil water during the time of collection, it is sufficient to just preserve the living specimens in ethanol or 60% or higher alcohol. If collecting flying insects, they can be preserved accordingly. Here, it is even less important to use boiling water first. Indoors as well as outdoors, it is important to search the wider surroundings of the body, as the postfeeding larvae migrate away from the corpse to find a safe place for pupation. Indoors, one should search under rugs, pillows, paper on the floor, behind floorboards and so on. The larvae have a tendency to fall down on purpose, so a hiding place on the floor is most likely. Outside, it can be difficult to determine how far the migrating larvae might have wandered, a search radius between 2 and 10  m is recommended (Amendt et al. 2011).

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Collecting

Most important in forensic casework involving entomological evidence is the identification of the insect species collected in association with the corpse or its surroundings. Correct identification of the insects collected is the foundation of all further estimations. One quick and easy way is the morphological identification using appropriate identification keys. The standard reference for the identification of blowflies is the book Blowflies (Diptera, Calliphoridae) of Fennoscandia and Denmark (Fauna Entomologica Scandinavica) by K. Rognes (1991). Another standard reference is A Manual of Forensic Entomology by K. G. V. Smith (1986): it includes an identification key for adults and larvae associated with carrion. The only equipment necessary for morphological-­species determination is a dissecting microscope with a proper light source. In general, it is easier to identify adult insect specimens than larval stages using their morphological features. It is advisable to have someone with a strong background in entomology perform identification. However, it may be impossible to identify an insect by means of its morphology, for example, due to damage; it might then be possible to use molecular identification tools. To ensure correct species identification, established molecular methods were transferred to the forensic field (Sperling et al. 1994; Stevens and Wall 1996; Stevens and Wall 1997; Wallman and Adams 1997; Benecke 1998). Analysis of mitochondrial DNA (mtDNA) and particularly of the cytochrome oxidase I (COI) gene appeared to be a useful tool in species identification among the subfamilies of Calliphoridae (Harvey et  al. 2003, 2008; Wallman et  al. 2005; Wells and Williams 2005; Wells et al. 2007; Reibe et al. 2009). The molecular method can be a reliable alternative to the morphological one, but the latter is cheaper, quicker and safer, if an expert performed it.

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7.4.3 Collecting, storing and identification of insects from a corpse

Storage The insects can be stored in ethanol for a long time. They do not have to be stored in a cooler or freezer once they are preserved in

7.4.4  Estimation of postmortem interval The term ‘postmortem interval’ can be misleading when using it in combination with forensic entomology. With the help of the age of insect larvae feeding on a corpse, it can be estimated how long the corpse has been infested by insects. However, a person can already be infested when he is still alive; for example, a neglected person with severe wounds on a smoker’s leg (Benecke et al. 2004), or as reported from soldiers in the First World War, wounds that would have led to amputation when the blowfly larvae had not fed the necrotic tissue to clean the wounds (Buechner 1965). By contrast, a person can be long dead and the larval age can still be a few days, when the corpse has been stored in a place with no access for the insects and has been put outside just a few days prior to finding. Therefore, one must take into account every aspect before defining about a postmortem interval. In the latter

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Corpses in houses or apartments are frequently found in late stages of decay infested by larvae of Calliphoridae, Phoridae, Muscidae or Sarcophagidae (Reibe and Madea 2010b). In such cases, the person was usually socially isolated, leading to delayed discovery of their bodies (Archer et al. 2005), and a postmortem interval determination is aggravated, since it is unclear how promptly the insects found the body and started laying eggs. Bodies can be colonised by insects in several different locations including poorly accessible environments (Goff 1991; Reibe et al. 2008). To estimate the colonisation period, it is therefore important to know how soon the insects can obtain access from outdoors (Anderson 2011; Pohjoismäki et al. 2010; Reibe and Madea 2010a).

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In forensic casework, two different methods are frequently used to calculate a postmortem interval. The first uses isomegalen or isomorphen diagrams, by which the lengths or the developmental stage of the larvae are combined as a function of time and mean ambient temperature in a single diagram (Grassberger and Reiter 2001). According to its originators, this method is optimal only if the body and therefore the larvae were not undergoing fluctuating temperatures, for example, in an enclosed environment where the temperature was nearly constant. Unfortunately, isomegalen and isomorphen diagrams are not available for all forensic-­ relevant insect species.

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case, if the insect evidence does not fit the decomposition stage of the corpse, the police can use that additional piece of information about how long the corpse has been available for insects. However, from the insect evidence alone, no information can be actually given about the postmortem interval in the latter case. One always has to be aware of possible sources of error. A postmortem interval estimation can be done by relating certain species to decomposition stages in a timely manner, and therefore achieve a rough idea about the colonisation time. Another method is to calculate the age of, for example, blowfly larvae feeding on the corpse and thereby giving a minimum time interval the person has been dead. This is only possible if the calculation is done for larvae of the first colonisation wave. If the life cycle of the first larvae has already been completed and the adult flies have started a new infestation of the corpse, it is only safe to calculate the time span for the first completed life cycle. The biological basis for the calculation of the larval age is their temperature-­dependent development. Larval growth rate depends on its body temperature, which is directly influenced by environmental conditions as the ambient temperature and the heat generated by maggot aggregations (Slone and Gruner 2007). Also, each species has its own temperature-­dependent growth rate. Insects are poikilothermic organisms. Their body temperature changes with ambient temperature. The metabolic rate of poikilotherm animals shows the same temperature dependency as the reaction kinetics in a biochemical system: the rate of a chemical reaction is increased twofold for each rise of 10 °C in temperature (van’t Hoff ’s reaction rate–temperature rule) (Wehner and Gehring 1995). For insect development, a relationship between ambient temperature and duration of developmental processes is long known. There is a temperature zone where the development rate is optimal for each species; furthermore, temperature thresholds exist below or above that optimum where no development will take place. One prerequisite for estimating a postmortem interval using larval age is to estimate the temperature regime of the time and place of larval development. One method to approximate the temperature is to record the temperature at the desired place for a few days and compare the values to data recorded at the nearest weather station. A regression analysis is applied to both data sets to get a formula that can then transform the data from the weather station covering the desired time frame into the temperature values that most likely influenced the developing larvae. The second requirement for the calculation of larval age as a function of temperature is the correct identification of the species and the third is the determination of the developmental progress. This means determination of either the stage or the length of the maggots is required, depending on the reference data set and the method that is used for the further calculation. Before discussing calculating larval ages, the time span between exposure of a corpse and infestation by insects should be considered, as it has to be added to the larval age in order to get a more precise result of the postmortem interval estimation.

Accumulated degree days or hours method The second method of calculating a postmortem interval estimates the accumulated degree days or hours (ADD or ADH). ADH values represent a certain number of ‘energy hours’ that are necessary for the development of insect larvae. The degree day or degree hour concept assumes that the developmental rate is proportional to the temperature within a certain species-­specific temperature range (overview in Higley and Haskell 2009). However, the relationship of temperature and development rate (reciprocal of development time) is typically curvilinear at high and low temperatures and linear only in between, which limits the use of the method. The formula for calculating ADH is given as follows:

ADH T

0 ,

where T is the development time (hours for ADH, days for ADD), θ is the ambient temperature and the minimum developmental threshold temperature θ0 is a species-­specific value (the so-­called development zero), which is the x-­intercept (i.e., an extrapolation of the linear approximation of the reciprocal of time for development). This value has no biological meaning: it is the mathematical consequence of using a linear regression analysis (Higley and Haskell 2009).

MEDICAL ASPECTS OF DEATH

determine the results’ reliability; especially when the latter is being used in real cases in court. It is also important to include sources of error to make sure a standard error can be calculated to avoid pseudo-­precise results.

7.4.5  Case reports Case 1 In this case report, the first developmental cycle of the blowfly Lucilia sericata had been completed when the body was found. Therefore, the calculation was based on another species, M. scalaris, which is additionally known for being associated with indoor bodies.

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One basic assumption for using the ADH/ADD method is that the ADH or ADD value for completing a developmental stage stays constant within certain temperature thresholds. For example, a developmental duration for reaching a certain stage in 14 days at 25  °C results in 238 ADD when a base temperature of 8  °C is assumed. A developmental duration for the same developmental stage of 19 days at 21 °C results in 231 ADD; both ADD values are in the same range. So, the necessary amount of ‘energy hours’ to reach the specific developmental stage is equal, whether it was 25 °C or 21 °C. With existing data for larval development, one can calculate the ADH/ADD values needed for certain stages to be completed and then easily calculate the larval age in a real case. The limits, however, of the ADH method lie within the temperature ranges when the reciprocal development is not linear anymore. In these areas, the basic condition for the method is not fulfilled and the method should not be used.

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A third approach to calculate larval age is to model larval development in dependency of temperature using more complex assumptions than a linear relationship (Ikemoto and Takai 2000; Tarone and Foran 2008; Reibe et al. 2010). It is important to sample large amounts of data for the model to find the best possible fit. Also, the results of the model should be tested and compared to the conventional methods performing large screening experiments with dead piglets and known postmortem intervals to

A woman was found dead in her apartment in January 2009. She had not been seen for approximately 40 days. She was lying on a mattress on the floor. The window in the room was closed and the shutters were down. All other windows and shutters in the rest of the flat were closed, apart from a tiny window in the bathroom. Next to her several pills were found. She was clothed in a t-­shirt, thin capri pants and woollen socks. Most of the body was mummified, and the face was partially skeletonised (Figure 7.4.4a,b).

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Figure 7.4.4  Case 1: (a) and (b) State of the corpse. (c) Open abdominal cavity. (d) Close-­up inside abdominal cavity showing hundreds of larvae and pupae of Megaselia scalaris.

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Entomological evidence Empty pupal cases of L. sericata were found adhering to the woollen socks. A live specimen of the beetle N. rufipes was crawling out of the clothes during autopsy. Dead adult specimens were collected, as well as empty pupal cases of M. scalaris. When the abdominal cavity was opened, thousands of larvae and pupae of M. scalaris were revealed (Figure 7.4.4c,d).

brought to the institute in a large plastic bag. Blowfly larvae were already crawling out of the bag. The bag was opened on 3 September 2008 at approximately 4:00 p.m. (Figure 7.4.5b). The head was found 1 day later, and more parts of the torso and the legs were found on 8 September 2008. When the parts were found, it was unclear who the person was and if or how long they had been missing.

Entomological evidence

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We identified three different species of the family of flies: Lucilia caesar, Calliphora vicina and Sarcophaga sp. All three species are typically early visitors of corpses. C. vicina and L. caesar are additionally described as preferring shadowy areas. Neither in the plastic bag nor on the body parts nor at the finding site pupal cases were found.

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For larval-­age calculation, we decided to use larvae of C. vicina for two reasons. Firstly, we had generated our own developmental data for C. vicina originating from Bonn, which is always better than using data from other countries or climatic regions. Secondly, we could use our own larval-­age calculator (Reibe et al. 2010) in which we had already implemented the data for C. vicina. As temperature data, we used data from the Meteorological Institute of the University of Bonn. We did not apply a regression analysis because we had only the chance to collect reference temperature data for 5 hours on one afternoon; this was not enough data for the regression analysis, as the nightly temperatures differed greatly from the temperatures at daytime. Instead, we set the error on the temperature data to 2 °C, as we wanted to avoid a pseudo-­precise result. The result for the age calculation of the average 12-­mm-­long larvae was 115 ± 5 hours.

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The temperature in the room when the corpse was found was 21 °C. It was assumed that the temperature remained almost constant, as the windows were closed and the shutters were down. L. sericata takes 19  days to complete its life cycle (author’s own developmental data (Reibe); data of Grassberger and Reiter (2001) suggest 16 days). M. scalaris completes development at a 12:12 photoperiod after about 37 days (Trumble and Pienkowski 1979). Since the individual’s missing period was about 40 days, postmortem interval determination using developmental data of M. scalaris was much more accurate than the postmortem interval calculated with data for L. sericata. The missing interval will most likely be very close to the real postmortem interval, as a suicide note was found dated shortly before Christmas.

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In September 2008, several body pieces were found in a tree-­ covered area not far from the city centre of Bonn, Germany. The first pieces brought to the Institute of Legal Medicine were parts of the torso of a woman. When the parts were found, they were lying openly under large trees (Figure  7.4.5a), but they were

Figure 7.4.5  Case 2: (a) Area where the torso of the body was found. (b) Torso of the body packed in a plastic bag with several fly larvae attached.

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References and further reading

sic studies: Validation and use in practice. Forensic Science International 98(3): 157–168. Benecke, M. and Lessig, R. (2001). Child neglect and forensic entomology. Forensic Science International 120 (1–2): 155–159. Benecke, M., Josephi, E., and Zweihoff, R. (2004) Neglect of the elderly: forensic entomology cases and considerations. Forensic Science International 146 (Suppl): 195–199. Bharti, M. and Singh, D. (2003). Insect faunal succession on decaying rabbit carcasses in Punjab, India. Journal of Forensic Sciences 48 (5): 1133–1143. Browne, L.B., Bartell, R.J. and Shorey, H.H. (1969). Pheromone-­mediated behaviour leading to group oviposition in the blowfly Lucilia cuprina. Journal of Insect Physiology 15: 1003–1014. Buechner, F. (1965) Pläne und Fügungen. Lebenserinnerungen eines deutschen Hochschullehrers. München: Urban und Schwarzenberg. Denlinger, D.L. (1994) Metamorphosis behaviour of flies. Annual Review of Entomology 39: 243–266. Eberhardt, T.L. and Elliot, D.A. (2008). A preliminary investigation of insect colonisation and succession on remains in New Zealand. Forensic Science International 176 (2–3): 217–223. Fabritius, K. and Klunker, R. (1991). Die Larven-­und Puparienparasitoide von synanthropen Fliegen in Europa. Merkblätter über angewandte Parasitenkunde und Schädlingsbekämpfung 32(1): 1–24. Goff, M.L. (1991). Comparison of insect species associated with decomposing remains recovered inside dwellings and outdoors on the Island of Oahu, Hawaii. Journal of Forensic Science 36 (3): 748–753. Goff, M.L. (1993). Estimation of postmortem interval using arthropod development and successional patterns. Forensic Science Review 5 (2): 81–94. Grassberger, M. and Frank, C. (2004). Initial study of arthropod succession on pig carrion in a central European urban habitat. Journal of Medical Entomology 41 (3): 511–523. Grassberger, M. and Reiter, C. (2001). Effect of temperature on Lucilia sericata (Diptera: Calliphoridae) development with special reference diagram. Forensic Science to the isomegalen-­and isomorphen-­ International 120 (1–2): 32–36. Greenberg, B. (1990). Behavior of postfeeding larvae of some Calliphoridae and a Muscid (Diptera). Annals of the Entomological Society of America 83: 1210–1214. Greene, G. (1996). Rearing techniques for Creophilus maxillosus, a predator of fly larvae in cattle feedlots. Journal of Economic Entomology 89 (4): 848–851. Harvey, M.L., Dadour, I.R. and Gaudieri, S. (2003). Mitochondrial DNA cytochrome oxidase I gene: potential for distinction between immature stages of some forensically important fly species (Diptera) in western Australia. Forensic Science International 131 (2–3): 134–139. Harvey, M.L., Gaudieri, S., Villet, M.H. and Dadour, I.R. (2008). A global study of forensically significant calliphorids: Implications for identification. Forensic Science International 177 (1): 66–76. Higley L.G. and Haskell N.H. (2009). Forensic Entomology, The Utility of Arthropods in Legal Investigations, 2nd edn, pp. 389–405. Boca Raton, FL: CRC Press. Ikemoto, T. and Takai, K. (2000). A new linearized formula for the law of total effective temperature and the evaluation of line-­fitting methods with both variables subject to error. Environment Entomology 29: 671–682.

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That led to the 29 August 2008 in the evening as a starting point for oviposition of blowflies. Due to the heavy dismembering of the body and the resulting amount of blood, it can be assumed that the blowflies were highly stimulated to deposit their eggs. The identity of the woman was proven later via dental records. She was a teacher reported missing in the week before 29 August 2008. In the week after the findings, a man was arrested in the Netherlands who appeared covered in blood at a doorstep of a family wanting to use the phone. He was disoriented and confessed to the police the murder of his German girlfriend. He reported that on Friday, 29 August 2008, he got into a fight with his girlfriend and killed her inside her own house with a knife and also started dismembering her. He wanted to hide her body in the garden and started digging a hole in the evening. The hole was already filled up after putting the legs inside, so the offender decided to transport the rest of the body parts to the nearby tree-­covered area, where people usually walk their dogs. To sum up, the events started on the afternoon of 29 August 2008 and went on until the morning hours of 30 August 2008, so the calculated larval age represents the real postmortem interval very nicely.

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Adams, Z. and Hall, M. (2003). Methods used for the killing and preservation of blowfly larvae, and their effect on post-­mortem larval length. Forensic Science International 138 (1–3): 50–61. Amendt, J., Richards, C.S., Campobasso, C.P. et al. (2011). Forensic entomology: applications and limitations. Forensic Science, Medicine, and Pathology 7(4): 379–392. Anderson, G.S. (2011). Comparison of decomposition rates and faunal colonization of carrion in indoor and outdoor environments. Journal of Forensic Sciences 56 (1): 136–142. Anderson, G.S. and VanLaerhoven, S.L. (1996). Initial studies on insect succession on Carrion in southwestern British Columbia. Journal of Forensic Science 41: 617–625. Archer, M. (2003). Annual variation in arrival and departure times of carrion insects at carcasses: Implications for Succession Studies in Forensic Entomology. Australian Journal of Zoology 51: 569–576. Archer, M.S., Bassed, R.B., Briggs, C.A. and Lynch, M.J. (2005). Social isolation and delayed discovery of bodies in houses: the value of forensic pathology, anthropology, odontology and entomology in the medico-­legal investigation. Forensic Science International 151 (2–3): 259–265. Arnaldos, I., Romera, E., García, M.D. and Luna, A. (2001). An initial study on the succession of sarcosaprophagous Diptera (Insecta) on carrion in the southeastern Iberian peninsula. International Journal of Legal Medicine 114 (3): 156–162. Arnott, S. and Turner, B. (2008). Post-­feeding larval behaviour in the blowfly, Calliphora vicina: Effects on post-­mortem interval estimates. Forensic Science International 177 (2–3): 162–167. Benecke, M. (1998). Random amplified polymorphic DNA (RAPD) typing of necrophageous insects (Diptera, Coleoptera) in criminal foren-

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Stevens, J. and Wall, R. (1996). Species, sub-­species and hybrid populations of the blowflies Lucilia cuprina and Lucilia sericata (Diptera: Calliphoridae). Proceedings of the Biological Society 263 (1375): 1335–1341. Stevens, J. and Wall, R. (1997). Genetic variation in populations of the blowflies Lucilia cuprina and Lucilia sericata (Diptera: Calliphoridae). Random amplified polymorphic DNA analysis and mitochondrial DNA sequences. Biochemical Systematics and Ecology 25: 81–97. Tao, S. (1927). A comparative study of the early larval stages of some common flies. American Journal of Epidemiology 7: 735–761. Tarone, A.M. and Foran, D.R. (2008). Generalized additive models and Lucilia sericata growth: assessing confidence intervals and error rates in forensic entomology. Journal of Forensic Sciences 53 (4): 942–948. Trumble, J. and Pienkowski, R. (1979). Development and survival of Megaselia scalaris (Diptera: Phoridae) at selected temperatures and photoperiods. Proceedings of the Entomological Society of Washington 81 (2): 207–210. Wallman, J. and Adams, M. (1997). Molecular systematics of Australian carrion-­ breeding blowflies of the genus Calliphora (Diptera: Calliphoridae). Australian Journal of Zoology 45: 337–356. Wallman, J.F., Leys, R. and Hagendoorn, K. (2005). Molecular systematics of Australian carrion-­breeding blowflies (Diptera: Calliphoridae) based on mitochondrial DNA. Invertebrate Systematics 19: 1–15. Watson, E.J. and Carlton, C.E. (2005). Insect succession and decomposition of wildlife carcasses during fall and winter in Louisiana. Journal of Medical Entomology 42 (2): 193–203. Wehner, R. and Gehring, W. (1995). Zoologie. Berlin: Thieme. Wells, J.D., Wall, R. and Stevens, J.R. (2007). Phylogenetic analysis of forensically important Lucilia flies based on cytochrome oxidase I sequence: a cautionary tale for forensic species determination. International Journal of Legal Medicine, Science and Law 121: 229–233. Wells, J. and Williams, D. (2005). Validation of a DNA-­based method for identifying Chrysomyinae (Diptera: Calliphoridae) used in a death investigation. International Journal of Legal Medicine 1–8.

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Kalinová, B., Podskalská, H., Růžička, J. and Hoskovec, M. (2009). Irresistible bouquet of death—­ how are burying beetles (Coleoptera: Silphidae: Nicrophorus) attracted by carcasses. Naturwissenschaften 96 (8): 889–899. Lane, R.P. (1975). An investigation into blowfly (Diptera: Calliphoridae) succession on corpses. Journal of Natural History 9: 581–588. Lowne, B. (1890). The Anatomy, Physiology, Morphology and Development of the Blow-­f ly. London: RH Porter. Manlove, J.D. and Disney, R.H.L. (2008). The use of Megaselia abdita (Diptera: Phoridae) in forensic entomology. Forensic Science International 175 (1): 83–84. Matthews, R. (1974). Biology of Braconidae. Annual Review of Entomology 19: 15–32. Matuszewski, S., Bajerlein, D., Konwerski, S. and Szpila, K. (2008). An initial study of insect succession and carrion decomposition in various forest habitats of Central Europe. Forensic Science International 180 (2–3): 61–69. Norris, K. (1965). The bionomics of blow flies. Annual Review of Entomology 10: 47–68. Pohjoismäki, J.L.O., Karhunen, P.J., Goebeler, S. et  al. (2010). Indoors forensic entomology: colonization of human remains in closed environments by specific species of sarcosaprophagous flies. Forensic Science International 199 (1–3): 38–42. Putmann, R. (1977). Dynamics of the blowfly, Calliphora erytrocephala, within carrion. Journal of Animal Ecology 46: 854–866. Reibe, S., Doetinchem, P.V. and Madea, B. (2010) A new simulation-­ based model for calculating post-­mortem intervals using developmental data for Lucilia sericata (Dipt.: Calliphoridae). Parasitology Research 107 (1): 9–16. Reibe, S. and Madea, B. (2010a). How promptly do blowflies colonise fresh carcasses? A study comparing indoor with outdoor locations. Forensic Science International 195 (1–3): 52–57. Reibe, S. and Madea, B. (2010b). Use of Megaselia scalaris (Diptera: Phoridae) for post-­mortem interval estimation indoors. Parasitology Research 106 (3): 637–640. Reibe, S., Strehler, M., Mayer, F. et  al. (2008). Dumping of corpses in compost bins—­two forensic entomological case reports. Archiv für Kriminologie 222 (5–6): 195–201. Reibe, S., Schmitz, J. and Madea, B. (2009). Molecular identification of forensically important blowfly species (Diptera: Calliphoridae) from Germany. Parasitology Research 106 (1): 257–261. Rognes, K. (1991). Blowflies (Diptera, Calliphoridae) of Fennoscandia and Denmark. Leiden: Scandinavian Science Press. Scott, M.P. (1998). The ecology and behavior of burying beetles. Annual Review of Entomology 43: 595–618. Slone, D. and Gruner, S. (2007). Thermoregulation in larval aggregations of carrion-­ feeding blow flies (Diptera: Calliphoridae). Journal of Medical Entomology 44 (3): 516–523. Smith, K.E. and Wall, R. (1997). The use of carrion as breeding sites by the blowfly Lucilia sericata and other Calliphoridae. Medical Veterinary Entomology 11 (1): 38–44. Smith, K.G.V. (1986). A Manual of Forensic Entomology. London: Trustees of the British Museum (Natural History). Sperling, F.A., Anderson, G.S. and Hickey, D.A. (1994). A DNA-­based approach to the identification of insect species used for postmortem interval estimation. Journal of Forensic Sciences 39 (2): 418–427.

7.5  Postmortem Injuries Natural biological as well as artefactual changes that a human body or its remains may undergo after death are complex and mostly unpredictable, since these phenomena are influenced by a broad range of variables (Tsokos 2005). Postmortem artefacts may mimic antemortem injuries for the unaware. Hence, the wrong conclusions may lead the investigative inquiries into the wrong direction, or, in the worst case, to a miscarriage of justice. On the one hand, some postmortem changes may render a careful external examination of a corpse difficult; for example, putrefactive skin changes may conceal cutaneous injuries due to external violence against the body before death. On the other hand, when found in curious death scene scenarios, postmortem injuries due to animal depredation or following resuscitation procedures may be interpreted falsely (Tsokos et al. 1999; Byard et al. 2002; Tsokos 2005).

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7.5.2 Putrefactive changes that may mimic antemortem injuries

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Putrefaction is a common problem encountered in forensic pathology that may result in considerable distortion and modification of tissues. After death, when homeostasis ceases, anaerobic bacteria (mostly Clostridia and Proteus species) migrate from the gut into blood vessels and into tissue where they multiply and spread throughout the whole body. Since open wounds are a portal of entry for microorganisms from the outside environment, those people who die from or with wounds that are wide open and extending far down within the tissues and body cavities show accelerated rates of putrefaction.

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After death, postmortem evaporation of tissue fluids leads to drying up of mucosal surfaces, for example, of the lips, the tip of the tongue, the glans of the penis, the glans of the clitoris or the pudendal lips, thus resulting in a hardened, light to dark brownish appearance of the affected mucosa. A dark brownish, sometimes blackish, appearance of parts of the tip of the tongue due to postmortem desiccation should not be confused with an epileptic tongue bite. Postmortem drying of mucosal surfaces occurs more rapidly and is more pronounced under dry and warm environmental conditions. If the eyes remain open after death, the area of the sclera exposed to air dries out, leaving a brownish-­blackish, occasionally reddish band-­ like scleral desiccation zone (Figure  7.5.1) referred to as ‘tache noir’ in the European forensic literature.

The characteristic bloating of a putrefied body as reflected by swelling of the face, distension of the abdomen and distension of the penis and scrotum (Figure 7.5.2a) is a result of bacterial gas formation. On palpation, crepitus is often noticed. Differential diagnoses of putrefactive gas formation into tissues include vital subcutaneous emphysema due to blunt impact trauma, iatrogenic manipulation or tension pneumothorax. Putrefactive skin changes may mask traumatic injuries, and, vice versa, a brownish-­black discolouration of the skin as a result of putrefaction may be misinterpreted as a sign of antemortem injury. Especially in prolonged postmortem intervals, the manifestation of putrefaction can cause interpretational problems when the skin of the deceased acquires a brownish-­blackish appearance that may conceal, for example, current marks, defensive injuries or puncture marks (Figure 7.5.2b). At later stages of putrefaction, the neck and face may become grotesquely bloated and gas formation within the pharynx and the floor of the mouth can cause considerable protrusion of the tongue (Figure 7.5.2c) that should not be misinterpreted as projection of the tongue due to upwards pressure on larynx and root of the tongue in cases of ligature strangulation or hanging. The purging of putrefactive fluid from mouth and nostrils (Figure 7.5.2d,e) may be misinterpreted as effusion of blood due to a violent mode of death. Purging of putrefactive fluid is the result of an increase of intrathoracic pressure due to bacterial gas formation within the thoracic cavities in putrefied bodies leading to leakage of liquefied tissue stained by haemolysis. When putrefactive-­fluid-­filled skin blisters (Figure 7.5.2f) are torn open, usually following application of pressure of friction, skin slippage (Figure 7.5.2g) results. Venous marbling, a branching outline of an arborescent greenish-­brownish pattern of the skin (Figure 7.5.2h) due to haemolysis in combination with postmortem bacterial invasion of subcutaneous venous vessels, may appear occasionally as bruising to the inexperienced (Figure 7.5.2i,j). Especially when circumscribed and not diffusely distributed over the whole body surface, discoloured areas of putrefaction may be confused with bruises at any stage of the process of putrefaction. Unusual skin lesions caused by the disruption and dehiscence of healing surgical wounds can be created by putrefaction and may mimic incised or stab wounds (Byard et al. 2005, 2006). This possibility should be considered especially when symmetrical, cleanly incised wounds are found in a putrefied body with bloating.

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7.5.3 Maceration

Figure 7.5.1  Scleral desiccation. The area of the sclera exposed to air has dried out, leaving a brownish-­reddish, band-­like zone that should not be confused with scleral haemorrhage.

The term maceration is strictly defined to sterile autolysis of an unborn foetus that has died in utero enclosed within the amniotic cavity. Despite this fact, in some textbooks, the term maceration is also used for skin slippage phenomena in putrefied bodies. The most prominent finding is skin slippage with underlying brownish-­blackish discolouration of tissue. If the amniotic cavity

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Figure 7.5.2  (a) Distension of the penis and scrotum in a putrefied body. (b) Brownish-­blackish appearance of the skin that may mask traumatic injuries such as current marks, defensive injuries or puncture marks. (c) Postmortem protrusion of the tongue due to putrefactive gas formation and purging of putrefactive fluid from the nostrils. (d) Postmortem purging of putrefactive fluid from the mouth. (e) Purging of putrefactive fluid from the mouth and nostrils that should not be misinterpreted as effusion of blood due to a violent mode of death. (f) Putrefactive-­fluid-­filled skin blisters. (g) Skin slippage in a putrefied body. (h) Venous marbling. (i) Venous marbling that may appear as bruising to the inexperienced. (j) Venous marbling.

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In bodies recovered from water (and also those found in moist environments), the skin of the hands and of the feet shows a whitish discolouration of the epidermis associated with swelling, wrinkling and skin detachment up to a glove-­ like peeling (Figure 7.5.3a). The extent of this finding, referred to as washerwoman’s skin, depends on the period of time a body was exposed to water, but is highly variable because of environmental factors (especially water temperature: high water temperature leads to acceleration of this phenomenon). The formation of washerwoman’s skin starts on the palms of the hands and feet where the horny layer of the skin is thicker than anywhere else on the body (Lunetta and Modell 2005). The finding of washerwoman’s skin is a pure postmortem phenomenon as a result of soaking of the horny layer of the epidermis. On bodies floating in water, ship’s propellers produce characteristic large, incised cuts lacking any adjacent soft-­tissue haemorrhage (Figure 7.5.3b). The decision whether or not such wounds were actually sustained post mortem or while the person was still alive is occasionally difficult, since even vitally acquired wounds appear bloodless when the body has spent a considerable time span in water due to washing out of blood from open wounds (Figure 7.5.3c). The propellers of large vessels may lead to amputation of limbs or even total dismemberment of the body that has to be differentiated from such cases where a body was thrown into water after mutilation and dismemberment in order to hide body parts and/or to make identification more difficult (Lunetta and Modell 2005). The common posture of bodies floating in water is in a prone position with the head, hands and feet hanging downwards, so that the nose, forehead, knees, backs of the hands, tips of the fingers and toes may sustain abrasions in shallow water where these body parts come in contact with the bottom (Figure 7.5.3d–f). In drowning victims or people whose death led to their deposition in water or when a body has been dumped in an aquatic environment, postmortem artefacts on the body surface due to aquatic living structures are often observed. Depending on the given aquatic environment, a broad variety of marine-­living structures is involved in tissue destruction of bodies recovered from water (Byard et  al. 2002; Lunetta and Modell 2005; Tsokos 2005). Crustaceans are considered the most effective tissue removers in water, typically leaving oval to round, crater-­like dermal lesions of varying size (Figure 7.5.3g,h). Sea lice (Natatolana woodjonesi), approximately 2.5-­cm-­long isopods that live on the surface of sand in shallow waters, are found in aquatic environments worldwide. They

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7.5.4 Postmortem injuries in bodies recovered from water

are active swimmers when searching for food. Sea lice can reduce a dead fish to a skeleton in a few hours. Sea lice may cause bizarre mutilation of submerged bodies (Byard et  al. 2002) (Figure 7.5.3i,j). Although rarely observed in daily routine, starfish may be responsible for superficial dermal haematomas found on bodies recovered from water. These are caused by their peculiar feeding mechanism, namely, sucking. When inflicted in the early postmortem interval, these haematomas have been reported to be easily mistaken for vitally sustained haematomas. When such starfish-­induced injuries are present upon a body that was recovered from water or when starfish themselves are found adhering to the corpse, this finding indicates that the body must have been, at least for a short period of time, located at the bottom of a body of water, since starfish are not able to swim and are only found at the bottom of their particular aquatic living space. Shark bites occur every so often in intact and fragmented bodies found at sea. Shark bites tend to have cleanly incised edges with significant loss of skin and underlying soft tissue. The pattern of typical shark bites corresponding to the triangular shape of their teeth can often be clearly identified. If the attack was not witnessed and only parts of the body are recovered, it is usually impossible to determine from the injuries whether or not death was the result of the shark attack with leaching of blood into water and bloodless appearance of the wound or if the injuries merely represent postmortem feeding on human remains (Byard et al. 2002). Adipocere formation (Figure  7.5.4), manifesting as a grey waxy substance deriving from the body fat, alters both the surface of the body as well as the postmortem injuries caused by marine life forms (Lunetta and Modell 2005).

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has been opened prior to the delivery of a stillborn foetus, bacterial putrefaction will alter the morphological picture of maceration. The presence of maceration without any putrefactive changes in a recently delivered child is indicative of stillbirth.

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7.5.5  Postmortem injuries in fire victims In burned bodies, morphological findings cover a broad spectrum, depending mainly on the temperature applied to the body, the duration of exposure of the body to heat and if the body is directly affected by open flames (Bohnert 2004). Artefactual changes in burned bodies may simulate antemortem injuries; for example, epidural haematoma caused by the heat. Heat-­induced postmortem defects of the calvarium may be misinterpreted as a gunshot-­exit wound. Cratering of the external table (‘external bevelling’) of flat bones such as the calvarium is a well-­known phenomenon of gunshot-­exit wounds. However, a similar finding with external bevelling can be observed occasionally in charred bodies as a pure postmortem injury (Bohnert et al. 1997; Hausmann and Betz 2002; Tsokos 2011) (Figure 7.5.5a). Skin splitting in burned bodies showing no signs of soot deposits or other heat-­mediated effects is another postmortem injury seen in corpses recovered from fire scenes (Figure 7.5.5b). These skin splittings originate during the cooling process of the body after the fire has stopped.

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Figure 7.5.3  (a) Washerwoman’s skin in a body recovered from water. (b) Postmortem injury following a cut from a ship’s propeller. (c) Wound of postmortem origin in a body recovered from water. Wounds, even those acquired ante mortem, appear bloodless when the body has spent a considerable time span in water. (d) Postmortem floating injuries on the face. (e) Postmortem floating injuries on the face. (f) Postmortem floating injuries on the back of the hand. (g) Postmortem injuries caused by crustaceans in a body recovered from water. (h) Postmortem injuries caused by crustaceans. These wounds are typically oval to round, crater-­like and of varying size. (i) Facial postmortem injuries caused by sea lice (Natatolana woodjonesi). Courtesy of Professor Jules Kieser, Otago, New Zealand. (j) Auricular postmortem defects caused by sea lice (N. woodjonesi). Source: Courtesy of Professor Jules Kieser, Otago, New Zealand.

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Heat-­mediated changes mimicking washerwoman’s skin are another postmortem artefact in fire victims. Usually, washerwoman’s skin is seen after prolonged exposure to water in drowning deaths. However, corresponding skin changes with a whitish discolouration and wrinkled detachment of the epidermis can also be found on the hands and feet in fire victims (Bohnert and Pollak 2003).

7.5.6  Postmortem animal depredation

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Postmortem animal depredation is a substantial part of the taphonomic processes a body undergoes after death. While the occurrence and presentation of such injuries are well known to

forensic pathologists, the morphological appearance of these injuries may be misinterpreted by police officers or members of other investigating authorities. Postmortem injuries may be inflicted by all kinds of animals irrespective of their size or environmental origin either from land, sea or air. The discrimination between antemortem injury and postmortem injury generally presents no difficulties due to the total absence of haemorrhage and reddening in the tissue adjacent to the wound margins as well as the lack of any vital reaction under the microscope in the latter cases. Low ambient temperature has, in addition to slowing the onset and extent of postmortem changes, considerable impact on a delayed manifestation of odour of the body, thus making the human remains less olfactorily absorbing for carnivores and rodents. The most effective tissue removers are insects and rodents (Patel 1994; Tsokos et al. 1999). Skin holes and soft-­tissue defects, for example, made by maggots, especially when overlapped by tissue shrinkage due to mummification with resulting enlargement of the defects, can mimic gunshot wounds or other mechanical tissue defects that were sustained before death, for example, as a result of stabbing with sharp-­pointed instruments such as knives or scissors. Skin and soft-­tissue artefacts caused by rodents may occur as early as within the first hour post mortem (Tsokos et  al. 1999). In the majority of injuries inflicted post mortem by rodents, the wounds have a circular appearance and the wound margins are finely serrated showing irregular edges (Figure 7.5.6a–c). Very small parallel series of cutaneous lacerations deriving from the biting action of the upper and lower pairs of the rodents’ incisors are diagnostic for rodent activity. However, the determination of a distinct rodent species (e.g., rats, mice) solely based on the morphological appearance of the damage to skin and soft tissue is often unconvincing. The finding of rodent excrement as the connecting link for the

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Figure 7.5.4  Adipocere formation in a body recovered from water.

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Figure 7.5.5  (a) External bevelling of the calvarium in a charred body. This is a pure postmortem artefact. Note that the excavation of the outer table of the cranium shows no charring or soot deposits in contrast to the surrounding parts of the occipital bone, thus indicating that the occipital defect occurred post mortem during the postmortem cooling process. (b) Postmortem skin splitting in a burned body.

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Figure 7.5.6  (a) Postmortem facial injuries inflicted post mortem by rodents. (b) Postmortem injuries inflicted post mortem by rodents. The wound margins are finely serrated and show irregular edges. (c) Postmortem injuries inflicted post mortem by rodents. (d) Postmortem defects on the forearm inflicted post mortem by rodents. (e) Postmortem injuries inflicted post mortem by a domestic dog. (f) Postmortem injuries inflicted post mortem by a domestic dog. (g) Claw-­induced linear scratch-­type abrasions in the vicinity of the damaged skin areas due to postmortem animal interference by a domestic dog. (h) Postmortem skin lesions caused by ants.

diagnosis of rodent interference to human remains has been proposed, thus providing the possibility of distinguishing DNA sequences of the animal itself from those of food residues of human origin (Höss et al. 1992). A broad range of carnivores can be involved in the postmortem destruction of corpses located in open spaces or indoors

(e.g., wild animals such as foxes and big cats or domestic animals such as dogs and cats). The wound margins caused by carnivores often appear more regular than those caused by rodents, and V-­shaped or rhomboid punctured wounds are often seen upon the intact skin in the immediate vicinity to the actual wound margins (Figure  7.5.6e,f). Such stab-­wound-­like defects represent

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Figure 7.5.7  (a) Postmortem skin lesions caused by cockroaches. (b) Pseudo-­ligature mark due to contact blanching of livor mortis. (c) Vibices in livor mortis. (d) Postmortem skin injury with the typical yellowish-­brownish to golden, shiny appearance. (e) Leaking out of gastric juice and bile post mortem. (f) Skin lesions caused by external cardiopulmonary resuscitation. (g) Defibrillation marks.

canine tooth marks of carnivore origin. An additional criterion for animal depredation by carnivores is the presence of claw-­ induced linear scratch-­type abrasions in the vicinity of the damaged skin areas (Figure 7.5.6g). Insects producing corrosive secretions (e.g., ants) may cause postmortem skin lesions that, when located upon the neck, may look like cutaneous marks due to manual strangulation (Figure 7.5.6h). Cockroaches produce superficial dermal abrasions that may mimic skin diseases. Injuries of cockroach ori-

gin are typically observed on body regions formerly protected by tunnel-­like layers of clothing such as the arms and legs (Figure 7.5.7a).

7.5.7  Pseudo-­ligature marks In the obese deceased, wrinkling of the skin of the neck may produce contact blanching of livor mortis (Figure 7.5.7b) that

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7.5.12 Injuries due to resuscitation procedures Resuscitation attempts require invasive iatrogenic manipulations on the patient such as external cardiopulmonary resuscitation (CPR), external defibrillation, endotracheal intubation and cannulation of peripheral and central vessels for the application of drugs. These resuscitation procedures may leave a broad pattern of antemortem, perimortem and postmortem injuries upon a deceased’s body (Buschmann and Tsokos 2009). Frequent consequences of resuscitation attempts are retropharyngeal bleedings, tooth damage and lesions of the mucosa of the larynx and trachea caused by intubation attempts. Fractures of the ribs and the sternum are the most frequent injuries seen after unsuccessful resuscitation caused by CPR. These injuries must be distinguished by the forensic pathologist from other causes of chest wall trauma; for example, attack or accident. Resuscitation-­related injuries of the thorax wall are only rarely observed in children because the infantile chest is of higher elasticity than that of the elderly. Bony chest injuries can be connected with superficial skin lesions caused by external CPR in the sternal area (Figure 7.5.7f). Another frequent finding following unsuccessful resuscitation attempts are dried-­up cutaneous defibrillation marks upon the chest (Figure  7.5.7g). If ribs are fractured during resuscitation measures (Figure 7.5.8a), this can cause perforation of the heart (Figure  7.5.8b) with subsequent pericardial tamponade seen at autopsy (Figure 7.5.8c). In addition, cases of aortic rupture from CPR, particularly located in the pars descendens, have been described (Buschmann and Tsokos 2009). As might be expected, visceral structures are less often affected from resuscitation-­ related trauma than thoracic structures because mechanical resuscitation attempts focus primarily on the heart and chest. However, upper belly organs such as liver

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Vibices are postmortem haemorrhages that are dot-­shaped, occasionally confluent and of bluish-­ blackish colour. They result from postmortem hypostasis leading to mechanical rupture of subcutaneous smaller vessels in areas of postmortem lividity due to pooling of erythrocytes in these vascular compartments under the influence of gravity during formation of livor mortis. Vibices are most exclusively found in obese corpses. In sparse livor mortis formation, for example, in the very old or after considerable loss of blood prior to death, vibices are usually absent. Vibices may be mistaken for true haemorrhages or haematomas, respectively, or, on the thorax, may mimic petechiae due to increased intrathoracic pressure. However, the presence of vibices is always solely restricted to areas of postmortem lividity (Figure 7.5.7c), which allows their correct interpretation as such.

Running out of gastric juice and bile during the handling or transportation of the body (Figure 7.5.7e) may produce postmortem skin alterations occasionally resembling those caused by chemical agents containing corrosives. Leaking out of urine post mortem may cause extensive skin damage to an infant upon the perigenital skin areas. The same is the case when the infant’s skin was in contact with a urine-­soaked diaper post mortem. One has to be careful to differentiate these postmortem skin changes from vitally acquired alterations and not to interpret them uncritically as signs of neglect prior to death (Evans 2001). Histologically, no vital reaction will be detected in such skin alterations.

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7.5.11 Skin changes produced by postmortem regurgitation of gastric juice and postmortem urine leaking

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Abrasions and lacerations upon the skin may be produced by manipulation of the body during postmortem handling, transportation or storage (Rutty 2001). These postmortem injuries are relatively easily distinguishable from vital injuries by their light yellowish-­brownish to golden, shiny appearance (Figure 7.5.7d). These postmortem skin injuries result from loss of the barrier function of the epidermal layer of the skin with subsequent evaporation of tissue fluid. In doubtful cases, incision of cutaneous injuries of postmortem origin will reveal a hardened, slightly flattened area on cut sections without any haemorrhage in the underlying soft tissue.

7.5.10 Postmortem injuries due to embalming Incisions made to aspirate body fluids and to instil embalming fluid are typical findings on the outside of the body following embalming procedures. The metal trocar through which the fluid is injected leaves puncture marks, depending in their size and shape on the diameter and type of trocar, in the inner organs such as the heart, liver, stomach and bowel.

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Figure 7.5.8  (a) Rib fractures following unsuccessful resuscitation measures. (b) Rupture of the left atrium of the heart following unsuccessful resuscitation measures. (c) Pericardial tamponade. (d) Liver ruptures following forceful resuscitation procedures.

Figure 7.5.9  Gastric air insufflation due to incorrect intubation.

(Figure 7.5.8d), stomach and spleen can be injured (Buschmann and Tsokos 2009). Incorrect intubation of the trachea with the endotracheal tube inserted into the oesophagus may lead to air insufflation into the stomach with subsequent gastric overstretching (Figure 7.5.9), in the worst case followed by gastric rupture.

References

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Estimation of the Time Since Death in the Early Postmortem Period, 2nd edn, pp. 215–225., London: Edward Arnold. Madea, B. (2002). Muscle and tissue changes after death. In: C. Henßge, B. Knight, T. Krompecher, B. Madea, and L. Nokes (eds.), The Estimation of the Time Since Death in the Early Postmortem Period, pp. 134–208. London: Edward Arnold. Madea, B. (2002). Estimation of duration of immersion. Nordisk Rettsmedisin 8: 4–10. Madea, B. (2005). Is there recent progress in the estimation of the postmortem interval by means of thanatochemistry? Forensic Science International 151: 139–149. Madea, B. (2006). Die Ärztliche Leichenschau 2 Auflage. Berlin, Heidelberg, New York: Springer. Madea, B. (2007). Praxis Rechtsmedizin: Befunderhebung, Rekonstruktion und Begutachtung, 2 Auflage. Berlin, Heidelberg, New York: Springer. Madea, B. (2007). Praxis Rechtsmedizin. Befunderhebung, Rekonstruktion, Begutachtung, 2 Auflage. Berlin, Heidelberg, New York: Springer. Madea, B. (2009). Death: time of. In: A. Jamieson and A. Moenssens (eds.), Wiley Encyclopaedia of Forensic Sciences, Vol. 2, 697–716. Chichester: John Wiley & Sons Ltd. Madea, B. (2009). Medical malpractice. In: A. Jamieson and A. Moenssens (eds.), Wiley Encyclopaedia of Forensic Sciences, Vol. 4, 1689–1709. Chichester: John Wiley & Sons Ltd. Madea, B. (2009). Time of death determination. In: A. Jamieson and A. Moenssens (eds.), Wiley Encyclopaedia of Forensic Sciences, Vol. 5, 2466–2479. Chichester: John Wiley & Sons Ltd. Madea, B. (2014). Handbook of Forensic Medicine. Chichester: Wiley. Madea, B. (2015). Rechtsmedizin. Befunderhebung, Rekonstruktion, Begutachtung. 3rd edn. Berlin: Springer. Madea, B. (ed.) (2015). Estimation of the Time Since Death, 3rd edn. Boca Raton, FL: CRC Press, Taylor & Francis. Madea, B. (2015). Preface. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn. Boca Raton, FL: CRC Press, Taylor & Francis. Madea, B. (2015). Supravitality in tissues. Definitions. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 17–24. Boca Raton, FL: CRC Press, Taylor & Francis. Madea, B. (2015). Postmortem mechanical excitation of skeletal muscle. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd ed., pp. 24–25. Boca Raton, FL: CRC Press, Taylor & Francis. Madea, B. (2015). Postmortem electrical excitability of skeletal muscle in casework. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 25–31. Boca Raton, London, New York: CRC Press, Taylor & Francis. Madea, B. (2015). Longitudinal studies of electrical excitability. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 31–46. Boca Raton, FL: CRC Press, Taylor & Francis. Madea, B. (2015). Postmortem lividity. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 59–62. Boca Raton, FL: CRC Press, Taylor & Francis. Madea, B. (2015). Cerebrospinal fluid chemistry. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 186–188. Boca Raton, FL: CRC Press, Taylor & Francis. Madea, B. (2015). Immunohistochemical methods as an aid in estimating the time since death In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 223–226. Boca Raton, FL: CRC Press, Taylor & Francis.

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whole animals using 1H-­ MR spectroscopy  – preliminary results. International Journal of Legal Medicine 125: 741–744. Mußhoff, F. and Madea, B. (2015). H3  – Magnetic-­ Resonance-­ Spectroscopy. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 203–212. Boca Raton, FL: CRC Press, Taylor & Francis. Naeve, W. (1978). Gerichtliche Medizin für Polizeibeamte. Heidelberg: Kriminalstik Verlag. Ondrizola, A., Riancho, J.A., de la Vega, R. et al. (2013). miRNA analysis in vitreous humor to determine the time of death: a proof-­of-­concept pilot study. International Journal of Legal Medicine 127 (3): 573–578. Ortmann, J., Markwert, P., and Madea, B. (2016). Precision of estimating the time since death by vitreous potassium – Comparison of 5 different equations. Forensic Science International 269: 1–7. Ortmann, J., Doberentz, E., and Madea, B. (2017). Immunohistochemical methods as an aid in estimating the time since death. Forensic Science International 273: 71–79. Padosch, S.A., Dettmeyer, R., Kröner, L. et al. (2005). An unusual occupational accident: fall into a sewage plant tank with a lethal outcome. In: Forensic Science International 149: 39–45. Patscheider, H. and Hartmann, H. (1993) Leitfaden der Gerichtsmedizin 3. Auflage, Bern, Göttingen, Toronto, Seattle: Verlag Hans Huber. Petersohn, F. (1972). Rechtsmedizinische Feststellungen an einer 3 Jahre in Plastik gehüllten Kindesleiche. In: Beiträge Gerichtliche Medizin 29: 351–358. Pieper, P. (2002). Moorleichen. In: H. Beck, D. Geuenich, and H. Steuer (eds.), Reallexikon der Germanischen Altertumskunde, Band 20, pp. 222–229. Berlin, New York: Walter De Gruyter. Pittner, S., Ehrenfellner, B., Zissler, A. et al. (2017). First application of a protein-­based approach for time since death estimation. International Journal of Legal Medicine 131: 479–483. Potente, S. (2015). Practical Casework. Conditional probability in death time estimation. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, 237–248. Boca Raton FL: CRC Press, Taylor & Francis. Pounder, J. (2000). Postmortem interval. In: J.A. Siegel, P.J. Saukko, and G.C. Knupfer (eds.), Encyclopaedia of Forensic Sciences, Vol. 3, 1167– 1172. San Diego: Academic Press. Preuß, J., Brünig, J., and Madea, B. (2004). Artifizielle Enterdigung durch Witterungseinflüsse. In: Rechtsmedizin 14: 14–19. Reh, H. (1967). Anhaltspunkte für die Bestimmung der Wasserliegezeit. Deutsche Zeitschrift für die gesamte gerichtliche Medizin 59: 235–245. Reh, H. (1969). Diagnostik des Ertrinkungstodes und Bestimmung der Wasserzeit. Düsseldorf: Triltsch. Reibe S., Doetinchem, P.V., and Madea, B. (2010). A new simulation-­ based model for calculating post-­mortem intervals using developmental data for Lucilia sericata (Dipt.: Calliphhoridae). Parasitology Research 107 (1): 9–16. Reibe, S. and Madea, B. (2010a). How promptly do blowflieds colonise fresh carcasses? A study comparing indoor with outdoor locations. Forensic Science International 195 (1): 52–57. Reibe, S. and Madea, B. (2010b). Use of Megaselia scalaris (Dipt.: Phoridae) for post-­mortem interval estimation indoors. Parasitology Research 106 (3): 637–640. Reibe, S. (2015). Forensic Entomolgy. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 249–257. Boca Raton, FL: CRC Press, Taylor & Francis Group.

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Madea, B. and Henßge, C. (2015). General remarks on estimation the time since death. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 1–7. Boca Raton, FL: CRC Press, Taylor & Francis. Madea, B. and Kernbach-­Wighton, G. (2015). Autolysis, putrefactive changes and postmortem chemistry. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 153–160. Boca Raton, FL: CRC Press, Taylor & Francis. Madea, B. and Henßge, C. (2015a). Eye changes after death. In: B Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 161–186. Boca Raton FL: CRC Press, Taylor & Francis. Madea, B and Henßge, C. (2015b). Cerebrospinal fluid chemistry. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 186–189. Boca Raton, FL: CRC Press, Taylor & Francis Group. Madea, B. and Knight, B. (2015). Gastric contents and time since death. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn. Boca Raton, FL: CRC Press, Taylor & Francis,. 213–222. Madea, B., Stockhausen, S., and Doberentz, E. (2016). Bestimmung der Wasserliegezeit nach Reh – weitere Untersuchungen zur Prüfung der Reliabilität. Archiv für Kriminologie 237: 1–2. Madea, B., Ortmann, J., and Doberentz, E. (2019). Estimation of the time since death – Even methods with a low precision may be helpful in forensic casework. Forensic Science International 302. doi: 10.1016/j. forsciint.2019.109879. Epub 2019 Jul 27. Mallach, H.J. (1964). Zur Frage der Todeszeitbestimmung. Berl Med 18: 577–582. Mallach, H.J. and Mittmeyer, H.J. (1971). Totenstarre und Totenflecke. Zeitschrift fur Rechtsmedizin 69: 70–78. Mathur, A. and Agrawal, Y.K. (2011). An overview of methods used for estimation of time since death. Australian Journal of Forensic Sciences 43(4): 275–285. Marshall, T.K. and Hoare, F.E. (1962). I Estimating the time of death. The rectal cooling after death and its mathematical expression. II The use of the cooling formula in the study of postmortem body cooling. III The use of the body temperature in estimating the time of death. Journal of Forest Science 7: 56–81, 189–210, 211–221. Mihailovic, Z., Atanasijevic, T., Popovic, V. et al. (2012). Estimation of the postmortem interval by analyzing potassium in the vitreous humor: could repetitive sampling enhance accuracy? American Journal of Forensic Medicine and Pathology 33: 400–403. Munoz, J.I., Suárez-­Penaranda, J.M., Otero, X.L. et al. (2001). A new perspective in the estimation of post-­mortem interval (PMI) based on vitreous. Journal of Forensic Science 46 (2): 209–214. Munoz Barús, J.I., Suárez-­Penaranda, J.M., Otero, X.L. et  al. (2002). Improved estimation of postmortem interval based on differential behavior of vitreous potassium and hypoxanthine in death by hanging. Forensic Science International 125: 67–74. Munoz-­Barús, J.I., Febero-­Bande, M., and Cadarso-­Suárez, C. (2008). Flexible regression models for estimating postmortem interval (PMI) in forensic medicine. Statistics in Medicine 27: 5026–5038. Munoz-­Barús, J.I., Rodriguez-­Calvo, M.S., Suárez-­Penaranda, J.M. et al. (2010). PMICALC: an R code-­based software for estimating post-­ mortem interval (PMI) compatible with Windows, Mac and Linux operating systems. Forensic Science International 194: 49–52. Musshoff, F., Klotzbach, H., Block, W. et al. (2011). Comparison of post-­ mortem metabolic changes in sheep brain tissue in isolated heads and

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Spindler, K., Wilfing, H., Rastbichler-­Zissernig, E. et al. (1996). A Global Survey for the Status and the Techniques of Conservation. Wien, New York: Springer. Spitz, W.U., Spitz, D.J., and Fischer, R.S. (2006) Spitz and Fisher’s Medicolegal Investigation of Death: Guidelines for the Application of Pathology to Crime Investigation. Springfield, Ill: Charles C. Thomas. Stephens, R.J. and Richards, R.G. (1987). Vitreous humour chemistry: the use of potassium concentration for the prediction of the postmortem interval. Journal of Forensic Science 32: 503–509. Sturner, W.Q. (1963). The vitreous humour: postmortem potassium changes. Lancet 1: 807–808. Sturner, W.Q. and Gantner, G.E. (1964). The postmortem interval: a study of potassium in the vitreous humor. American Journal of Clinical Pathology 42: 137–144. Tagliaro, F., Manetto, G., Cittadini, F. et al. (1999). Capillary zone electrophoresis of potassium in human vitreous humour: validation of a new method. Journal of Chromatography B 733: 273–279. Thierauf, A., Musshoff, F., and Madea, B. (200)). Post-­mortem biochemical investigations of vitreous humor. Forensic Science International 192: 78–82. Tröger, H.D., Baur, C., and Spann, K.W. (1987). Mageninhalt und Todeszeitbestimmung. Lübeck: Schmidt-­Römhild. Tsokos, M. (2005). Post mortem changes and artefacts during the early post mortem interval. In: M. Tsokos (ed.), Forensic Pathology Reviews, pp. 183–237. Totowa, New Jersey: Humana. Tumrana, N.K., Ambadea, V.N., and Dongreb, A.P. (2104).Thanatochemistry: study of vitreous humor potassium. Alexandria Journal of Medicine 50 (4): 365–368. Van den Oever, R. (1976). A review of the literature as to the present possibilities and limitations in estimating the time of death. Medicine, Science and the Law 16: 269–276. Van der Sanden, W. (1996). Mumien aus dem Moor. Die vor-­und frühgeschichtlichen Moorleichen aus Nordwesteuropa. Amsterdam: Batavian Lion International. Warther, S. (2013). Postmortale mechanische Erregbarkeit der Skelettmuskulatur (idiomuskulärer Wulst) zur Eingrenzung des Todeszeitpunkts. MD Thesis University Hamburg. Warther, S., Sehner, S., Raupach, T. et al. (2012). Estimation of the time since death: postmortem contraction of human skeletal muscles following mechanical stimulation (idiomuscular contraction). International Journal of Legal Medicine 126: 399–405. Wehner, F., Wehner, H.-­ D., Schieffer, M.C., and Subke, J. (1999). Delimination of the time since death by immunohistochemical detection of insulin in pancreatic ß-­cells, Forensic Science International 105: 161–169. Wehner, F., Wehner, H.-­D., Schieffer, M.C., and Subke, J. (2000). Delimination of the time since death by immunohistochemical detection of immunohistoch. Forensic Science International 110: 199–206. Wehner, F., Wehner, H.-­D., and Subke, J. (2001a). Delimination of the time since death by immunohistochemical detection of calcitonin. Forensic Science International 122: 89–94. Wehner, F., Wehner, H.-­D., and Subke, J. (2001b). Delimination of the time since death by immunohistochemical detection of glucagon in pancreatic α-­cells. Forensic Science International 124: 192–199.

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Rodriguez, W.C. and Bass, W.M. (1985). Decomposition of buried bodies and methods that may aid in their location. In: Journal of Forensic Sciences, 30 (30): 836–852. Rognum, T.O., Hauge, S., Oyasaeter, S., and Saugstad, O.D. (1991). A new biochemical method for estimation of post-­mortem time. Forensic Science International 51: 139–146. Rognum, T.O., Holmen, S., Musse, M.A. et al. (2016). Estimation of time since death by vitreous humor hypoxanthine, potassium, and ambient temperature. Forensic Science International 262: 160–165. Rosendahl, W. and Doppes, D. (2015). Rdiocarbon dating. Basic principles and applications. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 259–267. Boca Raton, FL: CRC Press. Rothschild, M.A., Schmidt, V., and Pedal, I. (1996). Leichenlipid: Unterschiedliche Entstehungsarten erschweren zusätzlich die Abschätzung der Leichenliegezeit. In: Archiv für Kriminologie, 197: 165–174. Rutty, G. (2015). The use of temperatures recorded from the external auditory canal for the estimation of the postmortem interval. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 134–151. Boca Raton, FL: CRC Press. Rutty, G. and Morgan B (2015). Cross-­sectional imaging and the post-­ mortem interval. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 269–276. Boca Raton, FL: CRC Press. Sampaio-­Silva, F., Magalhaes, T., Carvalho, F. et  al. (2013). Profilingo f RNA degradation for estimation of post mortem interval. PLOS ONE 8(2): e56507. Scheurer, E., Ith, M., Dietrich, R. et al. (2003). Statistical evaluation of 1 H-­MR spectra of the brain in situ for quantitative determination of postmortem intervals (PMI). Proceedings of the International Society for Magnetic Resonance in Medicine 11: 569. Scheurer, E., Ith, M., Dietrich, D. et al. (2005). Statistical evaluation of time-­dependent metabolite concentrations: Estimation of postmortem intervals based on in situ 1H MRS of the brain, NMR in Biomedicine, 18 (3): 163–172. Schleyer, F. (1958). Postmortale klinisch-­ chemische Diagnostik und Todeszeitbestimmung mit chemischen und physikalischen Methoden. Stuttfart, Germany, Thieme. Schleyer, F. (1963). Determination of the time since death in the early postmortem interval. In: F. Lundquist (ed.), Methods of Forensic Science, Vol. 2, pp. 253–293. New York: John Wiley & Sons Interscience Publishers. Schleyer, F. (1973). Wie zuverlässig ist die Kaliumbestimmung im Glaskörperinhalt als Mittel zur Todeszeitschätzung? Zeitschrift fur Rechtsmedizin 71: 281–288. Schoning, O. and Strafuss, A.C. (1980). Determining time of death of a dog by analysing blood, cerebrospinal fluid and vitreous humour collected postmortem. American Journal of Veterinary Research 41: 955–957. Siddamsetty, A.K., Verma, S.K., Kohli, A. et al. (2014). Estimation of time since death form electrolyte, glucose and calcium analysis of postmortem vitreous humour in semi-­arid climate. Medicine, Science and the Law 54: 158–166. Spindler, K. (1993). Der Mann im Eis: Die Ötztaler Mumie verrät die Geheimnisse der Steinzeit. München: C Bertelsmann. Spindler, K., Rastbichler-­Zissernig, E., Wilfing, H. et  al. (1995). Der Mann im Eis, Neue Funde und Ergebnisse. Wien, New York: Springer.

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Zhou, B., Zhang, L., Zhang, G. et al. (2007). The determination of potassium concentration in vitreous humor by low pressure ion chromatography and its application in the estimation of postmortem interval. Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences 852: 278–281. Zilg, B (2015). Postmortem analyses of vitreous fluid. MD Thesis Department of Oncology-­Pathology. Karolinska Institutet, Stockholm Sweden. Zilg, B., Bernard, S., Alkass, K. et al. (2015). A new model for the estimation of time of death from vitreous potassium levels corrected for age and temperature. Forensic Science International 254: 158–166. Zilg, B., Alkass, K., Berg, S., and Druid, H (2016). Interpretation of postmortem vitreous concentrations of sodium and chloride. Forensic Science International 263: 107–113. Zubakov, D., Kokshoorn, M., Kloosterman, A., and Kayser, M. (2009). New markers for old stains: stable mRNA markers for blood and saliva identification from up to 16-­years-­old stainds. International Journal of Legal Medicine 123 (1): 71–74.

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Wehner, F. (2009). Die Eingrenzung der Leichenliegezeit im spätpostmortalen Intervall  – Neue Ansätze mittels immunhistochemischen Verfahren. Medizinische Welt 60: 402–406. Wehner, F., Wehner, H.D., and Claßen, I. (2002) Eingrenzung der Leichenliegezeit mittels immunhistochemischen Nachweises von Cystatin-­C. 81. Jahrestagung der Deutschen Gesellschaft für Rechtsmedizin, Rostock. Wehner, F., Wehner, H.D., and Hahnenberger, K. (2003) Delimitation of the time of death by immunhistochemical detection of Somatostatin. Third European Academy of Forensic Science Triennial Meeting, Istanbul, Turkey. Wieczorek, A. and Rosendahl, W. (eds.) (2007). Mumien – Der Traum vom ewigen Leben. Mainz: Philipp von Zabern. Wieczorek, A. and Rosendahl, W. (eds.) (2010). Mummies of the World. Munich, Berlin, London, New York: Prestel. Yang, M., Li, H., Yang, T. et al. (2018). A study on the estimation of postmortem interval based on environmental temperature and concentrations of substance in vitreous humor. Journal of Forensic Science 63 (3): 745–751.

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of Catholic theology (e.g., the Reformation period) and severe pandemic issues (e.g., pestilence) and continued to grow in the second half of the 19th century. As the traditional burning of the corpse in the open flames of a funeral pyre was increasingly considered as being impious, the first incinerator in Europe was built in 1876 in Milan, Italy, and the first crematorium in Germany was set up in 1878 in Gotha, Thuringia (Holck 2005). At first, architects tried to avoid chimneys for the cremation buildings, so that the optical appearance of the crematoria as ‘factories’ would be avoided.

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The burning of corpses is one of the oldest known ways to dispose of human remains. Archaeological excavations indicate that the first known cremations took place as early as 3000 BC in Europe and the Middle East. In ancient times, the burning of corpses in the open flames of a funeral pyre was the most common method, except for the Egyptians who considered fire as an animal which they were not allowed to feed by sacrificing corpses to it. Also in ancient Greek and Rome, entombing was the most common way of cremation, which later changed due to the propagation of Christianity, because ‘man is made of earth and shall turn to earth’ (1 Moses 2:7 and 1 Moses 3:19), and earth interment was considered as an ideal way to the end of resurrection. Thus, during Christian medieval times, death by burning at the stake/cremation was reserved only for miscreants such as heretics and witches – not only to bring them to death by the ‘scavenging’ force of fire but also to prevent them from resurrection. For faithful Christians, cremation was strictly forbidden after death as ordered by Charles the Great in 785 AD. In 1886, cremation was prohibited again by the Catholic Church, and was to be punished with excommunication. Even in 1917, in the Codex Iuris Canonici, a Catholic burial was to be neglected after cremation. Catholic Christians are allowed to be cremated from 8 May 1963 by the Catholic Church, and, until now, Protestant Christians prefer cremation when compared to faithful Catholics (Matschke and Tsokos 2000). Nevertheless, renaissance of ancient cremation ideas started as early as in medieval times due to slight but expanding negligence

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Handbook of Forensic Medicine, Volume 1, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

8.2  Cremation in the modern day Cremation rates in Europe have constantly increased since the beginning of the 20th century due to changes of sociocultural and religious attitudes; it must not remain unmentioned in this context that crematoria were also integral components of the concentration camps of German National Socialism. Nowadays, there are more than 1000 crematories in Europe, and in 2006, the total number of cremations in Europe was more than 1,500,000. Cremation is a trend that is expected to continue and even grow in the near future. Currently, the cremation frequency throughout the Western world reaches approximately 85% of all burials in larger cities, since the commercial cremation process requires fewer goods and services and virtually no associated long-­term maintenance. For surviving kin who bear the financial burden of the funeral, the incineration of the relative might be the most cost-­effective and sometimes the only choice.

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Modern crematoria are highly specified engineered buildings, and incineration of a body occurs under controlled conditions where the body is subjected to a continuous source of heat. Although technical specifications may differ from crematory to crematory, the incinerator usually consists of two chambers heated by gas, electricity or oil, and is built of high-­grade steel plate, lined inside with heavy refractory tile or brick. The primary incinerator operates first at preheated temperatures between 650 and 700 °C before the casket with the body inside is transported into the furnace via conveyor to prevent excessive heat loss. From the moment the casket is placed inside the primary incinerator, an exothermal reaction occurs with a sharp increase of the stove temperature, and the subsequent burning of the casket and the body will initially increase a surplus of heat: the casket is ignited by the stored heat of the refractory lining of the furnace and the heat generated by the combustion. Furthermore, the burning of the shroud and the casket brings the amount of energy needed to keep the combustion going. The fire is supported solely by supplying hot air during this phase of the incineration which lasts about 40–50  minutes (Figure 8.1a,b). It ends with the blowing of the wood ash. Then gas burners are engaged, and the stove temperature increases up to 1200 °C in order to incinerate not only the remaining

ingredients but also smoke and odour that emerge during the cremation process. Then the remains are automatically transferred to the second incinerator (‘after chamber’ or ‘afterburner chamber’) to cool down. The whole cremation process takes about 90–120 minutes, depending on the given temperature, body mass, water content of the body, sex, age and other factors, including the presence of obesity or wasting diseases at death (Mayne Correia 1997; DiMaio and DiMaio 2001; Prahlow 2010). A study on the changes of the body occurring during cremation found that at temperatures between 670 and 810 °C, after 20 minutes, the calvaria was defleshed and fissures of the tabula externa were noticed. The body cavities burst open after approximately 30  minutes, the internal organs were severely shrunken after 40 minutes and the extremities were destroyed after 50 minutes. The shrunken and destroyed torso broke apart after 60–90 minutes (Bohnert et al. 1998). Organs and soft tissues are completely burned to fume and around 5% of the body weight remains as ‘ashes’ consisting of small or sometimes even larger calcined bone pieces, implanted structures and casket components. The unambiguous and individual assignment of the remains is ensured by a fireproof brick (ceramic plate mark) with a number, placed before cremation on the coffin or the body, as well as the fact that the ovens in a crematorium are not designed to burn more than one coffin at the same time. From the remaining ‘ashes’, iron parts such as brackets of the coffin or medical implants (staples, nails, screws, embolism filters, etc.) are sorted out magnetically, and gold and titanium implants are rejected

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Figure 8.1  (a) and (b) Contemporary cremation of a body in different stages in the primary incinerator. Source: Courtesy of Patrik Budenz.

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­ rocessed ‘cremained’ human ‘ashes’ may potentially exhibit a p composition comparable to human bones when analysed by particle-­induced X-­ray. Also, examination of the ‘cremains’ by means of thermogravimetry or electron microscopy can be successful. Incinerated teeth change morphologically, as the enamel and coronal dentin quickly dehydrates, the tooth structure is splintered off and the residual root structure appears chalky and extremely brittle. Nevertheless, postincineration radiographs indicate that the teeth maintain their radicular structure and the obturation materials contained within the teeth are not pulverised. In cases of cremated and subsequently processed teeth with restorative resins, usage of X-­ray fluorescence analysis as a tool in forensic odontology may also allow identification, emphasising the significance of the role of restorative resins for identification purposes, even after the extreme circumstances of incineration (Bush et al. 2007; Bonavilla et al. 2008; Schultz et al. 2008). Altogether, the analysis of cremated human remains is difficult, not always promising and requires an interdisciplinary approach of at least forensic scientists, anthropologists and forensic odontologists.

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8.5 Conclusions The cremation of a human body is a highly reductive taphonomic process that represents one of the most extreme examples of postmortem alteration. The thorough reduction and fragmentation of cremated human remains often leaves only little biological evidence of diagnostic value and among the three most common ways to entomb human remains – earth interment, burial at sea and cremation (incineration). Cremation poses the most difficulties to forensic practitioners because the body itself as evidence is completely lost. This fact led to the statutory practice of a second external examination of corpses before cremation in Germany. For example, as early as 1934, German law provided that a second external examination before cremation is mandatory to ensure that evidence specifically for an uncertain or non-­natural manner of death can be examined by the investigating authorities. This procedure is still scheduled in German Federal Law in 15 federal states (except for the state of Bavaria), leading to detection of unnatural causes of death (including manslaughter and homicide) in approximately 1% of cases by external examination before cremation. This ‘second look’ is carried out by forensic pathologists in the crematorium (Figure 8.2). After scrutinising the death certification, a systematic external examination of the body is conducted. Medicolegal diagnosis from the external examination must not contradict the cause and manner of death given on the death certification. If a dissent is found, cremation is not performed, and the investigation authorities will be made aware of the diagnosis found at the external examination and the circumstances of the case by the forensic practitioner. Usually, the public prosecutor then orders a medicolegal autopsy to clarify cause, manner and circumstances of death.

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before the subsequent crushing of remaining small bone pieces in a ballot-­box filling machine. Processing of the cremated bone pieces by mechanical or manual blunt force is also possible and may enable the correlation between the processing method used and the traces found on the cremated and pulverised bone pieces. The fine ‘ashes’ (actually burned and pulverised bone fragments of several millimetres) and the ceramic plate mark are filled into a cinerary urn which is then sealed. Outside on the cinerary urn, the name of the crematory, the name of the deceased, his or her birth, death and cremation dates and identification number are embossed. During the cremation process, the pollutants emitted by the combustion of organic matter with the presence of other trace elements are combustion gases NOx, CO, SO2, and particulate matter (solid particles suspended in a gas as a by-­product of the combustion processes), heavy metals and polychlorinated dibenzo-­p-­dioxins and dibenzofurans. Compared to emissions from other emission sources such as factories or waste incinerators (municipal, hazardous and medical), emissions from crematories are significantly lower and generally non-­threatening to the environment, but cremation-­related emissions of mercury should not be underestimated: mercury is unstable at cremation temperatures and free mercury metal is highly volatile. The average human body contains approximately 6  mg of mercury due to incorporation during lifetime. As a further source of mercury, teeth fillings with dental amalgam usually contain more than 0.5 g of mercury. Studies have found that as much as 200 μg/m3 of mercury is emitted during the cremation process of one human body with dental amalgam fillings: this reaches the accepted level of mercury emission (Mari and Domingo 2010). Another potential environmental limitation exists for corpses that are contaminated with radioactive substances. These bodies should not be cremated, as radioactive nuclides can spread after incineration along with the fumes. Although waste gases from the muffle are transported via the firing chamber and the recuperator and are subsequently purified by chemical or electric filters, radioactive nuclides can pass these filters. Furthermore, radioactive ‘ashes’ pose several problems of disposal (Aggrawal 2005).

8.4  Remnants of cremation As explained, remnants of cremation are only ‘ashes’ (actually burned and pulverised bone fragments of several millimetres); various implanted structures, such as dental restoration or orthopaedic hardware that are not burned but removed before further processing of the ashes, may allow identification after the cremation process (Baldwin and May 2005). While current cremation methods leave only crushed ‘cremains’ without any visible bone or teeth structures, older cremation procedures which operate at lower temperatures may leave fragments of calcined bones or teeth which can then be subject to forensic anthropological examination. Nevertheless, also currently cremated and

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Figure 8.2  Examination room at a crematory; findings at external examination before cremation must apply to the death certification.

developed techniques for the preservation of corpses and thus belongs to the earliest known cultures that embalmed their deceased. Starting from natural mummification due to the arid climate, techniques of artificial mummification developed, which included deconstruction of the body with removal of the internal organs and musculature and subsequent reassembly of the skeleton and filling of the body cavities with grass and ashes (Arriaza 1995; Aufderheide 2003; Tellenbach V and Tellenbach M 2007; Marquet et al. 2012). The Egyptian mummies might be of greater fame. In the early phases of the Egyptian advanced civilisation, similar to the Andean peoples, natural conditions with great aridity led to the formation of natural mummies (which may have been the reason why the conservation of the body as a central element was only integrated into religious systems in the first place), and from there on a constantly changing practice of elaborate artificial mummification developed. The implementation was the task of trained specialists, mostly priests, and the entire procedure, including drying phases, should ideally last about 70  days (Aufderheide  2003; Pommerening  2007; Aufderheide  2009; Mayer 2012). An essential element was the removal of internal organs, usually via a left flank cut. Lung, liver, stomach and intestine were separately embalmed in special vessels, the canopic jars. The heart, as the seat of the soul, was considered too elementary to be removed from the body, while the brain was disposed of. The removal of the brain was usually done by driving a special hook through the nose, breaking through the lamina cribrosa (Aufderheide 2003; Pommerening 2007; Robinson 2012). The retroperitoneal organs often, but not always, remained in

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8.6 Embalming

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The desire to preserve the human body after death and to protect it from decay is old, the motives for this vary. In ancient advanced civilisations, the best-­known example being Ancient Egypt, it was mainly religious motives that led people to look for ways to preserve corpses. Until modern times, the preserved body of an influential personality may also serve political purposes, as the example of Lenin shows. The sometimes enormous collections of anatomical and pathological specimens, some of which are still preserved, bear witness to the desire to preserve findings of scientific interest, partly out of honest scientific motivation, partly certainly also out of sensationalism. Nowadays, embalmings are carried out in the course of funeral preparations, on the one hand, to give relatives the opportunity to say farewell unaffected by changes to the corpse. On the other hand, preserved bodies are also an essential part of medical education and training. This may be within the framework of anatomy courses for students or special operation courses for doctors. A successful embalming can make forensic examinations of a corpse considerably more difficult and present forensic experts with special challenges.

8.6.1  Short history of embalming Naturally, only a brief, incomplete insight into the long history of embalming in the various regions of the world can be provided here. Already in the period 5000–6000 BC, the Chinchorro culture in the area of today’s Northern Chile and Southern Peru

the Roman and Greek world (Brenner 2014). A rare example is known from Spain, where approximately 5000-­year-­old bones covered with large amounts of powdered vermillion delivered over long distances were discovered (Martin-­Gil et al. 1995). In the Middle Ages, the need for preservation of a corpse arose particularly in the case of deceased dignitaries who had died (or fallen) far from the desired burial place and had to be transported over long distances. For this purpose, sometimes drastic measures were taken which, in addition to organ removal, could include dissection of the corpse and maceration of the bones (Wunn  2007; Mayer  2012). Public ceremonies were sometimes held for important personalities and rulers, which also required the preservation of the body (Wunn  2007; Trompette and Lemonnier 2009). During the Renaissance, medical progress led to a growing interest in more subtle embalming techniques to preserve bodies for scientific purposes (Trompette and Lemonnier 2009; Brenner  2014; Mayer  2012). Instructions for embalming have been handed down from the 16th century by the Dutchman Petrus Forestus (1522–1597) and the Frenchman Ambroise Paré (1510–1590). Both of them performed eviscerations and used herbal mixtures (including myrrh, wormwood and marjoram). Paré also describes the rubbing of the corpse with turpentine and fragrant oils. After William Harvey (1578–1657) described the blood circulation in 1628, it was used by the Dutchmen Jan Swammerdam (1637–1680), Frederik Ruysch (1638–1731) and Stephen Blanchard (1650–1720) for embalming corpses by means of vessel injections. In the following centuries, the techniques were refined, whereby regionally different characteristics can be observed. An overview can be found with Mayer (Mayer 2012). Here, only a few other milestones of embalming will be mentioned: the Italian Giuseppe Tranchina (1797–1837) carried out the first documented embalming in modern times, in which no evisceration was carried out and only a vascular injection (with a solution containing arsenic) was used. The Frenchman Jean-­Nicolas Gannal (1721–1783) should also be mentioned, who conducted and published the first scientific studies on embalming and is sometimes considered the father of modern embalming, but was opposed by representatives of the medical science (Trompette and Lemonnier 2009; Mayer 2012). Alfredo Salafia (1869–1933), who also embalmed the body of the two-­ year-­ old girl Rosalia Lombardo and became famous for his exhibition in the Catacombs of Palermo, was probably one of the first to use formaldehyde-­containing solutions (Piombino-­Mascali et al. 2009). The breakthrough of modern embalming for the purpose of reverent and hygienic farewells and burials is attributed to the American Civil War. In the USA, an embalming culture or a specialty known as thanatopractice for funeral purposes developed with the founding of its own institutes and schools beyond the medical sciences (Mayer 2012). A detailed analysis of the conflict between medical science and commercial use of embalming can be found in Trompette and Lemonnier (2009).

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the corpse. Whether this was due to a special purpose or practical considerations is not clear (Aufderheide  2003, Brier and Wade  2001). The removal of organs through the described accesses is anything but trivial in its practical implementation, as attempts at reconstruction have demonstrated (Brier and Wade  2001). The body was then dried. For this purpose, the cleaned body cavities were tamponed with natron-­soaked bundles and the body was stored on and covered with natron. After the drying phase and cleaning with natron, the body was rubbed with resins (possibly mixed with beeswax) and the body cavities were tamponed with resin-­soaked cloths (sometimes also with sawdust). Finally, the body was wrapped in resin-­ soaked linen (Aufderheide  2003; Aufderheide  2009). This procedure would be exemplary for the time of the New Kingdom and can by no means be considered a standard for all Egyptian mummies of all epochs. For example, the composition of the substance mixtures used for embalming was also subject to changes, as chemical analyses showed (Buckley and Evershed 2001). Bodies have also been embalmed in other regions of the world, such as in South and Central America, Oceania, Australia or by the Guanches on the Canary Islands (Aufderheide  2003; Mayer  2012). Exemplary here are the mummies from China, which are not so frequent in numbers compared to Egypt, but are nevertheless impressive: the necessity of preserving the deceased arose in particular from the very long time between death and burial (sometimes years in the case of emperors). In contrast to Ancient Egypt, no intensive manipulation of the corpse and in particular no removal of organs or opening of body cavities took place. The corpse was usually cleaned, bathed in baths containing alcohol, temporarily stored on ice (depending on the wealth of the deceased) and wrapped in numerous layers of cloth. The equipment of the burial chambers, the lining of the chambers with mercury and the use of bactericidal incense and thick-­walled, coated wooden coffins supported the preservation as well as regionally specific climatic conditions and soil conditions. The best-­known example is probably the Lady of Dai, a princess, who died around 170 BC. Her body not only contained all her organs, but also had movable limbs and a soft consistency. At the time of discovery, the corpse was found in a brownish liquid whose role has not yet been conclusively clarified (Werning 2007). Alexander the Great is a famous example of temporary preservation of corpses in ancient times. His body is said to have been preserved in honey after his death in Babylon for transport to Alexandria (Brenner 2014). As shown, beeswax was also used in Egypt for artificial mummification. The preservative properties of the highly sugary and thus dehydrating honey have been experimentally confirmed in recent times (Sharquie and Najim  2004), but Aufderheide (Aufderheide  2003) conclusively states that the historical sources are not very meaningful in the case of Alexander and that the possibility of preserving whole bodies in honey has not yet been proven. In prehistoric and ancient Europe, however, embalming was less common overall, although there were exceptions in

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Embalming techniques have a great importance in medical science. On the one hand, teaching on fixed cadavers is an essential part of the anatomical education of medical students. On the other hand, conserved body donations are also used in further medical education; for example, in training programmes for surgeons. In addition to the prevention of tissue decay and the necessary hygienic protection, the tissue structure plays an important role in this context in order to convey a haptically as “real” feeling as possible. Furthermore, the agents used should of course have as little impact on the health of those working with the body as possible. Since the 19th century, there have been countless attempts to develop the most suitable agents for this purpose, which meet the abovementioned requirements in their combination. Most of them are based on formaldehyde, eventually in combination with other substances. A detailed overview of the development can be found with Brenner (Brenner 2014). Formaldehyde (CH2O) is a gas from the substance group of aldehydes which dissolves well in water and forms methylene hydrates. The liquid known and sold as formalin contains 35–40% formaldehyde, whereby formaldehyde is present in the form of small polymers. Larger polymers form a whitish powder: paraformaldehyde. Formalin contains about 10% methanol to prevent the precipitation of paraformaldehyde. Formaldehyde forms methylene bridges between adjacent proteins, preferably between nitrogen atoms, and thus leads to tissue fixation (Kiernan 2000). Formaldehyde solutions are effective antiseptics and reliably prevent bacterial decay. They fix tissue well, but for practical exercises, they have the disadvantage of hardening the tissue and discolouration. Various approaches have been developed to achieve a haptic and visual impression that comes close to living tissue. One of the most widespread is probably the Thiel fixation method. Here, solution mixtures are used that contain, in addition to formalin, ammonium nitrate, potassium nitrate, boric acid and denatured ethyl alcohol, among others. Different solutions are used for different purposes (perfusion of the vascular system, hollow organs, brain, storage of the body, etc.). This method keeps the natural colour of tissues and a softer consistency (Thiel 1992; Thiel 2002). For didactic and scientific purposes, the production of durable preparations also plays an important role. Since these are of rather

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With regard to the frequency of embalming in the context of burials and farewells, there are great regional differences. While in the USA, for example, thanatopractice is of great importance and exists as an independent discipline, embalming is not very common in Germany, while in neighbouring France, it has increased significantly since the second half of the 20th century (Trompette and Lemonnier 2009). Apart from cosmetic reasons (whereby the measures for the aesthetic preparation of the corpse that fall within the scope of thanatopractice far exceed pure embalming), hygienic considerations also play a role, especially with regard to infection protection. Before the actual embalming, a thorough cleaning and disinfection of the body surfaces as well as first cosmetic measures (e.g., shave) take place. The mouth is usually closed with special suture techniques, the eyes are closed (if necessary, with the help of eye caps or cotton) and body orifices are closed with cotton cloths soaked in, for example, phenol. Special techniques are required to deal with injuries, decay, the consequences of medical measures or even autopsies. Details can be found in textbooks on thanatopractice (e.g., Mayer 2012). The embalming liquids used are often complex mixtures whose exact composition varies. They usually contain preservatives (formaldehyde and other aldehydes such as glutaraldehydes, alcohols, phenols or salts) and germicides (often ammonium compounds). Depending on the area of application, modifying substances such as buffers or anticoagulants, and also colourants or fragrances can be added. The embalming fluid dissolved in water is usually injected into the arterial leg of the vascular system (arterial embalming; Figure 8.3) during or after the blood has been drained via the venous leg. Since perfusion cannot, by its very nature, be achieved evenly in all parts of the body and organs, putrefaction changes occur at different rates. Manual techniques and storage are used to counteract this. The embalming solution is often injected at different parts of the body. The choice of the artery depends on a number of variables, not least on the experience and preferences of the

person performing the procedure. Attempts are made to counteract the occurrence of discolouration or oedema. The incision sites are usually treated with an absorbent solution and skin incisions are closed with tissue adhesive. In a further step (cavity embalming), body cavities and hollow organs are punctured using a trocar, liquids and gases are aspirated and preservative solution is introduced. Sometimes the head cavity is also infiltrated transnasally with a trocar. In addition, preservative solution can be applied subdermally (hypodermic embalming) or superficially (surface embalming) (Hanzlick 1994; Mayer 2012).

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Figure 8.3  Arterial embalming in progress.

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Specific difficulties arise in the dissection of embalmed bodies. Here, reference is made to forensic sections of embalmed bodies – the sectioning of mummies or other historical finds carried out for scientific reasons naturally requires a specific procedure. The fact that the possible absence of organs restricts the findings (e.g., in the case of dissected corpses transferred from abroad) does not require further explanation. However, even pure embalming without organ or tissue removal can cause difficulties in the examination. The fixation solution can, as in the case of formalin, cause irritation to the eyes, respiratory tract and skin of

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the examiners and may even be carcinogenic, thus posing a long-­ term health risk. Since the fixation changes the haptic impression decisively, this must be taken into account when performing the dissection in order to avoid accidental injuries in case of unaccustomed tissue properties. The haptic impression (e.g., the consistency of the liver) is not reliable. Calcifications as in arteriosclerosis can of course still be detected. The colouration of body surfaces and organs changes, which can make it difficult to visually identify haematomas or pathological changes in organs (Figure  8.4). Formaldehyde and blood, for example, take on a grey colour when mixed (Hanzlick 1994). The cleaning of the corpse destroys not only possible traces on the surface of the body, but also under the fingernails, which are cut. Injuries are often tried to make unrecognisable in the course of funeral preparation. Skin cuts are closed, haematomas are cosmetically changed. Shaving the face can cause small injuries that can be misinterpreted. The injury pattern presented to the examiner on a corpse that has undergone thanatopractical treatment is therefore very different from the original findings. On the other hand, injuries can also happen that can lead to misinterpretations: the skin incisions made for the application of the embalming solution should be easily distinguished from injuries that occurred during lifetime by their typical localisation, closure and lack of vital signs. Injection of the solution by means of a trocar into the trunk cavity and the hollow organs causes organ injuries; in case of transnasal access to the head cavity, ethmoid breakthroughs occur. Embalming solution injected into the neck vessels with appropriate pressure can lead to bleeding into the soft tissues in the neck area. Pre-­existing oedema or gas accumulations in the facial area may be relieved by the mortician during preparation for the funeral and then evade detection during dissection, but the procedure can lead to blood leaking into the

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little relevance for forensic purposes, we will only briefly discuss this here. In addition to the use of paraffin and polyethylene glycol (PEG), the plastination process developed by Gunther von Hagens in Heidelberg in the late 1970s deserves special mention. In a rough outline, the process, which is established in various variants, consists of a sequence of different working steps. After fixing the body, for example, with formalin, and subsequent ­preparation according to the purpose of the image, the body is dehydrated in a deep-­frozen acetone bath and then degreased. The essential step is the replacement of the intermediate medium (e.g., acetone) by the selected fixation substance (e.g., silicone S10) in a vacuum. In the case of silicone, curing would be ­performed by gassing (von Hagens 1979; von Hagens et al. 1987). The process, which has been varied and refined in recent decades, yields impressive plastinates which, in addition to their use,  ­ especially in medical education, have also reached an ­audience of millions in the context of large, although not uncontroversial, exhibitions.

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(a) Figure 8.4  (a) Embalmed heart in situ and (b) un-­embalmed heart in situ in comparison.

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Aufderheide  2003). After leaving arsenic, it is still true that components of the embalming fluid can simulate poisoning, for example, methanol intoxication, if they are not known (Powers et  al. 2019). When carrying out toxicological tests, it must always be taken into account that the blood is largely replaced or at least strongly diluted. Other body fluids (e.g., urine) are usually removed. A supplementary analysis of the embalming fluid is recommended, since washed out analytes can sometimes be detected here. Formaldehyde is also highly reactive and can influence analytes via methylation, for example. This should always be taken into account when interpreting the results. The stability of different substances varies greatly and also depends on various variables such as the time since embalming, the composition of the embalming fluid, the change in pH value, etc. The extraction of the analytes can cause difficulties and an influence on the analytical methods or reagents should also be considered (Skopp 2004; Skopp 2010; Nikolaou et al. 2013; Takayasu 2013; Rohrig 2019). DNA analysis from embalmed tissue is also problematic, as formaldehyde (and possibly other substances used) damages the DNS. The choice of suitable material is important, and success has been achieved with bone marrow, nails and muscle tissue, for example (Wheeler et al. 2017). Possibilities of improving methods to optimise results are the subject of current research (Gielda and Rigg 2017).

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t­issue, which can be mistaken for vital haematomas. A similar situation can also be observed on the scrotum, for example, when hydrocells are mechanically corrected or embalming solution is applied via trocar. The injection of embalming solution can lead to the displacement of blood clots present premortem or formed after death into the right heart or pulmonary circulation, thus simulating a pulmonary artery embolism. Attempts to remove clots from the large veins can lead to injuries, including artificial pericardial tamponades. The artificially created pressure can lead to rupture of aneurysms. Aortic dissections caused by embalming have also been reported (Rivers  1978; Hanzlick  1994; Williams and Davison 2014; Guenevere et al. 2016). The authors have examined a case in which the potential difficulties occurred in an impressive accumulation. Three bodies and numerous body parts were brought to autopsy, which were confiscated by the authorities in a private institution that offered preparatory courses for medical studies, because the operator of the institution did not have permission for this and the documents required under German law were not available. The bodies, originating from the USA and inlaid in formalin, showed numerous and sometimes rough manipulations (severing of limbs, removal of individual organs, performed medical exercises, etc.). While some injuries were due to embalming (closed accesses to the vessels for arterial embalming), others were apparently the result of practical medical exercises, while in others the purpose was difficult to understand. In addition to the extensive manipulations, the consequences of improper storage with partial decay and disfiguring discolouration of the tissues also became apparent. This extremely limited the diagnostic findings (Figure 8.5). In one case, however, pulmonary artery embolism was found to be the cause of death (Windgassen et al. 2017). The problem of toxicological analyses of material from embalmed corpses already shows a historical example. Gannal was able to exculpate an accused female from the suspicion of poisoning her husband with arsenic by showing that the embalming solution used contained arsenic (Mayer  2012;

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References and further reading

Figure 8.5  Embalmed body after several manipulations such as organ removals and injection of colouring fluid.

Aggrawal, A. (2005). Terrorism: Nuclear and biological. In: J. Payne-­ James, R.W. Byard, T.S. Corey and C. Henderson (eds.), Encyclopedia of Forensic and Legal Medicine, Vol. IV, pp. 277–289. Amsterdam: Elsevier Academic Press. Arriaza, B.T. (1995). Chinchorro bioarcheology: Chronology and mummy seriation. Latin American Antiquity 6: 35–55. Aufderheide, A.C. (2003). The Scientific Study of Mummies. Cambridge: Cambridge University Press. Aufderheide, A.C. (2009). Reflections of bizarre mummification practices on mummies at Egypt’s Dakhleh oasis: A review. Anthropologischer Anzeiger 67: 385–390. Baldwin, H.B. and May, C.P. (2005). Exhumation. In: J. Payne-­James, R.W. Byard, T.S. Corey and C. Henderson (eds.), Encyclopedia of Forensic and Legal Medicine, Vol. II, pp. 281–284. Amsterdam: Elsevier Academic Press. Bohnert, M., Rost, T. and Pollak, S. (1998). The degree of destruction of human bodies in relation to the duration of the fire. Forensic Science International 95: 11–21. Bonavilla, J.D., Bush, M.A., Bush, P.J. and Pantera, E.A. (2008). Identification of incinerated root canal filling materials after exposure to high heat incineration. Journal of Forensic Sciences 53: 412–418. Brenner, E. (2014). Human body preservation – Old and new techniques. Journal of Anatomy 224: 316–344. Brier, B. and Wade, R.S. (2001). Surgical procedures during ancient Egyptian mummification. Chungara (Ariaca) 33: 117–123.

Prahlow, J. (2010). Burns and fire-­related deaths. In: J. Prahlow (ed.). Forensic Pathology for Police, Death Investigators, Attorneys, and Forensic Scientists, pp. 481–500. Totowa, NJ: Humana Press. Rivers, R.L. (1978). Embalming artifacts. Journal of Forensic Sciences 23: 531–535. Robinson, A. (2012). The art of medicine – Jean-­Francois Champollion and Ancient Egyptian Embalming. Lancet 379: 1782–1783. Rohrig, T.P. (2019). Postmortem Toxicology, pp. 123–139. London: Academic Press. Schultz, J.J., Warren, M.W. and Krigbaum, J.S. (2008). Analysis of human cremains: Gross and chemical methods. In: C.W. Schmidt and S.A. Symes (eds.), The Analysis of Burned Human Remains, pp. 75–94. Amsterdam: Elsevier Academic Press. Sharquie, K.E. and Najim, R.A. (2004). Embalming with honey. Saudi Medical Journal 25: 1755–1756. Skopp, G. (2004). Preanalytic aspects in postmortem toxicology. Forensic Science International 142: 75–100. Skopp, G. (2010). Postmortem toxicology. Forensic Science, Medicine and Pathology 6: 314–325. Takayasu, T. (2013). Toxicological analyses of medications and chemicals in formalin-­fixed tissues and formalin solutions: A review. Journal of Analytical Toxicology 37: 615–621. Tellenbach, V. and Tellenbach, M. (2007). Mumien im Andenraum  – Präsenz der Verstorbenen. In: A. Wieczorek, M. Tellenbach and W. Rosendahl (eds.), Mumien – Der Traum vom ewigen Leben. Reiss-­ Engelhorn-­Museen Manheim u. Verlag Philipp von Zabern, Mainz. S 95–112. Thiel, W. (1992). Die Konservierung ganzer Leichen in natürlichen Farben. Annals of Anatomy 174: 185–195. Thiel, W. (2002). Ergänzung für die Konservierung ganzer Leichen nach W. Thiel. Annals of Anatomy 184: 267–269. Trompette, P. and Lemonniere, M. (2009). Funeral embalming: The transformation of a medical innovation. Science Studies 22: 9–30. Von Hagens, G. (1979). Impregnation of Soft Biological Specimens with Thermosetting Resins and Elastomers. Anatomical Record 194: 247–256. Von Hagens, G., Tiedemann, K. and Kriz, W. (1987). The current Potential of Plastination. Anatomy and Embryology 175: 411–421. Werning, J. (2007). Mumien in China. In: A. Wieczorek, M. Tellenbach and W. Rosendahl (eds.), Mumien  – Der Traum vom ewigen Leben, S 123–135. Reiss-­Engelhorn-­Museen Manheim u. Verlag Philipp von Zabern, Mainz. Wheeler, A., Czado, N., Gangitano, D. et al. (2017). Comparison of DNA yield and STR success rates from different tissues in embalmed bodies. International Journal of Legal Medicine 131: 61–66. Williams, E.J. and Davison, A. (2014). Autopsy findings in bodies repatriated to the UK. Medicine, Science and the Law 54: 139–150. Windgassen, M., Larscheid, P. and Tsokos, M. (2017). Leichen als Direktimport: Ein Beitrag über die rechtlichen Grenzen der Nutzung menschlicher Leichen zu Lehrzwecken. Archiv für Kriminologie 242: 97–109. Wunn, I. (2007). Mumien in Klöstern und Kirchen  – Mönche, Päpste und Fürsten. In: A. Wieczorek, M. Tellenbach and W. Rosendahl (eds.), Mumien  – Der Traum vom ewigen Leben, S 144–153. Reiss-­ Engelhorn-­Museen Manheim u. Verlag Philipp von Zabern, Mainz.

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Buckley, S.A. and Evershed, R.P. (2001). Organic chemistry of embalming agents in Pharaonic and Graeco-­Roman mummies. Nature 413: 837–841. Bush, M.A., Miller, R.G., Prutsman-­Pfeiffer, J. and Bush, P.J. (2007). Identification through X-­ray fluorescence analysis of dental restorative resin materials: A comprehensive study of noncremated, crecremated individuals. Journal of Forensic mated, and processed-­ Sciences 52: 157–165. DiMaio, V.J.M. and DiMaio, D.J. (2001). Fire deaths. In: V.J.M. DiMaio and D.J. DiMaio (eds.), Forensic Pathology, 2nd edn, pp. 367–388. Boca Raton, FL: CRC Press. Gielda, L. and Rigg, S. (2017). Extraction of amplifiable DNA from embalmed human cadaver tissue. BMC Research Notes 10: 737–741. Guenevere, R., Husain, M., McGoey, R. et al. (2016). Postmortem aortic dissection: An artifact of the embalming process. Journal of Forensic Sciences 61: 246–249. Hanzlick, R. (1994). Embalming, body preparation, burial, and disinterment  – an overview for forensic pathologists. American Journal of Forensic Medicine and Pathology 15: 122–131. Holck, P. (2005). Cremated bones. In: J. Payne-­James, R.W. Byard, T.S. Corey and C. Henderson (eds.), Encyclopedia of Forensic and Legal Medicine, Vol. I, pp. 281–284. Amsterdam: Elsevier Academic Press. Kiernan, J.A. (2000). Formaldehyde, Formalin, Paraformaladehyde and Glutaraldehyde: What they are and what they do. Microscopy Today 1: 8–12. Mari, M. and Domingo, J.L. (2010). Toxic emissions from crematories: A review. Environment International 36, 131–137. Marquet, P.A., Santoro, C.M., Latorre, C. et  al. (2012). Emergence of social complexity among coastal hunter-­gatherers in the Atacama Desert of Northern Chile. Proceedings of the National Academy of Sciences 109: 14754–14760. Martin-­Gil, J., Martin-­Gil, F.J., Delibes-­de-­Castro, G. et al. (1995). The first known use of Vermillion. Experientia 51: 759–761. Matschke, J. and Tsokos, M. (2000). A short review in the history of cremation. In: K. Püschel and M. Tsokos (eds.), External Examination Before Cremation  – Research in Legal Medicine, Vol. 22, pp. 29–37. Lübeck: Schmidt-­Römhild. Mayer, R.G. (2012). Embalming. History, Theory, and Practice, 5th edn. New York: McGraw-­Hill Medical. Mayne Correia, P.M. (1997). Fire modification of bone: A review of the literature. In: Haglung, W.D. and Sorg, M.H. (eds.), Forensic Taphonomy – The Postmortem Fate of Human Remains, pp. 275–293. Boca Raton, FL: CRC Press. Nikolaou, P., Papoutsis, I., Dona, A. et al. (2013) Toxicological analysis of formalin-­ fixed or embalmed tissues: A review. Forensic Science International 223: 312–319. Piombino-­Mascali, D., Aufderheide, A.C., Johnson-­Williams, M. et  al. (2009). The Salafi method rediscovered. Virchows Archiv 454: 355–357. Pommerening, T. (2007). Mumien, Mumifizierungstechnik und Totenkult im Alten Ägypten  – eine chronologische Übersicht. In: A. Wieczorek, M. Tellenbach and W. Rosendahl (eds.), Mumien – Der Traum vom ewigen Leben, S 71–88. Reiss-­ Engelhorn-­ Museen Manheim u. Verlag Philipp von Zabern, Mainz. Powers, R.H., Criss, B.E., Topmiller, R.G. et al. (2019). Methanol Detected in a Subdural Hematoma as an Embalming Artifact. Journal of Forensic Sciences 64: 946–949.

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valuable, although it should be kept in mind that new information that is obtained later on may change the picture. When arriving at the scene, the forensic pathologist should discuss the situation with the scene investigators in order to agree on appropriate personal protection equipment, and also to obtain information regarding the protection of the scene; which areas have been cleared, and which areas might not as yet be accessible. It is also important to know if the body has been moved by people present at the time of death, by first responders, paramedics, or sometimes by a general practitioner who has certified the death. If so, furniture and other items might also have been moved. If resuscitation attempts have been carried out, there might be marks on the chest, around the nose and mouth, and venipuncture marks on the neck and arms; hence, blood and saliva next to the body may be explained by such interventions. All changes to the original appearance of the scene should be considered when hypothetical scenarios are drawn up. In retrospect, some of the conclusions drawn during the scene investigation might turn out to be wrong, hence it is a good practice to critically review initial observations and conclusions at the scene if and when the death investigation is completed in order to learn more from the personal experiences. It is desirable that a forensic pathologist is called in to participate in a scene investigation liberally, and preferably in an early phase, whenever the primary actions have been taken by the investigators to protect the body and the scene. The advantage of an early participation is that the methods for time of death estimation perform better at shorter postmortem intervals and that the conditions at the scene can be observed before various things have been moved or even removed. However, in many situations, the deceased has previously been pronounced dead by a general

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The basic questions that the crime scene investigators should answer are: Has a crime been committed? If so, how, when, and where was it committed, and who is the victim and who is the perpetrator? The crime scene investigator will rarely be able to answer all these questions, but the findings may be critical for the further death investigation and many times evidence from the scene may be important if and when the case goes to court. The forensic pathologist can assist the scene investigators with professional observations that can add to other findings at the scene, and may be requested to testify in court, to answer questions both regarding the scene investigation and the subsequent autopsy findings. The scene of death may be very variable, but most often we imagine a house or an apartment where a dead person is discovered, and also imagine that the body is not severely decomposed. However, even if the text in this section often will assume such a situation, the forensic pathologist on call should be prepared for visiting places that are quite different. Hence, warm clothes will be warranted if the body is found in snow and rubber boots if the scene is a swamp. It is also a good idea to have something to eat before heading off, since eating and drinking should be avoided at the scene, and sometimes the police requests participation of the forensic pathologist for an extended time as the crime scene work progresses. There may be a number of practical issues to be sorted out before going to the scene, such as the exact location, if there is more than one dead body, and the possibility of health hazards, such as contagious disease, noxious gases, or explosives. A brief account of the circumstances surrounding death is also

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Handbook of Forensic Medicine, Volume 1, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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Most forensic pathology departments or medical examiner’s offices will typically have a suitcase with examination equipment that can be used by the doctor on call. Depending on the duties, this suitcase may include equipment for the examination of living victims and suspects as well as equipment for crime scene visits, or it may exclusively contain items that are useful at the scene; for instance, those listed in Box 9.1. If several colleagues share the same suitcase, a good habit is to make sure that all items are available before ­starting a scheduled on-­call duty; for example, checking that the thermometer and square-­wave generator are working properly. The personal protection equipment is basically used to safeguard evidence, for example, transfer of foreign DNA, but it may be equally important to reduce the risk of health hazards. Often the scene investigating team will offer a coverall, face mask, gloves, shoe covers, swabs, and various light sources, but it is better to bring extras than discover that something is missing, particularly if the scene is at a remote place. Moreover, in some situations, and particularly in developing countries, there are no scene investigators but only regular police officers at the scene with scant equipment. Having said that, the typical scenario is that there is in fact a team of crime scene investigators which has arrived at the scene in a well-­equipped van, so the focus of the forensic pathologist will then be on the examination items that relate to time since death examination and sampling of biological material from the dead body. The forensic pathologist may take advantage of this in

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different ways; the investigators may, for example, use a CrimeScope that can pinpoint spots of saliva or semen on the skin that the forensic pathologist can collect with swabs. Even if the scene investigators will take a large number of photos, these may not be available when the forensic pathologist returns to the forensic medicine department and writes the report. Hence, it is a good practice for the forensic pathologist to bring a digital camera and take photos on his/her own to be used along with sketches and notes in order to remember details and also for conclusions, for example, regarding overall injury pattern. This camera should preferably be equipped with a zoom objective and a macro function to produce photos that can show minute details of important injuries. Sometimes high-­resolution shots can actually be magnified to such degree that small features not observed with a regular magnifying lens can be discovered. A dictaphone can many times help to momentarily document observations. A pen and a notepad require two hands, whereas a dictaphone can be operated with one hand while the other is used for examination. The personal protection equipment may sometimes be extensive if and when biohazards are suspected. It is then important that the different parts fit well and do not disturb the work; for instance, a face shield as well as glasses above a face mask may become misty and obstruct a good vision. Furthermore, it is important that the pathologist repeatedly checks the apron, sleeves, and gloves for stains of blood or other fluids in order to avoid transfer of traces. In particular, the gloves should be changed frequently. A number of potential risks exist to the investigators, depending on the nature of the scene and the circumstances. The forensic pathologist may have certain knowledge about the severity of certain relevant pathogens, whereas the crime scene investigators will have more experience regarding weapons, explosives, noxious gases, and electricity hazards. It is important that the forensic pathologist and the scene investigators discuss these matters in the early phase, so that appropriate precautions are taken during the examination of the scene. A reflex hammer may not often be used, although it can be useful for testing of idiomuscular reaction, even if a specially designed stick might be better for this purpose. Likewise, a stethoscope will rarely be used, but in situations where there are no convincing livores and a barbiturate poisoning is considered, it will be critical to know if the person actually is dead. Tweezers with a good grip is desirable when pulling out the conjunctivae for inspection, and for the examination of certain wounds as well as for picking up hair strands and variable particles present on the body surface. The tweezers and other instruments that have been in contact with such trace evidence should be cleaned when reused to avoid transfer of material. A plastic bag for waste items as well as a small container for sharp objects in reach when examining the body will often prove useful. Small plastic tubes with sterile water can be used to moist s­ terile swabs for collection of dried stains and also for cleaning purposes. If not otherwise indicated, sterile swabs, moistened or not, should be put back into the original package and appropriately labeled. If completely soaked, they might need to let dry to avoid going moldy, but this can usually wait until they are unpacked at the crime lab. The sterile swab packages should also be checked for their

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practitioner, who initially was notified about the death. For those readers who are general practitioners, this chapter contains some useful information regarding signs that may justify the reporting of the death to the police, or, conversely, findings and information that rather indicate a natural death. Occasionally, even the forensic pathologist may be asked to go to a scene at which there are no (as yet) technical staff from the police, and in those situations, particular caution to avoid changes to the scene should be exercised.

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Box 9.1  Equipment for scene investigation.

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x Coverall, apron, face mask, goggles, latex gloves, shoe covers x Calibrated thermometer, square-­wave generator, Mydriaticum Roche, acetylcholine, syringes, needles x Stethoscope, otoscope, reflex hammer, tweezers (Adlercreutz and anatomical), precision scissors, scalpel x Sterile swabs, small paper bags, test tubes and containers, redline bags x Tubes with sterile water, sterile NaCl solution, and containers with formalin x Field drug tests x Camera, dictaphone, ruler, male/female body charts, envelopes x Handheld digital microscope, magnifying lens, torch/headlight

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Box 9.2  Tasks for the forensic pathologist at the scene. Confirmation of death

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Identification of the dead body

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Estimation of the time of death

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External examination of the body

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Documentation of injuries/signs of exposures

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Examination of the scene with focus on explanations for the death

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Giving opinion on the possible cause and manner of death

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Making observations

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Assisting the scene investigators with advice

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e­ xperience, since a number of people have usually already been at the scene and looked at, or even tried to resuscitate, the presumably dead person, so by the time the forensic pathologist arrives lividity has usually been established. The tasks for the forensic pathologist serving at the scene may vary substantially depending on the circumstances. The most common tasks are summarized in Box 9.2. In most jurisdictions, the work at the scene is led by the police, represented by specially educated and trained crime scene investigators, and the role of the forensic pathologist will depend on the demands of the scene investigation team, and their need for medical expert assistance will in turn be dependent on the nature of the scene and the circumstances. It is important that the forensic pathologist checks with the scene investigators what areas are secured, what trace evidences have been, or not yet been, collected and documented in order to avoid interfering with the technical examination of the scene. Further, the forensic pathologist and scene investigators should also discuss which examinations should be done at the scene and which ones can wait until the body has been transported to the forensic medicine department. In some cases, a CT scan or a prioritized autopsy is considered more important to clarify a number of critical questions, whereas a comprehensive scene investigation to secure trace evidence on the body and the scene might be considered more important in other situations. After having certified the death, a fairly common task is to assist in the efforts to establish the identity, if unknown. In some cases, the examination of the body may reveal tattoos and scars that the police consider conclusive when they have reference information about the person. In other cases, the body stature, signs of chronic illnesses, or tooth characteristics may give clues when there is only one or a few likely matching persons. Even if comparison of dental radiographs is the most common method used in forensic identification work, an inspection of the mouth may disclose characteristics such as missing teeth, prominent maxillary or mandibular prognathism or unusual fillings that are considered to be sufficient for a preliminary identification. In cases where there is no clue as to the identity, particularly when the body is severely decomposed or skeletonized, the estimation of the age and determination of the sex will be important to limit the search for possible matches. A visual examination of the body

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expiration date, since this might be an issue later on if the chain of custody and the routines are questioned. The same is true for atropine and other agents used for pupillary reactivity. The digital thermometer should have a probe equipped with a detector, such as a platinum-­100 electrode, located as close to the tip as possible and a length that allows for measurement of the rectal temperature about 8 cm inside the anus in order to record the core body temperature. At least once a year the thermometer should be calibrated at a certified temperature laboratory, implying that the forensic pathology department needs to have at least two instruments in order to always have one appropriately working thermometer. Most probes can be used for accurate readings of both the rectal temperature and ambient temperature in air or on surfaces, providing a one decimal reporting over a wide range of temperatures. For assessment of electrical excitability of skeletal muscles, a square-­wave generator is recommended. Different models are available on the market, and some forensic departments have developed their own. The most common generators produce constant current rectangular impulses of 30  mA with a 10-­ms duration repeated every 50 ms. The voltage and the design of the electrodes may vary; hence, the reference time intervals for the reaction patterns may differ between devices. There are today handheld digital microscopes with high magnification and circumferential, adjustable LED lamps that allow for detailed examination of surface details such as minute insulin injection marks. Although originally introduced with USB connection, there are now a number of these devices which allow for snapshots and a wireless operation with a smartphone, tablet, or laptop. Having said that, a regular high-­quality magnifying lens is still an item that often can disclose important evidence. Rapid field test kits may be of interest in some cases, for example, to detect drugs of abuse, and several of these can be used on saliva/oral fluid, urine, or blood. Even though toxicology in most cases will be performed later on with advanced and validated methods, preliminary results at the scene might lend support (or not) for the presumption that the deceased was a drug addict. There are actually also rapid test kits for various infectious diseases, allergy, and heart conditions. Hence, such test results may give information about the deceased subject that may add to other observations when considering possible causes and manner of death. In addition to such rapid tests, there are portable analytical systems, for example, blood gas instruments used by medical staff in some ambulance helicopters, that can give immediate readings of electrolytes, glucose, pH, urea, etc., but such results are rarely urgently needed at the scene. However, if a benchtop instrument is available at the forensic medicine department, results obtained a few hours later can be valuable for the further death investigation.

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9.3  Tasks for the forensic pathologist If there is any chance that the person is still alive, or can be resuscitated, all actions to preserve life should be given priority. Such a situation is very unlikely that the forensic pathologist will

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the bullet path in the body. If there is only one gunshot wound, then both the position of the entrance wound and the direction of the bullet path may assist in the differentiation between a suicide and a homicide (Druid 1997; Karger et al. 2002). In contrast to several other forms of forensic casework, such as laboratory analysis, it is impossible to establish a standard protocol for all procedures to be carried out at the scene, because of the variation of the conditions and situations. Hence, some of the tasks listed in Box 9.2 will not apply because a number of questions are already resolved, and represent so-­called “species facti,” whereas other questions may not be possible to answer, for example, the cause of death when only a few skeletal remains are found. Hence, it is difficult to determine what actions the forensic pathologist should take, and rather the work should be based on the needs expressed by the police. Having said that, the observations that the forensic pathologist does at the scene may add new questions leading to additional tasks for both the investigating team and himself/herself. Further, even if no standard protocol for the whole scene investigation can be established, certain examinations, and collection of trace evidence, should of course follow established procedures if applicable. The forensic pathologist should take notes frequently, use sketches, and take own photographs with the consent of the scene investigators, in order to describe details correctly to avoid drawing inferences that are not compatible with the actual findings at the scene. After some time, the memories of certain details will fade and then the documentation (written notes, dictates, and photos) at the scene will become very valuable.

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at the scene may sometimes allow for a determination of the sex, whereas the age is usually very difficult to estimate, and the forensic pathologist should refrain from giving narrow ranges. If requested to estimate the time since death, the methods described in Chapters 7.1–7.5 are recommended. In Section 9.4, some aspects on the procedures are provided. In general, it is recommended that the examination for time since death is performed as soon as possible for several reasons; the re-­establishment of rigidity needs some time to pass before evaluation, most methods will perform better at shorter intervals, and both the scene investigation team as well as the head of the death investigation at the police will typically want to know a time frame for the death as soon as possible, for example, as a basis for decisions to arrest or not to arrest a suspect. One of the most common questions for the forensic pathologist is to interpret major injuries, and how these may have been sustained. A conspicuous injury can be the sole reason for the appearance at the scene of a death that otherwise is not suspicious. Hence, a comparison of the injuries with weapons and other objects, protruding edges and corners of furniture, wall moldings, and thresholds, and with bloodstain patterns is often warranted. The question may also be raised regarding the age of injuries, and if all, or some, of them might have been sustained at a different place. Actually, in some cases, it might turn out that the scene of death is not the scene of crime. It is therefore important that the forensic pathologist and the investigating team keep an open-­minded discussion about various observations during the investigation and avoid to get stuck with one hypothetical scenario. In some cases when the cause of death is obvious, the task will rather be to give an opinion on the most likely manner of death, particularly differentiation of homicide versus suicide, or homicide versus accident. In other cases, both the cause of death and manner of death are unclear, and then additional alternatives are added, such as different forms of natural death. A common situation is the finding of a large amount of blood at the scene assumed to come from the mouth and/or nostrils. Then an examination of the nature of the blood (e.g., blackish-­brown tinges suggesting a gastric ulcer as the origin) and an inspection of the mouth (large bite marks on the tongue) and the facial skeleton (instability of the nose and maxilla) are important. Similarly, an examination of the surroundings (places where a trauma may have occurred, hard objects that may have been used to cause injuries) as well as a review of known diseases of the dead person (liver cirrhosis, abdominal pain) can provide valuable clues. In cases of hanging, the focus will often be to identify signs of intravitality of the suspension mark on the neck, and possible unexpected other injuries on the skin of the neck. Furthermore, it is important to carefully study the suspension mark and how well it matches the ligature. In many cases, a ligature has been removed and resuscitation attempts performed, which requires a reconstruction of the original position and orientation of the ligature and of the body. In cases of gunshot wounds, the crime scene investigators will often be able to make a reconstruction of the bullet trajectories, but the forensic pathologist may then have the task to identify entrances and exits on the body and how these are connected in order to determine

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9.4  Time of death The methods recommended for time since death estimation to be used in the early postmortem interval are described in some detail in Chapters 7.1–7.5. Most of them have been used for several decades, and ever since publications of their use in combination were ­published (Henssge  1988; Henssge et  al. 1988; Madea  2016), such examination protocols have been extensively applied in several countries. As already pointed out, most methods will perform better at shorter postmortem intervals, and it is therefore usually a good practice to conduct the time of death estimation as soon as possible. The order in which examinations and tests for time of death estimation are performed is very dependent on the conditions at the scene. If certain parts of the body first need to be subjected to trace evidence collection, for example, in suspected sexual assault cases, certain time of death methods may have to wait. If no such concerns are at hand, usually a test of skeletal muscle rigidity of one arm and one leg may be the first choice, since this allows for a testing of re-­establishment during the time when the forensic pathologist remains at the scene. An early testing for Zsako’s reflexes may also be justified if the postmortem interval is believed to be very short. Furthermore, sampling of vitreous humor is a very quick procedure and will not cause any delay for other tests. Lividity is also easy to examine with a thumb pressure. Examination

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many blood vessels that can be injured. Hence, the eyelids, the sclera, and the conjunctivae should be inspected and documented before testing the musculus orbicularis oculi muscle, and the same applies for other areas where the excitability is tested. The thickness of the electrodes may vary and also their shape; some are thin and rounded with a very pointing tip and others are somewhat blade-­shaped with less pointing tip. The thicker the electrode, the larger area of the muscle will be affected by the electric current, and also the voltage may vary between wave generators. It is therefore important to compare the reactivity with reference intervals for the particular equipment that is used. Even though there is a large body of literature about methods for time of death estimation, the results will to some extent be dependent on the investigator. It is recommended that the forensic pathologist repeatedly applies the different methods for time of death estimation on cases with a known postmortem interval (and known ambient temperature and other factors that may impact on the results) whenever there are opportunities to do so. In particular, the impact of clothing – none, or one, or several layers – and combined effect of surface and air temperature can be difficult to estimate, and the more the cases examined, the better the forensic pathologist will become to use the pertinent correction factor for the temperature method.

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for pupillary reactions, body and ambient t­emperature, and electrical excitability may however take a little longer time and require preceding examination of these body regions. Concerns may be raised as to the introduction of a probe into the anus for rectal temperature if sexual assault is suspected. This problem may be resolved by putting the probe into a finger of a glove from a “sterile” single-­package. When removing the probe, the glove can be turned inside out so that any material in the anus and rectum such as sperm or other foreign material can be secured and later analyzed. Of course, such a procedure can imply that material outside of anus follows as the probe covered with the finger of the glove is introduced, but by having two co-­workers lifting each leg, the anus can usually be well exposed and contamination from the surrounding skin avoided. In the early postmortem interval, the measurement of rectal and ambient temperature remains the most practiced method for time of death estimation and hence there should be strong reasons for omitting taking the rectal temperature at the scene if the time of death is considered important for the death investigation. Some professionals advocate the use of ear temperature, liver temperature, or brain temperature, and sometimes these may be an alternative, for example, if the body has undergone a rectum amputation, but there are at present less reference data for these temperature recordings and both liver and brain temperature measurements will cause physical damage to the tissues. When measuring the ambient temperature, it may be necessary to record both the temperature on the floor and in the ambient air to obtain a representative figure of the temperature that the body has been exposed to since death. This temperature then might need to be adjusted depending on if doors or windows have been opened, or closed, during the postmortem intervals, for example, by the person(s) who discovered the body, or by first-­responders. Hence, several recalculations of the postmortem interval may be necessary due to additional information obtained later on. Vitreous fluid is easy to collect with a syringe and an 18-­gauge needle inserted at the lateral canthus of the eye bulb. The tip of the needle can usually be seen through the pupil. A small amount of fluid, for example, 0.2 ml, from each eye can be collected in the same syringe and transferred to a clean Eppendorf tube for analysis at any clinical chemistry lab. The concentration of potassium and other electrolytes in the tube will remain the same for appreciable time. If the forensic medicine department has a benchtop instrument such as a blood gas analyzer, the fluid in the syringe can be directly injected and a result obtained within one minute or so. Repeated studies on treatment of the vitreous fluid have shown that centrifugation and sonication steps are not necessary (Zilg et al. 2009; Zilg et al. 2015). If the body is lying with the head down so that the tissues in the face have become congested post mortem, there is a possibility that the sampling can cause a local hemorrhage at the lateral aspect of the eye, so it is advisable that the eye, conjunctivae, and eyelids are inspected and documented before the sampling. As compared to vitreous sampling, bleedings are more common after testing for electromuscular reactivity. Since the electrodes are inserted at some depth into the muscles, there are

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9.5  Examination of the body

As previously pointed out, the scene investigators and the forensic pathologist should discuss possible health hazards and take pertinent precautions before the body is examined. For instance, at autopsy, there is a very low risk of contracting an infection from a dead body, since the interval from death to autopsy often is a few days, and by that time there will not be sufficient amounts of active virus particles to cause an infection. Having said that, there are reports about transfer of contagious diseases from lifeless persons to first-­responders/ambulance personnel exposed during resuscitation attempts and in particular to the staff during intubation. At the scene, the forensic pathologist might need to look at details on the body, so if the person recently has died and may have a dangerous infectious disease, particular caution should be exercised. However, even so, the risk is still low except in countries with a fair prevalence of hemorrhagic fever in the population. The external examination of the body should ideally include all parts of the body surface and the natural orifices. To this end, the clothes need to be taken off. However, in certain cases, the investigators want to undress the body at the morgue or at the forensic medicine department to have better control of the match between various injuries on the clothes and the skin, or to protect certain stains and other trace evidence on the clothes. Whenever the body is turned around at the scene for the sake of the external examination, the forensic pathologist should keep in mind that blood may leak out from wounds and produce new blood-­flow patterns on the skin or clothes and pools of blood on the floor. One possible option to consider is to put adhesive or transparent

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insignificant, but with a position that could indicate that they represent defense injuries and neck compression, respectively. Similar reasoning may apply regarding other manners of death; cut wounds on the palmar side of the wrists are often not as deep to cause death, but will in most cases suggest a suicide, although by a different method, such as intoxication. Examination of the scene of a fire is often challenging, particularly since the apron, gloves, protective sleeves, and shoe covers easily get sooty, implying that the personal protection equipment, particularly the gloves, needs to be changed repeatedly, interrupting the investigation. Furthermore, injuries on a charred body may be difficult to identify. The examination of the body at the scene therefore often will be somewhat limited and focused on features that indicate that the person was alive during the fire, such as soot in nostrils and mouth, crow’s feet sign next to the eyes, and bright red lividity. When the body is to be turned around to examine the undersurface or later on lifted into a body bag, and again lifted onto a stretcher, care may be needed to avoid causing injuries if the body has become brittle. Outdoor scenes are less common and may pose various problems; in an urban setting, there may be a need to protect the scene from curious people, and even a tent may be necessary to put up; an examination may have to be done at low temperatures when a body has been found in the snow; and, by contrast, mosquitoes may be the main problem if the body is discovered in a moist subtropical forest. In these situations, decisions often are made to limit the external examination to the most relevant body regions and then give priority to a more thorough examination in a nearby morgue or at the forensic medicine department. For example, the case of a woman who was eventually considered to have been killed by a moose was described by Gudmannsson et al. (2018). The woman was found dead in a forested area in the summer and scene investigators and a forensic pathologist visited the scene. However, the investigation was carried out in the dark, and during heavy rain. Due to multiple deep wounds, and the finding of flail chest upon autopsy, the police suspected that the husband had killed her by means of a riding lawn mower, but later on, trace material from the body proved to originate from a moose. It may be speculated that if the scene had not been assumed just to be a spot on which the body had been dumped but rather the scene of death, a more careful re-­examination might have revealed many moose footprints and alerted the investigators to consider a moose attack already in the early phase. One question may be raised regarding the possibility of incapacitation, that is, if the injuries observed are so severe that the person was unable to move. Sometimes this is important in order to pinpoint the position and/or activities of the person at the time when the injuries were sustained. There are published reports describing subjects being shot or stabbed in the chest, who were able to walk or even run for quite some distance despite large wounds of the heart observed at autopsy. Similarly, delayed death is a well-­known feature of a proportion of persons developing an acute subdural bleeding. At the scene, the problem will be that many injuries that cause death are not visible, or only partly visible upon the external examination. It is thus recommended that

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film over such wounds to prevent leakage. These can then be removed at the forensic medicine department and be saved for further examination if needed. The most important task when doing the external examination is of course to look for various injuries due to blunt force, sharp force, firearms, cold, fire, electricity, and more, but also to look for signs of other exposures, for example, gases such as carbon monoxide. Intoxications by pharmaceutical drugs and drugs of abuse are common and may be very difficult to exclude, since most of them leave no specific traces on the body. One important exception are opioid intoxications, in which white, or sometimes blood-­tinged, foam may protrude from the nostrils and mouth (and at autopsy frothy liquid usually will be present in the lower airways) mimicking the foam seen in many drowning victims. In cases subjected to resuscitation, the foam may have been removed, and the police may then need to ask the people first present at the scene about their observations. It is also important to document absence of injuries, since certain preliminary hypotheses might require that a particular body area should display injuries. The external examination may also reveal signs of disease. Small, rounded bleedings on the abdomen and/or thighs may be caused by repeated insulin injections, suggesting that the deceased was a diabetic. Buffalo hump, plethora, and rounded face along with hirsutism may indicate a chronic use of cortisone because of asthma or rheumatoid arthritis, and in such cases, bruises may in part be explained by this medication. Pitting edema looks and feels just the same as in a living subject and suggests that the person suffered from cardiac failure. There are a number of additional diseases that may be suspected from changes of the body exterior, but focus should of course be on such medical conditions that could lead to fatal complications and represent possible causes of death to be considered just like alternatives due to unnatural causes. If there are many injuries, the character, distribution, and pattern of the injuries are all important to consider rather than putting too much efforts on describing them in detail. The investigation team will usually want to know how and when the injuries have been sustained, and if they may be self-­inflicted or caused by another person. To this end, it is important to open-­mindedly consider a number of possible alternatives that are compatible also with, for example, the position of the body, the characteristics at the scene, and the known circumstances. Since new information can be added later on, it is advisable not to exclude certain alternatives too swiftly, but at the same time provide the scene investigators with opinions about the likelihood of different scenarios. The examination may be general and inclusive in obscure deaths where no particular hypothesis of the cause of death has been postulated, but where circumstances (e.g., the finding of a young woman who has been repeatedly threatened by her ex-­ boyfriend) are the reason for a careful death investigation. Then particular efforts should be put in examining details that could provide clues. A small abrasion at a high position in the scalp may not seem significant, but should warrant an explanation if above the hat brim line, and might be the site of impact of a blow that could have caused a subdural hematoma. There might also be minor injuries on the arms or the neck, which as such also are

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Case example 1

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A middle-­aged couple had come home from a party and while the man was brushing his teeth in the bathroom, he heard a bump from the adjacent hallway and looked out to find his wife lying on the floor. He helped her up to the bedroom on the second floor. Next morning, both woke up, talked briefly, and then fell asleep again. A couple of hours later the man found his wife lifeless, and she was pronounced dead upon the arrival of ambulance staff. At autopsy, there were fractures of the left superior cornu of the thyroid bone, a fracture of the fifth cervical vertebra, a sternal fracture, one rib fracture on the right side, and three rib fractures on the left side, and almost 2 l of blood in the pleurae. In addition, signs of an impact to the back of the head and localized hemorrhages in the soft meninges were also observed. The cause of death was considered to be blood loss. The man was arrested and a reconstruction at the scene was arranged. At that time the husband stated that he had a vague memory of having heard several bumps while he was in the bathroom, suggesting that his wife might have tripped and fallen in the steep stairway (Figure 9.1). After some additional investigation, the prosecutor decided to close the case, because of the lack of a specific scenario that could represent a criminal act. When the suspicion of possible homicide was raised, the house had been cleaned and there was no useful trace evidence to be found. This example raises the question as to how many injuries can be sustained at multiple body regions during a fall in a stairway. Even if tests with a doll are carried out, there will be a large number of variations of impacts as a body tumbles down to be considered; hence, suspected fall in stairs seemingly remains a challenge to the forensic community, but a careful investigation of the untouched scene may disclose impact sites that may be compared to injuries found on the person.

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The focus for the forensic pathologist at the scene is of course the dead body, but there are several reasons for an overall examination of the scene to generate a preliminary hypothesis of the sequences of events that preceded the death. As previously pointed out, in most situations, there is a leader of the scene investigation team who will be responsible for all actions taken at the scene, so it is important that the forensic pathologist should follow his/her directions in order not to interfere with the technical investigation. Occasionally, a pathologist makes findings at autopsy, and may ask the police to visit the scene. Most doctors who do that will experience that even if the place has been well described in the police report, and even if several photos have been available, the visit to the scene will present many specifics that were not appreciated from the documents and photographs reviewed. Hence, scene investigation should be considered as a privilege, since many question marks can be straightened and certain imagined scenarios can get support, whereas others might become less likely. In particular, the visit to the scene will provide a true 3D view and a better appreciation of distances between various objects. Hence, just standing and walking around at the scene, the space available, or not, for various movements of the deceased person and possible suspect(s), can be much better appreciated than by reviewing documents and photographs. Usually, the scene investigators will provide the forensic pathologist with a brief account of the circumstances and their findings so far. Then the pathologist will be asked to examine the body, and usually the scene investigators will assist, and may in parallel collect trace evidence and make sure that the illumination is appropriate. Depending on the findings, the forensic pathologist may want to search for possible sites of impact that may match various injuries observed. Such spots may already have been identified by the scene investigators, but might not be considered by the forensic pathologist to be a satisfactory explanation. The advantage of the presence of both a trained physician and experienced scene investigators is obvious, since the combined competences can be very fruitful. Whereas the scene investigators are experts on technical aspects, understand what to search for, and may easily identify small and large pieces of evidence materials, the forensic pathologist will contribute with substantial medical knowledge including anatomy, pathophysiology, relevant medical conditions, toxicology of drugs, and traumatology. Hence, the scene investigators may indicate a bloodstain pattern and explain the origin of the blood, while the forensic pathologist can make a statement regarding the most likely injury out of several wounds. Sometimes tramline injuries are observed, and then the task will be to search for long, hard objects such as sticks, rods, and

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bats, but even parts of a chair may be considered. Falls in stairs represent a common situation, but often it is very difficult to reconstruct how different parts of the body might have impacted on the steps, the handrail, the walls, and finally the floor, if reached. Then the scene investigators may contribute by a detailed examination of sites of impact, particularly if the skin or clothes have been stained with fresh blood that has produced contact marks.

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the forensic pathologist exercises caution when providing an opinion on this at the scene, since the autopsy may reveal a different panorama of internal injuries than initially assumed.

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Figure 9.1  A steep and winding stairway in which the woman in case example 1 was supposed to have fallen.

MEDICAL ASPECTS OF DEATH

Box 9.3  Samples that may be collected. x x x x x x x x x

Vitreous fluid Femoral blood Urine (puncture) Nasal, oral swabs Genital and anal swabs Hair samples Nail scrapings/clips Stains on the skin (semen, saliva, etc.) Tissue fragments

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clearly labeled with ­information about the type of sample, the sampling site, and the date. If the identity of the dead person is known, then the person’s name should also be noted if the specimen is collected from the body. Most of the sampling at the scene will be the task for the scene investigators, but often the forensic pathologist is asked to take care of the sampling of material present of the skin and from the body orifices. Box  9.3 displays some samples that the forensic pathologist might consider to collect at the scene. Material from the nails may either be collected with a specially designed small semi-­sharp stick from the underlying of the free ends, or be collected by clipping the free ends of the nails. The material from the right hand and left hand should be put in separate containers, and in certain situations the clippings from each nail may be put in separate containers. Material from the nails should be collected before fingerprint testing. For fibers, loose hair strands, and similar small material, special transparent tapes are preferably used. It is rarely possible to examine hair strands in some detail, but if a high-­quality loupe or even a handheld digital microscope (see Section 9.2) is available, it may be possible for a trained expert to differentiate hair strands that originate from the head of the dead person from, pubic hair and guard hair from pets. A close microscopic examination of loose hair strands may also reveal if the hair has fallen off or pulled out. In the latter case, follicular tissue (root sheath cells adhering to the root) may be identified. In cases of suspected sexual assault, it is recommended that a so-­called rape kit is used. This usually contains everything that may be needed for all sampling including a comb for collection of possible foreign hair or other material in the pubic region, a set of swabs pre-­labeled with sampling sites and sampling types, and test tubes for collection of blood and urine (which may be more appropriate for examination of living subjects) along with detailed instructions and a checklist for the examination procedure. The standard routine is to put all the samples in their cover packages back into the rape-­kit box.

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A common situation is that a fatal intoxication may be considered in a case of obscure death. If the person is a drug addict, a search for injection marks on the body, not limited to the antecubital fossa but at several other body regions, may be justified, but equally important is the search for paraphernalia such as needles, syringes, spoons, aluminum foils, lighters, redline bags, powders, rolled cigarette paper, and even nasal sprays that may suggest a recent intake of hazardous drugs. It is furthermore very common in Western countries that people, even when at good health, keep various medications at home, in the kitchen, or in the bathroom. The task will then be to identify drugs that may cause a fatal intoxication, and also to search for signs of a recent intake of a large number of pills. In outdoor settings, there may be many different things to consider. Regarding the time of death estimation, the ambient temperature might have varied considerably, and very soon several species of flies will find the body and lay eggs, and the maggots that follow will often destroy select tissues at a fast rate. Even if there is an entomologist contracted by the police, he or she might not be on call and therefore it is advantageous if the forensic pathologist has some basic knowledge about common species of flies and bugs, and their development stages. Similarly, if various injuries are found, it will prove valuable if the forensic pathologist is familiar with bite marks caused by rodents in order to separate such injuries from antemortem injuries (Tsokos et  al. 1999). Furthermore, a doctor interested in botany may upon the examination of the body observe leaves from trees or bushes that are absent at scene, suggesting that the body has been moved. In addition to all the possible causes of death that can be considered in a regular indoor setting, some alternatives can be added for persons found dead out in the nature such as heat or cold exposures, lightning, and snakebites.

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Samples from the dead body may be collected for various purposes, for example, vitreous fluid can be collected for time of death estimation by analysis of the potassium concentration, but most of the sampling will regard trace evidence. To this end, sterile swabs are extensively used, but occasionally in cases of explosions, there might be suspected small pieces of tissue that can be put in a small container with formalin for microscopic examination, or perhaps also split to allow for DNA analysis. The investigation team might have a polilight device or a CrimeScope which may help to identify possible stains of semen or saliva on the body. Dried blood, semen, and saliva stains are best removed from a surface with a sterile cotton-­tipped swab lightly moistened with distilled water from a dropper bottle. Stains that are wet can be collected without moistening the swab. When using the swab to collect stains, a part of the cotton tip should be left intact in order to be used as a control area upon analysis at the crime lab. Unless otherwise indicated, the swabs are put back into their original packaging, which should be

9.8 Documentation It is recommended that the forensic pathologist takes written notes or uses a handheld dictaphone to record findings in real time rather than waiting until the scene investigation is c­ ompleted,

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In most instances, the forensic pathologist is asked to give an opinion about the cause and manner of death. For the scene investigators, the manner of death may be of prime importance for their decisions about how much efforts they should put in the examination of the scene. If suspicions of a crime can be dispelled from the doctor’s findings at the scene, the scene investigators may limit their examination of the scene and give priority to other urgent cases being investigated in parallel. In some cases, the cause of death is obvious, such as massive blood loss due to deep cut wounds to the neck, but the manner of death may be either homicide or suicide. The results of the examination that the forensic pathologist performs may not always be sufficient to determine the manner of death, but a combination of the pathologist’s and the crime investigators’ findings is needed to establish the most likely manner of death. Figure 9.2 illustrates a case where the suspicions of a criminal act changed over the course of the death investigation. The sequence of events is briefly described in the following text.

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Figure 9.2  The degree of suspicion of a crime that different observations and information during a death investigation raised in case example 2. Points 1–3 refer to the time for the scene investigation, and points 4–8 to the autopsy and additional information requiring further analysis.

Case example 2

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since it may be difficult to remember all observations and their particulars. Sketches may also be helpful, for example, to mark the position and characters of injuries of interest. If the scene investigators give their consent, the forensic pathologist may take own photographs, which can be reviewed before photos taken by the scene investigators are obtained. However, many photos may turn out to be of suboptimal quality, and it is recommended that forensic pathologists either take a course in photography or study various Internet sites for guidance regarding, for example, shutter, and aperture settings and learn how to best obtain representative colors, since this may be important for the interpretation of bruises. Whereas crime scene investigators are likely to have an obligation to always provide a written report of their scene investigation in most jurisdictions, such requirements may not apply to all forensic doctors visiting the scene. However, it should be kept in mind that the opinions expressed to the scene investigators, regarding cause and manner of death and how various injuries likely have been sustained, may impact on different decisions that the police make. Hence, it is advisable that such opinions expressed at the scene also are documented in writing, since this allows for a transparency of communication and obviates possible disagreements later on as to what was communicated at the scene. At a minimum, the written documentation should include all tests (and their results) for time since death examination, if performed, and the most relevant findings as well as a list of all samples collected. It should also be stated which samples have been given to the scene investigators and which ones have been brought to the forensic pathology department. It is important that the documentation can be used to prove an appropriate chain of custody.

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All of the events described here are displayed graphically in Figure 9.2 in terms of their impact on suspicion of foul play. (1) A middle-­aged woman was found dead in her apartment by her husband, who calls an ambulance. The woman shows livores and is stiff, hence the paramedics do not do anything, but instead notify the police. (2) The police officers find the woman lying in a couch almost naked and there are numerous large bruises all over the body. The husband, who is drunk and confused, is arrested. (3) The forensic pathologist on call is requested to come to the scene, and confirms that there are quite a large number of bruises, but that most of them are old and at positions rather suggesting accidental falls. (4) The neighbors report that the couple used to constantly drink a lot of alcohol, but that they had not heard any fight. 5). The autopsy does not reveal any fractures or any internal injuries, and there is no sign of significant blood loss. The only major finding is jaundice and a massive fatty liver. The husband is released from custody. (6) Toxicology shows a femoral blood alcohol level of 2.7 promille, but no drugs. The case is signed out as an alcohol-­related death. The day after the husband was released from custody, he is found dead in the apartment. The autopsy showed a massive subdural hemorrhage of somewhat uncertain age. At this point you may consider that the victim rather was the murderer and the suspect the murder victim, although there was no firm evidence at the scene or from the autopsy to support this notion. (7) Five years later, the son contacts the police and wants to confess that he had “killed his parents.” He was unaware of their causes of death. He explains that he had added a cleaning agent into their spirits to make them distasteful, hoping that the couple then would abstain from drinking alcohol. The product contained ethylene glycol. 8). There was no material left for toxicological analysis, but the microscopic slides were available. Renewed examination of these slides with polarized light from the kidneys did however not reveal any birefringent crystals.

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In summary, there is no standard protocol that can be followed for the forensic pathologist appearing at the scene. In most jurisdictions, the leader of the crime scene investigation team will be formally responsible for all measures and action taken at the scene. However, in practice, the forensic pathologist and scene investigators will typically establish a close collaboration and jointly make various decisions in order to make the investigative process as productive as possible. The tasks may vary depending on the circumstances and the conditions at the scene, but may to some extent also be determined by the leader of the death investigation at the police, a coroner, or a prosecutor. The forensic pathologist should be prepared for variable conditions and assignments, and should conduct the work carefully and with an open mind.

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9.11  Bloodstain pattern analysis 9.11.1  General aspects

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fluid, it behaves more or less like water rather than highly viscous fluids, and results from raindrop research can typically be applied on blood drop behavior. Blood is influenced by cohesive forces such as the viscosity and surface tension as well as disruptive forces (e.g., air resistance, gravity, and applied forces). These inherent properties will always be the same regardless of external forces acting on the blood, meaning that the effects of any given impact can be fairly well predicted. First of all, blood droplets in flight have a spherical shape, even if those detaching from a surface by the influence of gravity initially have a teardrop-­like shape. During flight, the droplet will display oscillations that abate with time as the mass of the droplet stabilizes. The drop will strive to maintain the spherical shape as long as it travels in air. The bloodstain pattern that we see is the final result of the impact of a mass of blood that collides with a surface, but is preceded by a few steps during which the blood will change shape, see Figure 9.3. The images in the left column in Figure 9.3 show the phases for a falling drop that impacts a surface at 90°. In the early contact phase, the lowermost part of the blood drop will expand outward, while the remainder of the drop maintains a rounded shape, forming a dome. The blood mass will then continue to expand radially as the remainder of the blood on top is added, making the surface area covered larger than the diameter of the drop during flight. The displacement phase implies that the blood mass of the drop has completely expanded outward and the surface area has become larger. Actually, this surface area will remain unchanged in the two subsequent phases and hence represent the final size of the stain. The lateral expansion during the displacement phase is substantial; a 4-­mm droplet can cause an 11-­mm stain. The degree of lateral displacement is dependent on the velocity of the drop and hence drops falling from a height will create a larger stain than a drop of equal size falling a short distance. At the boundary rim of the collapsed droplet, short spines or protuberances are formed directed outward and upward. The third step is called the dispersion phase during which blood is forced further lateral, but also upward causing a rise of the rims and protuberances. Upon a 90° impact, the blood mass may resemble a blossom. If the force impacting the blood mass is sufficient, some of the protuberances may spawn satellite drops that detach from the rim. This phenomenon requires that the force overcomes the inherent cohesive forces of the blood mass. The last phase is called retraction and results from the surface tension striving to pull the fluid back into a single form. This may lead to a well-­ demarcated rounded stain, but if the forces causing outward shifts in the blood mass overcome the surface tension, spines will be protruding from the margins and satellite drops may be formed. The size of individual stains is important for the interpretation of blood deposits. The size of the stain caused by the impact of a drop of blood will primarily depend on the size of the drop. The size of drops falling passively will mainly be determined by the character and size of the area from which they detach. The size of free-­flying blood droplets is dependent on the initial wounding agent (force applied). Hence, the higher the force, the smaller size

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In some cases, the only reason for reporting the death to the police may be the finding of blood at the scene. If the death concerns a decomposed body, it may be soon discovered that the assumed bloodshed instead represented putrefaction fluid flowing from the mouth and nose, and if the death otherwise was not suspicious a scene investigation might not be performed. Similarly, if there actually are bloodstain patterns, but the examination of the dead body and the scene suggests a gastrointestinal bleeding, further investigation at the scene might be considered unnecessary. However, if there is a suspicion of crime and there are conspicuous bloodstain patterns, a thorough examination of these may provide valuable information for the death investigation. Scene investigators are usually well trained in bloodstain pattern recognition and interpretation, but their expertise may vary, and in some situations, it may be necessary for the forensic pathologist to study bloodstains in some detail to assess their possible relation to injuries observed on the body. In order to do so, some basic understanding of blood behavior when passively or forcibly released from injury sites and when impacting various targets is valuable. This subsection therefore contains a description of some common bloodstain patterns, their characteristics, and underlying mechanisms.

9.11.2  Blood properties and behavior Blood is a colloidal fluid with a viscosity of 4 poise (1 poise = 0.1 kg/m/s). Water has a viscosity of 1 poise, whereas the viscosity of ketchup is 50,000 poise. Hence, even if blood is a ­non-­Newtonian

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Contact/ Collapse

Displacement

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of the droplets released. Hence, when assessing a bloodstain pattern, the preponderant size of the individual stains is used to classify the spatter as low-­, medium-­, or high-­velocity impact spatter. There is an overlap between the groups, but medium-­velocity impact spatter is characterized by the presence of stains with a preponderant size of 1–4 mm resulting from forces up to about 7.5 m/s (e.g., produced when a perpetrator is repeatedly boxing or kicking a victim). In high-­velocity impact spatter, requiring a force in excess of 30 m/s, the preponderant stain size is less than 1 mm, and sometimes the stains are so small that a mist-­like pattern is produced (Figure 9.4). Drops falling passively onto a surface, for example, from wounded veins, will cause much larger stains than drops ejected from a wounded artery under a high pressure. An additional important principle is that falling droplets of the same size will produce larger stains during a longer impact of gravity according to an asymptotic curve (Figure 9.5). In practice, most drops falling passively down to the floor from a wounded person will have a diameter between 12 and 17  mm. The stains rarely become more than 22 mm even when the drop falls from heights exceeding 200 cm, since the air resistance then will impede the gravitational acceleration. The size of a stain will thus depend on the size of the free-­flying droplet and its terminal velocity as it impacts its target. Hence, a small droplet formed after a heavy blow to the head of a victim and that hits a nearby surface will produce a larger stain than a drop that has traveled a long distance before impacting a similar surface, provided that they had the same size in the air. The fate of drops impacting the surface at more acute angles is shown in the middle and right columns of Figure  9.3. The same principles for each of the phases as in the left column apply, but since the gravity will act on a larger mass at one part

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Figure 9.3  The phases that falling drops undergo as they impact a target at different angles. Source: Illustration: Eszter Nagy, London. See text for details.

Figure 9.4  A schematic of a forward spatter from a gunshot wound. Source: Illustration: Eszter Nagy, London.

of the blood mass, one or more longer spines may be formed, and drops will more easily detach at the lower end of the main drop. Please note that as the protruding end of a spine becomes overstretched, a thinner connecting string will be created, and when the spine breaks, the spine and the satellite stain will have their pointing ends toward each other. This means that the

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s­ atellite stains will show a rounded forward edge in contrast to stains impacting from a distance at an angle which will have a pointing forward edge. From Figure 9.3, it can also be appreciated that the distribution of spines and satellite drops will focus more as the angle becomes more acute. However, when assessing the angle of deposition regarding stains on objects, it should be kept in mind that the object that the drops have impacted may have been moved after the deposition; for example, furniture that partly or completely overturned during a fight (see Figure 9.6). Although the final result of the impact of a mass of blood on a target can be well predicted from the properties and behavior of blood, the characteristics of the surface are also important. Most of the bloodstain appearance described here will be true for a smooth, nonporous surface. However, rough surfaces (even appearing smooth at a quick glance) can give rise to a more irregular pattern. Further, there are porous nonabsorbent and porous absorbent surfaces. The former can for instance be asphalt and the latter cardboard or a blanket. Hence, regarding a blood drop impacting a target at an acute angle, irregularities of the surface may hinder the release of satellite stains going forward and to the left, but allow the release of satellite drops going to the right. However, such effects will be at random, and by studying the characteristics of all stains in an area, the directionality can usually be determined even for patterns on quite rough surfaces such as asphalt. Porous absorbent materials such as a wool sweater pose an additional problem, since droplets can travel deep into the material without leaving any clues as to the angle of impact. In the health care, we are using different measures of blood coagulation such as activated thromboplastin time and prothrombin time. For bloodstains, a few morphological alterations due to the activation of the coagulation can be observed after deposition. Initiation is reported to start after 15 s to 1.5 min, and no flowback 5–20 min after bloodshed. Clot retraction, defined as serum separation from the fibrin mass, has been reported to

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Figure 9.5  The impact of the height from which blood drops are falling on the resulting stain size.

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Figure 9.6  Blood drops have dripped on a cupboard as it was lying down, and blood flows have been created as the cupboard was raised. The schematic to the right shows flows from an oversaturated smear. Source: Illustration: Eszter Nagy, London.

begin from 30 min to 1.5 h after bloodshed (Wonder 1985). Of course, the clotting of a deposited blood mass is affected by several factors; for example, the blood coagulation capacity of the blood (e.g., anticoagulation medication and liver disease), the size of the deposit, and the ambient temperature. In addition to coagulation, morphological changes of a bloodstain will be caused by other factors; in particular, evaporation. The

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obtaining a time frame for a suspected or obvious homicide in the early phase of an investigation, it is expected that more techniques for age estimation of bloodstains at the scene will be developed.

9.11.3  Types of bloodstains

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There are several classification systems that exist in bloodstain pattern analysis of bloodstain patterns, and modifications of them appear in the literature. One simple classification is based on the deposition of blood on a surface by passive forces, applied forces, and direct contact (Figure 9.8). Bevel and Gardner (2008) state that all of the classification systems, regardless of their perspective, rely on a few basic mechanisms for their formation, which are shown in Box 9.4. There are numerous types of bloodstain patterns defined in textbooks and journal publications, and a selection of the most important types are described in the following text and summarized in Table 9.1. Several of the different bloodstain pattern types are present at the scene in Figure 9.9.

Gravitation

Bloodstain patterns

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Contact

Drip

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Smear

Drip trail

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Wipe

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Gush

Swipe

Splash

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Blood flow

Exhaled blood

Pool of blood Figure 9.8  A simple classification of different types of bloodstain patterns.

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drying of a bloodstain typically commences at the margins, and a skeletonization effect will appear upon wiping the stain implying that a ring will be seen on the outside boundary of the stain after 30–60 s (Figure 9.7). The ring will then gradually become thicker with time. An undisturbed stain of a single drop will become completely dry after 20–90 minutes, depending on the thickness and environmental factors. Extreme high and low temperatures will affect the drying time, but under normal indoor conditions, the impact of temperature is very small. By contrast, airflow across the drop has a profound effect, and this has to be taken into account particularly at outdoor crime scenes or when stains are found close to air-­conditioning outflow or fans. If the blood is diluted, usually with water, the drying time will be prolonged. Age estimation of bloodstains may be important to pinpoint a time frame for the infliction of injuries causing a bloodshed on the floor, walls, and furniture of a homicide scene. If a time of death estimation is performed, this time may be different from the time of the injuries, since there might be a survival time. General changes of a bloodstain after deposition include the color change from bright red to dark red, the clotting and drying phenomena, and later on deposition of dust. Recently, some developments have been done to allow for age estimation with Fourier transform infrared (FTIR) spectroscopy; Zhang et  al. (2017) reported repeatable spectral changes at defined times since deposition with such instrumentation looking at both rat blood and human blood. FTIR requires very small samples and today there are handheld instruments that actually can be used in the field. An even more exciting approach is imaging analysis that can be installed as an application on a smartphone. Hence, Shin et  al. (2017) published such a smartphone software for colorimetric analysis of bloodstains to estimate their age. In a subsequent article, the same group described six different stages of changes of a bloodstain that could be assessed: coagulation, gelation, edge desiccation, center desiccation, crack propagation, and final desiccation (Choi et al. 2019). To this end, the blood pool analysis could be used for the first three stages, the crack ratio analysis for the first six stages, and colorimetric analysis for all six stages. The authors also reported effects of different surface materials with low to high absorptive properties. Given the importance of

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Box 9.4  Basic mechanisms behind bloodstain patterns. x Blood dispersed from a point source by a force (e.g., impact patterns, expectorate) x Blood ejected over time from an object in motion (e.g., cast-­off patterns) x Blood ejected in volume under pressure (e.g., spurt and gush patterns) x Blood dispersed through the air as a function of gravity (e.g., drip patterns, drip trails) x Blood that accumulates or flows on a surface (e.g., pools, flows) x Blood that is deposited through transfer (e.g., smears, pattern transfers)

Figure 9.7  A wipe across bloodstains that have started to dry.

Source: Bevel and Gardner (2008).

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Table 9.1  Different types of bloodstains, their characteristics, and the underlying mechanisms of formation. Criteria

Underlying mechanism

Drip

• Rounded or elliptic (oval) shape • 4–25 mm in diameter • Clearly demarcated

Blood drips from a wounded person or a bloodied object without any other influence than gravity

Drip trail

• A series of adjacent drops forming a linear or nonlinear track • Leads from one position to another • Individual stains of similar size

Repeated dripping of blood from a person or an object moving from one position to another

Blood into blood

• The primary blood volume is larger than a drop • Satellite stains elliptic or rounded and show a random distribution of shape and directional angles

Blood drips into each other or into an existing volume of blood, or fluid. Fluid dripping into a volume of liquid blood can cause a similar pattern

Splash

• A group of stains radially distributed from a pool of blood • Gradual change of shape with distance from main stain • Usually similar size of stains

Small stains are formed radially around a pool of liquid blood impacted by an object or a part of a body (e.g., by stepping on a large volume of blood). A larger volume of blood falling down by gravity can also cause a splash

Blood flow

• The primary blood volume is usually larger than a drop • Usually about 5-­mm-­wide streams

Blood flows downward in streams (or occasionally in other directions by strong winds). Adjusts to the surface by spreading further

Pool of blood

• Primary blood volume larger than a drop • Adjusts passively to the shape of the surface without spreading further • Often well-­demarcated margins • Not absorbed/not completely absorbed by the surface

An accumulation of liquid blood that is adjusted to the underlying surface by gravity

Impact spatter

• A group of related small stains, usually 5 mm or less in diameter • Arranged radially from the source • Adjacent stains almost parallel • Gradual change in impact angle outward • Variable sizes of parent stains, but consistent throughout the pattern

A mass of blood is broken up at a point/area source by a force, for example, gunshot or blunt force, producing small, stable droplets than on impact results in small stains

Cast-­off stain

• • • •

Blood is released from an object or part of a body during a swinging movement

Gush

• Primary blood volume much larger than a drop • Adjacent spines and spatters radiating from the stain • Large elliptical spatter stains surrounding the stain

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A group of stains Linear distribution Adjacent stains almost parallel Gradual change in impact angle and/or direction

A large volume of blood is ejected and will be partly separated in the air and produce both large and small masses of blood. When impacting the target, secondary patterns are typically produced. Gush is usually considered to require a stronger initial force than splash

• A group of stains with a primary volume larger than a drop • Flows, spines, and satellite stains from individual stains • May form linear or wave-­like trails

Blood under pressure is ejected, for example, from a breached artery

Exhaled blood

• Group of stains of variable sizes • Possible dilution • Air bubbles and mucus strands

Blood introduced into the upper or lower airways, for example, due to facial or chest injuries is exhaled, often by coughing

Void

• An empty area within a bloodstain pattern where blood is expected

An object or a part of a body shields/covers a part of an area that is otherwise impacted by blood

Smear

• • • • •

Smears are formed by transfer of blood from one object to another without lateral movement. However, some experts consider wipes and swipes as subgroups of smears

Spurt

Transfer by direct contact Usually distinct borders May be a repeated contact Contains no streaks or featherings May show a pattern from the object

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Table 9.1  (Continued) Type of stain

Criteria

Underlying mechanism

Wipe

• Pre-­existing blood can be discerned • Displaced blood from the original boundary • Striations and feathered boundary, dried outer ring, and uneven thickness may be seen

An object or a part of a body comes in contact with a pre-­existing stain of liquid blood and displaces its blood laterally

Swipe

• • • • •

Caused by fluid blood that is transferred from a bloodied object or part of a body that moves across a surface area without pre-­existing blood

Saturation stain

• Usually well-­demarcated margins • Variable shape, determined by the surface topography • Is absorbed by porous material

Liquid blood accumulates in an absorbing material

Fly spots

• Very small, often rounded, sometimes elliptical spots • Often occur in groups and may form irregular lines • May be present behind shielding objects

Blood regurgitated or excreted by flies that settles down on walls or a ceiling

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Transfer by direct contact Decreasing blood volume at one end Usually diffuse borders Striations inside the stain No pre-­existing bloodstain can be detected

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Figure 9.9  A made-­up crime scene where the victim has been beaten in the head with a hard object, sustained repeated beating against the back of her head, and been stabbed in the neck. She has exhaled blood on the wall and plinth, and waved with her arms across the large pool of blood and also caused a smear on the plinth. The perpetrator has caused swipes on the drawers and pulled her back by holding her feet causing wide striations in the pool of blood.

Drips are produced by gravity acting on a mass of blood, for example, from a wound and usually range 10–20 mm in diameter, although they may be even as small as 3  mm and as large as 25 mm. If falling on a horizontal and smooth surface, they will typically be rounded with even boundary, and when present in a group, they are usually randomly distributed. The size will depend on the height as previously discussed, hence larger stains will represent the impact of drops that have fallen from some height and continuously accelerated during the course in the air. A small undisturbed drip stain is seen on the floor in Figure  9.10, just

next to two larger stains which have been altered and represent wipes. A drip trail indicates the movement of a wounded person or a bloodied object carried around. The trail may be straight or winding depending on the movement of the source of blood across the scene, but the starting and end points can sometimes be disclosed as well as the direction, for example, if the person has been moving quickly (see Figure 9.11). A blood into blood stain may be found at the end of a drip trail, for instance, because a wounded person walking away fell down over a couch or just stopped for some reason. When not

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Figure 9.10  An undisturbed seemingly completely dry stain and adjacent wipes in different directions across two larger stains which then still contained liquid blood.

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Figure 9.12  Blood into blood pattern.

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blood ejected, which upon impact can produce large primary stains from which secondary spines as well as radiating satellite stains are generated. Whenever there is an accumulation of blood, for example, at the surface of a wound, a point will be reached when gravity overcomes the surface tension of the mass of blood, and either drops or a flow of blood is formed depending on the presence of a surface for the blood to travel. The blood streams typically average about 5 mm and will follow a path that is directed by both the gravity and possible hindering structures on the surface. As can be seen in Figure 9.9, the smear on the plinth to the right, the flows are not completely perpendicular even though the surface is seemingly smooth. This is because the impact of gravity is not sufficient for the particular mass of blood to overcome its downward travel when meeting hindering small irregularities on the surface. Even individual drops may break up upon impact and result in flows as seen on the wall and the drawers in Figure 9.9. A pool of blood is commonly seen next to or under a part of a body lying on the floor of a crime scene. Such pools of blood are usually created by drops and flows of blood originating from one or more larger wounds. The shape of the pool of blood will be determined by the structure of the surface. Figure  9.9 shows a very large pool of blood which has first been disturbed by movements of the victim’s arms, and later by the action of the perpetrator when dragging the victim backward from the spot. Within undisturbed areas of the pool of blood, the distinct margins can be appreciated. Impact stain pattern is produced when a mass of blood is forced away from the body, for example, by blunt force (blows and kicks) or a gunshot, and then breaks up into stable droplets that initially have a medium or high velocity. This implies small droplet sizes, usually 2 mm or less, which when impacting a target will produce stains that are 5 mm or less in diameter. The stains will distribute radially from the source and hence impact

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Figure 9.11  Drops falling down from a wounded man while standing, walking fast, and running.

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moving, the drops from the wound will then fall into each other creating a volume of blood with increasing size. An example is shown in Figure 9.12. Even though the drops have not all fallen exactly in the center because of movement of the source of origin of the drops, the random distribution of satellite stains with variable sizes and shapes can be observed around the developing pool of blood. When such a pool of blood has been formed, a splash pattern may be created as a person steps on it (Figure 9.13). The spatter distributes radially from the area of impact in the direction of impact. The volumes of blood released from the pool of blood will vary and result in a mixed pattern composed of spines of different sizes and a spatter with highly variable sizes and shapes of the individual stains. Although not all of them will display a directionality, this becomes more evident at the periphery of the pattern. The variable pattern is explained by larger masses of

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Figure 9.14  A schematic of a convergence analysis.

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the target at variable angles. This means that the angle of impact of the individual stains always will change outward, but not necessarily at increasingly more acute angles, since the center of the pattern might not be at a right angle of the surface. The higher the velocity of the blood droplets upon impact, the more satellite stains will be produced. To the left in Figure 9.9, a medium-­ velocity impact pattern has been experimentally produced on the lower part of the wall and on the floor. Note the elongated shape of the individual parent stains and presence of minute satellite stains. High-­velocity impacts are typically produced by forward spatter from a gunshot wound, and sometimes the stains will be so small that a mist-­like pattern is produced (Figure 9.4). In order to determine the origin of an impact pattern, convergence analysis is used (see Figure 9.14). First, the direction of the individual stains is determined, and by grouping those with a similar shape along lines, the point of convergence can often fairly well be determined. By careful examination of an impact spatter pattern, several points of convergence may be identified (Figure 9.15). Figure 9.16 shows how such an analysis is performed in the field. Hence, such analysis can determine the minimum number of impacts. It should be pointed out that one single-­impact pattern considered to have been produced by beating with a hard object implies at least one impact, because the first blow will not cause a spatter even if it causes a large laceration of the skin. A complication to the convergence analysis is the influence of air resistance and gravity on the droplets traveling a longer distance through the air. However, again the careful analysis of the direction of the individual stains on the surface can often allow for a discrimination between possible flight paths and impossible ones (Figure 9.17). Cast-­off stains are produced when a bloodied object or a part of a body, such as a hand, is swung in the air, causing the release of blood during a forceful movement. This results in a group of stains forming a straight line (Figure 9.18). If the blood is distributed on different surfaces such as the ceiling, walls, and furniture,

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Figure 9.13  A schematic of a splash pattern when stepping hard on a pool of blood. Source: Illustration: Eszter Nagy, London.

Figure 9.15  A schematic of a convergence analysis resulting in two points of convergence.

it may not always immediately be appreciated that it is a linear distribution and that they all are coming from the same source. A cast-­off pattern is shown on the wall in Figure  9.9, going from

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Figure 9.16  Application of convergence analysis of a blood spatter on a wall.

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Figure 9.18  A schematic of the production of a cast-­off pattern as a blood-­bearing bat is swung in the air.

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Below solid line: Possible flight paths

produced by an excessive force acting on a large volume of blood, and splash when a mass of blood passively is falling down or ejected by a moderate force. Gush pattern will also show the mixed spatter pattern that is described for splash in the earlier text, but may display a distribution of stains further away when the velocity upon impact is substantially higher. When arteries are breached, blood may be ejected at a high speed from the wound, and this stream of droplets may produce an area of densely packed small stains if the victim is not moving, or a trail of small stains as the victim bends or moves around. Such patterns are called spurts. Due to the high speed of the droplets, these typically produce numerous spines and satellite stains. An artery that is completely cut off might not produce a spurt pattern because of a rapid retraction of the separated parts of the vessel, but rather result in an accumulation of blood which then may generate a gush pattern. Blunt-­force impacts to the face, and cut wounds or gunshot wounds to the face, neck, or the chest, may lead to significant bleedings to the nose, mouth, and airways. This blood may then, to a variable extent, mix with saliva and mucus of the lower airways, and upon forceful expiration an expectorate spatter pattern can be produced. This is characterized by a group of stains of variable sizes, and which may show signs of dilution. Mucus strands between individual stains are an even more convincing feature that allows for an identification of this type of spatter pattern. Three such patterns are seen at the lower part of the wall and on the plinth in Figure 9.9. Void implies an empty area next to a bloodstain pattern caused by the shielding of a person or an object at the time blood was traveling in the air. The object may be a door, and then it can be determined if the door was open, and at what angle, when, for example, a gunshot was fired. However, sometimes the shielding is from an object that has been removed from the scene, and then it might be a challenge to determine the object from the void area.

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Figure 9.17  Possible paths of blood drops impacting on a wall.

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right to left, slightly elevated. There will be a gradual change in the shape of the individual stains, as their angle will change along the line. This cast-­ off pattern was produced by swinging a ­bloodied knife. The rounded group of bloodstains to the right was caused by the initial release of blood drops from the knife, as it was quickly moved toward the wall immediately before the swinging movement and hence does not represent a cessation cast-­off pattern. This can promptly be excluded when examining the shape of the individual bloodstains along the line, and the position of their satellite stains. Kunz et  al. (2017) performed cast-­off experiments with a blood-­bearing object and stated that certain conclusions may be drawn about the intensity of the swing and the position of the person swinging the object, but pointed out that caution should be exercised regarding such interpretations, since several influencing factors may be difficult to evaluate in practice at a scene. Gush pattern and splash spatter are sometimes used interchangeably, but most experts favor the use of gush patterns when

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In a wide sense, a smear is any stain or pattern caused by the transfer of blood from one object onto another, implying a direct contact. Smears are characterized by a contiguous boundary and a diminished volume of blood across the body of the stain (as opposed to most other stains and in particular a pool of blood). A displacement of blood can be clearly observed. In Figure 9.9, an example of a smear is found on the plinth below the drawers to the right. The blood is deposited by contact of the forearm and hand of the victim. The female victim had fairly long hair, so we can exclude contact by her head, since there are no striations or feathering in the stain. Smears caused by a direct contact of a bloodied object without lateral movement may show the shape and pattern of the bloodied surface, for example, a hand or the outsole of a shoe, but in many cases the stain is unspecific. Furthermore, events during a violent crime often follow a rapid sequence that does not allow for the deposition of stains without some degree of lateral movement. The term wipe is defined as the disruption of a pre-­existing liquid/not completely dry bloodstain by a lateral movement. Depending on the surface of the touching object or part of body, the wipe can display striations of the body of the stain and feathering at the end. However, if the disturbance is caused by the contact by a very smooth surface acting with a gentle pressure, no markings may be seen. Figure  9.7 shows a wipe from recently deposited bloodstains that have skeletonized. If a bloodstain is disturbed even earlier after its deposition, it might not be possible to tell if the pattern is a wipe or a swipe. A swipe implies the transfer of blood from a bloodied object or a part of the body to another surface with no pre-­existing blood. Depending on the characteristics of the touching object, there might be striations in the body of the swipe. The direction of the swipe might not always be possible to determine, but feathering can be decisive as well as accumulation of blood at the point where the object separates from the surface. Examples of experimentally produced swipes are shown in Figure 9.19. A saturation stain implies a deposition of blood on an absorbent surface. The more absorbent material, the more of the blood mass will continue into it rather than sideways, and hence the margins will be well demarcated. Most physicians have been taught that external bloodshed may be overestimated and internal bleedings underestimated. This is true also at a crime scene; however, one common deposition site for blood are mattresses, which by saturation can absorb large volumes of blood even if the surface area is not so large, and remain an elusive explanation for the missing blood of victim considered to have died from a fatal blood loss. Flies and other insects at a scene may produce peculiar bloodstain patterns. Small, usually rounded spots of blood that are irregularly distributed on areas where a spatter from other common mechanisms is not expected can be explained by deposits of blood from flies which can produce them via regurgitation or excretion, and by transfer from their feet. Such depositions should be suspected, particularly if there are shielding objects in front of an area of bloodstain pattern consisting of very small individual stains.

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Figure 9.19  Swipes experimentally created by a bloodied piece of paper tissue (nos. 1, 6, and 10); a bloodied thumb (nos. 1–3); a piece of carpet (nos. 5 and 7); and two bloodied fingers (No. 8 and 9). The blue arrows indicate the direction and the yellow rings the points where the press is released and the bloodied object detaches from the surface. Note the accumulation of the blood at the point where the pressure is released, which occurs when there is still some blood left on the object. The striations from the rough surface of the paper tissue and carpet are conspicuous. The thinning is somewhat variable and will depend on the pressure, which often is not constant.

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9.11.4  Pattern analysis and conclusions In summary, the well-­ characterized properties and general behavior of blood upon passive and active movements allow for certain predictions of the spatter produced by different events, and, conversely, the prediction of the mechanisms behind a particular bloodstain pattern. Having said that, bloodstain pattern analysis involves several pitfalls, and the examination of various patterns is often thorough and time-­consuming. Even though this work is the responsibility of the crime scene investigators, many times the forensic pathologist with expertise in medical matters may contribute to the interpretation, particularly after having examined the injuries present on the dead body.

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Alkass, K., Saitoh, H., Buchholz, B.A. et al. (2013). Analysis of radiocarbon, stable isotopes and DNA in teeth to facilitate identification of unknown decedents. PLoS One 8 (7): e69597. Anderson, G.S. (2000). Minimum and maximum development rates of some forensically important Calliphoridae (Diptera). Journal of Forensic Science 45 (4): 824–832. Andersson, M.G., Ceciliason, A.S., Sandler, H. and Mostad, P. (2019). Application of the Bayesian framework for forensic interpretation to casework involving postmortem interval estimates of decomposed human remains. Forensic Science International 301: 402–414. Bevel, T. and Gardner, R.M. (ed.) (2008). Bloodstain Pattern Analysis with an Introduction to Crime Scene Reconstruction. Boca Raton, FL, USA: Taylor & Francis. Buschmann, C.T. and Tsokos, M. (2009). Frequent and rare complications of resuscitation attempts. Intensive Care Medicine 35 (3): 397–404. Byard, R.W., James, R.A. and Gilbert, J.D. (2002). Diagnostic problems associated with cadaveric trauma from animal activity. American Journal of Forensic Medicine and Pathology 23 (3): 238–244. Ceciliason, A.S., Andersson, M.G., Lindstrom, A. and Sandler, H. (2018). Quantifying human decomposition in an indoor setting and implications for postmortem interval estimation. Forensic Science International 283: 180–189. Choi, W., Shin, J., Hyun, K.A. et al. (2019). Highly sensitive and accurate estimation of bloodstain age using smartphone. Biosensors and Bioelectronics 130: 414–419. Comiskey, P.M., Yarin, A.L. and Attinger, D. (2019). Implications of two backward blood spatter models based on fluid dynamics for bloodstain pattern analysis. Forensic Science International 301: 299–305. Druid, H. (1997). Site of entrance wound and direction of bullet path in firearm fatalities as indicators of homicide versus suicide. Forensic Science International 88 (2): 147–162. Edelman, G.J. and Aalders, M.C. (2018). Photogrammetry using visible, infrared, hyperspectral and thermal imaging of crime scenes. Forensic Science International 292: 181–189. Edelman, G., van Leeuwen, T.G. and Aalders, M.C. (2012a). Hyperspectral imaging for the age estimation of bloodstains at the crime scene. Forensic Science International 223 (1–3): 72–77. Edelman, G.J., Gaston, E., van Leeuwen, T.G. et al. (2012b). Hyperspectral imaging for non-­contact analysis of forensic traces. Forensic Science International 223 (1–3): 28–39. Edelman, G.J., van Leeuwen, T.G. and Aalders, M.C. (2015). Visualization of latent bloodstains using visible reflectance hyperspectral imaging and chemometrics. Journal of Forensic Science 60 Suppl 1: S188–192. Fang, C., Liu, X., Zhao, J. et al. (2020). Age estimation using bloodstain miRNAs based on massive parallel sequencing and machine learning: A pilot study. Forensic Science International Genetics 47: 102300. Galloway, A., Birkby, W.H., Jones, A.M. et  al. (1989). Decay rates of human remains in an arid environment. Journal of Forensic Science 34 (3): 607–616. Gibson-­Daw, G., Albani, P., Gassmann, M. and McCord, B. (2017). Rapid microfluidic analysis of a Y-­STR multiplex for screening of forensic samples. Analytical and Bioanalytical Chemistry 409 (4): 939–947.

Gudmannsson, P., Berge, J., Druid, H. et al. (2018). A unique fatal moose attack mimicking homicide. Journal of Forensic Science 63 (2): 622–625. Hall, J.E. and Hall, M.E. (ed.) (2020). Guyton and Hall Textbook of Medical Physiology. Philadelphia, PA, Elsevier. Harvey, M.L., Gaudieri, S., Villet, M.H. and Dadour, I.R. (2008). A global study of forensically significant calliphorids: Implications for identification. Forensic Science International 177 (1): 66–76. Henssge, C. (1988). Death time estimation in case work. I. The rectal temperature time of death nomogram. Forensic Science International 38 (3–4): 209–236. Henssge, C., Madea, B. and Gallenkemper, E. (1988). Death time estimation in case work. II. Integration of different methods. Forensic Science International 39 (1): 77–87. Herr, D.M., Newton, N.C., Santrach, P.J. et al. (1995). Airborne and rescue point-­of-­care testing. American Journal of Clinical Pathology 104 (4 Suppl 1): S54–58. James, S.H., Kish, P.E. and Sutton, T.P. (ed.) (2005). Principles of Bloodstain Pattern Analysis. Theory and Practice. Boca Raton, FL, USA: Taylor & Francis. Johnson, M.D., Schaffner, W., Atkinson, J. and Pierce, M.A. (1997). Autopsy risk and acquisition of human immunodeficiency virus infection: A case report and reappraisal. Archives of Pathology & Laboratory Medicine 121 (1): 64–66. Karger, B., Billeb, E., Koops, E. and Brinkmann, B. (2002). Autopsy features relevant for discrimination between suicidal and homicidal gunshot injuries. International Journal of Legal Medicine 116 (5): 273–278. Kunz, S.N., Adamec, J. and Grove, C. (2017). Analyzing the dynamics and morphology of cast-­off pattern at different speed levels using high-­speed digital video imaging. Journal of Forensic Science 62 (2): 428–434. Kunz, S.N., Brandtner, H. and Meyer, H. (2013). Unusual blood spatter patterns on the firearm and hand: A backspatter analysis to reconstruct the position and orientation of a firearm. Forensic Science International 228 (1–3): e54–57. Kunz, S.N., Brandtner, H. and Meyer, H.J. (2015). Characteristics of backspatter on the firearm and shooting hand – an experimental analysis of close-­range gunshots. Journal of Forensic Science 60 (1): 166–170. Kunz, S.N., Klawonn, T. and Grove, C. (2014). [Possibilities and limitations of forensic bloodstain pattern analysis]. Wiener Medizinische Wochenschrift 164 (17–18): 358–362. Lin, H., Zhang, Y., Wang, Q. et al. (2017). Estimation of the age of human bloodstains under the simulated indoor and outdoor crime scene conditions by ATR-­FTIR spectroscopy. Scientific Reports 7 (1): 13254. Madea, B. (2016). Methods for determining time of death. Forensic Science, Medicine and Pathology 12 (4): 451–485. Madea, B., Krompecher, T., Knight, B. et  al. (2002). Estimation of the Time Since Death in the Early Postmortem Period, 2nd edn. Boca Raton, FL: CRC Press, Taylor & Francis. Megyesi, M.S., Nawrocki, S.P. and Haskell, N.H. (2005). Using accumulated degree-­days to estimate the postmortem interval from decomposed human remains. Journal of Forensic Science 50 (3): 618–626. Nikolic, S., Atanasijevic, T., Micic, J. et al. (2004). Amount of postmortem bleeding: an experimental autopsy study. American Journal of Forensic Medicine and Pathology 25 (1): 20–22.

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Ubelaker, D.H., Thomas, C. and Olson, J.E. (2015). The impact of age at death on the lag time of radiocarbon values in human bone. Forensic Science International 251: 56–60. van Daalen, M.A., de Kat, D.S., Oude Grotebevelsborg, B.F. et al (2017). An aquatic decomposition scoring method to potentially predict the postmortem submersion interval of bodies recovered from the North Sea. Journal of Forensic Science 62 (2): 369–373. Virkler, K. and Lednev, I.K. (2009). Analysis of body fluids for forensic purposes: from laboratory testing to non-­destructive rapid confirmatory identification at a crime scene. Forensic Science International 188 (1–3): 1–17. Wonder, A.K. (1985). What is blood? International Association of Bloodstain Pattern Analysts News 2(2). Zhang, Y., Wang, Q., Li, B. et  al. (2017). Changes in attenuated total reflection Fourier transform infrared spectra as blood dries out. Journal of Forensic Science 62 (3): 761–767. Zilg, B., Alkass, K., Berg, S. and Druid, H. (2009). Postmortem identification of hyperglycemia. Forensic Science International 185 (1–3): 89–95. Zilg, B., Bernard, S., Alkass, K. et al. (2015). A new model for the estimation of time of death from vitreous potassium levels corrected for age and temperature. Forensic Science International 254: 158–166.

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Ortmann, J., Markwerth, P. and Madea, B. (2016). Precision of estimating the time since death by vitreous potassium – comparison of 5 different equations. Forensic Science International 269: 1–7. Pizzola, P.A., Roth, S. and De Forest, P.R. (1986a). Blood droplet dynamics–I. Journal of Forensic Science 31 (1): 36–49. Pizzola, P.A., Roth, S. and De Forest, P.R. (1986b). Blood droplet dynamics–II. Journal of Forensic Science 31 (1): 50–64. Saferstein, R. (2009). Forensic Science: From the Crime Scene to the Crime Lab, 2nd edn. New York: Pearson Education (US). Saukko, P. and Knight, B. (2014). Knight´s Forensic Pathology. Boca Raton, FL: CRC Press, Taylor & Francis group. Shin, J.C.S., Yang, J.S., Song, J. et  al. (2017). Smart forensic phone: Colorimetric analysis of a bloodstain for age estimation using a smartphone. Sensors and Actuators B: Chemical 243: 221–225. Slaughter, L., Brown, C.R., Crowley, S. and Peck, R. (1997). Patterns of genital injury in female sexual assault victims. American Journal of Obstetrics and Gynecology 176(3): 609–616. Tsokos, M., Matschke, J., Gehl, A. et al. (1999). Skin and soft tissue artifacts due to postmortem damage caused by rodents. Forensic Science International 104 (1): 47–57.

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s­ uspicious death. This will guarantee the right of the deceased to have the truth told. It will also help in the punishment of perpetrators and hopefully in the prevention of further crimes. The medicolegal autopsy also has a research potential, as many cases of interest to society are examined by the forensic expert who will achieve an overview of a lot of case categories, such as sudden unexpected death, intoxications, road traffic accidents, child abuse, battered women, homicides, various types of accidents, occupational disease and work accidents among others. It is an obvious obligation for the forensic expert to use the knowledge to publicise the observations and thus inform society of possible preventive measures. Clinical autopsies have a multitude of inherent benefits. New diseases are discovered, the statistics concerning the cause of death and the spectre of disease will find a sound basis in the autopsy; clinicians will be able to adjust their diagnoses and treatment when they are faced with disease processes, and autopsies also give an opportunity for clinicians to develop new techniques such as angiography. The surgeon may practise operational skills, including trying new operations on corpses before live patients. The clinical autopsy will also give an opportunity to explore disease processes macroscopically and microscopically, and surgical pathology has developed a multitude of new methods, including histochemistry and molecular genetics. In some countries, the number of autopsies is decreasing, especially if the relatives are asked for permission. In such cases, the relatives will often fail to

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It is only quite recently that the autopsy has gained widespread use. The present technique was in many respects produced and developed in 19th-­century Germany. As the autopsy was from the beginning seen as a tool in the development and the investigation of disease processes, it took a turn towards the upholding of justice during the second half of the 20th century. The forensic autopsy is now known from the media as a tool to give justice to the individual. The Western idea of the importance of the rights of the individual is reflected in the fact that the number of medicolegal autopsies worldwide have increased tremendously during the last 50 years. Everybody is aware that if an autopsy has not been performed, the cause of death and manner of death are only established with much uncertainty. Forensic activity at an international level has proven the autopsy to be an efficient tool for the documentation of physical violence committed by individuals, gangs or even the authorities such as the military or the police. The result of an autopsy has a high level of truth, as it is a physical documentation not leaving too much room for individual interpretations. It is furthermore easy to document the lesions using photography, scanning, X-­ray or histology. The medicolegal autopsy is applied in even more countries as an indispensable examination in the case of

Handbook of Forensic Medicine, Volume 1, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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differentiate from the terms used for the autopsy. In cases that do not require the attention of the judicial system, it is performed by the attending physician or GP, while deaths that merit the closer attention of the police will usually be performed by specialists, such as the district medical officer, a forensic pathologist or in the Anglo-­Saxon countries and their former colonies by the ancient system of the coroner. Common to both is that the physician must not have any family or similar close relation to the deceased, since this might raise questions of impartiality. The demonstration of the signs of death is similar in the two different groups of deaths, but the question of the identity differs. When patients die in hospital or when the GP is called to the deathbed of a patient he or she has known for years, the identity of the deceased is seldom in question. The opposite is true when the district medical officer (DMO) or coroner is called to service. He or she cannot be expected to know the deceased, so in this case the responsibility for the identification of the body rests with the police. If they are in doubt, they may request the assistance of the forensic pathologist, odontologist and so on, and so identification becomes one of the reasons for performing the autopsy, in this case a forensic autopsy. The hospital physician or the GP will know the deceased well and there will be no reason to perform the meticulous external examination used in the case of suspicious deaths by the DMO or coroner. It suffices in most cases to demonstrate the signs of death such as rigor and/or lividity. Conversely, the DMO or coroner will have no knowledge at all of the deceased and must therefore devote the necessary time to study police reports, medical records and so on, before beginning the meticulous external examination

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see the importance for society and will feel a need to protect the deceased. This is an unfortunate development, and there is a need to promote understanding of the benefits of autopsies. In case of a suspected crime, the relatives usually do not have much possibility of hindering a medicolegal autopsy. Therefore, the medicolegal autopsies are stable or increasing in number in most countries. Autopsies may be described as either forensic or clinical autopsies, the latter also known as hospital autopsies. They may be very similar in principle, but the emphasis is different. The forensic autopsy must serve the needs of the judicial system with a special view to penal or insurance purposes, while the clinical autopsy aims at demonstrating the diagnosis and treatment offered to the patient, whether in hospital or in the clinic of the general practitioner (GP) (Figure 10.1.1). Before any autopsy, be it forensic or clinical, death must be established with certainty, and in most countries, a death certificate must be presented to the pathologist. The criteria for the determination of death are regulated by law and differ from country to country. In addition, most countries have different procedures to handle the deaths that may end up with either a forensic or clinical autopsy. Common for the two is that the identity of the deceased must be ascertained, if that is not indeed the purpose of the autopsy, and the cause of death and the manner of death must be established, again this may be the purpose of an autopsy. The examination of the deceased is variously described in different countries; in the Nordic and Germanic countries, the words ‘sighting of the body’ – ligsyn, Leichenschau – are used to

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Figure 10.1.1  The autopsy room showing the unitary table design combining evisceration and dissection areas, and spacious and abundant daylight from high windows (not shown).

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in Denmark, the Ministry of Justice issued a recommendation in 1995 concerning the performance of forensic autopsies, and this recommendation, written by a committee composed of the three professors of forensic medicine among others, is strictly adhered to (Ministry of Justice 1995). The description provided in the following text is to a large extent based on this work, with the addition of special procedures and the development of new techniques. The recommendations also contain a proposal for a unified autopsy report, and this is followed by the institutes as well, with the inevitable variation due to tradition, and a similar procedure is found in Sweden (National Board of Forensic Medicine 1994).

10.2.1  External examination

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The external examination is an important part of the medicolegal autopsy procedure. The findings are often crucial to the solution of the case and require great care, sense of detail and a thorough documentation. The forensic autopsy in Denmark is routinely preceded by postmortem computed tomography (PMCT), see Chapter 10.6.

Clothing

The clothes and their condition may yield important information that may explain what happened in the particular case. When handling an unidentified body, details of the clothes, size, colour, make and logos may contribute to the identification, although it will rarely be enough to make the final decision. Note must be made of the fit of the clothes, and special note must be made of well-­known irregularities of clothing, such as the open fly buttons in drowned sailors falling overboard while relieving themselves. In the case of violent death and when there is a suspicion of foul play, a detailed examination is necessary. This will often be supplemented by the examination in the police technical laboratory. In some cases, such as gunshot deaths, the finding of powder residue may decide the estimation of range, or in stab wounds, the clothes may give evidence that may lead to the correct answer, or when neglected, may unnecessarily lead the investigators down the wrong track. The clothing must be removed with care, avoiding contamination from the surroundings or by the investigators. It must be removed intact and if it has been torn by, for example, emergency room personnel during attempts at resuscitation, such tears must be recorded. All traces of foreign bodies, dirt, hairs and fibres, insects, blood or semen, particles of paint, tears or holes must be described in detail and photographed. Use of ultraviolet light may in some cases reveal invisible stains. There will usually be police technical staff at the scene, and they will, when necessary, collect evidence and make the photographic documentation. The corpse is photographed fully dressed and subsequently photographed as each piece of clothing is removed. Clothing, valuables and personal effects will undergo a more thorough examination in the police laboratory, but this

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of the body. The DMO will in particular look for signs of trauma or disease. An examination of the eyes, conjunctivae and eyelids, as well as the mucous membranes of the mouth, is obligatory to rule out strangulation. The complete body, front and back as well as all the natural orifices are inspected. The death certificate is an integral part of the examination, as it forms the legal documentation of death and may be used in a court of law. From the point of view of the police, the manner of death is by far the most important factor. If it is a natural death, the police will not care very much if the cause of death is pulmonary embolism or coronary thrombosis, no matter how relevant it may seem for the attending physician, unless there is a question of suspected medical malpractice. While the suspicion of homicide will most everywhere lead to a forensic autopsy, the practice in the other manners of death differs very much from country to country. In countries such as Denmark, the progression from inspection to autopsy is strictly governed by the police, so obvious suicides – which are of no interest to the police – will not lead to an autopsy. Similarly, the police will not request autopsy in cases of accidents where no one may be blamed, and there is thus no legal action needed. This is in contrast to the Nordic countries where both manners of death will require a forensic autopsy in nearly all cases. No hard and fast rule can be applied. Readers must investigate the rules of their country for specific information.

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A forensic autopsy is normally requested by the police, the coroner or in the Latin countries by that quaint institution ‘the investigating judge’. They may in some countries be requested by insurance companies as well. They are an important part of the police investigation of suspicious deaths. Such deaths must be reported by the physician attending the death according to the local rules and regulations. In Denmark, for example, all deaths that may be caused by a criminal act (homicide), accidents and suicides, people found dead unattended, sudden unexpected deaths, deaths due to occupational disease, deaths due to suspected medical malpractice and deaths in prisons and so on must be reported to the police. They will then decide if an examination by the DMO and the police is warranted, and will in return decide if a forensic autopsy is necessary. Apart from deaths due to occupational diseases, where the state occupational disease office requests autopsies, the police have the right of decision. The Danish Health Act requires a forensic autopsy if the death is or is suspected to be due to a criminal act, if the manner of death is not established by the DMO and police or if the cause of death is not established and the autopsy is deemed necessary from the judicial point of view. In some medical conditions, a forensic autopsy is mandatory, such as in drug addicts. A harmonisation of forensic autopsies in European countries has been proposed (Council of Europe 2000). The rules for ­forensic autopsies are governed differently in different countries:

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All lesions must be described carefully and in great detail. They must be photographed and/or sketched on a preprinted form, the latter being particularly useful in court where the photographs may be frightening to the layperson and yielding little information except to the experts. The site, size, direction and details concerning the edge and depth must be recorded. If there are multiple lesions, numbering will be useful, and it will be most convenient if the numbering in the text is the same as the numbering in the accompanying photographs. In case of gunshot wounds, it is important to describe changes around the skin lesion, particularly the gunshot residue, if any. When there is more than one wound channel, a probe can be used to illustrate the direction of the shot, but this procedure must never be used until the whole wound channel has been explored. Introducing a probe blindly into a wound will in many cases create a via falsa that may confuse more than it enlightens. It may be difficult to determine the direction of the shot in non-­ perforating gunshot wounds into body cavities such as the pleura or abdomen because of the short wound channel before the bullet disappears into the cavity. While it may be very difficult to establish the range from bullet wounds from pistols and rifles, it can be ascertained much more easily in shotgun wounds. The pathologist will provide precise data of the distribution of the pellets, and the police ballisticians will then be able to recreate the shot using the same weapon, the same ammunition and the same barrel in case of twin or triple-­barrelled shotguns. This is specialist work for the police, but the experienced pathologist may often be able to give an indication of range and type of weapon. Only by working closely with the police ballistic laboratory, will it be possible to contribute qualified information.

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The standard description of the corpse will always contain information of sex, estimated age if not known, height and weight and race or colour of the skin. Furthermore, the site of lividity, vibices and notable colour changes such as the cherry-­red lividity in carbon monoxide poisoning provides information. Postmortem rigor is verified by flexing and extending some of the larger joints. Some other postmortem changes may contribute to the establishment of the time of death, such as the discolouration of the skin, bullae with discoloured contents, a reticular pattern of cutaneous veins, greenish discolouration of the skin of the abdomen and so on. The body temperature will rarely be of any use for this purpose at the time of the autopsy, since the difference between the core temperature of the corpse and the ambient temperature will have disappeared in about 24 hours after death. With the exception of emergency homicide autopsies, we are rarely able to perform autopsies sooner than that.

External lesions

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earlier occasion. It is particularly important to note the position of the tracheal tube in case of death under anaesthesia, in the trachea or unintentionally in the oesophagus.

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must not tempt the pathologist to neglect his or her own appraisal of the material. If the clothing and effects are retained by the mortuary staff for later delivery to the relatives, these must be carefully documented, preferably with photos so that there will be no reason to doubt either the chain of custody or the honesty of the staff.

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Special identifying features

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Naevi, tattoos, scars, deformities, length and colour of the hair, colour of the eyes, dentures, contact lenses and so on may contribute to the identification of an unidentified corpse and must be carefully recorded. This information may also unexpectedly be of use in a later criminal investigation. When recording the identifying features of an unidentified body, particularly in a mass casualty situation, special forms are used, the best known being the Interpol Disaster Victim Identification Forms (Interpol 2013). The most important information collected in the autopsy room to be used in an identification situation is the dental status. While the pathologist and the autopsy room technician will assist in the registration of the dental status, the forensic odontologist will be called in to perform this procedure. In the case of hospital deaths or death after medical intervention, be it as a complication to a therapeutic or diagnostic intervention or after resuscitative measures have been unsuccessful, all medical equipment that has been used in the patient, such as catheters, intravenous drips, tracheal tubes, pacemakers and so on, must not be removed before the autopsy. This must be impressed on the hospital staff which will often make an effort to make the patient as presentable as possible before being shown to the relatives, but crucial evidence may otherwise be lost this way. There must be a detailed description of all medical equipment attached to the patient at autopsy, including the way it has been applied. Puncture wounds, bandages, skin damage and so on, which may have been caused by the medical intervention, must be distinguished from injury and manipulation performed at an

Conduct of the external examination The external examination should be conducted in a systematic fashion, starting with the hair of the head and ending with the soles of the feet. Lesions should be described with reference to relevant points of location such as the distance from the centre line of the body or the sole of the foot or the distance to well-­ defined anatomical landmarks such as the jugular incisure. Haemorrhage from the external orifices, the mouth, nose and ears, must be registered. The lips may be swollen and may contain mucosal lesions with haemorrhage which may be easily overlooked if one does not take particular care. Foreign bodies in the mouth, such as residue of edibles, may be a terminal phenomenon, but may also indicate aspiration. Puncture marks, be they the result of medical treatment or self-­inflicted, must be noted. Such marks are usually found in the regions of the elbows, hands or feet, but are also often found in the inguinal regions or on the side of the neck. If blood can be squeezed out from such a wound,

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The examination of the head will begin with an overview of the anatomy, noting any asymmetries of the bony structures, after which a careful study of the details of the head begins. Traditionally and logically, the hair or lack of hair is first described, then the length, curvature and the distribution, noting if there is balding, and if so, which type. The colour of the hair is a less reliable parameter, dying of the hair being the norm among women and now often seen among men as well. If a description is necessary for identification purposes, care must be taken to know the antemortem information of colour and length as recent as possible to minimise the risk of confusion. The hair must be examined for stains and foreign bodies, and blood spatter must be described with the hair in the position most likely at the time of staining. The hairy covering of the head, the beard, moustache or other facial hair may be quite distinctive, but is also subject to change in size, shape and colour. Having described the hair, the skin covered by hair and covering the skull must be searched for lesions and identifying features such as tumours and scars. It must be accepted that this may be difficult. Lesions with significant bleeding into the tissues may only be recognised when looking at the galea from the inside, and may then be identified in the skin. If there is any significant swelling such as a haematoma, the pre-­autopsy computed tomography (CT) scan of the head may give a clue to the origin. One should continue in a methodical fashion with the forehead, the cheeks and the skin covering the mandible as well as the skin of the front and back of the neck. Apart from the usual subjects for examination, one should note the presence or absence of petechial haemorrhages behind the ears. The eyes should be examined starting with the eyelids where petechiae are looked for on the outside as well as the inside of the eyelids. The upper eyelids should be double twitched to make sure that the entire mucous membrane is examined, at the same time looking for petechiae in the conjunctiva. The colour of the conjunctiva should be noted, icterus often being seen here.

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Beware of the possibility of an artificial eye which may look deceptively natural, but which will be revealed by touch. The nose is often the subject of blunt injury, the symmetry should be observed and palpatory fractures should be found at this time. For identification, any special features of the nose should be noted, size, colour, tumours and signs of disease. Piercing of the nose is common in teenagers or young adults. The mouth may be divided into the lips, the buccal cavities, the teeth and the oral cavity which may or may not be described with the internal organs, according to the routine of the institute. The lips are covered with skin and mucous membranes and are as prone to blunt injury as is the nose. Penetrating wounds of the lips from the teeth may be seen. The colour of the lips is of little value; they are much influenced by congestion and will in women often be covered by lipstick of the most variable colours. The inside of the lips should be checked for lesions and particularly the upper and the lower lip bands are subject to tears in blunt injury such as is seen in fist fights and child abuse. The teeth must be given a preliminary examination stating the number of teeth present, the state of the teeth and if major prosthetic work has been performed. A general description will suffice, any closer examination should be left to the forensic odontologist, and the wait is well worth it, considering the much better information that will result. The tongue is usually described with the internal organs, but since it is usually divided at the tongue band, it is wise to have a look during the external examination, since the evisceration may conceal evidence of injury.

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it is probably recent, but it is advisable to make an incision and dissect the vessel to be completely certain. Older puncture marks are often surrounded by a discolouration of the skin and a hardening, sometimes with distinct scar tissue. This is a common finding in drug addicts. Piercings are also found regularly among the younger population, particularly in the face, ears and nipples, but are also seen in the genitalia. The piercings may be useful when identifying a corpse with unknown identity. When examining the eyes, it is essential to confirm or exclude the presence of petechiae which may indicate strangulation in one of the forms this may take. The petechiae may vary in number and intensity, from few discrete ones to prominent haemorrhages that may be confluent at times. Petechiae must also be sought in the skin of the face, on the eyelids or in the mouth. A similar investigation must be performed on the neck to make sure that excoriations, nail marks or haemorrhages are not overlooked.

Neck Injury to the neck, particularly in connection with suffocation in its various forms, is so prominent in forensic pathology that particular attention must be paid to this region. The marks and injuries in suffocation and particularly in strangulation will be dealt with in another chapter, but since it is during the external examination that suspicion may arise of such a mechanism of death, they shall be described in general terms. Injury to the neck may leave finger marks, nail marks and strangulation marks. Finger marks are prominent in manual strangulation and will usually be placed on the sides of the neck, their number and distribution may give a clue to the position of the hands of the assailant and must therefore be described and photographed in detail. A drawing will be useful. Nail marks may originate from the victim as well as the assailant, the former while trying to ward off the attack. Nail marks from the victim are also seen in ligature strangulation when the victim tries to ease the pressure from the ligature. In cases of typical hanging, the suspension mark is located high on the neck, usually above the level of the larynx (‘Adam’s apple’), continuous in the front, but after creating a figure of an inverted V in the back of the neck, interrupted at the site of the noose. In atypical hanging, the V will be on the side of the head. There is typically only one linear mark in hanging; multiple marks must give rise to suspicion that what appears to be a

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noted. Hernias must also be looked for, since a strangulated hernia may cause ileus. If there is a suspicion of sexual assault, sampling from the vagina, rectum and mouth must be done at the earliest occasion, even when a special examination of the genital organs is planned (see the subsection ‘Sexual trauma’, under Section 10.2.3).

Back While the front of the torso is usually divided into thorax and abdomen, the back is more conveniently treated as a whole. When inspecting the back, evidence of congenital or acquired spinal malformations may be visible. The back will usually present the best view of the lividity, and its disposition must be noted, whether it is consistent with the history, or if the distribution could make one suspect foul play in having had the body moved after death. The history may justify a dissection of the skin of the back to search for haematomas, particularly in cases of suspected torture or other breaches of international humanitarian law.

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s­ uicidal hanging may actually be an attempt to cover up murder by ligature strangulation. The mark in a case of a typical hanging will have no or only minute vital reaction, since the circulation is effectively stopped immediately when the noose has been tightened. There are also no or very few petechial haemorrhages. In ligature strangulation, there is a much wider variation in the lesions. The ligatures may be of different thickness from the piano-­wire type in a torture scenario to the broad scarves used in impulsive murders in the family or among acquaintances. The mark may be single or quite often double or more when the assailant winds it several times around the neck. Since it is difficult to produce an instantaneous stop of the circulation, petechiae are prominent, and many petechiae in a case claimed to be suicidal hanging should cause suspicion of foul play. Evidence of treatment such as indwelling catheters in the cervical veins have hopefully been left in place by the nursing staff, and they should not be removed until the internal examination has verified that they were placed correctly.

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The extremities

The thorax, due to its size, is the site of numerous lesions in cases of blunt as well as sharp violence, including gunshot wounds, but it may also be the site of disease. The first consideration is the shape of the thorax, whether it is barrel shaped as in chronic obstructive lung disease or excavated as in congenital disease. The stability of the thoracic cage must be tested; considerable thoracic injury may be hidden under a seemingly normal surface. The skin covering the thorax may show evidence of disease in the shape of scars, pigmentation and cutaneous tumours. If fat embolism is suspected, the axilla is searched for petechiae. Self-­inflicted injury will often be seen on the thorax in addition to the arms and legs, typically within reach of the victims themselves. Pacemakers are usually implanted on the upper left quadrant. The removal must be noted in the autopsy report. The breasts must be routinely palpated for tumours and haematomas.

The extremities will often be the subject of trauma such as lesions contracted during a fight. Cuts to the forearms and hands are particularly important in knife/sharp violence, and the legs are the prime target for injuries in traffic accidents, such as the Messerer fracture from direct impact on the lower legs. The arms will often be the site of injection marks in drug addicts, and scars from self-­ inflicted incision wounds will be found particularly on the skin of the forearms, but also on the legs where they are within comfortable reach. Material from the nails is collected, either by cutting the nails or by scraping them with a wooden sampler, and the 10 samples are stored in individually marked containers.

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Like the thorax, the abdomen due to its size is often the site of the same kinds of injury as noted in the preceding text. In addition, the presence of effusions, such as ascites, may be suspected by the form and size of the abdomen, and preparations must be made to collect this upon opening. The skin of the abdomen may bear evidence of disease in the form of varicose veins and spider naevi in liver disease and striae distensae may suggest disturbances in metabolism such as use of anabolic steroids. Scars from operations, particularly such that lay before the introduction of laparoscopic operations, may be prominent and may give cause for ventral hernias.

External genitalia The external genitalia are searched for evidence of trauma. The urethra may be catheterised to make sure that urine is collected for analysis, and diseases such as hydrocele and varicocele testis are

10.2.2  Internal examination The complete forensic examination includes a full examination of all internal organs. This is necessary not only in order to locate possible lesions but also to establish macroscopically visible disease and secure specimens for a microscopic examination which may be of significant value for the evaluation of the cause of death. The body cavities are opened with one section covering the thoracic and abdominal cavity and one covering the head. While the body cavities are usually opened first, there are cases, such as in strangulation, when the opening of the head first may create a bloodless field. This is useful for the dissection of the neck structures with haemorrhages in the tissues such as the neck musculature and adjacent to the hyoid bone or thyroid horns, when broken.

Evisceration The body is opened by a section starting from below the ears and ending at the public symphysis, avoiding the umbilicus and whatever lesions and scars may be in the midline of the body (Figure 10.2.1).

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The incision may be continued on the side of the neck using one out of a number of techniques. Some continue the long incision on one side of the neck only, thereby making the suturing easier at the cost of some accessibility, some prefer a transverse incision along the upper margin of the clavicles to a point behind the ears. This gives superior accessibility to the neck organs, but will need more diligent and time-­consuming suturing to produce a satisfactory result from the point of view of the relatives. Having made the incision, the front of the thorax is dissected laying open the front of the thoracic cage and the sternum. This is when a test for the presence of pneumothorax may be done creating a pocket between the skin and the thoracic wall, filling it with water and when puncturing the pleura noting if air escapes. The abdomen is opened from the lower end of the sternum downwards, care being taken to collect fluid in the abdomen, if present. The dissection of the front of the thorax is now continued downwards so that there is access to the lower margin of the thoracic cage. The joints between the sternum and the clavicles are transected, and the ribs are cut with a special pair of scissors or using a knife that can cut the ribs, thus creating a triangular plate with the tip upwards. This plate is then dissected from the mediastinum beginning from below, taking care not to open the pericardium accidentally. The thoracic and abdominal organs are now in full view from the front and are inspected before the dissection continues, noting adhesions, malformations such as situs inversus and signs of injury. For ease of access, the duodenum is now located and a double ligature placed around it at the ligament of Treitz, thus preventing spillage of intestinal contents. The mesentery is cut all along the duodenum, jejunum and ileum, and the ileocaecal communication is freed up to the end of the ascending colon, the mesentery of the transverse colon is cut and the descending colon is freed from the posterior abdominal wall. The mesentery of the sigmoid colon is cut, and a double ligature placed as distally as possible,

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Figure 10.2.1  Y-­shaped incision for evisceration. Courtesy of T. Hansen.

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again to avoid spillage of colonic contents. The sigmoid colon and the rectum are divided, and the entire intestinal canal may be removed in toto and is examined externally and, if necessary, opened in its total length. The presence or absence of the appendix is noted as is Meckel’s diverticulum. One may now return to the organs of the neck. The skin of the neck is dissected from the organs of the neck up to the level of the lower margin of the mandible. The bottom of the oral cavity is opened by incising along the inside of the mandible from below, freeing the tongue together with the glottis and larynx in one block. Holding the tongue with one hand, the organs of the neck are dissected from the spine, and this is continued all the way to the level of the diaphragm. When transecting the vessels to the arms, blood from the upper extremities may flow from the cut surface, and this should be collected, anticipating problems of getting enough blood for analysis from vessels of the lower extremities. In case of adhesions between the pleural membranes, sharp dissection may be necessary, and care must be taken not to injure the lungs. The diaphragm is then divided along the thoracic wall and the kidneys may be freed from the posterior abdominal wall in the same procedure. The final procedure in the removal of the internal organs is the removal of the pelvic contents. The urinary bladder is freed from the pubic symphysis manually, taking care not to damage the bladder wall risking the loss of urine for analysis. The manual dissection is continued to the level of the prostate or uterine cervix and continued around the rectum. It is now possible to insert a knife between the symphysis and the bladder and transect the urethra and rectum or urethra and vagina, respectively. One may then continue the sharp dissection of the abdominal organs, in casu the abdominal aorta, all the way to the lower part of the pelvis, and all of the internal organs may be removed for dissection. Having opened the body cavities, the spinal column is split open with a chisel in order to visualise fractures or metastases to the bone marrow. To reduce the internal organs into a more manageable size, they are divided into three major blocks and the heart. The block is turned with the backside upwards, and the aorta is opened in its entirety from the arcus aortae to the pelvic bifurcature and the opening continued as far as possible. The renal arteries are opened, taking care to notice any stenosis that might explain arterial hypertension. The pathologist now has the choice of either dividing the aorta above the level of the intestinal arteries and freeing it up to the arcus or dissecting it in its entirety from the iliac arteries to the arcus in one piece. Both methods have their merits, and the authors (Knudsen and Thomsen) use both according to the situation. The diaphragm is then freed, and the suprarenal glands are removed. Many remove the spleen at this point, for convenience, being careful not to open the stomach at the same time, an obvious risk considering the anatomy. Having now easy access to the portal region from behind, it is also convenient to locate the portal vein at this time and make an incision to reveal portal thrombosis, if any. The kidneys may then be released bluntly from the back of the peritoneum and lifting them by the renal arteries and veins, the tissues connecting them may

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­ ulling the frontal lobes upwards and then using a scalpel cutting p the optic nerves, making it possible to make an incision in the tentorium along the edge of the os petrosa. It is then possible to lift the brain so far that the medulla oblongata may be divided at the level of the foramen magnum. The brain may then be removed, weighed and the dissection continued. Intracranial haematomas must be described as far as localisation, coagulation of the blood and the presence or absence of a limiting membrane are concerned, and the amount of blood must be measured, if at all possible. If there is no trauma that may explain the haematoma, the blood vessels must be inspected and carefully dissected with a view to congenital abnormalities or aneurysms. As a minimum, the circle of Willis must be isolated and all major branches identified, particular attention being paid to bifurcatures which are the most common sites for aneurysms. This must be performed in connection with the autopsy, since the fixation necessary for a later neuropathological examination of the brain will make it much more difficult to isolate the vessels. The source of a traumatic subarachnoid haemorrhage may be difficult to find at a routine autopsy, since it often originates from the vertebral arteries and requires a dissection of the cervical spine for a sufficient examination. Radiological examination with contrast injection in the vertebral arteries may be useful. In intracranial haematomas in children, particularly subdural haematomas, an examination of the eyes with a view to retinal haemorrhages, as seen in shaken baby syndrome, should be performed (Levin 2010). Having removed the major arteries of the circle of Willis, the brainstem is divided, creating a smaller block from the pons, medulla and cerebellum. The cerebellum is divided in the midline, opening the fourth ventricle. The cerebellar hemispheres are sliced from the sides, and finally a number of transverse cuts are made in the pons and medulla with a view to demonstrate haematomas or tumours. The brain is then cut in approximately 1 cm slices from the frontal lobes, care being taken to keep the slices absolutely parallel, an important point if a comparison between the two sides of the brain is to be meaningful. For special examination of the brain and the spinal cord, neuropathological expertise is required, and the examination should be performed in cooperation with a qualified neuropathologist. Problems often encountered are alcohol-­related changes in the central nervous system, age estimation of subdural haematomas, the relevance of degenerative changes, cerebral causes of sudden unexpected death, metabolic disturbances, infections, malformations and so on. It is the responsibility of the forensic pathologist to make the final evaluation of the findings and relate the changes in the central nervous system and pathological findings to whatever other findings, analyses and antemortem information are available.

TO

be cut, and the urogenital block may be divided from the area of the pancreas. The arteries to the intestines are cut in the same process, the abdominal aorta having been removed or not, see in the preceding text. The whole block is now turned with the neck towards the pathologist and the oesophagus opened from above to just above the stomach. The oesophagus is then cut loose just below the level of the laryngeal cartilage and dissected from the airways and folded downwards. Holding the neck organs in one hand, the thoracic organs are folded upwards so the heart is protruding. The pericardium is opened with an inverted Y-­shaped incision, taking care to collect effusions such as blood from a ruptured myocardium. A finger is then inserted in the sinus transversus and the aorta and the pulmonary artery cut as distally as possible. As the pulmonary artery is cut, any emboli must be observed and collected. The inferior and superior venae cavae are cut as distally as possible as are the four pulmonary veins. It is not acceptable to open the atria; all the vessels supplying them must be dissected individually. After the heart has been removed, the block containing neck, lungs and airways is separated from the abdominal organs, care being taken to ensure that the oesophagus is not cut, an accident that will often earn the well-­deserved scorn of the senior colleague supervising the procedure. The internal organs are now cut into manageable sizes, and the dissection of the remaining internal organs may commence.

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U

Head

FO

R

C

O

N

TR IB

The skin of the calvarium is divided by a section going from one ear to the other in the frontal plane. This will contribute to a stable head after the skull has been sawed off in a horizontal plane. Haemorrhage in the scalp is frequently seen after blunt injury to the head, be it intentional or after a fall following a stroke or similar. The extent and localisation must be noted and correlated to external injury. Haematomas and slight swelling may occur without external injuries and may be overlooked if only a limited examination is performed. Skull fracture is often seen after blunt injury, especially in traffic accidents, but also extensively in gunshot wounds, particularly with an entry wound in the mouth. It is important to pull off the dura completely to reveal fracture lines which may otherwise be hidden in the base of the skull as well as in the cupola. It may be indicated to open the sinuses and the medial ear in meningitis or septicaemia of unknown origin. In case of extensive skull fractures, such as after high-­velocity gunshot wounds, a better view of entry and exit wounds may be had by gluing together fragments to present the whole of the skull, but the use of three-­dimensional (3D) presentations after CT scans may be useful if the scalp has contained the fragments. In an intact skull, a comparison of the outer and inner diameters of the skull fractures may help to identify entry and exit wounds, the hole in the table first penetrated being smaller than the second one, as a rule. When the top of the skull has been removed and the brain is laid open, it is released from the base of the skull by carefully

Neck The organs of the neck are the first to receive attention when handling the block containing them, the lungs and the airways. The block is placed with the back upwards and the soft parts are

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FO

R

C

O

N

TR IB

U

LY

N

O

SE

TO

Before opening the chest, the presence or absence of a pneumothorax should be established. Radiological examination is to be preferred, but as a substitute or as a supplement the pleural cavities may be examined for pneumothorax. Haemorrhage and fractures of the thorax are particularly common in traffic accidents and falls from a great height, but may be a result of even minor trauma, especially in the elderly. The number and site of the fractures must be noted as well as the presence of blood or fluid in the pleural cavities. Fractures of the sternum and the ribs are common after external cardiac resuscitation attempts, but these usually do not show vital reaction, being postmortem phenomena. If the cardiac resuscitation is particularly effective, a blood pressure sufficient to create a seeming vital circulation may be successful, so this must be evaluated against the information from the medical report of the case. Small air bubbles may be found in the mediastinal adipose tissue after resuscitation, if large force has been used. This may be confused with subcutaneous emphysema which is felt crispy. Normally, the lungs will not cover the front of the mediastinum, but this may be the case in drowning where the lungs may be immersed in water, and pressure may leave impressions of the pathologist’s fingertips on the surface of the lungs. Pleural adhesions, emphysema, lacerations of the lung tissue, subpleural haemorrhage or petechiae must be noted, if present. The airways are opened from behind cutting through the muscular part of the trachea and the large bronchi as far as possible. If they are wide enough to allow opening with a medium-­sized pair of scissors until close to the lung surface, this is traditionally taken as an indication of bronchiectases, but this is very subjective. The airways must be searched for foreign bodies such as gastric contents, and, in cases of fire, soot must be looked for, since this is indicative that the deceased has breathed after the start of the fire. The pulmonary arteries must be opened as far as possible to look for pulmonary emboli, particularly in young women who have used oral contraceptives.

U

Chest

In penetrating injury to the lungs, it can be difficult to establish both the depth/length of the wound channel and its direction. An attempt to evaluate these factors must be made before removing the organs. It must be realised that the organs in the prone condition may be situated differently from the upright condition. The pericardium may contain fluid or blood, the latter in cardiac rupture after a myocardial infarction or after a puncture wound. A haemopericardium may be lethal if it exceeds 400 mL in adults. The heart is measured transversely along the sinus and from the coronaries to the apex. The coronary arteries are opened, taking care not to displace thrombi or more rarely emboli, or the technique using transverse sections in their full length may be used, according to local preferences. The heart is then sectioned in horizontal slices from the apex to the level approximately 1  cm below the tip of the major papillary muscles. The cavities are opened in the direction of the blood flow. The previous technique of connecting the superior and inferior venae cavae is now obsolete. Instead, the right atrium should be opened by cutting from the inferior vena cava into the tip of the auricle, leaving the ostium of the superior vena cava intact. This will display the inside of the atrium as well as the old method and has the advantage that sampling for examination of the sinus node is greatly facilitated. The ostium of the pulmonary artery is opened and the pulmonary valves inspected for number and pathology. The pulmonary veins are then opened and a cut made from the left atrium into the left ventricle as close to the septum as possible. The left ventricle is then cut open parallel to the left anterior descending coronary artery, taking care not to divide the left coronary artery in the bifurcature, but dividing the circumflex artery 0.5 cm distally to the bifurcature and then turning left into the aorta. Sections parallel to the surface may be made in the wall of the left ventricle and the septum according to local practice. Finally, the heart is weighed after having been emptied from coagula (Basso et al. 2017). In case of sudden unexpected death in children and young adults, the examination must include possible pathological processes in the cardiac conduction system. This is time-­consuming, and the sampling will result in a large number of sections which will require special stains and may present a considerable workload to the laboratory technicians as well as the forensic pathologists. The process is described in handbooks of cardiac pathology, and the reader is referred to those for particulars (Basso et  al. 2017; Gulino 2003). The macroscopic examination may reveal changes consistent with arrythmogenic right ventricular dysplasia (ARVD), now called arrythmogenic cardiomyopathy (ACM) where the wall of the right ventricle has been transformed to a thin fibrous and fatty membrane, but less pronounced cases may require extensive microscopic examination to demonstrate fibrous and fatty tissue in the myocardium. This diagnosis may be crucial to genetic counselling of the relatives and should not be neglected because of the work involved. Samples for genetic studies must also be collected, such as biopsies from the tendon of Achilles, as soon as possible after death, and a procedure for handling these samples must be made in advance. Copious sampling of myocardial tissue for microscopy and microbiological

R

­ issected from the laryngeal cartilages, exposing the upper and d lower horns of the thyroid cartilage, the thyrohyoid ligaments and the horns of the hyoid bone. In order to demonstrate the hyoid bone in its entirety, a large knife is inserted from behind, below the hyoid horns, but above the epiglottis, and cutting to the level of the body of the hyoid bone which will be elegantly displayed. It is necessary to perform a careful dissection of the tissues of the neck when there is a suspicion of strangulation or blunt injury to the neck, even if there are no external signs. Subcutaneous haemorrhages and haematomas in the neck muscles, haemorrhage in or fracture of the hyoid bone, thyroid cartilage or cricoid cartilage indicate trauma to the neck. Fractures are much more common in adults, the tissues being much more elastic in young people. Injury to cartilage may be a result of the evisceration, but the injury will then be without vital reaction such as haemorrhage. Venous haemorrhage behind the oesophagus may be seen as a postmortem artefact.

MEDICAL ASPECTS OF DEATH

LY

10.2.3  Special procedures

N

10.2.3.1 Trauma Neck

In case of trauma to the neck, particularly when strangulation in one of its forms is suspected, special attention must be paid to this region. Since evidence of haemorrhage in the organs of the neck may be blurred by postmortem bleeding due to evisceration, it is recommended that the brain be removed first. The organs of the neck are left in place when removing the thoracic organs by transecting them at the level of the clavicle. These two steps will empty the vessels of the neck, and any haematomas may be assumed to have been inflicted in vivo. The skin of the neck is then freed from the platysma and any lesions noted. Then the platysma and the layers of fat below it are removed, exposing the muscles at the front of the neck. These are then isolated one by one, beginning with the sternocleidomastoid muscles that are cut at their lower margin along the upper surface of the clavicle and folded upwards and backwards. It is now possible to view the passage of the large vessels of the neck, the carotids and the external jugular veins. These may now be isolated and examined for lesions from knife/ sharp violence to the neck. A probe should not be inserted until the vessels are laid free, but then it may be instructive for photographic documentation to use a probe of different colour from the surrounding organs to demonstrate the lesion. The muscles on the front of the neck connecting the base of the skull and the laryngeal skeleton with the upper thoracic aperture can now be isolated beginning with the muscles between the thyroid cartilage and the sternum and then the muscles from the base of the skull to the thyroid cartilage. Removing the muscles of the front of the laryngeal skeleton will also reveal the thyroid gland. Having laid free the neck from the front, the organs may be freed from the lower margin of the lower jaw in the manner described in the preceding text, and the organs of the neck may be removed en bloc after which the organs may be treated as described in the preceding text.

TR IB

U

TO

During the evisceration, fluid or blood in the abdominal cavity must be collected and measured, and sent for microbiological examination, particularly if there is a suspicion of peritonitis. Ascites is usually due to disease such as cirrhosis of the liver, cardiac failure or peritoneal carcinomatosis. Alternatively, haemorrhage is usually due to blunt or penetrating trauma. There will as a rule be external signs, such as haematoma, of trauma to the abdominal wall. The organs including omentum and the retroperitoneum must be inspected before the organs are removed for dissection. The origin of haemorrhage may be lesions of the liver or the spleen. Lesions of the liver may be due to attempts at resuscitation, but this will only rarely cause haemorrhage, and the lesions will be without vital reaction. Penetrating injury may involve several organs, and since the intestines are very mobile, lesions of the small intestine may be found at a distance from the lesion in the abdominal wall. The examination of the genitalia in cases of suspected sexual assault requires special care and must include a diligent search for haemorrhage, excoriations, superficial lesions of the mucous membranes and scar tissue in the vulva/vagina and anus. It is advisable to resect all of the perineum en bloc in order to facilitate a detailed study of the region. Photography and sampling must follow strict guidelines to ensure completeness.

O

Abdominal cavity

the actual identification is usually done by comparing radiological material such as bitewings or panoramic radiograph. In cases of fire, the dental examination will be the primary method and compared to DNA the fastest method of identification. The forensic odontologist may also be able to provide valuable information in bite marks. When only the skeleton is available for examination, anthropological expertise will often be necessary to establish sex, age, ancestry and so on of the victim, and to assist the forensic pathologist in the study of the bones.

SE

e­ xamination may also be necessary when there is a suspicion of myocarditis which may not be apparent at autopsy, but where the diagnosis rests on the histological examination or the culture of bacteria or virus.

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192

Skeleton and spine

FO

R

C

O

N

The study of fractures, including old healed fractures, is of particular interest in traffic accidents and in cases of suspected battered baby syndrome. It is easy to find fractures of the intervertebral discs and haemorrhage in the fractures with the naked eye as well as fractures of the extremities with visible angling. It may be necessary to perform a radiological examination, now commonly performed as a full-­body CT scan. This will provide excellent documentation for diagnosis as well as for use in court, should this turn out to be necessary. In gunshot lesions to the skeleton, fragmentation is most often the result, although a single hole may be seen, particularly in flat bones. The former lesion may lead to widespread injury which may be confused with lesions from other causes. In the skull, the entry wound will often have a typical cone shape with the greater diameter on the inside, while the exit wound will have the larger diameter on the outside. Both wounds will thus be cone-­shaped with the narrow part pointing towards the weapon. Dental investigation is performed by trained forensic odontologists, and such examinations are of particular value in identification. It is still the method of choice when good antemortem data are available as is the case in most Western countries. While the dental chart is useful for screening in cases with many ­victims,

Air embolism If air embolism is suspected, such as in diving deaths, the diagnosis may be made using a number of precautions. Since whole-­ body CT scan is now often the rule, this will help identifying air

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Sexual trauma

C

Newborn children

O

LY

N

O

N

TR IB

U

TO

In the case of sexual assaults with a fatal outcome, it may be necessary to perform a special examination of the genital organs, particularly in the female victim. The incision that has opened the abdomen is continued in two branches encircling the genital area including the anus. The anterior margin of the pelvis may be sawn through or may be dissected free of the external genitalia which may then be removed together with the internal genital organs and the rectum/anus. The female external genitalia are then examined from below, and thorough sampling is performed in addition to the sampling made during the external examination. The other genital organs are then examined as described in the preceding text.

SE

Fat embolism may be seen in a number of traumatic conditions, classically in fractures of the long bones, but is also described in trauma to the subcutaneous fat. It may be seen macroscopically as petechial haemorrhages in the brain (‘purpura cerebri’), but needs special techniques to demonstrate microscopically. The fat is soluble in the usual fixation procedures for microscopy, but if frozen sections are used they may be stained with special fat stains such as Oil Red O or Sudan Black. Lungs, kidneys and the brain are selected for this examination. In many cases, bone marrow embolism may be seen in routinely stained sections, especially in the lungs. It is not an uncommon finding after external cardiac massage, which will usually produce sternal and/or rib fractures, sending bone marrow particles into the circulation.

U

Fat embolism

descent or lack of descent of the testes in the male and the development of the large and small labia in the female. The hair and the fontanelles described as is the shape and size of the cranium. The child is measured according to the local procedures: the classic measurements of the cranium being the minimum. Special attention is paid to the development of the bony structures on the CT scan or on the whole-­body X-­ray which should always be performed in child deaths. The internal examination is mostly similar to that of the adult with the necessary variations due to the age of the child. The abdominal wall is opened in such a way that it will allow the study of the umbilical vessels, and their patency is noted. The oesophagus and the airways are examined with a view to potential broncho-­oesophageal fistulae. During the examination of the lungs, small samples are tried in water for their ability to float indicating that breathing has taken place. This is an unreliable test, however. The opening of the heart should take place with the heart in situ, beginning with investigation of the atria and the ventricles for communications between them. An incision is made in the right and left atria, respectively, and a probe introduced into the ventricles and in a retrograde fashion into the venae cavae and the pulmonary veins. Only when the pathologist has made sure that the cardiac vessels are normal or has made a note of variations is the heart removed, noting the precise number of vessels and their location. The heart should be removed taking into consideration the possibility of a persistent ductus arteriosus or a coarctation of the aorta which may be in close approximation of the site of the pulmonary artery and the aorta lying close together. Due to the partial development of the brain of the newborn, it should be removed with great caution, after removing of the cranial bones in pieces. If the cause of death is not obvious, the brain should be examined by a pathologist with special interest and qualifications in paediatrics and/or neuropathology.

R

in the vessels. If not, then conventional radiography should be used. The organ most likely to suffer is the heart, and therefore it must be opened under water, in order to identify and sample the air in the heart chambers. The heart may be opened using a syringe connected to a container with water for collecting the air for analysis.

FO

R

The investigation of death in the newborn follows the same main directions as described in the preceding text, but there are obviously a number of special considerations. In hospital cases, the child will usually be presented in hospital clothes, but in cases of children dying at home or found dead, the clothes must be examined as they are in adults. Special care must be directed at the possibility of strangulation, accidentally, for example, by the umbilical cord or pieces of clothing or intentionally in homicide. The external examination must include mention of stains or residue of vernix caseosa or meconium. The umbilicus is measured and the manner of division noted when it is not a hospital case, and the placenta must be examined, if present, for signs of acute or chronic placental insufficiency such as infarction or infection. The development of the child is judged by the length of the nails, the development of the nasal or aural cartilages, the

10.3  Clinical Autopsy Peter J. T. Knudsen and Jørgen L. Thomsen For many years, the tradition has been to perform an autopsy after death in hospital. The knowledge about the disease obtained in this way has been crucial to our understanding. Despite the enormous technical advances in our diagnostic proficiency, there are still a lot of cases in which the cause of death is not elucidated until an autopsy has been performed. In the Nordic countries as well as in most other Western countries, the autopsy rate has been declining in the last decades and is now below 10% in Denmark and Norway, a far cry from the 40% plus that was the rule a few years ago. This is unfortunate and will have long-­term negative influence on the reliability of the death statistics, the medical quality assurance programmes, training and research, postgraduate training and the legal rights of the individual. Among the causes of this development is a declining interest among ­clinicians

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willing to maintain the number of autopsies that could support valid death statistics. The exception to this rule is paediatric autopsies where parents, paediatricians and pathologists all agree that the autopsy is necessary to find a cause of death of the child and most importantly to prevent a recurrence, if possible, in the next pregnancy. The autopsy is an important part of medical quality assurance. In principle, all deaths should be followed by an autopsy – ‘the patient has a right to be autopsied if he dies in hospital’ – including deaths in the home, but there are a number of deaths where an autopsy is particularly relevant as follows: 1. Postoperative deaths. 2. Death after an unknown or undiagnosed disease. 3. Dementia in patients 75% with or without healed myocardial infarction

Minor anomalies of the coronary arteries from the aorta (RCA from the left sinus, LCA from the right without interarterial course, high take-­off from the tubular portion, LCx originating from the right sinus or RCA, coronary ostial plication, fibromuscular dysplasia, intramural small vessel disease)

Hemopericardium due to aortic or cardiac rupture

Anomalous origin of the LCA from the right sinus and interarterial course

Intramyocardial course of a coronary artery (myocardial bridge)

Mitral valve papillary muscle or chordae tendineae rupture with acute mitral valve incompetence and pulmonary edema

Cardiomyopathies (hypertrophic, arrhythmogenic right ventricular, dilated, others)

Focal myocarditis, hypertensive heart disease, idiopathic left ventricular hypertrophy

Acute coronary occlusion due to thrombosis, dissection, or embolism

Myxoid degeneration of the mitral valve with prolapse, with atrial dilatation or left ventricular hypertrophy and intact chordae

Myxoid degeneration of the mitral valve with prolapse, without atrial dilatation or left ventricular hypertrophy and intact chordae

Anomalous origin of the coronary artery from the pulmonary trunk

Aortic stenosis with left ventricular hypertrophy

Neoplasm/thrombus obstructing the valvular orifice

ECG-­documented ventricular pre-­excitation (Wolff–Parkinson–White syndrome, Lown–Ganong–Levine syndrome)

Atrial septum lipoma

Thrombotic block of the valvular prosthesis

ECG-­documented sinoatrial or AV block

AV node cystic tumor without ECG evidence of AV block, conducting system disease without ECG documentation

Laceration/dehiscence/poppet escape of the valvular prosthesis with acute valvular insufficiency

Congenital heart diseases, operated

R

U

N

O

SE

Congenital heart diseases, unoperated with or without Eisenmenger syndrome

U

TO

Dystrophic calcification of the membranous septum (±mitral annulus/aortic valve)

TR IB

Massive acute myocarditis

LY

Certain

N

AV: atrioventricular; ECG: electrocardiogram; LCA: left coronary artery; LCx: left circumflex branch; RCA: right coronary artery. Source: Data from Basso et al. (2008).

FO

R

C

O

Most cases of sudden death after the age of 35–40 are due to coronary arteriosclerosis. By contrast, most cases of sudden death before the age of 35 are due to coronary anomalies (anomalous origin and course of coronary arteries), ruptured vessels (aorta), cardiac hypertrophy, cardiomyopathies, or arrhythmias, the latter having a strong genetic background (structural cardiomyopathies such as hypertrophy, arrhythmogenic, dilated, restrictive, and noncompaction cardiomyopathy; channelopathies such as Brugada syndrome, long QT syndrome, short QT syndrome, and Wolff–Parkinson–White (WPW) syndrome (Figure  10.7.2). In a large molecular autopsy series of sudden unexplained death (n = 49), Tester et al. were able to show that more than one-­third of cases hosted a presumably pathogenic cardiac channel mutation (Tester and Ackermann 2006). In a cohort of sudden infant death syndrome (SIDS) in 5–10% of cases, mutations were found after postmortem cardiac channel genetic testing (Tester and Ackermann 2005). With mutation detection, screening the working group of Carracedo was also successful in identifying several mutations

both in adults and in children (Bríon et al. 2009; Rodríguez-­Calvo et al. 2008). Although the incidence of the SIDS declined in the last two decades, it is still the leading cause of death for infants aged  between 1  month and 1 year in developed countries (Krous  et  al. 2009). After the identification of avoidable risk factors such as the prone sleeping position, bottle-­feeding, smoking, and overheating, subsequent reduction-­of-­the-­risk campaigns have been initiated which led to a decline of the prevalence of SIDS. However, the pathophysiology is still unknown. Since hypertrophic cardiomyopathy (HCM) is one of the most prevalent causes of sudden cardiac death in younger adults (below the age of 35), it was hypothesized that HCM genes may be implicated in the development of sudden death in infants. The working group in Santiago de Compostela developed a MassARRAY genotyping platform analyzing more than 600 HCM patients. Meanwhile, 140 cases of SIDS have been studied by using SEQUENOM MassARRAY TM system and a total of 14 samples were detected as carrying genetic variants in

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Sudden cardiac death

Channelopathies

Long QT syndrome (LQTS)

Arrhythmogenic right ventricular dysplasia (ARVD)

Brugada syndrome (BrS)

N

Hypertrophic cardiomyopathy (HCM)

LY

Myocardial disease

SE

O

Catecholaminergic Polymorphic Ventricular Tachycardia (CPVT)

U

Short QT syndrome (SQTS)

TO

R

Figure 10.7.2  Genetic cause of sudden death. Source: According to Rodríguez-­Calvo et al. (2008).

The  mother was convicted for homicide of the fourth child; however, subsequent genetic testing revealed a previously not described mutation in the LQTS-­ associated KCNH2 gene. Although the functional significance of this particular mutation was not known (obviously expression analysis was not carried out), the jury found the finding sufficient to raise reasonable doubt to the question of guilt.

TR IB

U

four different genes (Bríon et al. 2009). Based on these preliminary results, HCM mutations may have a relation with SIDS; however, functional studies have to be carried out to get information on the functional significance of these mutations in structural normal hearts.

Evidential value of new mutations

FO

R

C

O

N

An interesting case concerning the evidential value of new mutations was reported by Rognum (Bjarkøy case) (see Madea et  al. 2010). In Norway, a mother lost her four infants ­subsequently between 1992 and 1997. In the cases of first two infants, the diagnosis of the cause of death was SIDS. In case of the third child, the cause of death was pneumonia due to aspiration of amniotic fluid. In the cases of first three children, no death scene investigation was carried out. After the fourth child had died, suspicion was raised that all deaths were probably due to Munchausen syndrome by proxy, since the mother had always been the only p ­ erson present. Many pathologists worldwide have seen similar cases with subsequent deaths of children. After the death of the fourth child, a plastic bag was found in the kitchen garbage with the lip mark and DNA from the baby and fingerprints of the mother. The mother admitted having had the bag over the babies’ heads, but only two days before death. Munchausen by proxy was suspected and the cause of death probably suffocation supported by the finding of   massive intra-­alveolar hemorrhage in all lung sections.

Diagnosis of infection Molecular pathological techniques are also of great importance in the diagnosis of infectious diseases and have been successfully applied also on very old tissue; for example, mummy tissue. In forensic medicine, these techniques have been applied for the diagnosis of, for example, myocarditis (Krous et al. 2009; Madea et al. 2010; Madea and Drexler 2011). Frequent causes of viral myocarditis are especially adenoviruses, enteroviruses, Epstein–Barr virus, influenza virus, parvovirus B19, and cytomegalovirus. Besides clinical history and clinical examination, the diagnosis of myocarditis is based on the investigation of myocardial tissue in living on endomyocardial biopsies. The endomyocardial biopsies are searched for viruses by PCR, reverse transcriptase-­polymerase chain reaction (RT-­PCR), and in situ hybridization; furthermore, immunohistological investigations are used. There is, however, no correlation between molecular pathological and immunohistochemical techniques; that is, positive virus findings may be seen in morphological normal hearts and vice versa

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(a)

CASE 1 - CHRONIC INTOXICATION CYP2C19

NORDOXEPIN

–DOXEPIN CYP2D6

CASE 1

POST MORTEM TOXICOLOGY –doxepin 2.4 mg/l (therapeutic 0.03-0.15 mg/l) –nordoxepin 2.9 mg/l –same yr: nordoxepin 0.2-1.4 mg/l in other autopsies

SE

•

O

N

- DOXEPIN INTOXICATION

U

•

INTERPRETATION: ACCIDENTAL, CHRONIC INTOXIATION

R

34 control cases MR 1.5 - 25 suicidal poisonings (n=23) MR > 3.8

U

TR IB

Case

Controls

Suicides

MR 1

2

3

4....

25

Figure 10.7.3  (a) and (b) Fatal doxepin intoxication, the manner of death remained unclear (suicide/accident). The postmortem DNA analysis revealed two CYP2D6 alleles, which were both not functional. Manner of death accidental. Source: From Koski  et al. (2007).

FO

R

C

O

N

MR = 0.83

• •

TO

There is no doubt that pharmacogenetics as a diagnostic tool has the potential to improve patient therapy, especially individualized drug therapy. Especially, genomic variance of phase I and phase II drug-­metabolizing enzymes plays an important role in an individual dose adjustment according to a patient’s genotype. Polymorphisms of the cytochrome P450 2D6  have great influence on the metabolism of many drugs, some being of great forensic relevance as the analgesics codeine, tramadol, hydrocodone, and oxycodone. Further candidate genes are, for example, μ-­opioid receptors. Meanwhile, a considerable amount on genetic variance and their impact on pain and analgesia has been published which is not only of great interest concerning individualized medication and side effects of drug therapy but may also be of forensic interest concerning the interpretation of serum levels. Molecular pathological methods are not only of help in determining the cause but also the manner of death, for example, in the differential diagnosis of accident/suicide or even accident/ homicide. This differential diagnosis may be of utmost importance also for the regulation of insurance compensations. Koski et  al. (2007) reported the case of a 43-­year-­old male alcoholic with suicidal tendencies who was found dead; the cause of death was doxepin intoxication; however, the manner of death remained unclear (suicide/accident). The postmortem DNA analysis revealed two CYP2D6 alleles, which were both not functional (CYP2D6*3/*4). Therefore, doxepin could only be metabolized to nordoxepin, but not to 2-­ hydroxydoxepin or 2-­hydroxynordoxepin. Correspondingly, high doxepin and especially high nordoxepin concentrations were found with a ratio of 0.83. In suicidal poisonings, the ratio is much higher; therefore, an accidental chronic intoxication could be assumed (Figure 10.7.3a,b). In another case reported by Koren et al. (2006), postmortem DNA analysis was important to exclude homicidal morphine intoxication (Figure 10.7.4a,b). Concerned was a full term healthy male infant with intermittent difficulties in breastfeeding and lethargy, starting on day 7. On day 12, it presented with gray skin and decreased milk intake;

2-HYDROXY-NORDOXEPIN

–2-HYDROXYDOXEPIN

(b)

10.7.3  Manner of death

CYP2D6

LY

(Madea 2009). Positive virus findings in the myocardium can of course explain the cause of death, especially if signs of myocarditis such as myocyte necrosis, myocytolysis, and interstitial lymphomonocytic infiltrates are visible. There are several case reports in the literature on positive myocardial virus findings in cases of sudden death. For SIDS, it was even claimed that positive virus findings (especially enteroviruses, but also adenoviruses or parvovirus B19) might be the cause of death in nearly 30% of cases. However, these findings could not be confirmed by succeeding investigations and may be due to methodical problems (Krous et  al. 2009; Madea 2009; Madea and Drexler 2011).

on day 13, it was found dead. The autopsy revealed no anatomical cause of death; however, a blood morphine level of 70 ng/ml was found. Neonates breastfed by mothers receiving codeine typically have morphine serum concentrations of 0–2.2 ng/ml. The mother received a combination preparation of codeine 30  mg and paracetamol, which she took for two weeks. She stored milk at day 10 which had a morphine concentration of 87 ng/ml (normally 1.9–20.5 ng/ml). Of course, the question was for the reason of the intoxication (accidental/homicidal). Genotype analysis revealed that the mother was an ultrarapid metabolizer (CYP2D6*2A/*2×2) consistent with an increased formation of morphine from codeine. Thus, cause of death was accidental morphine intoxication. Molecular biological methods are also of importance for the assessment of side effects during medical treatment. Different

PART II  

MEDICAL ASPECTS OF DEATH

CASE

(b) (a)

CASE

- MORPHINE POISONING IN A BREASFED NEONATE

- MORPHINE POISONING IN A BREASFED NEONATE

• Genetics • Mother: CYP2D6*2A / *2x2 (UM) • Grandfather, father, son: CYP2D6*1/*2 (EM)

• Autopsy –no anatomical anomalies –toxicology: blood morphine 70 ng/ml –mother received codeine 30 mg + paracetamol 500 mg preparate –due to episotomy pain x 2 / day, after two days half dosage (somnolence and constipation) –normally breastfed blood conc 0-2.2 ng/ml –mother stored milk on day 10 (conc. 87 ng/ml, normally 1.9-20.5 ng/ml)

EM EM Clinical picture and laboratory test are consistant with death due to morphine intoxication

UM Consistent with increased formation of morphine from Codeine and with somnolence and constipation

EM

LY

228

O

N

Figure 10.7.4  (a) and (b) Morphine poisoning in a breastfed neonate. Genotype analysis revealed that the mother was an ultrarapid metabolizer with an increased formation of morphine from codeine. Source: From Koren et al. (2006).

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10.7.4  RNA analysis

U

Identification of body fluids

R

Postmortem RNA profiling plays an important role in forensic medicine, for example, concerning human postmortem tissue identification or examination of gene expression levels at the time of death. The examination of gene expression at the time of death may be of great importance concerning the determination of cause and circumstances of death, survival time, ­pathophysiological conditions of disease and injury, duration of terminal episode, etc. (Ishida et  al. 2000, 2002; Bauer and Patzelt 2002; Bauer and Patzelt 2003; Bauer et al. 2003a, 2003b; Bauer 2007; Ikematsu et  al. 2007, 2008; Courts and Madea 2011c, 2012; Grabmüller, Madea and Courts 2015; Grabmüller et  al. 2015, 2017, 2018). Although RNA is known to be quite unstable, RNA could be extracted in adequate quality from tissue samples collected post mortem. However, a main point in the interpretation of gene expression levels regarding postmortem degradation is the standardization on the expression of endogenous control genes. RNA analysis offers insight into diseases and mechanisms leading to death and could develop into a valuable tool for diagnosis of the cause of death in forensic pathology. Other possible applications include the determination of the age of wounds and injuries and the postmortem interval (Bauer et al. 2003a, 2003b; Courts et al. 2015). The molecular identification of body fluids by analysis of cell-­specific mRNA expression already represents a new technique supplementing DNA analysis in forensic cases (Bauer and Patzelt 2002; Bauer 2007). The detection of epithelial cells in dry bloodstains by RT-­PCR is based on cell-­and tissue-­ specific gene expression. For instance, matrix metalloproteinase (MMP) mRNA could be detected in endometrium, but not in blood and other epithelia. This was confirmed in further studies

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CYP2D6 genotypes may determine in comparison to the wild-­ type (extensive metabolizer = EM) a missing (poor metabolizer = PM) deteriorated (intermediate metabolizer = IM) or an increased enzyme function (ultrarapid metabolizer = UM). This means that CYP2D6 genetic variations determine for instance plasma levels of tramadol and its metabolites, the O-­desmethyltramadol enantiomers. At the μ-­opiate receptor (MOR1), active M1 metabolite (+)O-­desmethyltramadol is of great importance for analgesia. If plasma concentrations of tramadol and M1 metabolites depend on the CYP2D6 genotypes, especially ultrarapid metabolizers may suffer from respiratory depression in postoperative analgesia (Musshoff et al. 2008, 2010; Stamer et al. 2007). Meanwhile algorithms have been developed in which cases of  pharmacogenomic investigations should be carried out (Figure 10.7.5). As always in forensic medicine, a close collaboration between pathologist, toxicologist, and geneticist is necessary for the interpretation of the results (Figure 10.7.6). In the literature, several cases have been described with a difficult differential diagnosis between homicide and poisoning and very high pethidine doses for palliative reasons. Daldrup and Lehmann (see Madea et  al. 2010) reported the case of a 88-­year-­old woman who died in the house of one of her physicians. She was treated with very high doses of pethidine (300 mg) in an infusion, and additionally with promethazine. She died within a short time after beginning of the infusion. Suspicion was raised, since the doctor presented a forged testament. In venous blood of the deceased, high concentrations of pethidine (6.4 mg/l) were found. In a first trial, the doctor was convicted of murder and sentenced to a long imprisonment (11 years). In a second trial, he pleaded to have administered a high dose of pethidine for pain relief and was acquitted. This case illustrates that in similar cases of claimed palliative care, genetic studies may be of importance too.

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Forensic pathology review

Comprehensive drug screening

A. Is there an elevated (toxic) drug concentration?

Covariables to consider

J. Was suicide the intent of the decedent?

Drug interactions (Micromedex)

D.

Drug metabolized by polymorphic enzyme

E.

Sample site (peripheral, heart, etc.)

F.

F. Fallani s intervals (I, II, III, IV)

G.

Case-Hx/death scene investigation

H.

Medical Hx (medications/drugs of abuse)

I.

Autopsy findings

J.

Intent (suicide)

N

LY

High metabolic ratio (acute vs. chronic)

C.

R

No

TO

Yes

B.

O

Covariables considered A-J

Toxic drug concentration

SE

Toxicology case review

No

A.

U

Yes

E. Was peripheral blood used?

U

Yes

TR IB

No

C/H. Are any other drugs detected?

F. Post-mortem redistribution may cause elevated concentration

No

O

N

Perform tissue levels and/or alternative blood sources

C

C. Drug-drug interaction may cause elevated concentration

D. Is the drug metabolized by a polymorphic enzyme?

R FO

Yes

No

Yes

Perform genotyping. May show genetic predisposition for toxicity Finalize death certification Figure 10.7.5  Milwaukee pharmacogenomic algorithm for forensic toxicology. Source: From Wong et al. (2006).

MEDICAL ASPECTS OF DEATH

Autopsy

Pathologists

Toxicologist Geneticist

Autopsy data

DNA Laboratory

Toxicology Laboratory

Parental drug

Toxicologist Geneticist Pathologist

Metabolite(s)

Genotyping

LY

PART II  

Pharmacogenetic interference

O

Forensic interpretation

N

230

SE

Relevance to death

Pathologists

degree of RNA fragmentation, which is significantly correlated with the postmortem interval. Overall, RNA degradation is a slow process under the conditions investigated experimentally, and significant differences, which might be used for determination of the post-­mortem interval (PMI), do not occur prior to 3–4 days post mortem. However, as with other laboratory methods for the determination of the time since death, rather large confidence intervals have to be considered so that the use of this method in forensic casework is limited (Bauer et  al. 2003a, 2003b).

C

O

N

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U

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with artificial menstrual bloodstains indicating that the detection of MMP expression in bloodstains may serve as a forensic marker for menstrual blood (Bauer and Patzelt 2002). Cell-­specific gene expression can not only be used to identify menstrual blood but the basic nuclear proteins protamines 1 and 2 are suitable markers for spermatozoa identification because they are exclusively expressed in the haploid genome. Protamine mRNA can be detected in semen stains by the highly sensitive RT-­PCR. With seminested PCR, 10–100 spermatozoa are theoretically sufficient to provide positive amplification results; with hot-­start PCR, at least 100–1000 cells are required corresponding to an average semen volume of 0.01–0.1 microliter (Bauer and Patzelt 2003).

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U

Figure 10.7.6  Collaboration between pathologist, toxicologist, and geneticist in pharmacogenetics. Source: From Sajantila et al. (2006).

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Age of bloodstains and postmortem interval

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In  vitro RNA degradation is a complex and nonlinear process which can serve as indicator for the quality and age of stains. Bauer et al. developed a semiquantitative duplex RT-­PCR assay, which in combination with competitive RT-­PCR using an external standard allows quantification of RNA degradation levels. Using this method, they investigated 106 bloodstains stored up to 15 years. The distribution of peak-­area quotients of standard and target messenger mRNA as measured by laser-­induced fluorescence capillary electrophoresis was closely correlated with the age of samples. Bloodstains with age difference of five years and more exhibit statistical significance variances in peak-­area quotients. Bauer et  al. were able to show that fatty acid synthase (FASN) multiplex PCR is a suitable method to quantify the

Wound-­age estimation Vital reactions are defined as local reaction of tissue at the site of damage. Typical vital reactions are, for example, an inflammatory reaction or expression patterns of adhesion molecules. Today, based on systematic postmortem quantitative analysis of mRNA transcripts, molecular biological methods are applied for wound-­age estimation. Animal experimental studies have already suggested time-­ dependent expressions of cytokine and chemokine mRNAs after skin injury. However, for the interpretation of the validity of these changes (besides the problems of quantitative analysis of mRNA transcripts), the manifestation time of these local vital reactions has to be considered in comparison to the duration of the agonal period. From many studies on immunohistochemical parameters of vitality, it is well known that there is a correlation between the manifestation time of various criteria of vitality and their

231

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Novel RNA technology involving real-­time RT-­PCR has provided a practical procedure for quantitative postmortem mRNA analysis. Postmortem mRNA quantification is an important tool in the study of the pathophysiology of diseases and traumas leading to death. Further applications are wound-­age estimation and wound-­healing processes, but also systemic responses to violence or environmental hazards. In opiate addiction, the MOR1 is the primary site of action for morphine in the most commonly used opioids. The MOR1 expression in animal and human brain has intensively been studied. The expression of the human MOR1 in postmortem human brain tissue was examined using real-­time PCR technology. Tissue samples from 11 fatalities due to opiate overdose and five normal subjects with different causes of death were analyzed in order to elucidate whether chronic opiate abuse is followed by a regulation of MOR1 expression. In each case, nine selected brain regions (thalamus, caudate nucleus, hypothalamus, ventral tegmental area, hippocampus, amygdala, frontal cortex, nucleus accumbens, and putamen) were evaluated. The MOR1-­mRNA level was determined relative to the housekeeping gene β2-­ microglobulin. While in most regions the MOR1-­mRNA levels in the brain of addicts were not different from the control group (with varying levels between 0 and 15% of housekeeping gene level), in the brains of three drug-­ related fatalities, an enormous increase was encountered in the thalamus where the MOR1-­mRNA level amounted for up to 10 000% of the measured housekeeping gene level. The results obtained by toxicological hair analysis in the group of drug-­ related fatalities indicate that the enormous thalamic MOR1-­ expression is primarily found in individuals who died from acute heroin overdose, but did not show signs of a substantial chronic administration of the drug. Further studies have to be performed to evaluate if the observed MOR1-­mRNA upregulation in the thalamus in a subpopulation of acute lethal intoxications mirrors a state of functional hypersensitivity associated with the occurrence of death (Becker et al. 2004).

U

MOR1 receptor mRNA expression in human brains of drug-­related fatalities

method and to detect and validate RNA signatures for different kinds of biological stains. However, mRNA stability and susceptibility to degradation have always been issues for mRNA-­based gene expression analysis and it was shown that impaired mRNA integrity, represented by the RNA integrity number (RIN), has an influence on the reproducibility of results by introducing a variable extent of bias. This is particularly intricate for forensic routine applications using mRNA, because biological stains from casework are often challenged by moisture, UV light, temperature, suboptimal environmental pH, etc., potentially degrading mRNA beyond usability. Although evidence concerning mRNA stability in forensic settings is controversial, there can be no doubt that single-­ stranded RNA transcripts of considerable length are less stable and more susceptible to degradation by physical and chemical strain and especially by ubiquitous RNAses than for instance a DNA molecule of comparable length. miRNA profiling has serious advantages as compared to mRNA profiling. First, due to their tiny size of about 22 nt, mature miRNAs are much more stable than mRNAs, which is of paramount importance in forensic settings, as it renders mature miRNAs decidedly less susceptible against fractionation by chemical or physical strain. This also applies for FFPE tissue samples, which can be of pivotal importance to forensic casework, but in which intense nucleic acid fractionation occurs. Not only has miRNA recovery from FFPE tissues been shown to be feasible and to lead to valid profiling results but it even outperforms FFPE tissue mRNA expression profiling by achieving a higher degree of resemblance to fresh tissue profiles and therefore significantly better correlation.

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validity as vital parameters. This means that the shorter the manifestation time, the lower the validity as vital parameter, since induction of supravital reactions seems to be possible even post mortem. All investigations on vitality and wound-­ age estimation based on molecular pathological methods have to keep this in mind.

10.7.5  Micro-­RNA Today, body-­fluid identification via specific mRNA quantification is an established standard technique in many forensic laboratories. Much work has been done to improve single aspects of this

10.7.6 Responsibilities of pathologists toward relatives of the deceased With the postmortem detection of mutations, the forensic pathologist also has medical responsibilities regarding a potential risk of family members of the deceased and an appropriate genetic counseling is necessary (Rodríguez-­Calvo et al. 2008; Kauferstein et al. 2009a; Oliva et al. 2010; Oliva and Pascali 2010; Robb 2010; Kauferstein et al. 2015). As far as with molecular autopsies a reasonable cause of death has been identified, the forensic pathologist also has the duty to inform the family of all autopsy results (Figure 10.7.7). Since in inherited conditions such as HCM or the long QT syndrome, other family members may also be at risk and may benefit from preventive strategies. Genetic counseling in sudden cardiac death is an integral part of the care provided to patients and their families and there should be a close cooperation between pathologist, medical geneticist, and cardiologist. The primary task of the forensic pathologist is to think of a genetic background, especially in cases of sudden unexpected death, and to reserve proper material for further genetic analysis.

232

PART II  

Forensic Autopsy SUD/SCD

Store DNA

10ml blood in EDTA or 1 cm3 fresh spleen of liver or 2 cm3 muscle or skin Refer to Cardiac Genetics Service (CGS)

Notify family members about possible hereditary nature of findings*

CGS

Genetic counsellor

Family history assessment

N

LY

Cardiac assessment

O

No abnormalities

SE

Hereditary cardiac disorder diagnosed

U

DISCHARGE**

CARDIOLOGY CLINIC

R

CGS

TO

Postautopsy care pathways service in cases of young or adult SUD or SCD where an inherited heart disease is suspected: *Add suitable statement to postmorten report: SUD:“Unexplained death may be caused by inherited cardiac disease. The deceased individual‘s relatives may therefore be at risk. Please refer the deceased individual‘s next of kin to Cardiac Genetics Service.“ or SCD: „Death has been caused by a cardiac disease which may have a genetic basis. The deceased individual‘s relatives may therefore also be at risk. Please refer the deceased individual‘s next of kin to Cardiac Genetics Service.“ ‚Ongoing surveillance seems indadvisable; there is a need for written information reporting the individual that he/she will need to be reassessed if he/she develops key symptoms, or if the family history changes

MEDICAL ASPECTS OF DEATH

Cardiac management

Cardiac management

U

Diagnostic genetic testing

ARRHYTHMIA SERVICE

TR IB

Figure 10.7.7  Flowchart of care of relatives of deceased in cases of young or adult sudden cardiac death with genetic background. Source: From Oliva and Pascali (2010).

N

References

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Ackermann, M.J., Tester, D.J., Porter, C.J., and Edwards, W.D. (1999). Molecular diagnosis of the inherited Long-­QT syndrome in a woman who died after near-­drowning. New England Journal of Medicine 341: 1121–1125. Baasner, A., Dettmeyer, R., Graebe, M. et al. (2003). PCR-­based diagnosis of enterovirus and parvovirus B19 in paraffin-­embedded heart tissue of children with suspected sudden infant death syndrome (SIDS). Laboratory Investigation 83: 1451–1455. Basso, C., Burke, M., Fornes, P. et  al. (2008). Guidelines for autopsy investigation of sudden cardiac death. Virchows Archiv 452: 11–18. Basso, C., Calabrese, F., Corrado, D., and Thiene, G. (2001). Postmortem diagnosis in sudden cardiac death victims: macroscopic, microscopic and molecular findings. Cardiovascular Research 50: 290–300. Bauer, M. (2007). RNA in forensic science. Forensic Science International: Genetics 1: 69–74. Bauer, M. and Patzelt, D. (2002). Evaluation of mRNA markers for the identification of menstrual blood. Journal of Forensic Sciences 47: 1278–1283.

Bauer, M. and Patzelt, D. (2003). Protatmine mRNA as molecular marker for spermatozoa in forensic stains. International Journal of Legal Medicine 117: 175–179. Bauer, M., Polzin, S., Gramlich, I., and Patzelt, D. (2003a). Quantification of mRNA degradation as possible indicator of postmortem interval – a pilot study. Legal Medicine 5: 220–227. Bauer, M., Polzin, S., and Patzelt, D. (2003b). Quantification of RNA degradation by semi-­quantitative duplex and competitive RT-­PCR: a possible indicator of the age of bloodstains? Forensic Science International 138: 94–103. Becker, J., Schmidt, P., Mußhoff, F. et al. (2004). MOR1 receptor mRNA expression in human brains of drug related fatalities – a real-­time PCR quantification. Forensic Science International 140: 13–20. Behr, E.R., Dalageorgou, C., Christiansen, M. et  al. (2008). Sudden arrhythmic death syndrome: familial evaluation identifies inheritable heart disease in the majority of families. European Heart Journal 29: 1670–1680. Blakey, G. and Farkass, D. (2009). General approach to molecular pathology. In: Cagle, P.T. and Alan, T.C. (eds.), Basic Concepts of Molecular Pathology. Molecular Pathology Library 2, pp. 61–68. Springer Science and Business Media.

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contrast” doped ballistic models. Forensic Science, Medicine and Pathology 11: 365–375. Grabmüller, M., Schyma, C., Madea, B. et  al. (2017). RNA/DNA co-­ analysis on aged bloodstains from adhesive tapes used for gunshot residue collection from hands. FSMP 13: 161–169. He, Y.J., Brockmöller, J., Schmidt, H. et  al. (2008). CYP2D6 ultrarapid metabolism and morphine/codeine ratios in blood: was it codeine or heroin? Journal of Analytical Toxicology 32: 178–182. Holmgren, P. and Ahlner, J. (2006). Pharmacogenomics for forensic toxicology: Swedish experience. In: S.H.Y. Wong, M.W. Linder, and R. Valdes (eds.), Pharmacogenomics and Proteomics. Enabling the Practice of Personalized Medicine, pp. 295–300. Washington DC: AACC Press. Hund, J.L. and Dacic, S. (2009). Applications in anatomic pathology. In: P.T. Cagle and T.C. Alan (eds.), Basic Concepts of Molecular Pathology. Molecular Pathology Library 2: 69–72. Springer Science and Business Media. Ikematsu, K., Takahashi, H., Kondo, T. et al. (2008). Temporal expression of immediate early gene mRNA during the supravital reaction in mouse brain and lung after mechanical asphyxiation. Forensic Science International 179: 152–156. Ikematsu, K., Tsuda, R., and Nakasono, I. (2006). Gene response of mouse skin to pressure injury in the neck region. Legal Medicine 8: 128–131. Ikematsu, K., Tsuda, R., Tsuruya, S., and Nakasono, I. (2007). Identification of novel genes expressed in hypoxic brain condition by fluorescence differential display. Forensic Science International 169: 168–172. Ingles, J. and Semsarian, C. (2007). Sudden cardiac death in the young: A clinical genetic approach. Internal Medicine Journal 37: 32–37. Ishida, K., Zhu, B.-­L., and Maeda, H. (2000). Novel approach to quantitative reverse transcription PCR assay of mRNA component in autopsy material using the TaqMan fluorogenic detection system: Dynamics of pulmonary surfactant apoprotein A. Forensic Science International 113: 127–131. Ishida, K., Zhu, B.-­L., and Maeda, H. (2002). A quantitative RT-­PCR assay of surfactant-­associated protein A1 and A 2 mRNA transcripts as a diagnostic tool for acute asphyxial death. Legal Medicine 4: 7–12. Jannetto, P.J., Wong, S.H., Gock, S.B. et al. (2002). Pharmacogenomics as molecular autopsy for postmortem forensic toxicology: Genotyping Cytochrome P450 2D6 for oxycodone cases. Journal of Analytical Toxicology 26: 438–447. Jeffreys, A.H., Wilson, V., and Thein, S.L. (1985). Hypervariable “minisatellite” regions in human DNA. Nature 314: 67–73. Jin, M., Gock, S.B., Jannetto, P.J. et  al. (2005). Pharmacogenomics as molecular autopsy for forensic toxicology: Genotyping cytochrome 450 3A4*1B and 3A5*3 for 25 Fentanyl Cases. Journal of Analytical Toxicology 29: 590–598. Junge, A., Dettmeyer, R., and Madea, B. (2008). Identification of biological samples in a case of contamination of a cytological slide preparation. Journal of Forensic Sciences 53 (3): 739–741. Junge, A., Steevens, M., and Madea, B. (2001). Successful DNA typing of an urine sample in a doping control case using human mitochondrial DNA analysis. Journal of Forensic Science 47: 5–13.

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Bríon, M., Allegue, C., Gil, R. et al. (2009). Involvement of hypertrophic cardiomyopathy genes in sudden infant death syndrome (SIDS). Forensic Science International: Genetic Supplement Series 2: 495–496. Brugada, R. (2010). Clinical Approach to Sudden Cardiac Death Syndrome. London, Dordrecht, Heidelberg, New York: Springer. Cagle, P.T. and Alan, T.C. (eds.) (2009). Basic Concepts of Molecular Pathology. Molecular Pathology Library 2, Springer Science and Business Media. Carturan, E., Tester, D.J., Brost, B.C. et al. (2008). Postmortem genetic testing for conventional autopsy-­negative sudden unexplained death. American Journal of Clinical Pathology 129: 391–397. Chugh, S.S., Senashova, O., Watts, A. et al. (2004). Postmortem molecular screening in unexplained sudden death. Journal of the American College of Cardiology 43: 1625–1629. Corrado, D., Basso, C., and Thiene, G. (2001). Sudden cardiac death in young people with apparently normal heart. Cardiovascular Research 50: 399–408. Courts, C., Grabmüller, M., and Madea, B. (2013a). Dysregulation of heart and brain specific micro-­RNA in sudden infant death syndrome. Forensic Science International 228: 70–74. Courts, C., Grabmüller, M., and Madea, B. (2013b). Functional single-­ nucleotide variant of HSPD1  in Sudden Infant Death Syndrome. Pediatric Research 74: 380–383. Courts, C., Grabmüller, M., and Madea, B. (2013c). Monoamine oxidase A gene polymorphism and the pathogenesis of sudden infant death syndrome. Journal of Pediatrics 163 (1): 89–93. Courts, C. and Madea, B. (2010). Genetics of the sudden infant death syndrome. Molecular Pathology in Forensic Medicine. Forensic Science International 203: 25–33. Courts, C. and Madea, B. (2011a). No association of IL-­10 promoter SNP  – 592 and -­1082 and SIDS. Forensic Science International 204: 179–181. Courts, C. and Madea, B. (2011b). Significant Association of TH01 Allele 9.3 and SIDS. Journal of Forensic Science 56 (2): 415–417. Courts, C. and Madea, B. (2011c). Specific Micro-­RNA Signatures for the Detection of Saliva and Blood in Forensic Body-­fluid Identification. Journal of Forensic Science 56 (6): 1464–1470. Courts, C. and Madea, B. (2012). Ribonukleinsäure. Bedeutung in der Forensischen Molekularbiologie. Rechtsmedizin 22: 135–144. Courts, C., Sauer, E., Hofmann, Y. et al. (2015). Assessment of STR typing success rate in soft tissue from putrefied bodies based on a quantitative grading system for putrefaction. Journal of Forensic Sciences 60: 1016–1021. Grabmüller, M., Courts, C., Madea, B. et  al. (2018). RNA/DNA co-­ analysis from bloodstains on aged polyvinyl-­alcohol gloves prepared for securing evidence from the hands of victims of fatal gunshot injuries. International Journal of Legal Medicine 132 (1): 53–66. Grabmüller, M., Madea, B., and Courts, C. (2015). Comparative evaluation of different extraction and quantification methods for forensic RNA analysis. Forensic Science International: Genetics 16: 195–202. Grabmüller, M., Schyma, C., Euteneuer, J. et  al. (2015). Simultaneous analysis of nuclear and mitochondrial DNA, mRNA and miRNA from backspatter from inside parts of firearms generated by shotsat “triple

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Oliva, A. and Pascali, V. (2010). Sudden cardiac death in forensic pathology. In: Brugada, R. (ed.), Clinical Approach to Sudden Cardiac Death Syndrome, pp. 91–110. London, Dordrecht, Heidelberg, New  York: Springer. Reibe, S., Schmidt, J., and Madea, B. (2009). Molecular identification of forensically important blow flight species (dipthera: calliphoridae) from Germany. Parasitology Research, online first, 28. October 2009. Richter, N. (1903). Gerichtsärztliche Diagnostik und Technik. Leipzig: Hirzel Verlag. Robb, L. (2010). Genetic counselling in cardiovascular conditions. In: R. Brugada (ed.), Clinical Approach to Sudden Cardiac Death Syndrome, pp. 327–335. London, Dordrecht, Heidelberg, New York: Springer. Rodríguez-­Calvo, M., Brion, M., Aleegue, C. et  al. (2008). Molecular genetics of sudden cardiac death. Forensic Science International 182: 1–12. Sajantila, A., Lunetta, P., and Öjanperä, I. (2006). Postmortem pharmacogenetics: toward molecular autopsies. In: S.H.Y. Wong, M.W. Linder, R. Valdes (eds.), Pharmacogenomics and Proteomics. Enabling the Practice of Personalized Medicine, pp. 301–310. Washington DC: AACC Press. Sauer, E., Babion, I., Madea, B., and Courts, C. (2015). An evidence based strategy for normalization of quantitative PCR data from miRNA expression analysis in forensic organ tissue identification. Forensic Science International: Genetics 13: 217–223. Sauer, E., Madea, B., and Courts, C. (2013). An evidence based strategy for normalization of quantitative PCR data, from miRNA expression analysis in forensically relevant body fluids. Forensic Science International: Genetics 11: 174–181. Shojania, K.G., Burton, E.C., McDonald, K.M., and Goldman, L. (2003). Changes in rates of autopsy detected diagnostic errors over time. A systematic review. JAMA 289: 2849–2856. Smith-­Zagone, M.J., Pulliam, J.F., and Farkass, D.H. (2009). Molecular pathology methods. In: D.G.B. Leonard (ed.), Molecular Pathology in Clinical Practice: Genetics, pp. 15–39. Stamer, U.M., Mußhoff, F., Kobilay, M. et al. (2007). Concentrations of tramadol and o-­desmethyltramadol-­enantiomers in different CYP2D6 genotypes. Clinical Pharmacology Therapeutics 82 (1): 41–47. Tester, D.J. and Ackermann, M.J. (2005). Sudden infant death syndrome: How significant are the cardiac channelopathies? Cardiovascular Research 37: 388–396. Tester, D.J. and Ackermann, M.J. (2006) The role of molecular autopsy in unexplained sudden cardiac death. Current Opinion in Cardiology 21: 166–172. Tester, D.J. and Ackermann, M.J. (2007). Postmortem long QT syndrome genetic testing for sudden unexplained death in the young. Journal of the American College of Cardiology 49 (2): 240–246. Wong, S.H.Y., Gock, S.B., Run-­Zhang, S. et al. (2006). Pharmacogenomics as an aspect of molecular autopsy for forensic pathology/toxicology. In: S.H.Y. Wong, M.W. Linder, R. Valdes (eds.), Pharmacogenomics and Proteomics. Enabling the Practice of Personalized Medicine, pp. 311–318. Washington DC: AACC Press. Wong, S.H.Y., Linder, M.W., and Valdes, R. (2006). Pharmacogenomics and Proteomics. Enabling the Practice of Personalized Medicine. Washington DC: AACC Press.

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Kauferstein, S., Kiehne, N., Neumann, T. et  al. (2009a). Plötzlicher Herztod bei jungen Menschen durch kardiale Gendefekte. Deutsches Ärzteblatt 106 (4): 41–47. Kauferstein, S., Kiehne, N., Neumann, T. et  al. (2009b). Cardiac gene defects can cause sudden cardiac death in young people. Deutsches Ärzteblatt International 106 (4): 41–47. Kauferstein, S., Madea, B., and Courts, C. (2015). Forensische Molekularpathologie. CME Beitrag. Rechtsmedizin 24 (6): 513–532. Koren, G., Cairns, J., Chitayat, D. et al. (2006). Pharmacogenetics of morprescribed phine poisoning in a breastfed neonate of a codeine-­ mother. Lancet 368: 704. Koski, A., Ojanperä, I., Sistonen, J. et al. (2007). A fatal doxepin poisoning associated with a defective CYP2D6 genotype. American Journal of Forensic Medicine and Pathology 28: 259–261. Koski, A., Sistonen, J., Ojanperä, I. et al. (2006). CYP2D6 and CYP2C19 genotypes and amitriptyline metabolite ratios in a series of medicolegal autopsies. Forensic Science International 158: 177–183. Krous, H.F., Ferandos, C., Masoumi, H. et al. (2009). Myocardial inflammation, cellular death, and viral detection in sudden infant death caused by SIDS, suffocation or myocarditis. Pediatric Research 66 (1): 17–21 Levo, A., Koski, A., Ojanperä, I. et al. (2003). Post-­mortem SNP analysis of CYP2D6 gene reveals correlation between genotype and opioid drug (tramadol) metabolite ratios in blood. Forensic Science International 135: 9–15. Madea, B. (2009). Sudden death, especially in infancy – improvement of diagnosis by biochemistry, immunohistochemistry and molecular pathology. Legal Medicine 11: S36–S42. Madea, B. and Drexler, J. (2011). Virusätiologie des Sudden Infant Death Syndrome. In: G. Darai, M. Handermann, H.G. Sonntag C.A. Tidona, L. Zöller (Hrsg.), Lexikon der Infektionskrankheiten des Menschen – Erreger, Symptome, Diagnose, Therapie und Prophylaxe, 4. Auflage S. 931–962. Berlin, Heidelberg, New York: Springer-­Verlag. Madea, B. Saukko, P., Oliva, A., and Musshoff, F. (2010). Molecular pathology in forensic medicine. Forensic Science International 203: 3–14. Mehling, L.M., Piper, T., Lott, S. et al. (2017). Several possible toxicological and genetic tools for the extension of the detection window after GHB intake. Toxichem Krimtech 41, Band 84, Heft 3, 184–190. Mehling, L.M., Piper, T., Spottke, A. et al. (2017). GHB-­O-­ß-­glucoronide in blood and urine is not a suitable tool for the extension of the detection window after GHB intake. Forensic Toxicology 35 (2): 263–274. Musshoff, F., Madea, B., Stüber, F., and Stamer, U. (2008). Plasma levels of tramadol and o-­desmethyltramadol-­enantiomers in patients with different CYP2D6 genotypes. In: F. Pragstand R. Aderjan, GTFCh Symposium 2007, Aktuelle Beiträge zur forensischen und klinischen Toxikologie. Fahreignung – K.-­o.-­Mittel – Toxikokinetic – Analytische Methoden. Bad Vilbl 156–161. Musshoff, F., Stamer, U., and Madea, B. (2010). Pharmacogenetics and forensic toxicology. Molecular Pathology in Forensic Medicine. Forensic Science International 203: 53–62. Oliva, A., Brugada, R., D’Aloja, E. et al. (2010). State of the art in forensic investigation of sudden cardiac death (SCD). American Journal of Forensic Medicine and Pathology.

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Forensic microbiology is broadly defined as the discipline of applying scientific methods to the analysis of microbiological evidence in criminal and civil cases for investigative purposes (Schmedes et al. 2016). In the context of autopsy and determining the cause of death in which infectious disease may have been a factor, forensic microbiology can be important for identifying causative agents. There is a long history of forensic microbiology used by pathologists, well before there was a formalized forensic microbiology discipline. Louis Pasteur and Robert Koch, using an artificial culture medium, invented bacterial culturing. Subsequently, culture services became available in the 1890s (Uruburu 2003; Boundy-­Mills et  al. 2015; Casaregola et al. 2016; Bonnet et al. 2020), and the value of postmortem investigation of microorganisms was evaluated as early as in 1895, when Achard and Phulpin worked on the origins of the microorganisms found in dead bodies (Achard and Phulpin 1895). Thereafter, the general value, sampling techniques, antemortem–postmortem comparisons, and related interpretation issues of bacterial cultures have been pursued and debated from early 1900s until recently (Gradwohl 1904; Fredette 1916; Giordano and Barnes 1922; Burn 1933; Kurtin 1958; Carpente and Wilkins 1964; O’Toole et al. 1965; Wood et al. 1965; Roberts 1969; Koneman and Davis 1974; MacDonald 2010). The basic concepts and challenges of postmortem microbiology were delineated in such studies. In the last decade, new technological innovations have laid the ground for more detailed understanding of the human microbiome in general (Turnbaugh et al. 2007; Methé et al. 2012) and consequently enhanced the capabilities of the field of forensic microbiology (Giampaoli et  al. 2012; Metcalf et al. 2013, 2016; Yasuhiro et al. 2015; Schmedes et al. 2016; Javan et al. 2016; Blondeau et al. 2019) (Figure 10.8.1). Early forensic microbiology work focused on how bacterial cultures can aid in determining the cause of death or specifying the etiologic agent behind the infectious death in a postmortem investigation. These studies addressed several pressing issues. The microbial differences between the samples taken during hospital treatment and those taken at the autopsy were of keen interest (Wood et al. 1965). The need for multiorgan sampling during the autopsy was considered (Kurtin 1958). Also, the possibility of

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false positive results or sample contamination, and the importance of correct sampling techniques to avoid these issues were studied (O’Toole et al. 1965). Forensic microbiology is now in a new era, mostly due to modern laboratory techniques developed in the past decade, but also due to the enhanced understanding of the human body. Humans are composed not only of their own (human) cells, but an estimated 10× more of our cells that make up the human body ecosystem are of microbial origin. Since 2005 (Margulies et al. 2005; Shendure et al. 2005), high-­ throughput DNA-­sequencing methods, also known as massively parallel sequencing or next-­generation sequencing (NGS), and bioinformatics advances allow the characterization of microorganisms for a variety of forensic applications. Behind these capabilities are two major discoveries in basic biology and physiology. First, the sequencing of the entire human genome, which was a massive undertaking simultaneously achieved by the public funded Human Genome Project (HGP) (https://www.genome. gov/) and by a privately funded group. This project disclosed that human cellular and organ functions are encoded by approximately 20 000 protein genes. Equally as important was the finding that humans, during and after birth, quickly gain over a million additional genes via the establishment of their microbiomes, which host at least the same amount, if not an order of magnitude greater number of cells than the human cells of a body. The data show that a vast majority of commensal bacteria reside in the human colon with an estimate of 1014 bacteria, followed by the skin with an estimate of 1012 bacteria, and the rest of the body being populated by less than 1012 bacteria. These data stem from the ongoing Human Microbiome Project (HMP) (Turnbaugh et  al. 2007) (https:// www.hmpdacc.org/), which was based originally on the medical community’s interest in the microbes associated with the human body and the important roles they play in health and disease. Within the HMP, the collection and characterization of the diversity and function of the human microbiome have constantly increased (Huttenhower et al. 2012; Methé et al. 2012). Due to the developments described in the preceding text, modern forensic microbiology has also gained prominence and is now a wider discipline with an active academic research component and routine applications for analysis of microbial evidence at a depth and resolution that some time ago was unthinkable. The application areas in forensic microbiology vary and include determining cause of death, sexual assault investigations, predicting geographical origin of unidentified bodies, and tracking of pathogens in suspected bioterrorism and bio crimes, to name a few applications (Table 10.8.1).

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1895 Archard & Phulpin 1st postmortem microbiology

1860 Louis Pasteur 1st liquid medium bacterial culture

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1916 Fredette Concept of agonal spread

1904 Gradwohl Concept of postmortem transmigration

1965 O’Toole et al. Concept of sterile autopsy 1964 Carpenter & Wilkins 1st review of postmortem Microbiology in large cohort (> 2000 cases)

1994 First use of HIV sequence analysis in a rape case

1974 Koneman et al. Clinical importance of postmortem microbiology

2001 Anthrax letters in USA 2001 Human genome sequence

1998 development of MLST

Figure 10.8.1  Important milestones in forensic microbiology from 1860s to 2020.

2008 Human microbiome project

2016 Microbial clock and Postmotem interval

2014 ESGFOR study 2000’s development of (meta)barcoding, group founded metagenomics, and 2nd-3rd generation sequencing

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Table 10.8.1  Examples of application areas in forensic microbiology. Investigation of the cause of death Estimation of postmortem interval Investigations of forensic archeology and mass graves Estimation of geographical origins of unidentified cadavers Transmission of pathogens related to forensic or medicolegal queries Characterization of body fluids and identification in criminal cases Linkage of a crime scene or a victim with the assailant

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Figure 10.8.2  The human postmortem microbiome. The epinecrotic microbiome consists of skin and body orifices, including the mouth and gastrointestinal tract. The thanatomicrobiome includes all internal organs without anatomical access from the surrounding environment. A body with advanced putrefaction has components of both of these microbiome systems.

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The classical use of investigation of specific microbes or microbial communities in a forensic or medicolegal context is the antemortem versus postmortem presence or absence of microbes. In healthy living individuals, normal physiology and hemostasis are in balance with the human microbiome. This balance is broken when a pathogen invades the human body and causes infectious disease. Typically, pathogens are introduced into the body through body orifices (eyes, nose, mouth, genitals, and anal routes) or open wounds or injuries. The current spectrum of clinically significant and fatal microbes is mostly known, and thus postmortem microbial investigations are directed by these known microorganisms. Emerging microbial disease, epidemics, and potential pandemics, which are of interest to public health institutions, also are of concern to the medicolegal community.

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Postmortem changes can produce artifacts in the microbiomes that are present ante mortem. The longer the postmortem interval (PMI) is, the greater is the alteration of the microbiome. An underlying principle of forensic microbiology and autopsy is that the microbiology on the body and its close environment, that is, skin and surroundings, should be separated from the one in the body, that is, in the internal organs and body fluids. These two microbial systems are the epinecrotic microbiome and thanatomicrobiome, respectively (Figure 10.8.2). Due to the changes in the human body after death, the postmortem microbiology findings can be a result of various developments. Microbial findings in a postmortem sample representing an infectious agent, possibly causing the death or contributing to it, may have alternate explanations for the presence of the microorganism(s). The major critique toward a genuine positive finding is that the microbes found in the body post mortem are due to agonal spread of the microbes, postmortem translocation of those microbes, and/or contamination. These factors may lead to an interpretation of postmortem microbial findings that is in fact a false positive or a false negative.

For the scenarios mentioned in the preceding text, one should take into consideration the following topics to understand the actions that may occur post mortem. A genuine positive finding is a case in which microbes invade the body and infect the target organ(s) or body fluid(s) before an individual’s death. It is noteworthy that the mere presence of microbes does not necessarily imply a fatal infectious disease. Episodes of bacteremia can occur without significant symptoms and evidence of inflammation with associated tissue damage. However, if bacteremia occurred before death and symptoms and an infectious agent were present post mortem, the disease can be considered to be at least a contributing factor to the cause of death. The association of symptoms, signs of a specific disease, and inflammation give greater importance to the presence of the specific microbe. A genuinely positive microbial isolate still is growth (culture) of a recognized pathogen. “Agonal spread” was introduced by Fredette in 1916, as a theoretical concept, in which bacteria invade the bloodstream during the agonal period or during resuscitation. The consequent “terminal infection” is due to spontaneous bacterial invasion into the bloodstream, and there is no observable tissue or organ destruction. The basis for this observation is that the integrity of the mucosal surfaces is compromised by ischemia or hypoxia, allowing a leakage of different microbial species into internal organs. The invasion arises in the body areas where microbes naturally

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teria are introduced into the blood, body fluid, or organ when the sample is obtained. Typically, commensal bacteria from the skin can appear in the organ sample as a false positive finding due to inadequate sampling causing contamination. A way to reduce contamination has been proposed with the means of “sterile autopsy” (O’Toole et al. 1965; Minckler et al. 1966). This concept included sterile morgue facilities with bodies cleaned as if being prepared for a surgical operation, staff wearing similar personal protective equipment as in the operation theater, and autopsy dissections performed with controlled airflow and use of sterile surgical instruments. The postmortem interval was kept short (20 beats/min or PaCO2 < 32 mmHg >12,000 or 10% band forms

PART III  

TRAUMATOLOGY AND VIOLENT DEATH

Both sepsis and SIRS can result in isolated or multiorgan failure and death, with sepsis accounting for 50% mortality despite modern intensive therapy care. In relation to the cause of SIRS, following trauma there is a surge in the stress hormone response mediated by the h ­ ypothalamic– pituitary–adrenal axis which leads to the catecholamine stress response. An overly vigorous pro-­inflammatory, hyperinflammatory host defence response may lead to SIRS with organ injury due to uncontrolled immune activation (Lenz et al. 2007). Compensatory anti-­ inflammatory response syndrome or CARS is a hypoinflammatory response to trauma which can lead to post-­traumatic immunosuppression (Keel and Trentz  2005). This in turn results in an increased susceptibility to infection and systemic septic complications.

18.2  Sepsis and multiple organ failure 18.2.1 Sepsis Human sepsis is a spectrum of pathophysiological changes in the host system resulting from a generalised activation and systemic expression of the host’s inflammatory pathways in response to infection. Normally, pro-­inflammatory mediators such as tumour necrosis factor-­α (TNF-­α), interleukin-­1 (IL-­1), IL-­6 and IL-­8 are released in response to infection or injury and/or ischaemia, to eliminate pathogens and to promote wound healing. This response is then downregulated by the release of anti-­inflammatory mediators (e.g., IL-­1 receptor antagonist (IL-­1ra) and IL-­10), resulting in the restoration of homeostasis. In sepsis, however, local defence mechanisms are insufficient to eliminate the infectious agent and overstimulation of the host’s immune effect or cells occurs. This overwhelming systemic pro-­inflammatory reaction is frequently followed by an overactive compensatory anti-­inflammatory mediator release. The severity of sepsis is proportional to the intensity of the host’s immune and metabolic response to infection. When the balance between pro-­and anti-­inflammatory responses is lost, immunological imbalance and massive systemic inflammation result (Adrie and Pinsky 2000). Sepsis occurs in approximately 1% of all hospital inpatients and accounts for between 20% and 30% of intensive care unit admissions. Despite modern techniques of resuscitation and organ support, septic shock continues to have a mortality rate of approximately 50%.

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An imbalance between SIRS and CARS has been suggested as a cause for multiorgan dysfunction syndrome (MODS) also known as multiorgan failure (MOF). Endothelial cell damage, dysfunction of vascular permeability, capillary leakage, microcirculatory disturbances with cellular hypoxia, parenchymal cell apoptosis and necrosis are all involved in the pathophysiology underlying MODS (Keel and Trentz 2005). MODS is split into primary (early) and secondary (late) types. Examples of primary MODS are cerebral oedema of head injury or ARDS of thoracic injury. The clinical presentation of secondary MODS is dependent upon the organ system affected and is scored using two clinical scoring systems: the Goris score for multiorgan failure or the Marshall score for MODS. The sequential organ failure assessment (SOFA) score can be used to describe the dysfunction of seven systems (i.e., respiratory, cardiovascular, renal, hepatic, gastrointestinal, haematological and central nervous systems). These scores are used to assess injured patients at risk of death (Keel and Trentz 2005).

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gastrointestinal haemorrhage and acute renal failure due to the presence of acute tubular necrosis.

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Necrotising fasciitis is an infection of the fascia planes. It has been reported to result in death, usually about 4 days post trauma, following both sharp force and blunt trauma (Rutty and Busuttil 2000). It can affect virtually any part of the body with the affected part typically going black in colour. Apart from the local infection and necrosis, there can be an associated toxaemia or septicaemia; hence, postmortem blood cultures may be negative for the causative organism.

Miscellaneous complications of trauma Finally, there are a number of local and systemic pathologies that can arise as late complications of trauma which can lead to the death of the patient, either on their own or as part of multisystem failure. These include compartment syndrome, disseminated intravascular coagulation (DIC), physiological stress ulcers of the gastric and duodenal mucosa which may lead to fatal

Definitions and terms

Systemic inflammatory response syndrome The term sepsis, in popular usage, implies a clinical response arising from infection. However, it is apparent that a similar, or even identical, deleterious generalised systemic inflammatory reaction can arise in the absence of infection in response to a variety of life-­threatening clinical conditions (e.g., major trauma, burns, extensive surgical procedures, protracted haemorrhagic or cardiac shock or pancreatitis) (Bone  1996; Weigand et  al. 2004). Figure 18.16 shows the possible reaction patterns of the host response after an initial event leading to injury and/or infection.

Sepsis, severe sepsis and septic shock The condition of SIRS can have either a non-­infectious or infectious origin. When triggered by infection, the term ‘sepsis’ is used. Infection leading to sepsis may be bacterial, fungal, parasitic, protozoan or viral in origin.

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considered pathognomonic for an underlying septic condition in the deceased do not exist. The overwhelming majority of autopsy and microscopic findings in sepsis-­related cell and tissue injury, induced by germs or their products and mediated by a broad cascade of endogenous inflammatory mediators, are neither specific nor sensitive for sepsis and, as a result, lack evidence when considered as isolated findings. Nevertheless, the detection of diverse potentially sepsis-­ induced pathological alterations by routine histological examination can be considered characteristic to a certain degree within the framework of the entire case history and may, therefore, add relevant information to the postmortem elucidation of potentially sepsis-­related fatalities. However, many of the inflammatory organ changes found in sepsis-­related fatalities can also be demonstrated in a large proportion of clinical conditions going along with SIRS or in individuals following prolonged ischaemia, thus reflecting the unspecific reaction pattern of organs and tissues to various endogenous and exogenous noxae. Initiated by a variety of causes and triggered by endogenous mediators, shock events can lead to hypoperfusion with subsequent hypoxia and accumulation of various metabolites resulting in the development of so-­called ‘shock lesions’ that are not specific for shock, as they are found also in ischaemic episodes of other causes with or without underlying SIRS. As a consequence of DIC in sepsis, petechial or more extended haemorrhages can be seen on the skin or on mucocutaneous surfaces and serous membranes or in parenchymal organs by gross examination. The internal organs may be both the focus of sepsis as well as the target of sepsis-­induced tissue alterations, and it is important to distinguish between a primary infectious organ alteration (septic focus) and secondary lesions (septicopyaemic abscesses) as a direct result of bacterial spread from the initial focus on the one hand and tertiary organ alterations (unspecific shock lesions, inflammatory changes in internal organs far away from the initial focus) on the other; the latter initiated by a variety of causes and triggered by a wide range of endogenous mediators in the sequel of the systemic inflammatory cascade.

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Figure 18.16  Possible reaction patterns of the host response after an initial event leading to injury and/or infection. SIRS, systemic inflammatory response syndrome. Source: Adapted from Bone (1996) and Weigand et al. (2004).

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Sepsis and other critical illnesses produce a biphasic inflammatory (immunological, hormonal and metabolic) response. Sepsis, severe sepsis and septic shock represent the increasingly severe stages of the same disease (Bone 1994). ‘Severe sepsis’ is defined by a deterioration of organ function in the presence of hypotension, organ dysfunction and hypoperfusion. The term ‘septic shock’ is reserved for severe sepsis with hypotension despite fluid resuscitation and resultant perfusion abnormalities. The progression from sepsis to severe sepsis and to septic shock is a continuum, reflecting the host’s inflammatory response to infection. During this process, an increasing proportion of patients develop the ARDS, DIC, organ dysfunction syndrome and multiple organ failure.

Pathology

The majority of pathoanatomical textbooks and manuals devote little attention, if any, to pathomorphological organ alterations in sepsis. This is because the clinical pathologist is hardly ever in the position to set up a primary diagnosis of sepsis post mortem. A thorough microscopic examination and toxicological analysis are necessary to rule out concomitant diseases and/or intoxications, respectively, that may have contributed to fatal outcome in a given case. Apart from septicopyaemic abscess formations in internal organs, distinct pathomorphological alterations that can be

Selected organs

Lung The morphological alterations of the lung in sepsis are a consequence of pathophysiological changes defined by the term ARDS. At gross inspection, the lungs in ARDS usually appear to be of a gloomy bluish-­reddish colour and the cut surfaces of the lungs are commonly wet. At microscopic examination, especially in septic shock, there is often marked platelet aggregation with fibrin deposits in the pulmonary vessels (Figure  18.17). The occurrence of microthromboses and megakaryocytes in the pulmonary microvasculature has been reported to appear more frequently in septic shock than in shock of other origin (Tsokos 2005, 2007).

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Figure 18.17  Microthrombosis adjacent to a pulmonary vessel wall in a sepsis death.

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Figure 18.19  Septicopyaemic abscess within the myocardium.

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coronary vascular tone and myocardial contraction rate as well as release of myocardial depressant factor. Septic shock is often especially associated with severe left ventricular dysfunction. Therefore, in cases of septic shock, the left ventricle is often dilated at autopsy with the apex of the heart appearing rounded and the ventricular wall having a flaccid appearance (Müller-­Höcker and Haerty 1993). It was proposed that an interstitial myocarditis can be found in nearly one-­third of sepsis-­related fatalities by histological means (Fernandes et al. 1994). If present in the myocardium, septicopyaemic abscesses (Figure  18.19) are most often located in subendocardial regions of the right ventricle (Tsokos 2005).

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Figure 18.18  Leukocyte sticking of the lungs in septic shock, showing engorgement of the pulmonary vessels and extensive intravascular leukostasis.

The histological finding of pulmonary trapping of polymorphonuclear granulocytes (so-­called ‘leucocyte sticking’; Figure 18.18) reflected by vascular engorgement and extensive leukostasis, most often in the total absence of any interstitial or intra-­alveolar inflammatory reaction, is a striking phenomenon usually seen more often in cases of rapidly fatal septic shock than in sepsis with a prolonged clinical course.

Heart Sepsis leads to depression of myocardial function. This is attributable to a number of mechanisms including haemodynamic alterations, development of myocardial ischaemia, changes in

Liver Despite the still high incidences of cholecystitis, appendicitis and diverticulitis (which are frequently sources of bacterial infection in the liver), pyaemic abscesses of the liver are a relatively rare finding in developed countries. Histologically, leukostasis of neutrophils in the liver sinusoids, formation of intrasinusoidal fibrin aggregations (Figure  18.20) and intraparenchymal haemorrhages are more frequent and more pronounced in septic than hypovolaemic or traumatic shock. However, liver cell necrosis, a common sequel of prolonged hypovolaemic shock, is rarely seen in sepsis. Whilst neutrophils constitute only 1–2% of non-­parenchymal cells in the non-­ septic individual, a dramatic 10–20-­ fold increase in neutrophils occurs within hours following the onset of sepsis. Therefore, especially in acute septic shock, an extensive accumulation of neutrophils (leukostasis) in the liver sinusoids is regarded as a characteristic histological finding (Dinges et al. 1993). A leukocytic infiltration of the acinocentral areas has been described as an infrequent finding in the liver in sepsis (Caruana

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Surprisingly, only a paucity of studies has dealt with the neuropathology of sepsis so far, coming to some discordant findings and to variable incidences in similar lesions. Histologically, a circumscribed loss of neurons in the hippocampus formation has been described in humans dying of septic shock (Evans and Krausz 1994). However, this finding simply reflects a protracted circulatory failure before death and is therefore highly unspecific. In a retrospective neuropathological study, 12 fatal cases of encephalopathy associated with sepsis were investigated. The facultative histomorphological features of septic encephalopathy were described as follows: cerebral infarcts (17%), brain purpura and multiple small haemorrhages (17%), septicopyaemic microabscesses (67%) (Figure 18.21), proliferation of astrocytes and microglia in the cerebral cortex (17%) and central pontine myelinolysis (17%) (Jackson et al. 1985). Every forensic pathologist will agree that these findings, apart from septicopyaemic abscesses, are highly unproductive for establishing the postmortem diagnosis of sepsis. Proliferation of astrocytes and microglial cells is a highly unspecific finding, which may reflect various types of metabolic disorders including ischaemia. Central pontine myelinolysis, a rare neurological disorder defined by symmetrical demyelination in the central base of the pons, is sporadically found in autopsy cases of alcoholism and malnutrition. Pendlebury et al. (1989) reviewed 2107 consecutive autopsies with neuropathological examination and identified 92 cases with pathological evidence for infection involving the central nervous system. Of these, 35 took the form of multiple microabscesses. An underlying sepsis was often present and the lungs were the most frequent site of primary infection with Staphylococcus aureus and Candida albicans being the most frequently identified causative organisms (Pendlebury et al. 1989).

Spleen

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In sepsis, the spleen is usually enlarged and swollen, and the capsule has a tense appearance and is easily accidently torn open during evisceration. The cut surface shows a soft and hyperaemic parenchyma with a reddish-­greyish, sometimes muddy-­brownish appearance. At gross examination, the pathologist’s attention should also focus on the splenic vein, since a septic thrombophlebitis of this vessel can cause pylephlebitic liver abscesses that either spread through neighbouring tissue or via the haematogenous route. The term ‘acute splenitis’ (‘septic spleen’) refers to a soft, runny consistency of the splenic pulp draining from cut sections, histologically corresponding to an increased number of neutrophils and macrophages. The concept of acute splenitis as a postmortem marker of systemic infection is generally accepted and, in this author’s experience (Tsokos), it is the most frequent finding in sepsis-­related fatalities, seen in >90% of cases. Septicopyaemic abscesses of considerable size were a frequent finding in the pre-­antibiotic era, but are rare now. However, various exogenous or endogenous factors such as inappropriate or lack of specific antibiotics, immunocompromise brought about by the severity of an underlying debilitating illness (e.g., malignant diseases, metabolic disorders or immunodeficiency) or concomitant treatment with immunosuppressive agents prior to death may still show up with septicopyaemic abscesses in the spleen.

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Figure 18.20  Fibrin aggregations within the liver sinusoids in sepsis.

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Brain At autopsy, cerebral infarction is a quite uncharacteristic finding in sepsis-­ related fatalities, presenting in 17–26% of cases.

Figure 18.21  Septicopyaemic microabscess in the brain.

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of fibrin thrombi depends upon the time between the onset of DIC and death. The postmortem finding of intravascular microthrombi is more common in individuals who died a few hours after the onset of septic shock than in those who survived the onset of septic shock for days. Despite the obvious clinical manifestation of DIC in the living patient, fibrin thrombi may not be histologically detectable in all cases as a result of postmortem fibrinolysis.

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The development of ARDS is relatively rare in pure hypovolaemic shock events without underlying infection or trauma. At gross inspection, the lungs in ARDS usually appear to be in a gloomy bluish-­reddish colour and the organ weight is increased due to pulmonary oedema, congestion and pulmonary trapping of inflammatory cells. The cut surfaces of the lungs are commonly wet due to accumulation of protein-­rich oedema fluid in the alveolar spaces and interstitial oedema, the amount of muddy-­greyish fluid draining from the cut sections being highly dependent on the quantity of intravenous infusions administered prior to death.

Heart

Subendocardial haemorrhages are a striking feature seen on many occasions in forensic autopsy practice. Subendocardial haemorrhages are frequently seen following haemorrhagic shock, craniocerebral trauma, stroke, preceding resuscitation efforts and intoxications. Subendocardial haemorrhages are most exclusively limited to the endocardium of the left ventricle. Hypercontraction bands and contraction band necrosis (Figure  18.23) as well as elongated and undulated

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One of the most characteristic morphological findings in sepsis-­ related deaths is unilateral or bilateral bleeding in the adrenal cortex. Bleeding of the adrenals (Figure 18.22) may vary in size from tiny focal haemorrhages visible only at microscopy up to total haemorrhagic infarction (‘adrenal apoplexy’) easily detectable at gross examination. To this author’s experience (Tsokos), adrenal haemorrhages are, in addition to acute splenitis, the most frequent finding in sepsis-­ related fatalities. Bilateral adrenal haemorrhage in conjunction with skin bleedings due to DIC and meningitis is classically associated with Waterhouse–Friderichsen syndrome (WFS), most commonly secondary to meningococcal or pneumococcal sepsis.

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Figure 18.22  Massive haemorrhages of the adrenal gland in a sepsis fatality.

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Initiated by a variety of causes and triggered by endogenous mediators, shock events can lead to hypoperfusion with subsequent hypoxia and accumulation of various metabolites resulting in the development of so-­called ‘shock lesions’ that are not specific for shock, as they are found also in ischaemic episodes of other causes in multiple organ failure.

Pathology As a consequence of DIC in multiple organ failure, petechial or more extended haemorrhages can be seen on the skin or on mucocutaneous surfaces and serous membranes or in parenchymal organs by gross examination. In DIC, microthrombi formation may present histologically to a lesser or greater extent in capillaries, arteries and veins of all sizes in each and every tissue and organ (Tsokos 2005, 2007). The frequency

Figure 18.23  Myocardial contraction band necrosis in a case of multiple organ failure.

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cardiomyocytes, often separated by a marked interstitial oedema (depending in its extent to a certain degree on the amount of intravenous infusions administered prior to death), are another characteristic but unspecific finding seen in multiple organ failure deaths.

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At gross inspection, the liver in shock is enlarged showing a tense Glisson’s capsule and rounded edges. The weight of the liver is often increased in multiple organ failure due to both interstitial oedema and accumulation of leukocytes.

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Figure 18.25  Acute tubular necrosis, showing dilated tubules with flattened basophilic and vacuolar endothelium.

Figure 18.26  Shock lesions within the gastrointestinal mucosa, with dot-­shaped fresh haemorrhages.

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Gross pathology of the kidneys in shock and multiple organ failure, respectively, includes bilateral swelling with tense capsules and a pronounced, dark red (congested) medulla contrasting to the paleness of the cortex (Figure 18.24). Acute tubular necrosis is the most frequent form of parenchymal acute renal failure. Acute tubular necrosis showing up as dilated tubules with flattened basophilic and vacuolar endothelium (Figure  18.25) may be induced by sepsis, ischaemia-­ reperfusion and nephrotoxic drugs and is therefore frequently encountered in various forensic autopsy cases. In advanced stages of autolysis, the differentiation between antemortem shock-­induced kidney changes and pure postmortem phenomena is usually not only difficult but often also impossible.

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As a consequence of DIC, microthrombi formation can be detected in the glomerular capillaries in a large proportion of cases. Another frequent, though unspecific finding is the intravascular accumulation of blood and bone marrow cells in the vasa recta of the renal medulla, giving strong evidence of shock prior to death.

Gastrointestinal tract

Figure 18.24  Shock kidney.

Subserous and submucous petechial haemorrhages, erosions and acute ulcers visible with the naked eye at autopsy are the most common shock lesions in the gastrointestinal tract seen in multiple organ failure (Figure 18.26). In addition, fibrin thrombi may be seen histologically in smaller vessels of the bowel mucosa and submucosa.

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Beth Zimmer, M., Nantwi, K. and Goshgarian, H.G. (2007). Effect of spinal cord injury on the respiratory system: basic research and current clinical treatment options. Journal of Spinal Cord Medicine 30: 319–330. Bone, R.C. (1994). Sepsis and its complications: The clinical problem. Critical Care Medicine 22: 8–11. Bone, R.C. (1996). Immunologic dissonance: a continuing evolution in our understanding of the systemic inflammatory response syndrome (SIRS) and the multiple organ dysfunction syndrome (MODS). Annals of Internal Medicine 125: 680–687. BTS (British Thoracic Society) (2008). Emergency Oxygen Guideline Group. Guideline for emergency oxygen use in adult patients. Thorax 63 (Suppl VI): 1–68. BTS (British Thoracic Society) (2017). BTS guideline for oxygen use in adults in healthcare and emergency settings. Thorax 72 (Suppl I): 1–90. Burton, J.L. and Rutty, G.N. (2010). Dissection of the internal organs. In: J.L. Burton and G.N. Rutty (eds.), The Hospital Autopsy. A Manual of Fundamental Autopsy Practice, pp. 145–146. London: Hodder Arnold. Caruana, J.A., Jr, Montes, M., Camara, D.S. et al. (1982). Functional and histopathologic changes in the liver during sepsis. Surgery, Gynecology and Obstetrics 154: 653–656. Cohn, S.M. (1997). Pulmonary contusion: Review of the clinical entity. Journal of Trauma 42: 973–979. Danzl, D.F. and Pozos, R.S. (1994). Accidental hypothermia. New England Journal of Medicine 331: 1756–1760. Dimsdale, J.E., Hartley, L.H., Guiney, T. et al. (1984). Postexercise peril. Plasma catecholamines and exercise. Journal of the American Medical Association 251: 630–632. Dinges, H.P., Schlag, G. and Redl, H. (1993). Morphology of the liver in shock. In: G. Schlag and H. Redl (eds.), Pathophysiology of Shock, Sepsis, and Organ Failure, pp. 257–264. Berlin: Springer. Doberentz, E. and Madea, B. (2019). Traumatic carotid sinus reflex and postmortem investigations of the glomus caroticum in cases of pressure to the neck. In: G.N. Rutty (ed.), Essentials of Autopsy Practice. Reviews, Updates and Advances, pp. 67–88. Cham: Springer. Dreyfuss, D. and Saumon, G. (1998). Ventilator-­induced lung injury. American Journal of Respiratory Critical Care Medicine 157: 289–323. Egger, C., Bize, P., Vaucher, P. et al. (2012). Distribution of artifactual gas mortem multidetector computed tomography (MDCT). on post-­ International Journal of Legal Medicine 126: 3–12. Evans, T.J. and Krausz, T. (1994). Pathogenesis and pathology of shock. In: P.P. Anthony and R.N.M. MacSween (eds.), Recent Advances in Histopathology, pp. 21–47 Edinburgh: Churchill Livingstone. Fernandes, C.J., Jr, Iervolino, M., Neves, R.A. et  al. (1994). Interstitial myocarditis in sepsis. American Journal of Cardiology 74: 958. Filograna, L., Bolliger, S.A., Spendlove, D. et al. (2010). Diagnosis of fatal pulmonary fat embolism with minimally invasive virtual autopsy and post-­mortem biopsy. Legal Medicine (Tokyo) 12: 233–237. Furuya, Y. (1981). Experimental traumatic asphyxia (1). Grades of thoracic compression and mortality. Igaku Kenkyu 51: 117–119. Giertsen, J.C., Sandstad, E., Morild, I. et al. (1988). An explosive decompression accident. American Journal of Forensic Medical Pathology 9: 94–101. Ho, A.M. and Ling, E. (1999). Systemic air embolism after lung trauma. Anesthesiology 90: 564–575.

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Bleeding of the skin is a clinically well-­known manifestation of DIC in the course of multiple organ failure with or without sepsis (Figure  18.27). Sometimes these bleedings appear in a discrete petechial pattern; occasionally, they take the shape of extensive, confluent haemorrhages. In multiple organ failure death, almost exclusively associated with liver failure, jaundice of the skin, sclerae and conjunctivae observed at external examination is also common.

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Figure 18.27  Bleeding of the submucosa of the bowel in multiple organ failure.

References and further reading

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Acosta, J.A., Yang, J.C., Winchell, R.J. et al. (1998). Lethal injuries and time to death in a level I trauma centre. Journal of the American College of Surgery 186: 528–533. Adrie, C. and Pinsky, M.R. (2000). The inflammatory balance in human sepsis. Intensive Care Medicine 26: 364–375. Al-­Sarraj, S., Fegan-­Earl, A., Ugbade, A. et  al. (2012). Focal traumatic brain stem injury is a rare type of head injury resulting from assault: A forensic neuropathological study. Journal of Forensic Legal Medicine 19: 144–151. Armstrong, E.J. (2011). Drowning: A diagnosis of exclusion. In: E.J. Armstrong and K.L. Erskine (eds.), Water-­related Death Investigation. Practical Methods and Forensic Applications, pp. 1–26. London: CRC Press. Arnaud, F., Tomori, T., Teranishi, K. et al. (2008). Evaluation of chest seal performance in a swine model: Comparison of Asherman vs. Bolin seal. Injury 39: 1082–1088. Bajanowski, T., West, A. and Brinkmann, B. (1998). Proof of fatal air embolism. International Journal of Legal Medicine 111: 208–211. BALTS (Battlefield Advanced Life Support) (2001). Shock. Journal of the Royal Army Medical Corps 147: 187–194. Baumann, A., Audibert, G., McDonnell, J. and Mertes, P.M. (2007). Neurogenic pulmonary oedema. Acta Anaesthesiologica Scandinavica 51: 447–455.

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NSCISC (National Spinal Cord Injury Statistical Center) (2019). Spinal Cord Injury Facts and Figures at a Glance. https://www.nscisc.uab. edu/Public/Facts%20and%20Figures%202019%20-­%20Final.pdf last visited April 2020. Ogawa, S., Akutsu, K., Sugimoto, R. et al. (1963). Physiological studies on “coking” in judo with reference to the hypophysio-­adrenocortical system. Bulletin of the Association for the Scientific Studies on Judo, Kodokan report 2: 107–114. Pang, D. and Wilberger, J.E., Jr (1982). Spinal cord injury without radiographic abnormalities in children. Journal of Neurosurgery 57: 114–129. Patel, N.Y. and Riherd, J.M. (2011). Focused assessment with sonography for trauma: Methods, accuracy, and indications. Surgical Clinics of North America 91: 195–207. Pendlebury, W.W., Perl, D.P. and Munoz, D.G. (1989). Multiple microabscesses in the central nervous system: A clinicopathologic study. Journal of Neuropathological Experimental Neurology 48: 290–300. Polson, C.J. (1963). Essentials of Forensic Medicine. Oxford: Pergamon Press. Raschka, C., Koch, H.J. and Rau, R. (2002). Influence of choking and arm lock technique in judo on the acoustic reflex threshold (art) in healthy well-­trained male and female Jodka. Nagoya Journal of Medical Science 65: 29–36. RC (Resuscitation Council) UK (2010). Guidelines. http://www.resus. org.uk/pages/guide.htm#updates (last accessed 16 April 2013). RC (Resuscitation Council) UK (2015). Guidelines. https://www.resus. org.uk/resuscitation-­guidelines/ last visited April 2020. Richards, J.R. and McGahan, J.P. (2017). Focused assessment with Sonography in Trauma (FAST) in 2017: What radiologists can learn. Radiology 283: 30–48. Rodriguez, G., Francione, S., Gardella, M. et al. (1991). Judo and choking: EEG and regional cerebral blood flow findings. Journal of Sports Medicine and Physical Fitness 31: 605–610. Rossen, R., Kabat, H. and Anderson, J. (1943). Acute arrest of cerebral circulation in man. Archives of Neurology and Psychiatry 50: 510–528. Rutty, G.N. and Busuttil, A. (2000). Necrotizing fasciitis: reports of three fatal cases simulating and resulting from assaults. American Journal of Forensic Medical Pathology 21: 151–154. Rutty, G.N. (2003). Fire deaths. In: J. Payne-­James, A. Busuttil and W. Smock (eds.), Forensic Medicine. Clinical and Pathological Aspects, pp. 349–73. London: GMM. Rutty, G.N. (2004). The pathology of shock verses post-­mortem change. In: G.N. Rutty (ed.), Essentials of Autopsy Practice. Recent Advances, Topics and Developments, pp. 93–128. London: Springer. Rutty, G.N., Cary, N. and Lawler, W. (2017). Death in crowds. In: G.N. Rutty (ed.), Essentials of Autopsy Practice. Reviews, Updates and Advances. London: Springer, pp. 46–48. Ruwanpura, R., Kumara, K.S., Jayawardane, H. and de Alwis, L.B. (2008). Suicidal bomb explosions in Sri Lanka. American Journal of Disaster Medicine 3: 47–51. Sauaia, A., Moore, F., Moore, E. et al. (1995). Epidemiology of trauma deaths: a reassessment. Journal of Trauma: Injury, Infection Critical Care 38: 185–193. Sauvageau, A. (2014). Death by hanging. In: G.N. Rutty (ed.), Essentials of Autopsy Practice. Advances, Updates and Emerging Technologies, Vol 6. London: Springer.

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Hopkins, I.H.G., Pountney, S.J., Hayes, P. and Sheppard, M.A. (1993). Crowd pressure monitoring. In: R.A. Smith J.F. Dickie (eds.), Engineering for Crowd Safety, pp. 389–398. London, Elsevier. Jackson, A.C., Gilbert, J.J., Young, G.B. and Bolton, C.F. (1985). The encephalopathy of sepsis. Canadian Journal of Neurological Science 12: 303–307. Keel, M. and Trentz, O. (2005). Pathophysiology of polytrauma. Injury, International Journal of Care Injured 36: 691–709. Klatzo, I. (1967). Neuropathological aspects of brain edema. Journal of Neuropathology and Experimental Neurology 26: 1–13. Knight, B. (1991). Complication of injuries. In: B. Knight (ed.), Forensic Pathology, pp. 339–351. London: Edward Arnold. Leech, C., Porter, K., Steyn, R. et al. (2017). The pre-­hospital management of life-­threatening chest injuries: A consensus statement from the Faculty of Pre-­ Hospital Care, Royal College of Surgeons of Edinburgh. Trauma 19: 54–62. Lenz, A., Franklin, G.A. and Cheadle, W.G. (2007). Systemic inflammation after trauma. Injury, International Journal of Care Injured 38: 1336–1345. Levy, V. and Rao, V.J. (1988). Survival time in gunshot and stab wound victims. American Journal of Forensic Medical Pathology 9: 215–217. Lewis, T. (1932) Vasovagal syncope and the carotid sinus mechanism. British Medical Journal 1: 873–876. Marcus, B.J., Collins, K.A. and Harley, R.A. (2005). Ancillary studies in amniotic fluid embolism: A case report and review of the literature. American Journal of Forensic Medical Pathology 26: 92–95. Marik, P.E. (2011). Pulmonary aspiration syndromes. Current Opinion in Pulmonary Medicine 17: 148–154. Maron, B.J. and Estes, N.A.M. (2010) Commotio Cordis. New England Journal of Medicine 362, 917–27. Mason, J.K. (1968). Pulmonary fat and bone marrow embolism as an indication of ante-­mortem violence. Medicine Science and the Law 8: 200–206. McArthur, B.J. (2006). Damage control surgery for the patient who has experienced multiple traumatic injuries. Association of Peri-­Operative Registered Nurses Journal 84: 991–1000. McCrory, P. and Berkovic, S.F. (1998). Second impact syndrome. Neurology 50: 677–683. McCrory, P. (2001) Does second impact syndrome exist? Clinical Journal of Sports Medicine 11: 144–149. McNarry, A.F. and Goldhill, D.R. (2004). Simple bedside assessment of level of consciousness: comparison of two simple assessment scales with the Glasgow Coma scale. Anaesthesia 59: 34–37. Mellor, S.G. and Cooper, G.J. (1989). Analysis of 828 servicemen killed or injured by explosion in Northern Ireland 1970–1984: the hostile action casualty system. British Journal of Surgery 76: 1006–1010. Menezes, R.G., Fatima, H., Hussain, S.A., et al. (2017). Commotio cordis: A review. Medicine, Science and the Law 57(3): 146–151. Mikhail, J. (1999). The trauma triad of death: Hypothermia, acidosis, and coagulopathy. American Association of Critical-­ care Nurses Clinical Issues 10: 85–94. Müller-­Höcker, J. and Haerty, W. (1993). Pathomorphological aspects of the heart in septic patients. In: G. Schlag and H. Redl (eds.), Pathophysiology of Shock, Sepsis, and Organ Failure, pp. 853–858. Berlin: Springer. NSCISC (National Spinal Cord Injury Statistical Center) (2006). Spinal cord injury: Facts and figures at a glance. Journal of Spinal Cord Medicine 29, 89–90.

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Tsokos, M. (2006). Pathology of sepsis. In: Rutty, G.N. (ed.), Essentials of Autopsy Practice. Current Methods and Modern Trends, pp. 39–86. London: Springer. Tsokos, M. (2007). Postmortem diagnosis of sepsis. Forensic Science International 165: 155–164. Udobi, K.F., Childs, E.D. and Touijer, K. (2003). Acute respiratory distress syndrome. American FamilyPhysician 67: 315–322. Veevers, A.E., Lawler, W. and Rutty, G.N. (2009). Walk and die: An unusual presentation of head injury. Journal of Forensic Science 54: 1466–1469. Weigand, M.A., Horner, C., Bardenheuer, H.J. and Bouchon, A. (2004). The systemic inflammatory response syndrome. Best Practice Research in Clinical Anaesthesiology 18: 455–475. Williams, J.S., Minken, S.L. and Adams, J.T. (1968). Traumatic asphyxia – reappraised. Annals of Surgery 167: 384–392. Wright, R.K. and Davis, J.H. (1980). The investigation of electrical deaths: A report of 220 fatalities. Journal of Forensic Science 25: 514–521. Yunoki, K., Sasaki, R., Taguchi, A. et al. (2016). Successful recovery without any neurological complication after intraoperative cardiopulmonary resuscitation for an extended period of time in the lateral position: a case report. JA Clinical Reports 2 (1): 7. doi:10.1186/ s40981-­016-­0036-­7 Zimmer, G., Miltner, E. and Mattern, R. (1994). Capacity to act after stab and cutting injury. Archiv für Kriminologie 194: 95–104.

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Saunders, S., Kotecha, D., Morgan, B. et al. (2010). Demonstrating the origin of cardiac air embolism using post-­mortem computed tomography; an illustrated case. Legal Medicine (Tokyo) 13: 79–82. Schrag, B., Mangin, P., Vaucher, P. and Bollmann, M.D. (2012). Death caused by cardioinhibitory reflex. What experts believe. American Journal of Forensic Medical Pathology 33: 8–12. Schrag, B., Vaucher, P., Bollmann, M.D. and Mangin, P. (2011). Death caused by cardioinhibitory reflex cardiac arrest -­a systematic review of cases. Forensic Science International 207: 77–83. Sharma, S., Singh, M., Wani, I.H. et al. (2009). Adult spinal cord injury without radiographic abnormalities (SCIWORA): Clinical and radiological correlations. Journal of Clinical Medical Research 1: 165–172. Tadler, S.C. and Burton, J.H. (1999). Intrathoracic stomach presenting as acute tension gastrothorax. American Journal of Emergency Medicine 17: 370–371. Thoresen, S.O. and Rognum, T.O. (1986). Survival time and acting capability after fatal injury by sharp weapons. Forensic Science International 31: 181–187. Tien, H., Chu, P.T.Y. and Brenneman, F. (2004). Causes of death following multiple trauma. Current Orthopaedics 18: 304–310. Tsokos, M. (2005). Pathology of sepsis. In: Rutty, G.N. (ed.). Essentials of Autopsy Practice. Current methods and modern trends, pp. 39–86. London: Springer.

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Vital signs are findings which allow the conclusion to be made about the vital origin of a trauma (e.g., bloodstain pattern in form of arterial blood spray, aspiration or swallowing of blood). Local vital reactions at the site of trauma must be differentiated from systemic vital reactions of the great organ systems (especially circulatory and respiratory systems; Box 19.1.2). In fatal injuries, the survival time after trauma is often short, shorter than the manifestation time of local vital reactions at the site of injury. The manifestation of a local vital reaction at the site of injury (e.g., immigration of polymorphonuclear leucocytes) requires some time. Systemic vital reactions, especially of the circulatory and respiratory systems, develop very fast and are often the main proof of the vitality of injuries. Several influencing factors on the time course of local vital reactions are known (Box 19.1.3). Vital reactions have been a main research topic in forensic medicine for a long period and many renowned forensic pathologists have devoted important papers to this field. The research area ranges from macroscopically visible organ reactions (e.g., haemorrhages of the tongue and Simon’s haemorrhages of the intervertebral discs), over tissue alterations (enzyme histochemistry, later on immunohistochemistry with a wide range of enzymes and other analytes, and molecular pathology) to biochemical responses to injury. Especially in the fields of immunohistochemistry and molecular pathology, much progress has been achieved in the last years (e.g., heat-­shock proteins or positive aquaporin 3 staining in mechanical skin trauma). Furthermore, 20 years after its implementation, postmortem imaging also contributes to the detection and visualisation of vital signs.

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Burkhard Madea, Elke Doberentz, Christian Jackowski, Wolfgang Grellner and Toshikazu Kondo

19.1.1 Introduction

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Vital reactions have been a main research topic of European. Numerous European textbooks include larger sections on vital reactions. According to Bernard Knight, the ‘vital reaction’ is a dubiously valid phenomenon, mainly due to the fact that death and dying are processes, not events (Saukko and Knight 2004). This section describes the basic concepts concerning vital reactions which are of value in daily forensic practice and for future research. The question whether or not an injury was inflicted during life is one of the most important subjects in forensic medicine. As mentioned, because death and dying are processes and not events, the term ‘vital reaction’ (VR) may only constitute a dubiously valid phenomenon in the perimortal period. Any effects in, at or by the body following trauma are named ‘vital reactions’, allowing the conclusion that trauma was really inflicted during life. The term vital reaction can be subdivided into vital reactions, vital processes and vital signs (VS) (Box 19.1.1). Vital reactions are local reactions of tissues at the sites of damages. Contrary to this, vital processes are reactions involving the whole organism and not only local cells and tissues. This reaction of the organism requires functions of the nervous, respiratory and cardiovascular systems.

Handbook of Forensic Medicine, Volume 1, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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Circulation

Box 19.1.1  Definitions of vital reactions, vital processes and vital signs.

Haemorrhages

Vital reactions

Local reactions of tissue at the site of damage

Vital processes

Reaction of the whole organism; not only cells and tissues are damaged. This reaction of the organism requires function of the nervous, respiratory and cardiovascular systems

Vital signs

Findings which allow a conclusion of vital origin (bloodstain pattern in form of arterial blood spray; aspiration or swallowing of blood)

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Source: According to Orsos (1935).

Exsanguination/petechial haemorrhages/ embolisms: air, fat, tissue, bone marrow and foreign bodies (for example, bullet fragments)

Respiratory system

Aspiration (chyme, foreign bodies, blood, brain tissue, soot, water or other fluids) Alveolar–capillary diffusion (gas; detection of gas in the circulation) Emphysema acuta (for example, emphysema aquosum seen in drowning) Emphysema of the skin

Gastrointestinal tract

Vomiting/swallowing Peristaltic transport of stomach contents Absorption/resorption of detectable substances Gastric mucosal erosions (for example, Wischnewsky ulcers seen in hypothermia)

Endocrine glands

Agonochemical stress reaction with increase of the catecholamine levels

Nervous system

Crow’s-­feet-­like pattern/secretion of saliva and mucus

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Box 19.1.3  Possible influencing factors on the time course of local vital reactions after tissue damage.

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General factors

Exogenous factors

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Box 19.1.2  Vital reactions of different organ systems (systemic vital reactions).

Local factors

Internal or external haemorrhages are seen in a variety of traumatic deaths. They are mainly due to lesions of arteries, veins or capillaries. Diapedetic haemorrhages or haemorrhages due to coagulation disorders are comparatively rare. Haemorrhages due to lesions of arteries or veins are normally seen at the site of trauma (Table 19.1.1). Capillary haemorrhages such as petechial haemorrhages or ecchymoses can also be seen distant from the site of injury (Figure 19.1.1). Petechial haemorrhages are due to an intravascular rise of pressure with an increasing transvascular pressure gradient from the inside to the outside of the vessel. The higher the intravascular pressure is, the shorter the manifestation time of petechial haemorrhages. Petechial haemorrhages are of great medicolegal significance regarding the following questions: • Vitality of injury (e.g., pressure to the neck). • Was the compression of the neck life-­threatening? • Priority of injuries (e.g., first pressure to the neck, then ­stabbing, or vice versa). • Distinguishing the type of pressure to the neck between reflex death and manual or ligature strangulation. Intensive subcutaneous haemorrhages may also be present in postmortem trauma (Figure 19.1.2), and considerable amounts of blood may leak from wounds post mortem. However, exsanguination due to anaemia does not occur. Exsanguination of a body due to anaemia of the internal organs is always a vital reaction. Minor extent and intensity of postmortem lividity, and anaemia of the body, the skin and mucous membranes, together with subendocardial bleeding, are typical findings of fatal haemorrhage. Further typical vital haemorrhages are the so-­called ‘Simon’s haemorrhages’ which are found over the lumbar spine in cases of typical hanging;

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• Hereditary factors, age, gender • Nutritional status • Additional diseases • Endocrine influences • Vegetative influences • Metabolic disorders • Pharmaceuticals • Vital hypothermia/hyperthermia

Table 19.1.1  Vital reactions – haemorrhages. Phenomenon

Mechanism

Detection

Postmortem origin

Macroscopic Yes Lesions of vessels, Arterial/ blood extravasation venous corresponding to haemorrhage the pressure drop intravasal/ extravasal Capillary

Intracapillary pressure rise caused by congestion with increase of the transvascular pressure gradient

Macroscopic Yes

The degree of haemorrhage depends on the size of the injured vessel, the blood pressure and the resistance to the blood that is streaming.

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haemorrhages of the tongue in long-­lasting manual or ligature strangulation, congestion, cyanosis and petechial haemorrhages above the ligature in atypical hanging and intramedullary haemorrhage of the cervical spinal cord are due to contusion of the neural tissue (Figures 19.1.3 and 19.1.4).

Embolism

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Fat embolism  Fat embolism mostly originates from destroyed subcutaneous fatty tissue as a direct result of trauma. Twenty to thirty grams of fat can result in a fatal fat embolism. Causes of death are either right-­heart failure due to blockage of pulmonary vessels or fat embolism of the brain. Fat embolism can always be identified histologically. A classic grading of pulmonary fat embolism was recommended by Falzi et al. (1964) (Table 19.1.3).

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Amniotic fluid embolism  Amniotic fluid embolism is a rare but possibly lethal complication during pregnancy and birth. It is due to a direct communication between amniotic cavity and maternal venous system. The histological diagnosis is based on the detection of amniotic fluid components such as skin particles, meconium, lanugo hair and mucus in peripheral pulmonary vessels. Additionally, early shock alterations such as platelet aggregates and microthromboses can be found. The diagnosis can also be made by immunohistochemical proof of cytokeratin in fetal skin particles.

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Figure 19.1.1  (a) and (b) Petechiae of the face, the eyelids and the conjunctivae. (c) Petechiae of the mucosa of the larynx due to thoracic compression.

Figure 19.1.2  Postmortem subcutaneous haemorrhages due to investigation of the idiomuscular pad, showing tram-­like haemorrhages up to 8 hours post mortem. Source: Reproduced with permission from Dotzauer (1958).

Cell and  tissue embolism  Cell and tissue embolism is often found after trauma with injury (laceration) of internal organs with spreading of organ and tissue parts via the bloodstream into the lungs. Air embolism  Routine X-­ ray examination reveals that air embolism is far more common in medicolegal autopsies than generally expected. In particular, this phenomenon is found in cases of craniocerebral trauma with rupture of the sinus as site of air entry. These cases are comparatively rarely seen today in cases of illegal abortion, but may occasionally occur as a result of medical malpractice (e.g., neck surgery with opening of cervical veins and high pressure infusion). Volumes of about 70 mL of air may cause immediate death due to air embolism. A characteristic autopsy finding is an acute dilatation of the right ventricle with no or only some foamy blood seen in the ventricle. The diagnosis of air embolism is made by chest X-­rays taken routinely prior to autopsy or by a special autopsy technique with opening of the right and left ventricles under water (technique according to Richter 1905). In the case of air embolism, the escaping amount of air is either described with regard to volume, the duration of the escape and the volume of gas bubbles or the air being analysed by gas chromatography. The presence of plenty of gas in the right heart chamber in the absence

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Figure 19.1.3  Haemorrhages. (a) Haemorrhages of the intervertebral discs (Simon’s haemorrhages) in case of typical hanging. (b) Haemorrhages of the tongue in a case of long-­lasting ligature strangulation. (c) Haemorrhages of the cervical medulla due to a temporary cavity in an extradural shot to the neck. (d) Cyanosis and swelling of the face, massive congestion and petechial haemorrhages in an atypical hanging.

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of putrefaction is considered a valid proof of air embolism. The discussion about the importance of gas analytical findings for the detection of air embolism has not yet been concluded; however, gas analysis will be necessary in the future in appropriate cases. A normal composition of air at 760 mmHg (1013 hPa) and 15 °C is the reference for gas analytical findings in a body: nitrogen 78.1 vol%, oxygen 20.9 vol%, argon 0.9 vol% and other gases with concentrations much less than 1  vol%; for example, carbon dioxide 0.03  vol%, hydrogen 0.00005 vol%, methane 0.0002 vol% and sulphur dioxide 0.0–0.0001 vol%. Typical components of putrefactive gas are carbon dioxide and hydrogen, but it could also contain methane and hydrogen sulphide. Pedal et  al. (1987) have recommended carbon dioxide concentrations of less than 15%, nitrogen concentrations higher

than 70% and a ratio (carbon dioxide divided by nitrogen) of less than 0.2 as reliable gas analytical criteria for a diagnosis of air embolism. According to these authors, the measured volumes, oxygen concentrations and the ratio of carbon dioxide to oxygen do not show a distinct differentiation between air embolism and putrefaction.

Respiration Besides the circulatory system, vital reactions of the respiratory system are also very important (e.g., for the differential diagnosis between live birth and stillbirth). Important vital reactions of the respiratory system include acute pulmonary emphysema and aspiration/inhalation (Figures 19.1.6 and 19.1.7). In cases of neck

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Figure 19.1.4  (a) Subendocardial haemorrhages in a case of bleeding to death. (b) Shock kidneys with pale cortex and sharp demarcation to the renal medulla.

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Table 19.1.2  Vital reactions–embolisms. Mechanism

Detection

Embolism

Intact circulation

Thromboembolism

Coagulation, intact circulation

Bullet embolism

TO

R

Phenomenon

Postmortem origin

No

Entry of the projectile into the vascular system, intact circulation

Macroscopic

No, or spreading by resuscitation

Fat embolism

Traumatic migration of fat or fat marrow into the circulatory system, intact circulation

Histological

No differential diagnosis of nontraumatic fat embolism in case of acute liver dystrophy

Air embolism

Intake of air/injection of air into the vascular system, intact circulation

Radiological, test for air embolism (Richter), gas analysis

Putrefaction gas

Amniotic fluid embolism

Entry of amniotic fluid into the venous vascular system, intact circulation

Histological

No

Tissue embolism

Traumatisation of tissues with irruption into the vascular system, intact circulation

Macroscopic/histological

No

Histological

Resuscitation

Spreading of bone marrow, intact circulation

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Bone marrow embolism

R

C

O

N

TR IB

U

Macroscopic/histological

Table 19.1.3  Grading of fat embolism (FE). FE grade

Type of embolism

Localisation

0 = no FE

Punctiform

Possibly only rarely, not in every visual field

1 = mild FE

Drop-­shaped

Rarely, in every visual field

2 = obvious FE

Lake-­shaped or sausage-­shaped

Multiple, detectable in every visual field

3 = massive FE Antler-­shaped

Source: According to Falzi et al. (1964).

Plentiful, everywhere, in every visual field

compression, an increase in airflow resistance is observed. This is characterised by an inspiratory stridor. Due to an increase of pCO2, an acceleration and a deepening of respiration are noted leading to acute pulmonary emphysema, possibly also due to interstitial emphysema. Acute pulmonary emphysema is also found in cases of drowning. It loses its diagnostic value in cases of resuscitation with artificial ventilation or with putrefaction.

Aspiration/inhalation Aspiration of liquids and solids is an important sign of vitality in cases of aspiration of blood, soot, gastric contents and

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drowning. Only inhalation into the deeper parts of the bronchial tree can prove vitality. Postmortem fluids might also flow passively into the trachea and the main bronchi (Table 19.1.4).

N

LY

Blood aspiration  Blood aspiration can be diagnosed by subpleural foci or on the cut surfaces of the lung by a leopard-­skin-­like pattern (Figure 19.1.8). However, similar findings could be produced also post mortem by artificial ventilation with a respiration bag (Dräger) after different amounts of blood were filled into the trachea. The findings produced post mortem could not be differentiated from vital blood aspiration. Inhaled blood and other aspirated fluids are mixed with air during ventilation. Blood-­coloured foam or foamy fluid is formed in the airways and the oral cavity. Drowned bodies show foam and froth protruding from the mouth if the respiratory orifices are not below the water surface. This froth and also blood-­ coloured foam originate from a mixture of aspirated fluid, air and bronchial secretion (Figure 19.1.9).

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O

Figure 19.1.5  Fulminant massive pulmonary thromboembolism.

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Figure 19.1.6  (a) Massive pulmonary emphysema in case of drowning. The margins of the lungs cover the mediastinum nearly totally (sometimes even overlapping). (b) Massive pulmonary emphysema in a homicide by drowning with numerous subpleural haemorrhages.

Figure 19.1.7  (a) and (b) Floating of lung and small intestine in a positive hydrostatic test where the infant’s survival time after birth was more than 12 hours.

CHAPTER 19  

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Vital Reactions

Table 19.1.4  Vital reactions – aspiration/inhalation. Phenomenon

Mechanism

Detection

Postmortem origin

Aspiration In case of apnoea only by insufflation (artificial ventilation)

Inhalation of gas/fluids, penetration of liquids and solid particles into the respiratory tract during inspiration 1. Bleeding penetrating into the tracheobronchial system 2. Ventilation

Macroscopic/histological

In case of apnoea only by insufflation/ artificial ventilation

Brain tissue aspiration

1. Head injury with basal skull fracture 2. Ventilation

Macroscopic/histological

Spontaneous

Soot

1. Soot production during fire/ smouldering fire 2. Ventilation

Macroscopic/histological

If charring is extensive with charring defects to the trachea and the lungs, artificial findings are possible

Inhalation of hot gases

1. Inhalation of hot air/steam 2. Thermic mucous membrane damage

Microscopic

Fluids (drowning)

1. Submersion, originated in water 2. Ventilation

Macroscopic

In case of apnoea only by insufflation; mucous membrane damage at least partly possible post mortem Penetrating also possible for corpuscular components by hydrostatic pressure (depending on water depth)

FO

R

C

O

N

TR IB

U

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O

N

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Blood aspiration

Figure 19.1.8  (a) and (b) Blood aspiration with leopard-­skin-­like pattern subpleurally and on the cut surface of the lung.

Soot inhalation  Soot inhalation into finer trachea branches within the lungs proves vital in burning during life. In cases of severely burnt bodies with opening of the trachea and the thoracic cavity, artificial findings of soot may be found. Histologically, soot may be found in the respiratory tract and the finest bronchi, often combined with extensive mucus secretion. Soot particles embedded in mucus are a proof of vitality. In cases of heat inhalation, capillary and venous congestion with microhaemorrhages of the tracheal submucosa, elongation of cells and nuclei and mucosal oedema with ectasia of lymphatic vessels may be found. In cases of burning, inhalation of carbon monoxide and hydrogen cyanide is a further proof of vitality (Figure 19.1.10).

Acute emphysema  An important VR of the respiratory system is the acute pulmonary emphysema. For example, in cases of neck compression, an increase in airflow resistance is observed. This is characterised by an inspiratory stridor. Due to an increase of pCO2, an acceleration and a deepening of the respiration are promoted leading to acute pulmonary emphysema, possibly also to interstitial emphysema. Acute pulmonary emphysema is also found in cases of drowning where it is called emphysema aquosum. Emphysema alterations lose their diagnostic value in cases with resuscitation including artificial ventilation or with advanced putrefaction.

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• No significant increase in the number of alveolar macrophages or polynuclear giant cells can be observed in cases of strangulation/ throttling or chest compression. The duration of the asphyxiation process, at least in most of these forensic autopsy cases, does not seem to be long enough to enable the development of pulmonary polynuclear giant cells, as described in experimental animals with considerably longer intervals of hypoxia. An own recent study underlines the fact that the preterminal hypoxia, being observed in a usual medicolegal scenario (maximum 20 minutes in cases investigated), does not suffice to induce changes in the populations of pulmonary inflammatory cell lines.

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Digestion

N

Swallowing

TR IB

U

U

TO

Histological alterations in the lungs  There is still controversial discussion on the issue, whether the appearance and number of macrophages and giant cells in pulmonary tissue can support the diagnosis and type of, for example, fatal asphyxia, in particular with regard to the survival time (rapid or protracted asphyxia). Janssen described a mobilisation and proliferation of alveolar cells with the detection of giant cells under special conditions. Later, other authors also showed the presence of these cells in other causes of death. The following practical conclusions were drawn (Betz et al. 1994):

R

Figure 19.1.9  Froth extruding from the mouth and nose in a drowned person.

SE

O

Swallowing is defined as the voluntarily induced propulsory reflectory peristaltic transport of fluid or food into the stomach. Swallowing of blood, foreign objects, tissue components, teeth, drowning fluid and soot occurs intravitally. However, drowning fluid only in the stomach alone is no evidence of drowning, since due to hydrostatic pressure water can get into the stomach even post mortem. In cases of drowning, there is a typical separation of the gastric content into three phases with a foamy phase on top, a liquid phase in the middle and a solid phase at the bottom (so-­ called Wydler’s sign). Mucosal tears of the gastric mucosa caused by overexpansion of the stomach due to swallowed drowning fluid or to emesis under water against resistance are also vital reactions. Like inhaled air, swallowed air is considered as a vital sign in newborns in the absence of putrefactive changes.

• Polynuclear giant cells can also occur in the lungs of healthy,

Aquaporins are membrane water channels that are part of the control of the water content of cells. They are expressed in many epithelial and endothelial cells. Studies showed that

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O

N

non-­asphyxiated individuals, and considerable variations in the number, in particular, of monoculear alveolar macrophages can be found.

VR/VS related to structural proteins

Figure 19.1.10  Soot aspiration (a) macroscopically and (b) microscopically, together with mucous and epithelial hyperaemia of the submucosal vessels due to inhalation trauma.

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CHAPTER 19  

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O

Figure 19.1.11  (a) Aquaporin 3 staining; gunshot entrance wound. Aquaporin 3 staining 100% positive × 400. (b) Aquaporin 3 staining; frost erythema. Aquaporin 3 staining 100% positive × 400. (c) Aquaporin 3 staining; excoriation 100% positive staining of the epidermis × 400. (d) Aquaporin 3 staining, incised wound 100% positive staining of epidermis × 400.

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aquaporin 3 (AQ3) is considered as a vulnerable marker of antemortem neck compression (Figure  19.1.11). Own studies revealed that AQ3 can be used as a vitality marker for all kinds of antemortem skin injuries. There is no significant difference between the ligature group on one hand and other forms of external violence to the skin on the other hand.

Structural proteins in circulation Structural damages cause a depletion of structural proteins at the tissue affected and an increase of these proteins in the circulation. Clinical biomarker of structural damage such as troponin for myocardium may be analysed post mortem as well and has shown to be stable for a shorter postmortem interval. As a further biochemical vital reaction, increased myoglobin concentration in cardiac blood in fatal electrocution was

discussed. These increased values were thought to be due to myocytolysis and lack of myoglobin into the blood. However, further investigations revealed that the myoglobin concentration in cardiac blood increases with increasing postmortem interval ­ (PMI). There was no correlation with fatal electrocution.

VR/VS related to regulatory proteins Heat shock proteins (HSPs) are a group of proteins that belong to molecular chaperones. HSPs were first described in 1962. They have a cytoprotective function and support the cell to survive in (sub)lethal conditions. HSPs control protein biosynthesis by supporting accurate protein synthesis, preventing aggregation of newly folded proteins and stabilising various cell structures. HSPs are expressed in cells when there is exposure to variable stressful stimuli. HSP expression is for instance stimulated by hypothermia

TRAUMATOLOGY AND VIOLENT DEATH

Turillazzi et al. (2010) investigated the immunohistochemical expression of a panel of cytokines and inflammatory cells in skin specimens of autopsy cases of death due to hanging to discuss their significance in assessing whether hanging mark and signs occurred before or after the death of the victim. They concluded that tryptase, IL-­15 and CD15 appeared to be reliable parameters in the determination of the vitality of ligature marks.

Regulatory proteins in circulation

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Since more than 70 years, many investigations have been carried out on various biochemical analytes as vitality markers, especially lactate, catecholamines, thyroglobulin, etc. Berg especially studied the suitability of catecholamines as vitality markers in strangulation. In external asphyxiation, high catecholamine values were found in comparison to reflex death (Table 19.1.5).

N

TR IB

U

TO

R

U

SE

or hyperthermia, as well as by every other external and internal physiological stress. Important HSPs are HSP27 and HSP70, named according to their molecular weight. Expression of HSPs has been extensively studied in renal tissue, especially in cases of hypothermia and hyperthermia (Figure  19.1.12). HSPs are expressed in renal tissue in cases of lethal hypothermia. In a study group of 100 cases of death due to hypothermia, renal glomerular HSP70 expression was observed in 80% of cases and renal tubular expression was found in 89% of cases compared with a control group. In fire-­related fatalities, an extensive and different expression pattern of HSP27 and HSP70 according to the survival time can be observed. HSP27 is rapidly expressed in short-­term survivors and HSP70 is higher in long-­term survivors. In fire-­related fatalities, especially a very rapid HSP27 expression can be found in the pulmonary tissue, but also in renal tissue.

N

PART III  

O

358

C

O

Figure 19.1.12  Heat shock proteins in fire death. (a) Positive expression of the trachea and soot aspiration, and (b) positive expression of intrapulmonary tissue.

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Table 19.1.5  Catecholamine concentration in cadaveric blood (serum in ng/mL) in 111 cases. Calculation of significance of pair differences (adrenaline–noradrenaline) according to Wilcoxon. Adrenaline

Noradrenaline

Difference

Duration of terminal episode

Cause of death

m

s

m

s

A − NA

n

P

I short

External asphyxiation Internal asphyxiation Fatal haemorrhage Cardiac death

197 105 128 135

142 73 143 88

175 134 98 195

145 119 141 148

22 −29 30 −60

30 12 12 16

0.1 0.1 0.05 0.1

II long

Craniocerebral trauma Intoxication

242 200

253 127

285 129

266 108

−43 −29

21 10

III missing

Reflex death, bolus death, sudden cardiac death

11

6

12

9

−1

10

0.1   0.1

m = mean value. Source: From Berg and Bonte (1973).

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N

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Especially in myocardial ischaemia, postmortem magnetic resonance (PMMR) being very sensitive to pH value changes and oedematous alterations has proven to visualise ischaemic myocardium already when macromorphology and micromorphology remain negative. Thereby, PMMR especially on higher field strengths such as used on 3T scanners supports the detection and visualisation of the vital ischaemic lesion causing the lethal ventricular arrhythmic event, which is often not to be proved with morphological techniques (Figure 19.1.13). The autopsy detection of cerebral ischaemia or haemorrhage may also be challenging in cases with an advanced stage of

LY

VR/VS in postmortem imaging

putrefaction. When brain liquefaction started already, macromorphology and histology may not allow for a significant assessment of the brain tissue anymore. However, experiences with PMMR have shown that as long as the skull and dura remain intact, the cerebral microanatomy remains far longer assessable by PMMR compared to dissection approaches (Figure 19.1.14). Especially postmortem magnetic resonance imaging (PMMRI) is very sensitive to fatty tissue, as it has high T1w and T2w signals due to its long hydrogen-­containing chains. To visualise fat embolism as cause of death and as vital sign by PMMRI has been challenged by two research groups so far. Massive accumulation of intravascular fat has been demonstrated already, but the use of PM imaging to securely detect and quantify fat embolism also within the precapillary arterioles requires a spatial resolution, which exceeds the resolution limits of today’s 1.5 and 3T scanners. However, further developments of the MR technique are expected to allow an imaging-­based quantification of the embolised fat content throughout the entire lungs. That will provide a more objective and general assessment of the degree of fat embolism compared to the histological assessment techniques used today. And it will also ensure that minor fat accumulations within the pulmonary circulation do not remain undetected at the histological level and can support the assessment of vitality even when routine histology is negative. Especially postmortem computed tomography (PMCT) has the advantage of easy and secure gas detection, whereas autopsy requires some suspicion in advance to decide for specific detection techniques to verify, for example, gaseous emboli. Thereby, the broad implementation of PMCT prior to forensic autopsy revived the value of gaseous findings related to the cause of death as well as related to the vitality of sustained injuries (Figure 19.1.15). All gas-­related findings are detected meanwhile in any case independent from any existing or non-­existing suspicion. Even tiny amounts of pathological intravascular gas accumulations, which are likely to remain undetected at autopsy, may be interpreted as a sign for an ongoing circulation at the time of injury. Also, a quantification of pathological gas accumulations may be easily performed using the image-­reading software solutions coming along with the computed tomography (CT) scanners. Extravasal gas accumulations have also been described as possible vital signs when related to ongoing ventilation. For example, in hanging cases, ascending mediastinal emphysema up to the strangulation mark may only be explained by the forced ventilation attempts against increased resistance causing acute alveolar ruptures. Starting at the most peripheral bronchioles, the gas finds its way along the peribronchial tissue into the mediastinum and upwards into the soft tissues of the neck. Since these interstitial gas accumulations may easily be overlooked at autopsy, PMCT has increased the detectability of this ventilation-­related vital sign. However, when interpreting pathological intravasal and extravasal gas accumulations as possible vital signs, the occurrence of mechanical resuscitation attempts has to be taken into account, as these are also known to cause pathological gas accumulations. Furthermore, with increasing postmortem interval,

N

Furthermore, the catecholamine values are correlating with the duration of terminal episode. Craniocaudal concentration differences of the catecholamine concentration between left ventricular and sinus blood were described. However, this finding could not be confirmed in further investigations.

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CHAPTER 19  

Figure 19.1.13  Peracute ischaemic myocardial lesion without any myocardial finding at autopsy. (a) T2w short-­axis image presenting with a focal hypointensity within the septum. (b) 90% stenosis within the dominating and the septum supplying coronary.

360

TRAUMATOLOGY AND VIOLENT DEATH

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TO

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N

Figure 19.1.14  Vital intracranial haemorrhage in an almost liquefied brain. (a) Intracranial flattening of the brain due to liquefaction in axial T2w MR image. (b) Sharply outlined hypointensity representing a haemorrhage in the left parieto-­occipital region. (c) Liquefied brain at autopsy. (d) Attempt to visualise the haemorrhage within the liquefied brain.

Summary

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R

C

putrefaction gas appears within the vascular system. The attenuation behaviour is similar to, for example, air; however, the dissemination of putrefaction gas may differ from vital embolic gas distributions. PMCT may also show tiny aspirated fragments not to be found at autopsy, such as bony fragments in skull trauma (Figure 19.1.16).

Vital signs and vital reactions in forensic practice: 1.  Blunt forces • Severe haemorrhages. • Fat embolism. • Tissue embolism. • Discolouration (yellowish, greenish, brownish) haemorrhages. • Active exsanguination. • Active and passive defence injuries.

of

• • • •

Blood and tissue aspiration. Soft-­tissue emphysema. (Tension) pneumothorax. Histological and immunohistochemical vitality markers. 2.  Sharp forces • Gas embolism. • Active exsanguination. • Blood aspiration. • Active and passive defence injuries. • Soft-­tissue emphysema. • (Tension) pneumothorax. • Histological and immunohistochemical vitality. 3.  Strangulation 3.1 Ligature strangulation • Petechiae. • Cranial congestion. • Haemorrhages at strangulation mark. • Haemorrhages around fractures of laryngeal skeleton.

361

Vital Reactions

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CHAPTER 19  

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Figure 19.1.16  Axial PMCT image of the thorax in case of a severe head trauma. The arrow points to a tiny piece of bone aspirated from the damaged skull base. Note the air embolism within the pulmonary artery as well.

• Self-­inflicted injuries close to the manual strangula-

U

tion marks (nail marks).

• Aquaporin 3 expression.

N

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3.3 Hanging • Petechiae (cave: in free-­suspension hangings, petechiae are a rarity and may indicate a different lethal strangulation prior to the hanging). • Haemorrhages beneath strangulation mark. • Intracutaneous haemorrhages between two strangulations marks. • Haemorrhages at clavicular insertion of sternocleidomastoid muscle. • Simon’s haemorrhages at the lumbar discs. • Ascending mediastinal emphysema. • Saliva excretion. • Aquaporin 3 expression under strangulation mark. 4.  Asphyxiation • Increased catecholamines. • Water condensation (e.g., inside a head-­covering plastic bag). • Acute emphysema. • Subpleural haemorrhages. 5.  Hyperthermia/fire • Soot aspiration. • Soot swallowing. • Thermal inhalation trauma. • Carbon monoxide and other fire-­related gases in blood. • Heat shock proteins. • Enhanced smile wrinkles. • Fatty degeneration of tubular epithelium. • Positive HSP expression in hot water drowning. 6.  Hypothermia • Wischnewsky spots in stomach. • Haemorrhages in the body core muscles (M. psoas).

C

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Figure 19.1.15  Air embolism as seen using PMCT. (a) Axial PMCT image with air-­filled right heart chamber. (b) Completely air-­filled pulmonary artery.

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• Aquaporin 3 expression under strangulation mark. • Intracutaneous haemorrhages between two strangulations marks.

• Defence injuries. • Self-­inflicted injuries close to the strangulation mark (nail marks).

• Further histological and immunohistochemical vital-

ity markers. 3.2 Manual strangulation • Petechiae. • Cranial congestion. • Subcutaneous and intramuscular haemorrhages beneath manual strangulation mark. • Haemorrhages around fractures of laryngeal skeleton. • Defence injuries.

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• Heat shock proteins. • Frost erythema.

7.  Water/drowning • Acute emphysema (emphysema aquosum). • Frothy tracheal content and froth extruding from mouth and nose.

• Swallowing of drowning fluid. • Diatoms in lung periphery.

N

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References and further reading

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8.  Electricity • Indirect signs of cardiac failure such as congestion. 9.  Gunshots • Gas embolism. • Fat embolism. • Foreign body embolism. • Tissue embolism. • Active exsanguination. • Severe haemorrhages along the wound channel. • Severe haemorrhages along the temporary cavity. • Active and passive defence injuries. • Blood and tissue aspiration. • Soft-­tissue emphysema. • (Tension) pneumothorax.

Doberentz, E. and Madea, B. (2018). Heat Shock Protein Expression in Various Tissues in Thermal Stress. In: A. Asea and P. Kaur (eds.), Regulation of Heat Shock Protein Responses. Heat Shock Proteins, Vol 14, pp. 429–461. Berlin Heidelberg New York: Springer. Doberentz, E., Führing, S. and Madea, B. (2015). Sudden infant death syndrome (SIDS) – no significant expression of heat shock proteins (HSP27, HSP70). FSMP 12 (1): 33–39. Doberentz, E., Führing, S. and Madea, B. (2016). Hsp27 and 70 expression in the heart, lung and kidney in SIDS. Romanian Journal of Legal Medicine 24: 247–252. Doberentz, E., Genneper, L., Böker, D. et al. (2014). Expression of Heat Shock Proteins (Hsp) 27 and 70 in various organ systems in cases of death due to fire. International Journal of Legal Medicine 128: 967–978. Doberentz, E., Genneper, L., Wagner, R. and Madea, B. (2017). Expression times for Hsp27 and Hsp70 as an indicator of thermal stress during death due to fire. International Journal of Legal Medicine 131 (6): 1707–1718. Doberentz, E., Markwerth, P., Wagner, R. and Madea, B. (2017) Expression of Hsp27 and 70 and vacuolization in pituitary glands in cases of fatal hypothermia. FSMP 13 (3): 312–316. Dotzauer, G. (1958). Idiomuskulärer Wulst und postmortale Blutung. Deutsche Zeitschrift für die Gesamte Gerichtliche Medizin 46: 761–771. Falzi, G., Henn, H.E. and Spann, W. (1964). Über pulmonale Fettembolie. Münchener Medizinische Wochenschrift 106: 978–981. Grellner, W. and Madea, B. (1994). Pulmonary micromorphology in fatal strangulations. Forensic Science International 67: 109–125. Grellner, W. and Madea, B. (2002). Role of pulmonary macrophages and giant cells in fatal asphyxia – comment on “Is the appearance of macrophages in pulmonary tissue related to time of asphyxia?”. Letter to the Editor. Forensic Science International 127: 243–244. Grellner, W., Madea, B., Kruppenbacher, J.P. and Dimmeler, S. (1996). Interleukin-­1 (IL-­1 ) and N-­formyl-­methionyl-­leucyl-­phenyl-­ alanine (FMLP) as potential inducers of supravital chemotaxis. International Journal of Legal Medicine 109: 130–133. Grellner, W., Vieler, S. and Madea, B. (2005). Transforming growth factors (TGF-­a und TGF-­b1) in the determination of vitality and wound age: immunohistochemical study on human skin wounds. Forensic Science International 153: 174–180. Grellner, W.and Madea, B. (1996). Immunohistochemical characterization of alveolar macrophages and pulmonary giant cells in fatal asphyxia. Forensic Science International 79: 205–221. Gutjahr, E. and Madea, B. (2019). Diagnose einer gewaltsamen Erstickung: Re-­Evaluation der Spezifität makroskopischer und histomorphologischer Befunde. CME-­Beitrag Teil 1. Rechtsmedizin CME Zertifizierte Fortbildung 30: 55–63. Gutjahr, E. and Madea, B. (2019). Inflammatory reaction patterns of the lung as a response to alveolar hypoxia and their significance for the diagnosis of asphyxiation. Forensic Science International 297: 315–325. Hejna, P. and Rejtradová, O. (2010). Bleedings into the anterior aspect of the intervertebral disks in the lumbar region of the spine as a diagnostic sign of hanging. Journal of Forensic Sciences 55 (2): 428–431. Helpap, B. (1987). Leitfaden der Allgemeinen Entzündungslehre. Berlin: Springer Verlag.

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Berg, S. (1963). Physiologisch-­chemische Befunde im Leichenblut als Ausdruck des Todesgeschehens. Deutsche Zeitschrift für die Gesamte Gerichtliche Medizin 54: 136–149. Berg, S. (1966). Adrenalin-­und Noradrenalinwerte im Blut bei gewaltsamen Todesursachen. Deutsche Zeitschrift für die Gesamte Gerichtliche Medizin 57: 179–183. Berg, S. (1975). Vitale Reaktionen und Zeitschätzung. In: B. Mueller (Hrsg), Gerichtliche Medizin Band 1, pp. 326–340. Berlin, Heidelberg, New York: Springer. Berg, S. and Bonte, R. (1973). Katecholaminwerte im Leichenblut und Liquor bei verschiedenen Agonieformen. Zeitschrift fur Rechtsmedizin 72: 56–62. Betz, P. (1996). Neue Methoden zur histologischen Altersbestimmung menschlicher Hautwunden. Arbeitsmethoden der medizinischen und  naturwissenschaftlichen Kriminalistik. Bd. 20. Lübeck: Schmidt-­Römhild. Betz, P., Beier, G. and Eisenmenger, W. (1994). Pulmonary giant cells and traumatic asphyxia. International Journal of Legal Medicine 106: 258–261. Betz, P., Nerlich, A., Penning, R. and Eisenmenger, W. (1993). Pulmonary giant cells and their significance for the diagnosis of asphyxiation. International Journal of Legal Medicine 106: 156–159. Brinkmann, B., Fechner, G. and Püschel, K. (1984). Identification of mechanical asphyxiation in cases of attempted masking of the ­homicide. Forensic Science International 26: 235–245. Brook, O.R., Hirshenbaum, A., Talor, E. and Engel, A. (2012). Arterial air  emboli on computed tomography (CT) autopsy. Injury 43 (9): 1556–1561.

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Kernbach-­Wighton, G., Sprung, R. and Saternus, K.S. (2002). Zum Katecholaminspiegel bei Unterkühlung. 11. Frühjahrstagung der DGRM Region Nord, 2002, Potsdam. Kondo, T. and Ishida, Y. (2010). Molecular pathology of wound healing. Forensic Science International 203: 93–98. Legas Pérez, I., Falcón, M., Gimenez, M. et al. (2017). Diagnosis of vitality in skin wounds in the ligature marks resulting from suicide hanging. American Journal of Forensic Medicine and Pathology 38 (3): 211–218. Madea, B. (2020). Asphyxiation, Suffocation and Neck Pressure Deaths. Boca Raton FL: CRC Press Taylor & Francis Group. Madea, B. and Grellner, W. (2002). Vitale Reaktionen. Teil 1. Rechtsmedizin 12: 378–394. Madea, B. and Grellner, W. (2003). Vitale Reaktionen. Teil 2. Rechtsmedizin 13: 32–48. Madea, B., Doberentz, E. and Jackowski, C. (2019). Vital reactions – An updated overview. Forensic Science International 305: 110029. doi: 10.1016/j.forsciint.2019.110029. Epub 2019 Oct 31. Madea, B., Herrmann, N., Käferstein, H. and Sticht, G. (1994). Hypoxanthine in vitreous humor and cerebrospinal fluid – Marker of postmortem interval and prolonged vital hypoxia? Forensic Science International 65: 19–31. Madea, B., Saukko, P., Oliva, A. and Musshoff, F. (2010). Molecular pathology in forensic medicine  – Introduction. Forensic Science International 203: 3–14. Madea, B., Wagner, R., Markwerth, P. and Doberentz, E. (2017). Heat shock protein expression in cardiac tissue in amphetamine related deaths. Romanian Journal of Legal Medicine 25: 8–13. Maeda, H., Zhu, B., Ishikawa, T. and Michiue, T. (2010). Forensic molecular pathology in violent deaths. Forensic Science International 2003: 83–92. Müller, E., Eulitz, J. and Lobers, W. (1990). Zum Wert des Thyreoglobulinblutspiegels für die Diagnose von Strangulationen. In: B. Brinkmann and K. Püschel (Hrsg.), Ersticken, pp. 64–69. Berlin Heidelberg New York: Springer-­Verlag. Nikolic, S., Zivkovic, V., Jukovic, F. et al. (2009). Simon’s bleeding: A possible mechanism of appearance and forensic importance  – a prospective autopsy study. International Journal of Legal Medicine 123: 293–297. Orsos, F. (1935). Die vitalen Reaktionen und ihre gerichtsmedizinische Bedeutung. Beitrage Zur Pathologischen Anatomie 95: 162–237. Pakanen, L., Kortelainen, M.L., Särkioja, T. and Porvari, K. (2011). Increased adrenaline to noradrenaline ratio is a superior indicator of antemortem hypothermia compared with separate catecholamine concentrations. Journal of Forensic Sciences 56 (5): 1213–1218. Pedal, E., Mosmayer, A., Mallach, H.J. and Oehmichen, W. (1987). Luftembolie oder Fäulnis? Gasanalytische Befunde und ihre Interpretation. Zeitschrift fur Rechtsmedizin 99: 151–167. Pollak, S., Vycudilik, W., Reiter, C. et al. (1987). Großtropfiges Fett im Blut des rechten Ventrikels. Zeitschrift fur Rechtsmedizin 99: 109–119. Preuss, J., Dettmeyer, R., Poster, S. et al. (2008). The expression of heat shock protein 70  in kidneys in cases of death due to hypothermia. Forensic Science International 176: 248–252. Püschel, K. (1982). Vitale Reaktionen zum Beweis des Todes durch Strangulation. Habilitationsschrift: Univ. Hamburg.

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Ishida, Y., Nosaka, M., Shimada, E. et al. (2018). Forensic application of epidermal AQP3 expression to determination of wound vitality in human compressed neck skin. International Journal of Legal Medicine 132 (5): 1375–1380. Ishida, Y., Nosaka, M., Shimada, E. et al. (2018). Immunohistochemical analysis on Aquaporin-­1 and Aquaporin-­3 in skin wounds from the aspects of wound age determination. International Journal of Legal Medicine 132(1): 237–242. Ishikawa, T., Kondo, T. and Maeda, H. (2016). Innovative Concepts and Technology for Medico-­legal Investigation of Death: How to learn about human life from the deceased. The Medico-­legal consultation and Postmortem Investigation Support Center (MLCPI-­SC), Osaka, Japan, c/o Department of Legal Medicine, Osaka City University Medical Schook Asahi-­ machi 1-­ 4-­ 3, Abeno-­ ku, Osaka 545-­ 8585, Japan, Yoyodo Printing Kashida Ltd. 1-­2-­14, Gamou, Joto-­ku, Osaka 536-­0016, Japan. Jackowski, C. (2013). Special issue on postmortem imaging. Forensic Science International 225 (1–3): 1–2. Jackowski, C., Christe, A., Sonnenschein, M. et al. (2006). Postmortem unenhanced magnetic resonance imaging of myocardial infarction in correlation to histological infarction age characterization. European Heart Journal 27 (20): 2459–2467. Jackowski, C., Hofmann, K., Schwendener, N. et  al. (2012). Coronary thrombus and peracute myocardial infarction visualized by unenhanced postmortem MRI prior to autopsy. Forensic Science International 214 (1–3): e16–e19. Jackowski, C., Schwendener, N., Grabherr, S. and Persson, A. (2013). Post-­mortem cardiac 3-­T magnetic resonance imaging: visualization of sudden cardiac death? Journals of the American College of Cardiology 62 (7): 617–629. Jackowski, C., Sonnenschein, M., Thali, M.J. et al. (2007). Intrahepatic gas at postmortem computed tomography: forensic experience as a potential guide for in vivo trauma imaging. Journal of Trauma 62 (4): 979–988. Jackowski, C., Thali, M., Sonnenschein, M. et al. (2004). Visualization and quantification of air embolism structure by processing postmortem MSCT data. Journal of Forensic Sciences 49 (6): ­ 1339–1342. Jackowski, C., Warntjes, M.J.B., Berge, J. et al. (2011). Magnetic r­ esonance imaging goes postmortem: Noninvasive detection and assessment of myocardial infarction by postmortem MRI. European Radiology 21 (1): 70–78. Janssen, W. (1963). Riesenzellbildung bei Erstickung. Deutsche Zeitschrift für die Gesamte Gerichtliche Medizin 54: 200–210. Janssen, W. (1977). Forensische Histologie. Arbeitsmethoden der medizinischen und naturwissenschaftlichen Kriminalistik. Lübeck: Schmidt-­Römhild. Janssen, W. and Bärtschi, G. (1964). Vitale und supravitale Reaktionen der Alveolarzellen nach protrahiertem Sauerstoffmangel. Deutsche Zeitschrift für die Gesamte Gerichtliche Medizin 55: 47–60. Kauert, G., Schoppek, B. and Eisenmenger, W. (1990). Zur Frage der strangulationsbedingten kraniokaudalen Konzentrationsdifferenz biochemischer Parameter am Beispiel der Katecholamine. In: B. Brinkmann and K. Püschel (Hrsg.), Ersticken, pp. 53–57. Springer-­ Verlag, Berlin Heidelberg New York.

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to several months and even years. Concerning very short intervals, the issue of wound age is nearly identical with the issue of vitality. The forensic pathologist can be confronted with the estimation of wound age in association with murder, manslaughter, bodily harm with fatal consequences (primarily survived), accidents and further constellations. Most of the research results were gained by studying open wounds of the skin. There is little information on the time-­dependent alteration of blunt-­force injuries. The most difficult question is to differentiate between wounds inflicted shortly before and shortly after death. This coincides with processes in the late agonal stage and the early supravital period. Generally, various influencing factors have to be considered in the time course of wound healing, which is comparable with inflammatory processes (Box 19.2.1). The main questions for the forensic pathologist are as follows: • Was an injury caused during lifetime and not in the supravital or postmortem period (vitality)? • How is it possible to differentiate between vital and (early) postmortem induction? • How long was the survival time after the infliction of the injury (wound-­age estimation)?

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Püschel, K., Türk, E. and Lach, H. (2004). Asphyxia-­related deaths. Forensic Science International 144: 211–214. Richter, M. (1905) Gerichtsärztliche Diagnostik und Technik. Leipzig: Hirzel. Ritz-­Timme, S., Eckelt, N., Schmittke, E. and Thomsen, H. (1998). Genesis and diagnostic value of leucocyte and platelet accumuliations arround “airbubbles” in blood after veinous air embolism. International Journal of Legal Medicine 111: 22–26. Saukko, P. and Knight, B. (2004) Knight´s Forensic Pathology, 3rd ed. London: Arnold, pp. 136–221. Schulz, F., Buschmann, C., Braun, C. et al. (2011). Haemorrhages into the  back and auxiliary breathing muscles after death by hanging. International Journal of Legal Medicine 125: 863–871. Sebastian, R., Chau, E., Fillmore, P. et al. (2015). Epidermal aquaporin-­3 is increased in the cutaneous burn wound. Burns 41: 843–847. Shiotani, S., Ueno, Y., Atake, S. et al. (2010). Nontraumatic postmortem computed tomographic demonstration of cerebral gas embolism following cardiopulmonary resuscitation. Japanese Journal of Radiology 28 (1): 1–7. Sigrist, T. (1987). Untersuchungen zur vitalen Reaktion der Skelettmuskulatur. Beiträge zur Gerichtlichen Medizin 45: 87–101. Simon, A. (1968a). Vitale Reaktionen im Bereich der Lendenwirbelsäule beim Erhängen. Wiss Z Univ Halle 17: 591–597. Suyama, H., Nakasono, I., Yoshitake, T. et  al. (1982). Significance of haemorrhages in central parts of the tongue found in medicolegal autopsy. Journal of Forensic Science International 20: 265–268. Turillazzi, E., Vacchiano, G., Luna-­Maldonado, A. et al. (2010). Tryptase, CD15 and IL-­15 as reliable markers for the determination of soft and  hard ligature marks vitality. Histology and Histopathology 25: 1539–1546. Yen, K., Plattner, T. and Dirnhofer, R. (2005). Retrograde blood ­aspiration: A vital reaction. Forensic Science International 154 (1): 13–18. Yen, K., Thali, M.J., Ghayev, E. et al. (2005). Strangulation signs: initial correlation of MRI, MSCT, and forensic neck findings. Journal of Magnetic Resonance Imaging 22 (4): 501–510.

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In most cases, postmortem tissue samples are available for ­investigation purposes (skin or organ lesions). Therefore, for a long time, morphologically orientated techniques have been the ­methods of first choice and are still mainly used: • Routine histology is done with paraffin embedding of tissue samples and staining with haematoxylin and eosin (H&E). This method allows a rough differentiation of cell types and tissue structures. Immigrating granulocytes, macrophages or fibroblasts can frequently and easily be detected. For a special differentiation of substructures (e.g., connective tissue, ­ haemosiderophages),

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19.2  Wound-­Age Estimation: General Introduction and Methods Wolfgang Grellner

19.2.1 Introduction

Vitality and wound healing or wound-­age estimation are phenomena which are closely related to each other, in particular with regard to skin lesions. According to Oehmichen (1990), there is overlapping in the early phase of repair processes following trauma: alterations within the first 30  minutes can be regarded as both vital reactions and signs of wound healing. Both phenomena are central issues in daily forensic work as well as main research fields. Wound age describes the time interval between the infliction of an injury and the time of death. It can be considered as the survival time of the individual following a physical injury. Forensically relevant wound-­age periods can vary from minutes

Box 19.2.1  Factors influencing the time course of inflammation or wound healing. Local factors • Type and intensity of trauma • Severity and extension of damage • Type of tissue • Temperature • Circulation Systemic factors • Hereditary factors • Age • Sex • Nutritional state • Diseases • Endocrinopathy • Metabolic disturbances Exogenous factors Drugs Source: After Helpap (1987).

Table 19.2.1  Examples of wound-­age estimation by ­immunohistochemical methods in human-­skin lesions.

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Regular detection or marked reaction

Longest detection

Antigen/ marker

Earliest detection

TGF-­β1 TGF-­α

Minutes Approximately 10 minutes

30–60 minutes 30–60 minutes

Fibronectin

Approximately 10–20 minutes

>4 hours

IL-­1β IL-­6 TNF-­α

15 minutes 20 minutes 15 minutes

30–60 minutes 60–90 minutes 60–90 minutes

ICAM-­1 VCAM-­1 E-­selectin L-­selectin

50 minutes 30 minutes 30 minutes 30 minutes

Tenascin

2–3 days

From 5 days

Months

Collagen III

2–3 days

From 6 days

Months

Collagen V

3 days

From 6–7 days

Months

Collagen VI

3 days

From 6–7 days

Months

4–6 days

From 7 days

Months

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>2 hours >1.5 hours >1–1.5 hours >1.5 hours

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there are a variety of further staining methods (e.g., elastic van Gieson stain (EVG), Prussian blue stain). • Enzyme histochemical methods detect enzymes of fibroblasts at their location in the tissue section (e.g., esterase, acid phosphatase, adenosine triphosphatase (ATPase)). Formerly, they were used to define wounds with an age of about several hours. However, the methods proved to be too unreliable and showed a high rate of negative cases even after intervals of hours. Therefore, they did not find large dissemination and now seem to be obsolete. • Immunohistochemistry is the method of choice in modern investigations in vitality and wound age. This method serves for the identification of special substructures within tissues (antigen–antibody reaction) and has to be modified according  to individual issues. As a first step, it must be checked whether or not an antibody can be used on formalin-­fixed and paraffin-­embedded tissue sections, or if native frozen material is required. A further critical step is the dilution of antibodies and reagents which usually must be tested (the recommended dilution is often not suitable). Numerous parameters have been tested with respect to their time-­ dependent appearance in wound repair (extract in Table 19.2.1). Most of these factors are expressed within days after traumatisation. Modern markers such as cytokines cover the time span of minutes to hours. Apart from morphological methods, numerous attempts were made to establish biochemical methods, in particular for the evaluation of vitality (time intervals up to approximately 30 minutes). Such quantitative analyses can be performed in wound fluids or tissue extracts of wounds. Modern techniques are frequently based on immunological tests such as enzyme-­linked immunosorbent assay (ELISA). Due to considerable methodological efforts and high standard deviations of results, biochemical techniques have so far failed to achieve an essential breakthrough. Whereas the methods discussed so far serve for the detection of vitality and wound markers on the protein level, it is also possible to analyse the earlier stages of a reaction on the mRNA level. For this purpose, there are morphological methods such as in situ hybridisation and molecular biological techniques such as reverse transcription polymerase chain reaction (RT-­PCR) (review in Kondo and Ishida 2010). For special investigations (with modern markers), it is frequently necessary and also useful to collect not only wound material but also reference tissue from comparable (e.g., contralateral) uninjured regions. Selection of the appropriate method depends on the aims and the experience of the investigator (Table 19.2.2). In many cases, the scientific knowledge is available; however, it is not transferred to daily casework due to technical or financial reasons.

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Myofibroblasts Laminin HSPG Collagen IV Smooth muscle cell actin

Approximately 1.5 days Approximately 1.5 days 4 days 5 days

Months

7 days 11 days 12 days

Months Months Months

Months Months Months

Marker of macrophages RM 3/1 25 F 9 G 16/1 Epithelial basal membrane Fragments (laminin, HSPG, collagen IV, collagen VII) Complete restitution

4 days

From 13 days

8 days

From 21 days

13 days

From 23 days

Approximately 21 days

Keratin 5

19.2.3  Results and interpretation Histological and immunohistochemical studies can be evaluated qualitatively, semi-­quantitatively or quantitatively: • Qualitative detection means the presence or absence of special features or markers.

Complete staining of basal layer

HSPG, heparan sulphate proteoglycan; ICAM, intercellular adhesion molecule; IL, interleukin; TGF, transforming growth factor; TNF, tumour necrosis factor; VCAM, vascular cell adhesion molecule.

TRAUMATOLOGY AND VIOLENT DEATH

Level

Method

Cellular structures

Routine histology Special staining methods

Protein/antigen

Immunohistochemistry Quantitative biochemical methods

mRNA

In situ hybridisation Reverse transcription polymerase chain reaction (RT-­PCR)

• Semi-­quantitative detection requires a scale for the (relative)

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intensity of a histological phenomenon or an immunohistochemical staining. For example, the distribution of positive epidermal cells can be assessed semi-­quantitatively in separate epidermal layers based on per cent ranges (e.g., 50%). The degree of reactivity of other histological

structures (e.g., corium, vessels, sweat glands, subepidermal cells) may be classified as negative, slightly positive, moderately positive or strongly positive. • Quantitative studies include cell counts or counts of special signals, mainly in the sense of counts per microscopic visual field. For example, 10 or 15 fields are counted and then the average value is calculated for one field. Biochemical studies must strictly follow the test protocol for the appropriate marker. The abovementioned ELISA requires standard solutions with known concentrations and establishment of a standard curve. Statistical evaluation completes the biochemical (and morphological) analyses. It is not possible to present all results of wound-­age estimation published so far in this chapter. By means of the relatively simple routine histology (H&E and additional standard stainings), the following basic phenomena can be assessed: • Detection of (first) neutrophil granulocytes (approximately 20–30 minutes) (Figure 19.2.1). • Increase of granulocytes and macrophages (hours) (Figure 19.2.2).

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Figure 19.2.1  (a) Haematoma of the scalp with a wound age of many hours, showing infiltration of neutrophil granulocytes (H&E, 200×). (b) Haematoma of the scalp with a wound age of many hours, showing infiltration of neutrophil granulocytes (naphthol AS-­D chloroacetate esterase (NASDCE), 200×).

Figure 19.2.2  (a) Haematoma of the scalp with a wound age of many hours, showing infiltration of macrophages (immunohistochemistry with CD68, 200×). (b) Haematoma of the temporal muscle with a wound age of many hours to several days, showing infiltration of neutrophil granulocytes (NASDCE, 200×).

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• Detection of haemosiderophages (3  days) (Figures  19.2.3

detection of a marker with a special method). It is mostly not possible to give time intervals or maximum time spans. Furthermore, it must be remembered that many results in literature data are based on cell culture or animal experiments. A critical evaluation of such data is necessary. The highest reliability can usually be expected from studies with human autopsy material.

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and 19.2.4). A typical survey of further results with the help of modern immunohistochemistry is shown in Table 19.2.1. It is a characteristic feature of wound-­age estimation that – in comparison to literature data from experimental studies  – only a minimum wound age can be determined (earliest

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Figure 19.2.3  (a) Haematoma of the temporal muscle with a wound age of many hours to several days, showing infiltration of macrophages (immunohistochemistry with CD68, 200×). (b) Haematoma of the anterior thorax with a wound age of at least 3 days, showing infiltration of haemosiderophages (Prussian blue, 200×).

Figure 19.2.4  (a) Soft tissue between the ribs, showing granulation tissue in an older injury (H&E, 100×). (b) Soft tissue between fractured ribs in an older injury, showing granulation tissue mixed with fresh haematoma due to a blunt force injury (H&E, 100×). (c) Soft tissue between fractured ribs in an older injury with granulation tissue and infiltration of haemosiderophages (Prussian blue, 200×). (d) Soft tissue between fractured ribs in an older injury with cell-­rich scar tissue (H&E, 100×).

TRAUMATOLOGY AND VIOLENT DEATH

19.3.2 Molecular biology of skin wound healing

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Disruption of the epidermal cells releases prestored inflammatory cytokines such as interleukin-­1 (IL-­1) and tumour necrosis factor-­α (TNF-­α) from keratinocytes. After the extravasation of blood constituents, the resulting clot induces haemostasis and provides a matrix for the influx of inflammatory cells. Platelets also secrete growth factors and chemokines. These substances promote the recruitment of inflammatory leucocytes to the site of injury. Infiltrating neutrophils remove contaminating bacteria. Subsequently, monocytes are differentiated to macrophages, and recruited at the injured sites. Macrophages play an important role in augmenting the inflammatory response and tissue debridement.

Re-­epithelialisation, granulation tissue formation and neovascularisation Within hours of injury, the release of epidermal growth factor (EGF), transforming growth factor-­ α (TGF-­ α) and fibroblast growth factor (FGF) acts to stimulate epithelial cell migration and proliferation, resulting in the start of re-­epithelialisation. Upon completeness of wound closure, keratinocytes undergo stratification and differentiation to restore the barrier. The formation of granulation tissue and angiogenesis starts at the wound space approximately 4  days after injury. Macrophages not only augment inflammatory responses but also secrete angiogenic growth factors such as vascular endothelial growth factor (VEGF) and FGF. Neovascularisation is essential for the synthesis, deposition and organisation of a new extracellular matrix (ECM). Under the guidance of TGF-­ β and platelet-­ derived growth factor (PDGF), fibroblasts are phenotypically converted into myofibroblasts expressing α-­smooth muscle actin (α-­SMA).

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Inflammation

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Betz, P. (1994) Histological and enzyme histochemical parameters for the age estimation of human skin wounds. Int J Legal Med 107, 60–68. Betz, P. (1995) Immunohistochemical parameters for the age estimation of human skin wounds. Am J Forensic Med Pathol 16, 203–209. Grellner, W. (2002) Time-­dependent immunohistochemical detection of proinflammatory cytokines (IL-­ 1β, IL-­ 6, TNF-­ α) in human skin wounds. Forensic Sci Int 130, 90–96. Grellner, W. (2004) Zytokine und Adhäsionsmoleküle in der rechtsmedizinischen Vitalitäts-­und Wundaltersbestimmung. Aachen: Shaker. Helpap, B. (1987) Leitfaden der allgemeinen Entzündungslehre. Berlin, Heidelberg, New York: Springer. Kondo, T. and Ishida, Y. (2010) Molecular pathology of wound healing. Forensic Sci Int 203, 93–98. Oehmichen, M. (1990) Die Wundheilung. Berlin, Heidelberg: Springer. Oehmichen, M. and Kirchner, H. (1996) The Wound Healing Process – Forensic Pathological Aspects. Lübeck: Schmidt-­Römhild.

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Toshikazu Kondo

19.3.1 Introduction

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In forensic practice, wound-­age examination is one of the most important matters. When lots of wounds are found in an autopsy case, forensic pathologists have to determine the causal relationship between the cause of death and those wounds. Subsequently, forensic pathologists are always required to estimate how long before each injury was inflicted, and to judge which injury directly related to the death. This is called ‘wound-­age estimation’, a classical but still modern and indispensable topic. Wound-­age estimation is closely associated with skin wound healing. Skin wound healing starts immediately after injury and consists of three phases: inflammation, proliferation and maturation. These phases proceed with complicated but well-­ organised interaction between various tissues and cells. During the inflammatory phase, platelet aggregation at the injury site is followed by infiltration of leucocytes such as neutrophils, macrophages and T lymphocytes into the wound site. In the proliferative phase, re-­ epithelialisation and newly formed granulation tissue begin to cover the wound area to complete tissue repair. Angiogenesis is indispensable for sustaining granulation tissue. These histopathological consequences are evaluated as local vital reactions. In molecular pathology, the cell–cell interaction through bioactive molecules is an essential event during skin wound healing. Leucocyte recruitment and activation are regulated by adhesion molecules, inflammatory cytokines and chemokines, and the proliferation of granulation tissues with neovascularisation is well controlled by growth factors and extracellular matrices. Unfortunately, conventional histological and histochemical methods are not able to detect these molecules. Thus, immunohistochemical techniques opened a new field of wound-­age estimation.

Tissue remodelling Frameworks of collagen and elastin fibres replace the granulation tissue, followed by saturation with proteoglycans and glycoproteins; this is followed by tissue remodelling involving the synthesis of new collagen mediated by TGF-­β. Collagen remodelling is dependent on continued synthesis and catabolism of collagen by several macrophage-­derived matrix metalloproteinases.

19.3.3  Markers for wound age estimation Adhesion molecules The accumulation of leucocytes such as neutrophils and macrophages at the injured sites is a hallmark in the inflammatory phase of skin wound healing. In the initial step of leucocyte migration, adherence of leucocytes to endothelial cells through adhesion molecules is the most important event, implying that age adhesion molecules are sensitive markers for wound-­

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CXC chemokine ligand 8 (CXCL8), monocyte chemoattractant protein-­1 (MCP-­1)/CC chemokine ligand 2 (CCL2) and macrophage inflammatory protein-­1α (MIP-­1α)/CCL3 are also possibly useful as markers of wound-­age estimation (Table 19.3.2). In particular, a ratio of >50% for IL-­8  indicates a wound age of 1–4 days. A ratio of >30% for MCP-­1 or >40% for MIP-­1α could be detected in skin wounds with a wound age of at least 1  day (Kondo et al. 2002a).

Growth factors

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During skin wound healing, several growth factors are involved in angiogenesis and granulation tissue formation in the proliferative phase. VEGF and bFGF are potent angiogenic growth factors. The time-­dependent expression of both VEGF and bFGF was found in a murine skin wound model, suggesting that these growth factors can become useful markers for wound age estimation (Takamiya et al. 2002, 2003). Actually, VEGF was expressed on macrophages and myofibroblasts in human skin wounds, and VEGF-­positive ratios of >50% possibly indicated a wound age of 7–21 days (Table 19.3.2) (Hayashi et al. 2004). TGF-­α promotes the proliferation of epidermal cells. TGF-­β1 is produced by macrophages, fibroblasts, keratinocytes and platelets. TGF-­β1 helps initiate granulation tissue formation by increasing the expression of genes associated with ECM formation including fibronectin, the fibronectin receptor, collagen and protease inhibitors. Both TGF-­α and TGF-­β1 expressions were enhanced within the first hour after an injury, suggesting that they can be used in the estimation of wound vitality and wound age (Grellner et al. 2005).

Table 19.3.1  Adhesion molecules. Earliest

Latest

P-­selectin

A few minutes

E-­selectin

1 hour

ICAM-­1

1.5 hours

VCAM-­1

3 hours

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Cytokines have many biological functions in the immune, central nervous, endocrine and haematopoietic systems. Pro-­ inflammatory cytokines such as IL-­1, IL-­6 and TNF-­α contribute to the promotion of inflammatory reactions. Additionally, cytokines that act towards leucocytes as chemoattractants are called chemokines. These pro-­ inflammatory cytokines and chemokines were upregulated at both protein and mRNA levels at the injury sites, implying that they could become useful markers for wound-­age estimation (Kondo and Ohshima 1996; Mori et al. 2002; Lin et  al. 2003; Ishida et  al. 2004, 2006, 2008a). Several groups including ourselves have investigated the expression of these cytokines and chemokines in human skin wounds with known different ages. IL-­1α-­positive infiltrating cells such as neutrophils, macrophages and fibroblasts could be detected immunohistochemically. As shown in Table 19.3.2, a ratio of IL-­ 1α-­positive cells considerably exceeding 30% indicates a wound age ranging from 4 hours to 1 day (Kondo et al. 1999). Our immunohistochemical study suggested that chemokines such as IL-­8/

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estimation (Dreßler et  al. 1997, 1999a, 1999b). Actually, the strong immunopositive reaction for P-­selectin can be observed a few minutes at the earliest and 7 hours at the latest. The immunoreactivity of intercellular adhesion molecule-­1 (ICAM-­1), vascular cell adhesion molecule-­1 (VCAM-­1) and E-­selectin can be detected 1–3 hours at the earliest after injury (Table 19.3.1).

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ICAM, intercellular adhesion molecule; VCAM, vascular cell adhesion molecule.

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Table 19.3.2  Cytokines, growth factors and stress protein. Wound age

>30%

4 hours to 1 day

>50%

1–4 days

MCP-­1

>30%

1–7 days

MIP-­1α

>40%

1–9 days

VEGF

>50%

7–14 days

Ubiquitin

>30%

7–21 days

ORP-­150

>40%

7–21 days

COX-­2

>50%

8 hours to 2 days

IL-­1α IL-­8

Positive ratio

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COX, cyclooxygenase; IL, interleukin; MCP, monocyte chemoattractant protein; MIP, macrophage inflammatory protein; ORP, oxygen-­regulated protein; VEGF, vascular endothelial growth factor. Source: Based on Hayashi et al. (2004).

Stress proteins Ubiquitin (Ub) is a highly conserved protein with a molecular weight of 8500 and is present in all eukaryotes. Ub is a member of the heat shock protein family, which is rapidly induced by various types of stimuli such as hyperthermia and chemical or mechanical stress. Ub is also available as a marker for wound-­age estimation. Ub-­positive ratios, considerably exceeding 30%, possibly indicate a wound age of 7–14  days (Kondo et  al. 2002b) (Table  19.3.2). Moreover, oxygen-­regulated protein 150 (ORP-­150), a molecular chaperone, can modulate the intracellular transport of VEGF from the endoplasmic reticulum to the Golgi apparatus, eventually contributing to wound healing. Actually, an ORP-­150-­positive ratio of more than 40% possibly indicates a wound age of 7 days or more (Ishida et al. 2008b). The combined examination of ORP-­ 150 and VEGF may be more reliable for wound-­age estimation.

Cyclooxygenase Prostaglandin E2 (PGE2) and prostaglandin I2 (PGI2) are known to be pro-­inflammatory mediators. Cyclooxygenase (COX), the key enzyme, is required for the conversion of arachidonic acid to prostaglandins. Two COX enzymes, COX-­ 1 and COX-­ 2, are responsible for the production of prostaglandin H2 (PGH2), the first step in prostanoid biosynthesis. Although COX-­1 is constitutively expressed under physiological conditions, COX-­ 2 is

TRAUMATOLOGY AND VIOLENT DEATH

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Endothelial progenitor cells (EPCs)

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Peripheral blood of adults contains bone-­marrow-­derived cells with properties for endothelial differentiation. They are termed endothelial progenitor (precursor) cells (EPCs). EPCs are positive for certain cell surface markers, such as CD133, CD34 and low concentrations of VEGF receptor-­2 (VEGFR-­2). EPCs can participate in angiogenesis in ischaemic and damaged tissues. The author (Kondo) and team preliminary examined the appearance of EPCs in murine skin wound healing (Ishida et al. 2012a). Actually, EPCs can be detected in human skin wounds and the time-­dependent appearances of EPCs are applied to wound-­age determination (Ishida et  al. 2015a). The initial appearance of EPCs is a wound aged 2 days, and the EPC number is increased in accordance with wound age. In particular, the EPC number of over 20 indicates a wound age of 7–12 days (Ishida et al. 2015a). It is proposed that the detection of EPC would give significant information for wound-­age estimation.

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During the proliferative phase of skin wound healing, several different types of collagens are de novo synthesised in order to replace necrotic tissue and to rebuild skin structure. Betz and colleagues (Betz et al. 1992b, 1992c, 1993a, 1993b) performed a series of immunohistochemical studies on several types of collagens in human skin wound samples with various wound ages. In accordance with different wound ages, the typical morphological findings of strongly staining network-­like structures associated with fibroblasts for each collagen subtype were closely related with different wound ages: collagen III: about 2–3 days; collagens V and VI: >3 days; and collagen I: >5 days. Moreover, basement membrane components such as collagens IV and VII, laminin and heparan sulphate proteoglycan are useful for wound-­age estimation (Table 19.3.3). Fibronectin, a multifunctional cell adhesion protein, can support the adhesion and migration of fibroblasts, keratinocytes and endothelial cells (Hayashi et al. 2004). Fibronectin is presumed to be the most sensitive marker for wound-­age estimation, as evidenced by immunopositive reaction for fibronectin detected in wound samples aged a few minutes (Betz et al. 1992a) (Table 19.3.3). However, fibronectin-­positive reaction might be found in postmortem incised wounds of porcine-­ skin-­ like vital wounds (Grellner et al. 1998). Practically, forensic pathologists should pay more attention to the evaluation of fibronectin immunoreactions.

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a new population of bone-­ marrow-­ derived progenitor cells, expressed both leucocyte (CD45, CD13) and mesenchymal antigens (collagen I, fibronectin). Fibrocytes have potential for the differentiation of myofibroblasts, playing a crucial role in fibrosis and wound healing. In unwounded specimens, CD45+/Col-­I+ fibrocytes are not detected. Dual-­positive fibrocytes were initially detected in skin wounds with a postinfliction interval of 4 days, and their number increased in lesions with more advanced wound age. A fibrocyte number of over 15 strongly suggests a wound age of 9–14 days (Ishida et al. 2009).

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enhanced under inflammatory conditions. With regard to the practical applicability with forensic safety, COX-­2-­positive ratios, considerably exceeding a ratio of 50%, indicate a wound age of 8 hours to 2 days. Collectively, COX-­2 would be a useful marker for the estimation of early wound age (Ishida et  al. 2012b) (Table 19.3.2).

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Markers

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Table 19.3.3  Extracellular matrix.

Earliest

Routine

Latest

2–3 days

6 days

Months

Collagen V

3 days

6 days

Months

Collagen VI

3 days

6 days

Months

Collagen I

5 days

6 days

Months

Laminin in myofibroblast

1.5 days

6 days

Months

HSPG in myofibroblast

1.5 days

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Fibronectin

10–20 minutes

4 hours

Months

Collagen III

HSPG, heparan sulphate proteoglycan.

Aquaporins (AQPs) AQPs are a family of channel proteins that play important roles in water transport to maintain the osmolality in the body. AQPs are expressed in ubiquitous tissue and cells such as erythrocytes, epithelial and endothelial cells as well as keratinocytes. In the forensic field, recent studies implied that organ-­ specific AQP expression would be useful for the differentiation between fresh and salt water drowning (An et  al. 2010, 2011; Hayashi et  al. 2009). AQP1 and AQP3 are expressed by endothelial cells and keratinocytes, respectively. The detection of AQP1+ vessel area and AQP3+ keratinocytes would be informative for wound-­age determination. Moreover, there is objective accuracy of wound-­ age determination (Ishida et al. 2018a). Moreover, AQP3+ signals in keratinocytes are more evident in compressed regions of neck skin as compared to non-­compressed regions. Thus, AQP3 can be considered a valuable marker to diagnose the trace of antemortem compression (Ishida et al. 2018b).

Dendritic cells (DCs) DCs can play essential roles in innate and adaptive immune systems in various organs. Double positive cells of CD11c and HLA-­ DRα are determined as DCs. In human skin wounds, DCs are recruited into wound sites. Actually, the presence of DCs in a skin

19.3.4  Application of reverse transcription polymerase chain reaction analysis

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Forensic pathologists consider that the detection of mRNA is inappropriate in forensic autopsy samples because of mRNA degradation with an increase of postmortem interval. However, reverse transcription polymerase chain reaction (RT-­PCR) analyses can detect mRNA of certain molecules in forensic autopsy samples with early postmortem intervals. After incision in mice, local IL-­6 mRNA level peaked at 6 hours, whilst the peak levels of mRNA for IL-­1α, IL-­1β and TNF-­α occurred between 48 and 72 hours (Sato and Ohshima 2000). By contrast, IL-­10 is one of the anti-­inflammatory cytokines; IL-­10 mRNA started to increase at 15 minutes, indicating that IL-­10 is the most sensitive cytokine (Ohshima and Sato 1998). In another group, the expression of IL-­1β mRNA showed that significant increase occurred at > : 0–20

Figure 19.4.1  Ratio of bone formation and resorption during life.

maintenance 20–50

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Table 19.4.1  Bone-­healing process after fracture.

References and further reading

3 days

Increasing of circulating endothelial progenitor cells

≥3 days

Neovascularization and activation of fibroblasts

14–21 days

Increasing of chondrocyte and osteoblast activity

≥21 days

Bone maturation

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In the complex process of bone fracture healing, various biological factors (different hormones and hormone receptors such as receptors for parathyroid hormone (PTH) and PTH-­related peptide, vitamin D3, calcitonin receptor, estrogens and progestins, androgens, thyroid hormone, glucocorticoid, diabetes and insulin as well as a number of growth factors such as insulin-­ like growth factor, platelet-­ derived growth factor, vascular endothelial growth factors, fibroblast growth factor, transforming growth factor-­ß) and mechanical factors (e.g., mechanical stress, hydrostatic pressure) take a great influence on the differentiation of fibroblasts and chondrocytes and on the mineralization of the extracellular matrix and consequently on bone fracture healing. To ensure a rapid healing as mentioned in Table 19.4.1 and to avoid complications, fractures are stabilized by surgical internal or external fixation with plates and screws in case of dislocated or segmental fractures. Postfracture complication (necrosis, infection, infarction, instability) reduces and prolongs the healing process. A time frame for the healing process in these cases cannot be given.

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2 days

Amiethab, A. (2020). Fracture Healing. https://www.orthobullets.com/ user/574 Bahney, Ch., Zondervan, R., Allison, P. et al. (2018). Cellular Biology of Fracture Healing. https://doi.org/10.1002/jor.24170 Bilezikian, J., Raisz, L. and Rodan, G. (2002). Principles of Bone Biology, Vol. 1, S 8. Cambridge: Academic Press. Delling, G. (1997) Skelettsystem. In: Remmele W. (eds) Pathologie 5. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-­ 3-­ 642­59232-­4_4 Ghiasi, M., Chen, J., Vaziri, A. et  al. (2017). Bone Reports. https//:creativecommons.org/licenses/by-­nc-­nd/4.0/ Hempfling, H. and Krenn, V. (2016). Schadenbeurteilung am Bewegungssystem, Vol 1. De Gruyter. Klein, M., Bonar, F., Freemont, T. et al. (2011). Traumatic and circulatory alterations of bone. In: ARP Atlas of Nontumor Pathology, Bd. 9, p. 308, Virginia. Lang, G. (2013). Histotechnik, Praxislehrbuch für die biomedizinische Analytik, 2nd ed. Berlin: Springer. Madea, B. et al. (2015). Thanatologie. In: B. Madea (eds.), Rechtsmedizin. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-­ 3-­ 662-­ 43500-­7_3 Mommsen, U., Holzrichter, D. and Schumpelick, V. (1991). Frakturen. In: V. Schumpelick N. Bleese and U. Mommsen (eds.), Chirurgie, p. 684. Stuttgart: Ferdinand Enke Verlag. Sheen, J.R. and Garla, V.V. (2020, Jan). Fracture Healing Overview. [Updated 2019 Nov 25]. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing: https://www.ncbi.nlm.nih.gov/books/NBK551678/ Plenk, H. Jr (1989). Knochengewebe und Zähne. In: P. Böck (ed.), Romeis Mikroskopische Technik, 17th edn, p. 539; Urban und Schwarzenberg; München, Wien, Baltimore (ISBN: 3-­541-­11227-­1). Werner, M. (2016). Knochenfrakturen. In: K. Amann R. Kain and G. Klöppel (eds.), Pathologie. Berlin, Heidelberg: Springer. https://doi. org/10.1007/978-­3-­642-­04566-­0_44 https://www.unimedizin-­mainz. de/fileadmin/kliniken/virologie/Dokumente/methoden.pdf

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Burkhard Madea, Stefan Pollak, Annette Thierauf-­Emberger, Veronique Henn, Christoph Meissner, Manfred Oehmichen, and Peter Mygind Leth findings and described causes of events are consistent. This is an important task, especially in cases of, for example, child abuse. Primary causes of death due to mechanical violence include the following: • Destruction of vital organs (brain, spinal cord, lungs, heart). • Mechanical handicaps of the functioning of vital organs. • Bleeding to death. • Embolism (air embolism, fat embolism). • Suffocation. Mechanical destruction of the vital organs occurs, for example, in cases of multiple trauma caused by traffic accidents, falls from height or railway accidents. Mechanical handicaps of the functioning of vital organs are, for example, intracranial haematomas (epidural or subdural), bilateral pneumothorax (e.g., multiple rib fractures with collapse of the lungs), thoracic compression (Perthes thoracic compression) and pericardiac tamponade. In case where the victim survives, the primary trauma, typical secondary causes of death may be: • Infections (wound infections, sepsis). • Embolism. • Circulatory shock (haemorrhagic or traumatic shock). • Burn injuries.

Burkhard Madea

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In the area of forensic medicine, several types of wounds are differentiated according to their origin: blunt-­force injury, sharp-­force injury, gunshot wounds, strangulation, injuries due to heat and coldness, etc. (Table 20.1.1; Figures 20.1.1–20.1.9). Different sorts of trauma cause characteristic patterns of injury and it is often the task of the forensic pathologist to reconstruct the causal mechanism of the injury based on the wound pattern. A detailed description of wounds includes: the size and exact localisation of injuries (not just on the skull, but where on the skull – above or below the hat brim line, at the vertex, at the front or at the occiput) and, of course, a precise description of the single injuries concerning, for example: • Length. • Size. • Diameter. • Wound morphology (wound edges, wound surface, wound angle, adaptable wound or any tissue defects). • Discolouration (haematomas: colour, demarcation from the surrounding tissue). Furthermore, the wound morphology has to be compared and associated to the case history to reconsider whether injury

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Handbook of Forensic Medicine, Volume 1, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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Table 20.1.1  Different types of external violence, wounds and trauma.

Electrical injury

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Low pressure (Caisson’s disease), high pressure (high-­altitude disease) Natural electricity (lightning), technical electricity (electrocution) Electromagnetic and corpuscular radiation Organic/inorganic substances

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Hyperthermia (fire death, inhalation injuries, scalding burns, chemical burns), hypothermia

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Deprivation of food and fluids Abnormal low or high temperatures Barotrauma

Blunt force/sharp force: stab wounds, incised wounds, shot wounds, shotgun wounds, bruising Strangulation (hanging, ligature strangulation, manual strangulation), smothering, choking, mechanical asphyxia, oxygen deficiency in the air, drowning Starvation/dehydration

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Mechanical trauma Suffocation

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Figure 20.1.1  Different types of injuries (blunt-­force injury, stab wounds, shot wounds, arrow wounds) from an early 16th century book of surgery by Hans von Gersdorff (Feldbuch der Wundartzney, Straßburg 1528).

Figure 20.1.2  Different types of blunt force injuries. (a) Abrasions. (b) Patterned intradermal bruising caused by kicking with profiled shoe sole. (c) and (d) Laceration. (e) Subcutaneous haematoma of the chest.

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Figure 20.1.3  Sharp-­force injury. (a)–(c) Stab wounds. (d) Postmortem incised wound of the neck. (e) Suicidal incised wounds of the forearm and wrist.

Figure 20.1.4  Semi-­sharp force injuries. (a) Amputation of fingers by pincers. (b) Propeller injury by boat propeller. (c) Decapitation by a train.

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Figure 20.1.5  Gunshot entrance wounds. (a) With massive sooting. (b) With tiny burns from propellant flakes. (c) and (d) Contact wound with muscle mark. (e) Entrance wound with abrasion, but no grease staining (shot through clothing).

Figure 20.1.6  (a)–(c) Frost erythema by hypothermia.

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Figure 20.1.7  Injuries due to heat. (a) and (b) Peeling of the epidermis by scalding. (c) Soot deposits on the skin. (d) Postmortem shrinking of the skin. (e) Third-­degree burns, which were survived for some days.

Figure 20.1.8  Electrical injuries with electrical marks, electrical blisters and burns, and drying of skin in a case of death by lightning.

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Figure 20.1.9  Strangulation marks. (a) Manual and ligature strangulation. (b) and (c) Ligature strangulation. (d) Hanging.

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20.2.1 Introduction

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Blunt trauma is often contrasted with penetrating trauma, in which an object such as a knife or a bullet enters the body. The term itself is used to describe physical damage due to mechanical force applied either by the impact of a moving blunt object or by the movement of the body against a hard surface. These mechanisms result in the transfer of kinetic energy with consecutive tissue deformation and injury (mainly by compression, traction, torsion and shear stresses). In addition, inertial forces due to rapid acceleration or deceleration may act on the body. Blunt force injuries occur in criminal assaults (e.g., blows with a blunt or edged implement, punch or kick), physical child abuse, domestic violence, traffic and industrial accidents, suicides (e.g., a jump from a height, intentional railway collision) or accidental falls.

20.2.2  Blunt injuries to the integument Abrasions Abrasions are superficial injuries to the skin characterised by traumatic removal, detachment or destruction of the epider-

mis, mostly caused by friction and/or pressure. In so-­called tangential or brush abrasions, a lateral rubbing action scrapes off the superficial layers of the skin (e.g., if the body slides across a rough surface) and leaves a denuded corium, which is initially covered with serosanguineous fluid. In fresh grazes, the direction of impact can often be determined by abraded epidermal shreds, which remain attached to the end of the scrape. Later, the tissue fluid dries out and forms a brownish scab. If the lesion does not reach the dermis, it heals within several days without scarring. After detachment of the scab, the regrown epidermis initially lacks the pigmentation of the surrounding skin, so that the formerly abraded area appears light until melanin has been synthesised as a result of the exposure to UV-­B radiation. Infliction just before or after death results in a leathery (‘parchment-­ like’) appearance with yellowish-­ brown, reddish-­ brown or dark-­ brown discolouration (Figure  20.2.1). When investigating dead bodies, it should be borne in mind that abrasions are easily overlooked as long as the epidermis-­free areas have not changed their initial pink colour due to drying. The sole presence of an abrasion does not prove that it has been inflicted in life, unless it goes along with an underlying haematoma or with signs of wound healing. Another type of abrasion is caused by a vertical impact to the skin (so-­called pressure or crushing abrasion). In such cases, the injuring object may be reflected by the shape of the skin injury, so that the patterned abrasion can be regarded as

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example, handcuffs leave linear, much thinner train-­track-­like marks on the wrists. Under certain circumstances, the form and location of abrasions may be indicative of a special trauma; for example, crescent-­shaped fingernail marks and/or scratches in manual strangulation; strip-­ like, linear excoriations with subsequent parching in cases of hanging and ligature strangulation; seat-­belt marks in vehicle occupants after head-­on collisions or human bite marks (typically showing two semicircular bows with their concavities facing each other). In contact shots, the muzzle imprint is a patterned pressure abrasion reflecting the contours of the barrel end and other constructional parts situated near the muzzle (e.g., foresight, bolt spring rod). Stab wounds may be accompanied by a distinctive contusion mark, if the penetrating knife is vigorously pushed in up to the handle. Skin artefacts resulting from resuscitation procedures have to be taken into consideration when performing an external body examination. The forensic importance comes from the risk of misinterpretation, as some findings may suggest bodily harm or even homicide. To give a single example, fingernail marks and scratches located on the face and neck can simulate smothering or throttling even if they were inflicted during mouth-­to-­mouth resuscitation. Precordial abrasions are frequently seen after manual chest compression. Mechanical devices as used by professional health carers for providing circulatory support may leave a patterned imprint mark (e.g., from the suction cup and the pressure pad). In case of a traffic victim run over by a car, the body may be wedged between the vehicle’s underbody and the road surface. When dragged along, the tangential impact first causes brush abrasions followed by progressing loss of the exposed soft tissues and finally the underlying bones are ground forming plane boundary surfaces.

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an imprint of the causative object, often depicting edges or ­protruding parts (e.g., incisors and canines in bite marks). Other examples would be: excoriations inflicted with a bunch of keys, a pistol grip, a belt buckle or an externally threaded tube, a fall onto a metal grid or a blow against skin covered with a ribbed textile. Although this list is necessarily incomplete, most excoriations do not have a characteristic shape. The location of an abrasion always indicates the site of contact or impact. If the skin was affected before death, the resulting injury often consists of both an abrasion and a concomitant bruise (compare with Figure 20.2.1a). Photogrammetry can be used to obtain a three-­dimensional image of an abrasion and to compare the data with an object that may have caused the injury. Photogrammetry supplies reliable evidence even if the images are analysed later, and is not only suitable to assess abrasions, but also for the morphometrical classification of patterned contusions. Another method for an exact three-­dimensional documentation of curved wound surfaces is structured light three-­dimensional scanning. Fingernail scratches constitute a special group of abrasions and are particularly often found in sexual offences and attacks to the neck; they can be seen both on the victims and the offenders, if the victims defended themselves. Depending on the intensity of the abrading traumatisation, the appearance of such lesions may range from superficial reddening of the skin via shallow excoriations to deep, bleeding skin lesions extending down to the corium (Figure  20.2.2). Terminal epidermal tags indicating the direction of fingernail movement are easily lost and thus seen relatively seldom in surviving victims. Typical features of fingernail scratches are parallel abrasions (if two or more fingernails were raked across the skin simultaneously) of roughly equal width along their course, whereas, for

Figure 20.2.1  (a) Dark-­brown discolouration of a premortal excoriation with whitish shreds of epidermis indicating the direction of impact. Note the additional presence of bruises. (b) Brush abrasion with black particles from the asphalt surface rubbed into the skin on sliding along the road. The denuded corium shows reddish-­brown discolouration from drying.

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Figure 20.2.3  Mixed bruise consisting of intradermal (arrows) and subcutaneous (SC) haemorrhages. View from above (upper half of the image) and cut section (lower half).

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In light-­ skinned individuals, intradermal bruises may reflect the surface configuration of the impacting object. The skin squeezed into grooves will show intradermal bleeding, whereas the areas exposed to the elevated parts remain pale. Especially in falls from a height, in physical abuse or in traffic accidents, the texture of the clothing may produce a pattern of intradermal bruises corresponding to the weaving structure of a garment (Figure  20.2.4a) or the chain links of jewellery (Figure 20.2.4b). Patterned extravasations of this type are also seen in tyre-­tread marks, if an individual is run over by a wheel (Figure  20.2.5), and in bruises from vertical stamping with the ribbed sole of a shoe (Figure 20.2.6). Also, the coarse knitting pattern of a glove worn by the offender may be reflected as regularly arranged intradermal haemorrhages after a blow to the face. High pressure exercised on a piece of lace underwear may cause intradermal bleeding congruent with the gaps of the textile material. After a strangulation act, patterned intracutaneous haemorrhages may indicate that a similarly structured piece of clothing or a necklace had been interposed between the throttling hand and the victim’s neck (see Figure 20.2.4b). Some further causes for the formation of defined intradermal haematomas should also be mentioned: blows with a twisted clothes line or a rug beater causing negative imprints of the impacting object on the skin; kick with the knee to the cheek of a victim resulting in strip-­like bruises reflecting the depressions between the ribs of corduroy trousers and impact of the facial skin against a floor tile with a grooved profile. Groups of punctiform blood extravasations in the corium do not only occur as a result of vertical pressure exercised by objects with a profiled surface but also in tangential abrasions (e.g., the edge of a screwdriver sliding across the skin, scratches from fingernails and so-­called ‘coin rubbing’). The cause of such intradermal haemorrhages is the rupture of vessels due to shearing forces in the papillary layer of the dermis.

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Figure 20.2.2  (a) Fingernail abrasion on the lateral neck of a 33-­year-­old man who committed rape homicide 6 days before. (b) A partially healed injury caused by the victim in self-­defence.

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Contusions or bruises are extravasations of blood within the soft tissues originating from ruptured vessels as a result of blunt trauma (mostly caused by squeezing, but sometimes also by suction). In this context, only the contusions that are visible externally are considered. Textbooks usually differentiate between intradermal and subcutaneous bruises. In the first category, the haemorrhage is located directly under the epidermis, that is, in the corium. This kind of superficial haematoma is usually sharply defined and carmine red, whereas the more common bruises of the deeper subcutaneous layer have blurred edges and, at least initially, a bluish-­purple colour. Blood localised in the subcutis appears blue on the surface due to scattering processes in the dermis, as the blue wavelengths of the light are scattered (and thus reflected) to a greater extent than the red wavelengths. In practice, haematomas are often ‘mixed’ (i.e., they are composed of intracutaneous and subcutaneous portions) (Figure 20.2.3).

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Figure 20.2.4  (a) Patterned intradermal haemorrhages in the left shoulder region of a 49-­year-­old man who had been kicked. At the time of impact, the skin was covered with a knitted pullover. (b) Intracutaneous haemorrhages reflecting the links of the necklace, which was interposed during manual strangulation 1 day before (25-­year-­old woman).

Figure 20.2.5  (a) A 3-­year-­old child with tread marks on the right side of the head and neck after being run over by a car. Patterned intradermal bruises (arrows) depicting the grooved face of the tyre (b).

Groups of punctiform haemorrhages in the corium can also develop after the skin is exposed to negative pressure: typical examples for this phenomenon are sucking kisses on the neck (so-­called ‘love bites’, mostly associated with subcutaneous haematomas) and ‘cupping’. Reactive reddening of the skin due to local mechanical irritation is often accompanied by punctiform, intracutaneous haemorrhages. In the presence of additional structural tissue changes

such as patchy haematomas, post-­traumatic redness of the skin can be detected as late as 3 days after the incident. Petechial extravasations of blood due to congestion in cases of manual and/or ligature strangulation or traumatic asphyxia are discussed elsewhere (see Chapter  2.1). A further kind of skin petechiae not related to direct impact trauma may occur in victims of systemic fat embolism (FE), as explained elsewhere in this chapter.

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Figure 20.2.6  Left side of the face (a) showing both a subcutaneous haematoma (lower lid) and a patterned intradermal haemorrhage (zygomatic region) caused by a kick with the ribbed sole of a shoe (b).

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Figure 20.2.7  Anterior aspect of the trunk after removal of the skin at autopsy. Band-­shaped subcutaneous haematomas along the course of the seat belt (37-­year-­old front-­seat passenger injured in a head-­on collision).

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Subcutaneous bruises are usually non-­ patterned. Under favourable conditions, subcutaneous haematomas may show characteristic features allowing conclusions as to the circumstances of the injury, although – contrary to intracutaneous haematomas – no detailed and exact reproduction can be expected. However, in many cases, general statements as to the character of the causative implement are possible (such as that caused by a seat belt in Figure  20.2.7). A classical example for this type of injury is the tram-­like haematoma resulting from a blow with a rod, a stick, a broom handle, a pool cue or another elongated instrument. In retrospect, it seems remarkable that this distinctive injury pattern was described by Walcher as late as 1932. At the end of his paper entitled ‘On the local effect of blows with sticks, rods and similar objects with special emphasis on the presence of two track-­ like lines of hemorrhage’, the author wrote: ‘As this phenomenon has been ignored in the literature so far, it seems justified to address this issue’. Since then, the parallel tram-­like haematoma with an undamaged zone in between caused by a blow with a rod-­ like object is discussed in detail in every forensic textbook and atlas. Streak-­like subcutaneous haematomas are highly significant in child abuse (see Chapter 38.2). They are most common where rods, sticks, baseball bats and so on (i.e., objects with a round profile) were used, although the spectrum of potential tools is much wider and ranges from belts to ropes and electric cables (Figure 20.2.8). Because of their flexibility, the latter objects are often used as an open noose to deal a blow. A pliable striking tool such as a whip adapts to curved body parts causing a continuous

Figure 20.2.8  Tram-­like contusion (a) from a blow with an electric cable forming an open noose (b).

welt both on frontal and dorsal aspects of the affected region. Blows with a rigid implement cannot only produce parallel streaks but – on curved parts of the body – also oval haematomas to be distinguished from bite marks of similar configuration. Some authors emphasise another special form of subcutaneous haematoma, namely, the roundish bruises with a diameter of about 1–1.5 cm caused by local fingertip pressure (Figure 20.2.9).

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Figure 20.2.9  Grip marks on the medial aspects of the upper arms in a 35-­year-­old woman killed during physical abuse.

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itive traumatisation. Common sites of abusive bruises are the face, buttocks, trunk and arms. For details, see Chapter 38.2. orbital haematoma (‘black eye’) is induced either A peri-­ directly or indirectly as follows: 1. The most common cause is a direct blunt impact to the orbital region (e.g., by a fist blow or kick). Quite often, the contused skin is abraded and/or torn in the region of a bony support. The bleeding can also originate from adjacent injured structures (e.g., fracture of the nasal bone). 2. A second mechanism is that blood seeps down from a soft tissue lesion in the forehead to the loose tissue of the eyelids due to gravity (Figure 20.2.11a). 3. As a third alternative, a fracture of the base of the skull in the region of the thin orbital roofs is to be taken into consideration (Figure 20.2.11b,c). Depending on the causative mechanisms of formation, the time elapsing between traumatisation and manifestation of the discolouration differs: after direct blunt impact, haematomas in the eyelids develop almost instantly. In fractures of the orbital roofs, haematomas in the eyelids may be seen within 30 minutes after the trauma. If the haemorrhage originates from an injury to the soft tissue of the forehead, it takes at least 4 hours before discolouration of the eyelids occurs. Subcutaneous haematomas can shift or spread due to gravitation not only in the area of the forehead and orbita, but also in other regions with loose subcutaneous fatty tissue. In darkly pigmented individuals, contusions are sometimes invisible to the naked eye. In such cases, the use of infrared light sources can be helpful for diagnosing subcutaneous blood extravasations. Diaphanoscopy (also called transillumination) is another means to determine the presence of bruises which otherwise would remain undetected. When light is passed vertically through the skin from a halogen lamp, a lucent zone appears

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These ‘fingertip bruises’ or ‘six-­penny bruises’ are often seen in victims of child abuse and sexual assault. Apart from the roundish shape and minor size, grouped arrangement may indicate vigorous gripping or holding. Sites of predilection are the medial aspects of the upper arms where this form of haematoma may develop in elderly and helpless persons also as a result of normal manipulations such as manually supporting or handling patients. Human bites produce bruises forming two opposing arches (from the upper and lower jaws) with a pale centre; often the haematomas are combined with abrasions and/or lacerations, which may sometimes reflect individual teeth characteristics. Human bites are typically found in sexual assaults and child abuse. Of differential diagnostic significance are the love bites mentioned in the earlier text, especially as these haematomas are preferably located on the neck and can be mistaken for throttling marks. Distinctively shaped haematomas suggest that the injury was caused by an object having a similar appearance. This statement has to be qualified, however, as primarily uncharacteristic haematomas may undergo structuring later after being compressed over a prolonged period of time. Under such circumstances, pale areas or differences in intensity may develop within the skin discolourations, for example, after tying or applying a tight bandage. Victims of blunt force violence often sustain contusions from self-­defence, typically located on the ulnar aspects of the forearms and the back of the hands (Figure 20.2.10). A competent assessment of bruises is of utmost importance when differentiating between accidental and non-­ accidental (abusive, deliberately inflicted) injuries. In cases of physical child abuse, the main evaluation criteria comprise (1) any discrepancies between the story told by the carer and the findings in physical examination; (2) location, number and arrangement of bruises; (3) relation to motor development in early childhood; (4) any pattern typical of abuse (e.g., tram-­like contusions, slap injuries mirroring a handprint, bite marks); (5) different ages of infliction suggesting repet-

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Figure 20.2.10  Defence injuries on the hands of a 23-­year-­old woman killed by multiple blows with a hammer.

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around it whose diameter is narrower above a medium with a higher optical density (e.g., a subcutaneous haematoma). The examination is performed in a dark room with a small flashlight. In contrast to this, ultraviolet photography is not suitable to show dermal or subcutaneous haemorrhages, as the ultraviolet radiation is largely absorbed or reflected in the epidermis already, thus photographically ‘extinguishing’ tattoos and haematomas. However, ultraviolet photography is excellently suited to document bite marks, even if the bite injury itself has already healed. In certain body regions, haematomas may manifest themselves in a special manner. In elderly people with fair and atrophic skin, the extensor surfaces of the forearms and the back of the hands often show sharply defined, dark purplish blotches known as actinic or senile purpura, a common disorder attributed to chronic sun exposure resulting in damage to the dermal connective tissue (Figure 20.2.12). The blood extravasation into the cutis is caused by minor trauma, which often passed unnoticed. Actinic purpura must not be confused with defence injuries. Haematomas involving the anterior thoracic wall mostly omit the nipples and the surrounding areolae. The propagation of extravascular blood is obviously impeded by the special tissue texture of the nipple–areola complex. An extensive impact on the body, for instance, in a fall from a high bridge with subsequent impinging on the water surface, may produce a haematoma pattern depicting the contours of a bone underlying the contact area such as the femur. In such cases, the mechanism of haematoma formation is quite similar to that in tramline bruises from blows with a stick: The soft tissue overlying the bone is compressed by the impacting blunt trauma followed by an ultrafast sideward displacement of blood with consecutive extravasation on either side. Another peculiar finding worth mentioning is seen in the parietal region: When the head hair is pulled vigorously, mostly in child or spouse abuse, the scalp can be detached from the skull-

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Figure 20.2.11  (a) Laceration above the right eyebrow with concomitant bruising of the upper lid (33-­year-­old man who had fallen to the ground under the influence of drugs and alcohol). (b) Laceration of the left frontotemporal region with peri-­orbital haematoma due to fractures of the orbital roof (c) in a 79-­year-­old woman who had fallen down the stairs.

cap. The gap in between may be filled with blood which forms a subgaleal haematoma palpable in physical examination and typically accompanied by local hair loss. In general, bruises are regarded as a sign of vitality indicating that the contusion was inflicted prior to death. During life, the blood from ruptured vessels is forced into the soft tissue by active extravasation. Nevertheless, to a limited extent, postmortem formation of contusions is possible due to passive ooze of blood. In surviving victims, a deep bruise may not become apparent on the skin until several hours or even days later because of the slow percolation of free blood from the original site to superficial tissue layers. In a living/surviving person, the contusion undergoes a temporal series of colour changes usually proceeding from the periphery towards the centre. Initially, most subcutaneous bruises

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appear purple-­blue. As the haematoma resolves during the healing process, the haemoglobin released from the red blood cells is chemically degraded into other pigments such as haemosiderin, biliverdin and bilirubin. The colour changes  – usually in the course of several days – to green and yellow before it finally disappears (Figure 20.2.13). However, the rate of change is quite variable and depends on numerous additional factors, above all the extent of the bruise. Sharp or vague demarcation of the haematoma discolouration against the surrounding skin is no reliable criterion for the age of the lesion. Fresh subcutaneous haematomas appear often blurred through the overlying layers at first, the more so as they are frequently accompanied by local oedema in the first few days, which makes optical demarcation from intact skin all the more difficult. Thus, haematomas may show more pronounced contours and stronger colours after a few days than in the initial phase. Then, as the haematoma is absorbed and blanches, the margins become vague again (‘diffuse’). The statements contained in textbooks with regard to age-­ dependent colour changes of (subcutaneous) haematomas must be taken with caution. In view of the numerous variables (e.g., intensity of impacting force, vascularisation of affected tissue, vulnerability of local blood vessels, tissue density, volume of extravasation, individual structural differences), some authors even advise against macroscopical age determination by means of the colour quality: 1. Based on their own studies, Langlois and Gresham (1991) arrived at the conclusion that yellow is found only in haematomas with an injury age of more than 18 hours. Younger people developed a yellow discolouration sooner than persons over 65 years of age. 2. In three of 30 subjects with artificial haematomas, Tutsch-­ Bauer et al. (1981) found yellowish portions on the periphery already on day 1; on day 7, yellow was present in 93% of the

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Figure 20.2.13  A 47-­year-­old victim of domestic violence (blows and kicks) inflicted 2 days before. The haematomas are already turning yellow in the marginal parts.

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subjects. Around day 4, many of the subjects showed the colours blue, green, brown and yellow simultaneously, and on that day the haematomas also reached their maximum extension. In 1970, Lins and Hamper already recorded reflectance curves of artificial bruises and analysed the changes in colour in relation to time. They demonstrated that the discolourations in the orange-­yellow and bluish-­green range observed in haematomas slowly regressed within 4–6 days (as the bruised skin areas macroscopically blanched). Klein et al. (1995) measured reflectance using an ophthalmospectrometer in the spectral range of 430–700  nm in artificial bruises caused under standard conditions and in spectacle haematomas. At wavelength 580 nm (yellow), spectacle haematomas with an age difference of more than 2 days could be differentiated. A comparison of the reflectance behaviour over time showed an increase in reflectance in the spectral range of 540–580  nm (green/yellow) correlating with the observed blanching of the haematoma. The authors found that the reflectance curve was not only dependent on the age of the haematoma, but also on its intensity and the pigmentation of the skin. People who are not aware of Mongolian spots (slate grey naevi, congenital dermal melanocytosis) are prone to mistake this congenital skin condition for bruising. The spots are mostly located in the lumbosacral region and on the buttocks. Their occurrence is limited to early childhood. The prevalence is high in Asia and many other parts of the world. The skin discolourations are irregularly shaped and have a blue or blue-­grey hue. They are due to the presence of melanin-­containing cells in the dermis. The size of an intradermal or subcutaneous haematoma is not always indicative of the intensity of the force applied to the affected area. Elderly people with senile purpura or patients suffering from bleeding diathesis may get bruises from slight knocks or for other minor reasons. On the other hand, absence of an externally visible injury does not necessarily mean that there was no relevant trauma. Subcutaneous bruises of surviving victims are often followed by gravity shifting of the haemorrhage leading to a secondary downward movement and enlargement of the haematoma. This phenomenon has already been mentioned in the context of periorbital haematomas, but it is also seen in other body regions. Facial bruises due to fist blows can be taken as an example: Within a short period (several hours up to few days), the bluish skin discolouration may have spread to the previously unaffected neck (Figure 20.2.14). Secondary shifting of extravascular blood also applies to fracture haematomas. A special type of blunt injury to the soft tissues is frequently seen in pedestrians who were struck or run over by motor vehicles. Both the skin and the subcutaneous layer may be avulsed from the underlying fascia or bones by shearing forces so that a blood-­ filled pocket is formed, typically in combination with crush damage to the adjoining fatty tissue (Figure 20.2.15). If the covering skin is torn due to overstretching, an open décollement will result. Extensive stripping of the skin and subcutis from the muscles is also seen in high-­speed impact traumas (e.g., in victims of airplane crashes).

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support (e.g., the scalp, face, back of the hand, shins and elbows). When the force acts on the skin, the subcutaneous tissue is squeezed between the injuring object and the bony platform so that the integument is compressed and crushed until it tears and splits sideways. Differentiation of a laceration from a cut wound is not always easy. Lacerations with nearly cut-­like, smooth wound edges can be caused by the impact of an edged structure, the more so as the transition from blunt to cutting edges is fluent. Generally, lacerations are diagnosed with the help of the following criteria: 1. The edges of the tears are typically irregular and ragged (Figure 20.2.16). 2. The wound margins are crushed and bruised usually showing at least a narrow zone of abrasion. 3. Bridging tissue strands (vessels, nerves and fibres) may run from side to side of the wound (especially near the wound corners). 4. Even if the soft tissues are completely severed, the underlying bone does not show any sharp indentation. 5. In lacerations, the head hair remains usually intact, whereas it is often cut through if the wound is inflicted with a cutting object or a sharp-­edged heavy weapon. 6. Sometimes foreign material is deposited in the wound slit and helps to reveal the causative object. The wound slits may be linear (especially in blows with a narrow and edged instrument), Y-­ shaped or star-­ like. Semicircular or crescent-­shaped tears of the scalp are produced by blows with an edged instrument having a round contour such as the flat end of some hammers. If the impacting object hits the skin at an oblique angle, one of the edges will be ripped away resulting in unilateral undermining (undercutting, avulsion), which indicates the direction of the force. The abrasion surrounding the tear may correspond to the shape and dimensions of the impacting blunt instrument or – in the case of a fall to the ground – the area of contact. Janssen (1963) pointed out that the straight part of a linear laceration may correlate with the edge length of the object used. Already in 1938, Werkgartner had developed a subtle method to analyse lacerations and emphasised the significance of the impact marks left by the edges of the tool. Wound slits caused by the direct pressure of an edge are mostly associated with a narrow line of abrasion on the margins. Alternatively, any extensions radiating from the ends are not caused by direct pressure of the hitting edge, but by a wedge-­like effect. If the flat face of a blunt weapon impacts with the whole surface, the resulting laceration is typically irregular or stellate and accompanied by wide excoriations; straight and rectangular demarcations of the concomitant abrasion allow conclusions as to the size and contours of the striking surface. If the impacting object hits the skin at an oblique angle, one of the margins is not only crushed by the impacting edge, but also undermined. In lacerations from implements with a rectangular or square surface, the morphology varies depending on how the object impacts: with one edge, with a corner or vertically; this latter case produces a stellate laceration with an extensive abrasion reflecting the shape and size of the hitting surface. Blows with a rod-­like

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Figure 20.2.14  An 81-­year-­old woman who accidentally fell in her domestic environment. She sustained contusions of the left zygomatic and mandibular regions. The facial skeleton remained uninjured. The victim died one day later due to her pre-­existing ischaemic heart disease. The secondary downward movement of the facial haematomas led to an extensive bluish discolouration of the neck skin.

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Figure 20.2.15  Avulsion of skin and subcutis (closed décollement) on the lateral aspect of the left lower leg in a 73-­year-­old cyclist hit by a car. The blood-­filled pocket was exposed by a vertical incision.

Lacerations

Apart from excoriations and haematomas, lacerations are one of the most important sequelae of blunt traumatisation of the body surface. Lacerations are tears of the skin or of internal organs (see in the following text). On skin, they may be caused by blows from blunt objects (such as a hammer, a wrench, a crowbar, a whipped pistol, a rod, a toecap of heavy footwear or a fist); other lacerations are produced by impact from vehicles or by a fall to the ground. Lacerations occur most commonly in body regions where the integument directly overlies a firm bony base acting as

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Figure 20.2.16  (a) Laceration above the left eyebrow in a 22-­year-­old victim of a traffic accident (driver of a motor scooter, collision with an obstacle beside the road). The surrounding skin is abraded and parched reddish-­brown. (b) Lacerations in a 50-­year-­old man who had suffered kicks to the mouth.

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In victims of traffic accidents, injuries from breaking safety glass take a special position. Drivers, front-­seat passengers and rear-­seat occupants as well as pedestrians may suffer skin lesions from striking either the windscreen or a door window. The windscreen of modern cars is made of laminated glass which holds together when shattered, typically in the form of a spider web pattern. The tempered glass of side and rear windows disintegrates into numerous crumbly fragments with sharp edges. A breaking windscreen produces a great number of very shallow cuts, often in parallel-­serial arrangement. In addition, the whole contact area may show an epidermis-­free skin being prone to postmortem drying with subsequent reddish-­brown discolouration. Injuries from fragments of tempered glass are characterised by a so-­called ‘dicing pattern’. The small glass cubes of shattered door windows may penetrate the skin and get stuck in the wounds. The skin edges are mostly clean-­cut, but sometimes their margins are abraded. The presence and location of injuries from a side door window or a windscreen can help to differentiate between the driver and the front-­seat passenger. A forceful impact against the windscreen rim or an A-­pillar will cause a laceration, often accompanied by skull fractures. Seen from the criminalistic point of view, the transfer of traces is of special importance: On the one hand, the wound slits may contain splinters of glass. On the other hand, sparse blood staining of the windscreen and head hair entrapped amongst the glass fragments of a spider web fracture can be expected. In contrast to lacerations which are always located at the site of impact, skin severances due to overstretching are mostly found away from the area where the injuring object hits the body. Therefore, such tears lack any concomitant abrasion of the margins.

Figure 20.2.17  (a) Scalp laceration from a blow with an iron tube (b). The hair was removed at autopsy.

object such as a metal tube typically cause straight lacerations with wide zones of contusion and abrasion on the margins as well as diverging extensions on the ends (Figure 20.2.17). Occasionally, blunt injuries are inflicted by means of glass bottles or beer mugs. Blows with such objects are predominantly directed against the head, where they may cause swelling, haematomas or lacerations, but sometimes also lethal craniocerebral trauma. When the glass breaks upon impact, its edges may produce cut-­like wounds in addition.

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The head is a common target in assaults with blunt objects; other frequent causes of head injuries are traffic accidents, falls from a height and falls from a standing position. In fatal child abuse, the most common mode of death is head injury. The area of impact usually reveals injuries of the scalp or the facial skin; however, severe and even lethal traumatisation are not necessarily associated with local skin discolouration, marked swelling, excoriation and/or laceration. There may be no externally visible signs, especially in skin areas covered with cushioning hair and in cases of a fall onto a flat surface. An impact site on the vertex may suggest that the head sustained a blow, whereas in falls from standing positions, the scalp injuries are expected at the level of the brim of the hat. According to autopsy studies, there are many exceptions to this ‘rule’: Only ‘top of the head’ lacerations (in the dorsofrontal and parietal regions) are quite uncommon in falls from a standing position.

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20.2.3  Head injuries

Single or multiple linear fractures are caused either by a blow with a broad implement or an impact on a hard surface (e.g., due to a fall on the head). In such cases, the skull is deformed (flattening/indenting at the point of impact and outward bending/bulging in the periphery). The fracture lines originate where the bone is bent outward and therefore is exposed to traction forces exceeding the limits of the bone’s elasticity; from these extruded parts of the skull, the fractures extend towards the area of impact, but also in the opposite direction. For this reason, either of the ends is often in congruity with the impact injury of the scalp (local haematoma with/without concomitant abrasion and/or laceration). Several fracture lines may radiate outward from a central point of impact where the skull is often depressed and/or shattered to pieces forming a spider’s web or mosaic pattern consisting of radiating and circular fractures (Figure  20.2.19). The sequence of skull injuries may be determined according to Puppe’s rule: A later fracture does not cross a pre-­existing fracture line, but terminates when reaching an earlier one. Before fusion of the cranial sutures (i.e., in children and young adults), a fracture may travel along the seam resulting in diastasis (diastatic fractures, Figure 20.2.20). If a gaping fracture runs from one side of the cranial base to the other (mostly after lateral impact or side-­to-­side compression), this transverse type is called a hinge fracture (Figure  20.2.21) because of the independent movement of the front and rear halves of the skull base. Longitudinal fractures of the base of the skull frequently occur due to a fall on the occiput; in such instances, the linear fractures typically run through the posterior fossa either ending near the foramen magnum (Figure  20.2.22) or extending to the floor of the middle and anterior fossae. Alternatively, longitudinal fractures of the base can also be produced by impaction of the frontal region. Depending on its course and location, a base fracture may be followed by several clinical signs: bleeding from the ear (in fractures

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Skull fractures

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These may involve the cranial vault, the base of the skull and the facial skeleton. Though the presence of a skull fracture indicates severe traumatisation, the fracture itself rarely threatens the victim’s life. There are several types of skull fractures to be distinguished.

Figure 20.2.18  Inguinal regions of a 28-­year-­old female front-­ seat passenger fatally injured in a high-­speed head-­on collision. The victim was wearing a seat belt. Note the stretch-­mark-­like tears located symmetrically in both groins and just above.

Figure 20.2.19  Left parietal region of a 64-­year-­old man who had fallen down the stairs. The impact site shows a spider’s web fracture.

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Figure 20.2.20  (a) Skullcap of a 47-­year-­old man who died from craniocerebral trauma after a fall on the occiput under the influence of alcohol showing a linear diastatic fracture along the sagittal suture. The star indicates the site of impact. (b) Space-­occupying unilateral subdural haematoma covering the right hemisphere.

Figure 20.2.21  Transverse (side-­to-­side) hinge fracture dividing the base of the skull into a front and a rear half (61-­year-­old man killed in an airplane crash).

Figure 20.2.22  Base of the skull with a linear midline fracture in the posterior fossa extending down to the foramen magnum sustained in an unobserved fall on the occiput. Secondary fractures of the orbital roofs (still covered by the dura, which is dark coloured from underlying blood) can be seen in this 88-­year-­old man (status following repeated myocardial infarction and bypass surgery).

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of the temporal bone with concomitant haematotympanum and rupture of the eardrum, Figure 20.2.23); bleeding from the nose and mouth (in fractures involving paranasal sinuses, which provide a communication with the nasopharynx); periorbital haematoma (from fractures of the orbital roofs; compare with Figure 20.2.11c); leakage of cerebrospinal fluid coming out of the nose or the ear (if the dura is injured along the fracture) and bacterial infection of the meninges (by spread from the nasal cavity, the paranasal sinuses and the middle ear, especially when the fracture is accompanied by a tear of the dura). Some special types of skull fractures are mentioned briefly. A ring fracture is located in the posterior fossa and encircles the foramen magnum. It occurs in a fall from a height onto the victim’s feet or buttocks, so that the cervical spine is driven into the skull. Another mechanism takes effect in high-­speed head-­on collisions: the frontal impact abruptly decelerates the vehicle and the trunk of belt-­restrained passengers, whereas the head continues to move in the previous direction due to inertia resulting in hyperextension/hyperflexion of the craniocervical junction. Ring fractures of the skull base are also seen in motorcyclists. Bone impressions and depressed fractures are always localised at the point of impact where the head is struck with an object having a relatively small surface area such as a hammer, a protruding corner of a piece of furniture or another edged implement. The outline of a clean-­cut defect in the outer table may reproduce the shape and size of an edged instrument, for instance, in cases of pistol-­whipping. If only limited force is applied, the depressed fracture can be restricted either to the outer or, less often, to the inner table of the skull (the latter with inward displacement of the bone fragments; Figure  20.2.24). A depressed fracture from a blow striking the skullcap at an angle may be concentrically

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Figure 20.2.23  (a) Bleeding from the right ear in a 79-­year-­old woman after a fall down the stairs. (b) Fracture of the base of the skull running through the right petrous bone.

Figure 20.2.24  Impression fracture of the skullcap in a 26-­year-­ old man after blows with a heavy monkey wrench. (a) The outer table of the parietal bone shows only minor depression. (b) Two pieces of the inner table are displaced inward protruding into the cranial cavity.

terraced. The bone fragments of a depressed fracture are often dirt-­stained indicating an impact of an object with equal depositions on its surface (e.g., paint, grease, rust, soot; Figure 20.2.25). The diaphanoscopic postmortem examination of blunt impact injuries to the head sometimes reveals non-­diaphanous regions deriving from intraossary haematomas (‘bone bruising’). Their location coincides with corresponding injuries of the scalp such as contusions and lacerations so that conclusions can be drawn as

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A fracture of the base of the skull involving an air sinus (e.g., in the sphenoid bone) provides a communication between the cranial cavity and the nasopharynx permitting the passage of blood into the airways. If the victim has lost consciousness, aspiration of blood into the lungs will be the consequence, possibly followed by fatal asphyxiation (Figure 20.2.26). The same complication can be seen after blunt traumatisation of the visceral cranium. In contrast to adults, the flat cranial bones of infants are much more flexible. They are separated from each other by unossified and movable sutures. Therefore, fracture lines mostly end at sutures. If, however, a suture is crossed, the sections of the fracture are typically displaced laterally.

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to the area of impact. Hole fractures from bullets perforating a flat bone of the skull are mostly roundish and clean-­cut at the site of entrance, but bevelled out in a crater-­like manner at the exit site (see chapter 21.2, Handbook of Forensic Medicine, 2e). Blunt force applied to the occiput, mostly as a consequence of a fall on the back of the head, frequently causes independent fractures of the anterior cranial fossa such as cracks of the thin orbital roofs (secondary fractures) at the site of the contrecoup (compare with Figure 20.2.22).

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Common causes of facial trauma are vehicle crashes, interpersonal violence (e.g., blunt assaults such as blows from fists or objects, kicks), falls as well as industrial and sports accidents. Due to the use of seat belts and airbags, the incidences of maxillofacial trauma in drivers and front-­seat passengers have decreased considerably. The same is true for motorcyclists protected by helmets. Although most facial injuries are not life-­threatening by themselves, they are often associated with severe craniocerebral trauma. Besides, they may be followed by lethal complications, for instance, when the airways are obstructed either by bleeding or by swelling of surrounding tissues. In victims who lost consciousness due to additional brain concussion or contusion, death frequently results from blood aspiration with consecutive asphyxia. Apart from soft tissue injuries such as abrasions, bruises, lacerations of the facial skin and oral mucosa (Figure 20.2.27), facial trauma can also involve bones and teeth. Bone fractures are

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Figure 20.2.25  (a) External view of a cranial vault (after removal of the scalp) showing depressed fractures with blackish soiling of the bone fragments (detailed view). The foreign material originates from the underside of a rail vehicle (85-­year-­old man who suffered from dementia and was run over on a railway track). (b) Internal view of the cranial vault.

Figure 20.2.26  Blood aspiration following fractures of the skull base in a 16-­year-­old suicide jumping from a height. (a) Frontal aspect of the overinflated lungs with dark spots indicating subpleural blood infiltration. (b) Trachea and main bronchi filled with blood. (c) Cut surface of the pulmonary tissue showing dark foci of aspirated blood.

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A space-­occupying bleeding under, over and/or between the brain membranes is followed by local displacement of the brain and raised intracranial pressure with concomitant flattening of the cerebral hemispheres. Intracranial haematomas as well as traumatic brain swelling, which often accompanies head injuries, may result in transtentorial (uncal) herniation (in cases of supratentorial mass lesion, Figure 20.2.28) and/or herniation of the cerebellar tonsils which are forced into the foramen magnum leading to compression of the brainstem with secondary damage and failure of the medullary respiratory centres. From the clinical and forensic points of view, the possible occurrence of a so-­called ‘lucid’ or ‘latent’ interval has to be mentioned. After initial unconsciousness (due to cerebral concussion), there may be a symptomless period of several hours or even days before the victim becomes comatose again because of the increased haemorrhage leading to elevated intracranial pressure. Epidural (extradural) haemorrhages are located between the skull and the underlying dura mater which is stripped from the bone by bleeding from a torn vessel (Figure  20.2.29). Epidural haematomas have a typical disk-­or lens-­shaped appearance, which can be demonstrated by means of premortem or postmortem computed tomography (CT) scans (Figure  20.2.30). The most common site is the temporal and the adjacent parietal region where the branches of the middle meningeal artery are easily lacerated in the course of a transecting fracture line. Since the well-­adherent dura has to be avulsed from the bone, epidural haematomas originate more often from arterial bleeding than from venous bleeding (e.g., due to a torn dural sinus). In the great majority, an extradural haemorrhage is associated with a cranial fracture.

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mostly accompanied by local swelling and haematoma, deformity of the face, abnormally movable fragments and bleeding from the nose and/or mouth. Commonly injured bones include the nose, maxilla, mandible, zygoma, frontal bone and other bony structures forming the orbit. Traditionally, the midface fractures are categorised according to their location. Le Fort described three main types which are still used to classify these injuries. Le Fort I fractures run horizontally through the maxilla which is separated from the palate. Le Fort II fractures involve both halves of the maxilla by running symmetrically upward to the orbital rim and crossing the midline through the nasal bones so that the fracture’s route is comparable with a pyramid. Le Fort III fractures are characterised by a transverse craniofacial disjunction at the level of the orbital floors. Nevertheless, in practice, fracture patterns do not always follow the system introduced by Le Fort. Fractures of the mandible account for a high percentage of all facial fractures especially amongst young males. They mainly occur due to vehicle accidents, assaults, work-­related traumas, falls and sports accidents. Mandibular fractures may be simple (closed), compound (open) or comminuted. The anatomical regions frequently affected are the symphyseal area, the body and the mandibular angles, the right and left rami as well as the condylar and coronoid processes. In victims of physical child abuse, the face is a common target of blunt traumatisation, for example, by bitch-­slaps or other kinds of beating. Apart from local erythema with associated swelling and haematomas often forming a handprint, the affected children frequently show bruises and lacerations of the lips, a tear of the frenulum, damage to the teeth (avulsion from the sockets, loosening, breakage) and injuries to the ear (contusion of the pinna, eardrum rupture from barotrauma following a direct blow).

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Figure 20.2.27  Blunt facial trauma in a 60-­year-­old man due to multiple blows to the oral region. (a) Haematoma discolouration of the lips and nose. (b) Inner aspect of the upper lip with lacerations and bruising.

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Figure 20.2.28  Blunt craniocerebral trauma caused by interpersonal violence. The 35-­year-­old woman died after a lucid interval of several hours. (a) The unilateral subdural haematoma led to a shift of the midline structures from right to left; the close-­up view shows a torn bridging vein (source of bleeding). There was no skull fracture. (b) Underside of the cerebral hemispheres with transtentorial (uncal) herniation (arrows) and no cerebral contusions.

Figure 20.2.29  Death from a space-­occupying extradural (epidural) haematoma in a 49-­year-­old man who had hit the ground with the left side of the head and sustained a linear parietotemporal fracture. (a) Horizontal section showing the left-­sided epidural blood accumulation. (b) View from above after removal of the skullcap with the haematoma adhering to the outer surface of the dura mater. (c) Base of the skull still covered with the dura, which is detached by the extradural haemorrhage located in the left middle fossa. (d) Base of the skull after removal of the dura.

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An antemortem traumatic extradural haematoma must not be confused with a postmortem epidural thermal haematoma. The latter is seen in severely burnt bodies with a (at least partially) charred skull. It is caused by a heat-­induced shift of bloody fluid from the diploe and the venous sinuses. The thermal haematoma has a crumbly or fatty appearance and a brick-­red or brown colour (see Chapter 23.2). Subdural haematomas are intracranial bleedings located beneath the dura mater and above the arachnoid. Most often the haemorrhage arises from the tearing of overstretched bridging veins that traverse the subdural space between the surface of the cerebral hemispheres and the superior sagittal sinus (Figure  20.2.31, compare with Figure  20.2.28a). Other possible sources of subdural bleeding are injuries to venous sinuses or to the cerebral parenchyma (such as cerebral contusions with concomitant laceration of the arachnoid). The subdural haemorrhage usually covers one cerebral hemisphere in a cap-­ like manner from the parasagittal area via the lateral surface down to the basal fossas; on a horizontal section, it appears as a sickle-­ shaped accumulation of blood. In contrast to epidural haematomas, subdural haemorrhages are often not associated with skull fractures; additional damage to the brain tissue may also be absent. A high percentage of subdural bleedings is caused by acceleration or deceleration of the head, for instance, in falls when the head impacts on a hard surface, but also in traffic accidents and physical child abuse (battered-­ child syndrome and shaken baby syndrome, see Chapter 38.1). Apart from acute and subacute subdural haemorrhages (Figure 20.2.32), there are cases of prolonged haematoma

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Figure 20.2.30  CT scan of a 39-­year-­old man showing a large left-­sided extradural haematoma with concomitant shift of the midline to the right and brain swelling. The victim was hit by a fist blow to the face and fell on the back of the head. The extradural haematoma was associated with frontal contusions. The patient underwent neurosurgical craniotomy with removal of the space-­ occupying blood accumulation and survived with only minor neurological deficits.

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Figure 20.2.31  Acute subdural haematoma in a 44-­year-­old epileptic after an apparent fall to the ground showing a reflected dura with adhering blood accumulation on its (a) underside and (b) injured bridging veins. The immediate cause of death was aspiration of gastric contents.

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formation and organisation, mainly in elderly people and sometimes without a history of previous traumatisation. Such chronic subdural haematomas typically consist of brown and gelatinous blood accumulations adherent to the meninges and sometimes covered with a tough membrane. Traumatic subarachnoid bleeding may result from damage to the cortex such as brain contusion (e.g., contrecoup lesions, Figure 20.2.33), from penetrating injuries to the brain and as a consequence of vessel tears within the subarachnoid space. An extensive haemorrhage on the ventral surface and around the brainstem may arise from a laceration of an artery belonging to the circle of Willis or from another great vessel (such as a torn basilar and vertebral artery). In sudden deaths without any preceding trauma, subarachnoid haemorrhage is most often due to the spontaneous rupture of a so-­called berry aneurysm, which is typically located at bifurcations and branches of cerebral arteries belonging to the ‘circle of Willis’ (see chapter 34.3, Handbook of Forensic Medicine, 2e). The blood extravasation usually fills the basal cisterns and then spreads laterally over the cerebral hemispheres.

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Figure 20.2.32  Right-­sided subacute subdural haematoma (survival time: 4 months) in an 89-­year-­old woman who had suffered a craniocerebral trauma due to a fall down the stairs. (a) Reflected dura with reddish and brownish gelatinous adhesions on the underside. (b) Basal surface of the brain showing residues of former cerebral contusions (frontal and right parietal lobes).

Cerebral injuries ‘Concussion of the brain’ is a clinical diagnosis which means a disorder of cerebral function following immediately upon a (blunt) head injury. It is usually characterised by a transient loss of consciousness (initial coma); it is often combined with retrograde/anterograde amnesia and vegetative signs such as nausea and vomiting. In mere concussions, the unconsciousness lasts only for a relatively short time (less than 1 hour) and the brain tissue does not show any evidence of structural damage.

Figure 20.2.33  Contrecoup lesions of the brain covered with subarachnoid haemorrhage (frontal poles and left temporal lobe). The impact site was located in the right occipital region of this 81-­year-­old woman hit by a car, who survived for 3 days.

Nevertheless, even a simple cerebral concussion may be followed by the victim’s death, if the head trauma is joined by interfering mechanisms (for instance, drowning or aspiration of gastric contents during unconsciousness). Cerebral contusions are traumatic lesions of the brain frequently seen in the cortex and sometimes extending into the

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Figure 20.2.34  Extensive cerebral contusions in a head trauma due to a fall from a height (32-­year-­old man who died 4 days after an industrial accident). The impact site was located in the right parietal region. (a) Contralateral contrecoup lesions are mainly on the left frontal lobe. (b) Close-­up view of the contusion haemorrhages.

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Figure 20.2.35  Cut surfaces of brain areas showing contusion injuries (cortical haemorrhages at the gyral crests).

underlying white matter (Figure 20.2.34). Fresh contusion haemorrhages are mostly located on the crests of the gyri and composed of grouped streak-­like or punctate blood extravasations (Figure 20.2.35). The cortical lesions are often covered with subarachnoid haemorrhage. In contrast to cerebral contusions, the term ‘laceration’ means a major destruction of the anatomical context (for instance, mechanical separation of the tissue due to bone fragments or penetrating bullets). In the case of survival, the contusion haemorrhages are reabsorbed and assume a yellowish-­ brown appearance forming cystic scars. These remote contusions are often referred to as ‘plaque jaune’ (Figure  20.2.36, compare with Figure 20.2.32b).

Due to the injuring mechanism, most cerebral contusions occur in brain regions that are directly opposite to the point of impact. This contrecoup type of contusion is classically caused by a fall on the occiput, when the moving head is suddenly decelerated with the consequence that the inlying brain is damaged due to inertia. Contrecoup lesions hardly ever occur in early childhood, which can be explained by the age-­related pliability of the skull. In falls on the back of the head, the contrecoup areas of the brain (poles and undersurfaces of the frontal and temporal lobes) are subjected to an ultrashort negative pressure (‘cavitation’) resulting in vessel ruptures and cortical haemorrhages. Alternatively, the so-­called coup contusions arise at the area of

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prolonged coma and a fatal outcome even in the absence of an intracranial mass lesion. Macroscopically, DAI is characterised by small focal haemorrhages and salient cerebral swelling. Cerebral oedema is a frequent finding in significant head injuries. The formation of oedema is due to an increase in the fluid content of the brain, predominantly in the white matter. Post-­ traumatic oedema may be generalised (diffuse) or related to focal tissue damage (e.g., adjacent to an area of cerebral contusion or laceration). At autopsy, the weight of the brain is increased; the gyri are pale and flattened with shallow sulci in between. From the pathogenetic point of view, oedema is attributed to a heightened vascular permeability which in turn may be worsened by additional hypoxia. As with space-­occupying bleedings such as subdural or epidural haematomas, brain swelling is a common cause of raised intracranial pressure. The enlarged volume of the oedematous brain leads to a displacement of cerebral tissue downward through the midbrain opening resulting in grooving of the unci and/or hippocampal herniation. Expansion of the subtentorial brain leads to herniation of the cerebellar tonsils which are forced into the foramen magnum. Herniation with concomitant compression of the brainstem may be followed by secondary haemorrhages (localised in the midbrain and pons) and finally by lethal dysfunction of vital centres. Unilateral or bilateral pressure on the midbrain impairs its blood supply followed by oedema, necrosis and haemorrhage in the affected brain tissue (Figure  20.2.37). Multilocular or confluent haematomas resulting from secondary alterations must be differentiated from spontaneous cerebral bleedings, the more so as the latter ones are frequently located in the pons or elsewhere in the brainstem.

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impact due to the local deformation and compression of the brain. Even severe coup and contrecoup injuries are not necessarily associated with skull fractures. In victims with both coup and contrecoup lesions, the degree of contrecoup damage is usually more pronounced. Fracture contusions are localised in topographical correspondence to fracture lines and/or depressed fractures. In gunshot injuries to the head, cortical haemorrhages are frequently observed far away from the actual wound channel. This phenomenon can be explained by intense pressure variations induced by the energy release of the bullet when passing through the braincase. The cerebral contusions are mainly located at the gyral crests of the basal brain surfaces (see Chapter 21.2). Severe head traumas are often followed by post-­traumatic epilepsy, especially in victims with penetrating head injuries, depressed skull fractures, dural tears, intracranial haematomas and structural brain damage. Early post-­traumatic seizures occur within the first week, whereas late post-­traumatic epilepsy can develop at any point thereafter. Diffuse axonal injury (DAI) and the accompanying tissue tear haemorrhages are considered a consequence of shear and tensile strains from sudden acceleration/deceleration or rotational movements of the head. Overstretching of the nerve fibres in the white matter leads to axonal injury varying from temporary dysfunction to anatomical transection, the latter being followed by microscopically visible club-­shaped retraction balls on the axons. The sites of predilection include the corpus callosum, the parasagittal white matter, the superior peduncles and the rostral brainstem. In the course of the repair process, microglial cells proliferate in the areas of axon damage. In victims of substantial head injuries, especially after traffic accidents, diffuse axonal injuries may be responsible for

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Figure 20.2.36  Old (remote) cerebral contusions as incidental autopsy findings. Brownish discolouration (due to residual haemosiderin) and cystic alterations/defects at the gyral crests of the former contrecoup areas were seen in (a) the left temporal lobe and (b) the base of the frontal lobes with involvement of both olfactory nerves explaining post-­traumatic loss of the sense of smell.

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rotation, extension, extension-­rotation, vertical (axial) compression and lateral flexion, all of them associated with special fracture types and sometimes with (sub)luxations and dislocations/displacements. The so-­called hangman fracture is a traumatic spondylolisthesis of C2  with bilateral fractures through the pedicles due to hyperextension. In atlanto-­occipital dislocation, the ligamentous relationships between the occipital bone and the arch of C1 are torn by severe flexion or extension. Atlas fractures are caused by the impaction of the occipital condyles on the ring of C1. Amongst the odontoid process fractures of the axis, three types are distinguished based on the level of bone severance (I: avulsion of the tip; II: fracture of the base; III: fracture line extending into the body of axis). Depending on the degree of mechanical instability and/or dislocation, vertebral fractures may lead to spinal cord injuries with corresponding neurological symptoms. Figure 20.2.37  Pons with cerebellum lying behind it. Secondary haemorrhages are discernible on the cut surfaces of the pons. Severe craniocerebral trauma in a 47-­year-­old man (brain contusions with concomitant subarachnoid and space-­occupying subdural haematomas). Survival time: 27 hours.

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Non-­penetrating blunt force may damage the thoracic wall and/ or the chest organs. Rib fractures are caused either by direct or by indirect violence. In the first case, a localised force is applied and the underlying ribs are broken in the contact area (for instance, along the seat belt in head-­on collisions); the other (indirect) type of rib fracture occurs away from the impact, mainly due to compression of the chest. Rib fractures are frequently associated with complications that may be dangerous or even life-­threatening (Figure 20.2.38): 1. If a victim sustains numerous fractures, the ribcage loses its rigidity so that the injured section of the chest wall will not participate in the expansion of the thorax during inspiration with the result of paradoxical respiration (flail chest) and concomitant hypoxia. 2. Sharp pointed ends of the rib fragments may penetrate the pleura and lacerate the lung (Figure 20.2.39a) and/or the intercostal blood vessels with consecutive bleeding into the chest cavity (haemothorax). 3. A leak in the visceral pleura or a penetrating injury of the thoracic wall permits air to enter the pleural cavity (pneumothorax) so that the lung collapses, if it is not fixed to the chest wall by pre-­existing pleural adhesions. A valve-­like leakage in the pleura leads to a so-­called tension pneumothorax caused by an increasing pressure of trapped air in the pleural cavity and followed by a complete collapse of the affected lung and a shift of the mediastinum to the opposite side. 4. The presence of air bubbles in the subcutis (Figure 20.2.40) or in the mediastinum (subcutaneous/mediastinal emphysema) may derive from injuries of the trachea, the bronchi, the thoracic wall or the lungs by air entering the adjacent soft tissues. When air has accumulated beneath the skin, a characteristic crackling can be felt on touching (subcutaneous crepitation). Apart from the chest wall, gas bubbles may spread in the soft tissue of the neck and face with conspicuous bulging of the eyelids (Figure 29.2.41).

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20.2.4 Neck trauma

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Some aspects of blunt injuries to the neck are dealt with in other sections of this book (e.g., traffic injuries in Chapter  63.1 and strangulation in Chapter  22.1). Only a few general remarks are therefore made in this context. In victims sustaining blunt neck trauma, the soft tissues (e.g., integument, muscles, thyroid gland, vessels), the airways (e.g., larynx, trachea) and the vertebral column with its spinal cord may be affected. Non-­penetrating injuries to the anterior neck most often arise from motor vehicle accidents, assaults, falls (from height) and sports. In such instances, the laryngeal skeleton is compressed between an impacting object and the spine. When neck injuries are present, one always has to consider an intentional infliction by another person. This does not only apply to manual or ligature strangulation but also to blows and kicks. In fatal assaults by kicking and trampling, neck injuries can be seen in about 40% of cases with a high frequency of throat skeleton fractures. In non-­homicidal falls from height, blunt neck injuries (including haematomas as well as fractures of the hyoid bone and/or the thyroid cartilage) have been observed with a frequency up to 33%. Apart from bleeding with consecutive blood aspiration, size reduction of the airways (by endolaryngeal haematoma, oedema, mucosal disruption, displaced fractures) may be another cause of respiratory distress. Spinal injuries range from temporary and merely functional impairment due to ligament and muscle strains (distorsion) to vertebral fracture and dislocation, the latter ones often accompanied by spinal cord lesions. One-­third of the cervical spine fractures occur at the level of C2, and half at the level of C6 or C7. The causative mechanisms resulting in spine injuries include flexion-­

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Figure 20.2.38  Polytrauma in a 71-­year-­old man caused by a fall from a height. (a) Blood accumulation in the left thoracic cavity (haemothorax), rupture of the diaphragm with displacement of the gastric fundus into the left thoracic cavity and rupture of the liver and small intestine with leakage of chyme into the abdominal cavity. (b) Left-­sided serial rib fractures partly perforating the parietal pleura.

Figure 20.2.39  (a) Front view of the right lung with multiple lacerations from penetrating fragments of fractured ribs (arrows). (b) Dorsal aspect of the left lung showing deep lacerations of the lower lobe.

Blunt force injuries to the lung are mainly encountered as contusions or lacerations. A contusion is typically caused by a substantial impact on the chest with consecutive inward bending of the thoracic cage. In young victims, contusions are not necessarily accompanied by fractures of the ribs or the sternum because of the high pliability of the juvenile thoracic cage. From the morphological point of view, a contused lung shows bruising either as a subpleural suffusion or as an intrapulmonary haemorrhage. Lacerations of the lung (Figure 20.2.39b) can result when a severe

compressive or crushing force is applied to the chest so that the pulmonary tissue bursts or tears. Another possible mechanism is inward displacement of a fractured rib which impales the lung. Blunt traumatisation of the heart manifests itself as concussion, contusion or myocardial rupture. In most cases, the force is directly applied to the anterior chest, which compresses or crushes the heart between the sternum and the vertebral column. Bruises of the cardiac wall may be localised in the subepicardial fatty tissue (sometimes in combination with post-­traumatic

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sites of predisposition: in the descending part of its arcus (near the attachment of the ligamentum arteriosum, Figure  20.2.43b) or immediately above the cusps of the aortic valve (compare with Figure 20.2.42). Other locations (for instance, in association with a dislocated vertebral fracture) are rather rare. The laceration of the aorta may occur either as complete or partial transection. In the latter case, the outer layers of the vascular wall are not damaged; the intimal tears are often multiple, semicircular and parallel (so-­called ladder-­rung tears). If the trauma is survived at least for a short time, parietal thrombosis or post-­traumatic aneurysm may follow as secondary complications.

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coronary occlusion) or within the myocardium, which then appears dark red from interstitial haemorrhage. Lacerations of the heart are most often seen in the relatively thin right ventricle or in the atria; they are less common in the left ventricle, papillary muscles, cardiac valves and interatrial and interventricular septum (Figure 20.2.42). The risk of cardiac rupture is especially high during diastole, when the heart chambers are filled with blood and therefore easily burst on being exposed to a sudden compressive force. Such injuries usually have a fatal outcome either from massive blood loss and haemorrhagic shock (if the pericardial sac is torn and the blood pours into the pleural cavity) or from cardiac tamponade (blood accumulation in the pericardial sac resulting in insufficient filling of the cardiac chambers and impaired forward circulation). In very rare cases, the traumatic heart rupture may be delayed, if the blunt chest trauma first led only to a tear of the inner myocardial layers, which later perforated into the pericardial sac. Traumatic aortic ruptures typically occur in vehicular accidents and in falls from a height. The most important mechanism is sudden deceleration, possibly in combination with compression and/or shearing. Traction forces tear the aorta transversely at two

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Figure 20.2.40  Tissue emphysema with air bubbles in the chest wall (shown after removal of the skin and subcutis).

Figure 20.2.42  Anterior aspect of the heart of a 64-­year-­old car driver who died after a head-­on collision with a truck, showing transection of the ascending aorta and traumatic lacerations of the right and left ventricular walls.

Figure 20.2.41  Orbital region of a 71-­year-­old man who sustained blunt chest trauma with subtotal severance of the trachea and subsequent gas emphysema. The eyelids are bulged due to air bubbles in the subcutis (a) and in the subconjunctival layer (b).

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to the chest organs (lungs, heart and pericardium) as well as pneumothorax and haematothorax. The broken bones are mostly accompanied by only minor fracture haematomas. The probability of sustaining bone injuries from chest compression is particularly high in elderly patients. In fatalities, histological investigation of lung tissue quite often reveals fat embolism.

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Vertebral fractures can occur anywhere along the spine. A common location is the thoracolumbar region, which is affected in more than 60% of cases. The most frequent type of injury is the compression fracture in which the vertebral body collapses due to a sudden downward force often resulting in a wedge-­shaped deformation of the bone. Pre-­existent bone diseases such as osteoporosis and some (metastatic) spine tumours are associated with increased fragility, so that even minor trauma can be sufficient to break the weakened bone. In dislocations, the vertebrae are no longer in alignment, mostly as a consequence of overstretched/torn ligaments and often accompanied by concomitant fractures or severed disks. A dislocation of the spine is prone to instability with a high risk of compressing or even squeezing through the spinal cord. In the age group of infants and pre-­verbal children, rib fractures deserve special attention. If serious accidental traumatisation of the chest (e.g., in a traffic accident) and genetic/metabolic bone disorders can be excluded, rib fractures should always raise concern for maltreatment. In the context of physical abuse, rib fractures are either unilateral or bilateral and typically multiple. The paravertebral sections close to the ribs’ necks are sites of predilection in chest compression. Fractures of different age and the presence of other abuse-­related injuries are additional indicators of non-­accidental trauma. Callus formation and remodelling suggest an earlier infliction. On radiographs, the callus formation may appear as a string of beads. For further information, see Chapter 38.2. Cardiopulmonary resuscitation (CPR) involving chest compression inevitably bears a considerable risk of thoracic trauma, which has to be accepted to maintain a sufficient cardiac output. The spectrum of injuries potentially related to CPR comprises fractures of the thoracic cage (sternum and anterior ribs), damage

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Figure 20.2.43  Thoracic trauma due to a suicidal fall from a height in a 68-­year-­old man. (a) Spine fracture, right-­sided severance of the intercostal muscles, and serial rib fractures perforating the parietal pleura on the left side. (b) Traumatic rupture of the aorta (at the transition from the arcus to the descending part) with subsequent haemorrhage into the adjacent mediastinum.

20.2.6 Abdominal injuries and pelvic fractures Blunt force injuries of the abdomen are frequently seen in traffic and work accidents, in child and spouse abuse, in other criminal assaults (with kicking, stamping and punching) and also in suicidal falls from heights. The abdominal organs most vulnerable to blunt trauma are the solid liver and spleen on the one side and the mesentery on the other (see Figure 20.2.38a). Concomitant external signs of blunt traumatisation such as contusions or abrasions are by no means obligatory. Substantial injuries to the liver, the spleen and the mesentery always have to be regarded as life-­threatening and potentially fatal, especially in cases without rapid surgical treatment. The main reason is intraperitoneal bleeding from lacerations. Ruptures of the liver and spleen can be classified either as a transcapsular or a subcapsular laceration. In the first case, both the capsule and the parenchyma are injured so that the blood instantaneously pours into the peritoneal cavity (Figures  20.2.44 and 20.2.45). The second type of laceration is characterised by the initial formation of a subcapsular haematoma (Figures  20.2.46 and  20.2.47), which expands continuously and may cause a delayed rupture when the covering capsule tears due to overstretching (mostly several hours or even days after the trauma).

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Figure 20.2.44  Transcapsular lacerations of both hepatic lobes in a 48-­year-­old car driver who had a head-­on collision with a truck.

The stomach and the intestine are less susceptible to blunt traumatisation than the parenchymatous abdominal organs. The hollow viscera are more likely to rupture, if they are filled with food or fluid (Figure 20.2.48b). Another reason why the intestine or stomach may be prone to damage is squeezing of the organs between the indented abdominal wall and the lumbar vertebrae. Fatal outcomes from contusions or lacerations of the gastrointestinal tract are usually due to diffuse peritonitis. Renal injuries (Figure 20.2.49) are a relatively rare source of severe bleeding, since the kidneys are deeply located behind the peritoneum. Nevertheless, they can be ruptured by a heavy impact to the loin (e.g., in traffic accidents or assaults). Though the empty urinary bladder is placed within the pelvis, when filled it moves upwards and is therefore exposed to blunt traumatisation of the lower abdomen. Consequently, rupture of the empty bladder is expected to be extraperitoneal and

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Figure 20.2.45  Transcapsular lacerations of the spleen in a 40-­year-­old motorcyclist.

Figure 20.2.46  (a) Large haematoma beneath the still intact capsule of the liver (incidental autopsy finding) with no intra-­abdominal haemorrhage. (b) The subcapsular ruptures and blood accumulations were made visible by partial removal of the detached capsule. The causative trauma remained unclear (probably an unobserved fall in the bathroom where the victim was found dead). There were no resuscitation attempts.

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Figure 20.2.47  Delayed rupture of the spleen in a 45-­year-­old woman who had been physically abused 3 days before death (domestic violence). (a) The secondary rupture of the capsule was followed by fatal bleeding into the peritoneal cavity (2 L of blood). (b) The site and extent of the subcapsular/intraparenchymal haematoma can be seen on the cut surface. Autopsy revealed the additional presence of left-­sided rib fractures (ribs 7–10). In spite of persistent abdominal pain the victim refused to accept medical treatment.

accompanied by pelvic fractures, whereas a bladder distended with urine may rupture into the peritoneal cavity. Blunt traumatisation of a pregnant uterus is a possible cause of fetal death, mostly due to separation or rupture of the placenta. In young and middle-­aged people, great forces are needed to produce pelvic fractures. Therefore, these injuries are frequently seen in victims of motor vehicle crashes (including motorcyclists), after falls from a height or in persons who suffered severe compression of the lower trunk. However, in elderly people with an increased fragility of bones (usually due to osteoporosis), the most common cause of pelvic fracture is a simple fall to the ground. The causative mechanisms in pelvic fractures comprise lateral compression, anteroposterior compression, vertical shear

Figure 20.2.48  Homicidal death of an intoxicated 49-­year-­old man (blood alcohol concentration 2.87‰) who suffered blunt traumatisation of the abdomen by multiple blows and kicks. Lacerations can be seen of (a) the mesenterium and (b) the intestinal wall with consecutive haemoperitoneum (2 L of blood).

and a combination of these forces. Fractures of the pelvic ring may be stable if the sacroiliac complex is intact. A complete disruption of the sacroiliac joint is accompanied by both rotational and vertical instability. A heavy impact to the pubic region (e.g., in head-­on-­collisions of motorcyclists) disrupts the pelvic girdle in the symphyseal area so that the right and left halves of the pelvis are separated from each other (more at the front than at the rear, ‘open book fracture’). Pelvic fractures are often associated with severe bleeding into the surrounding soft tissues and therefore constitute a major risk of traumatic shock.

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In cases of fatal child abuse, lacerations of the abdominal organs with subsequent exsanguination are the second most common cause of death (Figure  20.2.50). As in adults, haematoma discolouration visible on external examination of the body is by no means obligatory. Therefore, at autopsy, the abdominal wall should be dissected layer by layer in order to ascertain any site of impact.

20.2.7 Injuries to the extremities

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Figure 20.2.49  Multiple lacerations of the kidney in a 16-­year-­old suicide jumping from a height.

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Apart from injuries to the skin and the subcutaneous layer, other anatomical structures such as the muscles, bones and joints may be involved in blunt force trauma. Extensive crushing of the soft tissues, the formation of blood-­filled cavities, comminuted fractures and severance of large vessels are frequent findings in victims of automobile–pedestrian accidents. The biomechanics and the morphology of impact traumas to the leg are dealt within the context of traffic injuries (see Chapter 63.1). A bending load is by far the most common mechanism of fracture for lower-­extremity bones. Bending may produce an oblique, transverse or wedge fracture in the shaft of long bones (see Chapter 16). Other possible mechanisms of fractures

Figure 20.2.50  Fatal child abuse of a three-­and-­a-­half-­year-­old boy with blunt traumatisation of the abdomen (fist blows). Anterior aspect of the trunk (a) before autopsy and (b) after removal of the skin. For better visualisation of the subcutaneous and intramuscular haematomas, the affected tissue has been incised. Rupture of (c) the liver and (d) the mesentery close to the ileocecal junction.

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Figure 20.2.52  Cut surface of the right lung with purulent bronchopneumonia in an 82-­year-­old woman who sustained severe craniocerebral trauma in an accident 6 days before death.

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are axial loading (e.g., in dashboard-­style impacts) and torsion (resulting in spiral fractures). The injury patterns of skin, soft tissues, joints and bones often contribute to reconstruction in traffic accidents. Bone bruises at the site of impact (e.g., in a lateral femur condyle) can be visualised either by autopsy or by magnetic resonance imaging (MRI). In children, intentional injuries caused by abuse have to be differentiated from sequelae of accidental traumatisation (see Chapter 38.2). In early childhood, physical abuse accounts for a high percentage of bone fractures. As already mentioned in the earlier text, this applies especially to the ribs, and also to the long bones of the extremities (radius and ulna, tibia and fibula and humerus and femur). With regard to age-­related features of paediatric bone injuries such as greenstick fractures, subperiosteal haematoma and calcifications, metaphyseal corner fractures and epiphyseal dislocations, reference is made to the pertinent literature. A special type of bone injury is seen both in children and adults when victims of a blunt force attack try to protect themselves by raising their hands or arms. A classical fracture site is the ulnar shaft, which is exposed in victims parrying a blow or kick (‘defence injury’). Slip, trip and fall accidents constitute a major public health concern. Especially amongst the elderly, injuries from simple falls to the ground are often followed by life-­threatening consequences (e.g., thromboembolism or pneumonia; Figures  20.2.51 and 20.2.52). Most femoral neck fractures and intertrochanteric/ subtrochanteric fractures are due to a fall or other minor trauma when the bone’s fragility is increased in patients with osteoporosis. In younger people with normal bones, hip fractures are caused by high-­energy trauma (e.g., motor vehicle accident, fall from a height, etc.). On external examination, the affected leg is typically shortened and rotated. Other fracture sites often seen in patients with osteoporosis are the distal radius (after a fall on the outstretched hand) and the proximal humerus.

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Figure 20.2.51  Fatal pulmonary thromboembolism following total hip replacement surgery in a 52-­year-­old woman; the latency period between trauma and death was 12 days.

20.2.8  Sequelae of blunt trauma Internal bleeding (from closed injuries) and external bleeding (from traumatic amputation, severe avulsion wounds and compound fractures) are important factors contributing to haemorrhagic shock and consecutive organ dysfunction (mainly affecting the lungs and kidneys, see Chapter 18.2). Local complications of blunt trauma in general and bone fractures in particular include the compartment syndrome (increased pressure within an injured body region enclosed by fascia resulting in deficient blood supply) and – mostly bacterial – infection of the damaged tissue. The spectrum of early systemic complications comprises conditions such as hypovolaemic shock, adult respiratory distress syndrome (ARDS), disseminated intravascular coagulation, fat embolism syndrome, sepsis and crush syndrome (traumatic rhabdomyolysis accompanied by circulatory shock and kidney failure ensuing from a crushing injury to muscle tissue). It has already been pointed out in the context of blunt head trauma that an interfering mechanism such as aspiration of blood and gastric contents may constitute the immediate cause of death. Venous air embolism is another potentially lethal complication of cranial trauma: If an open skull fracture is associated with a gaping lesion of a dural sinus, air may enter the injured vessel provided that the intravascular pressure is temporarily lower than the atmospheric pressure (e.g., due to deep inspiration). If this happens, gas is sucked in and transported to the right side of the heart, where it causes pump failure because, contrary to blood, air is compressible. In blunt head traumas with a rapid fatal outcome,

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at least in a large part. The lipids extracted from pulmonary tissue in cases of FE demonstrate an increase in the triglyceride fraction with a shift towards oleic acid. Thus, the composition of lipids in the lungs can be regarded as a further indication that the embolised fat predominantly derives from the adipose tissues. Considering pathophysiology, clinical symptoms and autopsy findings, a distinction has to be made between pulmonary and systemic fat embolism. Liquid fat released from injured tissue enters the venous vessels and is carried to the right heart and the pulmonary arteries, before it gets stuck in the capillaries of the alveolar septa. Large amounts of fat impacted in the lung vessels may increase the right ventricular load and impair gas exchange with subsequent respiratory insufficiency. In patients presenting with FES, pulmonary symptoms usually occur first. Nevertheless, pulmonary fat embolism mostly is a concomitant feature of mechanical trauma and not the actual cause of death. In fatalities with high-­grade FE, drops of fat can be seen in the venous heart blood even with the naked eye. When an uninjured body is exposed to postmortem burning, fat may also be visible in the blood of the right heart, but the pulmonary capillaries do not contain impacted fat globules typical of true FE. Systemic fat embolism is to be expected when fat globuli penetrate the lung capillaries and are distributed in the systemic circulation. Alternatively, embolised fat can pass through arteriovenous anastomoses or a cardiac shunt (e.g., a patent foramen ovale). The most vulnerable target organs of systemic FE are the brain, the kidneys and the myocardium. On external examination, petechial bleedings in the skin and conjunctivae may be indicative of systemic FE. At autopsy, punctate haemorrhages of the brain (‘purpura cerebri’) constitute the macro-­ morphological correlate of cerebral involvement (Figure 20.2.54). The underlying vascular obstruction by embolised fat globules entails the neurologic symptoms observed in FES. Histological examination reveals intravascular fat not only in the brain (predominately in the white matter), but also in the kidney glomeruli and in the capillaries between the myocardial fibres. In trauma patients, the manifestation of FES is mostly delayed in time (commonly between 24 and 72 hours post injury, but on rare occasions earlier as well).

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venous air embolism should therefore be taken into consideration as a potential mechanism causing peracute death. Another sequel to blunt trauma is pulmonary and systemic fat embolisms (caused by globules of fat, usually subsequent to bone fractures or damage of fatty tissues). Pulmonary fat embolism is a common phenomenon found in the great majority of fatalities from blunt trauma. By contrast, the clinical entity referred to as fat embolism syndrome (FES) is much less frequent in patients who sustained mechanical injury. Typical manifestations of FES are characterised by a triad of symptoms: respiratory distress, neurologic disturbances and petechial skin rash. The latter one is mostly distributed over the face, neck, anterior chest and the axillary regions. The clinical symptoms are usually ascribed to vascular obstruction and inflammatory response mechanisms (Figure 20.2.50). From the pathoanatomical point of view, FE means the presence of fat droplets impacted in peripheral vessels. The intravascular fat globules are demonstrated histologically in frozen sections using suitable fat staining methods (Figure  20.2.53). According to the mechanical formation theory, liquid fat from disrupted adipocytes gets access to venous vessels, especially in cases of long-­bone and multiple fractures as well as extensive damage to adipose tissue (e.g., in crush injuries). Nevertheless, it is undoubted that moderate FE can also occur without any history of blunt traumatisation. Vital burns, alcoholic fatty liver disease, osteomyelitis and acute pancreatitis can be cited as examples. Surgery involving fatty tissues and bones is often accompanied by FE. The same is true for chest compression in CPR. Analogous to other kinds of embolisation, FE is regarded as a sign of vitality, as it presupposes an adequate circulation (see Chapter 19.1). The frequent concurrence of pulmonary fat and bone marrow embolism suggests that both phenomena arise from injured bones,

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Figure 20.2.53  Pulmonary tissue of a 68-­year-­old man who died from blunt polytrauma due to a fall from a height (bilateral serial rib fractures and pelvic fracture; survival time: 7 hours). Oil Red O staining of a frozen section visualising numerous embolised lipid droplets in the small lung vessels recognisable by their deep red colour (magnification approximately 100×).

Figure 20.2.54  Cut surface of the right temporal brain lobe showing pinhead-­sized haemorrhages in the white matter (purpura cerebri) caused by systemic fat embolism. The presence of abundant fat droplets in the pulmonary and cerebral vessels was histologically confirmed; compare with Figure 20.2.53).

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Flaherty, E.G., Perez-­Rossello, J.M., Levine, M.A. and Hennrikus, W.L. (2014). Evaluating children with fractures for child physical abuse. Pediatrics 133: e477–e489. Klein, A., Rommeiß, S., Fischbacher, C., Jagemann, K.-­U. and Danzer, K. (1995) Estimating the age of hematomas in living subjects based on spectrometric measurements. In: Oehmichen, M. and Kirchner, H. (eds.), The Wound Healing Process – Forensic Pathological Aspects, pp. 283–291. Lübeck: Schmidt-­Römhild. Pollak, S. and Saukko, P. (2013). Blunt injury. In: J.A. Siegel, P.J. Saukko and M.M. Houck (eds.), Encyclopedia of Forensic Sciences, 2nd edn, Vol. 3, pp. 1–10. London: Academic Press. Pollak, S. and Vycudilik, W. (1981). Pulmonary lipids in fatal burns. Wiener Klinische Wochenschrift 93: 111–117. Pollak, S., Vycudilik, W., Reiter, C. et al. (1987). Large droplet lipids in blood in the right ventricle. Zeitschrift Rechtsmedizin 99: 109–119. Pircher, R., Pollak, S., Thierauf, A. et al. (2013). Modification of hematoma findings in the breast region. Forensic Science International 224: 33–36. Rothberg, D.L. and Makarewich, C.A. (2019). Fat embolism and fat embolism syndrome. Journal of American Academy of Orthopedic Surgeons 27: e346–e355. Saukko, P. and Knight, B. (2016). Knight’s Forensic Pathology, 4th edn, pp. 133–148, 167–228, 339–352, 475–495. Boca-­Raton, FL: CRC Press.

Body abuse and killing by kicking are perceived more and more frequently and occur differently from region to region, whereby social factors play a significant role, as the evaluation of the logs of 446 cases in different regions of Germany has shown. Violent offenses by kicking are mostly milieu-­typical acts of previously known, mostly male and lower-­social-­class people, which appear as a violent escalation after previous minor disputes (Heinke 2010a, b). Most of the time, disputes caused by drunk people that have gone into a scuffle ultimately lead to kicks against someone who fell in the dispute. The victim’s blood alcohol concentrations of 3.00‰ and more are not uncommon (in the cases examined in Hamburg, Greifswald and Halle, the average blood alcohol concentration was 2.16‰, the highest 4.5‰). The disputes usually take place without cause or for little reason. The reasons mentioned later are “desire for a quarrel” and the view that “homeless people and vagrants, foreigners or people with a ‘migration background,’ who certainly also appear as perpetrators, do not fit into society” and “homeless people take advantage of taxpayers.” The choice of the victim is often a “product of chance”; it is not uncommon for people with moral courage who want to help or arbitrate to become victims. Cases in which a victim is specifically sought out and—­sometimes over several days—­mistreated with blows and kicks are rarer. Mostly, drunk offenders claim that they no longer knew what they were doing due to the alcohol-­related cognitive impairment—­ that is, they were considerably restricted in their ability to control—­and that they kicked the victim indiscriminately. This admission is refuted if the injuries—­as in many cases—­are almost exclusively on the head, since the head–body relationship would suggest a different distribution pattern for the injuries “if you were kicked at random.” The trivialization of the expected severity of the injury on the grounds that people only kicked with sneakers and not with combat boots is refuted due to the development of strength that is caused by kicking and not by the shoe. From a legal point of view, the shod foot is now uniformly viewed as a weapon, and bodily harm caused by being kicked as a “life-­threatening” act. In experiment values, measurements of 350–850  N were reached by fist punching, and 500– 1200 N by kicks by men and women. The author summarized these results in one sentence: “The smallest measured values for kicks and the largest for punching overlap in both sexes. . .” (Figure 20.3.1). The results of the evaluation of the arrested “kick deaths” perpetrators showed that usually sneakers were worn. In one case, even kicks with “bare” feet were fatal. In a study conducted by the University of Kelaniya, Sri Lanka, kicks to the trunk were found in 27% of cases of torture (in many cases in combination with other torture methods). Decisive for the degree of injury are not the type and severity of the footwear worn and not the constitution of the perpetrator, but rather the force of stepping, the kicking technique and the point or points of impact on the victim’s body. Usually, such crime sequences lead to multiple traumas, but a single kick can be fatal.

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In cases of prolonged survival, intercurrent infection, pneumonia, multiorgan failure and pulmonary thromboembolism (originating from post-­traumatic venous thrombosis) are dangerous and often fatal complications of an originally non-­lethal injury (e.g., fracture of the femur neck).

20.3  Homicides by Kicking

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A 37-­year-­old man was found dead next to his bicycle in front of a cowshed in the morning after drinking copious amounts of alcohol in a restaurant the previous evening. At first, the situation indicated a fatal bicycle fall. However, the on-­site investigation by a forensic doctor set the course in a different direction, so that a forensic autopsy was ordered. The main finding of the autopsy was massive abdominal trauma with hemascos of 1400 mL due to rupture of the mesenteric vessels. The juxtaposition of broken ribs, broken nose bones, lacerations on the back of the head and multiple hematomas could not easily be explained by a bicycle fall and pointed to multiple massive blunt violence (probably kicks). The blood alcohol concentration was 2.62‰, and the urine alcohol concentration was 4.08‰. After one and a half years of investigative work, the perpetrators, three young people, were found who admitted to having punched and kicked the man out of boredom, the desire for an argument and because he never resisted. Definition Kicking to death is defined as the bringing about of fatal internal injuries by kicking the victim’s body.

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by showing appropriate dirt. An “endogenous” crumple zone effect occurs in upholstery through subcutaneous fatty tissue, so that the least indicative findings are to be expected on parts of the body that are covered by clothing and are rich in fat (Figure 20.3.2). The internal injuries are caused by different mechanisms. On the one hand, it is the direct action of force that can lead for example to subcutaneous fatty tissue bleeding, bone fractures and torn organs. On the other hand, it is the indirect force in form of an acceleration trauma that plays a major role, especially in head and brain injuries. Kicks in the head often lead to broken bones, with midface fractures (25% of cases) particularly frequently. In addition, fractures of the base of the skull (12%) and the roof of the skull (10%) give clear indications of the applied force. The acceleration experienced by the head of a person lying on the ground when kicked in the head averages 56 g, and exceeds the brain acceleration that a pedestrian experiences when the head hits the hood of a car at a collision speed of 36 km/h. The maximum values of 103 g roughly correspond to the head acceleration of a driver wearing a seat belt in a frontal crash at 50 km/h. The torques acting on the neck are around 16.3–40 Nm (maximum 125 Nm). The head acceleration and the simultaneous rotation of the head cause coup and contrecoup injuries to the brain as well as rotational trauma. Intrathoracic organ injuries fill the spectrum from, for example, minor lung contusions to heart ruptures.

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Figure 20.3.1  Force release in kicks and fist punches.

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Legally, both combat boots and sneakers are considered dangerous tools. The evaluation depends on the type of offense committed. A step on the toe of an injured person with a combat boot does not imply the use of a dangerous tool or a “life-­threatening” injury, whereas a kick with a sneaker on the head of a victim constitutes a threatening” act. In the BGH judgment of 24 September “life-­ 2009  – 4 StR 347/09 (Regional Court, Dortmund), the following statement was made: “The shoe on an offender’s foot is to be regarded as a dangerous tool within the meaning of § 244 I No.2 StGB if either a firm, heavy shoe is involved, or if the victim is kicked in particularly sensitive parts of the body with a ‘normal street shoe’ with force or at least violently.” (Heinke 2010b, issue 3, p. 151)

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20.3.2 Localization of injury and external finding patterns The external injury patterns are very different depending on the affected body region and direction of the force. In the case of orthograde hits, blunt violence on unpadded areas of the body (e.g., forehead), the weapon is often depicted one-­to-­one as a “negative impression” due to skin bleeding. Here, the mechanism known from striking with a stick or a stick-­like object, which leads to a double-­contoured injury, comes into play. If a region of the body is covered by thin-­layer clothing, the pattern of the worn textiles can be recognized within the imprint of the weapon. With thicker layers or wadding of the clothing, a so-­called crumple zone effect occurs, which often only shows indistinct to no traces of the external violence on the skin. Even the clothing alone can, for example, reflect the shoe-­sole profile

Figure 20.3.2  Kick marks on the neck and chest, and (inset) the murder tool.

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When examining living people, an ultrasound examination of the affected body regions can be helpful in order to detect externally undetectable hematomas. However, the findings are to be interpreted with caution, since the age of the hematomas cannot be specified for narrow periods of time using this method. Injuries to internal organs should also be considered if the picture is unremarkable and should be clarified by sonography. Possibly, a computed tomographic X-­ray examination and/or magnetic resonance tomography is insightful. When dissecting the deceased, skin and subcutaneous fatty tissue must be dissected in layers, so that only discreetly developed fatty tissue bleeding can be seen, which may hint at the instrument of crime (Figure 20.3.3). Often, it is only the meticulously documented individual injuries and their overall picture that exclude an accident as the cause of death, as seen in the case study mentioned in the earlier text.

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The breadth of intra-­abdominal injuries ranges from tears in the relatively unprotected liver capsule to central disintegration of the liver to contusions and ruptures of the relatively protected kidneys. The severity of the internal injury does not always correlate with the external injury pattern. Pre-­existing illnesses can significantly promote death through consequential damage, such as bleeding to death from cracked and crushed wounds in the case of pre-­existing blood clotting disorders as a result of alcohol-­toxic liver cirrhosis. Also with higher blood alcohol concentrations (especially values above 3.50 ‰), it should be considered whether alcohol intoxication has contributed significantly to the occurrence of death or it is a competitive cause of death. However, alcohol intoxication as the cause of death must always be negated if the injuries alone can informally explain the occurrence of death—­ regardless of whether the victim was intoxicated.

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Figure 20.3.3  Well-­recognizable arched hematoma in skin and subcutaneous fatty tissue after kick with a low shoe/loafer; dissection in layers.

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The easiest way to document orthograde hits on unpadded areas of the body is to “draw” the finding pattern on foil. This gives you a one-­to-­one image of the findings. Furthermore, a photographic documentation with and without a scale should be made by a suitably good recording technique and a person who is familiar with the method. The localization of the injuries on the body must always be traceable. In the case of padded body regions, “tracing” is usually unnecessary due to missing or only sketchy findings. But photo documentation should always be provided.

Important When examining living people, the possibility of internal organ injuries should be considered, even if the outer findings are ­ inconspicuous.

References and further reading Bodziak, W.J. (1990). Footwear Impression Evidence, pp. 126–130. New York Amsterdam London: Elsevier. Böhm, E. (1987). Zur Morphologie und Biomechanik von Trittverletzungen. Beiträge zur gerichtlichen Medizin 45: 319–329. Böhm, E. and Schmidt, B.U. (1987). Kriminelle und kinetische Energie bei Tötungshandlungen durch stumpfe Gewalt. Beiträge zur gerichtlichen Medizin 45: 331–338. Brüschweiler, W., Braun, M., Fuchser, H.J. and Dirnhofer, R. (1997). Photogrammetrische Auswertung von Haut-­und Weichteilwunden sowie Knochenverletzungen zur Bestimmung des Tatwerkzeuges  – grundlegende Aspekte. Rechtsmedizin 7: 76–83. Graß, H., Madea, B., Schmidt, P. and Glenewinkel, F. (1996). Zur Phänomenologie des Tretens und Tottretens. Archiv für Kriminologie 98: 73–78.

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Cellular elements of the CNS

The neuron

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A human brain contains about 1 × 1012 neurons with each of it receiving up to 30 000 presynaptic terminals (Tanner 1978). The task of nerve cells is to report stimuli from our environment or from inside the body to our brain and to process them in the neuronal network. For every microscopic examination, it is important to identify each cell type properly. The variety in shapes and sizes of neurons in different areas of the brain makes it difficult to define the characteristic morphological features of a nerve cell. Usually, the cell body or perikaryon of a neuron is pyramidal in shape such as the typical Betz cells of the cortex. Others such as the Purkinje cells of the cerebellum appear like pears, and finally there are neurons which resemble lymphocytes such as the granule cells of the cerebellar cortex. The perikaryon contains a lot of organelles for oxidative metabolism and protein synthesis. The special cytoskeleton consists of three components: neurofilaments, microtubules, and purified microtubuli. The latter are composed of alpha-­and beta-­tubulin and microtubule-­associated proteins. The microtubule-­associated protein (MAP2) has gained importance because it is able to aid in diagnosis of hypoxic damage of neurons, which may be useful in forensic routine investigations (Kühn et  al. 2005). Neurons are easily visualized microscopically by hematoxylin and eosin (H&E), Nissl, or silver staining as well as by a number of monoclonal antibodies. Besides the abovementioned feature, nerve cells are characterized by cell processes called axons and dendrites connecting one neuron with the other by synapses, and forming an intracranial network. Axons and dendrites are detectable by means of specific silver staining technique and immunohistochemistry. Axonal-­injury investigation is best achieved by beta-­amyloid precursor protein (β-­APP) staining or the silver staining technique, and facilitates the detection of injury by presenting disrupted axons, axonal balls, or axonal bulbs. The myelin sheath of myelinated axons can be demonstrated by Luxol fast blue staining or a specific antibody to myelin basic protein (MPB).

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Heinke, D.H. (2010a). Tottreten  – eine kriminalwissenschaftliche Untersuchung. Lengerich: Pabst Science Publishers. Heinke, D.H. (2010b). Das Deliktphänomen “Tottreten”  – Kriminalwissenschaftliche Erkenntnisse und Folgerungen für die rechtliche Bewertung –. beck-­online – NStZ: 119–125. Henn, V., Lignitz, E., Philipp, K.P. and Püschel, K. (2000). Zur Morphologie und Phänomenologie des Tottretens, Teil 1. Archiv für Kriminologie 205: 15–24. Henn, V., Lignitz, E., Philipp, K.P. and Püschel, K. (2000). Zur Morphologie und Phänomenologie des Tottretens, Teil 2. Archiv für Kriminologie 205: 65–74. Perera, P. (2007). Physical methods of torture and their sequelae: A Sri Lankan perspective. Journal of Forensic and Legal Medicine 14: 146–150. Reh, H. and Weiler, G. (1975). Zur Traumatologie des Tottretens. Beiträge zur gerichtlichen Medizin 33: 148–153. Schrader, S. (1933). Wunde und Werkzeug. Tödliche Schädelverletzung durch Fußtritte. Archiv für Kriminologie 92: 229–231. Steffen, W. (1989). Turnschuh als “gefährliches Werkzeug”. NJW 14: 920–921. Strauch, H., Wirth, I., Taymoorian, U. and Geserick, G. (2001). Kicking to death  – forensic and criminological aspects. Forensic Science International 123:165–171. Tröndle, H. and Fischer, T. (2003). Beck’sche Kurzkommentare, Strafgesetzbuch und Nebentexte. 50. neubearb. Aufl. Beck, München 1395–1397. Verhoff, M.A., Gehl, A., Kettner, M. et al. (2009). Digitale Forensische Fotodokumentation. Rechtsmedizin 19: 369–381.

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Forensic neuropathology is a specialized discipline of pathology and is required in cases where gross autopsy findings of the medical examiner’s casework suggest not only a death due to a trauma or disease of the head, skull, brain, spine, or spinal cord, but secondary alterations of the central nervous system (CNS) caused by physiologic responses or metabolic cascades to the primary event. The forensic neuropathologist is expected to provide answers regarding (i) the manner of death, (ii) the cause and time of death, (iii) information if neuropathological changes may have influenced the fatal mechanism, (iv) the type, the biomechanics, and time of the traumatic event, and, finally, (v) the ability of the victim to act at a given time point. In summary, it is the forensic neuropathologist’s task to give a detailed and plausible reconstruction of the events leading to death and to provide information about the time course of the neuropathological findings. More details regarding the morphology, the scientific background, and the literature are found in the textbook by Oehmichen et al. (2009a, see also Leestma, Oehmichen and König 2012).

The astrocytes Astrocytes are the most common cell type, outnumbering neurons by an order of magnitude. The astrocytes are essential to keep the highly differentiated neurons in their proper place and to maintain their environment. Astrocytes reveal long branched processes giving the cell a star-­like appearance. Nearly all astrocytes contain 10 μm intermediate filaments, expressing especially glial fibrillary acidic protein (GFAP). The cells are found in the gray as well in the white matter. Astrocytes are interconnected between each other via gap junctions and possess messenger systems that maintain essential communication between themselves and the neurons. Detection of different types of astrocytes and reactive astrocytes is best achieved by applying the GFAP antibody. Three subgroups of astrocytes can be distinguished, which can be

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mal microglia seem to be quite stable in the mature normal brain and may not be replaced by new bone-­marrow-­derived macrophages. Microglial cells as monocyte-­derived mononuclear phagocytes are the most important cells to diagnose pathological conditions of the brain and have been suggested to be a “sensor” of the pathological status of the human CNS (Graeber et al. 1997). Under pathological conditions, “activated microglia” appear, which possess the ability to proliferate locally, to emigrate to the site of injury, and to change morphologically and functionally. In normal CNS, microglia cells are detected by H&E staining as small bean-­formed nuclei. In human brain tissue with pathological alterations, activated microglia is best detectable in paraffin material using silver techniques or the monoclonal antibody LN-­3 as small cells with thin star-­like processes.

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Different types of brain damage such as edema, necrosis, as well as brain ischemia as a type of metabolic disturbance end all up in a final tissue reaction via a final common pathway. Since these cellular and tissue reactions are largely independent of a “specific” type of insult, they are, therefore, nonspecific. The basic principles of the individual types of reactions and their fundamental pathophysiological principles and morphology are to be outlined.

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differentiated by their morphology. Fibrous astrocytes are hosted in the white matter. They are stellate in structure with long, thin, poorly ramified processes that are smoothly surfaced. Protoplasmic astrocytes reside in the gray matter and exhibit ramified processes of a variable caliber. Radial astrocytes are observed in the white matter, disposed in a plane perpendicular to the axis of the ventricle. Their processes lack ramification and at least one process touches the pia mater, the others coursing through the gray matter. Reactive astrocytes are characterized by swelling of the cell body and an upregulation of GFAP and vimentin. If cytoplasm is abundant and the cell rounded, such reactive cells are termed fattened or gemistocytic astrocytes. This cell type reveals a homogeneous cytoplasm and a slightly enlarged nucleus with angular projections from which the processes arise. Astrocytes react to different neuropathological conditions such as trauma, infection, edema, infarction, seizure, or others. Laceration and hypoxic changes of the brain parenchyma produce a powerful glial response and induce also proliferation of connective tissue. Maximum numbers of proliferating GFAP-­positive astrocytes are observed on days 2.5 and 3 after the traumatic or ischemic event. At sites of brain tissue destruction, astrocytes form a scar in which they begin to shrink and finally disappear, leaving behind a dense meshwork of glial fibers. Excision of the brain parenchyma will leave a fluid-­filled space, the surrounding wall of which contains astrocytes with only slight reactive changes.

Increased intracranial volume

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Oligodendrocytes are small cells with a round or oval, relatively dense nucleus and a small rim of cytoplasm with a cell diameter of 6–8 μm. Oligodendrocytes can be demonstrated by silver technique, by antibodies to MBP, and by a panel of monoclonal antibodies. Oligodendrocytes possess ramified processes detectable by silver techniques. They also exhibit a structural polymorphism that reflects differences in function. This cell type is involved in the myelination and remyelinating processes and in immunological processes under pathological conditions. In injured CNS, oligodendrocyte precursor cells constitute a reactive glial population that goes through hypertrophy and mitosis under stimulation from an array of cytokines and growth factors. If there is demyelination, the precursor cells divide and differentiate into new oligodendrocytes that replace the ones that were lost. Activation and proliferation of these cells also occur in response to CNS damage, including excitotoxicity, mechanical injury, and viral infection.

The microglia Microglial cells have a mesenchymal origin and are observed as numerous as neurons in the CNS (Oehmichen 1978). The mesenchymal progenitor cells derived from the bone marrow (monocytes and/or macrophages) patrol the CNS continuously. Especially perivascular and meningeal “macrophage” populations are slowly replaced by hematogenous macrophages. By contrast, parenchy-

If brain volume increases, both blood and cerebrospinal fluid (CSF) are displaced by an increase of intracranial pressure (ICP). Consequent compression of the brain against the inelastic dura mater and the skull can lead to a lethal series of complications in clinical neurology. According to Miller and Ironside (1997), a panel of factors has an impact on brain volume. First, a disturbance of the blood– brain barrier (BBB) can lead to an increase in the fluid content, with a consequent increase in brain volume. The white matter of the brain is 68% water, the gray matter 80%. A rise in brain water content entails an increase in brain volume, that is, brain edema. Second, elevated cerebral blood volume, also known as “brain swelling,” is a congestive process. The intracranial blood volume can be raised by a number of factors such as: • Arterial hypertension. • Enhanced cerebral blood flow secondary to elevated cerebral perfusion pressure. • A decline in the cerebrovascular resistance of arterioles, capillaries, and postcapillary vessels due to hypercapnia. • Hypoxemia associated with severe elevation of arterial pressure. • Obstruction of the venous outflow of the brain. Third, an increase of the CSF pressure may result in an increased brain volume, for example, by an acute obstructive high-­pressure hydrocephalus. A fundamental distinction must be made between focal and global cerebral edema. The latter follows acute systemic

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the sodium–potassium membrane adenosine triphosphatase (ATPase) pump is disrupted, followed by a water accumulation within the cells due to an increase in intracellular sodium and loss of potassium. The increased brain volume leads to flattening of the gyri (Figure 20.4.1a) and compression of the ventricles. The ultimate result of the space-­occupying process is development of lateral and then downward herniation, visible at several loci at the falx cerebri (cingulate or subfalcine herniation), the tentorium cerebelli (lateral or uncal herniation), the thalamus/hypothalamus (central or diencephalic herniation, which may result in downward displacement and hemorrhage in the midbrain and pontine tegmentum), and at the foramen magnum (tonsillar herniation). A bilateral expanding supratentorial mass can cause herniation-­ induced notches (Figure 20.4.1b) as well as hemorrhages of the uncal area. This in turn exerts downward pressure on the medial part of the parahippocampal gyrus toward and through the tentorial

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ischemic or hypoxic events, for example, transitory cardiac arrest or chronic hypoxia (e.g., in respiratory diseases). Global cerebral edema may also be associated with metabolic diseases, intoxications, and inflammation. Focal cerebral edema results from focal tissue destruction due to infarction, traumatic hemorrhage, or tumor. This edema zone of infarction resembles penumbra surrounding the moon in full eclipse. Because these tissue changes are partly reversible, they are of considerable therapeutic interest. Two types of edema can be differentiated and have proved useful in distinguishing between various pathogenetic mechanisms and their sequelae (Klatzo 1967). The first type is called vasogenic edema and is related to a disturbance of the BBB. The second type of brain edema is cytotoxic in nature. The passage of ions and molecules of various sizes is controlled by lipid soluble substances in the endothelial wall and by ionic channels and active pumps like the sodium–potassium pump. As a result of energy failure,

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Figure 20.4.1  Gross autopsy findings of brain edema. (a) Flattening of the gyri, (b) uncal herniation, (c) notches of the cerebellar tonsills, and (d) pontine hemorrhage secondary to intracranial pressure.

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incisura. Herniation of the parahippocampal gyrus creates narrowing of the midbrain along its transverse axis and compression of the aqueduct. An early and fatal complication of expanding masses in the posterior cranial fossa is displacement of the cerebellar tonsils through the foramen magnum (Figure 20.4.1c). This may also be caused, however, by lesions occupying the supratentorial space. Morphologically, the tips of the tonsils exhibit hemorrhagic necrosis and grooving of the ventral surface of the medulla where it impinges on the anterior border of the foramen magnum. Both the fourth ventricle and aqueduct become compressed and displaced contralaterally. The expanding mass of the brain leads to brainstem compression with bleeding within the pons (Figure 20.4.1d).

develop which can be separated from the intact white matter at autopsy.

Hydrocephalus

Neuronal necrosis features irreversible changes of the cytoplasm (condensation, hydropic swelling, intense eosinophilia, loss of structure, homogenization) and of the nucleus (pyknosis, karyolysis, karyorrhexis). Necrotic tissue and cells always attract neutrophils, macrophages, and sometimes lymphocytes, and can activate the scavenger function of macrophages for elimination of myelin as well as cell debris. The infarcted area induced by prolonged ischemia displays macroscopically evident pallor on H&E, Nissl, and myelin staining within 3–5 hours as an indication of acidosis. A narrow halo of even greater pallor surrounds the necrotic area as well as a perifocal edema. Neurons become thorny and severely shrunken within 12–36 hours, with darkly staining incrustation of their pericellular structures. A survival time of 12 hours leads to homogenization of the cytoplasm and nuclear and cytoplasmic pallor of neurons. Between 36 and 48 hours, the neurons disappear except for total the nuclei. Within 1–2 hours, the necrotic tissue is characterized by an emigration of neutrophil leukocytes. Within 18 hours, the necrotic area exhibits extensive immigration and activation of microglial cells along the infarct margin. Hypertrophic astrocytes appear along the border zone within the brain parenchyma after 3–6 days. The infarct liquefies at its center and macrophages phagocytose the debris. The final stage of cortical necrosis is termed “laminar necrosis” associated with capillary proliferation, intense gliosis, and fibrosis during the final phase. The final stage of an ischemic involvement of the white matter, basal ganglia, and the thalamic nuclei is also a cystic necrosis. Ischemic damage of the hippocampal area is characterized by a segmental loss of neurons in the hippocampal cortex associated with compensatory microglial and astroglial activation.

Tissue decay and reactivity

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Hydrocephalus is characterized by abnormal accumulation of CSF within the cerebral ventricles and subarachnoid space. The CNS of the average adult contains a CSF volume of approximately 120–140 mL. An increase of fluid, over time, induces atrophy of the brain parenchyma and additional ventricular enlargement. CSF is formed from the choroid plexus at a rate that remains unchanged over a wide range of ICP values. The subarachnoid space and ventricular system are connected via the foramina of Luschka and Magendie in the basal cisterns and are absorbed by the arachnoid villi. External hydrocephalus ex vacuo involves diffuse loss of gray or white matter that gives rise to external atrophy, with dilatation of the subarachnoid space. Diffuse loss of white matter can cause expansion of the ventricular system, the so-­called internal hydrocephalus ex vacuo. Normal-­pressure hydrocephalus can also result from a hydrocephalus e vacuo, which is associated with primary ventricular system enlargement caused by white matter destruction. It is often found in victims of severe brain injury, in alcoholics, and vascular disease patients with multi-­ infarct dementia or other types of progressive degenerative brain disorders, especially age-­dependent dementia. Among the causes of obstructive hydrocephalus are mechanical brain injury (MBI), subarachnoid hematoma, meningitis, arachnoid fibrosis, Arnold–Chiari malformation, and inborn aqueductal stenosis, which may lead to an obstruction of the connecting foramina of Luschka and Magendie. A routine neuropathological examination reveals the following findings: External hydrocephalus exhibits dilatation of the subarachnoid space, with no increase in collagenous fibers or cellular elements, but an increase in CSF. Internal hydrocephalus typically features macroscopically an enlarged ventricular system, and microscopically an interstitial edema, disruption of the ependymal cells lining the ventricle, and axonal destruction in the periventricular white matter. Proliferating astrocytes and/or gliosis replace in part the interrupted ependymal cell line (glial nodules). In chronic hydrocephalus with high-­pressure hydrocephalus, a flattening of the gyral crests is seen. In addition, a small reactive glial zone around the ventricular system may

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Necrosis  Necrosis (for review, Lindenberg 1982) is commonly used to designate the death of tissue components. The most common cause of necrosis is ischemia. Other causes include mechanical injury (contusion necrosis), toxic agents (for example, by formic acid in methyl alcohol), heat (thermocoagulation), or freezing (cryosurgery). Whereas transient ischemia only destroys neurons and oligodendrocytes (incomplete necrosis or selective neuronal necrosis, see Scholz 1953), prolonged ischemia (“infarction”) gives rise to complete necrosis of all tissue components.

Apoptosis  Apoptosis is the programed death of a cell as regulated by specific death genes (for review, Clarke 1998). It initiates a delayed secondary death of neurons in response to environmental changes, deficient metabolic and trophic supply, and an altered gene transcription. About 4 hours after a traumatic event, apoptosis begins, and remains demonstrable for about 3 days. The characteristic

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Examination of the brain

Gross investigation of the CNS After removing the brain from the skull at autopsy, it should be fixed in an ample quantity of 10% buffered formalin (10× of brain volume) for 2–4 weeks, prior to the neuropathological examination. If the brain is examined fresh, about half of lesions will be missed compromising the reconstruction of pathophysiological alterations leading to injury or even death as well as reconstruction of the course of action. Generation of morphologic artifacts in the formalin solution can be avoided by adding salt, which allows the brain to float freely in the solution. The other possibility is to suspend the brain using a string tied to the basilar artery. First, the dura should be inspected for lacerations, epidural or subdural hemorrhages, or other findings. After weighing the brain, it is important to look at the basis of the brain to get an idea of the status of the vessels and to notice if there are basal signs of brain swelling. In cases of basal subarachnoid hemorrhages, it is mandatory to remove the unfixed coagulated blood to detect a ruptured aneurysm, otherwise an artificial rupture may be inflicted. Cerebrospinal fluid should be removed for toxicological analysis prior to fixation either and can be drawn from the large basal cisterna or by lumbar puncture through the intervertebral discs from the abdominal cavity. For microbiological, virological, and toxicological tests, it is recommended to remove small parts of tissue from regions without pathological findings such as the occipital lobe. For a professional neuropathological examination, it is of utmost importance to standardize the processing. After weighting the formalin-­fixed brain, the outer surface of the brain should be described in detail, including the formalin-­fixed dura. The outer examination should focus especially on signs of hemorrhages and brain swelling. Then, the brainstem including the cerebellum is removed from the cerebrum. Afterward, the brainstem is removed from both cerebellar hemispheres by cutting through the cerebellar peduncles. Both cerebellar hemispheres are then sectioned in a parasagittal plane. It is recommended to cut the cerebrum in about 0.5-­cm-­thick sections parallel to the forehead. Alternatively, the cerebrum can be cut parallel to a hat line, which

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Axonal injury  Axonal injury (AI) is characterized by an interruption of axonal flow. As a result of axonal damage due to ischemia or MBI, the axons will swell and develop bulbs. Moreover, AI induces an anterograde (Wallerian) and a retrograde degeneration of the injured axons. The terms “anterograde” and “retrograde” refer to the direction of conduction of the nerve impulse along the axon, that is, the degeneration following a focal damage proceeds in a centrifugal or a centripetal direction. The hindered anterograde flow of proteins along the axon can cause the morphology of AI, that is, swelling of axons up to bulbs. This phenomenon was once demonstrated by H&E staining and by silver staining techniques within 16–24 hours after a traumatic event. But since injured axons are selectively characterized by expression of β-­APP, it is now routinely confirmed within 105– 180  minutes. It is important to stress that AI is a nonspecific phenomenon also associated with mechanical trauma, acute intoxication, or ischemia. Moreover, there is a scientific discussion if the β-­APP reactive axons are reversibly or irreversibly injured.

The number and type of inflammatory cells in the CNS vary widely depending on the attracting stimulus or on their inherent ability to attack a CNS antigen. Polymorphonuclear leukocytes (PMNs) are the first circulating leukocytes to reach the site of injury and are capable to pass the BBB. The deleterious effect of neutrophil recruitment is the development of cerebral edema as has also been shown in models of MBI. After entering the CNS, the function of lymphocytes is to recognize their antigen. Under inflammatory conditions, microglial and perivascular cells constitute the chief antigen-­presenting cells. Under pathological conditions, monocytes emigrate into the brain parenchyma, where their morphology and antigenic characterization both change. They now participate in the immunological process as macrophages and express major histocompatibility complex (MHC) class II antigens.

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morphology of apoptosis exhibits cleavage of the internucleosomal chromatin that can be identified in situ using the terminal deoxynucleotidyl transferase dUTP nick-­ end labeling (TUNEL) method. Apoptosis causes pyknosis of the nucleus and condensation and shrinkage of the cell body. As it progresses, budding and karyorrhexis occur, and ultimately a breakup into clusters of apoptotic bodies. Reactive changes are extreme rarely detectable.

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AI is detectable as a focal and a diffuse phenomenon: a focal accumulation of AI is detectable in MBI as well as in ischemia caused by a regional arterial obstruction, while diffuse axonal injury (DAI) is induced by global ischemia and acceleration-­ induced brain injury.

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Regenerative capacity  It is accepted as common knowledge that plasticity-­associated molecular and structural events occur in the injured brain. These are at least partly responsible for functional recovery. Increases in dendritic arborization, spine density, and synaptogenesis in both peri-­injury and intact cortical areas are the potential morphological strategies that enable the brain to reorganize its neuronal circuits. It is also accepted that induced both retrograde degeneration and the axonal injury-­ bulbs and swelling of the proximal axonal stump are markers of a regeneration process. Neurons are capable to renew throughout life from endogenous stem cells, especially in the hippocampus. Inflammation  The brain itself is an immune organ, and the targets to be protected are neurons, axons, dendrites, and myelin. In the absence of protection, these cells become necrotic or succumb to apoptosis, are phagocytosed, and disappear. Astrocytes and microglial cells possess many immunological features marking them as important immunoregulatory cells and hallmarks of CNS inflammation.

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Spinal cord and cervical spine

In cases of blunt injury to the head, global permanent ischemia or suspected shaken baby syndrome is recommended to investigate the cervical spine including the occipitocervical junction to get additional information regarding the cause of death. The removed cervical spine is completely fixed in formalin for 2–4 weeks. After

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For a microscopic investigation, it is recommended to standardize this procedure either. As indicated in Figure 20.4.2, specific topographic regions from each brain should be taken in routine investigations. These regions include (1) a block of the cleft between first and second gyri, (2) the putamen and globus pallidus, (3) the thalamus, (4) the left and (5) right hippocampi, (6)

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Microscopy

the corpus callosum, (7) the cerebellum, (8) the pons, and (9) the medulla. In dependence of disease or gross lesions, additional blocks should be collected. Always, it is necessary to examine one edge of the injury as well as intact adjacent tissue. The paraffin-­embedding procedure of the selected blocks must avoid a rapid dehydration or overheating of paraffin, which can induce artificial shrinking of neuropil and cells. For the examination, paraffin-­embedded tissue blocks are cut in about 5-­μm-­thick pieces and transferred to glass slides. For routine investigations, the tissue is stained with H&E. To answer specialized forensic questions, it is often necessary to identify distinctive structures or cell types. Table 20.4.1 gives an overview of routine staining techniques, histochemical methods, or immunohistochemical techniques to obtain best results to identify a certain structure.

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is recommended in cases of extensive putrefaction because a transverse section (cf. Flechsig) of the complete brain is the only method to get any valuable information of the status of the brain. Postmortem imaging gains increasing importance because continued technical improvements have increased the sensitivity of imaging procedures. These techniques are more and more in demand because society grows less tolerant toward autopsy. Nevertheless, at the moment a forensic autopsy is the “gold standard,” but imaging techniques provide valuable supplementary information and biometric data in cases of MBI. In addition, computer tomographic (CT) scans and magnetic resonance imaging (MRI) scans are very important for biometric analysis and reconstruction, and are extremely useful for 3D imaging of skull fractures.

Figure 20.4.2  Collection of different topographc regions of the human brain to perform a routine microscopic examination in forensic neuropathology. It is recommended to analyze (1) the cleft between first and second gyrus, (2) the putamen and globus pallidus, (3) the thalamus, (4) the left and (5) right hippocampus, (6) the corpus callosum, (7) the cerebellum, (8) the pons, and (9) the medulla.

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Table 20.4.1  Routine staining methods and immunohistochemistry methods for the detection of different cells and structures in forensic neuropathology. Target

Routine methods

Immunohistochemistry

Neuron Neurons Axons (norm.) (pathol.) Dendrites Myelin Synapses Astrocytes

Cresyl violet, Nissl staining Cresyl violet Silver techniques

Neuron-­specific enolase (NSE)

Glial fibers Oligodendrocyte

Silver technique Silver technique

Microglia (norm.) (pathol.) Endothelial cells Reticulin fibers Polymorphonuclear leukocytes (PMNs)

Silver technique

Neurofilament protein (NFP) β-­amyloid precursor protein (βAPP) Microtubule-­associated protein (MAP II) Myelin basic protein (MBP) Synaptophysin Glial fibrillary acidic protein (GFAP) Vimentin S100 protein

Luxol fast blue

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A closed brain injury is characterized by an intact dura mater; by contrast, an open brain injury usually is characterized by a laceration of skin and dura mater. Penetrating brain injury is caused by bullets or other flying objects, by sharp or pointed implements, or bar-­like weapons. Closed brain injury results from the impact suffered in traffic or sports accidents, falls or explosions, from violent blows delivered by hands and feet/shoes, weapons, or objects falling on the head. With regard to their possible consequences, mechanical injuries of the coverings, for example, injuries of the scalp, skull, dura, and leptomeninges, must be distinguished from injuries of the brain parenchyma. Whereas scalp injuries entail the risk of complicated wound-­healing processes, which may prove fatal if secondary bacterial infection reaches intracranial structures, skull injuries involving bone fractures and/or bleeding can lead to space-­occupying processes with shifting of the brain and/or herniation, sometimes accompanied by brainstem hemorrhage. A distinction must also be made between trauma caused by local impact (contact force) and trauma associated with head motion, that is, with acceleration, deceleration, and rotational movements (noncontact or inertial force). Impact-­induced injuries of the brain are commonly associated with cortical hemorrhages on the side of impact (homolateral = “coup” contusion injuries) and/or focal hemorrhages on the opposite side (misleadingly called “contrecoup” contusion injuries), both known as “contusion” or “contusional hemorrhage” as well. Coup and contralateral cortical hemorrhages are characterized pathologically by pericapillar cortical hemorrhages. Injuries of the dura and leptomeninges may be associated with both impact and inertial forces. Distinction must be made between focal and diffuse brain injuries, as well as between primary mechanical injury and secondary changes, especially hypoxia, ischemia, and brain edema. Last but not least, we have to

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fixation, it is sawed along the midline in a sagittal direction. Vertebral bodies and vertebral discs are evaluated for fractures, hemorrhages, or luxations (e.g., in cases of hanging). For the detection of dural hemorrhages, the spinal cord and the dura are removed. The soft tissue including the spinal cord should be investigated regarding hemorrhages, scarring, necrosis, or cord compression. For the examination of the whole spinal cord, the total spine should be removed either anterior or posterior like it is practiced in routine pathology. Following fixation, the examination should begin with an inspection of the outer surface of the cord including coverings. The cervical cord and the total spinal cord are then cut in transversal planes and 0.5-­cm-­thick pieces are carefully investigated. If gross findings are detected, a microscopic examination has to follow. In some cases of normal gross findings, a microscopic analysis can give additional information on axonal injury, cell reactions, demyelination, etc. A comprehensive examination enables the neuropathologist to answer questions especially regarding vitality of inflicted lacerations or time point of the cervical alterations.

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Silver technique Naphthol AS-­D chloroacetate esterase

20.4.2  Mechanical injury of the head and spine Basic principles

Physical trauma Physical trauma means damage inflicted to tissue by an external force. Damage of the brain due to an impact of the head is differentiated according to whether the injury is penetrating or nonpenetrating, and the result is called open or closed craniocerebral injury.

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reactive neurons, traumatic AI reveals a local diffusely scattered pattern or groups of AI along the axis of the axon.

Types of mechanical head injury

Secondary lesions

The following types of morphological changes can be observed following injuries to the head and brain. Deformation of bones of the skull is caused by a fall or blow to the head. Depending on the velocity, mass, contact area, and hardness of the impacting object, there may be perforation fractures, depressed fractures, linear fractures, or skull deformation without fracture. Skull fracture is a contact injury. If the impact involves a small area of contact releasing a large amount of energy, the result may commonly be a perforation fracture or depressed skull fracture. On the other hand, linear fractures are observed, if the contact involves a large surface area. Linear skull fractures are usually caused by falls and not by blows. If tissue is pulled, drawn, or stretched beyond its tolerable limits, strain or tension injury results. Shearing forces are created when part of the brain moves at a different speed or angle than surrounding tissues. The consequence will be DAI and petechial hemorrhages, which can be demonstrated by light microscopy. Torsional stress is created when one part of an object is twisted in one direction, while another part remains motionless or is twisted in another direction. The result will be parenchymal laceration and hemorrhage such as gliding contusional injuries, both of which can be demonstrated at autopsy. An acute stopping of a moving head by striking the ground or a rapid impact of sufficient intensity leads to contusional hemorrhages caused by temporary high pressure at the pole of impact and negative pressure at the region opposite to the impact. If this negative pressure vanishes, collapsing gas bubbles within blood produce vessel damages and focal cortical hemorrhages (cavitation theory, Sellier and Unterharnscheidt 1963). Lacerations are actual tears, not only in the cortex but also in the white matter. Gliding contusional injuries of the white matter occur on the superior surfaces of the cerebral hemispheres, especially in the first parietal gyrus, when the medial parts of the temporal lobes are impacted against the dura mater and the cerebellum against the foramen magnum at the time of injury. This type of wounding is commonly caused by the shearing forces generated by acceleration. DAI and the accompanying tissue tear hemorrhages are considered to be due to high levels of global shear and tensile strains as well as strain rates as a result of rotational and translational head acceleration. These acceleration-­induced tissue strains are associated with local brain displacements within the deep white matter measured by Hardy et al. (2001) and calculated by Kleiven and Hardy (2002). DAI is biomechanically characterized by a noncontact trauma caused by inertial forces as a result of abrupt cranial deceleration or sudden angular motion of the head. DAI usually requires “rotational” acceleration of the brain. Focal axonal injury is also detectable in ischemic injury. Ischemic (vascular) AI is to be distinguished from traumatic AI: While vascular AI exhibits a zigzag, a wavy, or a focal pattern of β-­APP

All of the abovementioned primary injuries are capable of giving rise to secondary lesions, the most common being brain swelling and ischemia, both may lead to herniation. The consequence of an intracranial space-­occupying cortical hemorrhage and/or an edema-­induced intracranial increase in volume due to general or perifocal edema or hypoxic/ischemic edema is displacement of the brain parenchyma in a caudal direction. Space-­occupying processes as supratentorial hemorrhages, either subdural or epidural, may cause central transtentorial herniation. Such lesions displace the cerebral hemispheres and basal nuclei downward, pushing the diencephalon and adjacent midbrain through the tentorial notch. Uncal herniation (Figure  20.4.1b) results from lesions expanding in the lateral middle fossa or temporal lobe, displacing the medial edge of the uncus and hippocampal gyrus medially and over the ipsilateral edge of the tentorium cerebelli. Compression of the midbrain occurs, as well as compression or stretching of the contralateral or ipsilateral third cranial nerve. In its classic form, uncal herniation syndrome entails compression of the posterior cerebral artery and ipsilateral cerebral peduncle and stretching of the ipsilateral oculomotor nerve. The resulting symptoms include contralateral hemiparesis, and a dilated ipsilateral pupil that does not react to light. Compression of the opposite cerebral peduncle against the contralateral edge of the tentorium (Kernohan’s notch) will induce hemiplegia ipsilateral to the mass lesion. Such secondary changes, which are common for all types of intracranial space-­occupying processes, can produce focal hemorrhages in the corpus callosum, cingulate gyrus, hippocampal gyrus, and in the brainstem. Tonsillar herniation is caused by compression of the cerebellar tonsils into the foramen magnum. This is sometimes associated with brainstem compression with secondary bleeding within the midbrain or pons (Figure 20.4.1d). The brain itself can be affected secondarily without suffering primary injury: mechanical injury to other organs can induce a generalized hypoxia/ischemia with hypovolemic shock or transient asystole. The brain is also susceptible to the sequelae of cerebral fat embolism, pulmonary embolism, etc. Mechanical trauma to the CNS entails a threat of secondary injury to all vital organs, the loss of function of which can lead to death. The consequences of a secondary focal or global ischemia are described in Chapter 4. Fracture of the base of the skull entails a risk of hemorrhage and vomiting; aspiration of the vomit or blood can lead to asphyxiation. This is especially likely if the victim is comatose and unable to swallow.

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classify mechanical spinal cord injury according to the same criteria as mechanical head injury.

Forensic aspects Brain injuries may be caused by a violent impact to the head. The head strikes or is struck by an object, for example, by falling to the ground, being thrown against the dashboard of a car, or by receiving

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If mechanical head injury is followed by death of the victim, the precise cause of death has to be determined and/or whether there was a causal link between the traumatic event, the brain injury, and the death. In general, (minor) brain hemorrhage is not invariably lethal and should not be indiscriminately regarded as responsible for a death if found post mortem. Even a depressed fracture will not always lead directly to death or to neurological deficits. The brain injury itself must be severe enough to explain the death from the external event; if it is not, another cause of death must be sought. The causal link may be difficult to establish in cases with preexistent diseases such as atherosclerosis, cardiac decompensation, etc., as well as in cases with a long survival period. Decisions can be made possible by a detailed analysis of the clinical history as well as the injuries and careful scrutiny of information on the period after traumatic event including therapy. The extent of secondary changes (e.g., edema, hemorrhage, ischemia, thrombosis, embolism, etc.) occurring after impact partly is dependent upon time of survival. These alterations may in turn induce further acceleration of changes (edema, hemorrhages, hypoxia, anoxia, acidosis, embolism, etc.). A neural shock—­especially spinal shock—­must also be considered as a possible cause of death. The severity of an injury to the CNS thus depends not only on the strength of the external force, but also on other factors. These include the victim’s age and/or chronic diseases, the ambient temperature, chemical influences (e.g., drugs and alcohol), injuries to other organs (e.g., cardiorespiratory trauma), and the clinical sequelae.

bacterial infection that may spread into the intracranial cavity and cause intracranial abscess or meningitis. Cerebral trauma is nearly always accompanied by injury to the skin and soft tissue of the face or scalp. As a rule, any visible injury to the head (hemorrhage, laceration) is a marker of a traumatic event and may be suspected to lead to immediate or delayed functional disturbance of the brain. Therefore, in cases of suspected MBI at autopsy, the hair must be carefully shaved to expose the scalp for careful examination and documentation; the facial skin must be carefully dissected. In regard to localization of scalp injuries, the following rule of thumb may be applied for an upright person: a scalp injury located above the line of the hat brim, that is, toward the top of the head, is probably caused by a blow, and a scalp injury situated below the hat brim is probably due to a fall on an unwrinkled or smooth plane (Figure 20.4.3). Abrasion wounds caused by impacts perpendicular to the scalp often leave imprints that can provide evidence as to the type of weapon used. In cases of fall, the imprints may point to the nature of the object/surface struck. Additional abrasions may also be found on the scalp opposite the site of impact if the head has been pressed forcefully against the ground or other object. Contusion injury marked by pronounced swelling, a pressure mark, and/or a subcutaneous hemorrhage occurs at the site of impact. The swelling is due to edema and/or bleeding within the subcutaneous tissue and sometimes beneath the aponeurosis. If the force of the impact is sufficient, all layers of the scalp may be affected. At the area of the impact, the scalp shifts in relation to the surface of the skull or galea. The movement may cause tearing of connecting blood vessels in the scalp and between the scalp and the skull, resulting in a subcutaneous or subgaleal hematoma associated with an apparent laceration and/or bruising wound of

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a hammer blow. The resulting mechanical loading of the head and the resulting contact and inertial forces to the brain induce local near and remote strains and local and global movements generating vascular and brain tissue damage, causing focal and diffuse brain injuries (Gennarelli and Meaney 1996). Therefore, two different mechanisms are to be distinguished: Impact injury leads to local effects resulting from contact between the head and an object. The inflicted damage includes abrasion, contusion, and laceration of the scalp, skull fracture, epidural hematoma, subdural hematoma (SDH), and cortical hemorrhages. Acceleration injury results in intracranial pressure gradients and shearing and tensile forces to the brain and exhibits subdural hemorrhage, subarachnoid hemorrhage (SAH), intracerebral hemorrhage, gliding contusion injury, DAI, and cavitation.

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Blunt injury of the scalp Injury to the scalp and the face is usually caused by contact forces. Due to its high mechanical stability, the scalp is able to dissipate much of the energy released by the mechanical force of an impact. Hematomas, abrasions, and lacerations of the scalp are indicators of the location, severity, and number of impacts to the head, as well as of the shape and type of impacting object. A single scalp injury, however, may lead to fatal bleeding under special conditions. On the other hand, open scalp wounds entail a risk of

Figure 20.4.3  Rule of thumb: a scalp injury located above the line of the hat brim is probably caused by a blow; a scalp injury below that line is probably due to a fall.

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Compact bone is more rigid than trabecular bone and can withstand greater pressure, but less tension stress. Trabecular bone is more elastic and is well able to store and release energy. Skull fractures are the morphological result of contact violence, and only exceptionally of noncontact forces. Direct forces generated by local impact can cause cranial fractures, or, if they affect the base of the skull, basal skull fractures. Indirect forces released by a bullet can produce burst or tear drop fractures via the transfer of energy.

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Differences in fracture morphology can be attributed to differences in the underlying biomechanics. In case of a blow or a fall of an upright person on the ground, impact induces strains and deformations to the skull. If the impact energy exceeds the elastic deformation capacity of the skull, fractures will be produced. Local fractures close to the impact point are called direct fractures, and the global ones, remote from the impact area, are called secondary or indirect fractures. A fracture always starts (perpendicular to surface) on the bending-­ tension side and extends asymptotically to the bending-­pressure side. Therefore, bending fracture lines at the inner and outer compact layers are systematically displaced to each other. In skull loads by blunt impact, bending is always a locally induced mechanism at and around the contact area. If the area is small (16 cm2), a perforation of the skull rarely occurs. Since the formation and propagation of fracture occur perpendicularly to the local tensile strain direction, the typical outward appearance of bursting fracture is linear and finely jagged, exceeding radially from the impact region and/or extending far from it, while the fracture lines of the bending fractures normally are arched and smooth, circularly surrounding the contact area. According to their anatomic location, specific form, and mechanism of generation, specific types of fractures are observed. Linear calvarial fractures are bursting fractures resulting from stress distributed over the entire cranial vault. The skull cap is deformed as a whole and fractures as a whole, usually just exceeding, but remote from the impact region. Longitudinal compressing loads induce transverse tension and therefore longitudinal fractures (Figure 20.4.4a), just as transversal large blunt impacts, produce transverse burst fractures (Figure 20.4.4b). Bursting of

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the scalp. The severity of bleeding depends largely on the intensity of impact and the type of lacerated vessel and may involve isolated epidermal bleeding, subcutaneous bleeding, or bleeding within all the layers of the scalp. Laceration wounds of the scalp generally exhibit a part of total separation of tissue continuity surrounded by a zone of bruised tissue. The laceration injury may be linear, stellate, or Y-­shaped. The tear may either penetrate some or all of the layers of the scalp. Acute angled or tangential bruising may cause partial avulsion of the scalp. Bleeding from a laceration wound is diffuse and if prolonged it may culminate in fatal hemorrhagic shock. In most cases, the different types of wounds are combined because of the different forces which likewise influence the biomechanics of the scalp wounding. Pulling of the hair or shearing forces applied to the scalp may detach the scalp from the skull, which is termed avulsion injury. If the hair is pulled from behind, the skin in front will tear first, if pulled from the front, the occipital region tears first. If the hair on the top of the head is pulled, the skin will yield at the vertex and tear above the ears.

Figure 20.4.4  (a) A longitudinal basal skull fracture is the result of a load on the longitudinal axis. (b) A transverse basal skull fractue is the result of a load on the transverse axis.

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will be the result of this fracture system. If there are several impacts, the fracture systems might form a network, and according to “Puppe’s rule,” the later fractures will not cross the earlier ones but will terminate at the earlier ones, a reliable indication that the latter preceded the former (Figure 20.4.6). Basilar skull fractures are normally bursting fractures, and the causal mechanisms are the same as in linear calvarial fractures. The causal load can be a fall to the ground or being driven over by the wheel of a vehicle, a blunt impact to the neck, mandibular or facial impact, or an impact to parts of the occipital area and almost any type of diffuse impact to the vertex of the head. Whereas, a sagittal fracture at the base of the skull is the result of a load on the longitudinal axis, a load on the transverse axis gives rise to a transverse fracture (Figure 20.4.4a,b). A transversal basilar fracture which involves the petrous bone frequently leads to bleeding from the external auditory canal and can be manifested by this symptom. A basilar fracture of specific form and generation is the basilar skull ring fracture around the foramen magnum, which is created by axial tension or pressure as a local response resulting from contact or noncontact loading. The contact may be a mandibular impact or striking the ground with the top of the head. A noncontact loading may be an acceleration of the head away from the restrained body or striking the ground with the lower end of the spinal column, for example, by fall from a height.

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the cranial vault as a way to reduce induced tension stress may also lead to diastasis of suture lines even at a distance from the point of impact. Depressed fractures are direct bending fractures and result from the local impact of a blunt object to a small area of contact like a blow with a hammer. If the energy of a blow is low and exceeds just the local capacity of elastic deformation, the outer table is bent inwardly, and a dent fracture results as a superficial indentation of the outer table. The inner table is bent inwardly as well and tears. With increased impact energy, there results a depressed fracture with a dislocated, impressed area of the outer compact layer, frequently formed by the impacting instrument, surrounded by closely parallel fracture lines at the border region, which are called stair-­step fractures. Comminuted fractures are formed at the inner table, and a splinter pyramid always results, which projects into the interior of the skull, usually perforating the dura and penetrating into the subjacent brain. In instances of high-­energy impact and a small instrument, in which contact area does not exceed more than 16 cm2, the injuring object perforates the skull completely. Since a two-­dimensional bending fracture spreads out in a funnel shape from the exterior to the interior of the skull, a cone-­like fragment is punched out and propelled into the brain. This leads to a hole in the skull, sometimes shaped like the perforating instrument. Fractures like these are called penetration fractures or depressed fractures. Combined fractures are fracture systems consisting of combined bending and bursting, direct and indirect, near and remote fractures. If energy remains after local absorption has produced a bending or depressed fracture, this residual energy will be exhausted in the generation of circular bending fractures around the impacted area and linear burst fractures radiating in a star-­ like fashion from the center. The result is a fracture system called spider’s web fracture (Figure 20.4.5). A subarachnoid hemorrhage and a laceration of the brain structures beneath the contact area

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Dural hemorrhages

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Dural hemorrhages can be induced by both contact and noncontact loading. An isolated dural hemorrhage resulting from external mechanical force is not invariably accompanied by additional direct injury to the brain, especially if the hemorrhagic source is not cortical. If there is a gradual displacement of the brain caused by space-­occupying bleeding, secondary clinical symptoms will be delayed, that is, occur after a symptom-­free (lucid) interval, the length of the delay depending on the rate and amount of

Figure 20.4.5  Spiders web fracture of the skull as a result of combined bending and bursting.

Figure 20.4.6  Puppe’s rule: in cases of two or more impacts of the skull, the later fractures will not cross the earlier ones but will terminate at the earlier ones.

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Subdural hemorrhage

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SDH is defined as bleeding into the space between the dura and the arachnoid mater (Figure 20.4.8). The morphological picture is dominated by the intracranial shift of brain tissue and edema, usually becoming apparent in the contralateral hemisphere through obliteration of the gyral sulci and flattening of the gyral crests. SDH can occur bilaterally or unilaterally, in isolation or in combination with skull fractures, with or without cortical hemorrhages. Most SDHs are located over the cerebral convexities, associated with cortical hemorrhages, and result from tearing and stretching of the parasagittal bridging veins (Krauland 1961), which drain the surface of the cerebral hemispheres and the CSF into the dural venous sinuses. According to Krauland (1982), the sources of SDH are cortical hemorrhage (contusion injury) (60–70%), a tear in a bridging vein (2.5–20%), arterial injury (cortical part of the middle cerebral artery, 2.5–10%), and sinusoidal injury (4–9%). In 4–11% of cases, the source cannot be found, either because there

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EDH is defined as bleeding between the skull and the dura mater (Figure 20.4.7). The hemorrhage attaches tightly to the skull and dura mater at the borders of the sutures and has a lenticular, convex appearance in CT scans. The volume of blood varies greatly, but amounts to at least 100 mL in most fatal cases. EDH is caused by local impact loading of the head and is nearly always associated with contact injury and often with skull fractures. The incidence of EDH is 20 times higher in cases with skull fracture than in cases without fracture (Auer et al. 1989). EDH usually involves arterial bleeding (>50 %) and arises from injury to the middle meningeal artery and its branches near the impact site. In a typical case, EDH occurs when a fracture crosses the middle meningeal artery, which adheres tightly to the inner surface of the temporal bone. Edges of fractured bone can cause laceration of underlying dural arteries and, less frequently, veins. If a parenchymal lesion does occur, it may produce a concomitant SDH. If the mechanical loading does not induce immediate central nervous symptoms, the lucid interval between the generation of injury and appearance of the first clinical symptoms is commonly shorter than for SDH, in most cases lasting not more than a few hours (delayed EDH). In up to 30% of the cases, the interval can last up to days (Poon et al. 1992). Although primary mechanically induced intracerebral hemorrhage involving the brain parenchyma rarely occurs, DAI is often

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Epidural hemorrhage

seen in the brain parenchyma, especially in the corpus callosum and brainstem, sometimes without attendant hemorrhage. A space-­occupying EDH will displace the brain resulting in compression of the brainstem (herniation syndrome). Prognosis of EDH depends on the interval between onset of the space-­ occupying induced cerebral compression and the beginning of surgical decompression; this means that timely diagnosis and treatment are essential to a favorable prognosis. The patient’s life can only be saved by early surgical decompression. The mortality of EDH is high because even early surgical intervention does not guarantee the patient’s survival or complete recovery.

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bleeding. The dura hemorrhages are characterized by a clot on the outside, that is, epidural hemorrhage (EDH), or inner side of the dura mater, that is, SDH, both are often associated with an accompanying discrete or distinct subarachnoid hemorrhage. In most cases, an associated intradural hemorrhage will be observed as well as red blood cells (and siderophages) within the Pacchionian bodies.

Figure 20.4.7  Acute epidural hemorrhage.

Figure 20.4.8  Acute subdural hemorrhage.

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may result from the rupture of a cerebral arterial aneurysm or an intracerebral angioma into the subdural space. Recurrent spontaneous SDH has been observed (Matsuyama et  al. 1997) and is arterial in origin. In these cases, a compromised vessel tears spontaneously, often caused by a hypertensive peak in blood pressure or preexisting damage to an arterial wall.

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Biomechanics of  SDH  Biomechanical conditions sufficient to cause SDH can be generated by a blow or fall. About 30% of all cases of SDH involve isolated SDH without associated skull fracture or cortical hemorrhages, generally caused by acceleration (Gennarelli and Thibault 1982) or blow. A blow to the chin can induce rotational movement sufficient to cause tears of the bridging veins and/or cortical and basal arteries. The resulting SDH may be bilateral or may occur on the same or contralateral side of the impact. If angular acceleration is low and of long duration, as it happens in traffic accidents, strains are propagated deep within the brain and cause DAI. Increased acceleration can result in acute SDH combined with DAI and tissue-­ tear hemorrhages. Such acceleration can be produced by falls in which the head strikes a hard surface or a blow. Rotation is most likely to cause tearing of the bridging veins along a transverse or diagonal–frontal axis, the greatest displacement of the skull relative to the brain occurring along the midline. If there is rotation around the vertical axis, which is often associated with translational acceleration, the brain is displaced against the skull in the parietal region, injuring the cortical arteries and inducing parietal cortical hemorrhages.

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is no visible accompanying local SAH or because the traumatic event was inflicted long ago and clear signs of the source are not detectable due to scarring of the tissue. SDH is three to five times as common as EDH. Acute, subacute, and chronic courses can be differentiated clinically and morphologically. Special forms of SDH are hemorrhagic internal pachymeningitis and subdural hygroma (SDHy). Fresh coagulated blood between the arachnoid and dura mater is characteristic of acute SDH and can be diagnosed by clinicians or pathologists in the first three days following the traumatic event. The loosely coagulated blood drains off at autopsy, leaving no—­or only minimal—­visible residues on the inner surface of the dura and brain after formalin fixation. A SAH and/or displacement of the cerebrum with unilateral flattening of the hemispheres are present in most cases. A SAH is defined as bleeding within the subarachnoid space. A rapidly developing SDH becomes life-­threatening in adults once its volume has attained 50 mL (Di Maio and Di Maio 2001). Most cases of SDH are accompanied by laceration and/or contusion injuries of the cerebral cortex. The ruptured vessel can be detected in some cases of acute or even subacute SDH. On removal of the brain, an operating microscope and contrasting agents or dye may be applied for demonstration of the vessel’s rupture. The site of a venous (or arterial) rupture often exhibits small focal SAH, which should be examined microscopically to confirm (or exclude) primary vascular disease, especially if nonaccidental injury is suspected. A concomitant old SDH (chronic SDH, see in the following text) is often found, especially if the victim was an alcoholic, an elderly person, or in cases of abusive head trauma (AHT, see Chapter 3). Spontaneous or mechanically induced rebleeding into an old SDH is common. If the SDH develops slowly (subacute SDH)—­within >48–72 hours and 1 hour) and other neurological deficits, for example, paralyses, speech disturbances, convulsive disorders, etc., which can be temporary or of long duration. Morphological changes are quantified using the contusion index, which assesses both the depth and severity of cortical hemorrhages (Table 20.4.3), and the principle site(s) of injury (Adams et al. 1980, Adams et  al. 1985). Simultaneous injuries of other

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Figure 20.4.9  Findings in cases of gunshot to the brain: 3D CT reconstruction of (a) the entrance hole and (b) exit hole of the skull. (c) Entrance and (d) exit holes of the skull at autopsy. (e) Straight permanent missile track cavity of the brain. (f) Zones of injury around the permanent missile track indicating tissue damage secondary to the temporary cavitation.

organs such as contusion of the heart can aggravate the effect of brain-­injury-­induced ischemia via secondary disruption, for example, of the blood–brain barrier (brain edema). We have to differentiate between cortical hemorrhages and intracerebral hemorrhages, focal and diffuse injuries (Graham et al. 1995), and contact injury and injury inflicted by acceleration forces. In addition to these primary mechanically caused brain injuries, secondary diffuse injuries may also occur, including edema, ischemia, and vascular injuries.

Concussion injury and cortical hemorrhage A blow is likely to cause contusion injuries or cortical hemorrhages of the brain directly beneath the impact site (“coup” hemorrhages), whereas a fall tends to induce cortical hemorrhages at a location diametrically opposite the point of impact (“contrecoup” cortical hemorrhages). A fracture of the skull is not necessarily detectable. These contralateral cortical hemorrhages are the result of the negative pressure described in the earlier text and

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Table 20.4.3  Contusion index of hemorrhages of the human brain. Depth of mechanically induced hemorrhages (D) Not present Partial thickness of the cortex Full thickness of the cortex Extending into digitate white matter Extending into deep white matter

0 1 2 3 4 0 1 2 3

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Extension of hemorrhages (E) Not present Localized (one gyrus or two adjacent gyri) Moderately extensive (involvement of the greater part of the surface of one lobe) Extensive (more than the surface of the lobe involved)

Forensic medical experience shows that if an adult slips while walking and falls on his back without reacting to cushion the force of the fall and strikes the back of the head on a hard level surface, the force of the impact is just sufficient to cause a bursting skull fracture. In some cases, the impact forces are strong enough to generate a fracture of the thin roof of the orbits leading to a binocular hematoma. Fall-­induced cortical hemorrhages tend to occur at sites opposite of the site of impact. Since in falls the most common impact site is the occipital area, cortical bleeding is encountered mainly in the frontobasal area. Forward falls on a level plane are frequently accompanied by reflexive actions to break the fall, whereas falls to the side are often cushioned by the intervening shoulder. Those arguments may explain that contralateral cortical hemorrhages in the occipital lobe are very rare. When a temporal impact occurs, the contrecoup injury may not be on the contralateral hemisphere, but on the opposite side of the same hemisphere adjacent to the falx cerebri. A blow inflicts brain injuries mainly at the impact side (impact pole). If the skull remains intact, injury of the brain is due to local deformation (depression and recoil) of the bone. Acceleration is lower at the contralateral pole and the resulting injuries—­mainly caused by negative pressure—­are minimal or absent altogether. In contrast to falls, massive hemorrhages develop at the impact site and are limited not merely to the gyral crests, but can extend deep into the white matter. A blow to the head with small surface areas generally does not create the critical negative pressure at the contralateral pole. At the impact pole, a depressed skull fracture is inflicted. The fractured bone causes lacerations of the dura or leptomeninges as well as of the surface of the cortex. Depending on the force of the impact, destruction of brain tissue occurs with massive, space-­occupying hemorrhaging. Cortical hemorrhages located on the side of impact are a direct result of the forces delivered by the blow. The most common impact site is the parietal region. If the head is resting on a hard surface when struck, it cannot be accelerated, so contralateral cortical hemorrhages are not observed or usually minimal.

Contusion index (CI) = D x E (for each anatomical locator)

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not the result of an impact. In case of a fall, the contrecoup laceration usually is more pronounced than the coup on the impact side. Sometimes, no coup damage may be observed, but only contrecoup cortical hemorrhages. The dynamic load exerted on the brain by blunt impact to the head is generated by positive and negative pressures and shearing forces. Numerous hypotheses exist as to which of these loads are responsible for which types and localizations of brain injury (for review, Leestma 2008). One hypothesis that has found general acceptance is the cavitation theory. According to Sellier and Unterharnscheidt (1963; cf. also Bandak 1997), the foci of cortical bleeding at the contralateral pole are not due to compression (contusion) as a result of an impact at the contralateral site (“contrecoup”), but to the cavitations created by negative pressure. According to these authors, the cortical hemorrhages result from an external hard blunt impact with short impact time and sufficient force accelerating the skull, creating transient positive pressure at the impact site and—­depending on the degree of skull deformation—­a temporary zone of lowered pressure at the contralateral pole. If the vapor pressure remains below normal levels (negative pressure), gases (N, CO2, O2) dissolved in fluids such as CSF and blood are released as bubbles (cavitations), with consequent rupture of small vessels as a result of the following collapse of these bubbles. Other forces, however, also have an (additional) effect (Gennarelli and Thibault 1982), particularly rotational shear forces. Cortical hemorrhages occur with greater frequency at the frontal and temporal poles and on the inferior surfaces of the frontal and temporal lobes than they do elsewhere in the brain, and cortical hemorrhages are rare at the occipital poles. Acceleration and rotational forces may create shearing which results in tearing of axons and myelin sheaths, sometimes also of vessels in the depths of the white matter with consequent immediate loss of consciousness and simultaneous diffuse injury. Hemorrhages can also occur in deep brain areas distant from the site of impact, which may be due to shearing or tensile forces or as a result of localized motions within the brain.

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Concussion injury and contusion The lowest grade of brain injury means “cerebral concussion” or “commotio cerebri,” which produces a temporary loss of consciousness but no long-­lasting neuronal deficits. The morphological appearance is characterized at most by signs pointing to a slight increase in intracranial pressure. Marked or repeated mechanical injury of the brain produces submicroscopic mitochondrial swelling in the nerve cells and a major breakdown in the blood–brain barrier. Contact-­induced loading to the brain leading to tissue damage that induces cortical hemorrhages at the gyral crests is termed contusional injury. The term “contusion” is applied if the pia mater is not breached, the term “laceration” if it is torn. Contusional injuries and lacerations are both the result of the brain tissue movements within the cranium. Mechanical loading to the head gives rise to neurological deficits and visible morphological changes. The main injury is a hemorrhage, mainly of the

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contact, that is, by indirect forces. Brainstem injuries have a poor prognosis; only a few cases survive, most cases die shortly after the occurrence of a hemorrhage. According to Bratzke (1981) and Krauland (1982), mechanically induced (primary) hemorrhages at the base of the cerebral peduncles, at the base of the fourth ventricle, in the interpeduncular fossa, and/or in the rostral portion of the pons are primary loading-­induced (noncontact) hemorrhages. They are thought to result from rotational acceleration-­induced ruptures of small arteries and roots of Galen’s vein. Primary (contact) hemorrhages tend to be located at the margin of the midbrain (Bratzke 1981) and are often associated with fractures of the base of the skull. Acceleration loadings subject the brainstem to massive shearing forces, which can lead upon rotation to tears at the branching of the large basal arteries and thus to SAH. Secondary brainstem hemorrhages are caused by a rapid increase in axial pressure produced by supratentorial space-­ occupying processes such as epidural and subdural hemorrhages, intracerebral hemorrhages, or severe cerebral edema. Those expanding processes can unilaterally or bilaterally compress, displace, and distort the brainstem. A nonmechanically induced increase in axial pressure with a space-­occupying process such as edema, ischemia, tumor, or spontaneous hemorrhage may rarely result in brainstem hemorrhage. Such brainstem hemorrhages are also called “Duret hemorrhages.” Intraventricular hemorrhage requires extremely severe external violence to the head and carries a poor prognosis (LeRoux et al. 1992). In some instances, it is caused by impact along the sagittal plane of the head that increases the minor axis and thus the ventricular diameter. The resultant deformation of the skull can generate tensile strain of subependymal veins. Intraventricular hemorrhages are almost always associated with DAI and tissue-­ tear hemorrhages, suggesting an additional mechanism involving angular acceleration-­induced shear strain. In addition, the sharp edges of the dura mater may cause lacerations if the brain is pressed against it. These lacerations are generated by acceleration forces, leading to cutting or bruising of the brain (median part of the hippocampal area, corpus callosum). Second, a hemorrhage as well as an edema-­induced brain swelling may force the brain against the dura, resulting in herniation of brain tissue with consequent structural distortion, stretching, and compression injuries. Often—­especially after rotational acceleration—­a hemorrhage is found in the white matter of the first frontal lobe gyrus, that is, a so-­called gliding contusion injury. Moreover, axonal injury can almost always be demonstrated in cases with survival times of >2–3 hours, most frequently in both the corpus callosum and the rostral portion of the brainstem as an indication of DAI.

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gyral crests of the cerebral surface especially of the gyri of the frontal basis, partly spreading to adjacent white matter (Figure 20.4.10). Deformations of the skull creating a direct compressive strain exceeding the tolerance of the pial vessels and brain tissue will give rise to a contusional injury at the impact site. Cortical hemorrhages opposite the impact site bear a morphological resemblance to their counterparts near the site of impact. Cerebellar contusional injuries and hematomas result from substantial local pressure to the occipital area. The musculature of the neck, the massive occipital skull bone with its smooth and regular internal contours, the elastic tentorium of the cerebellum in combination with the “fluid cushions” of the subarachnoid cisterns, and fourth ventricle constitute a “buffer” that greatly reduces the energy acting upon the cerebellum. Dorsal cerebral and cerebellar contusional injuries and hematomas are therefore much less frequent than homolateral or contralateral (cortical) hemorrhages in other regions of the brain. If cerebellar cortical hemorrhages are detectable, they occur in the gyral crests of the cerebellar surface, usually at the occipital poles. Every infratentorial hemorrhage is capable of causing an obstructive hydrocephalus and herniation of cerebellar tissue into the foramen magnum and incisura tentorii. Therefore, the sequelae of posterior fossa hemorrhages may be fatal.

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Figure 20.4.10  Basal cortical hemorrhages of the brain.

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Intracerebral hemorrhages External mechanical-­ loading-­ induced primary intracerebral—­ including intraventricular—­ hemorrhages are more common than once thought. The presence of galeal hemorrhages, the proof of additional cortical hemorrhages, and/or the phenomenon of an association of homolateral and contralateral wounding of the brain gives an important hint for a mechanically induced intra­ cerebral hemorrhage. Primary impact-­ induced intracerebral hemorrhages are also to be differentiated from secondary, that is, delayed, “traumatic” intracerebral hemorrhages. Primary mechanically induced brainstem hemorrhages are to be distinguished from secondary herniation-­ induced hemorrhages. Primary brainstem hemorrhages may be induced either by contact (skull fracture contra brain tissue) or—­rarely—­by non-

Chronic traumatic encephalopathy Chronic traumatic encephalopathy (CTE) is a neurodegenerative disease which displays severe and irreparable brain damage and is caused by repeated blunt force injuries to the head over a period of years. CTE is found in contact sports such as boxing, rugby, or soccer, but also in victims of domestic violence. Because these alterations were first described in the brain of boxers, the disease was called punch drunk syndrome or dementia pugilistica. Boxers who have suffered repeated knockouts are especially likely to

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detected in individuals with substance abuse, amyotrophic lateral sclerosis, or other neurodegenerative diseases. The described findings indicate that CTE pathology may not be specific for repeated blunt force injury to the head (Iverson et al. 2019).

Estimating of the posttraumatic survival time of contusional hemorrhages

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The determination of time-­dependent processes of contusional hemorrhages may be important in some cases for classifying the time of the traumatic event. It is well known that reactive processes in sterile inflammations follow a regular time course somewhat resembling a chain reaction (Oehmichen 1990). Most of these cytological criteria are detected by routine staining (H&E, Prussian blue, Elastica van Gieson (EvG), etc.) and can be supplemented by additional immunohistochemical (e.g., CD68, GFAP, β-­APP, etc.) or histochemical reactions (naphthol AS-­D chloroacetate esterase (NASD-­ClAE), scarlet red, etc.). A survey of first appearance as well as additional statistical data and last appearance of a definite morphological phenomenon is given in Table 20.4.4. An actual survey on the time-­dependent histological alterations is given by Oehmichen et al. (2003), associated with important statistical data (for further information, especially regarding the interaction of macrophages, see Oehmichen et al. 1986; Oehmichen et  al. 2009b). In contusional hemorrhages, ­polymorphonuclear leukocytes can be demonstrated as soon as 5 minutes after traumatic brain injury (TBI). While the invasion

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develop both neurological symptoms and progressive dementia years after the last boxing-­related traumatic event. CTE can be classified as a tauopathy. A reliable diagnosis of the disease is only detectable by detailed analysis of postmortem brains. Helpful gross autopsy findings are reduction of brain weight with cerebral atrophy of frontal and temporal lobes, changes in the septum pellucidum with enlargement of the cavum, and fenestration of the interventricular septum, enlarged ventricles, and degeneration of the substantia nigra. CTE is associated with atrophy of the olfactory bulbs,  thalamus,  mammillary bodies, cerebellum, ­ and brainstem. In later stages of the disease, there may be marked atrophy of the  hippocampus,  amygdala, and entorhinal cortex (Corsellis et  al. 1973, McKee et  al. 2015). Microscopic analysis reveals neurofibrillary tangles (NFT) in the cortex and brainstem, widespread diffuse Aß deposits, astrocytic tangles, and neuritis in clusters around small blood vessels, especially in the depth of the sulci. Abnormalities in phosphorylated 43 kDa TAR DNA-­binding protein are found in most cases. Clinical symptoms of CTE include memory loss, cognitive impairment, changes in mood and behavior, and dementia in later stages. Four clinical stages (stages I–IV) of CTE-­based disability have been observed, which correspond to ongoing tau pathology in brain tissue, ranging from focal perivascular NFT accumulation in the frontal neocortex to severe tau deposition affecting widespread brain regions (McKee et al. 2015). In recent years, there has been emerging evidence that described neuropathological alterations are not specific to CTE or progressive as a result of repeated brain trauma, but have been also

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Table 20.4.4  Time course of microscopic findings in cases of contusional hemorrhages.

TR IB

Observation period

Estimated limits of confidence (Clopper and Pearson 1934) (expressed in percentage)

Distribution-­free tolerance intervals with 95% reliability (expressed in percentage)

First appearance

Last appearance

RBCs

1 min

150d

77.70%

72.6–82.3%

87.5%

Polymorphonuclear leukocytes

5 min

110d

43.09%

37.5–48.9%

98.0%

Macrophages (Ms)

11.5 h

58a

73.11%

67.8–78.0%

96.4%

RBC-­containing Ms

12 h

150d

47.85%

42.1–53.6%

97.9%

4.2 d

44a

53.11%

47.3–58.8%

96.8%

12 d

1a

11.84%

08.4–16.0%

97.1%

Lipid-­containing Ms

17 h

30a

53.64%

46.8–60.4%

87.5%

Fibroblasts

4.4 d

9a

34.10%

28.8–39.7%

96.0%

Endothelial cells

3.9 d

53a

48.85%

43.1–54.6%

95.5%

Collagenous fibers

5d

58a

38.69%

33.2–44.4%

96.9%

Gemistocytic astrocytes

7h

35a

45.57%

39.9–51.3%

96.0%

Fibrillary gliosis

6d

58a

28.52%

23.5–33.9%

96.6%

Siderin-­containing astrocytes

5d

53a

33.11%

27.9–38.7%

94.7%

Neuronal destruction

1 min

150d

60.00%

54.3–65.5%

95.4%

Neuronophagia

4.8 h

125d

21.97%

17.4–27.0%

97.4%

Axonal swelling

2.8 h

125d

38.78%

25.2–53.8%

93.1%

Axonal balls

15 h

44a

50.71%

44.7–56.7%

77.4%

Mineralization of neurons

6d

44a

11.88%

08.5–16.1%

96.7%

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Histomorphologic alterations

Relative frequency of the observation

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of macrophages is usually seen after a few hours, it takes between 4 and 6  days before proliferation of capillaries, fibroblasts, and collagenous fibers begins (Oehmichen et al. 2003). Whereas some of the cytological criteria are always present (relative frequency 100%), some are never (relative frequency 0%) present during a certain posttraumatic interval. If a criterion is present in a given case, all intervals with a relative frequency of 0% for this criterion can be excluded; if a criterion is not present, all intervals with a relative frequency of 100% for this criterion can be excluded. A comprehensive investigation of a panel of cytologic criteria allows an estimation of the posttraumatic interval and can be refined by applying a logistic regression analysis and an empirical weighting factor, because each criterion is of different “importance” (cf. Kleinbaum 1998).

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The spinal cord is protected from blunt force injury by the surrounding—­predominantly muscular—­soft tissue, the vertebrae, and the spinal dura mater. While the segmental architecture of the bony spinal column allows flexibility of movement, it exposes the spinal cord to perforating injury through the spaces between the vertebrae. The nerve fibers radiating off the spinal cord are additional loci minoris resistentiae where the nerve ­fibers themselves are vulnerable to mechanical loading such as traction and compression.

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Basic principles

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columbar junction the most common sites of mechanically induced damage. Four mechanisms of closed spine injury can be distinguished. The most frequent mechanisms, especially of cervical spine injury, are transverse shear, longitudinal shear, and tension ­created by ventroflexive and retroflexive forces. By contrast, compression is produced by longitudinal forces acting on the column in cases of falls on the head, or bottom, which may result in ­flattening of vertebral bodies and/or fracture of the end plates. This is the principal mechanism of injuries at the lumbar or thoracolumbar level. Another type of injury is caused by rotation generated by torsional forces resulting in unilateral or bilateral dislocation, fracture dislocation of the vertebral body, and/or its processes. Compressive or tensile stress to the spinal column by external violence can dislodge the spinal cord in the sagittal or lateral direction or cause it to rotate around the longitudinal or lateral axes. At least, combined mechanisms lead to closed injuries because pure flexion or extension forces are often insufficient to produce ligamentous rupture, dislocation, or fracture dislocation (Gosch et al. 1972). In most instances, blunt mechanical loading (compression) produces no externally visible changes in the spinal cord. Petechial hemorrhages usually begin at the center of the cord. This is followed by a central hemorrhagic necrosis, chromatolysis, and acute swelling of nerve cells. In later stages, tissue softening and phagocytosis occurs, followed by cavitation and partial replacement of destroyed neural tissue by glial–mesenchymal scarring. The spinal cord can suffer primary damage if surrounding bony and soft tissues directly inflict the nervous tissue and vasculature, as commonly seen in fracture-­dislocation of the spine. Spinal SDH is often seen in combination with vertebral fractures and subarachnoidal spinal bleeding, but dural bleeding in the spinal canal, especially SDH, is rare. Dural hemorrhages are usually caused by spinal column injuries. Mechanically induced and spontaneous spinal EDH, which are often associated with ruptures of the dorsal parts of the intervertebral discs, are usually venous and spread diffusely. Subarachnoid hemorrhages are caused by contusions, accelerations, or other types of mechanical violence to the spine or may be secondary due to intracranial subarachnoidal hemorrhage.

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Closed spinal cord injury

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Closed spinal injury usually results from the mechanical stress created by a broad area of impact to the spine. Even if the load is rather low, the injury commonly is associated with a fracture or dislocation of the spine (Figure 20.4.11). The static conditions of the spine and cord make the lower cervical region and the thora-

Figure 20.4.11  Mechanical injury of the spine showing a fracture of the spine and hemorrhages within the spinal cord.

Open spinal cord injury The hallmark of penetrating spinal cord trauma is perforation of the dura mater spinalis caused by stabbing or gunshot, by a needle, or even by displaced fragments of the vertebral bones. Open spinal trauma can result in complete or partial paralysis below the lesion. Infection is one of the major complications of open injuries of the spinal cord. Spinal subdural, epidural, or subarachnoid infections, abscesses, and empyemas can develop, the inflammation usually originating in local infectious processes, for example, osteomyelitis. Gunshot injuries of the spinal cord always involve injury of the bony column. Injury of the spinal cord itself may be caused by the projectile, by bony fragments, or by both. Once lodged in

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In cases involving significant brain trauma, it is imperative to perform a complete pathological examination on the neck, vertebral vessels, and paraspinal muscles, partly even on the spine and the spinal cord, with special emphasis on the craniocervical junction, because a combined trauma is often observed. Morphological changes associated with acceleration loading of the cervical spine include hemorrhaging into the muscle and fat tissue, ligamentous tears in the spinal column, disc lesions, dislocations and subluxations of the vertebral bodies and small joints, bone fractures, intramural dissection, laceration and/or thrombosis of the vertebral arteries, epidural and subdural hemorrhages, compression or laceration of the spinal cord, sometimes associated with spinal hemorrhage, axonal injury, demyelination, edema, and ischemic necrosis. The medulla oblongata, which contains the neuronal centers for regulation of respiration and circulation, is located at the craniocervical junction. Mechanical loading to the cervical spine therefore can result in acute death. Three different kinds of direct craniocerebral injuries are observed. Dislocation injury of the cervical spine is produced by forces acting in an anterior–posterior direction or, less often, in a lateral direction (Sances et al. 1986). Compression fractures are due to compression of the intervertebral discs or bursting of vertebral bodies. Bending loads create both tensile and compressive stresses, the former being much more likely than the latter to cause severe, often visible primary tearing of tissue and blood vessels as well as hemorrhages. Hyperextension and/or hyperflexion fractures are characterized by rupture of the rim of the vertebral body plates and/or by tearing and bleeding of the dorsal (= hyperflexion) and/or anterior parts (= hyperextension) of intervertebral discs. Lateral loads create similar injuries due to tensile stress and are located on the right or left side of the vertebral bodies and/or intervertebral discs. Rotational forces, that is, torque around the longitudinal z-­axis, can disrupt the ligaments and displace the vertebral body, injuries that are sometimes associated with fracture of the odontoid, but usually not associated with neurological deficits. The main causes of such injuries are traffic accidents, blows, and chiropractic manipulation. Vertical forces (along the z-­axis), for example, created by an impact on the top of the head, can lead to compressed fracture of the atlas with bursting and separation of the ring of the atlas—­often into four parts (Jefferson’s fracture).

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Injury of the cervical spine

Hyperextension forces (retroflexion) can cause fracturing of the dens, the force being transmitted via the anterior ring of the atlas with consequent failure of the dens. A force conveyed anterior to posterior through the skull to the anterior ring of the atlas is transmitted to the dens where a fracture is inflicted due to shearing forces. Fractures can also be produced by hyperflexion force (antero­ flexion) and result in displacement of the vertebral body. The so-­ called broken neck, that is, fracture of the dens axis, is comparatively rare. In most instances, it is caused by hyperflexion forward with retroversion of the dens fragments toward the medulla that leads to  respiratory failure and may lead to rapid death. Injuries of the middle and lower cervical spine are caused by axial compression, or by translational hyperextension, or hyperflexion, sometimes associated with rotational components. Among the sequelae is ­disruption of the anterior (hyperextension) or posterior (hyperflexion) portions of the annulus fibrosus, or even traction fracture and displacement of the anteroinferior triangular portion of the ­vertebral body. A special type of indirect injury is the so-­called whiplash injury. The term “whiplash injury” describes a complex of clinical symptoms caused by minor or moderate acceleration of the head against the trunk. Patients complain of neck pain with muscle spasm, limited motion, and/or loss of the normal cervical lordosis. Additionally, headache, numbness, or weakness in one or both arms, tinnitus, and vertigo are reported. The symptoms are subjective and nonspecific, that is, unsupported by objective findings. Whiplash injury is mainly seen in victims of frontal, lateral, or rear-­end traffic collisions. Because pathological features of whiplash injury are clinically not detectable in most cases by diagnostic tests, its occurrence has been questioned and is the subject of many medical expertise. Experiments on human cadavers, healthy volunteers, and animals have shown a variety of soft tissue injuries ranging from muscle tears or sprains, ligament tears, especially of the anterior annulus fibrosus, and small or large fractures of the cervical vertebrae (Hoffman et al. 1992), and support the existence of this whiplash injury. Most German experts think that a load to the cervical spine of Δv = 10 km/h or less cannot cause neurological alterations or long-­term injury with chronic symptoms. No predictions are possible concerning the sequelae of a load of Δv = >15 km/h. These results have been accepted by the German legal system (Griess 2000).

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the spinal canal, the projectile and/or its fragments can move within it and thus cause paresthesias and motor paralysis, possibly also lead poisoning, even after prolonged survival times. High-­velocity projectiles create shock waves that are capable of inflicting injury on tissue without actually penetrating it. Gunshot-­wound-­induced pathological changes of the spinal cord feature partial or complete transection of the cord. Histologically, the mechanical insult displays liquefaction, necrosis of the lacerated tissue, and hemorrhages with tissue destruction.

20.4.3  Pediatric neuropathology Postnatal mechanical brain injury

Basic principles The prevalence of head injuries over trunk and limb injuries in infants is attributed to the disproportionately large size of the infant’s head compared to the rest of the body. It is widely accepted that the brain of younger children is more vulnerable to injury than the brains of older children or adults, especially to

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deceleration-­impact injuries caused by falls or acceleration-­burst injuries resulting from blows to the head. Moreover, the impact response of the infant head also depends on the age-­dependent mechanical properties of the skull and sutures. In infants, contusional injuries in the sense of cortical hemorrhages usually extend along lines of force through the brain rather than being confined to the point of impact on the cortical surface. Such injuries may manifest as small, hemorrhagic tears at the junction of the gray matter and white matter, less often as areas of hemorrhages deeper within the white matter. The common result of these injuries is cerebral edema. In children under two, the three major causes of head injuries are falls from heights, physical abuse, and traffic accidents. The poor balance and muscular coordination typical of this age group are the principal causes of MBI.

extremely uncommon in newborns, EDH in children is usually explained by contact forces created during accidental falls. It is mainly associated with parietal linear fracture, but may also be caused by injury of the meningeal arteries or venous sinuses during a complicated breech presentation or forceps delivery; a fracture is not always present. The volume of EDH in infants is sometimes sufficient to induce anemia. The excessive volume may be explained by the elasticity of the skull and disjunction of the suture. The clinical picture is determined by loss of consciousness, usually after a lucid interval. Neurological deficits (hemiplegia, hemiparesis, etc.), pyramidal signs, and seizures are also observed. Early computed tomography scanning enables correct diagnosis and is associated with an excellent prognosis.

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Subdural collections

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Subdural collection (SDC) is an umbrella term comprising various findings in the subdural space (Wittschieber et al. 2019). This collection may be important in diagnostic radiology to estimate the age of onset of a SDC and to differentiate, if there may be hints for repeated AHT of an infant (see Chapter 3). The most common finding in toddlers and infants is SDH, which almost always results from mechanical violence, especially in infants under 2 years of age. In the latter group, it may result from unobserved falls or repeated shaking (see Chapter 3). The biomechanics and pathophysiology of SDH in children are similar to the cause and process of SDH in adults. Chronic subdural hematoma (cSDH) is relatively rare in infants (Hwang and Kim 2000) and is nowadays considered to be a separate entity (Squier and Mack 2009). At autopsy, a characteristic petroleum-­like fluid is detectable within the subdural space and pia mater. Performing a microscopic evaluation, the hallmark of cSDH is a neomembrane, which in the absence of an acute hemorrhage often comprises the only lesion and is detectable around 10 days after the traumatic event (Walter et al. 2009). Neomembranes form a thin layer of richly vascularized tissue of dull appearance that adheres to the inner surface of the dura mater and contains small tortuous blood vessels forming spiders or irregular patches. Neomembranes frequently exhibit multicentric fresh hemorrhages plus evidence of old hemorrhages in the form of rust-­ colored pigmentation (macrophages containing hemosiderin). They also can completely envelop the site of cSDH, one membrane in contact with the dura, the other with the arachnoid. The appearance of such a neomembrane represents an important finding to distinguish cSDH and subdural hygroma.

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In children, mechanical loading to the skull can cause disruption of the bones and localized disjunction or disruption of the sutures. Skull lesions are relatively rare in head injuries of infants. The most common type of skull lesion is linear fracture, observed in 70–80% of all cases (Harwood-­Nash et al. 1971). Linear fracture is usually recognizable as a thin linear defect, with the same spacing along the entire course of the trajectory. In more than 80% of cases, such fractures are due to a fall, in 5%, due to a traffic accident. Parietal linear fractures and EDH can be caused by short falls, which do not induce initial unconsciousness. A fall of just 1 meter is sometimes sufficient to cause skull fracture in adults and children, even if it is onto a padded surface. Growing fractures are characterized by occurrence in infancy or early childhood. This type of fracture is accompanied by dural tearing, brain injury beneath the fracture, and continuing growth of the fracture to form a cranial defect. The growing fracture is explained by a mechanically induced brain edema that pushes and holds apart the fragments of the soft cortical bone. Depressed fractures comprise only 25% of all skull fractures (Choux 1986). This form of skull lesion is much less common in young infants (24 hours. Ischemic brain damage may result from space-­occupying SDH or it may develop as a consequence of transitory cardiac or respiratory failure or arrest. In over the half of the cases, a white matter atrophy or porencephaly is demonstrable in cases with survival times of more than 10 days. In many cases of AHT, focal AI is present in distinctive topographic regions such as the pons, corpus callosum, the craniocervical junction, or within the radices of the cervical spinal cord (Oehmichen et  al. 2008, Matschke et  al. 2015). AI may be explained in part by a vasogenic event, in part by traumatic impact. It is important to stress that signs for a general DAI characterized by a diffuse distribution of axonal lesions within the white matter of whole the brain are not regularly present, despite it has been observed by some authors.

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The cause of death in AHT cases has been discussed extensively in recent years, but till now is not completely understood. The space-­occupying effect of SDH does not adequately explain death in all cases of AHT, and the lack of mechanically induced DAI in the brain and signs of generalized cervical cord or nerve root injuries point in another direction. Recent experimental studies have suggested that repeated mild head injuries owing to the vulnerability of the immature brain or mechanical and neurotoxic effects of subdural blood to the immature brain may play a causal role. The cause of death may be complex and different mechanisms have been discussed (Geddes et  al. 2001a, Geddes et  al. 2001b, Oehmichen et  al. 2008). It has been suggested that hypoxic–ischemic brain injury may be one probable cause of death caused by a multifactorial process, which includes a slight space-­occupying SDH, possibly strengthened by accompanying vasospasms resulting from an associated SAH; a primary medullary stress due to whiplashing, which may entail a reflective cardiac or respiratory disturbance secondary to overextension of the cervical cord associated with secondary reduction or transitory arrest of the respiratory or cardiac function. Recently, a very plausible hypothesis was postulated to which focal hypoxic damage of the central pattern generator in brainstem centers may be responsible for death due to AHT (Matschke et al. 2015).

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Cause of death

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nonparietal. Rib fractures, fractures of long bones, and metaphyseal corner fractures are frequently detectable in AHT (Hung 2020).

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Figure 20.4.13  (a) and (b) Epidural hemorrhage of the cervical cord inflicted probably as a result of shaking.

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The observed encephalopathy in AHT is supposed to be the result of primary as well as secondary alterations of the brain. After primary brain injury, vasogenic edema initially occurs, because water leaks through permeable capillary walls with an increase of brain water content. Diagnosis of brain edema indicating the encephalopathy is based on an increased brain weight, a tightened dura mater, flattened gyri, and compressed ventricles. Due to a failure of cell membrane pumps, a cytotoxic edema results, because extracellular water is shifted into the intracellular space (Orman et  al. 2020). This process leads to the microscopically observed hypoxic–ischemic brain injury with necrosis of nerve cells and an increase of microglia, which partly surround the damaged nerve cells like a rosette. As suggested in the following text, the encephalopathy is likely due to hypoxic–ischemic injury when focal damage in vital brainstem centers occurs (Matschke et  al. 2015). Usually, no cortical hemorrhages are detectable (Figure 20.4.12a).

Microscopy

In acute cases, the histological examination of the dura mater may reveal extravasal red blood cells without any cell reaction. In dependence of the survival time, the detection of PMNs and macrophages is associated with a delayed death more than 2–3 hours for leukocytes, 12–20 hours for macrophages, and more than 72–100 hours for siderin-­containing macrophages (Oehmichen containing et  al. 2009a). The demonstration of hemosiderin-­ macrophages in dura, subarachnoid space, or brain within an estimated survival time after acute traumatic event of less than 72

Sudden infant death SID is defined as the sudden death of an infant or young child, which is unexpected by history and in which a thorough postmortem examination fails to demonstrate an adequate cause of death. In accordance with the prevailing definition, SID victims exhibit no specific morphological findings that can account for the sudden, unexpected death, probably because SID is not an

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CHAPTER 20   Mechanical Trauma and Classification of Wounds

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misplaced or disorganized aggregations of mature or immature neuroepithelial cells are found in cerebellar tissue (Matschke et al. 2020). The observation that granule cell migration and differentiation are reduced may suggest a wider defect. Third, a focal granule bilamination of the dentate nucleus of the hippocampus has been described of about 40% of SID cases (Kinney et al. 2015). Dental bilamination is a maker of temporal lobe epilepsy and leads to a vulnerability to seizures. Therefore, a routine bilateral microscopic examination of the hippocampus is recommended in SID cases. Another important microscopic finding is the increase of the number of apoptotic cells in the hippocampus and the brainstem, reflecting significant neuronal damage and a functional loss of key brain regions (Waters et al. 1999). It is general accepted that SID may be associated with functional alterations leading to brain hypoxia or ischemia, sometimes as a recurrent process. It has been shown that SID victims reveal a reduced associated protein-­ 2-­ reactive neurons number of microtubule-­ which support the thesis that the cause of death may be due to respiratory or cardiac dysfunction, especially in the absence of any other organ pathology (Oehmichen et al. 2009c). Till now, it is not clear if the described neuropathological findings are the primary cause of SID or secondary to unknown underlying mechanisms. The described neuropathological findings may give a hint for diagnosis and probable causes of SID, but none of it is so far of diagnostic value alone.

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entity, but only a description of what has happened. SID is a diagnosis of exclusion, and should be discussed when police records and the medical history of the infant have been analyzed, a complete autopsy including microscopic examination and toxicological and microbiological analyses has been performed, and no obvious cause of death could be determined. Helpful for the diagnosis is the triple-­risk hypothesis, which includes a vulnerable infant in a sensitive developmental period, whose death is precipitated by external circumstances. Recently, there have been striking advances in the genetic analysis (e.g., channelopathies) and of external stressors (e.g., prone position) to elucidate the cause of death (Jeffery 2018), but these issues are not presented here. In this chapter, important and recent neuropathological alterations in SID cases are described. There are some distinct neuropathological findings which are detectable by routine staining methods or immunohistochemistry. A consistent observation is brainstem gliosis, which is easily detectable applying the GFAP antibody on formalin-­ fixed, paraffin-­embedded routine specimens (Sawaguchi et  al. 2003). This gliosis may be caused by a primary developmental abnormality or secondary by induced repetitive hypoxic/ischemic events due to a dysregulation of homeostatic reflexes (Paine et al. 2014). As a hint for metabolic disturbances with a failure to respond to external stressors appropriately, a reduced serotonin receptor binding in the arcuate nucleus and other nuclei of the brainstem has been described (Ozawa and Takashima 2002; Duncan et al. 2010). In line with these findings is the observation in animal experiments that elimination of serotoninergic 5-­hydroxytryptamine neurons leads to a delayed arousal when challenged with repeated episodes of hypoxia during sleep (Barrett et al. 2016). Moreover, the retardation of neuronal development in some nuclei of the brainstem of SID victims is supported by a recent proteome analysis, where signs of a delayed neurological maturation have been observed (Hunt et al. 2016). A high dendritic spine density, a decrease of tyrosine-­hydroxylase-­ positive catecholaminergic neurons, and other neurochemical abnormalities in distinct areas of the brainstem have all been interpreted as signs of neuronal retardation and hypofunction or immaturity of cardiorespiratory regulation (O’Kusky and Norman 1994; Ozawa and Takashima 2002; Machaalani and Waters 2014). Moreover, hypoplasia of the arcuate nucleus, which cells play a role in central respiratory chemoreception, has been discussed to contribute to sudden death (Kinney and Filiano 1995). Therefore, the fatal process underlying SID may involve disturbance of respiratory regulation of nuclei of the brainstem that results in clinically observable apnea attacks (Schulte et al. 1982). It has been observed that heterotopia of cells in different regions of the brain may play a role in the lethal process. At first, heterotopic leptomeningeal neurons are increased over the brainstem of SID victims in comparison to controls (Rickert et  al. 2009). Furthermore, there is evidence that the external granule cell layer of the cerebellum is deeper, denser, and persists longer in infants dying from SID (Cruz-­Sánchez et al. 1997). Additionally, cerebellar heterotopia in SID cases has been observed, where

20.4.4 Ischemia, hypoxia, and asphyxia Basic principles

The important target organ of ischemia, hypoxia, and asphyxia is the brain which utilizes about 20% of the totally consumed oxygen of the body. Neurons are the target cells of the brain damaged by a loss of oxygen. Microtubuli-­associated proteins play an important role in growth, differentiation, and plasticity of neurons. These proteins are very sensitive to ischemic events. Therefore, MAP2 can serve as a useful marker to detect ischemia in the brain.

Ischemia Ischemia is characterized by a transient or permanent stop of the general or regional cerebral blood flow. Primary ischemic brain damage as a result of brain perfusion interruption is followed by a change in the blood gas values and pH within the brain associated with acidosis and increase of lactate. As a result of these processes, an energy depletion occurs within a few seconds to minutes resulting in a disturbance of the K+/Na+ pump with a shift of the K+ ions into extracellular space as well as of Na+ ions and water from extracellular space into the cytoplasm, leading to an early cytotoxic edema. This leads to a decline of cerebral perfusion and a secondary disturbance of the blood–brain barrier. The sequela will be an overlapping vasogenic edema within 10–20 minutes, which in most cases masks the primary (cytotoxic) edema.

N O SE Figure 20.4.14  Microscopic findings of neurons in cases of brain ischemia. Arrows indicate the shrunken eosinophilic neurons.

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The edema itself aggravates the anoxic damage, thus giving rise to a vicious cycle. Simultaneous leukocyte–endothelial cell interaction within 10–20 minutes is accompanied by a release of toxic substances, especially the cytokines. Ischemic injuries are expressed in different brain areas, partly depending on the type and extent of arterial supply. In cases of global ischemia, damage usually begins in the triple watershed zone, at the intersection of the territories of the cerebral arteries. If ischemic damage is suspected, the fissure between the first and second gyri of the frontal lobe, the CA1 region of the hippocampus, the Purkinje cell layer at the cerebellum, as well as corpus striatum or, specifically, the globus pallidus should be examined carefully. The distribution of ischemic damage can vary with the type of damage. Ischemia can be divided into focal and global, and into transient and permanent ischemia. Permanent global ischemia constitutes brain death, and is described in Chapter 4. Transient global ischemia constitutes the typical clinical situation of cardiac arrest, where duration of 5 minutes is known to be critical in the development of irreversible necrotizing cerebral damage. Transient and focal ischemia constitute a continuum: if the duration of transient focal ischemia is long enough, irreversible necrosis takes place and it becomes irrelevant whether reperfusion takes place at later times, since an infarction has already completely developed. The duration of the development of neuronal necrosis in transient focal ischemia depends on the depth and duration of ischemia, as well as on factors as temperature, glucose level, and epileptiform activity. It is clear, however, that transient focal ischemia has a much longer tolerable duration (at least half an hour, depending on temperature, glucose levels, etc.) before necrosis develops than 5  minutes of transient global ischemia. The implication is that focal ischemic brain damage has a deleterious effect on other parts of the brain. As progressively more of the brain is rendered ischemic, the tolerable duration decreases. term survived The most important alterations after long-­ ischemia with consecutive reperfusion are necrosis of neurons, patches of demyelination, brain-­parenchymal loss, and a proliferation of microglia and astrocytes. For forensic purposes, it is crucial to know the duration of the interval before ischemic damage can first be detected by morphological criteria. The literature reports a wide variation, ranging from 7.5  minutes (Cizkova et  al. 2000) to 3 hours (Spielmeyer 1922) up to a maximum of 15–25 hours (Scholz 1951). Neurons are the best targets to detect ischemia because these cells are dependent on a sufficient oxygen supply. The common cytologic marker of an ischemic injured neuron in case of reperfusion is the acidophilic/eosinophilic alteration of the neuronal cytoplasm and the shrunken neuronal nucleus, which can be demonstrated using H&E stain (Figure 20.4.14). These neurons should not be confused with the prelethal dark neurons. The described alterations of nerve cells in cases of cortical ischemia are best visualized in the middle cortical laminae. Applying Nissl stain, a local or generalized permanent ischemia without reperfusion results in a so-­called “pale nerve cell injury” (Kalimo et al. 1982, Lipton 1999), marked by neuronal and astrocytic edema, dilatation of nuclear

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chromatin, and slight swelling of mitochondria and endoplasmic reticulum leading to a picture of a so-­called vacuolar degeneration. These neurons are definitively known to be necrotic for two reasons: first, they disappear from tissue sections in experiments studying differing survival times, and second, they show cell membrane breaks and mitochondrial flocculent densities on electron microscopy. Applying morphological methods, ischemic cytological alterations may be seen earliest after 10–30 minutes (Kühn et al. 2005; Inamura et al. 1987), often within 3–4 hours (Ikonomidou et  al. 1989) of survival time and in general only after several hours. As a consequence of reperfusion likewise in cases of permanent focal ischemia, homogenizing cell changes, so-­ called ghost neurons, appear, marked by an irregular chromatin clumping, a shrunken cytoplasm which may be fragmented with small vesicles and dense bodies. This type of cell change will arise about 24 hours after an ischemic event (Brown 1977) and 6–12 hours after permanent ischemia (Garcia et al. 1995). Focal cerebral ischemia finally is associated with a total loss of neurons in the region supplied by the occluded arteries/artery, while transient global ischemia yields progressive damage to selectively vulnerable populations of neurons (Smith et al. 1984). Another phenomenal characteristic associated with permanent focal ischemia is the process of apoptosis. This “programed neuronal death” will appear in the penumbra of focal lesions after one to several hours. Morphologically, the cells are characterized by uniformly condensed chromatin, apoptotic bodies, and darkened, pyknotic cytoplasm. In recent years, an additional set of antibodies gives rise to the possibility of detecting the functional

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Meanwhile, over a decade ago, the loss of MAP2 expression by neurons of the frontal cortex or hippocampus in acute ischemic/ asphyctic events in cases of strangulation, drowning, and carbon monoxide poisoning was demonstrated (Kühn et  al. 2005). In these cases, MAP2-­negative neurons were always detectable in a higher percentage compared to the control group. It is important to stress that even in the control cases not all nerve cells were labeled by the antibody, because only 10% of nerve cells were stained in the hippocampus, and up to 70% were marked in the frontal cortex. The most striking differences between control group and cases of ischemia were detected in the CA2, CA3, and CA4 areas (Kühn et al. 2005). In the same way, MAP2 is a very useful marker in cases of global permanent ischemia to confirm brain death on the morphological level (Maith 2012). To sum up, MAP2 seems to be a useful marker of very early neuronal damage in cases of short and longer intervals of survived ischemia.

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Hypoxia is defined by a reduction of the partial pressure of oxygen below the normal level. In contrast to ischemia, the cerebral blood flow is maintained or even increased. This difference has a few very important sequelae. Because of the continuous blood flow, waste products as lactate, acidosis, etc., do not accumulate. In case of a transient hypoxic exposition, the breakdown of neuronal function will be reversible. In hypoxia, the intracerebral vascular system always reacts functionally to a drop in pO2 and pH and an increase in both pCO2 and extracellular K+. Hypoxia is to be divided into decreased supply of oxygen to the body, decreased transport of oxygen (anemia), and decreased utilization of oxygen (histotoxic hypoxia). Recognition of the fact that hypoxia alone does not cause necrotizing brain damage is especially important in view of the fact that acute onset hypoxia may cause prolonged coma. In contrast to ischemia, a pure hypoxic coma of this kind is caused by synaptic alterations, but not by neuronal cell body necrosis. The distinction between hypoxia and ischemia is thus of paramount importance in determining prognosis of coma, though both types of oxygen deficits are merging into one another. If the heart has not stopped, and blood pressure has been well maintained, or if the heart has stopped for only a brief period and ventilation has been adequate, then widespread pan-­cortical necrosis and brain death are unlikely to have occurred. If such patients die, autopsy will reveal no significant necrosis in the brain (Auer and Sutherland 2002).

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loss of nerve cells under microscopic examination, among others, the loss of immunostaining of MAPs (see Yanagihara et al. 1990). The morphologic features distinguishing selective neuronal necrosis from pannecrosis apply in every brain region. Of all the brain regions, the hippocampus displays a most heuristic selective and specific vulnerability. Therefore, careful and standardized sampling of the hippocampus is of utmost importance. On the collected samples, the examiner should see the typical snail-­shell-­ like structure of the hippocampus including the dentate nucleus. The classic section of the hippocampus is seen when a coronal section is taken from the temporal lobe at the level of the lateral geniculate body. In successful preparations, the CA4 to CA1 regions are easily detectable on the microscopic slides. The structure of the C-­shape of the dentate nucleus should be easy to identify. On the opposite of the tip of the “C,” the observer will find the CA1 sector, followed by the very narrow CA2 cell band and CA3 area. In the hilus of the dentate nucleus, the CA4 area is presented. The CA1 sector is most sensitive to ischemia. In rare cases, acute ischemic CA1 necrosis can occur after only 2–4 minutes of global ischemia. After 2–4  minutes of global cerebral ischemia, some CA1 pyramidal cells undergo neuronal necrosis (Smith et al. 1984). The phenomenon of delayed neuronal death in humans is detectable within 2–4  days in the hippocampus (Petito et al. 1987, Horn and Schlote 1992). In the CA4 sector, the area of the dentate hilus, cells die after only small global ischemic insults, as they do in CA1 (Benveniste and Diemer 1988). The pathologist should examine the dentate hilus if there is suspicion of short periods of global cerebral ischemia, especially in children, where microglial activation may be the only detectable alteration (Del Bigio and Becker 1994). After CA1 cells are affected, progressively longer or more severe ischemic insults will cause recruitment of CA3 cells into the necrotic process. Finally, the dentate gyrus, very resistant to cerebral ischemia, is recruited in the most severe insults. In other brain regions 10–12 hours after an ischemic event, the first gross alterations are seen characterized by a loss of demarcation between white and gray matter, a gelatinous tissue appearance, and a reduction of tissue consistency (encephalomalacia). The cerebral cortex displays a reddish discoloration or a red band at the border between white and gray matter. After a survival time of some days, the pallidum will be softened and cystically transformed. Survival times of more than a week lead to a cortical atrophy of the cerebrum and cerebellum. The combination of ischemia and arterial hypotension may be associated with an involvement of the brainstem and the medulla. In cases of global brain ischemia, necrosis of the globus pallidus and pars reticulata of the substantia nigra can occur, similar to that seen in carbon monoxide poisoning. In some of these cases, necrosis of the thalamus may display. The brainstem may exhibit symmetrical necrosis of brainstem nuclei. The cerebellum is regularly affected in global cerebral ischemia, so are the Purkinje cell as one of the most vulnerable type of nerve cells. Their cell bodies display the typical pink appearance, the cell boundaries blur, and due to necrosis their numbers are reduced.

Asphyxia Conditions leading to asphyxia are defined by cut off of atmospheric oxygen (as in drowning or plastic bag on the head), while cerebral perfusion continues. Asphyxia is characterized by retention of CO2 and is associated with acidosis, hypotension, and occasionally hypoglycemia. From the forensic point of view, we have to distinguish four types of ischemia/asphyxia: suffocation, strangulation, smothering, and chemical asphyxia.

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Strangulation

Death by suffocation is caused by a reduction of the oxygen concentration in the respired atmosphere. The different types of suffocation are listed in Table 20.4.5. Environmental suffocation describes a lack of atmospheric oxygen concentration due to high-­altitude sickness as well as in situations of physical displacement of oxygen, for example, by gases such as carbon dioxide, nitrogen, or other. A mechanical occlusion of the external respiratory orifices may be caused by sand or by a plastic bag over the head, which may cling to the face by static electricity and occlude the external airways (accidental, suicidal, homicidal, etc.). These types of asphyxia will not be associated with any specific neuropathological findings at autopsy. Only in cases of drowning, the number of MAP2-­expressing neurons in the hippocampal region was reduced (Kühn et  al. 2005). The nonspecific finding will be in most cases a brain swelling or a brain edema which, however, will not be present in all victims of death by suffocation. In cases of delayed death after suffocation, the neuropathological findings may range from being totally negative to expressing the symptoms of brain death, each dependent on the duration of interruption of the oxygen supply and the time interval between the insult and resuscitation or blood reflow. Partly, the neuropathological findings resemble the alterations observed in cases of delayed death after strangulation.

Whenever pressure is applied to the neck, there are five types of mechanisms which may account for death in various combinations: venous obstruction, arterial occlusion, reflex mechanisms, airway collapse, and mechanical neck injury. A venous outflow obstruction of the head is most easily produced due to the thin walls and easy collapse of veins. This accounts for the commonly seen conjunctival and facial skin petechial hemorrhages in victims of manual strangulation, hanging, etc. The increase in venous pressure in cases of strangulation causes the almost ubiquitous lesions at death: congestion of the brain vessels and a minor subarachnoid hemorrhage (“smear”) as well as periventricular hemorrhages. If more force is used in constricting the neck, carotid artery compression can result. This is known to lead to loss of consciousness in roughly 7–10 seconds. In attempting to overcome a victim by choking or mugging, it is this period of carotid insufficiency that is needed by the perpetrator to produce unconsciousness and finally subdue the victim by obviating the possibility of further struggle and resistance. The neuropathology of acute death by hanging is nonspecific. In cases of ligature strangulation, there is usually no complete occlusion of the arterial vessels at the beginning of mechanical neck compression, the face will be congested, petechiae and scleral hemorrhages are common, as well as swelling of the tissues. The brain examination will give evidence of congestion and brain

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Table 20.4.5  Different forensic types of hypoxia, ischemia, and asphyxia.

Ischemia hypoxia

– Deprivation of oxygen

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Hypoxia

– Lack of oxygen in breathable environment

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Suffocation Environmental suffocation

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Spontaneous brain perfusion or reperfusion

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– Blockage of the external air passage

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– Blockage of the upper airways by foreign body (bolus, aspiration) – Manual strangulation: external compression of trachea

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Strangulation

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Ligature strangulation

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Manual strangulation

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Mugging

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Hanging

– Constricting band is tightened by the weight of the victim’s body – Ligature strangulation (Spanish method of judicial execution)

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Garotting

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Intermittent ischemia

Ischemia

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diac arrest for a time of 5–15  minutes will finally lead to an irreversible interruption of the cerebral blood flow of the cerebrum and cerebellum with the consequence of a respiratory arrest, while the circulation of the body may be stable. Anoxia without brain reperfusion in cases that survived, that is, cases on respirator, is more difficult to detect by morphological criteria. Prolonged intracranial circulatory arrest and increased intracranial pressure aggravated by vasogenic edema finally lead to the clinical and morphological criteria of brain death. The neuropathological alterations of a nonperfused brain take about 12 hours to develop, although they become more obvious in 24 hours, an important interval if the neuropathologist is called upon to confirm brain death. Within the first 24 hours, it can only be diagnosed by a massive increase in brain volume, but the increase is definitely a function of time on respirator. During this period, a demarcation of the optical nerve, the pituitary gland, and the cervical spinal cord can be discerned. At autopsy, the brain weight is highly increased in many cases. The brain structure seems macroscopically roughly preserved in nearly all cases, but appears occasionally extremely swollen, congested, and characterized by a flattening of the gyri and a dusky hue. The leptomeninges often show a quantitatively different extent of congestion and fibrin precipitation as an indication of different regional no-­reflow phenomena. A transtentorial herniation with uncal necrosis is a common finding. Softened herniated cerebellar tonsils (Figure  20.4.15) are commonly necrotic and fragmented as well with pieces of necrotic tonsillar tissue lodged along the spinal cord anywhere from the cervical segment to the cauda equina. The brainstem is often torn and is characterized by congestion and hemorrhages and sometimes fragmented. In addition, hemorrhagic softening of the upper few segments of the cervical spinal cord as well as of the hypophysis and the nervus opticus occurs as an indication of demarcation of the necrotic brain.

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Smothering may be done by a gag obstructing the nose and mouth or by a pillow which is placed over the face and pushed down, or—­in children—­by a hand. Abrasion injuries of the face will occur if the victim puts up a resistance. No other morphological features are seen at autopsy. The same negative pathological and neuropathological findings will result from choking, that is, obstruction of the respiratory passage. Most often choking is accidental in manner: unintentional inhalation of food or other foreign bodies, resulting in obstruction of the respiratory passage (death by bolus). Typical drowning is characterized by an immersion-­induced blockage of the external air passage as well as of the upper airways by water. The common sequelae will be retention of carbon dioxide and convulsions. Mechanical asphyxia is characterized by mechanical fixation of the chest (thorax compression), as well as by curare-­like drugs or-­brain induced primary respiratory arrest, that is, by acute morphine poisoning. In acute death, neuropathological findings are marked by congestion without any specific findings.

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edema. Specific findings are absolutely uncommon. The dynamic development of visible neuropathology in cases of strangulation is dependent on time, in the presence of recirculation of blood. Pale nerve cell injury may be observed in complete permanent ischemia without recirculation and is detectable 30–60 minutes after strangulation. Dark nerve cell injury may be detected in cases of incomplete ischemia or in cases of severe complete ischemia followed by reflow. Delayed death after strangulation is a rare event. There are no common findings, as each case is characterized by an individual history regarding age, biomechanics of strangulation, duration of strangulation, survival time, etc. In cases of survived hanging, cerebral infarction in the brain has been described, and is due to carotid artery compression. Delayed death after strangulation may reveal “brain death” after resuscitation, or regionally an elective parenchymal necrosis and hemorrhages in the lentiform nuclei, caudate, and internal capsule.

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Permanent global ischemia constitutes brain death. Brain death is clinically defined as the irreversible loss of function of the brain and can be diagnosed by three cardinal clinical findings such as coma, absence of brainstem reflexes, and apnea. The diagnosis of permanent global ischemia is a very important task in forensic neuropathology, especially when there are other possible causes of death. Permanent global ischemia is a direct result of a stop of the cerebral perfusion. This, in turn, usually also arises from either an increase in intracranial pressure or a decrease in the mean arterial blood pressure to bring the cerebral perfusion pressure to less than 45 mmHg for a protracted period. After some time has elapsed, the brain can never be reperfused due to thrombosis-­and edema-­ induced capillary occlusion, and “brain death” ensues. Under normothermic conditions, the sequelae of persistent brain ischemia by suffocation or strangulation as well as by car-

Figure 20.4.15  Cerebellar tonsillar necrosis in cases of brain death.

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Border-­zone reactions are characterized by leukocytic infiltrations accompanied by hemorrhages in the upper segments of the cervical spinal cord, in the optic nerve, and in the hypophysis. Reactivity of the leptomeninges of the spinal cord displays an infiltration of leukocytes within the subarachnoid space of the spinal cord. Sometime later, necrotic tissue components, especially cerebellar tissue, will appear within the subarachnoid space, partly mixed with leukocytes. A survey of the morphological alterations is given in Table 20.4.6. Clinical experience has shown that reactivation of the neuronal functions is only possible during the first 10–15 minutes of total cerebral ischemia under normothermic conditions, because nerve cells are extremely sensitive to any reduction in oxygen supply. Complete interruption of the blood and/or oxygen supply leads to time-­dependent changes shown in Table  20.4.7 (Grote 1980). Even though animal studies could demonstrate the rever­ sibility of ischemic nerve cell damage, in normothermic humans irreversible brain death regularly occurs within this interval of continuous anoxia. Some experimental results, however, will give hope that the ischemic process will not be irreversible. We learned by experiments of Hossmann and Kleihues (1973, Bodsch et al. 1986) that under certain circumstances of experimental post­ ischemic treatment, reperfusion will succeed—­and a recovery of neuronal function on the cellular level could be established—­still after a period of 20–60 minutes of total ischemia of the brain.

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Gross sections of the brain display a typical lead-­colored, grayish cortex, a compressed ventricular system, and sometimes focal hemorrhages in white and/or gray matter. On frontal sections of the brain, the ventricular system is extremely compressed and the borders of gray and white matters are blurred. During longer survival times on the respirator, the width of the cortical ribbon may be reduced, especially in the parietal lobes. Furthermore, an intracortical band of necrosis may display, which appears as a slit at the border between white and gray matter. Microscopy of the cortex and the hippocampus reveals a diffusely congested and edematous cortex with cortical neurons exhibiting various degrees of acute ischemic changes. Shrinkage of the nerve cell bodies is partly detectable with dark staining nuclei or a pink-­staining cytoplasm in hematoxylin and eosin preparations. Reactive cells, that is, leukocytes, macrophages, or reactive astrocytes, are often detected in the brain tissue. As a general rule, ischemic neuronal alterations are only observed after reperfusion lasting at least 2 hours. Primary ischemic infarction or hemorrhagic infarction results in primary or secondary occlusion of cerebral arteries. An important target to establish the diagnosis of permanent global ischemia is the hippocampus with its CA1, CA2, CA3, and CA4 segments. Applying immunohistochemistry, neurons of the hippocampus may exhibit a detectable depletion of MAP2 expression in permanent global ischemia, especially in the CA1 segment, where often no MAP2-­positive nerve cells are found. Therefore, this marker may aid to diagnose global ischemia. The loss of MAP2 immunoreactivity is not an irreversible process, because a decrease in MAP2 labeling occurs prior to notable neuronal cell loss and a partial restoration of MAP2 immunoreactivity was observed 24 hours post injury (Huh et  al. 2003). These data show that MAP2 is an early and sensitive marker for neuronal damage, but acute MAP2 loss may not necessarily presage neuronal death. In single cases, MAP2-­ expressing astrocytes can be demonstrated, especially in the CA4 and CA3 regions. In these cases, nearly all neurons are MAP2-­ negative. Using MAP2 as a marker alone, it is not possible to differentiate brain death cases from brains without signs of brain hypoxia/ischemia. Therefore, MAP2 is an important, but not a reliable morphological marker of brain death diagnosis. Nevertheless, classical morphological criteria of brain death in combination with MAP2 demonstration are able to confirm the clinical diagnosis in all cases. The striking features of the cerebellum are swelling and congestion; autolysis of the granule cell layer of the cerebellar cortex is often observed. The brainstem reveals almost always edema, secondary hemorrhages, infarction, necrosis, and/or neuronal loss. The mesencephalon is distorted by local edema or external compression by herniating tissue of the medial temporal structures or the anterior cerebellar lobes. The whole brain reveals congested vessels partly associated with intravascular precipitation of fibrin and subsequent loss of cellular staining in the absence of reactive changes. The brain death syndrome during the early phase will microscopically be characterized by border-­zone reactions (demarcation zone), reactivity, and alterations of the leptomeninges.

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Table 20.4.6  Neuropathological findings in brain death. Gross examination Increased brain weight (>12%) Extensive brain swelling Herniation (hippocampal, tonsillar) Brown discoloration Pink centrum semiovale due to poor fixation Poorly defined demarcation of gray and white matters Microscopic examination Structural effacement/washed out tissue picture due to poor staining Eosinophilia without tissue reaction Regressive neuronal changes Congestion Border-­zone alterations (hemorrhages, cell reactions) Anterior pituitary lobe C1/C3 cervical segments of the cord Displacement of cerebellar tissue of the subarachnoid space of the spinal column Infarction of the optic tract

Table 20.4.7  Functional impairment of the brain in dependence of the duration of the disruption of oxygen supply. 5s 8–12 s 20–30 s 8–10 min 4 min

Marked functional impairment of the CNS Total loss of function of the CNS (loss of consciousness) Formation of a flat EEG Resuscitation time of the brain Resuscitation time of the entire organism with asystolia

Source: According to Grote (1980).

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Cizkova, D., Vanicky, I., Ishikawa, T. and Marsala, M. (2000). Time course of brain neuronal degeneration and heat shock protein (72) expression following neck tourniquet-­induced cerebral ischemia in the rat. Cellular and Molecular Neurobiology 20: 367–381. Clarke, P.G.H. (1998). Apoptosis versus necrosis. In: E. Koliatsos and R.R. Ratou (eds.), From Cell Death and Diseases of the Nervous System, pp. 3–25. Totowa, NJ: Human Press Inc. Corsellis, J.A.N., Bruton, C.J. and Freeman-­Browne, D. (1973). The aftermath of boxing. Psychological Medicine 3: 270–303. Cruz-­Sánchez, F.F., Lucena, J., Ascaso, C. et al. (1997). Cerebellar cortex delayed maturation in sudden infant death syndrome. Journal of Neuropathology & Experimental Neurology 56: 340–346. Del Bigio, M.R. and Becker, L.E. (1994). Microglial aggregation in the dentate gyrus: A marker of mild hypoxic-­ischaemic brain insult in human infants. Neuropathology and Applied Neurobiology 20: 144–151. Di Maio, D.J. and Di Maio, V.J.M. (2001). Forensic Pathology. Boca Raton, London, New York, Washington: CRC Press. Di Maio, V.J.M. (1981). Penetration and perforation of skin by bullets and missiles: a review of the literature. American Journal of Forensic Medicine and Pathology 2: 107–110. Duhaime, A.C., Christian, C.W., Rorke, L.B. and Zimmerman, R.A. (1998). Nonaccidental head injury in infants, the” shaken baby syndrome”. New England Journal of Medicine 338: 1822–1829. Duhaime, A.C., Gennarelli, T.A., Thiebault, L.E. et al. (1987). The shaken baby syndrome: A clinical, pathological, and biomechanical study. Journal of Neurosurgery 66: 409–415. Duncan, J.R., Paterson, D.S., Hoffman, J.M. et al. (2010). Brainstem serotonergic deficiency in sudden infant death syndrome. JAMA 303: 430–437. Fanconi, M. and Lips, U. (2010). Shaken baby syndrome in Switzerland: results of a prospective follow-­up study, 2002-­2007. European Journal of Pediatrics 169: 1023–1028. Friede, R.L. (1989). Developmental Neuropathology. Berlin, Heidelberg, New York: Springer. Garcia, J.H., Liu, K.-­F. and Ho, K.-­L. (1995). Neuronal necrosis after middle cerebral artery occlusion in Wistar rats progresses at different time intervals in caudoputamen and the cortex. Stroke 26: 636–643. Geddes, J.F., Hackshaw, A.K., Vowles, G.H. et al. (2001a). Neuropathology of inflicted head injury in children: I Pattern of brain damage. Brain 124: 1290–1298. Geddes, J.F., Vowles, G.H., Nickols, C.D. et al. (2001b). Neuropathology of inflicted head injury in children: II Microscopic brain injury in infants. Brain 124: 1299–1306. Gennarelli, T.A. and Meaney, D.F. (1996). Mechanisms of primary head injury. In: R.H. Wilkins and S.S. Rengachary (eds.), Neurosurgery, Vol. 2, pp. 2611–2621. McGraw-­Hill, New York. Gennarelli, T.A. and Thibault, L.E. (1982). Biomechanics of acute subdural hematoma. Journal of Trauma 22: 680–686. Gosch, H.H., Gooding, E. and Schneider, R.C. (1972). An experimental study of cervical spine and cord injuries. Journal of Trauma 12: 570–576. Graeber, M.B., von Eitzen, U., Grasbon-­Frodl, E. et al. (1997). Microglia: a „sensor“ of pathology in the human CNS. In: M. Oehmichen and H.G. König (eds.), Neurotraumatology  – Biomechanic Aspects,

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Jeffery, H.E. (2018). Future directions in sudden unexpected death in infancy research. In: J.R. Duncan and R.W. Byard (eds.), SIDS Sudden Infant and Early Childhood Death: The Past, the Present and the Future. Adelaide (AU): University of Adelaide Press; Chapter 14. Kalimo, H., Olsson, Y., Paljärvi, L. and Söderfeldt, B. (1982). Structural changes in brain tissue under hypoxic ischemic conditions. Journal of Cerebral Blood Flow & Metabolism 2: 19–22. Kinney, H.C., Cryan, J.B., Haynes, R.L. et al. (2015). Dentate gyrus abnormalities in sudden unexplained death in infants: Morphological marker of underlying brain vulnerability. Acta Neuropathologica 129: 65–80. Kinney, H.C. and Filiano, J.J. (1995). The Ventral Medullary Surface in SIDS. A Review, pp 164–168. Oslo Copenhagen Stockholm Boston: Scand Univ Press. Klages, U. (1970). Spontaneous versus traumatic lethal subarachnoid hemorrhage. Zeitschrift fur Rechtsmedizin 67: 67–86. Klatzo, I. (1967). Neuropathological aspects of brain edema. Journal of Neuropathology & Experimental Neurology 26: 1–14. Kleinbaum, D.G. (1998). Logistic Regression. Berlin, Heidelberg, New York: Springer. Kleiven, S. and W.N. Hardy (2002). Correlation of an FE model of the human head with local brain motion – consequences for injury prediction. Stapp Car Crash Journal 46: 123–144. Krauland, W. (1961). Über die Quellen des akuten und chronischen subduralen Hämatoms. In: W. Bargmann and W. Doerr (Hrsg), Zwanglose Abhandlung aus dem Gebiet der normalen und pathologischen Anatomie. Stuttgart: Thieme. Krauland, W. (1981).The traumatic subarachnoidal hemorrhage. Zeitschrift fur Rechtsmedizin 87: 1–17. Krauland, W. (1982). Verletzungen der intrakraniellen Schlagadern. Berlin, Heidelberg, New York: Springer. Kühn, J., Meissner, C. and Oehmichen, M. (2005). Microtubule-­ associated protein 2 (MAP2) – a promising approach to diagnosis of forensic types of asphyxiation. Acta Neuropathologica (Berl) 110: 579–586. Leestma, J.E. (2008). Forensic Neuropahology. Boca Raton, London, New York: CRC Press. LeRoux, P.D., Haglund, M.M., Newell, D.W. et al. (1992). Intraventricular hemorrhage in blunt head trauma: an analysis of 43 cases. Journal of Neurosurgery 31: 678–685. Levin, H.S., Meyers, C.A., Grossmann, R.G. and Sarwar, M. (1981). Ventricular enlargement after closed head injury. Archives of Neurology 38: 623–629. Lindenberg, R. (1982). Tissue reactions in the gray matter of the central nervous system. In: W. Haymaker and R.D. Adams (eds.), Histology and Histopathology of the Nervous System, Vol. 1, pp. 973–1275. Springfield/IL: CC Thomas. Lipton, P. (1999). Ischemic cell death in brain neurons. Physiological Reviews 79: 1431–1567. Machaalani, R. and Waters, K.A. (2014). Neurochemical abnormalities in the brainstem of the sudden infant death syndrome (SIDS). Paediatric Respiratory Reviews 15: 293–300. Mack, J., Squier, W. and Eastman, J.T. (2009). Anatomy and development of the meninges: implications for subdural collections and CSF circulation. Pediatric Radiology 39: 200–210.

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Cytologic and Molecular Mechanisms, pp. 239–252. Lübeck: Schmidt-­Römhild. Graham, D.I., Adams, J.H., Nicoll, J.A.R. et al. (1995). The nature, distribution and causes of traumatic brain injury. Brain Pathology 5: 397–406. Syndrom bei kollisionsbedingter geringer Griess, A. (2000). HWS-­ Geschwindigkeitsänderung. Neue Juristische Wochenschrift 53: 877–878. Grote, J. (1980). Gewebsatmung. In: R.F. Schmidt and G. Thews (eds.), Physiologie des Menschen, pp. 558–571. Berlin, Heidelberg, New York: Springer. Hardy, W.N., Foster, C.D., Mason, M.J. et al. (2001). Investigation of head injury mechanisms using neutral density technology and high-­speed biplanar X-­ray. Stapp Car Crash Journal 45: 337–368. Harwood-­Nash, D.C., Hendrick, E.B. and Hudson, A.R. (1971). The significance of skull fracture in children: A study of 1187 patients. Radiology 101: 151–154. Herrmann, B., Banaschak, S., Dettmeyer, R. and Thyen, U. (2008). Kindesmisshandlung. Berlin, Heidelberg, Springer Medizin Verlag. Hoffman, J.R., Schriger, D.L., Mower, W. et al. (1992). Low-­risk criteria for cervical–spine radiography in blunt trauma: a prospective study. Annals of Emergency Medicine 21: 1454–1460. Horn, M. and Schlote, W. (1992). Delayed neuronal death and delayed neuronal recovery in the human brain following global ischemia. Acta Neuropathologica (Berl) 85: 79–87. Hossmann, K.-­A. and Kleihues, P. (1973). Reversibility of ischemic brain damage. Archives of Neurology 29: 375–384. Huh, J.W., Raghupati, R., Laurer, H.L. et  al. (2003). Transient loss of microtubule-­associated protein 2 immunoreactivity after moderate brain injury. Journal of Neurotrauma 20: 975–984. Humphreys, R.P., Hendrick, E.B. and Hoffmann, H.J. (1990). The head-­ injured child who “talks and dies”. A report of 4 cases. Child’s Nervous System 6: 139–142. Hung, K.L. (2020). Pediatric abusive head trauma. Biomedical Journal 43: 240–250. Hunt, N.J., Phillips, L., Waters, K.A. and Machaalani, R. (2016). Proteomic MALDI-­TOF/TOF-­IMS examination of peptide expression in the formalin fixed brainstem and changes in sudden infant death syndrome infants. Journal of Proteomics 138: 48–60. Hwang, S.K. and Kim, S.L. (2000). Infantile head injury, with special reference to the development of chronic subdural hematoma. Child’s Nervous System 16: 590–594. Hymel, K.P., Jenny, C. and Block, R.W. (2002). Intracranial hemorrhage and rebleeding in suspected victims of abusive head trauma: addressing the forensic controversies. Child Maltreatment 7: 329–348. Ikonomidou, C., Price, M.T. and Mosinger, J.L. (1989). Hypobaric-­ ischemic conditions produce glutamate-­like cytopathology in infant rat brain. Journal of Neuroscience 9: 1693–1700. Inamura, K., Olsson, Y. and Siesjö, B.K. (1987). Substantia nigra damage induced by ischemia in hyperglycemic rats. A light and electron microscope study. Acta Neuropathologica (Berl) 75: 131–139. Iverson, G.L., Gardner, A.J., Shultz, S.R. et  al. (2019). Chronic traumatic encephalopathy neuropathology might not be inexorably progressive or unique to repetitive neurotrauma. Brain 142: 3672–3693.

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Oehmichen, M., Woetzel, F. and Meissner, C. (2009c). Hypoxic-­ischemic changes in SIDS brains as demonstrated by a reduction in MAP2-­ reactive neurons. Acta Neuropathologica 117: 267–274. Oehmichen, M. and König, H.G. (2012). Trauma. In. G. Klöppel, H.H. Kreipe, and W. Remmele (eds.), Pathologie, Teilband: W. Paulusand J.M. Schröder (eds.), Neuropathologie, pp. 403–445. Berlin, Heidelberg: Springer. O’Kusky, J.R. and Norman, M.G. (1994). Sudden infant death syndrome: Increased synaptic density in the central reticular nucleus of the medulla. Journal of Neuropathology & Experimental Neurology 53: 263–271. Orman, G., Kralik, S.F., Meoded, A. et  al. (2020). MRI Findings in pediatric abusive head trauma: A review. Journal of Neuroimaging 30: 15–27. Ozawa, Y. and Takashima, S. (2002). Developmental neurotransmitter pathology in the brainstem of sudden infant death syndrome: A review and sleep position. Forensic Science International 130S: S53–S59. Paine, S.M., Jacques, T.S. and Sebire, N.J. (2014). Review: Neuropathological features of unexplained sudden unexpected death in infancy: Current evidence and controversies. Neuropathology and Applied Neurobiology 40: 364–384. Petito, C.K., Feldmann, E., Pulsinelli, W.A. and Plum, F. (1987). Delayed hippocampal damage in humans following cardiorespiratory arrest. Neurology 37: 1281–1286. Poon, W.S., Rehman, S.U., Poon, C.Y.F. and Li, A.K.C. (1992). Traumatic extradural hematoma of delayed onset is not a rarity. Neurosurgery 30: 681–686. Raimondi, A.J. and Hirschauer, J. (1986). Clinical criteria  – children’s coma score and outcome scale  – for decision making in managing head-­injured infants and toddlers. In: A.J. Raimondi, M. Choux, C. Di Rocco (eds.), Head Injuries in the Newborn and Infant, pp. 141–150. New York Berlin Heidelberg: Springer. Reisner, H. and Reisner, T. (1976). Über traumatisch bedingte cerebrale Gefäßthrombosen. Wiener Klinische Wochenschrift 88: 158–161. Rickert, C.H., Gros, O., Nolte, K.W. et al. (2009). Leptomeningeal neurons are a common finding in infants and are increased in sudden infant death syndrome. Acta Neuropathologica 117: 275–282. Rooks, V.J., Eaton, J.P., Ruess, L. et al. (2008). Prevalence and evolution of intracranial hemorrhage in asymptomatic term infants. AJNR American Journal of Neuroradiology 29: 1082–1089. Sances, A. Jr., Yoganandan, N., Maiman, D.J. et al. (1986). Spinal injuries with vertical impact. In: A. Sances Jr., D.J. Thomas, C.L. Ewing, S.J. Larson and F. Unterharnscheidt (eds.), Mechanisms of Head and Spine Trauma, pp. 305–348. Goshen, New York: Aloray Publ. Sawaguchi, T., Patricia, F., Kadhim, H. et al. (2003). Clinicopathological correlation between brainstem gliosis using GFAP as a marker and sleep apnea in the sudden infant death syndrome. Early Human Development 75 (Suppl): S3–11. Scholz, W. (1951). Die Krampfschädigungen des Gehirns. Berlin, Göttingen, Heidelberg: Springer. Scholz, W. (1953). Selective neuronal necrosis and its topistic patterns in hypoxemia and oligemia. Journal of Neuropathology & Experimental Neurology 12: 249.

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Maith, J.M. (2012). Neuropathologische Diagnose des Hirntodes. Lübeck: Schmidt-­Römhild. Matschke, J., Büttner, A., Bergmann, M. et  al. (2015). Encephalopathy and death in infants with abusive head trauma is due to hypoxic-­ ischemic injury following local brain trauma to vital brainstem centers. International Journal of Legal Medicine 129: 105–114. Matschke, J., Sperhake, J.P., Wilke, N. et al. (2020). Cerebellar heterotopia of infancy in sudden infant death syndrome: an observational neuropathological study of four cases. International Journal of Legal Medicine May 21. doi: 10.1007/s00414-­020-­02316-­x. Matsuyama, T., Shimomura, T., Okumura, Y. and Sakaki, T. (1997). Acute subdural hematoma due to rupture of cortical arteries: A study of the points of rupture in 19 cases. Surgical Neurology 47: 423–427. McKee, A.C., Stein, T.D., Kiernan, P.T. and Alvarez, V.E. (2015). The neuropathology of chronic traumatic encephalopathy. Brain Pathology 25: 350–364. Miller, J.D. and Ironside, J.W. (1997). Raised intracranial pressure, oedema and hydrocephalus. In: D.I. Graham and P.L. Lantos (eds.), Greenfield’s Neuropathology, pp. 157–195. London Sydney Auckland: Arnold. Morad, Y., Wygnansky-­Jaffe, T. and Levin, A.V. (2010). Retinal haemorrhage in abusive head trauma. Clinical & Experimental Ophthalmology 38: 514–520. Mori, K. (ed.) (1991). MRI of the Central Nervous System. Berlin, Heidelberg, New York: Springer. Oehmichen, M. (1978). Mononuclear Phagocytes in the Central Nervous System. Berlin, Heidelberg, New York: Springer. Oehmichen, M. (1990). Die Wundheilung. Theorie und Praxis der Chronomorphologie von Verletzungen in der forensischen Pathologie. Berlin, Heidelberg, New York: Springer. Oehmichen, M., Auer, R.N. and König, H.G. (2009a). Forensic Neuropathology and Associated Neurology. Berlin, Heidelberg, New York: Springer. Oehmichen, M., Eisenmenger, W., Raff, G. and Berghaus, G. (1986). Brain macrophages in human cortical contusions as indicator of survival period. Forensic Science International 30: 281–301. Oehmichen, M., Jakob, S., Mann, S. et al. (2009b). Macrophage subsets in mechanical brain injury (MBI)  – a contribution to timing of MBI based on immunohistochemical methods: A pilot study. Legal Medicine (Tokyo) 11: 118–124. Oehmichen, M, Meissner, C. and König, H.G. (2000). Brain injury after gunshot wounding: morphometric analysis of cell destruction caused by temporary cavitation. Journal of Neurotrauma 17: 155–162. Oehmichen, M., Schleiss, D., Pedal, I. et  al. (2008). Shaken baby syndrome: A re-­examination of the role of diffuse axonal injury as a cause of death. Acta Neuropathologica (Berl) 116: 317–329. Oehmichen, M., Theuerkauf, I. and Meissner, C. (1999). Is traumatic axonal injury (AI) associated with an early microglial activation? Application of a double labeling technique for simultaneous detection of microglia and AI. Acta Neuropathologica (Berl) 97: 491–494. Oehmichen, M., Walter, T. and Meissner, C. (2003). Time course of cortical hemorrhages following closed traumatic brain injury. Statistical analysis of posttraumatic histomorphological alterations. Journal of Neurotrauma 20: 87–103.

TRAUMATOLOGY AND VIOLENT DEATH

20.5  Sharp Force Injury Peter Mygind Leth

20.5.1 Introduction

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A medical examiner is often asked to perform an investigation of a deceased victim with stab wounds at the crime scene or in the autopsy room, or to investigate a survivor at a hospital. The topic of this chapter is relatively broad and includes a variety of cases ranging from cases of self-­inflicted cut or stab wounds to homicides and accidents. What binds the chapter together is the lesion type, which may be defined as sharp wounds caused by sharp stabbing or cutting objects. In this chapter, we travel from the crime scene to the autopsy room or hospital ward. In which situations do we encounter sharp violence, and how common is it? What are the clinical manifestations of sharp violence? How dangerous was the assault, and will the victim suffer permanent injury? Which investigations must be performed at the scene or at the autopsy table? How can you, based on the pattern of lesions and the individual lesion, determine the cause of the lesions and investigate, describe and document the findings? Lastly, how can we reduce the number of people that are permanently disabled or killed by stabbing?

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Schulte, F.J., Albani, M., Schnitzler, H. and Bentele, K. (1982). Neuronal control of neonatal respiration  – sleep apnea and the sudden infant death syndrome. Neuropediatrics 13: Suppl, 3–14. Sellier, K. and Kneubuehl, B.P. (1994). Wound Ballistics and the Scientific Background. Amsterdam, London, New York, Tokyo: Elsevier. Sellier, K. and Unterharnscheidt, F. (1963). Mechanik und Pathomorphologie der Hirnschäden nach stumpfer Gewalteinwirkung auf den Schädel. Hefte zur Unfallheilkunde, Heft 76. Berlin, Göttingen, Heidelberg: Springer. Smith, M.-­L., Auer, R.N. and Siesjö, B.K. (1984). The density and distribution of ischemic brain injury in the rat after 2–10 minutes of forebrain ischemia. Acta Neuropathologica (Berl) 64: 319–332. Spielmeyer, W. (1922). Histopathologie des Nervensystems, Vol. 1. Allgemeiner Teil. J. Berlin: Springer. Squier, W. and Mack, J. (2009). The neuropathology of infant subdural hemorrhage. Forensic Science International 187: 6–13. Strich, S.J. (1961). Shearing of nerve fibers as a cause of brain damage due to head injury. A pathological study of twenty cases. Lancet II: 443–448. Tanner, J.M. (1978). Growth and Development of the Brain in Foetus into Man, pp. 103–116. London: Open Books. Unterharnscheidt, F. (1993a). Pathologie des Nervensystems von Hirn und Rückenmark. In: W. Doerrand G. Seifert (eds.), Spezielle pathologische Anatomie, Vol. 13/VI A, pp. 36–470. Berlin Heidelberg New York Tokyo: Springer. Unterharnscheidt, F. (1993b). Traumatologie von Hirn und Rückenmark. Traumatische Schäden des Gehirns (forensische Pathologie). In: W. Doerr, G. Seifert, and E. Uehlinger (eds.), Spezielle pathologische Anatomie, Pathologie des Nervensystems, Vol. VI B. Berlin, Heidelberg, New: Springer. Walter, T., Meissner, C. and Oehmichen, M. (2009). Pathomorphological staging of subdural hemorrhages: statistical analysis of posttraumatic histomorphological alterations. Legal Medicine (Tokyo) 11 (Suppl 1): S56–62. Waters, K.A., Meehan, B., Huang, J.Q. et al. (1999). Neuronal apoptosis in sudden infant death syndrome. Pediatric Research 45: 166–172. Watts, P., Maguire, S., Kwok, T. et  al. (2013). Newborn retinal hemorrhages: A systematic review. JAAPOS 17: 70–78. Wittschieber, D., Karger, B., Niederstadt, T. et  al. (2015). Subdural hygromas in abusive head trauma: pathogenesis, diagnosis, and forensic implications. AJNR American Journal of Neuroradiology 36: 432–439. Wittschieber, D., Karger, B., Pfeiffer, H. and Hahnemann, M.L. (2019). Understanding subdural collections in pediatric abusive head trauma. AJNR American Journal of Neuroradiology 40: 388–395. Wright, J.N. (2017). CNS injuries in abusive head trauma. AJR American Journal of Roentgenology 208: 991–1001. Wright, J.N., Feyma, T.J., Ishak, G.E. et al. (2019). Subdural hemorrhage rebleeding in abused children: frequency, associations and clinical presentation. Pediatric Radiology 49: 1762–1772. Yanagihara, T., Brengman, J.M. and Mushynski, W.E. (1990). Differential vulnerability of microtubule components in cerebral ischemia. Acta Neuropathologica (Berl) 80: 499–505.

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20.5.2 Occurrence

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Assault involving knives is a recognised problem. In European countries, stabbing is one of the most common homicide methods (Ormstad et  al. 1986; Leth 2010; Thomsen et  al. 2019b). Deaths by sharp force, most of which are homicides, account for a small percentage of the deaths investigated by legal autopsies. The perpetrators are mostly young men who are under the influence of alcohol. The victim of a female perpetrator is usually an intimate partner. A significant proportion of homicides by stabbing involve domestic conflicts and other interpersonal conflicts between people acquainted with each other. Many are partner homicides with female victims (Leth 2008; Leth 2009; Au and Beh 2011; Thomsen et al. 2019a). In Europe, the majority of stabbings occur in private homes. Rogde et  al. (2000) determined from a series of 141 homicides by sharp force in Copenhagen and Oslo (Rogde et al. 2000) that 78% of the female victims and 49% of the male victims were killed in their own home. Knives are commonly used in armed robberies and street violence. Sharp force injury accounts for 10–20 % of clinical forensic examinations, and sharp force trauma is the second most common cause of injury after blunt trauma investigated in clinical forensic practice (Schmidt 2010). To slit your own throat or stab yourself in the breast is a violent and unusual suicide method that constitutes a low percentage of all suicides (Byard et al. 2002; Assuncao et al. 2009). Most of the victims are men and are, on average, somewhat older than victims of knife homicides (Byard et al. 2002; Brunel et al. 2010), and

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The danger associated with a stab wound depends on which organs or tissues are affected. The clinician should examine the body for a long stab channel that penetrates vital organs, which might be under a seemingly trivial entrance wound. Due to shifts in soft-­tissue layers, the wound tract may close, resulting in only minimal external bleeding. Hypovolemic shock from blood loss is the most common life-­threatening complication, followed by pneumothorax. Arterial haemorrhages from major vessels may quickly cause unconsciousness and then death. If the bleeding originates from small vessels, the survival time will be longer because interstitial fluid will move into the intravascular space to maintain an adequate blood volume. The blood loss may then be several litres before death occurs. Pale organs and sparse lividity are typical autopsy findings of acute anaemia. If a bleeding vessel is in contact with the respiratory tract, fatal aspiration of blood may occur (Figure 20.5.2). Cardiac tamponade subsequent to a stab wound will occur if the opening in the pericardial sac is sufficiently small that blood escapes more slowly than it is infused. This effect is seen when a narrow, pointed object is used in the stabbing. The direct effect of injury to vital organs, such as the heart or spinal cord, may also contribute to death. If the jugular vein is severed when the body is in an upright position, air may be sucked in and transported to the right ventricle, resulting in an air embolism that may be fatal. The pathologist may be required to assess the victim’s ability to function after having been stabbed. Lesions of the heart or lung may not result in immediate incapacitation. Depending on lesion severity and the amount of blood loss, the victim may be able to function for several minutes. Transection of the spinal cord, however, will lead to immediate incapacitation. Subacute or chronic complications, such as infection, the formation of a pseudoaneurysm or an arteriovenous fistula and the formation of other types of fistulas, artery dissection, hernias and adherences, may occur. Neurological complications or impaired movements may result if a nerve or a tendon is severed.

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Homicides within the family are often impulsive, and in such cases, any sharp implement nearby may be used, such as a kitchen knife or, occasionally, a pair of scissors. Sharp force suicide is often committed with a kitchen knife. By contrast, in cases of violence amongst strangers, it is more common for the perpetrator to have been carrying a knife, typically a flick knife or a pocketknife. Any type of agent with a sharp edge or point may be used as a weapon (Figure  20.5.1). In bar brawls, a broken bottle or drinking glass may be used, often primarily as a blunt instrument that may shatter on impact, causing both blunt and incised injuries. These items can also be used as primary cutting instruments, such as by holding a bottle at the neck or a smashed beer tankard by the handle. Other sharp instruments include razors, forks, chisels, screwdrivers, awls and various tools and gardening or agricultural implements. Swords, rapiers and cutlasses are mostly of historical interest, but machetes and parangs are common in some areas of the world. Instruments such as axes, spikes and machetes have sharp and blunt effects upon impact that cause a combination of cutting injuries, and crushing of underlying tissue and bone fractures. A laceration may result if the blade edge is not sharp; furthermore, an axe has both a blunt and a sharp side.

20.5.4  Cause of death

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The medical examiner should be informed of the type and measurements of the knife. The weapon should be taken to the autopsy; however, this may not be possible because trace evidence must be secured first.

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less often alcohol intoxicated (Karlsson 1998). Only a minority of the victims are suffering from a psychosis (Fukube et al. 2008). Ritualistic suicides by hara-­kiri are mostly of historical interest. Accidental death due to sharp force trauma is uncommon and usually caused by a fall into a glass door or window, aquarium or other type of architectural glass surface (Karger et al. 2001). Some accidental deaths are caused by motorised machinery.

20.5.5  The scene

Figure 20.5.1  A woman was murdered by her ex-­husband. The broken handle of a bottle opener is protruding from a stab wound to the temple. She has also been slashed in the face with a broken bottle and cut in the neck with a grill knife.

Crime-­scene technicians are responsible for managing the scene. They must secure trace evidence at the scene before the medical examiner is admitted. The medical examiner’s prime responsibility is the dead body, but there are some areas of shared responsibility regarding trace evidence on the body. The position of the body, the distribution of blood and other trace evidence around the body must be documented and secured before the body is handled.

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It must be determined if a stabbing is a suicide or a homicide, and this decision is usually based on features of the scene, the autopsy findings and other circumstances. Many suicidal stabbings occur in private and in the bedroom or bathroom (Gill and Catanese 2002), occasionally behind a locked door (Figure 20.5.3). There will be no signs of struggle, and a suicide note may be found. The deceased may even have had the courtesy to commit suicide in the bathtub. The authors have seen an example of a suicide by stabbing in which an anatomical atlas was placed on the coffee table facing the deceased, and the use of a mirror as guidance has also been described (Riddick et al. 1989). The knife is usually found near the deceased, occasionally in the hand or in the stab wound. However, in the case of a homicide, a perpetrator may have placed the knife in the victim’s hand to give the appearance of a suicide (Figure 20.5.4). In homicide cases, there will be signs of struggle that include tumbled or broken furniture, bottles and crushed glass. If the knife has been removed from the scene, suicide is unlikely. However, it must be determined whether a well-­meaning relative removed the knife. In homicide cases in a private home, the knife may have been cleaned and returned to the kitchen drawer. In some cases, the knife may have been removed and disposed of by the perpetrator. Blood traces may be informative about the events in the scene and must be carefully documented and analysed. Fingerprints or sole prints in blood are important pieces of trace evidence. Forensic technicians who are specialised in blood-­distribution patterns can estimate the site of the attack and the number of impacts. Blood traces on the body should be photographed before moving the body. Resuscitation efforts may cause blood to flow from stab wounds. Therefore, the police must carefully record any such interference with the body. The type, number and location of the victim’s injuries are of importance for the interpretation of the blood traces on the scene. The forensic technician and the medical examiner should collaborate closely and

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Figure 20.5.2  Stab track penetrating the trachea, resulting in blood aspiration.

Figure 20.5.3  A 35-­year-­old schizophrenic female committed suicide by cutting her wrist, neck and groin and stabbing her chest in the bath of a hotel room, resulting in this bloody scene.

exchange information, and the medical examiner should visit the scene (Karger et al. 2008). The clothing should be examined. If the knife has passed through the garments, one should examine the tears produced by the knife and compare them with the sharp lesions on the body of

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Figure 20.5.5  A stab wound to the back. The upper end is pointed and the lower end is rounded. This is from the case described in Figure 20.5.4.

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the deceased. The number of tears in the clothing does not necessarily equal the number of sharp lesions. The victim may have been stabbed through a fold in the clothing that caused two holes, only the clothing may have been pierced or there may have been no clothing covering the stab wound. Crime scene technicians usually perform a more detailed investigation of clothing and it should be specified in the forensic report that the investigation performed in connection with the autopsy is preliminary. People who commit suicide by stabbing may lift or move the garments aside to stab the exposed skin, but this is not always the case (Karger et al. 2000).

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Figure 20.5.4  A 37-­year-­old man was stabbed to death by his wife after a domestic dispute. The knife was placed in the victim’s left hand, but he was right-­handed. He had a stab wound to the chest and a stab wound to the back. His wife soon confessed.

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The police investigation and court proceedings are assisted by a reconstruction of the events. The medicolegal investigation may contribute a large amount of information. All fights should be visualised as dynamic confrontations in which the participants constantly move, and oversimplified interpretations should be avoided. The assessment is based on both the single lesion and the pattern of lesions. Lesions caused by sharp force may be divided into stabs and cuts. A stab wound is caused by thrusting a sharp implement into the body, which causes a wound that is deeper than it is wide, whereas incised wounds are caused by the moving of a knife tangentially across the skin surface, which causes a wound that is wider than it is deep.

The external investigation A systematic approach is necessary in the external investigation of a victim of sharp force violence, regardless of whether the victim is alive or dead. It is customary to begin with the head and

continue with the neck, torso and limbs. All stabs and their anatomical positions should be carefully measured, labelled with a number, plotted on an anatomical diagram and photographed with a ruler, both before and after the wound edges are drawn together with transparent tape. The distance from the foot sole should always be measured for a later reconstruction of the event.

The individual external stab wound Each lesion should be carefully examined and measured. The assessment of a stab wound may assist in the identification of the type of sharp implement used. Determining whether one or more knives have produced the injuries may be difficult, and caution must be employed in the assessment. Most stab wounds result from one-­edged knives. The entrance wound from such a knife will typically have clean-­cut edges that can be approximated with one pointed end and one end that has either a fish-­tail-­like split or a squared or rounded appearance (Figure 20.5.5), but both ends may appear pointed even when the blade is single-­edged. Based on the entrance wound morphology, it is indeterminate whether the knife blade was straight or serrated. However, a serrated blade may occasionally cause abrasions if the blade has been drawn tangentially over the skin (Figure 20.5.6). An abrasion from the hilt that reproduces the pattern of the guard may be seen (Kaliszan et al. 2011) and is evidence that the knife has been fully plunged into the victim’s body. Bowie knives have a serrated back edge

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Cuts may be deepest at the point of entry and shallower towards the distal end and end in a shallow scratch. The direction of the knife may thus be determinable; however, the cuts may sometimes be deepest at the exit. If more cuts have been applied to the same area, the cut that crosses the others will be the one that has been applied last. Tangentially applied force by a sharp weapon may cause significant undercutting. This is often seen on the scalp, where cut hair bulbs can be seen on the cut surface, or in defensive wounds on the fingers. Undercut scalp wounds may bleed profusely. A blunt impact close to a bony surface, especially the cranium, may cause the skin to split cleanly so that it looks like a cut, but bridging tissue will be found in the depth of the wound, and there will often be abraded edges. With sharp force, scoring on the bone surface may be observed. The medical examiner is frequently required to assess the amount of force used in a stabbing because this has important legal implications in determining an alleged assailant’s intent to cause harm. However, the severity of the wound is more dependent on the organs that are injured than on the amount of force used. Several experimental investigations have determined which factors are of importance for penetration (Gilchrist et al. 2008). The dynamics of a stabbing depend on the weight, geometry and sharpness of the weapon, movements of the assailant and the victim and the number of layers penetrated, including the clothing. The sharpness of the knife tip is the most important factor in skin penetration. The speed of the knife on impact is also important. The kinetic energy that the knife possesses is proportional to its mass and the square of its velocity. A higher maximal stabbing speed can be achieved in an overarm attack compared with an underarm attack. Skin is elastic and resistant to knife penetration. This is especially the case with thick skin and if the skin is cut perpendicular to the skin tension lines (O’Callaghan et al. 1999). However, skin is more easily penetrated if stretched out, such as on the chest wall. When the skin has been penetrated, the elastic energy stored in the stretched skin is released, and no increased force is required to penetrate the underlying soft tissue (O’Callaghan et al. 1999). The assessment of force is somewhat subjective. A simple grading of mild, moderate and severe force is commonly used. A mild level of force is associated with the penetration of skin and soft tissue; moderate force would require the penetration of calcified cartilage or rib bone and severe force would be typical of a knife affecting dense bone, such as the spine, with damage to the blade. Different types of knives require appreciably different levels of forces to obtain the same degree of injury (O’Callaghan et  al. 1999); thus, one should be cautious when expressing an opinion on the amount of force used.

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Figure 20.5.6  A 4-­year-­old girl was killed by her father. There are several cuts to the neck caused by a kitchen knife, and the skin abrasions were caused by a serrated blade.

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that causes the corresponding edge of the wound to be torn or ragged. Scissors may cause an entrance wound that is indistinguishable from that caused by a knife wound if the scissors are used in the open position. However, if the scissors are closed (unless the scissor blades overlap completely), an atypical lightning or z-­shaped appearance may be observed. A projecting hinge screw may cause a mark or tear in the lateral edges of the wound. Stab wounds are often gaping, especially if they are located over joints or are perpendicular to the skin tension lines or underlying muscle bundles. Untreated stab wounds, which may be seen in torture cases, will leave a permanent scar (Leth and Banner 2005). Stabs to the heart may be irregular because of heart contractions. The length of the stab lesions may suggest the width of the knife blade, but the skin is elastic and may retract slightly, which would decrease the size of the wound. Moreover, the contraction of underlying muscles may have the same effect. If the knife is withdrawn at a different angle, the wound may be extended in a linear manner. One must also take into account the taper of the blade and the depth of penetration. If the knife has been withdrawn along the same track along which it was inserted, the length of the wound will indicate the minimum width of the blade at the maximum penetration depth. If the knife has been thrust more than once into the same wound or if the knife has been twisted while withdrawn, the stab wound may be V-­shaped or irregular. A single stab rarely causes several wounds. For example, this may occur if the blade perforates the arm or a female breast before penetrating the thorax.

The wound track Wound tracks must be examined to determine the direction and depth of injuries to the internal structures. The investigation is best conducted by dissection and not by probing. A postmortem CT scan may be a valuable supplement to the investigation, as mentioned in the following text.

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Figure 20.5.7  A 39-­year-­old man was killed by his stepson as revenge for many years of mistreatment.

Table 20.5.1  Suicidal and homicidal case findings of sharp force injury.

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Assessing the blade length of a missing weapon may be important; however, there are difficulties associated with making this assessment. If the knife has been driven into the wound to the hilt, the length of the wound track may be longer than the length of the blade because tissues are compressed during stabbing and then recoil. This is common for stab wounds to the abdomen and less common in chest wounds. If the blade is not inserted completely into the wound, the stab channel may consequently be shorter than the blade. When assessing stab depth, adjustments should be made for the position of the internal organs that are determined by the posture at the time of the stabbing and respiration. The direction of the thrust is often of importance for event reconstruction. This may be assessed by careful dissection of the tissue layers from the surface downwards and performing a comparison of injured deep structures with the position of the surface wound. The forensic pathologist should determine the angle of the stab channel relative to the body. This may be assessed with a great degree of certainty if two bony structures have been struck along the track of the stab wound, such as a rib and a vertebra. In other cases, it may be more difficult to determine the angle. If the  body was upright when the stabbing occurred, the tract of the stab wound in the internal organs might shift upwards when the body is placed in the supine position on the autopsy table. The variance in the relationship between the internal and external structures during the respiratory cycle must be considered. The direction of the stab wound track in the victim’s body is usually the result of a dynamic situation in which both the victim and assailant move relative to each other. Therefore, an interpretation of how a stab may have arisen is usually not possible without further information. It is however often possible to determine if a given stab has or has not arisen in a specified manner. Event reconstruction may be helpful in selected cases. A reconstruction may occur at the scene and include the accused demonstrating his actions on a figurant of the same size as the victim with a knife replica.

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The numbers of lesions and patterns

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The number of sharp lesions may vary between cases. In homicide cases, often only one stab to the chest that includes heart or lung lesions is observed. If the stabbing is a result of a frontal confrontation with a right-­handed assailant, stab wounds are usually found on the left side of the chest (Ormstad et al. 1986). In some homicide cases, stabs or cuts may be informative about the perpetrator’s state of mind. There will be, on average, fewer sharp lesions if the perpetrator is a woman and often only one lesion. In homicide cases, the stabs and cuts are frequently aimed at the chest and neck (Ormstad et al. 1986), although the arms or hands may be injured because of the victim’s defensive actions, and the resulting pattern is more random than that observed in suicide cases. As the number of stab wounds increases, more body regions are involved, including the back. If a large number of lesions are observed, the term ‘overkill’ may be used (Figure  20.5.7). Overkill may indicate a personal conflict, but may also be observed in psychotic or sexually motivated homi-

Suicidal

Homicidal

Preferred anatomical regions: wrist, neck, thorax

More random anatomical distribution

Single or a few deep wounds

Any number of deep wounds

Stab tracts are often descending

Stab tracts in different directions

Hesitation wounds

No hesitation wounds

Regularly repeated wounds

Irregularly repeated wounds

No defensive lesions

Defensive lesions

Only accessible parts of the body are injured

Other parts of the body may be injured

Avoidance of areas with high sensitivity to pain

No avoidance of areas with high sensitivity to pain

Often extensive blood traces on the victim’s hands

Often less blood on the victim’s hands unless from defensive wounds

Usually no other signs of struggle Clothing may be spared

May be other signs of struggle Clothing almost never spared

cides. In cases of non-­lethal violence, other areas of the body may be the primary targets. An interview study from Scotland indicated that knife-­fight participants may strike at areas such as the stomach and buttocks to reduce injury and usually try to avoid slashing the opponent’s face (Bannister et  al. 2010). The differences between homicidal and suicidal stab wound patterns are listed in Table 20.5.1 and are described in the following text.

TRAUMATOLOGY AND VIOLENT DEATH

Sharp force injuries to different body regions

Head Sharp violence towards the head may cause lesions on the face or scalp. A stab wound rarely penetrates the cranium to the brain, usually through the thin bone in the temple or the eye. In cases of an attack with a broken bottle, many bleeding cuts to the face may result (Figure 20.5.1).

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Incised injuries to the neck are predominantly homicidal or suicidal and only occasionally accidental. Self-­inflicted neck cuts are characterised by many separate shallow cuts and linear abrasions and one or a few deeper lesions, which may sever the great neck vessels and the trachea or larynx. The shallow lesions are termed ‘tentative’ or ‘hesitation’ wounds (Figure 20.5.9). A self-­ inflicted neck cut by a right-­handed person will often begin high on the left side and pass downwards across the front to terminate at a lower level of the left side. The cut is usually deeper at the origin and then lessens as it reaches the end. Such wounds are often unusually clean-­cut because the skin is manually stretched out before the cut is made. Self-­inflicted incised wounds may be surprisingly deep and may occasionally graze the vertebrae. Homicidal neck incisions or stabs are more irregular and haphazardly placed and are often, but not always, deeper

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In the majority of suicides, a single or small number of wounds are observed, but numerous self-­inflicted wounds are occasionally seen (Madea and Schmidt 1993; Kaliszan et al. 2010). Multiple superficial self-­inflicted scratches, cuts or streaky reddening may be found in individuals, mostly women, who claim to have been victims of an attack (Figure 20.5.8) or in individuals suffering from mental diseases or personality disorders. Self-­inflicted wounds are also occasionally seen after real sexual assaults or long-­term abuse. These wounds are always observed in easily reachable body regions and are usually of minor severity. Sensitive body parts such as nipples and lips are avoided. The lesions are linear or slightly curved, equally shallow and of uniform shape. The lesions have a grouped and parallel or criss-­cross arrangement and are either distributed symmetrically or primarily on the non-­dominant side of the body. Damage to the clothes is absent or inconsistent (Figure 20.5.8). Self-­injurious behaviour may be a sign of an acute life crisis, and these individuals should be referred to proper psychiatric or psychological care. Injuries may be inflicted for the purpose of insurance fraud. Most often, a finger is cut off, usually on the non-­dominant side. The thumb or the index finger has the highest insurance compensation. In contrast to accidental finger amputations, the cut in cases of intentional amputation is usually perpendicular to the long axis of the finger and at the level of the proximal phalanx. Hesitation marks may be observed. The reconstructed position of the hand at the moment of injury may appear unnatural (i.e., ‘execution position’). However, it may be difficult to prove that an injury was self-­inflicted.

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Figure 20.5.8  A 45-­year-­old woman who claimed to be a rape victim displays multiple, superficial, self-­inflicted cuts. Note the inconsistency of tears in the clothing.

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Extremities Multiple parallel incisions to the ventral aspect of the wrist or elbows are typical of self-­inflicted injury (Figure 20.5.11). These injuries are primarily perpendicular to the wrist, but some may be longitudinal or oblique, and bilateral involvement is common (Byard et al. 2002). The incisions are usually shallow and rarely lethal, even when the radial or ulnar artery is severed (Leth 1996). Multiple linear scars after such self-­inflicted incisions may be seen in individuals who have died from other causes. These scars may be an indication of previous self-­inflicted harm. Defensive injuries most frequently present as cuts or stabs on the forearms and hands (Figure  20.5.12), and are traditionally divided into active and passive defensive injuries. Passive defensive injuries occur if the victim tries to ward off an attack by raising his/her arms, which causes stabs or cuts to the dorsal side of the hand, forearm and upper arm. Active defensive injuries occur when the victim grabs or clutches the blade, thereby cutting himself/herself in the palms and ventral surface of the fingers or in the web between the thumb and index finger. The differentiation

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(Figure 20.5.10). If the victim is a small child or has been sleeping or unconscious, it will be easier for the perpetrator to make a clean cut. In such cases, the neck may be almost completely severed or, in cases involving infants, completely severed. Accidental sharp injury to the neck is occasionally seen in traffic accidents or accidents involving glass, often windows.

Figure 20.5.11  Self-­inflicted cuts to the left arm. This image is from the case described in Figures 20.5.3 and 20.5.9.

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Figure 20.5.9  Self-­inflicted cuts to the neck. This image is from the case mentioned in Figure 20.5.3.

Figure 20.5.10  A 75-­year-­old man was murdered by his wife. Homicidal cuts to the neck and face can be seen.

Figure 20.5.12  Defensive wounds to the left hand. This image is from the case described in Figure 20.5.1.

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Figure 20.5.13  A 43-­year-­old homosexual man was stabbed in the neck, chest and abdomen by his lover. The stab wounds have a random distribution compared with the stab wounds seen in Figure 20.5.13.

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between active and passive defensive injuries should not be used too rigidly; the many possible interactions between the victim and perpetrator must be considered. In some autopsy studies, two-­thirds of defensive wounds to the arms and hands were seen on the left-­hand side, but this cannot be demonstrated in studies of victims who survived an attack (Schmidt 2010). Some survivors have only defensive injuries, which is most likely because the attacker primarily used a knife to threaten the victim. Defensive wounds are occasionally seen on the thighs or shins when protecting the abdomen and genitals. Defensive injuries are of medicolegal importance because they indicate that the victim was conscious and actively tried to defend against an attack. Defensive injuries may be completely absent in victims who were severely alcohol-­intoxicated at the time of the attack (Racette et al. 2008). The number of defensive wounds is high in individuals with many stab wounds to other parts of the body, most likely resulting from protracted fights (Rouse 1994). When investigating the perpetrator, one should investigate his/her hands for cuts that may have occurred if the perpetrator had lost his/her grip on a handle that had become slippery with blood. This may occur after hitting solid resistance, such as a bone with a knife that does not have an adequate hand guard. The blade may either wound the palm of the dominant hand, often in the web between the thumb and index finger, or the fingers, depending on how the edge was orientated and the handgrip. If the blade protrudes to the ulnar side of the fist with the edge side facing up, the cuts will be on the flexor side of the fingers in a step-­ like arrangement when the fingers are extended, with decreasing depths of the cuts in the ulnar-­radial direction. If tendons are severed and the tendon stumps are strongly retracted, this would suggest that the knife was held in a firm handgrip at the moment of injury (Schmidt 2010). By contrast, defensive wounds are seen on both dominant and non-­dominant hands and are predominantly located on the radial side, with the little finger less frequently affected.

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Chest

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Stabs in the thoracic region are commonly seen in homicides and occasionally in suicides. In homicide cases, it is characteristic for the ribs to be severed, and the stab patterns have a random quality (Figure 20.5.13). By contrast, in suicide cases, stabs are typically located between the ribs and have a horizontal orientation (Brunel et  al. 2010). There may be many stabs which are frequently shallow (i.e., hesitation stabs), but they have a regular pattern and are parallel. Because most people know that the heart is on the left side, the lesions will typically be in the left precordium (Figure 20.5.14).

Abdomen Stabs to the abdomen are common in homicides and occur occasionally in suicides in which the epigastrium is the preferred region. The author once saw a suicide victim with a stab

Figure 20.5.14  A 42-­year-­old woman was suffering from disseminated sclerosis and committed suicide by stabbing herself in the chest.

wound to the epigastrium. There were several ‘hesitation-­type’ stabs to the liver (Figure 20.5.15) and one deep stab to the heart, all of which were from the same entrance wound, but with different directions.

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Assuncao, L. A., Santos, A. and Magalhaes, T. (2009). Suicide By Sharp Force Injuries  – A Study In Oporto. Legal Medicine 11 (Suppl 1): S216–219. Au, K.I. and Beh, S.L. (2011). Injury patterns of sharp instrument homicides In Hong Kong. Forensic Science International, 204: 201–204. Bannister, J., Pickering, J., Batchelor, S. et al. (2010). Troublesom Youth Groups, Gangs And Knife Carrying in Scotland. Edinburgh: Scottish Government Social Research. Bolliger, S.A., Preiss, U., Glaeser, N. et  al. (2010). Radiological stab wound channel depiction with instillation of contrast medium. Legal Medicine, 12: 39–41. Brunel, C., Fermanian, C., Durigon, M. and De La Grandmaison, G.L. (2010). Homicidal and suicidal sharp force fatalities: Autopsy Parameters in relation to the manner of death. Forensic Science International 198: 150–154. Byard, R.W., Klitte, A., Gilbert, J.D. and James, R.A. (2002). Clinicopathologic features of fatal self-­ inflicted incised and stab wounds: A 20-­year study. The American Journal of Forensic Medicine and Pathology, 23: 15–18. Eades, C., Grimshaw, R., Silvestri, A. and Solomon, E. (2007). ‘Knife Crime’ A Review of Evidence. London: Centre For Crime And Justice Studies. Fukube, S., Hayashi, T., Ishida, Y. et al. (2008). Retrospective study on suicidal cases by sharp force injuries. Journal Of Forensic And Legal Medicine 15: 163–167. Gilchrist, M.D., Keenan, S., Curtis, M. et al. (2008). Measuring knife stab penetration into skin simulant using a novel biaxial tension device. Forensic Science International 177: 52–65. Gill, J.R. and Catanese, C. (2002). Sharp injury fatalities in New  York City. Journal of Forensic Sciences, 47: 554–557. Kaliszan, M., Karnecki, K., Akcan, R. and Jankowski, Z. (2011). Striated Abrasions from a knife with non-­serrated blade – identification of the

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Computed tomography (CT) can detect a high percentage of stab wounds, and it is often possible to determine the depth and direction of the wound track (Schnider et al. 2009; Leth and Christensen 2011; Leth 2015). The information gained from CT is often more reliable than the information that can be obtained from dissection or probing. Such invasive techniques change the anatomy, whereas CT shows the undisturbed anatomy. However, caution should be exerted when measuring the channel depth in soft tissues because the channel may collapse, causing measurements to be too small. Notably, not all stab wounds can be visualised on CT, especially if there are many closely grouped wounds and if the wounds are superficial. CT findings should be correlated with findings from the external examination. CT can easily detect skeletal injuries, which are indicators of thrust force. Glass fragments can usually be visualised by CT. Instillation of contrast medium in the stab wound tract has been experimentally investigated (Bolliger et al. 2010).

References

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20.5.7  Postmortem computed tomography

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Figure 20.5.15  A 53-­year-­old man committed suicide by stabbing. There was a single entrance wound in the epigastrium, several superficial hesitation-­like stabs to the liver (left image), and one deep stab to the heart (right image). He had also lighted some candles and turned on the gas, which caused an explosion and a fire. The body was severely charred, but it could be seen that the zipper of his overalls had been opened, exposing the skin.

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20.5.8 Prevention

The people who carry knives in public places are primarily socially disadvantaged young men with a criminal record (McVie 2010). Peer and family influences, as well as community disintegration, illicit drug activity and a lack of educational and employment opportunities, contribute to weapon-­carrying. Preventive strategies may focus on these socioeconomic factors and also include measures such as a ban on carrying knives in public places, stop and search powers, increased prison sentences, community ­education and awareness-­raising campaigns (Eades et  al. 2007; Bannister et al. 2010). The use of broken glass as a weapon, which is termed ‘glassing’, may be avoided by introducing tempered glass in bars and pubs.

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Madea, B. and Schmidt, P. (1993). [Unusual suicidal stab injuries]. Archiv Fur Kriminologie, 192: 137–148. Mcvie, S. (2010). Gang Membership and Knife Carrying: Findings from the Edinburgh Study of Youth Transitions and Crime. Scottish Government Social Research. O’callaghan, P.T., Jones, M.D., James, D.S. et al. (1999). Dynamics of stab wounds: Force required for penetration of various cadaveric human tissues. Forensic Science International 104: 173–178. Ormstad, K., Karlsson, T., Enkler, L. et al. (1986). Patterns in sharp force fatalities – A comprehensive forensic medical study. Journal of Forensic Sciences 31: 529–542. Racette, S., Kremer, C., Desjarlais, A. and Sauvageau, A. (2008). Suicidal and homicidal sharp force injury: A 5-­year retrospective comparative study of hesitation marks and defense wounds. Forensic Science, Medicine, And Pathology 4: 221–227. Riddick, L., Mussell, P.G. and Cumberland, G.D. (1989). The mirrors use in suicide. American Journal Of Forensic Medicine And Pathology 10: 14–16. Rogde, S., Hougen, H.P. and Poulsen, K. (2000). Homicide by sharp force in two Scandinavian capitals. Forensic Science International 109: 135–145. Rouse, D.A. (1994). Patterns of stab wounds –A 6 year study. Medicine Science and the Law 34: 67–71. Schmidt, U. (2010). Sharp force injuries in “clinical” forensic medicine. Forensic Science International 195: 1–5. Schnider, J., Thali, M.J., Ross, S. et al. (2009). Injuries due to sharp trauma detected by post-­mortem multislice computed tomography (Msct): A feasibility study. Legal Medicine (Tokyo), 11, 4–9. Thomsen, A.H., Hougen, H.P., Villesen, P. et  al. (2019a). Sharp force homicide in Denmark 1992–2016. Journal of Forensic Sciences 65: 833–839. Thomsen, A.H., Leth, P.M., Hougen, H.P. et  al. (2019b). Homicide in Denmark 1992-­2016. Forensic Science International 1: 275–282.

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instrument of crime on the basis of an experiment with material evidence. International Journal of Legal Medicine 125: 745–748. Kaliszan, M., Kernbach-­Wighton, G. and Bouhaidar, R. (2010). Multiple self-­inflicted stab wounds to neck, chest and abdomen as a unique manner of suicide. Journal Of Forensic Sciences 55: 822–825. Karger, B., Niemeyer, J. and Brinkmann, B. (2000). Suicides by sharp force: Typical and atypical features. International Journal of Legal Medicine 113: 259–262. Karger, B., Rand, S., Fracasso, T. and Pfeiffer, H. (2008). Bloodstain pattern analysis  – casework experience. Forensic Science International 181: 15–20. Karger, B., Rothschild, M.A. and Pfeiffer, H. (2001). Accidental sharp force fatalities  – beware of architectural glass, not knives. Forensic Science International 123: 135–139. Karlsson, T. (1998). Homicidal and suicidal sharp force fatalities in Stockholm, Sweden. Orientation of entrance wounds in stabs gives information in the classification. Forensic Science International 93: 21–32. Leth, P.M. (1996). [An accident with a hedge trimmer]. Nordisk Rettsmedisin 2: 1–12. Leth, P.M. (2008). Lethal stabbings in Southern Denmark. The Scandinavian Journal of Criminal Law And Criminology 3: 279–286. Leth, P.M. (2009). Intimate partner homicide. Forensic Science, Medicine, and Pathology 5: 199–203. Leth, P.M. (2010). Homicides in Southern Denmark. Homicide Studies 14: 419–435. Leth, P.M. (2015). Computed tomography in forensic medicine. Danish Medical Journal 62: 1–26. Leth, P.M. and Banner, J. (2005). Forensic medical examination of refugees who claim to have been tortured. American Journal of Forensic Medicine and Pathology 26: 125–130. Leth, P.M. and Christensen, M.R. (2011). Computerized tomography used for investigation of homicide victims. Scandinavian Journal Of Forensic Science 17: 11–16.

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Forensic Ballistics: Injuries from Gunshots, Explosives and Arrows

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Bernd Karger

barrel and out of the muzzle. For this, all firearms have some basic components (Figure 21.1).

Barrel

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The term ballistics originates from the Greek, originally meaning the science of the trajectories of thrown objects. Today, this term describes the science of gunshots, which can be subdivided into the three branches: interior (inside the firearm), exterior (from muzzle to target) and terminal (target) ballistics. In cases of biological targets, terminal ballistics is also called wound ballistics. Forensic ballistics can be briefly defined as the application of ballistics for forensic purposes. The major task of this heterogeneous discipline is the reconstruction of events producing a gunshot injury. For this purpose, basic knowledge of firearms and ammunition as well as of interior and exterior ballistics is necessary. This is especially true for wound ballistics. Every forensic pathologist should know how gunshot injuries are produced. Compared to other wounding agents relevant in traumatology, bullets have a special position due to their physical characteristics: the mass is small and the velocity is high. The resulting ­penetration mechanism and injury are rather unique and have to be understood to investigate gunshot incidents effectively.

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21.1 Introduction

21.2 Firearms

21.2.1  Basic components Conventional firearms follow a simple common principle: a propellant explodes inside a closed room and the resulting overpressures act on the rear of the bullet, thus accelerating it along the Handbook of Forensic Medicine, Volume 1, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

The barrel provides the track to accelerate the bullet. The bore can be smooth or it can have rifling (i.e., spiral grooves cut over the length of the interior surface of the barrel). The resulting spiral elevations, called lands, are raised 0.1–0.2 mm compared to the grooves, and press firmly into the exterior surface of the bullet. This guidance imparts a rotation to the bullet around its longitudinal axis, thus stabilising the bullet during free flight similar to a gyroscope. Rifled barrels therefore commonly fire bullets and smooth barrels fire spheres, mostly shot pellets. The barrel’s rear end enlarges to the chamber for loading the cartridge. The diameter of a barrel, in rifled barrels measured between lands, is called the calibre.

Breech and lock The breech (action) seals the rear end of the barrel/chamber tightly so that the cartridge case cannot move backwards during firing and no expanding detonation gases can escape. The breech can be opened for ejecting the spent case and loading a new one, but it has to be locked if the gun is ready for firing. This is done by the lock (bolt). The lock contains the cocking, triggering, ­firing and safety devices. For firing a gunshot, the chain of trigger, trigger bar, main spring, hammer and firing pin is activated. Pulling the trigger drops the hammer and thus the firing pin, which ignites the primer located at the base of the cartridge. The

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Short-­barrelled weapons are intended for short distances. In revolvers (Figure 21.3), there is a gap between cylinder and barrel approximately 0.1 mm wide where gases can escape. Most revolvers are made from a solid frame where the cylinder is mounted on a crane and can be swung out for loading the chambers. Cocking the hammer rotates the cylinder (revolver = to roll over, to rotate), which brings a new cartridge in line with the barrel. If the hammer, which drops the firing pin, is cocked manually before pulling the trigger, this is a single-­action mode of operation. In a double-­action revolver, one pull on the trigger cocks the hammer, rotates the cylinder and then drops the hammer in one continuous operation. The trigger-­pull weight in single-­action revolvers is reduced (approximately 20  N) compared to double-­action modes (approximately 80 N). What distinguishes revolvers from pistols in more practical terms is the simple and reliable mechanism. The most popular

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Figure 21.1  Sectional view of a pistol including the (1) trigger, (2) trigger bar and hammer, (3) firing pin and the magazine (inside the grip), (4) barrel and (5) slide and recoil spring. Source: Courtesy of Dr A. Wacker, Münster, Germany.

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s­hotguns) weapons. If loading of a new cartridge is performed automatically and firing is done manually, it is a self-­loading or semi-­automatic firearm. This applies to almost every pistol, that is why these are sometimes called semi-­automatics. If firing can also be done automatically (serial or fully automatic fire), this is a fully automatic weapon. Among long-­barrelled firearms, rifles have a rifled bore and shotguns have a smooth bore. Handguns commonly have a rifled bore and are subdivided according to the construction: in pistols, the barrel and the chamber are tightly connected or made from one piece, while in revolvers or revolving pistols, there are five or six chambers in a cylinder which is separated from the rear of the barrel by a fine gap. Finally, submachine guns are fully automatic weapons, constructed for pistol calibre ammunition, while machine guns chamber rifle cartridges.

Bundle revolver

- Single loader - Repeater/bolt-action - Semi-automatic - Fully automatic

Break-action/ drop-barrel/ tip-up (single loader)

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Figure 21.2  Classification of conventional firearms.

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safety device locks or disconnects one component of the trigger chain. The proof-­mark is frequently located at the breech.

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For aiming purposes, short-­barrelled firearms and shotguns are commonly equipped with a front and a rear sight which have to be in line. For precise distant gunshots of rifles, telescopic sights can be mounted.

21.2.2  Classification of small arms Firearms can be categorised according to various criteria (Figure  21.2). Contrary to long-­barrelled weapons, handguns (short-­barrelled) can be operated with one hand. Single-­shot weapons have to be loaded after each firing. Multi-­shot weapons commonly have a magazine. The loading process is done manually in bolt-­ action (mostly rifles) and break-­ action (mostly

Figure 21.3  Short-­barrelled firearms: .38 Special revolver by Smith & Wesson (top), .22 semi-­automatic pistol by Star (bottom). Source: Courtesy of Dr A. Wacker, Münster, Germany.

Forensic Ballistics: Injuries from Gunshots, Explosives and Arrows

Rifles

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Long-­barrelled firearms can tolerate higher gas pressures due to the more solid breechblock construction, and the longer barrels increase the precision. This allowed the construction of two different types of firearms: rifles and shotguns (Figure 21.4). Rifle actions can be divided into single-­shot, bolt-­action, self-­ loading and also pump-­action rifles where the breechblock is attached to a moveable fore-­end similar to a bolt handle. The rifled bore, long barrel and rather high muzzle velocities (approximately 600–1000 m/s) make the rifle a precise firearm for long distances (point shot). A reduced trigger-­pull weight increases the shooting accuracy. Therefore, many hunting rifles have a hair trigger (i.e., a second trigger) which, if pulled, reduces the trigger-­ pull weight from 20–30 N to less than 1 N. This makes a rifle with a hair trigger susceptible to accidental discharge. Assault rifles are military fully automatic rifles, the most famous being the Kalashnikov models AK-­47 and AK-­74.

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A large majority of pistols are self-­loading or semi-­automatic (see Figure  21.3), but there are single-­ shot pistols mainly intended for target shooting, and derringers (i.e., small tip-­up pistols). Semi-­automatic pistols have a slide with an integral breechblock. The expanding gases inside the chamber, or the recoil motion, move the slide and breechblock to the rear, which ejects the spent cartridge case through a port at the side of the slide and supplies at the same time a new round from the magazine inside the grip. The gun will be ready to fire 0.1–0.2 seconds after the last gunshot. Pistols can be fired in single-­or double-­ action mode. The magazine contains approximately 5–20 rounds, but a pistol can always hold one more cartridge inside the chamber.

Shotguns (see Figure  21.4) have a smooth bore and commonly fire a multitude of shot pellets or, in some instances, one solid lead projectile (shotgun slug). Shotguns are frequently single-­or double-­shot weapons with one or two barrels hinged to the frame for loading (break-­action gun, tip-­up shotgun), but there are also bolt-­action, self-­loading and pump-­action firearms. The calibre of shotguns is designated by pure numbers  – the smaller the number, the larger the bore. After leaving the muzzle, the small mass of individual shot pellets causes a pronounced deceleration in the air, and the pellets also show a dispersion crossways and alongside. Deceleration and crossways dispersion limit the effective range of shotguns to approximately 50  m depending on pellet size. Since dispersion allows covering of the target, shotguns are intended for small and moveable game at rather short distances (spread shot). The pellet dispersion can be modified by the choke, a narrowing or constriction of the muzzle end of the barrel which delays dispersion. There are full choke, 3 4 choke, 1 2 choke, 1 4 choke and cylinder-­ bore shotguns. Hunting weapons are rifles, shotguns or combined weapons. Shotgun/rifle combinations can consist of one shotgun and one rifle barrel (twin type) or two shotgun barrels and one rifle barrel (triplet type). The barrel combinations can be positioned side by side (double-­barrel version) or over-­and-­under.

SE

Pistols

Shotguns

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manufacturers are Colt and Smith & Wesson. Colt started manufacturing the famous Single Action Army model ­ (‘Peacemaker’) in 1873, but Colt and revolver are not synonyms.

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21.2.3  Special firearms and resulting injuries Black-­powder firearms Muzzle-­loading black-­powder firearms provided with a percussion cap were widely disseminated in the 19th century. Modern replicas are easily available in many countries. Most weapons fire lead spheres from rifled barrels. The rifling is not intended to impart a spin to a sphere, but to achieve a tight fit of the projectile inside the barrel, thus increasing the shooting accuracy and preventing gases from escaping. Black powder consists of sodium or potassium nitrate, charcoal and sulphur. The special characteristics such as slow and incomplete combustion result in a restricted muzzle velocity, in a gross excess of expanding gases and gunshot residues leaving the muzzle and in pronounced thermal effects in close-­range shots. The soft lead spheres commonly show considerable deformation in soft tissues, resulting in large permanent wound cavities. In contact or near-­contact gunshots, the expanding muzzle gases can contribute substantially to the overall wounding effect.

Air weapons Figure 21.4  Long-­barrelled firearms: semi-­automatic shotgun calibre 12/76 (top), 7 × 64 mm rifle including a telescopic sight (bottom). Source: Courtesy of Dr A. Wacker, Münster, Germany.

Air-­powered guns fire 4.5  mm (0.177) or .22 projectiles from rifled or smooth-­bore barrels. There are short-­and long-­barrelled weapons. Most 4.5 mm projectiles have a mass of 0.5 g, the shape

TRAUMATOLOGY AND VIOLENT DEATH

Captive-­bolt devices

Zip guns

O

N

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Zip guns are simple homemade firearms or conversions of blank guns to firearms. Construction and quality of the guns vary considerably. The simplest construction is a metal tube in which a conventional cartridge, mostly .22, is inserted, while complicated zip guns resemble conventional firearms. Some crude types constitute more of a danger to the person operating the gun than to an intended victim. Most frequently, the barrel tube can burst and the firing pin acts like a nail. Otherwise, the rear part of the apparatus or the complete zip gun can be hurled backwards violently. Most zip guns have an oversized chamber and barrel without rifling, resulting in reduced muzzle velocity, unstable bullet flight and considerable gas leakage from chamber and barrel. Consequently, pronounced soot deposits and heat effects occur frequently following close-­range gunshots, and rifling marks are lacking. A hand firing a zip gun can show extensive and atypical soot deposits and even corresponding skin ruptures. Low velocity and an unstable bullet flight make most zip guns a short-­range weapon.

TO

Captive-­bolt devices are used in cattle slaughtering, mostly in Middle-­European countries. A captive bolt, 7–10  cm long and 10.5 or 12 mm in diameter, is driven out of the muzzle by discharge of a blank cartridge, mostly of calibre 9 mm. Free flight of the bolt is prevented by the construction of the device. The bolt is commonly circular and the concave frontal surface has sharp edges. The frontal surface of the device has two to four gas vents. Injury therefore commonly presents as punched out skin and bone defects. The tabula interna can show internal bevelling. The skin defect is surrounded by a bullet wipe. The wound tract can be longer than the bolt and at the end there is a plug of bone and skin. In German-­speaking countries, a large number of suicides have been reported, but homicides also occur. Most captive-­bolt devices are used similarly to the original purpose by firing in contact with the frontal or parietal regions. In these cases, skin defects show soot deposits in the vicinity depending on the number and location of the gas vents. Since these can be divergent, the range of fire can be estimated from the location of the soot deposits in the case of non-­contact injuries.

sectional density of the nails can produce deep penetration of tissue similar to an arrow. In non-­contact shots, the nail/stud commonly tumbles and can only penetrate deeply if it strikes with the tip first.

SE

can be blunt (diabolo), oblong or spherical. The compressed air can be produced by different mechanisms. The air can be pumped into a storage chamber (pneumatic type), a powerful spring can be compressed by manual action (spring-­air-­compression type) or a disposable gas cartridge (CO2) is used as propellant (gas-­ compression type). Air weapons can produce muzzle velocities of approximately 70–220 m/s. Due to the low sectional density, the velocity loss in air and tissues is considerable. The border velocity of 4.5  mm spheres for penetration of the skin is about 100 m/s, and for penetration of the eye globe about 80 m/s. Most air-­powered projectiles therefore have the capacity to penetrate the skin or cornea from limited ranges of fire. Disruption of the eye globe has occurred repeatedly, as having penetrating injuries to the head, thorax and abdomen including fatalities. In particular, the thin temporal bone can be perforated. If clothing is involved, the ­combined textile–skin barrier is commonly not perforated by the projectile, producing a haematoma below the impact site.

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462

Pen guns and signal guns

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Pen guns are used to fire tear gas or to give a signal in case of emergency. Many pen guns are designed to resemble ballpoint or fountain pens, lighters or other devices. Generally, inside the pen there is a metallic cylinder housing a spring-­loaded metal rod, which acts as a firing pin. The open end of the cylinder is threaded. An interchangeable signal or tear-­gas cartridge or an additional threaded cylinder serving as an auxiliary barrel is fixed to this end. In the latter case, conventional live ammunition can be fired. Most barrels are smooth-­bore. The short barrel, the lack of rifling and the loose fit of the cartridge and bullet result in low muzzle velocities reaching one-­ third to one-­half of the original one; and there are no rifling marks present. Except for the effects of the low velocity (i.e., low penetration power and lack of bullet deformation), the use of an unconventional firearm cannot be expected. The cylinder barrels can break and accidental deaths due to dropping the pen gun on the ground with subsequent discharge of the round have occurred.

Stud or nail guns

These are industrial tools firing nails or studs into constructive material. The special blank cartridges of calibres 5.6–10 mm have different strengths. The nails can reach velocities of more than 150 m/s. Stud guns have a guard at the frontal surface which has to be pressed firmly against a surface so that the device can be fired. However, this safety mechanism can be overcome by depressing the safety guard with one hand or by a similar action. Most injuries are accidental after nails have perforated a structure or have ricocheted off a surface. After ricochets, the nails or studs are commonly bent. After contact shots, there can be an outer muzzle imprint from the safety guard and the high

So-­called non-­lethal projectiles In riot control, so-­called non-­lethal devices have been introduced. Some of these devices use projectiles accelerated by different weapon designs. However, the wounding effect of a projectile depends on the projectile itself but also on the point of impact and the course of the trajectory. Consequently, almost every projectile can be lethal in one case and non-­lethal in another case.

463

Forensic Ballistics: Injuries from Gunshots, Explosives and Arrows

cases have an extractor flange larger in diameter than the cartridge case body, while it is smaller in rimless cartridge cases. The rear base commonly carries a head stamp stating manufacturer and calibre. A selection of conventional cartridges is depicted in Figure 21.6.

21.3.2  Primer and propellant

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Rimfire ammunition is an old type in which the firing pin hits the cartridge case base (and the primer compound) eccentrically. Today, rimfire ammunition is almost exclusively produced in calibre 5.6 mm (.22), euphemistically called ‘small bore’. All other conventional cartridges are centrefire ammunition where a central pocket at the base contains the primer cap. The propellant is ignited via flash holes, Boxer and Berdan primers differ in the

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Large but lightweight rubber or plastic projectiles have a low sectional density and therefore commonly do not perforate. However, the rapid velocity loss in air is an issue, and the penetration capacity depends to a large extent on the clothing covering the skin. Therefore, perforating injuries to the head, thorax and abdomen have occurred as well as non-­perforating but severe contusions of the heart, lungs and brain. The calibre varies from 12 gauge to 60 mm, for example, MR-­35 Punch (calibre 35 mm). Furthermore, there are cartridges containing a number of rubber spheres for shotguns and larger calibres. Another variant is the projectile expanding in front of the muzzle from the muzzle gas pressure or the air resistance. The ‘Short Stop’ cartridges, for example, use a small sack containing pellets. Projectiles expanding during the impact avoid the problem of rapid velocity loss in air, but deformation is a process taking up a certain time period. Consequently, perforations can occur before deformation is complete. Conventional cartridges containing rubber and plastic bullets are also used for practising and indoor shooting.

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CHAPTER 21  

Projectile

21.3.1 Cartridge

Case

Powder/propellant

Primer

(a)

(b)

Figure 21.5  Sectional views of centrefire cartridges: (a) schematic illustration, and (b) rifle cartridge (left) and pistol cartridge (right).

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A conventional small-­arms cartridge (Figure 21.5) consists of a cartridge case, a bullet, a primer and the propellant (gunpowder). The bullet is fired when the firing pin hits the impact-­sensitive primer composition at the rear base of the case. The primer ignites the propellant inside the case and the resulting expanding gases act onto the rear base of the bullet, driving it from the case through the barrel and out of the muzzle. The cartridge case, which seals the combustion chamber additionally, is commonly made from brass. Rimmed and belted cartridge

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21.3 Ammunition

Figure 21.6  Overview of conventional cartridges. From left to right: 7 mm Remington Magnum, .308 Winchester, .223 Remington (rifle cartridges), .44 Remington Magnum, .357 Magnum, .38 Special (revolver cartridges), .45 Automatic Colt Pistol, 9 × 19 mm Luger (Parabellum), 7.65 mm Browning (pistol cartridges), .22 Long Rifle (rimfire cartridge) and 12/67.5 shotgun cartridge. Horizontal cartridge: oversized .470 Nitro Express 9.9 cm in length for hunting very large game.

TRAUMATOLOGY AND VIOLENT DEATH

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Calibre designations can be confusing for several reasons. Firstly, the term calibre refers to the diameter of the bullet or more precisely to the bore measured from land to land, but it also expresses dimensions (and content) of cartridges. Secondly, metric and Anglo-­American units are applied. Thirdly, additional expressions such as the length of the cartridge case or names are used. The term Magnum, for example, expresses that the propellant load is increased compared to a standard cartridge of the same diameter. Fourthly, shotgun calibre designations differ completely. For pistol ammunition (Table  21.1), the metric system is commonly used and due to the rapid loading mechanism, ­rimless cartridge cases including a groove for the extractor are Table 21.1  The mass and muzzle velocity of a selection of conventional pistol and revolver bullets. Approximate values from a broad range of different manufacturers are given. Energy values are deliberately not stated.

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There are several types of bullet constructions (Figure 21.7). The traditional lead sphere of muzzle-­loading black-­powder weapons was first replaced by oblong lead bullets in rifled firearms. Solid lead bullets are commonly made from antimony lead, but a variety of material is also used. A jacketed bullet has a lead (or steel) core and a thin jacket made from tombac-­plated steel, gilding metal or a similar stable material. The lead core provides the mass of the bullet, while the jacket increases the form stability. If the tip of the bullet is included in the jacketing, this is a full-­metal-­jacketed (FMJ) bullet. All military bullets are FMJ because deformation or fragmentation is not intended due to international treaties. Armour-­piercing bullets have a hard steel core such as tungsten. In partial metal-­jacketed ammunition, the jacket is open at the tip of the bullet to expose the lead core. Since the maximum pressures during perforation of a dense medium occur at this location, these bullets are intended to deform and possibly to fragment. The exposed lead tip of semi-­ jacketed hollow point bullets has a cavity in it, while the tips of semi-­jacketed soft-­point bullets are rounded, pointed or flattened. Tracer bullets are FMJ and have a tracer composition at the base. In addition, there is a great variety of special bullet c­ onstructions such as exploding bullets, shaped charges, fragmenting/frangible

21.3.4 Calibre

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21.3.3 Bullet

bullets, solid bullets from copper alloys or so-­called non-­penetrating bullets. For hunting purposes, various special bullet constructions such as Nosler, ABC or Brenneke Torpedo Ideal intended to deform and fragment have been developed. The configuration of the tips of bullets is important for interior, exterior and wound ballistic reasons. Among others, there are round-­ nose (conical), wadcutter (flat cylinder), semi-­ wadcutter and hollow-­point bullets.

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construction of the holes. In ‘experimental’ caseless ammunition, the propellant is located around the bullet as a single solid piece without a cartridge case. Flobert cartridges are a simple type where primer and propellant are identical. The primer composition used to contain lead and barium, but modern compositions are free from these elements. The classic propellant was black powder, burning rather slowly and incompletely, and thus showing a pronounced soot discharge, a large amount of flames and sparks and a low energy output. Modern propellants are smokeless nitro powder, either single-­ base ­powder/nitrocellulose powder or double-­base powder/nitroglycerin powder. Nitro powder is manufactured in different forms, which influence the combustion velocity.

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Calibre designation

(b)

(c)

(d)

Figure 21.7  Sectional view of basic bullet types. (a) Solid (lead) bullet. (b) Full-­metal-­jacketed bullet. (c) Semi-­jacketed bullet (‘soft point’). (d) Hollow point (semi-­jacketed) bullet.

Muzzle velocity (m/s)

.22 short (5.6 mm short)

1.8

290

.22 long (5.6 mm long)

1.8

360

.22 Long Rifle

2.5

330

.22 Magnum

2.6

615

6.35 mm Browning (.32 Automatic Colt Pistol)

3.2

230

7.63 mm Mauser

5.5

430

7.65 mm Browning

4.7

300

9 × 18 mm Police

6.1

300

9 × 19 mm Parabellum (Luger)

8.0

350

9 mm Makarov

6.1

340

14.9

260

7.62 mm Nagant

7.0

290

.32 Smith & Wesson

5.5

200

.38 Smith & Wesson

9.4

210

.38 Smith & Wesson Special

10.2

260

.357 Magnum

10.2

430

.41 Magnum

13.6

450

.44 Remington Magnum

15.6

440

.45 Automatic Colt Pistol

(a)

Bullet mass (g)

CHAPTER 21  

465

Forensic Ballistics: Injuries from Gunshots, Explosives and Arrows

Table 21.2  The mass and muzzle velocity of a selection of rifle bullets for military and hunting purposes or both. Again, these are approximate values depending on the actual load of different manufacturers. Energy values are not stated because they are secondary to mass and velocity. Calibre designation

Bullet mass (g)

5.45 × 39 mm Kalashnikov

3.45

7.62 × 39 mm Kalashnikov

8.0

710

5.56 × 45 mm NATO

4.0

930

7.62 × 51 mm NATO

9.5

830

.222 Remington

3.2

980

.243 Winchester

5.2/6.5

7 × 64 mm

11.2

850

.30 M1 Carbine

7.1

600

.30–06 (7.62 × 63 mm)

9.8

830

.30–30 Winchester

11.0

680

.303 British

11.2

750

.308 Winchester

9.6

830

.300 Winchester Magnum

9.7

1030

.375 H&H Magnum

17.5

830

Load of shot/pellets

Muzzle velocity (m/s)

Shot shell

900

Wad

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Powder/propellant

Primer

N

1050/960

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Figure 21.8  Sectional view of a conventional shotgun cartridge. Traditional wads are made from felt, while modern cartridges contain a plastic cup construction.

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Depending on the country, the size of the shot pellets varies from approximately 1 to 9 mm. A 12/70-­gauge shot shell contains approximately 230 pellets each 3 mm in diameter. Smaller pellets ( 20 days, really 25 days. Jan. 3.5° 32

2.

Distension of tissues by gas

35

April

May

June

July

Aug.

Sept.

Oct.

Nov.

Dec.

3.9°

5.8°

9.9°

13.0°

17.4°

18.6°

18.6°

17.3°

13.2°

8.8°

4.7°

25

16 (23)

9–10

4–5

2

1–2

2

3

4–5

10

17

10

4–5

2–3

2

3

3–4

7

10

17

25

35

25

35

25

5.

Hair loss

35

25

16 (23)

(14)

4–5

2

2

3

3–4

7

10

17

16 (23)

(16)

4–5

3

2

3

3–4

7

10

17

16 (23)

10–12

4–5

2–3

2–3

3

3–4

7

10

17

2h

(1)

3

3

3–4

11

17

28

Hands: beginning of wrinkling

(1)

(1) 28–30

(12 h)

Nails become loose

Over 35

(40) 30–32

23

(6 h)

16

5

16

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6. 7.

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IB

Discolouration of the body Peeling of the epidermis

16 (23)

TR

3. 4.

N

Median water temperature (°C) Marbling

March

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Ø 1.

Feb.

C

Month

8.

Peeling of skin in glove form

35

(45)

23

9.

Nail loss

Over 53

45

30 (40)

21

10.

Feet: beginning of wrinkling

(1)

(1)

(12 h)

(1)

2–3

2h

3

3

3–4

4

7

20

28

8

3

4

10

Over 11

20

Over 35

(6 h)

½h

2h

(1)

R

10

14

2h

Nails become loose

Over 53

40

26 (35)

17

10

5

3

4

8

12

17

28

Peeling of skin

Over 53

60

35

16

10

5

3

5–6

8–9

Over 11 (14)

20

28

13.

Nail loss

Over 53

Over 60

53

Over 35

Over 28

Over 10

3

Over 10

Over 10

Over 11

Over 20

Over 35

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11. 12.

Transudate in pleural cavities*

35

25 (40)

18 (35)

10

5

3–4

3

3

11

5

Over 20

Heart without blood

Over 39

32–34 (40)

23

14–15

9

4

3

3

5

11

20

28

16.

Brain liquefied

35

30 (40)

(23)

14–16

5

3–4

3

3

6

10

17

28

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14. 15.

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Minimum time of duration of immersion in days ( ); maximum time in days; * >500 mL in adults, >20 days, really 25 days 831/91. Source: Madea (2015).

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Table 22.4.3  Time of immersion >2–2 days, really 3–4 days.

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Month

Feb.

March

April

May

June

July

Aug.

Sept.

Oct.

Nov.

Dec.

3.9°

5.8°

9.9°

13.0°

17.4°

18.6°

18.6°

17.3°

13.2°

8.8

4.7°

25

16 (23)

9–10

4–5

2

1–2

2

3

4–5

10

17

Median water temperature (°C)

3.5°

1.

Marbling

32

Distension of tissues by gas

35

Discolouration of the body

35

25

25

16 (23)

N

2. 3.

O

Ø

C

Jan.

16 (23)

Peeling of the epidermis

35

25

Hair loss

35

25

6.

Hands: beginning of wrinkling

(1)

(1) 28–30

4–5

2–3

2

3

3–4

7

10

17

4–5

2

2

3

3–4

7

10

17

16 (23)

(16)

4–5

3

2

3

3–4

7

10

17

16 (23)

10–12

4–5

2–3

2–3

3

3–4

7

10

17

2h

(1)

TR

4. 5.

10 (14)

IB

(12 h)

Over 35

(40) 30–32

23

35

(45)

23

9.

Nail loss

Over 53

45

30 (40)

2h

16

5

2–3

3

3

3–4

11

17

28

16

10

3

3

3–4

4

7

20

28

21

14

8

3

4

10

Over 11

20

Over 35

(6 h)

½h

2h

(1)

10

5

3

4

8

12

17

28

10.

Feet: beginning of wrinkling

(1)

(1)

(12 h)

(1)

11.

Nails become loose

Over 53

40

26 (35)

17

2h

12.

Peeling of skin

Over 53

60

35

16

10

5

3

5–6

8–9

Over 11 (14)

20

28

13.

Nail loss

Over 53

Over 60

53

Over 35

Over 28

Over 10

3

Over 10

Over 10

Over 11

Over 20

Over 35

14.

Transudate in pleural cavities*

35

25 (40)

18 (35)

10

5

U

3

3

11

5

Over 20

SE

R

TO

Nails become loose Peeling of skin in glove form

U

7. 8.

(6 h)

3–4

15.

Heart without blood

Over 39

32–34 (40)

23

14–15

9

4

16.

Brain liquefied

35

30 (40)

(23)

14–16

5

3–4

3

3

5

11

20

28

3

3

6

10

17

28

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N

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Minimum time of duration of immersion in days ( ); maximum time in days; * >500 mL in adults, >2–3 days; really 3–4 days in June, water temperature is 21 °C; 411/90. Source: Madea (2015).

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CHAPTER 22   Asphyxiation

average water temperature of the river Rhine (°C)

24,0 22,0 20,0 18,0 16,0

(°C)

14,0 12,0 10,0

LY

8,0 6,0

2,0 0,0 February

March

April

May

June

REH: average water temperature of the river Rhine 1961-1964 (°C)

July

August

September October

November December

SE

January

O

N

4,0

REH: average water temperature of the river Rhine, Bonn 1961-1964 (°C)

U

average water temperature of the river Rhine, Bad Honnef/Rhein 1999-2007 (°C)

TR IB

U

TO

R

Figure 22.4.3  Average monthly water temperatures of the Rhine from 1961 to 1964 and from 1999 to 2007. Source: Madea (2015).

10,0

8,7

N

8,0

O

7,0

C

6,0

R

5,0 4,0

FO

water temperature river rhine (°C)

9,0

2,9

3,0 2,0 1,0

.1 2. 20 02 27 .1 2. 20 02 28 .1 2. 20 02 29 .1 2. 20 02 30 .1 2. 20 02 31 .1 2. 20 02 01 .0 1.2 00 02 2 .0 1.2 00 03 2 .0 1.2 00 04 2 .0 1.2 00 05 2 .0 1.2 00 06 2 .0 1.2 00 07 2 .0 1.2 00 08 2 .0 1.2 00 09 2 .0 1.2 00 10 2 .0 1.2 00 2 11 .0 1.2 00 2 12 .0 1.2 00 2

02 20

26

20 25

.1 2. 24

.1 2.

02

0,0

measuring point: Bad Honnef / rhine

average water temperature

Figure 22.4.4  Example of a fluctuation of water temperatures during a period of 20 days (original water temperatures of the river Rhine from 24 December 2002 to 12 January 2003). Source: Madea (2015).

TRAUMATOLOGY AND VIOLENT DEATH

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22.5 Death in High Altitude and Barotrauma Mattias Kettner

22.5.1  Physical background of high altitude The atmosphere of the earth is defined by its chemical composition. It is stratified into layers with nitrogen (approximately 78%), oxygen (approximately 21%), and argon (approximately 0.9%) as their main constituents. While the Karman ellipsoid represents the boundary of the homosphere or anthroposphere at a height of approximately 100 km above sea level, the physiological anthroposphere or troposphere reaches up to approximately 12 km above sea level. The stratosphere reaches from 12 to 50  km above sea level, where it passes into the mesosphere comprising the volume between 50 and 80 km above sea level. The thermosphere constitutes the outermost layer of earth’s atmosphere ranging from approximately 80 km to 700 km above sea level. While the gravitational field of earth is constant and thus different levels of the constituents could be expected in Maxwell–Boltzmann statistics, a zoning of constituents based on their molar masses is prevented by turbulences and vertical air movements within the layers. The troposphere shows a rotation-­based flattened ellipsoid form with an equatorial bulge, which may reach up to 18 km in thickness and has a minimal thickness between 6 and 8 km at the poles. Within the troposphere, there is a relatively stable vertical pressure gradient. Barometric pressure at a specific height above sea level can be calculated with the barometric formula, which uses the international standard atmosphere (ambient temperature of 15 °C, barometric pressure of 1013.25 hPa, temperature gradient of 0.65 K/100 m);

FO

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N

TR IB

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Brinkmann, B. and Madea, B. (2004). Handbuch Gerichtliche Medizin, Band I. Berlin, Heidelberg, New York: Springer. Doberentz, E. and Madea, B. (2010). Estimating the time of immersion of bodies found in water – an evaluation of a common method to estimate the minimum time interval of immersion. Revista Española de Medicina Legal 36: 40–50. Doberentz, E. and Madea, B. (2016). Estimation of duration of immersion. In: B. Madea (ed.), Estimation of the Time Since Death, 3rd edn, pp. 189–203. Boca Raton, FL: CRC Press Taylor & Francis Group. Dotzauer, G. and Tamaska, L. (1968). Hautveränderungen an Leichen. In: Marchionini (Hrsg), Handbuch der Haut-­und Geschlechtskrankheiten (Ergänzungswerk Bd 1, T1). Gee, D.J. (1985). Drowning. In: C.J. Polson, D.J. Gee and B. Knight (eds.), The Essentials of Forensic Medicine, 4th edn, pp. 421–448. Oxford, New York, Toronto: Pergamon Press. Haberda, A. and Hofmann, R.v. (1927). Lehrbuch der gerichtlichen Medizin, 11. Aufl. Berlin, Wien: Urban und Schwarzenberg. Henßge, C., Brinkmann, B. and Püschel, K. (1984). Todeszeitbestimmung durch Messung der Rektaltemperatur bei Wassersuspension der Leiche. Zeitschrift fur Rechtsmedizin 92: 255–276. Henßge, C., Knight, B., Krompecher, T. et al. (2007). The Estimation of the Time Since Death in the Early Postmortem Period, 2nd edn. London, Sydney, Auckland: Edward Arnold. Knight, B. (1982). Legal Aspects of Medical Practice, 3rd. edn. London, Melbourne, New York: Churchill Livingstone. Knight, B. (1996). Forensic Pathology, 2nd edn. London, Sydney, Auckland: Edward Arnold. Madea, B. (2002). Estimation of duration of immersion. Nordisk Rettsmedisin 8: 4–10. Madea, B. (2015). Praxis Rechtsmedizin. 3. Auflage, Berlin, Heidelberg, New York: Springer. Madea, B., Stockhausen S. and Doberentz, E. (2016). Estimation of the immersion time in drowned corpses – A further study on the reliability of the table of Reh. Archiv fur Kriminologie 237: 1–12. Mant, A.K. (1984). Taylor’s Principles and Practice of Medical Jurisprudence. London: Churchill Livingstone. Mueller, B. (1975). Gerichtliche Medizin. 2. Aufl., Bd. I. Berlin Heidelberg New York: Springer. Polson, C.J., Gee, D. and Knight, B. (1985). Essentials of Forensic Medicine. 4th edn. Oxford: Pergamon Press. Prokop, O. and Göhler, W. (1976). Forensische Medizin. 3. Aufl., Stuttgart New York: Gustav Fischer. Püschel, K. and Schneider, A. (1985). Die Waschhautentwicklung in Süß-­und Salzwasser bei unterschiedlichen Temperaturen. Zeitschrift fur Rechtsmedizin 95: 1–18. Reh, H. (1967). Anhaltspunkte für die Bestimmung der Wasserzeit. Deutsche Zeitschrift für die Gesamte Gerichtliche Medizin 59: 235–245. Reh, H. (1969). Diagnostik des Ertrinkungstodes und Bestimmung der Wasserzeit. Düsseldorf: Triltsch. Reh, H. (1984). Über den frühpostmortalen Verlauf der Waschhaut an den Fingern. Zeitschrift fur Rechtsmedizin 92: 183–188.

Reh, H., Haarhoff, K. and Vogt, C.D. (1977). Die Schätzung der Todeszeit bei Wasserleichen. Zeitschrift fur Rechtsmedizin 79: 261–266. Spitz, W.U. (1993). Medicolegal Investigation of Death, 3rd edn. Springfield, Illinois: Charles C. Thomas. Weber, W. (1978). Zur Waschhautbildung der Fingerbeeren. Zeitschrift fur Rechtsmedizin 81: 63–66. Weber, W. (1982). Flüssigkeitspenetration durch Leistenhaut. Zeitschrift fur Rechtsmedizin 88: 185–193. Weber, W. and Laufkötter, R. (1984). Stadien postmortaler Waschhautbildung  – Ergebnisse systematischer qualitativer und quantitativer experimenteller Untersuchungen. Zeitschrift fur Rechtsmedizin 92: 277–290.

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References and further reading

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(22.5.1)

The usage of the barometric formula (which employs a standard atmosphere) is somewhat limited, since individual variations (dependent on a given situation; for example, height above sea

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(22.5.3)



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This equation describing ideal gases can be applied for real gases such as those in anatomical cavities or gases solved in body fluids under normal conditions. With increasing altitude, barometric pressure decreases resulting in adiabatic expansion and cooling of −6.5 K/1000 m within the troposphere. While ambient air (gas) increases in volume at higher altitudes, respiratory volumes remain stable leading to a decrease of inhaled substance (oxygen) per breath. The fraction of inspired oxygen (FiO2) at sea level of 0.21 decreases to 0.07 at 8000  m height (e.g., in the Himalayan and Karakoram ranges) resulting in a decrease in partial pressure of inspired oxygen:

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those who have to acclimatize after acute exposure to high altitude and those who are long-­term dwellers in the respective regions of earth; for example, residents of the Andean, Tibetan, and other high-­altitude Central Asian regions as well as residents of the Ethiopian highlands. In these regions, populations have been settling for thousands of years allowing for genetic adaptation to occur as opposed to other regions of the world; for example, the Rocky Mountains, which have been permanently inhabited for less than 200 years (Moore 2017). With a decrease of oxygen levels in ambient air (and thus a decreasing PiO2), sensors activate transcriptional, translational, and post-­translational responses protecting the cell and the entire organism from the consequences of severe oxygen deprivation and O2-­limited metabolism. On a systemic level, human adaptation to hypoxic conditions is affected by an increase in metabolic activity and blood flow accompanied by an increase of lung ventilation. The underlying mechanism is a complex arterial chemotransduction system in the carotid and aortic bodies monitoring arterial oxygen and carbon dioxide levels. On a cellular level, numerous models of oxygen-­ sensing mechanisms have been ­discussed over the past decades (e.g., O2-­sensing heme proteins,  nicotinamide adenine dinucleotide phosphate  [NADPH] oxidases, O2-­sensitive ion channels, etc.). Although the precise mechanism remains to be elucidated, there is growing evidence of a decisive role of mitochondria releasing reactive oxygen species (ROS) under hypoxic conditions in O2 sensing in diverse cell types (Schumacker 2014). During short-­term acclimatization to high altitude (e.g., due to air lifting, etc.), the initial increase in pulmonary ventilation as an acute phase response to hypoxia (the so-­called hypoxic ventilatory response (HVR)) may lead to a hyperventilation-­mediated ventilatory depression due to a decrease of the PaCO2 in a second phase. In sustained hypoxic environments, a gradual rise of ventilation is then noted for hours to days. This sequence of physiological regulatory mechanism in adaptation has been termed ventilatory acclimatization to high altitude (VAH). During de-­ acclimatization, a gradual decline in ventilation to its normoxic level has been described, while blunting of HVR during long-­ term adaptation to high altitude may take years (Teppema and Berendsen 2014).

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level, climatic and seasonal influences, etc.) may result in inaccuracy of barometric pressure calculation as compared to a pressure measurement. The thermodynamic equation of the state of ideal gases relates the physical parameters of pressure, volume, temperature, and (amount of) substance using the (molar) gas constant. The Boyle–Mariotte law relates pressure and volume under isothermal conditions assuming a constant amount of substance. Thus, pressure and volume of an ideal gas are reciprocally proportional quantities with a constant product:

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At a given height above sea level, measured barometric pressures vary between geographical regions. The equatorial bulge and the resulting ellipsoid form of the troposphere lead to significantly lower barometric and thus partial pressures of inspired oxygen in polar regions as compared to equatorial regions (e.g., the partial pressure of inspired oxygen (PiO2) during an ascent to Mount McKinley (Denali) (height of 6190 m; 63° northern latitude) closely resembles that of an ascent to Mount Everest (height of 8848 m; 27° northern latitude) (Trübsbach et al. 2011).

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22.5.2  Adaptation to high altitudes Human adaptation (acute or long-­term) has been characterized as any features of structure, function, or behavior that increase the  ability to survive and reproduce in a given environment (Dobzhansky 1968). High-­ altitude adaptation challenges for mountaineers as well as long-­term dwellers mainly result from the reduction in barometric pressure and thus PiO2. While the decline in PiO2 is approximately (inversely) linearly correlated with higher altitude, O2 levels in the arterial blood show a distinct decline only after exceeding altitudes of 2500 m (which is basically caused by O2-­binding kinetics and the resulting hemoglobin–O2 dissociation curve). To understand adaptation processes and to differentiate the respective pathological entities accordingly, two main groups of people subjected to these ­processes can be identified:

22.5.3  Acute altitude illness (AAI) AAIs are typically diagnosed in patients with a history of prior rapid ascent to high altitudes; for example, tourists/mountaineers or working staff/soldiers. Despite the fact that altitudes necessary for the development of altitude-­related disease symptoms vary interindividually and some susceptible individuals may experience symptoms beginning at a height of approximately 1500  m above sea level, the term high altitude is usually defined as above 2500  m (Imray et  al. 2011). Although most patients after rapid exposure to high altitude show an onset of typical symptoms within hours after ascent, more symptoms may appear up to 5 days after an ascent to high altitude. While exposure to high altitude

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Table 22.5.1  The 1991 Lake Louise Consensus on the Definition of Altitude Illness defined the three main entities in AAI. 1991 Lake Louise International Hypoxia Symposium Consensus on the Definition of Altitude Illness AMS

In the setting of a recent gain in altitude, the presence of headache and at least one of the following symptoms: -­gastrointestinal (anorexia, nausea, or vomiting) -­fatigue or weakness -­dizziness or lightheadedness -­difficulty in sleeping

HACE

Can be considered “end-­stage” or severe AMS. In the setting of a recent gain in altitude, either:

In the setting of a recent gain in altitude, the presence of the following:

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-­the presence of a change in mental status and/or ataxia in a person with AMS -­or the presence of both mental status changes and ataxia in a person without AMS

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Symptoms (at least two): -­dyspnea at rest

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-­ cough -­weakness or decreased exercise performance -­chest tightness or congestion

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Signs (at least two):

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-­crackles or wheezing in at least one lung field -­central cyanosis

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-­ tachypnea -­ tachycardia

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Sources: Adapted from Sutton, J.R. et al. (1992); Luks, A.M. et al. (2017).

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Acute mountain sickness

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and the concomitant hypoxic environment will lead to hypoxia-­ related alterations of organ functions in all relevant organ systems, three main entities resulting from acute exposure to high altitude can be identified: acute mountain sickness (AMS), high-­altitude cerebral edema (HACE), and high-­altitude pulmonary edema (HAPE) (Table 22.5.1) (Sutton et al. 1992; Luks et al. 2017).

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AMS is a syndrome formed by rather nonspecific symptoms including high-­altitude headache (HAH, which may be absent in about 5% of the patients with AMS (Sampson et al. 1983) and up to 33% of patients developing HACE (Wu et  al. 2006)) and in addition either nausea, vomiting, anorexia, insomnia, dizziness, obnubilation, fatigue, or a combination of the latter symptoms (Marmura and Hernandez 2015). AMS is rather uncommon under 2500 m above sea level, but affects up to 75% of nonacclimatized travelers at an altitude of 3000  m (Prince et  al. 2020). HAH is common in patients with a history of (ultra)rapid ascent to high and very high altitudes and is defined as a headache presenting after rapid ascent to altitudes of >2500 m, which resolves within 24 h after descent. Low oxygen saturation, reduced fluid intake, exertion, and a medical history of migraine are risk factors to develop HAH (Marmura and Hernandez 2015).

In the typical course of the disease, symptoms begin with a 4–12 h delay after the ascent and regress over 1–3 days. The severity of AMS can be self-­assessed or assessed by others using the updated 2018  Lake Louise Acute Mountain Sickness Score (Table  22.5.2) (Roach 2018, for clinical assessment, also Sutton et al. 1992). In this scoring system, major symptoms of AMS are included offering a subclassification based on the severity of each symptom. A total score of 3 and more points including a headache score of at least one point is considered sufficient to diagnose AMS. The AMS clinical functional score at the bottom of the scoring sheet indicates affection of activities by AMS. When symptoms including nausea and headache progress not responding to first-­line anti-­emetics and analgesics or adequate measures cannot be taken (e.g., AMS clinical functional score of 2 and more points), AMS may progress to HACE(Wilson et al. 2009; Luks et al. 2017).

High-­altitude pulmonary edema HAPE is a fatal form of noncardiogenic edema resulting from hypoxia, typically during a rapid ascent to high altitudes either by nonacclimatized travelers or by acclimatized high-­altitude dwellers returning from low altitudes (re-­entry HAPE) (Luks et  al. 2017). Edema forms due to (severe) pulmonary hypertension as a result of vasoconstriction in hypobaric hypoxia (Mason 2000).

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Table 22.5.2  The 2018 Lake Louise Acute Mountain Sickness Score uses a self-­assessment questionnaire on major AMS symptoms (headache, gastrointestinal symptoms, fatigue and/or weakness, and dizziness (lightheadedness) to diagnose AMS and to classify the severity thereof. 2018 Lake Louise Acute Mountain Sickness Score Self-­report questionnaire

Gastrointestinal symptoms

Not at all

0

A mild headache

1

Moderate headache

2

Severe headache, incapacitating

3

Good appetite

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Headache

Score points

Poor appetite or nausea

1

Fatigue and/or weakness

Not tired or weak Mild fatigue/weakness

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Severe nausea and vomiting, incapacitating

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Moderate nausea or vomiting

Severe fatigue/weakness, incapacitating

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Symptom score:

1

Moderate fatigue/weakness

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AMS clinical functional score

Not at all Symptoms present, but did not force any change in activity or itinerary

0 1

My symptoms forced me to stop the ascent or to go down on my own power

2

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3

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Overall, if you had AMS symptoms, how did they affect your activities?

AMS clinical functional score

Sources: Adapted from Sutton, J.R. et al. (1992); Roach 2018.

The typical X-­ray image of the lungs with the characteristic patchy edema (Figure 22.5.1) has been explained by an inhomogeneous hypoxic vasoconstriction resulting in a situation where unconstricted vessels fail due to massive pressure exposition (Bärtsch and Roach 2001; West 2013). Clinically, HAPE is characterized by one or a combination of AMS symptoms combined with increasing dyspnea beginning within hours after arrival in high altitude. An initially dry cough under exertion turns p ­ roductive with frothy

white and later blood-­tinged sputum (West 2013). Upon auscultation crackles may be heard at the lung bases. Typical symptoms also include tachycardia, tachypnea, cyanosis, and mild pyrexia (West 2012; Luks et al. 2017). Incidences of HAPE vary substantially with regard to differing altitudes and study populations. They have been reported to range between 0.57% at 4500 m up to 15% at higher altitudes (Menon 1965; Bärtsch et al. 1990; Jemsen and Vincent 2020). Patients with a history of prior ascents with

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TRAUMATOLOGY AND VIOLENT DEATH

deep coma, psychiatric changes of varying degree, confusion, and ataxia of gait (Hackett and Roach 2004). HACE is usually an aggravatio per continuitatem secondary to severe AMS and occurs commonly with HAPE. Reported incidences vary substantially and have been described (conceding obvious differences in study designs, ascent rates, and data acquisition) ranging from 0.5% (varying rates of ascent in 5355 visitors to 4555  m in Tibet) (Bärtsch and Roach 2001) to 31% (Vedic pilgrims at 4300 m in Nepal) (Basnyat 2000). The typical course of the disease develops over 24–48 h after initial AMS symptoms have appeared. Typically, deterioration of consciousness with drowsiness, progressing lassitude, and evident confusion are seen alongside other neurological symptoms such as ataxic gait (Hackett and Roach 2004). HACE in its initial phase has been compared to a state of mild drunkenness (Clarke 1988). HACE patients with a need for hospitalization should be evaluated completely considering patient history and thorough physical as well as laboratory examinations including serum electrolytes, blood cell count, and renal function. Analysis of cerebrospinal fluid should be employed to rule out central nervous system (CNS) infections possibly mimicking HACE symptoms. CNS imaging using magnetic resonance tomography (e.g., fluid attenuated inversion recovery [FLAIR], weighted diffusion weighted imaging [DWI], susceptibility-­ imaging [SWI]) may be helpful if available. Etiologically, hypobaric hypoxia at higher altitudes causes an overall increase in cerebral blood flow (CBF) (despite the opposing effect of hypocapnia-­ caused cerebral vasoconstriction). Magnetic resonance (MR) studies show evidence of a vasogenic origin of the edema with increased permeability of the vascular endothelium, despite the fact that there is no clear correlation between increased CBF and HACE (or AMS) (West 2013). In MR examinations, there is increased signal intensity in white matter, markedly in the splenia of the corpora callosa (Hackett et  al. 1998). Furthermore, hypoxia-­induced cellular ion pump failure results in intracellular hypernatremia and thus osmolarity with subsequent influx of water resulting in cellular swelling in the sense of a cytotoxic edema (Fishman 1975; West 2013). The primary treatment measure in patients with HACE is a rapid descent to lower altitudes. Dexamethasone has been proven to help the cerebral symptoms (Ferrazzini et al. 1987), but has little or no effect on the physiological alterations (Levine et  al. 1989). As prehospital measures, oxygen therapy and hyperbaric bags may be tried. Once the hospital has been reached, standard intracerebral edema therapy can be applied (West 2013).

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Figure 22.5.1  (a) X-­ray of a 37-­year-­old male HAPE patient with patchy to confluent edema distribution. (b) Computed tomography section of a 27-­year-­old male patient with recurrent HAPE showing the typical patchy distribution of edema. Source: Courtesy of Prof. Bärtsch, from Bärtsch (2005).

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HAPE show a recurrence rate of up to 60%. The mortality rate has been reported to reach 11% when the patient is treated and up to 50% if no treatment is available. About half of the cases also show AMS, and up to 14% will develop concomitant high-­altitude cerebral edema (Jemsen and Vincent 2020). The optimal treatment is dependent on access to treatment facilities. Treatment possibilities include receiving supplemental oxygen, entering a hyperbaric chamber, and receiving a pulmonary vasodilator (Luks et al. 2017). In addition, phosphodiesterase inhibitors may be useful, while diuretics are contraindicated (West 2012). If there is no treatment available and in view of a severely ill current patient status, HAPE patients should descend to lower altitudes (Luks et al. 2017).

High-­altitude cerebral edema HACE is a severe and potentially fatal condition as a result of ascent to high altitudes in nonacclimatized travelers. Patients show marked disturbances of consciousness that may progress to

Mortality and autopsy findings in AAI Data on mortality rates of visitors to high altitudes are rather scarce and are usually collected for specific areas (e.g., Aconcagua (Westensee et al. 2013), Himalayas (Himalayan Data Base 2020), or Mount Everest as a part thereof (Huey et al. 2007)). Furthermore, only few of these data bases differentiate nontraumatic fatalities (Westensee et al. 2013). A review of fatalities on Mt. Aconcagua (6962 m) in a 12-­year period revealed 33 fatalities in the group of mountaineers with permissions to the summit (0.77 per 1000, or

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22.5.4  Chronic altitude illness (CAI)

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Approximately 140  million dwellers of high altitudes at more than 2500 m above sea level in different geographical regions of the earth are subject to chronic altitude-­related medical conditions. People in these regions (either as sojourners, e.g., spending years at high altitude for economic reasons, or as permanent residents)—­mainly the Andes, the Himalayas and Tibet, and the Ethiopian highlands, but also in parts of the Rocky Mountains or potentially any other populated high-­altitude area)—­live in a state of constant alveolar hypoxia and frequently exhibit severe hypoxemia resulting in concomitant pathological alterations (West 2017). To harmonize the terms used for these medical conditions, the International Society for Mountain Medicine (ISMM) issued a consensus statement in 2005, in which the terms “chronic mountain sickness” (CMS) and “high-­altitude pulmonary hypertension” (HAPH) were used (Leon-­Velarde 2005).

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In 1925, Carlos Monge Medrano reported the first case of polycythemia in a Peruvian patient living at Cerro de Pasco (Monge 1925) giving this entity its name. Since then, numerous case reports and series of patients have been reported from South America and also from other high-­altitude regions, for example, in the United States of America and Asia (for details, see West 2017). The ISMM has defined CMS in its 2005 consensus statement as “a clinical syndrome that occurs in natives and residents living for a long time above 2500  m. It is characterized by excessive erythrocytosis (Hb ≥19 dL−1 for females and ≥21 dL−1 for males), severe hypoxemia, and in some cases moderate or severe pulmonary hypertension, which may evolve into cor pulmonale, leading to congestive heart failure. The clinical picture of CMS gradually disappears after descending to low altitude and reappears after returning to high altitude” (Leon-­Velarde 2005). Excessive erythrocytosis may reach hematocrit values of up to 91% (Jefferson et al. 2002), resulting in a massive increase in viscosity and subsequently in a decrease in cerebral blood flow (Thomas et al. 1977). The typical clinical picture comprises neuropsychological symptoms including headache, fatigue, somnolence, and depression. CMS patients typically gain weight and show poor exercise tolerance. Characteristically, these symptoms correlate with altitude, and diminish or disappear after a descent to sea level to reappear after return to high altitude (West et  al. 2013). Macroscopically, a combination of “virtually black lips and wine-­ red mucosal surfaces” has been described in Andean natives (Heath and Williams 1995). In Caucasians, signs may be limited to those of hypoxic lung disease at sea level; for example, clubbed fingertips and congested conjunctivae (West et al. 2013). Avoiding altitude habitation with a rapid and permanent descent to lower altitudes is the best therapeutic strategy, which admittedly is not a therapeutic option at all for some high-­altitude dwellers, for example, based on financial or cultural reasons. Then, venesection may be an effective therapeutic option. In

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0.077%, 21.2% were attributed to high-­altitude illness). Within this subgroup, all patients had shown HAPE and none HACE before death occurred. For those climbers, a mean height of 5131 m at the time of death was calculated. However, these figures are somewhat limited, since 15.2% of deaths were ascertained secondary to disappearance and 12.1% of the death causes remained unknown (Westensee et al. 2013). In addition, it has to be questioned if there were further mountaineers displaying signs of AAI in the subgroup with traumatic causes of death, since a fall from a height may be the initial manifestation of neurological disturbances (Firth et al. 2008). In a recent review of fatalities concerning mountaineers on Mount Everest (Huey et al. 2001), a subset of mountaineers (first-­time climbers, on commercial route, in spring during nondisaster years) were examined. Of 111 cases with assigned causes of death included for the time span between 1990 and 2019, 68.4% were classified as nontraumatic (which includes deaths due to hypothermia, sudden death, and unclassified others besides death from AAI without specification of AAI-­related subdiagnoses). Another problem is the heterogeneity of postmortem examinations (if the bodies are not missing, which is the case in approximately 20% of all fatalities above base camp on Mount Everest from 1921 to 2006) (Firth 2008). Due to varying legal requirements of the countries of death as well as the home countries, only a minor subset of the deceased is subject to full medicolegal autopsy. This may challenge the validity of certified diagnoses per se, but one has to keep in mind that a full death investigation on scene is not possible without endangering involved personnel and that a full investigation after descent is a priori hampered by a multitude of reasons (expeditions are frequently multinational undertakings with a rather short time frame of stay after the descent for most involved mountaineers). Given a suitable case history (death in or during descent from high altitude or after return to lower altitude with an anamnesis of AMS/HAPE/HACE symptoms), an AAI-­related cause of death should be taken into consideration. While only few forensic medical experts live and work in regions comprising high-­altitude areas, a “lowland” medical examiner may encounter cases of fatal AAI in daily casework, when a deceased is transported from a high-­altitude spot to his home country. Knowledge of the typical findings in the respective cases may therefore be relevant. Autopsy findings typically resemble those of pulmonary and cerebral edema. The lungs show a severe diffuse edema with increased organ weights (mean 1682 g, weights ranging 1200– 3000 g in a study comprising 10 cases of HAPE). Further findings include froth-­and fluid-­filled airways (frequently bloodstained) and dilation of the right heart (Hultgren et al. 1997). These findings resembled those of the earlier studies (Arias-­Stella and Kruger 1963). Cerebral edema seen in some of the cases was attributed to hypoxia of HAPE rather than being an etiologic factor (Hultgren et  al. 1997). Autopsy findings in HACE include macroscopic aspects of cerebral edema, for example, cerebellar herniation and flattening of gyri, as well as white matter spongiosis and petechial hemorrhage of the brain (Singh et al. 1969; Dickinson et al. 1983). Funduscopic inspection may reveal papilledema as well as retinal hemorrhage (Hackett and Roach 2004).

TRAUMATOLOGY AND VIOLENT DEATH

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Mortality and autopsy findings in CMS/HAPH

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HAPH is a medical condition affecting populations that permanently live in high altitudes under hypoxic conditions. Studies examining populations inter alia on the Altiplano have shown prevalences of up to 18% (Hakim et  al. 1983; Audi et  al. 1991; Pasha and Newman 2010), while in general a prevalence of 5–10% is estimated (León-­Velarde et  al. 2005). Therefore, HAPH is a general public health problem in countries with a relevant proportion of the population living in higher altitudes. HAPH results from hypoxia-­induced pulmonary vasoconstriction (Euler and Liljestrand 1946) and subsequent vascular remodeling of the pulmonary precapillary vessels that protect the capillaries from exposure to high hydrostatic pressure and blood flow (Maggiorini and Léon-­Velarde 2003). On a cellular level, chronic hypoxia will cause malfunction of potassium and calcium channels leading to an increase in intracellular calcium, which results in proliferation of smooth muscle cells within the pulmonary vasculature (Mirrakhimov and Strohl 2016). The clinical course in patients with HAPH is comparable with other causes of pulmonary hypertension. Patients may display exertional dyspnea, cough, hemoptysis, fatigue, chest tightness, lower extremity swelling, and/or syncope. During examination, signs of right heart failure (e.g., loud P2, tricuspid regurgitation, jugular venous distention, peripheral edema, a right ventricular heave, and ascites) may be present (Mirrakhimov and  Strohl 2016; Ulloa and Cook 2020). For the diagnosis of HAPH, a typical clinical picture with signs of right heart failure in a person living at high altitude (>2500  m above sea level) may be supported by electrocardiogram (EKG) (showing findings of right ventricular hypertrophy, a right bundle branch block, and tall p-­waves in inferior leads), echocardiogram (right ventricular and atrial dilation and increase in pulmonary pressure, both in the absence of signs in case of left-­sided heart failure), chest X-­ray (possibly showing signs of cardiomegaly or pulmonary vascular congestion), pulmonary function tests (excluding other lung pathologies), and most importantly right-­sided heart catheterization and pulmonary angiography (which should be carried out at the level of altitude where the patient resides, if possible). Here, a mean pulmonary artery pressure of >30  mmHg and a systolic pulmonary artery pressure of >50 mmHg are diagnostic for pulmonary hypertension (Mirrakhimov and Strohl 2016; Ulloa and Cook 2020). A subgroup of HAPH patients consists of patients moving to higher altitudes for longer deployments; for example, Indian soldiers and Han Chinese in Tibet. Here, a condition sometimes referred to as subacute mountain disease can be observed. In a study of Indian soldiers deployed to high altitudes of >5000 m, patients presented with dyspnea, cough, and effort angina as signs of dependent edema. Upon examination after descent to low altitude, they displayed cardiomegaly with right ventricular

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enlargement and in 17 cases pericardial effusion. Pulmonary artery pressure was elevated at rest and under mild exercise (Anand et  al. 1990). In addition, in a Lhasa-­based study of 17 patients, liver enlargement and increased jugular venous pressure were noted (Pei et al. 1989). In a study comparing Han Chinese and Tibetan populations in Tibet, described clinical signs included cyanosis, edema of the face, crackles of the chest, tachycardia, tachypnea, and liver enlargement. In this study, up to 3.64% of examined children in the Han population suffered from HAPH (Ge and Helun 2001). Data on long-­term therapeutic strategies are rather scarce and somewhat unsatisfying as compared to other causes of pulmonary hypertension. Moving to a lower-­altitude residence can be regarded as a general treatment recommendation (Sime et  al. 1971), which may not be an option for a lot of high-­altitude dwellers for various reasons. Beneficial effects of treatment with phosphodiesterase inhibitors (Xu et  al. 2014), endothelin receptor blockers, and acetazolamide have been reported in some studies (Mirrakhimov and Strohl 2016). In the acute phase of treatment, venovenous extracorporeal membrane oxygenation (ECMO) has been suggested for severe cases. If an individual develops irreversible pulmonary hypertension, pulmonary transplantation may be the only treatment option (Taimeh 2019).

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addition, acetazolamide and medroxyprogesterone have been suggested for long-­term treatment (Kryger et  al. 1978; Richalet et al. 2005).

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CMS/HAPH fatalities are (as far as pertaining to published reports) rarely subject to autopsy studies. In a report by Arias-­ Stella et  al. from 1973, which reviews two earlier studies and reports a single new case, 4 cases of CMS/HAPH are described. Here, three types of CMS are differentiated based on a classification which is not congruent with the subsequently developed 2005 ISMM consensus statement. In these studies, all patients died of (severe) cardiac insufficiency/heart failure showing right (also left in a single case) ventricular hypertrophy and peripheral pulmonary arterial thickening. In addition, two patients suffered from obesity, and two other patients had scoliosis aggravating CMS pathology, with both conditions excluding patients from the later adopted diagnostic requirements for the classification as CMS cases. Studies performed on the Han Chinese and Tibetan population in Tibet characterized postmortem findings of infant fatalities revealing extreme medial hypertrophy of muscular pulmonary arteries and muscularization of pulmonary arterioles. Massive hypertrophy and dilation of the right ventricle and the pulmonary trunk were seen macroscopically (Sui et al. 1988). In a setting with a suitable case history of long-­term dwelling in high altitude and a medical history of CMS symptoms, a CMS fatality should be taken into consideration. Autoptical examination may show right ventricular hypertrophy and hypertrophy of the pulmonary trunk, pericardial effusion, liver enlargement, as well as histologically proven (peripheral) pulmonary arterial thickening alongside corresponding hemoglobin and hematocrit values. Upon external inspection, cyanosis and edema of the face as rather unspecific symptoms may be present.

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While AAIs/AMS and CMS are medical conditions evoked by a decline in oxygen in high altitudes and the resulting hypoxia-­ related pathologies for first-­time visitors as well as long-­term dwellers in these regions of earth, hypobaric barotrauma shows pathological conditions of the human body evoked by a change in ambient air pressure and concomitant reductive or expansive gas volume adaptations. In a hyperbaric setting (e.g., during scuba diving), pressure changes affecting pathological conditions in the human body take place rapidly within a few meters of descent or ascent and the time for acclimatization is thus rather short. By contrast, hypobaric changes while ascending to and descending from a high altitude as a mountaineer usually do not lead to pressure changes adequate to cause tissue damage, for example, in anatomic cavities. But in other settings, for example, as an occupant in air travel, high and very high altitudes may be reached within a short period of time. If cabin pressure is not adequately equalized or not equalized at all, comparative supersaturation with nitrogen may result in decompression sickness and hypobaric barotrauma. Other possible scenarios include hypobaric chamber exposition as well as scuba diving followed by taking a commercial flight within a short period of time. One of the first reports of altitude-­related decompression sickness pertains to a balloon flight in 1862, when the balloon was reported to have ascended up to 8840  m and the balloonists developed inter alia paralysis of arms and legs and sudden blindness followed by loss of consciousness (Glaisher 1862). Some parts of this report have raised questions of the typical character of the symptoms in later reports, but it gives a vivid impression of the dramatic problems arising from decompression sickness in high altitude. Patients experiencing decompression sickness from high-­ altitude exposure (or a simulation thereof) will show symptoms based on the localization that gas expansion takes place after saturation under normal or hyperbaric conditions. In aviation medicine, the most frequently affected organs and structures have received special symptom names. In studies on lifetime prevalences, 75.5% of U-­2 pilots stated to have experienced either joint pain (“the bends”), skin manifestations (“the creeps”), or neurologic manifestations (“the staggers”); while 12.7% reported that they had experienced at least one episode of decompression sickness that forced them to abort their mission (Bendrick et  al. 1996). Furthermore, respiratory (“the chokes”) symptoms have been estimated to have appeared in 5% of cases (Harding 1983). Hypobaric barotrauma may affect all anatomic cavities of the human body. While in some of them the reduction of excessive gas volumes during a rapid ascent (during a flight) is affected by shunts (e.g., pressure equalization of the lungs via the trachea, and of the middle ear via the Eustachian tube), other cavities are  not vented by shunts, for example, in dental disorders. Furthermore, regular anatomic shunts may be insufficient due to anatomic abnormalities or medical conditions (e.g., impaired pressure equalization in sinusitis patients).

If pressure equalization is not sufficiently possible, the initial response is pain in the affected anatomic structure (such as in aerodontalgia). During ultrarapid ascents without pressure equalization, even in the small intestine, expanding gas volumes may cause extensive pain and, subsequently, vasovagal syncopes (Harding 1983). In patients being transported via helicopter or plane, additional problems may arise. Here, patients suffering from pneumothoraces have to be closely monitored, since a rise in altitude of approximately 1500 m has been reported to increase pneumothorax volumes by 12.7–16.2% (Knotts et al. 2013). In addition, the possibility of a spontaneous pneumothorax during a flight has to be taken into consideration, especially in patients with a known medical history of prior pneumothoraces, chronic obstructive pulmonary disease (COPD), pulmonary fibrosis, and bullous emphysema (Lippert et  al. 1991). Furthermore, volume expansion of gas-­containing devices such as balloon cuffs and endotracheal tubes has to be considered in patients not able to follow instructions (DeLorenzo et al. 2017). In those patients, problematic factors for ear barotrauma such as infections of the upper respiratory tract or otitis media have to be assessed to avoid the respective barotrauma.

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22.5.5 Barotrauma in high altitude—­ hypobaric barotrauma

Anand, I.S., Malhotra, R.M., Chandrashekhar, Y. et al. (1990, Mar 10). Adult subacute mountain sickness – a syndrome of congestive heart failure in man at very high altitude. Lancet 335 (8689): 561–565. Arias-­Stella, J. and Kruger, H. (1963, Aug). Pathology of high altitude pulmonary edema. Archives of Pathology 76: 147–157. Arias-­Stella, J., Krüger, H. and Recavarren. S. (1973, Nov). Pathology of chronic mountain sickness. Thorax 28 (6): 701–708. Audi, S.H., Dawson, C.A., Rickaby, D.A. and Linehan, J.H. (1991, May). Localization of the sites of pulmonary vasomotion by use of arterial and venous occlusion. Journal of Applied Physiology (1985) 70 (5): 2126–2136. Bärtsch, P., Baumgartner, R.W., Waber, U. et  al. (1990, Sep  29). Comparison of carbon-­dioxide-­enriched, oxygen-­enriched, and normal air in treatment of acute mountain sickness. Lancet 336 (8718): 772–775. Bärtsch, P. and Roach, R.C. (2001). Acute mountain sickness and high-­ altitude cerebral edema. In: T.F. Hornbein and R.B. Schoene (eds.), High Altitude: An Exploration of Human Adaptation, pp. 731–776. New York: Marcel Dekker. Basnyat, B., Subedi, D., Sleggs, J. et al. (2000, Summer). Disoriented and ataxic pilgrims: An epidemiological study of acute mountain sickness and high-­altitude cerebral edema at a sacred lake at 4300  m in the Nepal Himalayas. Wilderness & Environmental Medicine 11 (2): 89–93. Bendrick, G.A., Ainscough, M.J., Pilmanis, A.A. and Bisson, R.U. (1996, Mar). Prevalence of decompression sickness among U-­ 2 pilots. Aviation, Space, and Environmental Medicine 67 (3): 199–206. Clarke, C. (1988). High altitude cerebral oedema. International Journal of Sports Medicine 9: 170–174.

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Knotts, D., Arthur, A.O., Holder, P. et al. (2013, May-­Jun). Pneumothorax volume expansion in helicopter emergency medical services transport. Air Medical Journal 32 (3): 138–143. Kryger, M., McCullough, R.E., Collins, D. et al. (1978, Mar). Treatment of excessive polycythemia of high altitude with respiratory stimulant drugs. American Review of Respiratory Disease 117 (3): 455–464. Kurtzman, R.A. and Caruso, J.L. (2018, Mar). High-­altitude illness death investigation. Academic Forensic Pathology 8 (1): 83–97. León-­Velarde, F., Maggiorini, M., Reeves, J.T. et  al. (2005, Summer). Consensus statement on chronic and subacute high altitude diseases. High Altitude Medicine & Biology 6 (2): 147–157. Levine, B.D., Yoshimura, K., Kobayashi, T. et  al. (1989, Dec 21). Dexamethasone in the treatment of acute mountain sickness. New England Journal of Medicine 321 (25): 1707–1713. Lippert, H.L., Lund, O., Blegvad, S. and Larsen, H.V. (1991, Mar). Independent risk factors for cumulative recurrence rate after first spontaneous pneumothorax. European Respiratory Journal 4 (3): 324–331. Luks, A.M., Swenson, E.R. and Bärtsch, P. (2017, Jan 31). Acute high-­ altitude sickness. European Respiratory Review 26 (143):160096. Maggiorini, M. and Léon-­Velarde, F. (2003, Dec). High-­altitude pulmonary hypertension: A pathophysiological entity to different diseases. European Respiratory Journal 22 (6): 1019–1025. Marmura, M.J. and Hernandez, P.B. (2015, May). High-­altitude headache. Current Pain and Headache Reports 19 (5): 483. Mason, N.P. (2000, Feb). The physiology of high altitude: an introduction to cardio-­respiratory changes occurring on ascent to altitude. Current Anaesthesia and Critical Care 11 (1): 34–41. Menon, N.D. (1965, Jul 8). High-­altitude pulmonary edema: A clinical study. New England Journal of Medicine 273: 66–73. Mirrakhimov, A.E. and Strohl, K.P. (2016, Feb 8). High-­altitude pulmonary hypertension: An update on disease pathogenesis and management. Open Cardiovascular Medicine Journal 10: 19–27. Monge Medrano, C. (1925). Sobre un caso de enfermedad de Vaquez – sindrome eritrémico de altura. Boletin de la Academia de Medicina. Lima:Sanmartí, 7 pp. Moore, L.G. (2017, Nov 1). Measuring high-­altitude adaptation. Journal of Applied Physiology (1985) 123 (5): 1371–1385. Noh, S.J. and Lee, H. (2018, Sep). rapidly progressing fatal high-­altitude illness in a patient with hyperthyroidism. High Altitude Medicine & Biology 19 (3): 288–290. Pasha, M.A. and Newman, J.H. (2010, Jun).  High-­altitude disorders: Pulmonary hypertension: pulmonary vascular disease: the global perspective.Chest 137 (6 Suppl): 13S–19S. Pei, S.X., Chen, X.J., Si Ren, B.Z. et  al. (1989 Jun). Chronic mountain sickness in Tibet. Quarterly Journal of Medicine 71 (266): 555–574. Prince, T.S., Thurman, J. and Huebner, K. (2020, Jan-­). Acute mountain sickness. [Updated 2020  Jul 17]. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing. Richalet, J.P., Rivera, M., Bouchet, P. et al. (2005, Dec 1). Acetazolamide: A treatment for chronic mountain sickness. American Journal of Respiratory and Critical Care Medicine 172 (11): 1427–1433. Epub 2005 Aug 26. Sampson, J.B., Cymerman, A., Burse, R.L. et al. (1983, Dec). Procedures for the measurement of acute mountain sickness. Aviation, Space, and Environmental Medicine 54 (12 Pt 1): 1063–1073.

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Delorenzo, A.J., Shepherd, M. and Jennings, P.A. (2017, Mar-­ Apr). Endotracheal cuff pressure changes during helicopter transport: A systematic review. Air Medical Journal 36 (2): 81–84. Dickinson, J., Heath, D., Gosney, J. and Williams, D. (1983, Sep). Altitude-­related deaths in seven trekkers in the Himalayas. Thorax 38 (9): 646–656. Dobzhansky, T. (1968). Adaptedness and fitness. In: R. Lewontin (ed.), Population Biology and Evolution, p. 111. Syracuse: Syracuse Univ. Press. Euler, U.S.V. and Liljestrand, G. (1946). Observations on the pulmonary arterial blood pressure in the cat. Acta Physiologica Scandinavica 12: 301–320. Ferrazzini, G., Maggiorini, M., Kriemler, S. et al. (1987, May 30). Successful treatment of acute mountain sickness with dexamethasone. British Medical Journal (Clinical Research Edition) 294 (6584): 1380–1382. Firth, P.G., Zheng, H., Windsor, J.S. et al. (2008, Dec 11). Mortality on Mount Everest, 1921-­2006: descriptive study. British Medical Journal 337: a2654. doi: 10.1136/bmj.a2654. Fishman, R.A. (1975). Brain edema. New England Journal of Medicine 293: 706–711. Ge, R.L. and Helun, G. (2001, Fall). Current concept of chronic mountain sickness: Pulmonary hypertension-­ related high-­ altitude heart disease. Wilderness & Environmental Medicine 12 (3): 190–194. Glaisher, J. (1862). Notes of effects experienced during recent balloon ascents. Lancet 2: 559–560. Hackett, P.H., Yarnell, P.R., Hill, R. et al. (1998). High-­altitude cerebral edema evaluated with magnetic resonance imaging: clinical correlation and pathophysiology. JAMA 280 (22): 1920–1925. Hackett, P.H. and Roach, R.C. (2004). High altitude cerebral edema. High Altitude Medicine & Biology 5: 136–146. Hakim, T.S., Michel, R.P., Minami, H. and Chang, H.K. (1983, May). Site of pulmonary hypoxic vasoconstriction studied with arterial and venous occlusion. Journal of Applied Physiology: Respiratory, Environmental and Exercise Physiology 54 (5): 1298–1302. Heath, D. and Williams, D.R. (1995). High-­Altitude Medicine and Pathology. Oxford: Oxford University Press. Himalayan Database (2020). https://www.himalayandatabase.com/ downloads.html, last accessed Dec. 12th 2020. Huey, R.B., Eguskitza, X. and Dillon, M. (2001). Mountaineering in thin air. Patterns of death and of weather at high altitude. Advances in Experimental Medicine and Biology 502: 225–236. Huey R.B., Salisbury R, Wang J.L., Mao M. (2007) Effects of age and gender on success and death of mountaineers on Mount Everest. Biol Lett. 3(5):498–500. doi:10.1098/rsbl.2007.0317 Hultgren, H.N., Wilson, R. and Kosek, J.C. (1997, Nov). Lung pathology in high-­ altitude pulmonary edema. Wilderness & Environmental Medicine 8 (4): 218–220. Imray, C., Booth, A., Wright, A. and Bradwell, A. (2011, Aug 15). Acute altitude illnesses. British Medical Journal 343: d4943. Jefferson, J.A., Escudero, E., Hurtado, M.E. et al. (2002, Feb 2). Excessive erythrocytosis, chronic mountain sickness, and serum cobalt levels. Lancet 359 (9304): 407–408. Jemsen, J.D. and Vincent, A.L. (2020, Jan-­). High altitude pulmonary edema [Updated 2020 Aug 16]. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing.

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Trübsbach, S.S., Pircher, I., Treml, B. et al. (2011, Feb). [Respiratory system at high altitude: pathophysiology and novel therapy options]. Wiener klinische Wochenschrift 123 (3–4): 67–77. Ulloa, N.A. and Cook, J. (2020 Jan-­). Altitude induced pulmonary hypertension. [Updated 2020 Sep  25]. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing;. Wilson, M.H., Newman, S. and Imray, C.H. (2009, Feb). The cerebral effects of ascent to high altitudes. Lancet Neurology 8 (2): 175–191. West, J.B., Schoene, R.B., Luks, A.M. and Milledge, J.S. (2013). High Altitude Medicine and Physiology, 5th edn. Boca Raton, USA: CRC Press. West, J.B. (2012, Dec). High-­altitude medicine. American Journal of Respiratory and Critical Care Medicine 186 (12): 1229–1237. West, J.B. (2017). Physiological effects of chronic hypoxia. New England Journal of Medicine 376: 1965–1971. Westensee, J., Rogé, I., Van Roo, J.D., Pesce, C., Batzli, S., Courtney, D.M., and Lazio, M.P. (2013). Mountaineering Fatalities on Aconcagua: 2001–2012. High Altitude Medicine & Biology 14 (3): 298–303. Wu, T., Ding, S., Liu, J. et al. (2006, Winter). Ataxia: An early indicator in high altitude cerebral edema. High Altitude Medicine & Biology 7 (4): 275–280. Xu, Y., Liu, Y., Liu, J. and Qian, G. (2014, Apr). Meta-­analysis of clinical efficacy of sildenafil, a phosphodiesterase type-­5 inhibitor on high altitude hypoxia and its complications. High Altitude Medicine & Biology 15( 1): 46–51.

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Schumacker, P.T. (2014). Cellular and molecular mechanisms of O2 sensing. In: E.R. Swenson and P. Bärtsch (eds.), High Altitude  – Human Adaptation to Hypoxia, pp. 1–22. Springer: New  York/ Heidelberg. Sime, F., Peñaloza, D., and Ruiz, L. (1971, Sep). Bradycardia, increased cardiac output, and reversal of pulmonary hypertension in altitude natives living at sea level. British Heart Journal 33 (5): 647–657. Singh, I., Khanna, P.K., Srivastava, M.C. et al. (1969, Jan 23). Acute mountain sickness. New England Journal of Medicine 280 (4): 175–184. Sui, G.J., Liu, Y.H., Cheng, X.S. et  al. (1988, Jun). Subacute infantile mountain sickness. Journal of Pathology 155 (2): 161–170. Sutton, J.R., Coates, G. and Houston, C.S. (1992). The Lake Louise Consensus on the definition and quantification of altitude illness. In: J.R. Sutton, G. Coates and C.S. Houston (eds.), Hypoxia and Mountain Medicine. Burlington: Queen City Printers. Taimeh, Z. (2019, Sep). Assessment and treatment of the failing right heart: Considerations for transplantation referral. Journal of Thoracic Disease 11 (Suppl 14): S1817–S1820. Teppema, L.J. and Berendsen, R.R. (2014). Control of breathing. In: E.R. Swenson and P. Bärtsch (eds.), High Altitude – Human Adaptation to Hypoxia, pp. 37–55. Springer: New York/Heidelberg. Thomas, D.J., du Boulay, G.H., Marshall, J. et al. (1977, Jul 23). Cerebral blood-­flow in polycythaemia. Lancet 2 (8030): 161–163.

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Injuries Due to Heat

depth of the tissue, and not only on the surface temperature. The deep tissue temperature depends on the thermal capacity and conductivity of the different layers of the tissue. With increasing radial penetration depth, the temperature within the tissue declines rapidly. Figure 23.1 shows the penetration of heat into tissues after scalding and contact burns for various durations and temperatures (scalding for 2 and 20 s, and contact burns at 70 °C for 10 and 60 s). The temperature–time profiles are shown for different depths of the tissues. In scalding, the tissue temperatures are higher than that in contact burns, although the time impacts are much lower. Extensive animal experiments and experiments with human volunteers on the effect of duration of heat impact, impacting temperature and degree of thermal damage of the skin, have been carried out (Moritz and Henriques 1947). In these experiments, exposure times and temperatures were varied systematically. Figure  23.2 shows the resulting temperature–time curve for temperatures between 44 and 70 °C (scald). The lowest water temperature causing scalds was 44 °C; however, to achieve irreversible damages, a heat impact of six hours was necessary. An exposure to 60 °C hot water for 3 s caused transient hyperaemia (first-­ degree damage). An exposure of 5 s caused complete necrosis of the epidermis. Similar threshold values identified for the impact of dry heat were shown to be considerably higher due to a different heat conductivity of the impacting media (Figure 23.3). Figure  23.4 shows the temperatures developing in the tracheobronchial system when hot air or steam affects the respiratory tract. In an experiment, dry air with a temperature of 350 °C affecting the respiratory tract caused ante-­mortem temperatures of 159–182 °C within the larynx and no temperature

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Burns are injuries primarily to the skin due to heat or high temperatures (Table 23.1). A burn may also be caused by electricity, chemicals, light and radiation. Scalds are injuries caused by hot liquids (water or oil) or gases (steam); in contrast to burns, hair is not damaged thermally in cases of scalding. Chemical burns, current accidents and contact burns (which are of special importance in forensic medicine) cause characteristic injuries. The following are of forensic relevance: • Accidental deaths within a fire • Deaths in cases of arson • Suicidal self-­immolation • Disposal of corpses • Homicides by burning Spilling hot liquids over someone or immersing someone in hot liquids is a case of scalding with special medicolegal interest. Scalds of a patient in a bathtub may also be of interest (breach of the duty of supervision).

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23.2  Burns from heat 23.2.1 Correlation between heat damage, temperature, time of exposure and depth of burns Local heat damage of the skin and mucosa mainly depends on the temperature and time of exposure. The degree of heat damage is principally dependent on the temperature developing in the Handbook of Forensic Medicine, Volume 2, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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PART III   TRAUMATOLOGY AND VIOLENT DEATH

Table 23.1  Classification of burn sources. Source

Examples

Heat

Chemical burns Scalds caused by hot damp air or steam or gases Hot materials and liquids Direct impact of flames

Radiation

Gadgets emitting heat (sun lamps, infrared lights, etc.) Radiation of atomic energy

Electric current

Contact with conductors Spark discharge (lightning)

°C

°C

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3

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N temperature 1 mm 2 mm 3 mm

60

2

1 mm 2 mm 3 mm 4 mm

after contact

50 40 30

1 minutes

1 2 minutes tissue heat penetration after 20 s impact of vapour

(b)

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temperature

70 after vaporisation

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after vaporisation

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Source: Madea and Schmidt (2015).

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Tissue heat penetration at 70°C dry heat, 10 s contact time

tissue heat penetration at 70°C dry heat, 60 s contact time

(c)

(d)

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Figure 23.1  Heat penetration into the tissue after scalding ((a) and (b)) and contact burns ((c) and (d)) of various durations (scalding 2 and 20 s and contact burns at 70 °C for 10 and 60 s). The temperature–time course is shown in the tissues at the depths of 0.5 mm (blue), 1 mm (yellow), 2 mm (green), 3 mm (red) and 4 mm (brown). Source: Original data from Allgöwer and Sigrist (1957) and Madea and Schmidt (2015).

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CHAPTER 23   INJURIES DUE TO HEAT

increase in the bifurcation (Moritz et  al. 1947). When dry air with a ­temperature of 550 °C was applied to the respiratory tract, temperatures of 267–327 °C were measured in the larynx and a temperature of 50 °C was measured in the bifurcation. Figure 23.4 also shows the temperatures developing within the larynx and bifurcation for fire and damp air (>100 °C). Damp

air causes a damaging temperature increase at clearly lower temperatures as compared to fire due to its higher heat conductivity. Because of this physical fact, in steam boiler explosions, not only is the skin damaged, but also the mucosa of the respiratory tract.

23.2.2  Burn wound classification °C

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There are a number of different burn wound classifications. The traditional system divides burns into first-­degree, second-­degree, third-­degree and fourth-­degree burns. This traditional system

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45

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10

103

100 1

2

5

104 s

10 30 60 1 2 3 4 5 minutes hours

100

0

2

4

6

8

10

I. 12

14 minutes

Progression of time temperature curves at transition to irreversible damage (curve I), to hyperaemic, (curve II) and to ischaemic damage, (curve III) (according to Moritz and Henriques (1947))

Figure 23.3  Temperature–time curve in cases of radiation with temperatures up to 500 °C. Curve I: threshold for reversible damage; curve II: threshold for hyperaemic damage; and curve III: threshold for ischaemic damage. Source: According to Moritz and Henriques (1947).

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Figure 23.2  Time–surface temperature thresholds at which cutaneous burning (scalding) occurs. The dashed line indicates the threshold at which irreversible epidermal injury of porcine skin occurs. The solid line indicates the threshold at which epidermal necrosis of porcine skin occurs. The results of critical experimental exposure of human skin specimen are indicated by points. Exposure time is given in seconds against the surface temperature of the skin. Source: Original data from Moritz and Henriques (1947).

II.

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Time–surface temperature thresholds at which cutaneous burning (scalding) occurs (according to Moritz and Henriques 1947)

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dry air

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350°C

267–327°C

159–182°C

50°C

0°C

FO

Fire

327–550°C

94–106°C

51–135°C

53–94°C

Figure 23.4  Temperatures in the larynx and bifurcation of the trachea in cases of inhalative trauma, for dry and damp air. Source: Original data from Moritz and Henriques (1947); modified according to Madea and Schmidt (2015).

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PART III   TRAUMATOLOGY AND VIOLENT DEATH

has today been replaced or combined with other systems reflecting the requirement of surgical intervention (superficial, superficial partial-­thickness and full-­thickness burns) (Figure 23.5 and Table 23.2).

First-­degree burns or superficial burns The characteristic finding of a first-­degree burn is erythema without blistering. The epidermis is intact, although some damaged cells might desquamate. The reddened skin is painful. Histologically, first-­degree burns show dilated, blood-­filled dermal vessels. Complete healing takes 8–10 days.

s­uperficial (papillary) dermis and may also involve the deep (reticular) dermis layer. The blisters are filled with serous liquid in the layer between the epidermis and dermis. In second-­degree superficial burns, only the epidermal and granular layers are destroyed whereas the basal cell layer remains intact. Healing takes place within two to three weeks. In deep, second-­degree burns, the epidermis, including the basal cell layer, is completely destroyed, whereas epidermal appendages (hair and perspiratory glands) stay intact and are the basis for multicentre re-­epithelialisation. The healing process is delayed and can last for more than three weeks.

O

N

Third-­degree burns are characterised by a coagulation necrosis of the entire epidermis and dermis with deletion of the epidermal appendages. The skin is dry, white and leather-­like. The sense of pain is destroyed.

SE

This type of burn is characterised by blistering. The skin is damp, reddened and painful. Second-­ degree burns involve the

LY

Third-­degree burns

Second-­degree burns or partial-­or full-­thickness burns

degree

U

layer

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epidermis

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IIb.

III.

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subcutis

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dermis

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IV.

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Figure 23.5  Degree of burns and dermal layers (Madea and Schmidt (2015)).

Table 23.2(a)  Degree of burn and main symptoms. Degree

Depth

Colour and appearance

Tissue structure

Capillary filling

Sense of pain

Healing

First degree

Superficially epidermal

Red

Normal

+

+

5–10 days

Second degree

Superficially dermal

Red, blisters

Oedematous

+

+

12–20 days, few scars

Deeply dermal

Pink or white, blisters

Thickened

+/-­

+/-­

20–60 days, scarring

Third degree

Transdermal

White, brown

Leathery

-­

-­

No spontaneous healing

Fourth degree

Subcutaneous

Charred

Skin is missing

-­

-­

No spontaneous healing

Source: Madea and Schmidt (2015).

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CHAPTER 23   INJURIES DUE TO HEAT

Table 23.2(b)  Burn wound classification. Degree of burn Superficial partial thickness (second degree)

Full thickness (third degree)

Full thickness (fourth degree)

Brief exposure to flash, flame or hot liquids

Intense radiant energy; scalding liquids, semi-­liquids (e.g. tar) or solids; flame

Prolonged contact with flame, scalding liquids; steam; hot objects; chemicals; electric current

Electric current; prolonged contact with flame (e.g. unconscious victim)

Limited damage to epithelium; skin intact

Epidermis destroyed; minimal damage to superficial layers of dermis; epidermal appendages intact

Epidermis destroyed; underlying dermis damaged; some epidermal appendages remain intact

Epidermis, dermis and epidermal appendages destroyed; injury through dermis

Epidermis, dermis and epidermal appendages destroyed; injury involves connective tissue, muscle and possibly bone

Wound characteristics

Erythematous; hypersensitive; no blister formation

Moist and weepy, pink or red, blisters, blanching, hypersensitive

Pale; decreased moistness; blanching absent or prolonged; intact sensation to deep pressure but not to pinprick

Dry, leather-­like; pale, mottled brown or red; thrombosed vessels visible; insensate

Dry; charred, mottled brown, white or red; no sensation; limited or no movement of involved extremities or digits

Treatment course

Complete healing within five days without scarring

Complete healing within 21 days with minimal or no scarring

Prolonged healing (often longer than 21 days); may require skin grafting to achieve complete healing with better functional outcome

Requires skin grafting

Requires skin grafting; amputation of involved extremities or digits likely

TR IB

Fourth-­degree burns

N

In fourth-­degree burns, the skin and subcutaneous fatty tissue as well as muscles and bones are affected. The tissue is charred.

O

23.2.3  Burnt surface area

R

C

The total body surface area (TBSA) of burns can be calculated by using the ‘rule of nines’ (Figure 23.6). It must be remembered that the head of an infant is proportionally bigger than that of an adult. The TBSA is of relevance for the prognosis of burnt areas over the body, which may be of importance in reconstructing the case (e.g. to ascertain what position the body was in at the beginning of a fire). The degree of burn (depth of burn) and the victim’s age are also important (Table 23.3). With increasing age (>40 years), the prognosis deteriorates, but newborns and infants are also especially endangered. If age and burn surface area of a second-­ degree or third-­degree burn (according to Burn Index) add up to 100, the chance of survival is 50% at best even with optimal therapy. According to clinical experiences, a Burn Index of 120, survival is improbable. Even today, adult patients with deep

FO

N

O

SE

U

Depth of injury

R

Prolonged ultraviolet light exposure; brief exposure to hot liquids

TO

Cause of injury

U

First degree

LY

Deep partial thickness (second degree)

Feature

burns of more than 30% of the body surface area have a poor prognosis, and those with burns of more than 50% of the body surface area survive in exceptional cases only. In cases of an additional inhalation trauma (indicated by burns of the face or the perioral region), the prognosis worsens.

23.2.4  Post-­mortem heat damages The destruction of a corpse within a fire is caused by the direct impact of heat and by creation of steam with extracellular, intracellular and transcellular altering from the liquid phase to the gaseous phase. The bodily fluids commence to boil (boiling point of water, i.e. approximately 100 °C) and vaporise (Figure  23.7). With continuing temperature rise, the steam expands further. The increase in body temperature results in heat fixation of the tissues, as well as temporary inflation (similar to putrefaction) caused by vaporisation. Bodily fluids and lymph are boiled and evaporated; the lips become swollen; the tongue protrudes; and liquids leak from the mouth and nose. Subsequently, the skin and bones shrink. Morphological findings in burnt bodies may cover a broad spectrum from nearly intact bodies with just some soot deposition on the skin to severely burnt bodies with extensive charring to skeletal remains without any soft tissues or total incineration.

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PART III   TRAUMATOLOGY AND VIOLENT DEATH

Adult body part

% of total body surface

arm head neck leg anterior trunk exterior trunk

9% 9% 1% 18% 18% 18%

9%

age

0–1

0–4

5–9

10–14

15

1%

A-1/2 head

9 1/2%

8 1/2%

6 1/2%

5 1/2%

4 1/2%

B-1/2 thigh

2 3/4%

3 1/4%

4%

4 1/4%

4 1/2%

C-1/2 leg

2 1/2%

2 1/2%

2 3/4%

3%

3 1/4%

18% 9%

9% 18%

A

3½%

A

Adult

1%

18% 18%

1%

2%

2%

2%

2% 13%

LY

13% 1½%

1½%

1½%

9%

18%

14%

O

C 3½%

4¾% B

4¾% B

C 3½%

C 3½%

1¾% 1¾%

9% 18% 14% 18% 18%

1¾% 1¾%

Estimation of the burned body surface according to the method by Lund and Browder

TR IB

U

14%

arm head and neck leg anterior trunk exterior trunk

C 3½%

1½%

R

18%

4¾% B

2½%

TO

9%

4¾% B

1½%

1½%

U

% of total body surface

body part

1½%

SE

Child

18%

1%

N

2½%

1½%

3½%

N

Figure 23.6  Rule of nines for the estimation of the burnt body surface: arm (9%), head (9%), neck 1%), leg (18%), anterior trunk (18%) and exterior trunk (18%). These rates vary, depending on the age of the child or young adult, as shown in this figure. Source: Lund and Browder.

O

Table 23.3  Mortality probability for various combinations of ages and burnt body surface (1 = 100%).

5–9

68+ 63–67 58–62 53–57 48–52 43–47 38–42 33–37 28–32 23–27 18–22 13–17 8–12 3–7 0–2

1 1 1 0.9 0.8 0.7 0.6 0.5 0.4 0.2 0.1 0 0 0 0

1 1 1 0.9 0.8 0.7 0.5 0.4 0.3 0.2 0.1 0 0 0 0

10–14

R

0–4

C

Age in years Burnt body surface in %

FO

1 1 0.9 0.8 0.7 0.6 0.5 0.3 0.2 0.1 0 0 0 0 0

15–19

20–24

25–29

30–34

35–39

40–44

45–49

50–54

55–59

60–64

65+

1 0.7 0.8 0.8 0.7 0.5 0.4 0.3 0.2 0.1 0 0 0 0 0

1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 0 0 0 0

1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 0 0 0 0

1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0.1 0 0 0 0

1 1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 0 0 0

1 1 1 0.8 0.8 0.7 0.5 0.4 0.3 0.2 0.1 0 0 0 0

1 1 1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 0 0

1 1 1 1 0.9 0.8 0.7 0.6 0.5 0.3 0.2 0.1 0 0 0

1 1 1 1 1 0.9 0.8 0.7 0.6 0.5 0.3 0.2 0.1 0 0

1 1 1 1 1 1 0.9 0.9 0.7 0.6 0.5 0.3 0.2 0.1 0

1 1 1 1 1 1 1 1 0.9 0.8 0.7 0.5 0.3 0.2 0.1

Source: After Herndon (1997) and Allgöwer et al. (1998).

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CHAPTER 23   INJURIES DUE TO HEAT

200

temperature at surface

b) a) 150

°C

temperature curve in the underlying epidermis a) specimen without fat tissue b) specimen with fat tissue (1,2 cm)

100

LY

temperature in the fat tissue (1,2 cm)

The appearance of a burnt body depends on temperature, duration of impact and type of heat transmission. When a person dies within a fire, heat transmission does not stop with death and post-­mortem heat effects may be dominant due to: • Burns of the exposed tissues with consumption of the tissue by fire • Changes in the content and distribution of tissue fluids and vaporisation of bodily fluids • Fixation of tissue due to fluid loss • Shrinking processes also due to fluid loss As a consequence of the shrinking process of tissues, the skin is tight and may split; petechial haemorrhages of the conjunctivae may be found. The loss of fluids causes a heat fixation of the skin and internal organs. The skin becomes brownish and leather-­ like; induration of the internal organs may also be observed. A pugilistic attitude is common, with the arms abducted in the shoulder joints and flexed in the elbow joint and the legs abducted in the hip joint and flexed in the knee joint. This position, which is more usually seen on supine bodies, is caused by heat shrinkage of the muscles and tendons. As there is a predominance of flexor muscles, it results in a flexion of the joints. As the bodily fluids vaporise and shrink, a protrusion of the tongue and lips is caused. In cases of fire, therefore, the teeth are protected for a comparatively long time and even in severely charred bodies odontological identification may be possible. However, after shrinkage of the lips, the teeth are exposed to fire and may also be destroyed. Characteristic findings of post-­ mortem heat damage (Table 23.4) include:

30

40

50

t Temperature curves at the surface and in the underlying epidermis (using mean values) of all examined body parts

U

TO

Figure 23.7  Temperature–time curve on the surface of the skin and subepidermally. On the surface of the skin, the surrounding temperature is quickly achieved; however, subepidermally, there is a plateau at 100 °C (vaporisation of tissue water with swelling of lips and protruding tongue). Source: Reproduced with permission from Madea and Schmidt (2015); original data from Sachs (1973).

SE

20

U

10

R

0 0

O

N

50

TR IB

Table 23.4  Effects of heat on the body and related external and internal findings. External findings

Internal findings

Burns

Burns of skin

Burns and consumption of internal organs and bones

Singeing of hair

Oedema, mucosal bleeding and detachment of the mucosa of airways

O

N

Effects of heat

Consumption by fire Skin blisters

C

Changes of content

FO

R

Distribution of tissue fluid

Heat fixation

Vaporisation of bodily fluids Rupture of abdominal wall with prolapse of intestinal loops Leakage of fluid from mouth and nose Heat haematoma Accumulations of fat in body cavities, vessels or heart

Leather-­like, brownish fixation of the skin

Induration of internal organs and muscles Fragmentation of erythrocytes

Shrinking of tissue

Tightening of skin

Shrinking of organs

Splitting of skin

“Puppet organs”

Protrusion of tongue Petechial haemorrhages of neck and head Pugilistic attitude Source: Reproduced with permission from Bohnert (2004).

614

Figure 23.9  Pugilistic attitude of a burnt body. Source: From the collective of the Insitute (own research).

U

Figure 23.8  Splitting of skin. Source: From the collective of the Insitute (own research).

TO

R

U

SE

O

N

LY

PART III   TRAUMATOLOGY AND VIOLENT DEATH

1. Splitting of the skin

FO

R

C

O

N

TR IB

Shrinkage can cause splitting of the skin, followed by bursting of the abdominal wall with opening of the cavity. Since these lacerations may be quite straight and smooth, they should not be mistaken for vital incised wounds (Figure 23.8). 2. Pugilistic or boxer’s attitude Heat-­ related flexion with shrinking of muscles and fibres causes a characteristic appearance of the corpse with abduction of the shoulder joints, flexion of the elbows and wrist joints, and extension of the legs and pes equinus (Figure 23.9). 3. Heat haematoma A direct impact of flames on the skull finally leads to an epidural accumulation of brick-­red or brownish, brittle, dry or mud-­like blood that might also be liquid or semiliquid. This post-­mortem epidural extravasate originates from an extrusion of blood in the skull, which is compressed against the dura mater, as well as from shrinking and ablation of the dura mater of the inner surface of the skull (Figure 23.10). When recovering a corpse from a fire scene, one of the main tasks is – besides clarification of identity, underlying diseases and cause of death – to find out whether the dead person had been alive when fire was starting or whether the person was already dead. Even if soft tissues and muscles are largely charred, these questions can often be answered because the inner organs are usually relatively well preserved (heat fixation caused by fluid loss) due to the radial temperature decline (towards the body core).

Vital signs and reactions 1. Soot inhalation

The inhalation of soot particles into the bifurcation of the bronchial tree is a definite vital reaction. General autopsy findings include soot in the respiratory tract including the fine bronchi and abundant mucous secretion (Figures  23.11 and 23.12). 2. Soot swallow Simultaneously, soot might be swallowed and particles be found in the stomach or upper part of the small intestine. 3. Smoke inhalation If someone is burnt alive, positive carbon monoxide (CO) findings are expected. Nevertheless, the arterial carboxyhaemoglobin (CO-­Hb) concentration depends on the age of the individual, as well as on the circumstances and site of the fire. In older victims, the concentration is often significantly lower than that in younger victims. In cases of fires in rooms, negative CO findings might occur despite closed doors and soot inhalation. Similarly, fires outdoors (often in suicidal self-­ immolation) may frequently result in low or negative CO-­Hb findings. Typical autopsy findings of CO poisoning are light-­ red-­ coloured blood and salmon-­ pink-­ coloured muscles. However, light-­red-­coloured blood may also be a consequence of the heat impact without a simultaneous increase of the CO-­ Hb concentration. When nitrogen-­containing polymers (of

615

TO

R

U

SE

O

N

LY

CHAPTER 23   INJURIES DUE TO HEAT

Figure 23.12  Histology of soot inhalation: soot mixed with mucus and desquamated epithelia; due to inhalative trauma congestion of submucous vessels. Source: From the collective of the Insitute (own research).

FO

R

C

O

N

TR IB

U

Figure 23.10  Heat haematoma showing epidermal haematoma over the right hemisphere, with a view into the skull from the occiput. Source: Courtesy of Professor G. Dotzauer.

Figure 23.11  Soot inhalation into the larynx (a) and trachea (b). Source: From the collective of the Insitute (own research).

616

PART III   TRAUMATOLOGY AND VIOLENT DEATH

Table 23.5  Fire gases and their effects. Set free during combustion or carbonisation of:

Gas

Symptoms of toxicity

Lethal concentration in 10 minutes (ppm)

Toxic gas Wool, silk, polyacrylonitriles, nylon, polyurethanes from mattresses, upholstered furniture, curtains, carpets, cars, airplanes and paper in various degrees

Respiratory poison, rapidly fatal

180

Carbon dioxide

Within open fire and smouldering, complete combustion of all organic substances (heavier than air)

Mucous membrane irritation, dyspnoea, convulsions and apnoea

80 000

Carbon monoxide

Incomplete combustion of all organic substances (lighter than air)

Haemotoxin, nausea, headaches, unconsciousness and apnoea

Acrolein

Polyolefins (overheating of edible fat) and cellulose products under low temperature (500

Isocyanates

Few from polyurethanes

Nitrous gases

Small amounts from textiles, larger amounts from cellulose nitrate and celluloid, and fertiliser

Hydrochloric acid

Sulphur dioxide

LY

Hydrocyanic acid

N

1000–2000

TO

R

U

SE

O

Lung irritants

100

Severe lung irritation after latency time, and may lead to immediate death or long-­term damage

>200

Cable insulation materials, such as polyvinyl chloride (PVC), chlorinated acrylics and hardened metals

Eye cauterisation, severe lung irritation and intensity of poisoning of bound hydrochloric acid higher than that of the appropriate quantity in the gaseous state

500

Sulphur-­containing compounds and their oxidation products

Strong irritant gas and incompatible in much smaller doses than the fatal ones

50–100

O

N

TR IB

U

Strong lung irritation

C

Source: Reproduced with permission from Daunderer (1982).

FO

R

natural or synthetic origin) are combusted, either nitric oxide or hydrogen cyanide (HCN) is produced depending on the amount of oxygen available. Fatal inhalatory cyanide intoxication may result after a longer-­lasting cyanide exposure if the HCN concentration is around 90 ppm, and after short exposure if the HCN concentration in the air has reached 180– 270  ppm. Fire gases and their effects are summarised in Table 23.5.

and the long-­term survivors have the highest expression levels in all cases. Hsp27 and Hsp70 are also expressed in renal tubules and renal vessels in fire-­related fatalities (Figure 23.13). A positive Hsp expression can also be observed in cardiovascular tissue. The immunohistochemical investigation of an Hsp expression can support the proof of vitality in cases of death related to fire.

Heat shock protein (Hsp) expression

23.2.5  Thermal injuries of the respiratory tract (inhalation trauma)

In pulmonary tissue, Hsp27 and Hsp70 are expressed within seconds and minutes after heat exposure. In fire-­related cases of pulmonary tissue, Hsp27 and Hsp70 expression in the bronchial tubes showed a similar expression pattern in all short-­term survivors. With increasing survival time, Hsp70 expression greatly increased,

Characteristic injuries of the respiratory tract are caused by inhalation of hot air. In cases of immediate death, a frazzled and coagulation necrosis of the membrane mucosa can be found in the nasopharynx, larynx and trachea. The ciliated epithelium of the trachea shows a cell and core elongation aligned towards the

617

N

LY

CHAPTER 23   INJURIES DUE TO HEAT

(b)

(d)

N

(c)

TR IB

U

TO

R

U

SE

O

(a)

R

C

O

Figure 23.13  (a) Pre-­mortem heat exposure with short survival time (fire death). Kidney: Glomeruli and tubular epithelium clearly positive for Hsp70 stain with a magnification of 200x. (b) Pre-­mortem heat exposure with short survival time (fire death). Kidney: Glomeruli and tubular epithelium positive for Hsp70 stain with a magnification of 400x (c); heat shock protein expression (fire death) of the trachea (d); heat shock protein expression (fire death) of the bronchial tubes. Source: From the collective of the Insitute (own research).

FO

lumen. Vital reactions are: (i) severe oedema of the mucosa with lymphatic ectasia; (ii) capillary and venous hyperaemia with microhaemorrhages; and (iii) intra-­ alveolar, interstitial septal and perivascular pulmonary oedema.

23.3  Causes of death Causes of death due to heat and fire can generally be divided into immediate death at the site of the fire and late death that occurs as a consequence of the fire. • Immediate deaths are caused by: ○○ Smoke inhalation ○○ Local injuries to the skin

• Subsequent deaths are due to: ○○ ○○

Burn disease (renal function impairment) Infectious complications of the tracheobronchial system (in cases of inhalation trauma) or of the skin lesions

23.3.1 Death caused by heat and flames without vital signs The frequency of these kinds of death is given in the literature with 3–10% of all deaths caused by burns. If natural death or homicides are excluded, the following causal factors have to be taken into consideration:

618

PART III   TRAUMATOLOGY AND VIOLENT DEATH

1. Cyanide intoxication: Dependent on the material burnt, cya-

15 ­minutes; the shin bones are charred after 25 minutes; the ribs and skull are charred after 20 minutes; and the thighs and shin bones are completely charred after 35 minutes. Extensive observations on the destruction of the skull and teeth have been carried out during cremations. Further observations on the time course of the destruction of whole human bodies during cremations at temperatures between 670 and 810°C have been published by Bohnert et al. (1998). The bodies showed the pugilistic attitude after about 10  minutes; after 20  minutes, the calvaria were free from any soft tissue and heat fractures of the tabula externa could be noticed. Body cavities became visible after approximately 30 minutes so that the organs were exposed, and 40 minutes after cremation had started, the internal organs were severely shrunken and showed a net, sponge-­like structure. After about 50  minutes, the extremities were destroyed to an extent leaving only the torso that broke apart after one and a half hours. The complete incineration of a human body took about two to three hours. At the end of a cremation with temperatures ranging from 800 to 1200 °C, only ashes weighing 1–3 kg were left. The effects of fire on the skull and trunk and extremities in different observational studies are shown in Tables 23.6 and 23.7. Temperatures during cremation are similar to those reached in house fires. Therefore, these observational studies can be used as the basis for the estimation of the duration of a fire in correlation to the charring of a body. There is agreement in the literature that it is very difficult to completely destroy a body by fire (e.g. attempting to dispose of the body of a victim of a crime). Severely burnt bodies are best classified by the Crow-­Glassman Scale (Table 23.8).

Table 23.6  Effects of fire on the skull. Temperature

O

SE

U

TR IB

U

TO

Any particular temperature causes a defined progression of incineration of a corpse. Thus, the duration of a fire can be derived from the degree of burning. Information on the duration of a fire might be of special importance if alibis or the validity of a suspect’s perception of a fire has to be checked. A corpse is fully burnt except for small bone pieces in modern crematoria after one to two hours. Only in rare cases, the correlation between the duration of a fire and the degree of charring gains forensic importance and only few reports on this issue can be found in the literature. According to Richards (1977), the arms are badly charred in a temperature of 680 °C after 10 minutes; the legs are badly charred after 14  minutes; the face and arm bones are charred after

R

23.3.2  Duration of a fire and associated degree of charring of a corpse

N

LY

nide intoxication can develop very quickly and, thus, increased CO-­Hb concentrations might not be found. 2. Flashfire: Death may be due to apnoea from a laryngeal spasm, bronchial spasm, vagal reflex or inhalative heat shock. 3. Oxygen deficiency: Oxygen depletion by the fire source may lead to death. 4. Heat shock: This involves reallocation of the circulating blood volume due to skin exposure to heat. 5. Heat rigour: Functional impairment of breathing is caused by sudden heat rigour of the thorax.

1000–1100 °C (Günther and Schmidt 1953)

8–10

Soft tissues of the face charred

20–25

O

C

670–810 °C (Bohnert et al. 1998) Skull cap free of soft tissue Soft tissues of the face charred

Forehead and vertex free of soft tissues, protruding facial bones and calcinated

Bones of face showing Skull showing

R

20

680 °C (Richards 1977)

FO

13–16

N

Time (min)

Sparse soft tissues remain in the face Heat fractures of the skull cap

Severe shrinkage of soft tissues at the skull, calvaria broken, brain superficially charred and destruction of prominent parts of the facial skull

30

Tabula externa of the calvaria crumbling

40

Brain showing, bones of the face begin to disintegrate, bones of the face largely destroyed and base of skull showing

45–75

Base of skull still intact and head sometimes separated from the trunk

Source: Reproduced with permission from Bohnert et al. (1998).

619

CHAPTER 23   INJURIES DUE TO HEAT

Table 23.7  Effects of fire on the trunk and extremities. Temperature Body region

Time (min)

680 °C (Richards 1977)

670– 810 °C (Bohnert et al. 1998)

Thorax

20

Ribs showing

Thorax muscles charred, and ribs and sternum showing

Abdomen

30

Thoracic and abdominal cavities exposed Organs blackened and shrunken

Legs

Shrunken, charred organs with bumpy surface

50

Organs largely consumed by the fire Arms badly charred

15

Bones of arms showing

Pugilistic attitude

Hands largely destroyed, ulna and radius partially showing, hands and distal forearms burnt away, forearms completely consumed and upper arms largely free of soft tissue

50

Arms burnt away

14

O

N

20

Legs badly charred

20

Carbonisation of muscles Shin bones showing

U

25

Tibia and distal femur free of soft tissue Complete thigh and shins bone showing

Calcinated stumps of the thighs

TR IB

Source: Reproduced with permission from Bohnert et al. (1998).

U

50

TO

35

R

30

Table 23.8  Crow-­Glassman Scale of burn-­related destruction of the corpse. Second-­degree burns, sometimes with singeing of the hair; visual identification possible

Level 2

Burns of varying severity, sometimes with thermal destruction/amputation of ears, genitals, hands or feet; visual identification may still be possible

Level 3

Consumption by the fire with partial amputation of arms and/or legs; cerebral cranium intact

Level 4

Bony lesions of the cerebral cranium; residual extremities still present

R

C

O

N

Level 1

Fragmented skeletal remains without soft tissue

FO

Level 5

LY

10

SE

Arms

40

Source: After Bohnert (2004).

23.3.3  Checklist for the examination of burnt bodies Adhesions of the scene of fire on the body 1. Distribution, extension and layer thickness of soot adhesions 2. Traces of oxidising agents (e.g. fire starter or accelerants and inflammable liquids)

3. Type, amount and distribution of incendiaries and hot materi-

als, especially highly heated parts of metals, melted glass or aluminium that might show a possible position of the victim.

Traces of melting construction material (e.g. ceiling liners) and burning materials, such as flammable liquids in suicides or fire accelerants in accidents 4. Oddments of objects that fell on the body, such as the ­construction material of collapsing buildings 5. Material adhesion on the hands

Clothing 1. Localisation, extension, form and direction of thermal damage (consider clothing layers)

2. Damages not caused by fire 3. If a reliable identification of a burnt corpse is not possible,

look for individual signs like size of garments, material, type, colours and labels, content of pockets (e.g. personal ­documents and pieces of jewellery) and watch (time)

Post-­mortem external examination 1. Post-­mortem lividity, are nailbeds coloured light red? 2. Thermal damage of scalp hair, eyebrows and hair of the beard 3. Crow’s feet 4. Soot in the nostrils and oral cavity 5. Localisation, surface area, depth and distribution pattern of

burns (congruity with clothes?); transition region towards skin that is not thermally damaged

620

PART III   TRAUMATOLOGY AND VIOLENT DEATH

LY

N

Case example 2

After a fire in the washhouse of a family home, the corpse of the house owner was found in the adjacent stairways in the cellar. During autopsy, second-­to third-­degree burns of approximately 90% of the body surface were found. Additionally, the examiners found gagging, captivation of the hands and lower legs, soot inhalation and soot swallowing. The CO-­Hb concentration was 6%. They also found an odour of petrol and wet textile remains. Petrol was detected in blood, lung and brain samples. Two days later, the deceased’s wife was arrested when she tried to leave Germany. She had second-­to third-­degree burns at the extensor sides of both forearms, both front sides of the lower legs and at the dorsum of the right foot. Further criminal investigations revealed that the woman was abnormally jealous and had planned (after an argument with her husband) to burn her husband. She used pepper spray to make him defenceless, and then captivated and gagged him. Subsequently, she soused him with petrol and burnt him.

U

TO

as swallowed soot, inhalation of soot and thermal damage of the epithelium of the respiratory tract due to an inhalative trauma. 2. Post-­mortem findings of heat and flame impacts, such as heat haematoma, joint separation caused by heat, fractures of bones and calcination in the area of burning impacts. 3. Injuries not caused by burning. 4. Pre-­existing diseases that might be a competing cause of death or decrease sensitivity to burns. 5. In case of no or only doubtful identification, consider individual characteristics, such as surgical ablation of organs, internal sexual characteristics, findings related to osteology and other findings (e.g. degenerative diseases) facilitating an age estimation.

O

1. Findings related to vitality during heat and flame impact, such

An almost fully charred corpse of a man was found in a totally burnt-­out car that was detected in a forest. The man could be identified as the owner of the car by characteristic osteosynthetic material. Autopsy revealed a blasted skull (induced by heat) and loss of the soft tissues of the cervix, thorax and extremities due to shrinkage. The remaining parts of the trachea and the bronchi were filled with a brownish crumbly substance that was histologically defined as blood coagulated due to heat impact. The blood alcohol concentration was 81 mg/100 ml and the test for CO-­Hb was negative. Police investigations were accordingly intensified, and a fire accelerant was detected in the car. Additionally, there were blood stains on the forest soil close to the car as well as in the apartment of the dead male. The blood stains that were found in the apartment indicated that a fight had taken place. The following circumstantial evidence lawsuit found the deceased’s wife and her lover guilty of having killed him by force against the neck and subsequently having simulated his suicide in order to start a life as a couple.

SE

Autopsy

Case example 1

U

ration, intracranial haematoma with signs of local heat or flame impact or adhesions of the scene of fire 7. Findings of post-­mortem heat impact, such as charring or a pugilistic attitude of the extremities 8. Injuries not caused by heat or flames 9. In cases of doubtful identification, consider individual characteristics, like height and weight (estimated), hair colour, colour of eyes, surgical scars, external sexual characteristics, amputations, piercings and tattoos.

R

6. Correlation of lacerations, bony fissures/fractures, joint sepa-

TR IB

Additional examination

1. Histological examinations especially of the local heat damages

FO

R

C

O

N

regarding vitality and depth of damage, and damage of the respiratory tract and the lungs regarding an inhalative trauma. 2. (Optionally) examination of ‘fractures’ by stereomicroscopy for differentiation between fractures caused mechanically and those caused thermally. 3. Toxicological examination of inhalation gas constituents (CO, hydrocyanic acid, hydrogen chloride and sulphur dioxide) and of central nervous system depressants and narcotics, and examination of blood regarding alcohol intoxication (reduced ability to act when the fire started). 4. Hair analysis for the verification of a possible chronic drug abuse (which might also help in individualisation).

23.3.4  Criminological aspects Regarding criminology, the following constellations have to be distinguished: • Accidents, for example falling asleep with a burning candle or cigarette, or children playing with fire. • Suicide, which is sometimes epidemic-­like. • Homicide, either by burning (which is rare) or by burning after another crime in order to conceal it.

Case example 3 A 49-­year-­old construction worker was working at a trash dump site. He was crushing stones and cement blocks with the excavator bucket of a large excavator. Hidden amongst the stones was a 1.8 t British air mine from World War II (type HC 4000 LB MK IV, with a length of 2.8 m and a diameter of 76 cm). This heavy bomb detonated in response to the construction work with an explosive charge of 1.3 t. The detonation and the resulting shock wave were far reaching, with bomb fragments and damaged windows found in buildings at a distance of 1.2 km from the explosion site. The excavator was completely burnt out. Fragments of the construction worker body were found inside the excavator cab. Small body parts and organ pieces were also found up to 7 m away from the excavator. The man’s head and thorax were never recovered. The autopsy showed no macroscopic signs of vitality. Chemical–toxicological analyses were negative and the concentration of carbon monoxide was within the physiological range. Immunohistochemistry of renal tissues that were recovered showed moderate Hsp70 expression in the vascular endothelium and renal tubules. This positive Hsp expression can be considered as a phenomenon of supravitality.

621

CHAPTER 23   INJURIES DUE TO HEAT

23.4 Scalding

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Cases of scalding are often caused by a breach of duty of supervision or custodial care (bathing children or persons in need of care with water being too hot). Usually, the accused try to exonerate themselves by admitting that the injured person was left alone for a very short period of time only and that the water had only a slightly raised temperature. Such defence pleadings can be verified by using a temperature–time curve for the causation of scalding (see Figure 23.2). Scalding is of special importance in clinical forensic medicine since 5–14% of all children’s scalds are thought to be caused by abuse (Figure  23.16). For immersion scalds, this rate is even higher with 12–55%. Differential diagnoses between accidental and non-­accidental scalds are given in Table  23.9. Reasons for clinical suspicion include:

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The body of a 43-­year-­old man was found dead after a house fire. According to his wife, the man and the wife had been in a dispute, and he had threatened to shoot himself, whereupon she left the house and went to a friend. The autopsy revealed third-­degree burns (Figure 23.14). Six gunshot wounds were found, two of which in the thorax were associated with injuries to the lungs and heart. Pale mucous membranes and organs as well as shock kidneys were found as signs of high blood loss. Signs of a vital burn were not found. There was no evidence of soot aspiration, soot ingestion or heat damage to the mucosa of the upper respiratory tract. The concentration of carbon monoxide was within the normal range. Immunohistochemistry for Hsp27 and Hsp70 in heart, lung and kidney tissues was negative (Figure 23.15). The wife finally admitted that she had shot her husband and then set the house on fire. The cause of death was fatal haemorrhage due to gunshot wounds. The body was burnt post-­mortem.

N

Case example 4

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N

Figure 23.14  Part of the corpse with burns.

Figure 23.16  Typical immersion injury showing a sharp margin between healthy and injured skin corresponding to the water level. Source: Chef. Vermutlich Kollektiv des Instituts.

Table 23.9  Differential diagnosis of accidental and non-­accidental scalding. Accidental scalds

Immersion

Unsteady pattern of the injury (scalds of different depths)

Steady depth of scald

Diffuse limitation between scalds and Sharp margin between healthy and affected skin (water healthy skin (at the borders, the level might be reproduced scalds are rather less pronounced on skin like a map) due to cooling of the water) Extremities show rather scalds in the In case of immersion of form of splashes (e.g. on feet) extremities: scalds in the form of gloves or socks

Figure 23.15  Kidney tissues. Negative Hsp27 staining in the tubules, glomeruli and vessels. Magnification 400x. Source: Chef. Vermutlich Kollektiv des Instituts.

In case of scald of the thorax, the scald In case of immersion of the face, there are no signs of is usually configured like an arrow running water (water running down the body) Source: After Madea and Schmidt (2015).

622

PART III   TRAUMATOLOGY AND VIOLENT DEATH

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Human beings are homeothermic with a body core temperature of approximately 37 °C (±0.5 °C). The body core temperature is regulated and kept constant by autologous thermoregulation. Thus, heat absorption and heat production on the one hand and heat output on the other hand have to be well balanced. Heat production depends on the energy exchange of the organism. Without activity, 50% of heat production originates from the internal organs and 20% from the muscles. With physical activity,

LY

23.5  Generalised heat damage and hyperthermia

N

•

heat production might considerably increase these rates. There are three main mechanisms causing heat loss: • Radiation • Conduction and convection • Evaporation by the skin and lungs An imbalance of heat production and heat loss causes systematic dysfunctions of the temperature balance (e.g. loss of salt, malfunction of circulation, pathological increase of body core temperature and heat radiation affecting the skull with local hyperthermia of the brain). These systematic dysfunctions (Table 23.10) are: • Heat cramps • Heat collapse (exhaustion) • Heatstroke • Sunstroke Typical scenarios resulting in death from high ambient temperatures include physical illness in elderly people during heatwaves and children left in cars. Infants left unattended in motor vehicles are at special risk of heatstroke because intravehicular temperatures can increase quickly to lethal values. Definitions of terms are given in Table 23.11.

O

• • •

injury. An inadequate or inconsistent case history. A denial that the injury was caused by heat. The injured person itself or a sibling is accused of having caused the injuries. The pain is trivialised or denied (by the parents or child).

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• A discrepancy between anamnestic information and type of

Table 23.10  Causes and symptoms of systemic heat injuries. Heat exhaustion

Heatstroke

Sunstroke

Cause

Heavy labour in radiating heat Dehydration Loss of sodium chloride

Malfunction of circulation Vasodilatation Dehydrogenation Subsidence of blood into legs (when upright) Decrease/drop of heart time volume and blood pressure

Abnormally high heat input and restricted heat output Pathological increase of body temperature up to 43 °C High humidity intensifies the processes

Direct impact of sun radiation on the bare head

Symptoms

Muscle cramps Exhaustion Nausea Decrease of urine production

Skin reddened, sweaty and dry mucosa, tantalising thirst, headache, feeling of dizziness, scotoma, tinnitus, paraesthesia and circulatory shock

‘Read stadium’ with red, dry skin as long as circulatory regulation is intact, then ‘grey stadium’ with myogenic heart insufficiency, cerebral symptoms with delirium or coma, unconsciousness, epileptic cramps and meningitis symptoms

Meningeal irritations Meningeal haemorrhages Cerebral purpura

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N

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Heat cramps

C

Source: Madea and Schmidt (2015).

Table 23.11  Glossary of terms in heat-­related illness.

Heat stress

Three or more consecutive days during which the air temperature is >32.2 °C

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Heatwave

Definition

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Condition

Perceived discomfort and physiological strain associated with exposure to a hot environment, especially during physical work

Heatstroke

Severe illness characterised by a core temperature >40 °C and central nervous system abnormalities, such as delirium, convulsions or coma resulting from exposure to environmental heat (classic heatstroke) or strenuous physical exercise (exertional heatstroke)

Heat exhaustion

Mild-­to-­moderate illness due to water or salt depletion that results from exposure to high environmental heat or strenuous physical exercise; signs and symptoms include intense thirst, weakness, discomfort, anxiety, dizziness, fainting and headache; core temperature may be normal, below normal or slightly elevated (>37 °C but 24 hours at 10 °C at a wind speed of 1 km h–­1. If two layers of clothing are worn and wind speed is 5  km h–­1, survival times are calculated at 4 hours at –­50 °C, 5.6 hours at –­40 °C, 8.6 hours at –­30 °C, 15.4 hours at –­20 °C and >24 hours at –­10 °C (Türk 2010). However, the survival time depends strongly on individual factors.

24.2.3  Final cause of death The final cause of death is either ventricular fibrillation or asystolia. Internal asphyxiation or hypoxia due to a left shifting of the oxygen–haemoglobin dissociation curve, failing of enzymes and electrolyte dysregulation may also contribute to the final cause of

629

Injuries Due to Cold

death. According to animal experiments, ventricular fibrillation seems to be more predominant compared to asystolia.

24.3  Clinical phases of hypothermia

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For didactical purposes, several phases of hypothermia are differentiated: phase 1, the excitatory phase; phase 2, an adynamic phase (exhaustion); phase 3, a paralytic phase; and phase 4, the phase of apparent death (Table  24.1). Another classification of signs and symptoms of hypothermia can be found in Table 24.2. Although body core temperatures are given for these different grades and phases, it has to be kept in mind that the clinical picture at any given body temperature may vary widely. The clinical

phases are mainly characterised by functional alterations, for example within the muscular system from shivering via a drop of the muscular tonus to a rise of muscular rigidity, within the cardiovascular system from tachycardia to sinus bradycardia to bradyarrhythmia, and within the pulmonary system from hyperventilation to depression of ventilation to bradypnoea. Haemodynamic and rheological alterations in hypothermia are of importance for the development of morphological changes, and are especially responsible for the rise of resistance due to vasoconstriction and the increase of blood viscosity. These functional changes are also of medicolegal importance as the muscular rigidity seen in hypothermia must not be mistaken for rigor mortis (Figure  24.2). After initial shivering in body core temperatures of about 33–30 °C, cold stiffening occurs. The differential diagnosis should be easily made since in m ­ uscular

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CHAPTER 24  

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Table 24.1  Clinical phases of hypothermia; in individual cases, the progression of clinical symptoms may vary. Phase 2 33–30 °C

Phase 3 30–27 °C

Muscular system

Shivering

Drop in muscular tonus

Rise of muscular rigidity

Heart

Tachycardia

Sinus bradycardia

Bradyarrhythmia

Circulatory system

Reduced perfusion of body surface

Rise of resistance due to vasoconstriction

Ventilation

Stimulation of respiration, hyperventilation

Central depression of ventilation

Nervous system

Raised vigilance, confusion, painful acra

Disorientation, apathy, passing off pain

Unconsciousness, loss of reflex

NA

‘Excitation’

‘Exhaustion’

‘Paralysis’

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Phase 1 36–33 °C

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Rise of resistance due to increased viscosity of the blood

Bradypnea, apnoeic pause, decrease in compliance

Phase 4 Below 27 °C Either  Further decrease of vital functions Or  Cardiocirculatory arrest due to ventricular fibrillation or asystolia, cessation of breathing, apnoe

‘Vita reducta’ – apparent death

Hypothermia grade

Core temperature

Mild

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N

Table 24.2  Signs and symptoms with grades of hypothermia.

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C

90–95 °F (32.2–35 °C)

Signs and symptoms • • • •

Confusion progressing to impaired judgement and apathy Unable to perform complex motor functions and ataxia ‘Umbles’ – stumbles, mumbles, fumbles and grumbles Vigorous shivering, tachycardia, tachypnoea, bronchospasm, peripheral vasoconstriction, increased cardiac output, hypertension, cold diuresis

Moderate

82.4–90 °F (28–32.2 °C)

• Dazed consciousness, irrational behaviour, slurred speech, delirium, hallucinations • Paradoxical undressing; hide and die • Hyporeflexia, rigidity, reduced shivering • Hypoventilation • Progressive decrease in cardiac output and pulse; atrial and ventricular cardiac dysrhythmias; J-­wave electrocardiogram (ECG) changes

Severe

72–82 °F (135  mmol/l and urea >40  mg/dl. Persisting imbalances also result in corresponding alterations within the CSF (osmotic gradient). Regarding the post-­mortem diagnosis of water and electrolyte imbalances, measurements of the pH are of no value. Estimations of electrolytes in CSF and vitreous humour can only be of limited meaningfulness. On the one hand, the pH strongly depends on the state of the body, and, on the other hand, liquor often becomes sanguinolent when it is obtained so that there may be considerable alterations especially to electrolytes. Centrifugation may be of certain help, but cannot remove all components originating from damaged erythrocytes. This is why liquor from the lateral ventricles should be obtained, because after 12–24 hours there are no differences to lumbar liquor.

O

blood from the heart do usually exceed 200 mg/dl (93 mg/dl urea-­ nitrogen) during the first 13 hours post-­mortem in the case of uraemia from all imaginable causes.

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PART IV  

35.4.5  Water and electrolyte imbalances

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The regulation of the water and electrolyte balance aims to maintain isotonia and isovolumia within the intravasal space. Sodium, chloride and bicarbonate show the highest extracellular concentrations, whereas potassium and phosphoric esters predominate in the intracellular space. Owing to the fact that the relation between extracellular fluid volume and water exchange is much lower in infants than in adults, water imbalances may develop much earlier and be life-­threatening. It is not rare that electrolyte imbalances occur due to other diseases such as diabetes mellitus, chronic alcoholism and nutritive disturbances. There are some types of dysregulations, which can lead to sudden unexpected death and may therefore be of forensic medical relevance. Isotonic dehydration is characterised by extracellular loss of sodium and water in isotonic relation, e.g., during the polyuric phase of acute and chronic renal failure, ­vomiting and diarrhoea, pancreatitis and peritonitis, and due to dermal loss (following burn injuries). The main mechanism of hypotonic dehydration is salt depletion together with an extracellular deficit of water. Delirium and convulsions are typical cerebral symptoms, which have to be considered as causes of sudden death. Hypertonic dehydration (with hypernatraemia) leads to a deficit of free water in the extracellular and also in the intracellular space and is caused, e.g., by a lack of water supply, dermal loss (sweating) and also via the lungs (e.g. hyperventilation from infections and fever), the kidneys (diabetic coma) and the gastro-­ intestinal tract (diarrhoea, vomiting). The typical morphology comprises tinting of the skin, sunken eyes, dry surface of the galea and/or dry cutting areas of organs. A biochemical pattern was proposed as diagnostic tool. The so-­called dehydration pattern consists of an elevation of sodium >155  mmol/l, chloride

R

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Sodium and chloride There is an extracellular decrease of sodium parallel to an increase of potassium (see the preceding text) post-­mortem. As a general rule, there is a variation of the sodium level within CSF mostly corresponding to the serum concentration (ca. 128–157 mmol/l), except situations with severe infections of the central nervous system. Without differentiation regarding the mechanisms of death, sodium levels in CSF and serum are usually found within the

CHAPTER 35   Postmortem Biochemistry as an Aid in Determining the Cause of Death

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Diagnosis

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The homoeostasis of calcium has an important impact onto the neuromuscular conduction. Hypocalcaemia (total Ca 1.3 mmol/l) are chronic osteolytic or endocrine processes in most of the cases, which may be the reason for sudden unexpected deaths via electrolyte imbalances with arrhythmias, somnolence and coma. Under post-­mortem conditions, the serum calcium concentration is constant for ca. 10 hours with a slight increase thereafter (normal range in healthy individuals: 1.96–2.60 mmol/l). The calcium contents of CSF reflect approximately the serum level of ionised calcium. In vitreous humour, calcium levels are much more stable and there is less influence of agonal and post-­mortem effects.

A state of high excitation is characterised by a massive release of catecholamines, especially in situations with mechanical restraints and also in cases of prolonged agony. Such stress situations can be classified by estimation of adrenaline and noradrenaline levels using high performance liquid chromatography (HPLC) in serum, CSF and vitreous humour. Analyses in different compartments are useful to achieve semi-­quantification of the intensity of stress and its impact on the mechanism of death. Particularly, increased noradrenaline levels in CSF and vitreous humour are indicative of a protracted stress reaction. The author’s research has revealed massively increased catecholamine concentrations, partly exceeding the normal ranges many times (adrenaline values in vitreous humour and CSF 100–8,000 ng/l; noradrenaline levels 4,000–70,000 ng/l (normal ranges in serum: adrenaline 20–120  ng/l and for noradrenaline 150–170  ng/l)). Especially, high noradrenaline levels indicate a longer duration of stress. Hypothermia can also cause a massive release of catecholamines in the sense of intense stress. The levels are within the ranges of high excitation with the noradrenaline concentrations being considerably higher than those of adrenaline (10-­to 32-­ fold) comparable to cases with prolonged agony. Contrary to this, adrenaline levels often exceed those of noradrenaline in death cases with short agony. Death due to hypothermia results in mean quotients adrenaline/noradrenaline considerably 1 are typical for short agony (e.g. myocardial infarction, head trauma) being indicative of higher adrenaline levels. Additional analyses of volatile substances (ethanol, methanol, propanol-­1, propanol-­2 and acetone) usually show elevated acetone concentrations in all compartments being indicative of hypothermia, but basically only in cases that are ethanol-­free. Acetone and propanol-­2 are then altered equally. If relevant alcoholisation is found, both substances can only be found in very low or physiological ranges that is indicative of an anti-­ lipolytic effect of ethanol (acetone >35 mg/l if the blood alcohol level is benzoylecgonine > morphine, 6-­acetylmorphine (0–34% of the initial concentration) Average % of drug concentration following B/P: 6-­acetylmorphine: 22%/48%; morphine: 41%/60%; dihydrocodeine: 47%/54%; cocaine: 34%/34%; benzoylecgonine: 50%/62% Diazepam: 60%; nordiazepam: 32%; 7-­aminoflunitrazepam: 38% THC: 65%; THC-­COOH: 80% THC: 64%/87%; THC-­COOH: 94%/89%

Skopp et al. 2000

49–100% 23–100%   9–80% 17–100% 53–100% 64–95% (n = 7) increase by 7–255% (n = 21), possibly due to differences in hair morphology

Petzel-­Witt et al. 2017

Skopp et al. 1997b

Skopp et al. 1997a

Yegles et al. 2000 Van Elsué 2018

Miolo et al. 2020

R

N

H

R + CH2 N

–CO2

R

O

R H

N

CH3

R O–

Figure 50.1  Eschweiler–Clarke reaction: methylation of a secondary amine moiety.

O

N

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with amphetamine-­ type drugs, barbiturates, benzodiazepines, and fenfluramine (Gannett et al. 2001; Tracy et al. 2001; Cingolani et  al.  2005; Tirumalai et  al.  2005). Interestingly, some drugs appear to be reasonably stable in formalin such as diazepam, chlorpromazine, and succinylcholine (Forney et  al. 1982; Nishigami et al. 1995). Tissue dehydration impacts quantification of drugs, in addition (Rohrig 2019).

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50.8  Instability of drugs and artificial formation during processing and analysis Although the history of a sample appears to be most relevant to degradation of a drug and/or production of artifacts, such processes may also take place during isolation and determination of a drug. Generally, even the most promising method should consider errors from all sources (Savoie et al. 2009). Reference standards are a prerequisite for quantitative analyses. Pharmaceutical grade standards may contain trace amounts of structural analogues related to the manufacturing process. For example, the allowable pharmaceutical impurity limit of codeine in morphine is 0.5% (Rohrig 2019). Chromatography has been the mainstay of drug analysis for many years. Drug analysis is mostly preceded by protein precipitation and/or extraction into an organic solvent for partial purification of a biological fluid or a tissue homogenate. Lipophilic compounds will readily be extracted by non-­polar solvents. The formation of methochloride adducts has been reported to occur during isolation of clozapine, olanzapine, and ofloxacin using dichloromethane (Mohammadi et  al.  2008). Phosgene, which may be present in chloroform, reacts to form carbamate derivatives during isolation of tricyclic antidepressants (Wester et al. 1981). The more polar solvents being partially miscible with water will also remove water-­soluble materials. As a result, drug conjugates are transferred into the organic phase. If phase II metabolites are subsequently hydrolyzed, erroneously high levels of the parent drug will result (Mauden et  al.  2000; Skopp and Pötsch 2002a). Conjugate instability is most common with labile conjugates such as N-­ or O-­acyl-­glucuronides; for example, silylation of co-­extracted THC-­COO gluc resulted in the formation of THC-­COOH (Skopp and Pötsch  2002a; Shipkova et  al.  2003). The analysis of solvents or volatiles such as chloroform is usually performed using blood. Whole blood should be preferred over

U

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O

C

R

FO

+CH2O –H2O

R

TO

only as a result of their volatility but also because of their instability. A loss of up to 25% of blood toluene has been observed in sealed glass tubes stored at room temperature for 7 days, and a still higher loss was noted in glass tubes with rubber stoppers (Saker et  al.  1991). A total loss of chloroform occurred from blood stored at room temperature, which has largely been attributed to chloroform being lost as a result of opening the storage tube. Chlordiazepoxide tends to form desoxychlordiazepoxide during storage and further degrades to nordiazepam, which also represents a metabolite and an artifact in the analysis of the parent drug by gas chromatography (GC) (Skopp  2004). Some drugs undergo interconversion during storage. ex  vivo cis/trans-­ isomerization has been observed with retinoids, which are prone to photochemical and oxidative degradation as well. Conversion of the trans-­to the respective cis-­isomer has been reported for the calcium channel antagonist lacidipine (Baranda et al. 2006). Cyanide is unstable in stored blood samples. A significant decrease in blood cyanide concentration has been attributed to mechanisms that include evaporation, thiocyanate formation, and reaction with specimen components. Time, temperature, and cyanide concentration are apparently important factors in these changes (Bakshi and Singh  2002, Rohrig  2019). Conversely, an average increase of 35% of cyanide in unconditioned samples­ (n = 14) following storage of 25–30 days has been shown. If sodium fluoride (2%) was added to postmortem samples upon their collection, the concentration of cyanide in pretreated samples did not differ from their initial concentrations (McAllister et al. 2011). Storage of oral fluid samples that have been spiked with benzodiazepines were stored in the collecting device (Dräger DCD 5000, Dräger, Lübeck, Germany) either without further treatment or following the addition of 950 μL methanol up to 14  days at ambient temperature. Changes of negative controls in the respective devices and of the spiked medium as well as of pure substances in methanol kept in Eppendorf tubes (Eppendorf, Berzdorf, Germany), respectively, were also moniWesseling-­ tored. Ratios of 85–115% between reference samples were used as a criterion of stability. Instability was already observed after 2 days of storage, and recovery was less than 60% for all benzodiazepines kept in the original device without methanol having been added. In samples stored with methanol as a preservative, concentrations decreased by 20% at the most. Benzodiazepines that are not stable in oral fluid are chlordiazepoxide, flunitrazepam, and clonazepam (Kempf et al. 2009). Sometimes it is necessary to perform an analysis on specimens stored in formaldehyde or having been collected from an embalmed body. Embalming fluid is based on formaldehyde with final concentrations of 5–20% at acidic or neutral pH values, and usually also contains alcohols, e.g. methanol as an antipolymerizing agent. Formaldehyde being a highly reactive chemical agent can mask, alter, or destroy a drug. Most likely, reaction pathways are through hydrolysis and/or methylation via the Eschweiler– Clarke reaction (Figure 50.1). For example, methylation of nortriptyline to amitriptyline occurs during formalin fixation and storage (Dettling et al. 1990). Methylation has also been observed

TOXICOLOGY

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PART VII  

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CHAPTER 50   Issues Affecting Interpretation: Stability and Artifacts

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Degradation of norbuprenorphine is faster than that of buprenorphine, which may serve as a likely explanation for discrepant results of hair analysis reported in the literature. The concentration of both analytes will be underestimated if recovered by acidic procedures (Cone et al. 1984; Skopp et al. 2011). Lactone–hydroxyl carboxylic acid interconversion presents a challenge in quantitative analysis, which typically occurs for statins (e.g. simvastatine). The parent drug is available as a ­γ-­lactone and readily metabolizes to the corresponding active ­β-­hydroxy acid. This metabolite can convert back to the lactone form at a low pH value while the parent compound readily undergoes hydrolysis under acidic as well as under basic conditions (Li et  al.  2010). Minimum interconversion was seen with simvastatine following extraction at a pH value of 4  maintaining the acidic analytes in a non-­ionic lipophilic form. A flash freezing step to separate the organic layer from the aqueous helped to retain the polar matrix in the frozen phase (Patel et al. 2008). It is often necessary to evaporate extracts in order to reconstitute them into small volumes for transfer to chromatography. Loss by adsorption or volatilization has been described for several drugs during extraction and evaporation. The loss of tricyclic antidepressants during sample processing was as high as 50%; the addition of 0.05% diethylamine before evaporation significantly increased the recovery of amitriptyline, nortriptyline, imipramine, desipramine, doxepin, and nordoxepin (Tserng et al. 1998). The adsorption losses can be prevented, for example by silanization of glassware or by including a solvent, such as amyl alcohol, as an additive to the extracting solvent or prior to evaporation. Volatilization of amphetamines can be solved by converting them to their non-­volatile hydrochloride salts (Chamberlain 1995). Gas chromatography coupled to mass spectrometry (GC-­MS) in the electron impact ionization mode is generally accepted for an unequivocal identification for most drugs. Trazodone and nefazodone, for example, are not suitable for measurement by GC due to thermal instability (Garg 2008). A source of chloroform in blood on headspace GC was from thermal decomposition of trichloroacetic acid at a temperature of 60°C (Flanagan and Pounder 2010). Trichloroacetic acid is a metabolite of the solvent trichloroethylene or of chloral hydrate with an elimination half-­ life of 3–5 days. Further valuable information on artifacts is covered by a few reviews (Skopp 2009; Drummer 2010; Rohrig 2019; Maurer 2020; Baselt 2020). Liquid chromatography coupled to mass spectrometry (LC-­ MS) has become a powerful tool in forensic toxicology. Today, tandem mass spectrometry (MS/MS) and high-­resolution mass spectrometry are preferably used. During instrumental analysis, conversion of analytes can arise from degradation occurring on the column due to the chosen mobile phase or the ion source. Labile molecules can fragment during the ionization process (in-­ source transformation) before entering the mass analyzer. This fragmentation is prominent with N-­oxides or conjugates and can result in an overestimation of the parent compound. Misinterpretation may also be the case if chromatography fails to separate the parent compound from its oxides or conjugates. For example, baseline separation of clozapine from its N-­oxide and

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plasma because removing serum or plasma from the sample already enhances the risk of loss of volatiles (Flanagan and Pounder  2010). Glucuronidase/arylsulphatase used for cleavage of phase II metabolites has been suggested as a source of a cyclodipeptide which has been assigned as a possible dihydroergotamine artifact in a screening procedure (Theobald et al. 2005). Enzymatic hydrolysis of urine from individuals taking oxazepam resulted in reductive formation of the drug to nordiazepam, and small amounts of diazepam were present in hydrolyzed samples after administration of temazepam (Rohrig 2019). In case of acyl glucuronides, acyl migration leads to glucuronides being no longer amenable to enzymatic cleavage selective for 1-­O-­substituted beta-­D-­glucopyranosiduronic acids (Maurer 2020). During a methanolysis deconjugation step, artifactual formation of 4-­androsten-­3,17-­dione and androsterone from the internal standards [16,16,17-­d3]-­testosterone, and [16,16,17-­d3]-­5α-­a ndrostane-­3α,17ß-­ diol has been observed. This is achieved through enzymatic oxidation of the 17ß-­hydroxyl function to a keto function by 17ß-­ hydroxysteroid dehydrogenase and 17-­ ­deuterium–hydrogen exchange at the 16C atom (Kwok et al. 2008). Norfluoxetine is degraded under acidic hydrolysis by ether cleavage and water elimination. Both tranylcypromine and the norfluoxetine artifact are isobaric compounds showing similar electron impact mass spectra with or without derivatization (Schwaninger et al. 2010). Buprenorphine and norbuprenorphine undergo an acid-­catalyzed rearrangement when exposed to acid and heat by overall elimination of a molecule of methanol with concurrent formation of a tetrahydrofuran ring (Figure  50.2).

OH

OH

+ HCl – H2O

O N

OMe

R=H

H

OMe

H

N

+ H3C H3C

CH 3 CH 3

Buprenorphine rbuprenorphine

R

R=

O

N

C

HO H3C H3C

R

O

R

FO

R

CH 3

CH 3

Rearrangement of a methyl group

OH O

N

OMe H3C H3C H3C

+

CH 3 Cyclisation

OH

R

O N

O H3C

CH3 CH3 CH3

Figure 50.2  Acid-­catalyzed rearrangement of buprenorphine.

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For multi-­analyte assays, it is most practical that all analytes are added to the stability samples. Stability samples are analyzed in an analytical run containing calibrators and quality control samples. Two concentration levels at a relevant high and low concentration which are typically equal to the high-­and low-­quality control levels used to determine method precision are recommended (van de Merbel et al. 2014). Analysis of comparison and stability samples should be performed in a random order. Also, a sufficient number of replicates with three repetitions as a minimum should be performed to obtain a reliable average result for the stability assessment. For methods with a large variability, the number of replicates should be increased; six respective replicates at two concentrations for both, the ‘fresh’ and the stability samples, preferably at the extremes of the concentration range derived from authentic samples will perform satisfactorily. If only two to three replicates are processed from each sample, resulting confidence intervals may be very wide or even unreliable (Hartmann et al. 1998). Whole blood stability testing may not be strictly necessary as long as plasma/serum stability under the same storage conditions has been demonstrated. However, for analytes with significant partitioning into the cellular phase, spiked while blood must be incubated for a sufficient period to assure that the analyte distribution between the cellular and fluid fraction is at equilibrium. For urine, the varying pH value of 4.0 to 8.5 must be taken into account, and experiments should be conducted at three different pH values. As a sediment may be formed during storage, attention should be paid to take a representative sample especially when the analyte binds to the sediment. Urine usually contains high amounts of phase II metabolites that can convert back to the parent compound. Therefore, study samples should either be spiked with these metabolites or the use of incurred samples should be considered. For tissues, stability studies can only be performed by preparing homogenates. Stability of the analyte stock solutions and derived solutions that are used to prepare calibration and quality control samples should be assessed as part of the validation procedure of the bioanalytical assay. It is independent of that of the reference standard material, where the expiry date is regularly available. It appears necessary to investigate stability on long-­term storage as well as on daily use at bench top. Stock solutions should be correctly made up also considering the analyte’s stability in the particular solvent. For example, cocaine or heroin should be dissolved in acetonitrile rather than in methanol. When possible, dilutions should be prepared in the same solvent or buffer as the provided stock solution. The stability assessment of stock solutions and derived solutions should cover the maximum period of their usage and be performed in the same solvent and container type used for the preparation of calibration standards or quality control samples (Li et al. 2010). Stability can be examined by preparing a fresh solution and comparing its concentration to that of a stored sample. The acceptable difference should be tighter (≤7% or 10%) than normally applied in other stability assessments (≤15%) as analytical results are largely influenced by changes of stock solutions (Nowatzke and Woolf  2007; U.S. Department of Health and

TR IB

50.9  Procedures for evaluating stability and artifacts

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N-­desmethyl metabolite is mandatory to allow a reliable determination of all three analytes. Chromatographic analysis of acyl ­glucuronides may also be confounded by the fact that these compounds are isomeric and have very similar LC retention times and mass spectral properties. Selectivity can be achieved in an LC-­MS/MS assay through a total of three stages of separation­ (Li et al. 2010). Lactonization of a carboxylic acid metabolite can generate the same precursor ion as the corresponding lactone, thus leading to an overestimation of this compound. If high ionization temperatures are employed, hydrogen–deuterium ­ exchange of deuterated internal standards can occur during the ionization process. Inaccuracy of LC-­MS assays may also be attributable to ionization matrix effects that may enhance or – in most cases – suppress the ionization yield (Sauvage et al. 2008; Vogeser and Seger 2010). In case of high-­resolution mass spectrometry, the mass deviation between the exact and measured mass should not exceed 5 ppm (Maurer 2020). Appropriate procedures for the analysis of postmortem species have been sufficiently covered by recent reviews (Drummer 2007, 2010). A special pretreatment or homogenization according to the specimen’s nature and/or a more sophisticated clean-­up extraction of putrefied or embalmed materials may be required for all forms of chromatography. The lack of suitability of routinely applied isolation methods may result in poor recovery and co-­extraction of interferences leading to signal overlap and ion suppression or enhancement (Drummer  2010; Rochholz and Sporkert 2016).

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There is some guidance on how to perform and evaluate stability experiments in practice (Peters 2007; U.S. Department of Health and Human Services Food and Drug Administration, Center for Drug Evaluation and Research (CDER), Center for Veterinary Medicine (CVM) 2018). The matrix used in a stability study should resemble that of the authentic specimens as closely as possible. However, even in elaborate guidance documents, changes of the blank matrix have not always been considered. A pool of authentic matrices may cover interindividual variability whereas stripping with active carbon or otherwise altered matrix is not recommended (Nowatzke and Woolf  2007; van den Merbel et al. 2014). For analytes being susceptible to enzymatic degradation, the use of sufficiently fresh matrix is recommended. Not only the same matrix should be used but also its source should be the same. It should be considered that the expression and activity of esterases, which are a common cause of instability, are different within and among species. For example, degradation of cisatracurium by rat plasma esterases is rapid relative to human plasma (Li et al. 2010). It is essential to use the same container type and preservatives or additives for comparison and exposed (stability) samples; changes of container type or of additives or preservatives will afford a re-­evaluation of stability (Hartmann et al. 1998).

CHAPTER 50   Issues Affecting Interpretation: Stability and Artifacts

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Both approaches mentioned above suffer from within-­run variation, while the second one suffers also from between-­run precision. Bias error may arise during the preparation of calibration standards and quality control samples. An alternative strategy may involve preparation of a single batch of fresh biological matrix spiked at specified levels and subdivided for use as ‘stability’ and ‘comparison’ samples, the latter being stored at a temperature lower than –130°C which implies cryopreservation in liquid nitrogen. Although these ‘comparison’ samples may be equivalent to freshly prepared samples, the possibility of changes related to both the matrix and the analyte cannot totally be excluded (Dadgar and Burnett 1995). Criteria for long-­term stability assessment have not specifically been defined. Concentration ratios between comparison and stability samples should differ by no more than ±15%; the 90% confidence interval should lie within an acceptance interval of 85–115% or 80–120%. For all types of stability testing, a visual evaluation – by a scatter diagram or a box plot – and an outlier test, if indicated, should be performed. The 90% confidence interval is considered most useful as it also accounts for the degree of scatter and covers decreases as well as increases (Peters 2007). Incurred samples are more complex in their composition than quality control samples. Metabolites, especially phase II metabolites, e.g. glucuronides or N-­oxides, may often cause failure in the analytical process due to their instability. Incurred sample reanalysis and incurred sample stability should therefore be part of the bioanalytical process to verify reliability of the reported sample analyte concentrations (Li et  al.  2010). The original and repeat analyses should use the same analytical method with calibration standards, quality control samples and sample aliquots being processed separately from those in the original run (U.S. Department of Health and Human Services Food and Drug Administration, Center for Drug Evaluation and Research (CDER), Center for Veterinary Medicine (CVM) 2018). When stability studies are performed on authentic samples, the use of paired comparisons to test for stability appears reasonable. In addition, estimation of the critical difference is a popular approach that is based on the performance of the respective assay. The critical difference d is calculated according to d = 2 • 21/2 • SD, with SD being the standard deviation of the day-­to-­day imprecision. Successive measurements cn and cn+1 are considered significantly different if the absolute difference between cn and cn+1 is greater than d (Stamm 1982). The reaction type involved in drug degradation can be derived from time-­dependent changes, which finally may guide to a more proper estimation of the drug level at the time of sampling. Hydrolysis of ester-­type drugs can be described as apparent first-­ order reaction kinetics, whereas oxidation reactions often comply with second-­order reaction kinetics. Investigations on the reaction type involved in the degradation of forensic-­relevant drugs have already been performed, for example for morphine, morphine glucuronides, cocaine, benzoylecgonine, ecgonine methyl ester, LSD, and 11-­nor-­9-­carboxy-­Δ-­9-­tetrahydrocannabinol glucuronide (Skopp and Pötsch  2002a; Skopp, Klingmann et  al. 2001; Skopp, Pötsch et al. 2001, 2002).

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Human Services Food and Drug Administration, Center for Drug Evaluation and Research (CDER), Center for Veterinary Medicine (CVM) 2018). Process or bench-­top stability should confirm that analyte degradation does not occur during sample preparation or extraction. Stability samples are allowed to remain on the bench top, typically at ambient temperature and for 4–24 hours (Nowatzke and Woolf 2007). Stability is then assessed in relation to freshly thawed stability samples; and both sets are analyzed against calibration standards to decide if a preservative regime has to be included. On-­instrument or processed sample stability does not need additional experiments if samples are bracketed by quality control samples. Data on the reanalysis of processed samples may be useful if analysis, including calibration standards and quality control samples, is interrupted. All processed samples should be allowed to remain on the instrument after the initial analysis, and measurement is run once again typically after 12–72 hours. Even if there is a difference between the two runs, agreement of quality control samples as calculated from the respective calibration curves can indicate that the entire analytical sequence has successfully been reanalyzed. Freeze–thaw stability evaluation should include a minimum of three cycles at two different concentrations. Samples should be stored in a freezer for at least 24 hours at –20°C. When completely thawed, samples should be refrozen for at least 12–24 hours (Hartmann et  al.  1998; Nowatzke and Woolf  2007; U.S. Department of Health and Human Services Food and Drug Administration, Center for Drug Evaluation and Research (CDER), Center for Veterinary Medicine (CVM) 2018). Samples that passed through all three cycles were recommended to be analyzed together with a set of samples that was subjected to a single freeze–thaw cycle. Unlike periodically testing that is usually performed during long-­term stability studies, it was suggested to evaluate freeze–thaw stability after the last cycle only (Hartmann et al. 1998). A difference greater than 15% may indicate freeze–thaw instability (Nowatzke and Woolf 2007). Generally, long-­term stability may be evaluated by: 1. Spiking a drug-­free matrix with an analyte at a defined level. After storage at specified conditions, either a second blank sample is spiked at a similar level or quality controls are used which are analyzed with stored samples using freshly prepared calibration standards. 2. Analyzing the spiked matrix immediately after its preparation. Aliquots are stored for one or more periods, before analysis is performed using freshly prepared calibration standards on each occasion. First of all, it is recommended to analyze replicate aliquots of stability samples against freshly prepared matrix standards within 24 hours of stability sample preparation to assess accuracy of the preparation of the stability samples. Once this initial analysis has been successful, for example that imprecision is 1) a compound is and it allows the distribution of drugs within the body to be estimated. Hydrophobic drugs with high partition coefficients are preferentially distributed to hydrophobic compartments such as lipid bilayers while hydrophilic drugs preferentially are found in hydrophilic compartments such as plasma. To measure the partition coefficient of ionizable drugs, the pH of the aqueous phase is adjusted to represent the water compartment (usually around pH 7). The logarithm of the ratio of the concentrations of the unionized compound in the solvents is called KD or log P: K Doct / wat

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The pH of the milieu in which the drug is solved (and thus its pKa value) has a great impact on the amount of absorption from the gastrointestinal tract. Additionally, many ingested compounds are weak acids or bases and are thus present in solution in both non-­ionized and ionized forms. As mentioned above, the non-­ ionized molecules tend to be lipid-­soluble (KD > 1) and cross membranes by passive diffusion, whereas the ionized forms have low lipid solubility (KD < 1) and cannot cross the cell membranes. The diffusion of weak electrolytes across membranes will thus be influenced by their pKa value and the pH gradient across the membrane. The rather low pH in the stomach allows weak acids to be absorbed as they are uncharged there. In contrast, weak bases are ionized and can therefore not be absorbed by passive diffusion. In the intestine, absorption is best for weak acids or weak bases with pKa values approximately between 3 and 8. In addition, the big surface of the intestine and the longer dwell time within this area making the intestinal absorption quantitatively most important in contrast to all other areas. Furthermore, the pH value in the duodenum/intestine is rather alkaline with values up to 8.5 further increasing the absorption of especially basic drugs, which are in consequence not protonated and can penetrate through membranes. Finally, alteration of urine or plasma pH can be used to influence drug concentration in certain body fluids in the case of e.g.

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Orally administered drugs enter the central blood circulation after enteral absorption and after passing through the liver. In both steps, compounds can already be metabolized by enzymes located in the enterocytes and/or hepatocytes. This part of metabolism is named first-­pass metabolism. The extent of first-­pass metabolism has a strong impact on the bioavailability. The hepatic metabolism is described in 51.7. In the following, the focus is on intestinal first-­pass metabolism. In the gut, oxidative metabolism is mainly mediated by CYP3A4. Also, phase II enzymes are located here such as the UDP-­glucuronyltransferases and UGT2B7. Gastrointestinal bioavailability (Fg) depends on the absorption and intestinal metabolism. For example, cyclosporine, midazolam, or verapamil undergo significant intestinal metabolism. Furthermore, non-­hepatic first-­pass metabolism is located in other tissues that may be involved in the absorption processes such as the lung and skin.

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The P-­glycoprotein transporter The ATP-­ binding cassette (ABC) transporter P-­ glycoprotein (P-gp), the multi-­drug resistant mdr-­1 gene product, can have a great impact on the enteral drug absorption. P-­gp represents an ATP-­dependent drug efflux pump for various compounds. It also often mediates the development of resistance to anticancer drugs, in particular cancer cells. Therefore, increased intestinal expression of P-­gp can reduce the absorption of drugs that are substrates for P-­gp. Thus, there is a reduced bioavailability and, in consequence, therapeutic plasma concentrations cannot be reached. On the other hand, decreased P-­gp expression may lead to higher blood plasma concentrations, and drug toxicity may occur.

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Following entry of a xenobiotic to the systemic blood circulation, distribution describes the reversible transfer of drug from one location to another within the body. The main compartments are plasma (5% of body weight), interstitial fluid (16%), intracellular fluid (35%), transcellular fluid (2%), and fat (20%). Alternatively, the composition can be summarized as follows: water/muscle (80%) and fat (20%). Distribution is influenced by many parameters such as passive diffusion across lipid membranes, carrier-­ mediated active transport processes, and protein binding in the blood and tissue. The drug distributed with time primarily in highly perfused organs such as liver, heart, and kidney but also the brain. Distribution is limited in slightly perfused organs such as muscle, fat, and peripheral organs. The drug is moved from the plasma to the tissue until equilibrium is reached. The VD (see also 7.5.2.1) of a drug is a property that quantifies the extent of distribution into tissues. As stated previously, most tissue membranes behave as typical lipid barriers allowing small lipophilic molecules to cross cell membranes. Drugs can accumulate by diffusion under the influence of pH gradients, binding to intracellular constituents, or partitioning into lipid depots. Active transport plays also an important role in the distribution process. If a drug enters a tissue by an active transport mechanism, its concentration in the tissue can be many times greater than that in plasma. Binding to plasma proteins can also influence distribution of xenobiotics. As only unbound drug is in equilibrium across membranes, a drug that is extensively and strongly bound to plasma proteins has only limited access to the tissues. A further important point is the pH value of the tissue, blood, or organ, into which the drug diffuses. An inflammatory response can reduce the local pH to 5.5 or below due to the damaged vasculature. Anti-­inflammatory drugs such as acetylsalicylic acid can diffuse and accumulate in the uncharged state and become active in the damaged cells and surrounding milieu. In summary, distribution is affected by similar parameters as the absorption, which are discussed below.

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to plasma proteins and only 3% of the amount is free and hence pharmacologically active. Additionally, only this unbound fraction of drugs can undergo drug elimination with respect to metabolism and renal filtration. As the protein binding is reversible, a chemical equilibrium exists between the bound and unbound fraction. Among the involved proteins, plasma albumin is the most important binding mainly acidic drugs such as the non-­steroidal anti-­inflammatory drugs ibuprofen or diclofenac. Basic drugs such as the tricyclic antidepressants doxepin and amitriptyline are mainly bound to β-­ globulin and acid glycoprotein. Protein saturation by a drug may lead to a non-­linear relation between dose and the free drug concentration. If a therapeutic drug concentration is near to the protein binding saturation, its free plasma concentration will increase after additional dosing. Hence, doubling the dose can therefore more than double the free concentration of drugs and therefore double the pharmacologically active concentration. Changes in the concentrations of free drug lead to changes in the volume of distribution because free drug may distribute into the tissues leading to a decrease in plasma concentration profile. Also, the bound part may act as a reservoir or depot from which the drug is slowly released. Since the unbound form is being metabolized and/or excreted from the body, the bound fraction will be released in order to maintain equilibrium. Furthermore, extensive protein binding can lead to competition between drugs for protein binding, which may lead to drug-­ drug interactions. For example, if digoxin is given, it will bind to the plasma proteins in the blood. If warfarin is also given, it can displace digoxin from the protein, thereby increasing the unbound fraction of digoxin. This will lead to an increased pharmacological effect of digoxin, since only the unbound fraction exhibits activity.

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Several compounds such as cyclosporine and digoxin are substrates of this transporter and their absorption and thus bioavailabilities are reduced due to their affinities to the P-­gp. Besides the intestine, P-­gp is extensively distributed and expressed in other areas in the body such as hepatocytes, renal proximal tubular cells, adrenal gland, and capillary endothelial cells being part of the blood brain barrier.

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51.6.1  Binding to plasma proteins Many drugs exist mainly in bound form when present at therapeutic concentrations. In consequence, the free part of the drug, which is actually the active part, can be less than 1%. For example, the bound fraction of the anticoagulant warfarin at therapeutic concentrations is 97%. This means that 97% of warfarin is bound

51.6.2  Drug reservoirs after distribution into body fat and other tissues The impact of drug reservoirs in body fat on the drug toxicity is demonstrated by the anesthetic drug thiopental. Accumulation of a drug in fatty tissue can be predicted by looking at its KD value. Thiopental for instance has a comparably high KD value and accumulates substantially in body fat. This has consequences on its pharmacokinetic behavior. After a bolus intravenous injection, thiopental reaches its maximum concentration in its target tissue (CNS) within 1–2  min after injection. Subsequently, the plasma concentration falls as the drug distributes into other tissues such as muscle. Its concentration in the brain changes inversely proportional to its plasma concentration. This leads to a rapid termination of anesthesia by redistribution rather than by elimination. Finally, thiopental is then slowly distributed into fatty tissues. In general, accumulation of drugs in body fat is limited due to its poor perfusion. Consequently, drugs are delivered to body fat rather slowly, and the theoretical equilibrium distribution between fat and body water is approached slowly. However, accumulation in body fat is

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firstly be found in the genotype of the individual. Genes for certain enzyme systems may be absent, deficient, or overexpressed. Besides the phase I enzyme family, also phase II enzymes can show polymorphism such as uridine diphosphate glucuronyltransferase (UGT) UGT1A1, thiopurine methyltransferases, arylamine N‑acetyl transferase (NAT)2, and glutathione S-transferases GSTM1 and GSTT1. However, the actual metabolic activity determines the phenotype which depends on the genotype and the level of expression. In addition, variations must be taken into consideration resulting from interactions with other drugs, food ingredients, and/or tobacco smoke ingredients. An example of a clinically relevant enzyme deficiency is the lack of glucose-­6-­phosphate dehydrogenase. It is a genetic defect that is inherited via the X-­chromosome and, therefore, manifests in men with one respective copy. In women, it will only be manifested if they are homozygous for the corresponding defect. In these cases, application of dapsone or primaquine may lead to hemolysis, because in erythrocytes NADPH decreases leading to the reduction of oxidized glutathione. Oxidized glutathione can lead to damage of erythrocytes, hemolysis, and resulting methemoglobinemia. Normally, NADPH is regenerated by oxidation of glucose-­6-­phosphate.

51.7.1  Examples on polymorphism relevant in forensic and clinical toxicology

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often significant for certain drugs such as general anesthetics, e.g. the aforementioned thiopental, or insecticides that are therefore poorly metabolized and can accumulate slowly but progressively in body fat. On repeated administration, fat and other poorly perfused tissues can accumulate large amounts of such drugs. These reservoirs are then capable of maintaining plasma and, hence, brain concentrations of e.g. thiopental at levels above those needed for anesthesia. Thus, a compound whose duration of action is limited by rapid redistribution from its site of action to storage sites can become long-­acting if accumulation sites are present. Besides body fat, there are other reservoirs possible that have pharmacological importance. For example, chloroquine, an antimalarial accumulates in melanin whereas amiodarone, an antidysrhythmic drug accumulates in liver and lung. With respect to protein binding and drug reservoirs, it is evident that individual factors such as gender, age, and body weight can dramatically influence the pharmacological effect of certain drugs at the same dose. The percentage of body fat for women is greater than that for men being around 3%–5% in men and 8–12% in women. Body fat is also much higher in obese people, being around 25%. In older people, the relative amount of body fat also increases. Protein binding is of course influenced by the total amount of binding protein (e.g. albumin) available. This total number, similar to the body fat, is also influenced by nutrition. Malnutrition for instance will lead to a decreased amount of albumin and other binding proteins available in the body.

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51.7 Metabolism

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Xenobiotics such as therapeutic drugs, drugs of abuse, and other toxic compounds can be chemically modified by various metabolizing enzymes. although the liver is the main metabolizing organ, other organs also metabolize drugs, particularly those with direct contact to the environment such as the gut, lungs, and kidneys. The metabolizing enzymes include cytochrome P450 oxygenases, UDP-­glucuronosyltransferases, sulfotransferases, and glutathione S-­transferases. The metabolism can be divided into phase I (functionalization) and phase II reactions (conjugation). The modified and thus more water-­soluble xenobiotics can then be excreted. Although most end products of metabolism are pharmacologically inactive, some drugs (so-­called pro-­drugs) need to be bioactivated to active metabolites (e.g. O-­ demethyl tramadol from tramadol). However, such bioactivation may also lead to toxic products (e.g. para-­naphtochinonimine, the toxic metabolite of paracetamol). For enzymes involved in these phases, polymorphisms are known. For phase I reactions, the cytochrome P450 (CYPs) are of particular importance and are present in different isoforms. Two of these isoforms, among others, CYP2C19 and CYP2D6 show polymorphisms that may affect the pharmacological potency or the toxicity of xenobiotics if metabolized mainly through one of these isoforms. So, genetic variability plays an important role in xenobiotic metabolism. The reasons for these differences can

The most important phase I enzyme family that is polymorphically expressed is the family of cytochrome P450 isoenzymes. For instance, the polymorphism of CYP2D6 was discovered in a pharmacokinetic study on sparteine, in which side effects of participants occurred to an uncommonly high degree. Among these subjects, the total clearance of sparteine was reduced, and almost entirely unchanged sparteine could be detected in urine. Since debrisoquine behaves similarly, the CYP2D6 polymorphism is also called sparteine/debrisoquine polymorphism. About 10% of the Caucasian population are slow metabolizers for CYP2D6 and are therefore called poor metabolizers (PM). The activity of the enzyme is markedly reduced due to various mutations or complete loss of the corresponding gene. The rest of the population belongs to the type of rapid (or normal) metabolizers (extensive metabolizers, EM) or to the type of the so-­called ultra-­rapid metabolizers. The toxicological relevance of this polymorphism is widespread since many potent drugs such as tricyclic antidepressants, neuroleptics, and opioids are metabolized by CYP2D6. Similarly as described in the preceding text, slow and fast metabolizers exist with respect to the CYP2C19 isoforms. For  example, diazepam is metabolized by this enzyme and is expected in slow metabolizers to show prolonged half-­lives and delayed elimination. Besides CYPs, most of the other metabolizing enzymes are expressed polymorphically. For example, monoamine oxidase A, catechol-­O-­methyltransferase, and tryptophane hydroxylase can also have an impact on pharmacotherapy.

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Methadone

Among the phase II enzymes, the polymorphism of the NAT catalyzing the transfer of acetyl groups from acetyl-­CoA to arylamines or hydrazines is well known in humans; the two NAT genes are known to encode NAT1 or NAT2. For both enzymes, a polymorphism is known leading to the phenotypes of slow and fast acetylation. The frequency of individual phenotypes has strong ethnic differences. About 40–70% of Caucasians are slow metabolizers, but only 10–20% of the Japanese or Chinese population. The toxicological relevance of NAT polymorphism should not to be underestimated. Since it plays no role in acute toxicity, there is evidence that bladder cancer induced by benzidine is much more frequent in slow metabolizers than in rapid metabolizers. This is attributed to decreased detoxification by acetylation. Besides NAT, there are other important phase II enzymes that are polymorphically expressed such as the UGT isoforms. Examples on their impact on the effect of certain drugs will be given in the following. Furthermore, the impact of other polymorphically expressed enzyme systems will be highlighted and exemplified on several therapeutically used opioids and certain drugs of abuse.

Methadone is mainly N-­demethylated followed by rearrangement by water elimination to the pharmacologically inactive 2-­et hylidene-­1,5-­dimethyl-­3,3-­diphenylpyrrolidine (EDDP). For a given dose, methadone plasma concentrations can vary widely between individuals, contributing to a high interindividual variability in response to treatment. CYP3A4 and CYP2B6 are the major CYP isoforms involved in methadone metabolism, with CYP2D6 contributing to a minor extent. In this context, it is important that CYP2B6 is one of the most polymorphic CYPs in humans with over 100 described single nucleotide polymorphisms, numerous complex haplotypes, and distinct ethnic frequencies. Moreover, 20-­to 250-­fold interindividual variations in CYP2B6 expression has been demonstrated, presumably due to transcriptional regulation and polymorphisms, being in part responsible for the high interindividual variability in the response to methadone treatment. A higher stereoselective metabolism (S>R) occurred in livers expressing high levels of CYP2B6 compared with CYP3A4 suggesting a significant role of CYP2B6, but not CYP3A, in the stereoselective metabolism and disposition of methadone in humans. Furthermore, methadone is a P-­gp substrate and, although there are inconsistent reports, ABCB1 polymorphisms may also contribute slightly to the interindividual variability of methadone kinetics and influence dose requirements. The large interindividual variability in the activity of CYP enzymes, as well as their potential for induction and inhibition, is responsible for a large part of the variability in methadone kinetics and observed plasma levels. For instance, in order to obtain methadone plasma concentrations of 250 ng/mL, doses of racemic methadone as low as 55 mg/day or as high as 921 mg/day can be required in a 70-­kg patient without any co-­medication, depending on its phenotype.

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Oxycodone undergoes N-­demethylation mainly by CYP3A4 and CYP3A5 and O-­ demethylation to oxymorphone by CYP2D6. Variability in the activity of these enzymes because of polymorphisms and/or drug-­drug interactions is well known and CYP2D6 activity was correlated with oxycodone experimental pain assessment. CYP2D6 ultra-­ rapid metabolizers experienced increased pharmacodynamic effects in contrast to CYP2D6 poor metabolizers. The modulation of CYP2D6 and CYP3A activities had clear effects on oxycodone pharmacodynamics and these effects were dependent on CYP2D6 polymorphism. An interesting case report described a patient who had a clinically significant drug interaction between oxycodone and the CYP3A4 inducer rifampin resulting in insufficient pain suppression. The urine drug test for oxycodone (but not detecting nor-­oxycodone) was negative suggesting non-­compliance by the patient. Phenotyping (determination of the complete oxycodone metabolite profile) and genotyping concerning CYP3A4/5 and 2D6 allowed the physicians to be confident that the patient was compliant with the medication hence his oxycodone dose was optimized to control pain.

Buprenorphine Buprenorphine is N-­dealkylated mainly by CYP3A4 and CYP2C8 to nor-­buprenorphine, the active metabolite. Since CYP3A4 is the main isoform involved, drug-­drug or herbal-­drug interactions are observed influencing the disposition of buprenorphine in humans. Buprenorphine and nor-­ buprenorphine undergo extensive phase II metabolism, catalyzed by UGTs, namely by UGT1A1, UGT2B7 (buprenorphine), and UGT1A3 (nor-­ buprenorphine). UGT1A1 accounts (in pooled HLM) for approximately 10 and 30% of buprenorphine and nor-­buprenorphine glucuronidation, respectively, whereas UGT2B7 is responsible for at least 40% of buprenorphine glucuronidation. However, it is assumed that UGT1A1 and UGT2B7 does not influence the disposition of buprenorphine in humans.

Tramadol Tramadol requires metabolism to O-­demethyl-­tramadol, the active metabolite providing analgesic effects via agonism at the μ-­ opioid receptor. Tramadol itself shows non-­ opioidergic

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Delta-­ 9-­ tetrahydrocannabinol (THC), the primary hallucinogenic constituent of Cannabis sativa, is subject to hepatic metabolism primarily by hydroxylation. The primary metabolite 11-­hydroxy-­THC (HO-­THC), which is pharmacologically active, is formed by CYP2C9 and further oxidized, probably by alcohol dehydrogenase or microsomal alcohol oxygenase, to the intermediate aldehyde 11-­oxo-­THC followed by oxidation to 11-­nor-­9-­ carboxy-­ THC (THC-­ COOH) catalyzed by a microsomal aldehyde oxygenase, a member of the CYP2C subfamily. After glucuronidation of the carboxy group, THC-­COOH is excreted in urine and can be detected up to 4 days after smoking 1 marijuana cigarette and up to 4 weeks after frequent use. Formation of hydroxy-­ THC and the epoxide are catalyzed by CYP3A4. 8-­ THC-­OH is conjugated by UGT1A9 and UGT1A10 isoforms whereas THC-­COOH is a substrate recognized by UGT1A1 and UGT1A3. The resulting glucuronide of THC-­COOH is the main metabolite found in urine, and thus these hepatic enzymes play a critical role in the metabolic clearance of cannabinoids.

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The metabolism of heroin to 6‑acetylmorphine (6‑AM) and morphine is catalyzed in the liver mainly by the human cholinesterase (hCE)-­1 and in part by hCE-­2, in the serum by pseudocholinesterase, and also non-­enzymatically in the serum. Morphine itself is conjugated mainly by UGT2B7 to the inactive metabolite morphine-­3-­glucuronide (M3G) and, to a less extent, to the pharmacological active compound morphine-­6-­glucuronide (M6G). Finally, morphine is N‑demethylated to nor morphine by hepatic CYP3A4 and to a lesser extent by CYP2C8. People with Gilbert’s syndrome, characterized by impaired glucuronidation due to a polymorphism in the gene encoding UGT1A1, do not show altered morphine clearance or difference in the plasma concentration versus time curves for M6G or M3G. The discussion on the effect of UGT2B7 polymorphisms on M6G and M3G formation and clearance is still controversial. For instance, modifications in ABCB1 are associated with significantly lower frequency of fatigue and the frequency of vomiting and the frequency of nausea is higher in patients with modifications of UGT2B7.

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inhibition of CYP3A4 activity by clarithromycin and voriconazole other medications and a transient reduction in renal function.

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analgesic effect by serotonin and norepinephrine reuptake inhibition. The O-­demethylation of tramadol is primarily catalyzed by CYP2D6, whereas N-­demethyl-­tramadol is formed by CYP2B6 and CYP3A4. The impact of CYP2D6 phenotype on tramadol pharmacokinetics is similar after single oral, multiple oral, and intravenous administration showing significant pharmacokinetic differences between EM and PM. Furthermore, metabolism by CYP2D6 seems to be stereospecific in the way that very little (+)-­O-­demethyl-­ tramadol is produced by CYP2D6 PM. The response to tramadol in patients is correlated to CYP2D6 genotypes and non-­response rates to pain medication increases fourfold in PM and thus this genotype is associated with poor efficacy of tramadol analgesia.

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Codeine (3-­methylmorphine) is metabolized to morphine by O-­ demethylation catalyzed by polymorphically expressed CYP2D6 as it is also the case for its congeners dihydrocodeine, ethylmorphine, hydrocodone, or oxycodone as described above. Some variants of the CYP2D6 gene increase metabolism of these drugs to their more potent metabolites, whereas others decrease metabolism, which may influence the analgesic potency and abuse liability. Finally, codeine is N-­ demethylated by CYP3A4 and conjugated equally by UGT2B4 and UGT2B7 to codeine-­ 6-­ glucuronide. Children of breastfeeding mothers who are CYP2D6 UMs combined with reduced UGT2B7 activity are at increased risk of potentially life-­threatening CNS depression. In the following, an interesting example for the impact of pharmacogenomics will be given. After small doses of codeine, a patient developed a life-­threatening opioid intoxication. CYP2D6 genotyping showed that the patient had three or more functional alleles, a finding consistent with ultra-­rapid metabolism of codeine. The authors attributed the toxicity to this genotype, in combination with

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Cocaine, the main alkaloid of Erythroxylum coca, is a powerful stimulant that is primarily metabolized by the 3 esterases pseudocholinesterase, human carboxylesterase-­1 (hCE-­1), and human carboxylesterase-­2 (hCE-­2). Cocaine is hydrolyzed mainly by hCE-­1 to benzoylecgonine, the primary metabolite excreted in urine, or by pseudocholinesterase and hCE-­2 to ecgonine methyl ester. In the presence of ethanol, hCE-­1 catalyzes transesterification of cocaine to cocaethylene, a toxic metabolite that can be further hydrolyzed by hCE-­1 to benzoylecgonine or by hCE-­2 to ecgonine ethyl ester. In contrast, it was shown recently that cocaine is not hydrolyzed by the human liver hCE1, neither as highly active recombinant protein nor as enzyme isolated from human liver or intestinal extracts. These results correlate well with computer-­assisted molecular modeling studies that suggest that hydrolysis of cocaine by hCE-1 would be unlikely to occur. Cocaine is a substrate of the intestinal ChE, as determined using both the recombinant protein and the tissue fractions. Again, these data are in agreement with the modeling results. The first step in the oxidative metabolism of cocaine is N-­demethylation to pharmacologically active norcocaine mainly by CYP3A4. Norcocaine is further metabolized by N–hydroxylation catalyzed by several CYP enzymes. There is widely differing activities of human cholinesterase variants with a tenfold lower binding efficiency for cocaine and tenfold lower catalytic efficiency. This leads to the possibility that individual responses to cocaine may be mediated in part by greater or lesser metabolic capacity, genetically determined variations in cholinesterase activity, or by the activity of the carboxylesterase.

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Amphetamine and Methamphetamine

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In general, six different chemical processes are important in the metabolism of amphetamines. These are mainly aromatic 4-­hydroxylation, N-­dealkylation, and oxidative deamination and to a minor extent, aliphatic beta-­hydroxylation, N-­oxidation, and conjugation of the nitrogen. Amphetamine is metabolized by the ­hydroxylation of the benzene ring at the 4-­position to 4-­hydro xyamphetamine, and beta-­hydroxylated to 4-­hydroxynorephedrine. Additionally, amphetamine is converted to phenylacetone and subsequently oxidized to benzoic acid. Methamphetamine is metabolized by hydroxylation and N-­ demethylation to form 4-­hydroxymethamphetamine, also known as pholedrine, and to amphetamine, respectively. Ring hydroxylation of amphetamine and methamphetamine is catalyzed by CYP2D6 and inhibitors of CYP2D isozymes can block approximately 90% of this reaction. The N-­demethylation of methamphetamine is also catalyzed by this isoform. Interestingly, amphetamine and methamphetamine are substrates as well as competitive inhibitors of CYP2D6. The oxidative deamination is catalyzed by monoamine oxidase (MAO). This deamination to phenylacetone appears to be also catalyzed by CYP2C hence playing a major role in the deamination of amphetamine. Beta-­ hydroxylation is carried out by dopamine-­ β-­ hydroxylase, which reacts stereoselectively with the S-­enantiomer of amphetamine and methamphetamine. The S-­enantiomers of amphetamines are generally more rapidly metabolized than corresponding R-­enantiomers due to enantioselective N-­demethylation and beta-­hydroxylation.

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3,4-­Methylenedioxymethamphetamine (MDMA) undergoes two main metabolic pathways. The predominant pathway in humans involves multiple CYP enzyme-­catalyzed O-­demethylenation of MDMA to 3,4-­dihydroxy-­methamphetamine (DHMA), followed by catechol-­ O-­methyltransferase (COMT)-­catalyzed O-­ methylation, primarily to 4-­hydroxy-­3-­methoxymethamphetami ne (HMMA). DHMA and HMMA may also be conjugated by UGT to DHMA 3-­ glucuronide, DHMA 4-­ glucuronide, and HMMA glucuronide, or by SULT to DHMA 3-­sulfate, DHMA 4-­sulfate, and HMMA sulfate. A minor pathway includes demethylation to 3,4-­methylenedioxyamphetamine (MDA) followed by demethylenation to 3,4-­ dihydroxyamphetamine (DHA), O-methylation to 4-­hydroxy-­3-­methoxyamphetamine (HMA), and respective conjugation. Besides this, MDMA is known to be a potent mechanism-­based inhibitor of CYP2D6  which is also assumed to influence MDMA-­ induced neurotoxicity. DHMA was also shown to inhibit its own metabolism as well as the methylation of dopamine. Additionally, different pharmacokinetic properties were observed for the two MDMA enantiomers. The S-­enantiomer is eliminated from plasma at a higher rate than the R-­enantiomer most likely explained by stereoselective metabolism. Concerning CYP-­ demethylenation to DHMA, the initial metabolic step, marked enantioselectivity toward S-­MDMA was observed for CYP2C19 and slight preferences for CYP2D6. COMT-­ methylation of DHMA to HMMA and CYP-­N-­demethylation to MDA demonstrate similar selectivity. HMMA glucuronidation by UGT1A9 is markedly stereoselective with preference for the formation of the S-­enantiomer glucuronide, whereas glucuronienantiomer. UGT2B15 and dation by UGT2B7 favors the R-­ UBT2B17 reveal only slight preferences for S-­HMMA. In human HMMA can be liver microsomes, slight preferences for S-­ observed. Sulfation of HMMA is mainly catalyzed by SULT1A3 and to a minor extent by SULT1E1. Enantiomeric preferences cannot be observed for SULT1A3 or human liver cytosol. On the other hand, the efficiency for S-­DHMA 3-­sulfate formation is twice as high as for its R-­enantiomer, both in SULT1A3 and human liver cytosol. One reason for this difference in enantioselectivity might be the position for sulfation. DHMA is mainly sulfated in position 3, whereas HMMA can only be sulfated in position 4. Monitoring of stereoselective elimination of MDMA and its main metabolites in vivo also exhibited changes in enantiomeric disposition over time. MDMA, DHMA, and HMMA sulfate revealed preferences for the R-­stereoisomer elimination,

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conversely all other metabolites showed more S-­isomer within the first 24 h after ingestion. In the later excretion phase (after 24 h), R/S ratios were >1 for all compounds. Generally, changes in the R/S ratios over time could be useful for estimation of ingestion time. The use of R/S cut-­off values for different MDMA metabolites was shown to be useful for rough estimation of ingestion time and to distinguish between recent (within 24 h) or earlier MDMA consumption in human urine. R/S cut-­offs ≥ 2 for MDMA, HMMA sulfate, and HMMA glucuronide, and ≥1 for MDA, HMMA, and DHMA sulfate correctly predicted time of ingestion in more than 87% of all samples.

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The main metabolite benzoylecgonine can be detected in urine for 2–3 days after an intranasal dose of 100 mg of cocaine. The detection time of cocaine itself in blood is approximately 4–6 h after a dose 20 mg and approximately 12 h after a dose of 100 mg. In the serum of chronic users, benzoylecgonine is detectable for 5 days on average.

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51.8 Excretion Xenobiotics can be eliminated from blood via two main routes: the so-­called renal and hepatic elimination. In the latter route, drug elimination consists of two processes, metabolism and renal or biliary excretion of the metabolites. More polar xenobiotics that are not reabsorbed in the kidneys are eliminated by direct renal filtration. Some volatile compounds such as ethanol or gaseous anesthetics leave the body mainly by exhalation. Smaller amounts of xenobiotics can be excreted also via sweat or milk. The latter can be risky for the breastfed baby.

Akhtar, N., Ahad, A., Khar, R.K. et  al. (2011). The emerging role of P-­glycoprotein inhibitors in drug delivery: A patent review. Expert Opinion on Therapeutic Patents 21 (4): 561–576. Caldwell, J., Gardner, I. and Swales, N. (1995). An introduction to drug disposition: the basic principles of absorption, distribution, metabolism, and excretion. Toxicologic Pathology 23 (2): 102–114. Crisafulli, C., Fabbri, C., Porcelli, S. et al. (2011). Pharmacogenetics of antidepressants. Frontiers in Pharmacology 2 (6): 1–6. Jancova, P., Anzenbacher, P. and Anzenbacherova, E. (2010). Phase II drug metabolizing enzymes. Biomedical papers of the Medical Faculty of the University Palacky, Olomouc, Czechoslovakia Republic 154 (2): 103–116. Meyer, M.R. (2018). Toxicokinetics of NPS: Update 2017. Handbook of Experimental Pharmacology 252: 441–459. Ritter, J.M., Flower, R.J., Henderson, G. et  al. (2019). Rang and Dale’s Pharmacology, 9th edn. Edinburgh: Elsevier. Ritschel, W.A. and Kearns, G.L. (2009). Handbook of Basic Pharmacokinetics Including Clinical Applications, 7th edn, Washington, DC: American Pharmacists Association. Smith, H.S. (2009). Opioid metabolism. Mayo Clinic Proceedings 84: 613–624.

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The impact of individual genetic variations is given in all stages of the LADME process. Such variations have to be considered especially when interpreting analytical results in CT and particularly in FT. They may have an impact on conclusions drawn from e.g. blood plasma concentrations or urinary concentrations of certain compounds with respect to the amount of ingestion. Furthermore, such variation may influence the time of detection of drugs and in consequence may lead to misinterpretation of the time of ingestion.

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51.9 Conclusions

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References

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Maurer, H.H., Sauer, C. and Theobald, D.S. (2006). Toxicokinetics of drugs of abuse: Current knowledge of the isoenzymes involved in the

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Further reading

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Hepatocytes can transfer xenobiotics from blood plasma to the bile for the purpose of elimination from systemic circulation. Mainly more bulky and hydrophobic drugs are underlying this biliary excretion. As depicted in Fig. 1, glucuronides or sulfates of certain drugs can be excreted into bile and transferred to the intestine. Endogenous bacteria play an important role in this enterohepatic circulation providing glucuronidases and arylsulfatases that hydrolyzes glucuronide and sulfate conjugates. The free compounds can then be reabsorbed. This process is named enterohepatic circulation. This recirculating drug can amount to about 20% of total drug in the body and therefore dramatically prolong drug action. This is for instance of importance for drugs such as digitoxin, tamoxifen, rifampicin, carbamazepine, barbiturates, and oral contraceptives. Moreover, beside therapeutic drugs, this is also of importance for some toxic compounds such as colchicine (ingredient of colchicum autumnale, also known as “meadow saffron”) or amatoxins from amanita phalloides.

human metabolism of tetrahydrocannabinol, cocaine, heroin, morphine, and codeine [review]. Therapeutic Drug Monitoring 28 (3): 447–453. Meyer, M.R. and Maurer, H.H. (2010). Metabolism of designer drugs of abuse: An updated review [review]. Current Drug Metabolism 11 (5): 468–482. Meyer, M.R. and Maurer, H.H. (2011). Absorption, distribution, metabolism and excretion pharmacogenomics of drugs of abuse [review]. Pharmacogenomics 12 (2): 215–233. Meyer, M.R. and Maurer, H.H. (2012). Metabolism of amphetamine derivatives and their derived designer drugs: An updated review [review]. Drug Metabolism Reviews in preparation.

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Toxicology of Specific Substances

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A.W. Jones, F. Musshoff, T. Krämer, A. E. Steuer, D. Gerostamoulos, O.H. Drummer, G. Drasch, M. Balikova, J. Beyer, H. Teixeira, M. Thevis, W. Schänzer, H. Druid, and G. Skopp

Light-­to-­moderate consumption of alcoholic beverages, such as 10–20 g of ethanol daily, or the amount contained in 1–2 small glasses of wine, should not have any adverse effects on a person’s health and longevity. Epidemiological surveys of people’s drinking habits support the notion that consumption of a glass or two of red wine daily is a useful prophylactic treatment against cardiovascular diseases, such as ischemic stroke (Mukamal et al. 2005). However, for 10–15% of the population, especially among men, moderate drinking often escalates into heavy drinking and dependence, having many negative consequences for health and longevity (Room et al. 2005). Elevated blood-­ethanol concentration is found in 40–60% of unnatural deaths, including all types of suicide (Holmgren and Jones 2010), accidental drownings (Pajunen et  al. 2018), and violent deaths in general (Cvetkovic et al. 2017). Scientific research on the disposition and fate of ethanol in the body began in the 1920s when a reliable analytical method was developed that allowed quantitative determinations in micro-­ volumes of blood (Widmark 1922). Hundreds of controlled alcohol dosing studies provided reliable information about factors influencing the absorption, distribution, metabolism, and elimination (ADME) of ethanol in the body. By contrast, human dosing studies with other aliphatic alcohols are problematic, owing to the toxicity of some of the metabolites produced (Kraut and Kurtz 2008). Nevertheless, useful information can be gleaned from clinical case reports of patients poisoned by drinking the more toxic alcohols, such as methanol or ethylene glycol (Henderson and Brubacher 2002; Kraut 2015). Such poisonings often occur

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Alan Wayne Jones

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According to Paracelsus (1493–1541), “What is there that is not poison? All things are poison, and nothing is without poison. Solely the dose determines that a thing is not a poison” (Deichmann et al. 1986). This much quoted axiom is appropriate to describe the aliphatic alcohols because trace amounts are produced endogenously by various enzymatic or dietary processes, whereas the same substances are highly dangerous to human health if and when ingested (Kraut and Mullins 2018). The alcohol of major interest in forensic toxicology is ethanol (CH3CH2OH) or grain alcohol because this is the psychoactive substance contained in all alcoholic beverages. During forensic-­ medical investigations of living and deceased persons, the drug most often identified in biological specimens submitted for analysis is ethanol (Jones 2019a, 2019b). Many crimes are committed when people are under the influence of alcohol, such as drunk driving (Jones et  al. 2019), drug-­ facilitated sexual assaults (Kerrigan 2010), domestic violence, vandalism, and other aggressive behavior (Dingwall 2013). Ethanol intoxication and drunkenness are underlying factors in many fatal poisonings, either alone or combined with other psychoactive substances ingested together with ethanol (Jones et al. 2011; Ketola and Ojanpera 2019). The quantitative analysis of ethanol in blood, urine, and other biological specimens is the most frequently requested service for forensic toxicology laboratories when sudden and unnatural deaths are investigated (Jones and Holmgren 2003). Handbook of Forensic Medicine, Volume 2, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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when people buy alcohol produced illegally, which contains dangerously high concentrations of methanol, and hundreds of deaths worldwide (Rostrup et al. 2016). Outbreaks of methanol poisoning tend to occur in economically deprived parts of the world where, for various reasons, legitimate alcoholic beverages are unavailable, thus tempting people to purchase clandestine alcoholic drinks (Paasma et al. 2007; Ahmed et al. 2017). Forensic practitioners need to appreciate the many factors influencing ADME of ethanol and the clinical features associated with consumption of the more toxic alcohols (Jones 2016; Perry et  al. 2017). The treatment strategy when dealing with poisoned patients depends on the nature of the alcohol consumed and the clinical symptoms of intoxication, including the concentration determined in a sample of blood and elapsed time since ­ingestion. Life-­saving treatment includes administration of an antidote, treatment of metabolic acidosis clearing toxins from the blood by hemodialysis (Hassanian-­Moghaddam et al. 2019). This article presents an update of the ALCOHOLS section from the first edition of Handbook of Forensic Medicine, although the title is now altered to ALIPHATIC ALCOHOLS. The main focus remains the same, namely the forensic pharmacology and toxicology of ethanol, because interpreting blood-­alcohol concentrations (BACs) is a major task for forensic practitioners. Furthermore, most nations enforce statutory BAC limits for driving (ranging from 0.20 to 0.80 g/L), which requires that the analytical methods are accurate, precise, and fit for legal purposes (Jones et al. 2019). During the prosecution of drunken drivers, a suspect’s BAC often needs to be translated into the amount of ethanol consumed or back-­ extrapolation of the BAC to an earlier time, such as the time of driving, which requires detailed knowledge of ethanol’s pharmacokinetics (Jones 2011).

52.2  Chemistry of alcohols

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The aliphatic alcohols represent a family of organic compounds that contain one or more hydroxyl groups (-­OH). The latter is bonded to a saturated or unsaturated carbon atom. Textbooks devoted to organic chemistry usually denote alcohols with the general formula R-­OH, where R stands for an alkyl group or hydrocarbon side chain and -­OH is the functional hydroxyl group. The chemical structures of aliphatic alcohols can be classified as primary, secondary, or tertiary depending on the number of alkyl groups bonded to the carbon atom joined to the hydroxyl group (see Table 52.1.1). Aliphatic alcohols can also be considered as derivatives of water with one of the hydrogen atoms replaced with a saturated hydrocarbon group or, alternatively, as hydrocarbons with one or more of the hydrogen atoms replaced by hydroxyl groups. Another way to classify aliphatic alcohols is by the number of hydroxyl groups in each molecule, such as mono-­hydroxy (ethanol and methanol), di-­ hydroxy (ethylene glycol and 1.4-­ butanediol), tri-­ hydroxy (glycerol), or poly-­hydroxy (mannitol and sorbitol). Alcohols undergo a wide range of chemical reactions as expected from the hydroxyl group in the molecule and examples include halogenation to produce alkyl halides, reduction to ethers and hydrocarbons, oxidation to aldehydes and carboxylic acids, and the formation of esters in reactions with acids. This chapter is concerned with one particular biochemical reaction, namely oxidative metabolism in the liver by two polymorphic enzymes alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) (Agarwal and Goedde 1987). The liver also contains a microsomal enzyme denoted as CYP2E1, which has a higher km than ADH and therefore becomes engaged in ethanol metabolism at high BAC, such as occurs in heavy drinkers (Zakhari 2006).

OH

OH

Methanol

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H3C

Mono-­hydroxy secondary alcohol

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Mono-­hydroxy primary alcohols

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Table 52.1.1  Chemical structures of low-­molecular weight aliphatic alcohols.

H3C

H3C CH3

Isopropanol 2-propanol

H3C

OH

n-Propanol

H3C

OH CH3

Di-­hydroxy alcohols HO

OH

OH OH

Glycerol

t-Butanol O

OH

Diethylene glycol OH

Tri-­hydroxy alcohol HO

Ethylene glycol

HO

Ethanol

H3C

Mono-­hydroxy tertiary alcohol

HO 1,4-Butanediol

OH

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Qualitative and quantitative determination of alcohols in biological fluids involves relatively simple analytical procedures, and the necessary equipment is generally available in all clinical and forensic toxicology laboratories (Emerson 2004). Although various analytical methods have been used over the years, the gold standard today for legal purposes is gas–liquid chromatography (GC) with a flame ionization detector (FID), either by direct liquid injection or by the headspace (HS) sampling technique (Jones 2020a). HS analysis entails dilution of an aliquot of blood or other liquid samples with an internal standard and allowing this to equilibrate in an airtight glass vial at a constant temperature of 50oC or 60oC for about 20 min (Seto 1994). Thereafter, a sample of the vapor phase above the blood is removed with a gastight syringe or some automated system and injected into the GC instrument for analysis. Any volatile substances in the blood sample distribute between the air and liquid phases in the vial according to their partition coefficients. The concentration in the air phase is directly proportional to the liquid phase concentration at equilibrium (Tiscione et al. 2013). Modern gas chromatographs are equipped with automated headspace sampling routines, which are necessary for use at laboratories with large workload of samples from police investigations of traffic crimes and postmortem examinations (Jones 2014). The main advantage of sampling the vapor in equilibrium with blood instead of injection of an aliquot of blood is that the injection block and GC column are not clogged by non-­volatile constituents of the blood, such as fats and proteins. Methods that require making liquid injections of blood specimens usually incorporate a heated glass liner or short pre-­column before the main GC column, which serves to trap non-­volatile constituents that would otherwise cause problems and deterioration of the stationary phase. A robust and well-­proven analytical procedure for the determination of ethanol in blood for legal purposes is described below (Jones and Schuberth 1989). The tubes containing anticoagulated blood samples are first carefully rotated five or six times to ensure that plasma fraction and red cells are well mixed. Thereafter, the stoppers on the tubes are removed and an aliquot

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52.4  Determination in body fluids

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The low-­molecular aliphatic alcohols discussed in this chapter are all completely miscible with water, and there is no evidence that they bind to plasma proteins. After ingestion, these alcohols distribute into the water compartment of the body and the concentrations reached in different organs and tissues are in direct proportion to their relative water contents. Table 52.1.2 summarizes the physicochemical properties of aliphatic alcohols discussed in this chapter.

(100 μL) of blood specimen is diluted 1 + 10  with an aqueous solution of an internal standard (IS). The HS-­GC-­FID method is applicable to many types of biological specimens, including urine and brain tissue (O’Neal et al. 1996). Dilution of the blood specimen with IS is done using diluter– dispenser equipment available from various manufacturers. Dilution of the blood specimen in a ratio of 1 + 10 is necessary to eliminate matrix effects thus allowing the use of aqueous ethanol solutions for calibration of the GC instrument and as quality control specimens (Alonso et al. 2013). If the blood specimens are from living subjects, such as apprehended drovers, then n-­ propanol is suitable as the IS, whereas t-­butanol is preferable for autopsy blood specimens because small amounts of n-­propanol might be produced in the body after death by fermentation processes (Levine et al. 2020). Identification of the volatile substance in the blood samples (qualitative analysis) is achieved by comparing their retention times (RT) with those of authentic substances analyzed under the same conditions. RT is defined as time elapsed after injection into the GC instrument until the appearance of a peak response on the gas chromatogram. The concentration of ethanol in blood (quantitative analysis) is done by measuring the areas under the peaks on the GC trace by electronic integration. The peak area ratios of ethanol-­ to-­ internal standard is then determined and plotted against concentration in the known standard to construct a calibration curve. The flame ionization detector (FID) is highly sensitive and gives a linear response over a wide range of concentration of the substance injected into the GC, e.g. 0.10 to 5.0 g/L for ethanol. Although not necessary for routine forensic blood-­ethanol determinations, the sensitivity of the HS-­GC analysis can be increased by adding an inorganic salt, such as 1–2 g of sodium carbonate or sodium chloride to saturate the blood sample (Machata 1975). This raises the vapor pressure of non-­electrolytes in the sample and leads to higher concentrations in the headspace vapor at equilibrium. The accuracy of the known strength ethanol standard ­solutions used to calibrate the instrument is important, and these should be purchased from a reliable and traceable source. Control of the calibration during an analytical run is done with in-­house prepared ethanol standards. These can be prepared from absolute ethanol by weighing and making a 10% w/v stock solution. The stock solution is kept in a refrigerator (4oC) and is stable for several months. From this, working standards are made by serial dilutions to give the required concentration range encountered in the biological samples. Because the concentrations of ethanol in blood are important in criminal prosecutions, such as drunk-­driving cases, quality assurance is important, and all determinations should be made in duplicate or triplicate on two different GC systems that yield ­different retention times for ethanol (Sklerov and Couper 2011). Otherwise, two independent methods of analysis might be used, such as GC analysis and enzymatic oxidation. The s­ tandard operating procedure for blood-­ ethanol analysis at legal medicine

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Table 52.1.2  Physicochemical properties of low-­molecular weight aliphatic alcohols often encountered in forensic science and legal medicine.

Chemical formulae Structure

32.04 CH3OH Primary aliphatic alcohol

71-­23-­8

60.09

62.07

106.1

(CH2OH)2

(C2H4OH)2O

Primary aliphatic alcohol

Primary aliphatic alcohol

Secondary aliphatic alcohol

Aliphatic diol

Aliphatic diol

Propyl alcohol

Rubbing alcohol

Antifreeze solvent

DEG

82.5

197

245

34

Solubility in water

Miscible

 Chemical abstract service registry number.  Measured at 20oC and atmospheric pressure.

R FO

−12.9

−10.4

1.11

1.12

16.0

16.0

16.5

15.1

14.3

0.6–0.7

0.6–0.7

0.6–0.7

0.6–0.7

0.6–0.7

24

18

18

17

10

Miscible

Miscible

Miscible

Miscible

Miscible

C

1

−88.5 0.785

TR IB

Serum osmolality mOsm/kg H20

−126.5

0.805

N

0.6–0.7

82.6

0.789

O

15.5

Volume of distribution, L/kg

−114.1

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78.5

Acidity (pKa)

111-­46-­6

(CH3)2CHOH

64.7 −95.8

Diethylene glycol

107-­21-­1

60.09

Boiling point oC

0.791

Ethylene glycol

CH3CH2CH2OH

Beverage or grain alcohol

Melting point C

67-­63-­0

46.07

Wood alcohol

Density g/mL2

Isopropanol

CH3CH2OH

Common name

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n-­propanol

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Molecular weight g/mol

Ethanol 64-­17-­5

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Methanol 65-­46-­1

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Table 52.1.3  Results of a declared external proficiency test among clinical chemistry laboratories in Sweden when required to determine toxic alcohols and acetone in plasma. Substance in plasma

N1

Target value g/L

All laboratories mean ± SD, g/L

CV %

Ethanol

40

2.44

2.41 ± 0.165

6.8

Methanol

31

1.92

1.84 ± 0.171

9.3

Isopropanol

29

0.72

0.69 ± 0.023

3.3

Acetone

25

1.85

1.66 ± 0.143

8.6

Ethylene glycol

16

1.24

1.26 ± 0.157

12.4

 Number of participating laboratories and all used gas chromatographic methods.

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deviation/mean × 100), were less than 10%, except for ethylene glycol, which is generally considered more difficult to determine (Jones and Hård 2004). An inter-­laboratory comparison of blood-­alcohol determinations at laboratories in Italy was recently published (Zamengo et al. 2019). The latter was also a declared trial and the laboratories knew that they were being evaluated, which has led to calls to organize “blind” proficiency studies in the future (Mejia et  al. 2020; Pierce and Cook 2020). Obtaining satisfactory results from participation in such proficiency testing schemes has become an important part of gaining laboratory accreditation. A wide range of biological specimens might be submitted for analysis in forensic casework, including blood, plasma/ serum, vitreous humor, urine, and cerebrospinal fluid, and the same HS-­ GC-­ FID method of analysis can be used (Thelander et  al. 2020). However, because these biological specimens contain different amounts of water, they are also expected to contain different concentrations of ethanol. Specimens of plasma/serum contain about 92% w/w water compared with 80% w/w for whole blood, so the serum/blood ratios of ethanol are expected to be 1.15:1 (range 1.10–1.20) depending on the hematocrit of the blood sample (Iffland et al. 1999). In a post-­mortem study, there was considerable uncertainty involved when BAC was estimated indirectly from the concentration of ethanol determined in an alternative biological specimen, such as bile, vitreous humor or CSF (Thelander et al. 2020).

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l­aboratories in Germany involves making two determinations by GC and two by an enzymatic method (Krause 2007). The average of the four determinations of BAC is then calculated and used as evidence in drunk-­driving cases. In other countries, such as Sweden, duplicate determinations of BACs are made by two technicians who work independently with two different GC instruments and column packing material. The stationary phases have different polarities and yield different RTs for ethanol. If the difference between the two results is less than 10%, the mean of the two determinations is calculated and a deduction made (e.g. 6%) to allow for analytical uncertainty in the method of analysis (Jones and Schuberth 1989). Some laboratories seem to be moving toward the use of a ­GC-­MS method for ethanol analysis in their routine forensic casework (Tiscione et al. 2011). In addition to a FID detector, a mass spectrometer (MS) detector is used to enhance the identification of ethanol by looking for its characteristic mass fragmentation pattern; m/z 31 (base peak), m/z 46 (molecular ion), and m/z 45 (Cordell et al. 2013). This improves the qualitative identification of ethanol compared with relying only on RT, although one might wonder whether this more sophisticated analytical technology is really necessary for a substance such as ethanol in blood, which is present at such high concentrations in most forensic cases (Xiao et al. 2014). Headspace sampling is not a practical alternative for determination of aliphatic diols because of their higher boiling points (see Table  52.1.2). However, liquid injection is possible or chemical derivatives can be made prior to analysis (Hlozek et  al. 2014). Advantages and limitations of the various methods used for the analysis of ethylene glycol in clinical and forensic chemistry laboratories were recently reviewed (Porter 2012). A simple method for the determination of ethylene glycol in postmortem blood entailed precipitation of the proteins, centrifugation, and injecting an aliquot of supernatant directly into a GC-­FID instrument fitted with a capillary column (Jonsson et  al. 1989). Another method suitable for the analysis of aliphatic diols is by making a chemical derivative, such as the butyl or phenylboronic ester, prior to the GC-­FID analysis (Porter and Rutter 2010). Alternative derivatives, such as the trimethylsilyl esters, have also been described for the analysis of aliphatic diols. The internal standard for GC-­FID analysis is usually another glycol, such as 1,2-­propanediol, 1,3-­propanediol, or 2,3-­butanediol. Table  52.1.3 presents the results of a declared inter-­ laboratory proficiency trial at clinical chemistry laboratories in Sweden where GC methods of analysis were used for analysis of plasma spiked with known amounts of various alcohols (Jones et al. 1995). All laboratories were aware that they were participating in an external proficiency test of their performance. A good agreement was seen between the mean analytical result reported by all laboratories and the target or known concentration of the alcohol prepared. The between-­laboratory precision, as reflected in coefficients of variation (standard

52.5  Disposition and fate in the body Because aliphatic alcohols are more soluble in water than in lipids (Table  52.1.2) and total body water makes up  50–60% of body weight, large amounts of the alcohols must be ingested to increase BAC and cause toxicity. For example, feelings of mild euphoria would require drinking a bolus dose of 10–20 g ethanol (10– 20,000 mg), which compares with 5–10 mg of diazepam for an

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Absorption of alcohols can occur via the skin, the lungs (inhalation), or by ingestion via the mouth. Although only very small quantities of ethanol can penetrate the skin and enter the pulmonary blood by inhalation, by far the most common route of administration is oral ingestion. On reaching the stomach and intestine (duodenum and jejunum), the alcohols are rapidly absorbed into the portal venous blood. They first need to pass through the liver, then move on to the heart and lungs, back to the heart, before being distributed throughout the entire body. Alcohols readily cross the blood-­brain barrier and distribute into body fluids and tissues according to their relative water contents. Urine, saliva, sweat, and cerebrospinal fluid (~100  % water) are expected to contain higher concentrations of ethanol than blood (~80% w/w water) after equilibration is reached (Jones 2008). Organs with a rich blood supply, such as heart, brain, and kidneys, rapidly equilibrate with the ethanol concentration in the arterial blood. By contrast, more poorly perfused tissue, such as the resting skeletal muscle, takes longer to reach diffusion equilibrium with ethanol in the arterial blood. This creates a concentration gradient between the concentrations of ethanol in arterial blood and venous blood returning to the heart. When a bolus dose (0.6 g/kg) of ethanol was consumed on an empty stomach, the arterial BAC was higher than the venous BAC by as much as 0.25 g/L and this  arterio-­venous difference gradually decreased as time after drinking increased. On reaching the post-­absorptive phase, the venous BAC was slightly higher than the arterial BAC by about 0.02–0.08 g/L (Jones et al. 2004). The distribution volume (Vd) of a drug represents the relationship between the amount of substance in the entire body to the concentration in a sampling compartment, such as blood or plasma. For water soluble drugs, such as ethanol, its distribution volume Vd can be calculated from information about the water content of the whole body (~50–60 % w/w) and the blood (~80% w/w), which suggests a range of values from 0.63 to 0.75  L/kg (Jones et  al. 1992). However, in any given individual, Vd will depend on the person’s age, gender, and body composition, ­especially the proportion of lean to fatty tissue in the body. People suffering from obesity are expected to have a smaller Vd for ­ethanol because they have less water/kg body weight to dilute

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Although under normal circumstances the endogenous concentrations of ethanol in blood and other body fluids lack any clinical or forensic significance, there are certain rare exceptions. Trace amounts of ethanol normally produced in the gut by fermentation of dietary carbohydrates undergoes first-­pass metabolism as the portal venous blood flows through the liver. However, there is a medical condition referred to as “auto-­ brewery syndrome (ABS)” or “gut-­fermentation syndrome.” This implies that the human body produces abnormally high concentrations of ethanol that overwhelm the oxidative capacity of liver enzymes. As a consequence, BAC increases, sometimes appreciably to cause the typical signs and symptoms of drunkenness without having consumed any alcoholic beverages. This medical condition was first reported in Japanese patients, many of whom suffered from various digestive problems, they had eaten carbohydrate-­rich meals and had also inherited a defective form of the liver enzyme aldehyde dehydrogenase, which is meant to metabolize acetaldehyde (Kaji et al. 1976). The prevalence of this medical condition in western society is not known precisely, although an increasing number of case reports appear in the medical literature (Akhavan et  al. 2019; Malik et al. 2019). The cause of ABS is linked to strains of yeast proliferating the gastrointestinal tract, which is often associated with gut pathology or an imbalance in the gut flora. Conditions such as short-­gut syndrome (Jansson-­Nettelbladt et al. 2006) and Crohn’s disease (Welch et al. 2016) resulted in the biosynthesis of higher-­than-­normal concentrations of endogenous ethanol. The confirmatory test for ABS is the accurate determination of ethanol in blood or breath after a glucose challenge test (Cordell and Kanodia 2015). Patient management is mainly based on treatment with antifungal drugs aimed at restoring the digestive flora and limiting the fermentation of dietary sugars (Saverimuttu et al. 2019). Alleged ABS is sometimes used as a defense argument in cases of drunk driving (Logan and Jones 2000). However, it is important to remember that fermentation is not an instantaneous process and people can metabolize between 6 and 8 g of

52.5.2 Absorption, distribution, and elimination

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ethanol per hour by metabolism in the liver. Furthermore, a theoretical calculation shows that a normal blood-­glucose content of 1 g/L after fermentation produces a BAC of about 0.5  g/L. During the fermentation process, large volumes of carbon dioxide gas are produced, which has to be emitted from the body in some way. Until more controlled studies are done, it would be wise to remain skeptical about the prevalence of ABS in ambulant healthy subjects who are capable of driving on the highway.

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anxiety disorder or 60–100 mg codeine as an analgesic or antitussive medication. The dosage form of ethanol depends on the type of alcoholic beverage consumed, and the concentrations of ethanol vary from 3 to 5 % v/v in beer, 8 to 14 % v/v in table wines, 15 to 22 % v/v in fortified wines and 35 to 55 % v/v in distilled spirits or liquor. Trace amounts of ethanol and methanol (0.5–1.5  mg/L) are produced naturally in the body by various enzymatic and/or micro-­biological reactions, such as fermentation of dietary carbohydrates in the colon (Sprung et  al. 1981; Haffner et  al. 1996). Other metabolic pathways also exist that generate trace amounts of acetaldehyde, and the latter gets reduced by alcohol dehydrogenase, to produce endogenous ethanol (Ostrovsky 1986).

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52.5.3  Alcohol metabolizing enzymes

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Enzymes are biological catalysis essential to permit intricate biochemical reactions to occur at a body temperature of 37oC. Drug metabolizing enzymes are mainly located in the liver and gut and function as a protection for the organism against xenobiotics and toxins ingested with the diet. The major biochemical pathway for removal of alcohols is oxidative metabolism by hepatic ADH, which is located in the cytosol fraction, and ALDH, which is located in the mitochondria. Ethanol is oxidized first to acetaldehyde by the action of ADH, and the acetaldehyde metabolite is further oxidized to acetic acid by the action of ALDH (Edenberg 2007). Low km ALDH ensures that the concentrations of the more toxic acetaldehyde metabolite in blood remains low during ethanol metabolism in most populations (e.g. Caucasians and African Americans). However, people of East Asian descent, such as Japanese, Chinese, Koreans, 30–40% of them inherit a defective form of the low km ALDH2*2 isoform (Crabb et  al. 1989). This makes them highly sensitive to acetaldehyde, and they tend to flush in the face and complain of tachycardia, nausea, and headaches after drinking small amounts of ethanol. This defective form of ALDH has been attributed to the substitution of a single amino acid, namely lysine substituted for glutamate in position 487 of the protein chain (Crabb et  al. 2004). Those diagnosed as being homozygous for this trait experience such unpleasant sensations after drinking alcoholic beverages that they become life-­ long abstainers, owing to the untoward effects and toxicity of ethanol’s metabolite acetaldehyde (Agarwal and Goedde 1992). Human ADH is encoded by different genes, and the associated alleles give enzymes with different characteristics, including Vmax and km and different specificities for substrates. These genetic differences in ADH probably account for the observed ethnic and racial differences in elimination rates of ethanol from blood and inter-­individual differences in the same population (Edenberg 2007). The ADH enzyme is polymorphic, and the principal variant involved in the oxidative metabolism of ethanol is class I ADH, which has a low km (0.05–0.1 g/L). This means that the enzyme is saturated with the substrate after the first few drinks (2 x km = 0.20 g/L) and provided BAC exceeds 0.2–0.3 g/L, the enzyme is working at its full capacity and the elimination kinetics of ethanol from blood is a zero-­order process (constant rate per unit time). Hepatic ADH is also responsible for the metabolism of other aliphatic alcohols, including methanol, 2-­propanol, and ethylene glycol, although the pharmacokinetics of these other alcohols is more in accordance with first-­order kinetics (rate increases with concentration in blood). This implies that a constant proportion of the drug is eliminated per unit time. Figure  52.1.2 shows chemical equations depicting the ADH mediated oxidative metabolism of ethanol, methanol, and isopropanol. Note that primary alcohols are oxidized first to aldehydes and then to carboxylic acids, whereas secondary alcohols, such as 2-­propanol, are converted to ketones. The acetone metabolite is

1.0

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ingested ethanol and therefore achieve a higher concentration in the blood (Jones 2007). The concentration–time profiles of ethanol are slightly different depending on whether samples of blood, plasma, saliva, or urine were used for analysis. The concentration–time (C–T) profiles derived from the analysis of plasma and serum (92 % w/w water) run on a higher level than whole blood (80% w/w water). The slope of the rectilinear elimination phase is slightly steeper if the ethanol was determined in plasma or serum compared to blood and the y-­intercept (C0) is also higher, which means that Vd of ethanol derived from plasma/serum kinetics is lower than for the analysis of whole blood. The distribution ratio for ethanol between serum and blood is 92/80 = 1.15:1, which is in good agreement with experimental studies on water distribution (Iffland et al. 1999). Figure  52.1.1 compares Vd of ethanol based on results from several hundred controlled alcohol dosing studies published in the literature. The raw data were carefully selected based on certain criteria, such as the minimum ethanol dose (0.30 g/kg), rapid ingestion on an empty stomach, and taking a sufficient number of blood samples in the post-­absorptive phase to permit calculating the slope of the elimination phase in a reliable way. The scatter plot in Figure 52.1.1 shows mean values of 0.70 L/kg (N = 173) for healthy men compared with 0.60 L/kg (N = 63) in healthy women (Maskell et al. 2019). Only relatively small amounts of aliphatic alcohols are excreted unchanged in breath, sweat, and urine. Roughly, 90% of the dose of ethanol and methanol undergoes metabolism in the liver, and the remaining 10% is excreted unchanged in breath, sweat, and urine. The aliphatic diols are not volatile enough at body t­emperature to be detected in the exhaled breath.

0.70 0.60

0.5 0.4 0.3 Males

Females

Figure 52.1.1  Scatter diagram and bar-­and-­whisker plots of the distribution volume of ethanol in healthy male and female subjects.

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Primary alcohols

Secondary alcohols

Ethanol

Methanol

NAD+

2-propanol NAD+

NAD+

ADH NADH

ADH

ADH NADH

NADH

Acetaldehyde NAD+

Formaldehyde

Acetone

NAD+

ALDH (rapid) NADH

CYP2E1 (slow)

ALDH (rapid)

H2O

CO2

N

CO2

Acetol

Formic acid

H2O

Other metabolites

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2016). The clinical efficacy of fomepizole for use in treating people admitted to hospital after poisoning with methanol or ethylene glycol has been reviewed elsewhere (Jacobsen and McMartin 1997). A scheme comparing the oxidative metabolism of ethanol and methanol is shown in Figure  52.1.3, and reduction of the coenzyme NAD+ into NADH, which has important metabolic consequences. The increased NADH/NAD+ ratio changes the redox state of the liver to a more reduced potential, which impacts on other NAD-­dependent biochemical reactions, leading to an increase in blood-­lactate and an accumulation of fat in hepatocytes. Also shown in Figure 52.1.3 are the chemical structures of two enzyme inhibitors, namely fomepizole (Antizol®), or 4-­methyl pyrazole and disulfiram (Antabuse®) an inhibitor of low km ALDH. The carboxylic acids formed (acetic and formic acids) undergo extrahepatic oxidation in peripheral tissues to carbon dioxide and water. Acetate enters the citric acid cycle via acetylCoA, whereas formic acid is more resistant to oxidation in humans, owing to a lack of folinic acid. Another important enzyme involved in the oxidative metabolism of ethanol is located in the microsomal fraction of hepatocytes, originally known as MEOS, which stands for microsomal ethanol oxidizing system. Later research identified MEOS as being a family of cytochrome P450 enzymes and CYP2E1 was the one involved in the oxidation of ethanol. Its higher km (0.6–0.8 g/L) compared with ADH (~0.1 g/L) means that CYP2E1 is more important in clearing ethanol from the blood in heavy drinkers when they reach high BAC (Lieber 2004). An interesting feature of CYP2E1 is that after a period of continuous heavy drinking its enzymatic activity is increased, a process known as enzyme induction (Oneta et al. 2002). This is the mechanism proposed to explain the faster elimination rates of ethanol (0.25–0.35 g/L per h) sometimes seen in alcoholics during detoxification (Keiding et al. 1983).

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only slowly oxidized in the liver, first by hydroxylation to acetol and then via the action of other enzymes to give methylglyoxal and 1,2-­propanediol as metabolites (Kalapos 1999). An important discovery in biological alcohol research was that certain nitrogen-­ containing heterocyclic compounds, such as pyrazole and its 4-­methyl derivative, functioned as competitive inhibitors of liver ADH in  vitro (Li and Theorell 1969). These compounds compete with ethanol for binding sites on the enzyme-­coenzyme complex and this slows the oxidative metabolism of ethanol to acetaldehyde. Later on, 4-­methyl pyrazole (4-­ MP) was developed into a therapeutic agent (fomepizole) trade name Antizol® and is used in clinical medicine to treat people poisoned with methanol or ethylene glycol (Brent et al. 2001). Prior to the development of fomepizole, the antidote of choice for the treatment of patients poisoned with methanol and/or ethylene glycol was intravenous infusion of ethanol (Roe 1955). Sterile solutions of ethanol (7–10% v/v in saline) were available at emergency departments and these were administered as a rapid infusion to reach a BAC of about 1.0 g/L and thereafter a continuous ethanol drip was provided (~100 mL/h) to provide 7–8 g ethanol per hour. This was sufficient to replace the amount lost through metabolism and ensure that the steady-­state BAC of 1.0 g/L was maintained during invasive treatment, such as hemodialysis. The higher affinity of the class I ADH for ethanol as a substrate meant that oxidation of the methanol and ethylene glycol into their more dangerous acid metabolites was prevented (Barceloux et al. 2002). Because ethanol is a depressant of the central nervous system (CNS), it is less suitable as an antidote for children who might have inadvertently consumed methanol or ethylene glycol (Roy et al. 2003). Ethanol is also contraindicated in patients suffering liver dysfunction (e.g. alcoholics with cirrhosis), so fomepizole is the first-­choice antidote, despite it being more expensive than ethanol (McMartin et al.

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Figure 52.1.2  Comparison of the oxidative metabolism of ethanol and methanol (primary alcohols) and isopropanol (secondary alcohol) by alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH). In the process, the coenzyme nicotinamide adenine dinucleotide NAD+ is reduced to NADH.

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Alcohol dehydrogenase (ADH) NAD+

Formaldehyde

NAD+

NADH

NAD+

Ethanol

NADH

NAD+

NADH

Formate

NADH

Acetaldehyde

Acetate Enzyme inhibitor

Enzyme inhibitor

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CH3

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H3C

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Fomepizole (Antizol®) 4-methyl pyrazole

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CH3

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Aldehyde dehydrogenase (ALDH)

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Figure 52.1.3  Scheme showing oxidative metabolism of ethanol and methanol in the liver via the ADH and ALDH enzymes and two drugs used as enzyme inhibitors, 4-­methyl pyrazole (ADH inhibitor), and disulfiram (ALDH inhibitor).

with isotope dilution methods (Jones et  al. 1992; Endres and Gruner 1994). The absorption of ethanol starts immediately after it reaches the stomach, although the rate of uptake into the blood occurs much faster from the upper part of the small intestine (duodenum and jejunum), owing to the larger absorption surface area provided by the villi and micro-­villi. Accordingly, the speed of ethanol absorption and the time of occurrence of the peak BAC depends on factors influencing gastric emptying, especially whether drinking occurs on an empty stomach or after a meal (Jones 2019a). The rate of absorption is highly variable, which impacts on the values of tmax and Cmax derived from blood-­a lcohol curves (Jones et al. 1991; Uemura et al. 2005). Hundreds of controlled drinking experiments show that the highest concentration of ethanol in blood (Cmax) occurs between 10 to 120 min after the end of drinking. Much depends on the dose of ethanol ingested, the type of beverage consumed (beer or spirits), the speed of drinking, and especially the fed or fasted state of the individual (Jones and Jonsson 1994). Because the rate of absorption of ethanol is initially faster than the rate of distribution and metabolism, the BAC increases. As time passes and the amount of ethanol in the stomach becomes smaller and smaller, the rate of absorption becomes equal to the rate of distribution and metabolism. The BAC will eventually reach its highest concentration and thereafter begin to decrease, which signifies that that elimination rate is now faster than absorption rate. The post-­ absorptive phase of the blood-­alcohol curve is indicated when arterial and venous BACs are the same (Jones et al. 2004). Most of the ethanol that enters the blood stream (90–98%) is eliminated by oxidative metabolism and only a small fraction

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52.6.1 Ethanol

At room temperature and atmospheric pressure, ethanol is a clear, colorless, and flammable liquid and its physicochemical properties are summarized in Table  52.1.2. As a pure solvent, ethanol has a characteristic and pleasant odor, a sharp taste and it burns in air with a blue flame. pKa of ethanol is 16.0, which means that molecules remain unionized at physiological pH (7.3–7.4). The high polarity and small molecular size of ethanol means that it easily penetrates cell membranes and tissues by passive diffusion, and this includes the blood-­brain barrier. Ethanol distributes into the TBW space, which represents about 50–60% of a person’s body weight. Ethanol has been used as a tracer to determine TBW by the dilution technique, and the results showed good ­agreement

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Table 52.1.4  Examples of toxic alcohols and their metabolites often encountered in clinical and forensic toxicology, their main metabolites, and ­comments about the toxic manifestations. Main metabolites

Comments and manifestations of toxicity

Methanol

Formaldehyde and formic acid

Toxic metabolites, formaldehyde and formic acid, are produced. The latter is a strong organic acid (pKa 3.77) causing metabolic acidosis, blurred vision and sometimes permanent blindness. Methanol poisoning is associated with lowering of pH in the blood, elevated osmolal gap initially, and later a higher anion gap. In postmortem specimens, the analysis of formate in blood verifies methanol-­related deaths.

Ethanol

Acetaldehyde and acetic acid

Ethanol intoxication increases the osmolal gap, although metabolic acidosis is mild and mainly caused by elevated lactic acid. The acetaldehyde metabolite is toxic but this is rapidly converted to acetic acid by low km ALDH. Blocking ALDH with disulfiram (Antabuse®) causes a range of unpleasant physiological effects. Drinking to reach blood-­ethanol concentrations between 3 and 4 g/L can cause death.

Isopropanol (2-­propanol)

Acetone, acetol, and propanediol

Isopropanol is sometimes used for intoxication purposes, and increases osmolal gap but not a dangerous metabolic acidosis. Poisoned patients become ketonic, owing to accumulation of the metabolite acetone, which has a longer elimination half-­life compared with the parent drug. Depressant effects on the CNS are similar to intoxication with ethanol.

Ethylene glycol

Glycoaldehyde, glycolic acid, glyoxylic acid, and oxalic acid

Diethylene glycol (DEG)

2-­hydroxyethoxy-­ acetaldehyde, 2-­hydroxyethoxyacetic acid and diglycolic acid

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γ-­hydroxybutyraldehyde and γ-­hydroxybutyric acid (GHB)

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1,4-­Butanediol

This antifreeze solvent is converted into toxic metabolites, mainly glycolic, which is mainly responsible for the metabolic acidosis. Initial effects of ethylene glycol resemble ethanol intoxication, and the osmolal gap is elevated. Oxalic acid metabolite reacts with intracellular calcium ions forming calcium oxalate crystals in the renal tubuli, leading to kidney failure.

(7.2 and counteract the metabolic acidosis caused by acid metabolites of the toxic alcohols. • Removal of any un-­metabolized methanol and its metabolites from the bloodstream by single-­pass hemodialysis. Note that infusion of the antidote needs to be continued during this treatment. • Administration of folinic acid to boost the metabolism of formate into carbon dioxide and water. Clusters of poisonings and fatalities from drinking methanol or ethylene glycol frequently occur in third world nations where conventional alcoholic drinks are expensive or not easily available for religious or other reasons (Jones 2021). Recent examples of methanol-­related deaths have been documented from Malaysia (Md Noor et  al. 2020), Turkey (Gulen et  al. 2020), Pakistan (Ahmed et al. 2017), and Iran (Aghababaeian et al. 2019). In these countries, many people purchase contraband liquor, which the manufacturers have spiked with methanol as a way to increase profits, thus providing a cheap substitute for conventional alcoholic drinks. Because of their late presentation for hospital emergency treatment, the long-­term prognosis for patients poisoned with methanol is poor. Many of those that purchase contraband liquor for intoxication purposes often suffer from an alcohol-­use disorder and are generally in poor health and nutrition anyway. A well-­ documented outbreak of methanol poisoning has occurred in Norway in 2002–2004 (Hovda et al. 2005) and another in Estonia in 2001 (Paasma et al. 2007). Both these outbreaks were closely followed to determine the factors that led to a full recovery and better long-term prognosis after methanol poisoning (Paasma et al. 2009, 2012). If the aliphatic alcohols discussed in this chapter are arranged in decreasing order of their inherent toxicity and potential danger to human health, the ordering would probably be methanol > ethylene glycol > 2,3-­butanediol > diethylene glycol > isopropanol > ethanol. However, in terms of annual deaths, because ethanol is a legal drug and also a major recreational drug of abuse, statistics from the World Health Organization show that there are about three million alcohol-­related deaths annually from various causes, which far exceeds deaths caused by ingestion of methanol and/or ethylene glycol (Spillane et al. 2020).

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can develop a dependency which leads to behavioural changes and drug seeking behaviour. They can also harm the health of consumers and possibly cause death when used to excess. The legislative authorities can enact exceptions and modifications to a law, especially regarding the use of substitution therapy for heroin addicts. In general, the term ‘illegal drugs’ does not comprise drugs that are available on prescription. However, prescription-­only drugs can be used illegally such as the benzodiazepines which are often misused and combined with illicit drugs. Generally, narcotics acts refer essentially to all illicit drugs, not just the narcotic analgesics. The drugs listed in the acts are especially dangerous because consumers may become addicted particularly if there is no medical indication or the dose used is in excess of that normally prescribed (e.g. morphine). Forensic toxicological laboratories should be in a position to analyse the identity and quantity of any active substance(s) in an exhibit (e.g. tablet or powder). Addiction means a psychical and partly physical state of an organism evoked by a pharmaceutically active substance. This state is characterized by behaviour patterns and reactions that always include the need to consume the substance continuously and periodically, either to induce its physical effects or to avoid negative consequences of non-­consumption. Physical addiction involves the desire repeatedly to consume substances. It is also characterized by a withdrawal syndrome following an interruption or cessation of consumption. Physical addiction often leads to an ever-­increasing dose as manifestation of tolerance develops. Tolerance is particularly pronounced for opioids and benzodiazepines. Tolerance and addiction do not have to occur simultaneously. Generally, there is only a moderate development of tolerance, and withdrawal symptoms are often not clearly expressed. Death that results from the use of drugs (often termed overdose) is caused by the use of doses that exceed the developed tolerance. Most drug overdose deaths involve the use of heroin, but increasingly, deaths are caused by other opioids, especially methadone and fentanyl or oxycodone. Additives or the blending of so-­called ‘street drugs’ can lead to enhanced mortality depending on the combination used. The concomitant use of other substances such as alcohol, benzodiazepines and others in addition to heroin is dangerous due to the increased risk of central nervous system (CNS) depression and adverse effects on respiration. Deaths from cocaine use occur less frequently since toxic effects such as an increase in blood pressure, and tachyarrhythmia as well as cerebral seizures are less dangerous to rather young people who represent the major part of cocaine consumers. Respiratory paralysis may occur when very high doses of cocaine are consumed. The percentage of deaths due to the use of amphetamines or ecstasy appears very low taking into consideration the presumed frequency of consumption. Deaths in relatively young people can occur if they have heart disease  – usually a result of prolonged cocaine or amphetamine use. Apart from circulatory collapse, hyperthermia is another dangerous effect of these drugs. Hyperthermia causes intravascular coagulation, rhabdomyolysis and consecutively renal failure. Although not all statistics appear consistent, the

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Wallage, H.R., and Watterson, J.H. (2008). Formic acid and methanol concentrations in death investigations. Journal of Analytical Toxicology 32: 241–247. Watson, P.E., Watson, I.D., and Batt, R.D. (1981). Prediction of blood alcohol concentrations in human subjects. Updating the Widmark equation. Journal of Studies on Alcohol 42: 547–556. Wax, P.M. (1995). Elixirs, diluents, and the passage of the 1938 federal food, drug and cosmetic act. Annals of Internal Medicine 122: 456–461. Welch, B.T., Coelho Prabhu, N., Walkoff, L., and Trenkner, S.W. (2016). brewery syndrome in the setting of long-­ standing Crohn’s Auto-­ disease: A case report and review of the literature. J Crohns Colitis 10: 1448–1450. Widmark, E.M.P. (1920). Studies in the concentration of indifferent narcotics in blood and tissues. Acta Medica Scandinavica 52: 87–164. Widmark, E.M.P. (1922). Eine mikromethode zur bestimmung von äthylalkohol im blut. Biochemische Zeitschrift 131: 473–484. Widmark, E.M.P. (1932). Die theoretischen grundlagen und die praktische verwendbarkeit der gerichtlich-­ medizinischen alkoholbestimmung. Berlin: Urban & Schwarzenberg. Wilkinson, P.K. (1980). Pharmacokinetics of ethanol: A review. Alcoholism, Clinical and Experimental Research 4: 6–21. Wood, D.M., Warren-­Gash, C., Ashraf, T. et al. (2008). Medical and legal confusion surrounding gamma-­hydroxybutyrate (GHB) and its precursors gamma-­butyrolactone (GBL) and 1,4-­butanediol (1,4-­BD). Quarterly Journal of Medicine 101: 23–29. Woolf, A.D. (1998). The Haitian diethylene glycol poisoning tragedy: A  dark wood revisited. Journal of the American Medical Association 279: 1215–1216. Xiao, H.T., He, L., Tong, R.S. et al. (2014). Rapid and sensitive headspace gas chromatography-­mass spectrometry method for the analysis of ethanol in the whole blood. Journal of Clinical Laboratory Analysis 28: 386–390. Zakhari, S. (2006). Overview: How is alcohol metabolized by the body? Alcohol Research Health 29: 245–254. Zamengo, L., Tedeschi, G., Frison, G. et al. (2019). Inter-­laboratory proficiency results of blood alcohol determinations at clinical and forensic laboratories in Italy. Forensic Science International 295: 213–218. Zvosec, D.L., Smith, S.W., McCutcheon, J.R. et al. (2001). Adverse events, including death, associated with the use of 1,4-­ butanediol. New England Journal of Medicine 344: 87–94. Zvosec, D.L., Smith, S.W., Porrata, T. et al. (2011). Case series of 226 gamma-­ hydroxybutyrate-­associated deaths: Lethal toxicity and trauma. American Journal of Emergency Medicine 29: 319–332.

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52.2 Illegal drugs Frank Musshoff An illegal drug or a so-­called ‘controlled substance’ is a drug whose use, sale or distribution is not allowed. What drugs are considered legal or illegal varies from country to country as do punishments. Drugs are often addictive, meaning that a person with regular use

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In cases of crimes perpetrated in order to be able to buy drugs later, it is questionable whether responsibility could be entirely diminished since there has to be the ability to control one’s actions when perpetrating the crime. In exceptional cases, it might be possible to conclude from the type and extent of the crime whether it was motivated by the perpetrator’s own drug addiction or not. In cases of crimes that are not associated with drug addiction, only the effects of the acute intoxication need to be examined.

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The generic term cannabis refers to different products of the plant Cannabis sativa. The dried leaves and dried flowering tops are called marijuana containing 4.0

Respiratory depression, coma

Source: Reproduced with permission from Madea and Musshoff (2009), © Deutsches Ärzteblatt, ÄrzteVerlag GmbH.

have a sleep promoting effect, like GHB. Again, like GHB, BDO in very high doses can cause coma and death. There is a substantial regional variability in patterns of illicit drug use and some drugs that are specific or unspecific to the demographic area. Not all drugs of interest can be specified here.

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A receptors: Classification on the basis of subunit structure and receptor function. Pharmacological Reviews 50 (2): 291–313. Barnett, V.T., Bergmann, F. Humphrey, H. and Chediak, J. (1992). Diffuse alveolar hemorrhage secondary to superwarfarin ingestion. Chest 102: 1301–1302. Barr, D. and Needham, L. (2002). Analytical methods for biological monitoring of exposure to pesticides: A review. Journal of Chromatography B 778: 5–29. Barraclough, B.M., Nelson, B., Bunch, J. and Sainsbury, P. (1971). Suicide and barbiturate prescribing. Journal of the Royal College of General Practitioners 21 (112): 645–653. Baruch, E., Vernon, L.F. and Hasbun, R.J. (2007). Intractable nausea caused by zolpidem withdrawal: A case report. Journal of Addiction Medicine 1 (1): 48–50. Baselt, R.C. (2011). Disposition of Toxic Drugs and Chemicals in Man, 9th edn. Foster City: Biomedical Publishing. Baselt, R.C., Yoshikawa, D., Chang J. and Li, J. (1993). Improved long-­ term stability of blood cocaine in evacuated collection tubes. Journal of Forensic Sciences 38: 935–937. Basu, A., Brown, S., Kirkham, N. et al. (2009). Coma blisters in 2 children on anticonvulsant medication. Journal of Child Neurology 24 (8): 1021–1025. Bauman, J.W., Madhu, C., McKim, J.M.J. et al. (1992). Introduction of hepatic metallothionein by paraquat. Toxicology and Applied Pharma­ cology 117: 223–241. Becquemont, L., Mouajjah, S., Escaffre, O. et  al. (1999). Cytochrome P-­450 3A4 and 2C8 are involved in zopiclone metabolism. Drug Metabolism and Disposition 27 (9): 1068–1073. Berkowitz, B.A., Ngai, S.H., Yang, J.C. et al. (1975). The disposition of morphine in surgical patients. Clinical Pharmacology and Therapeutics 17: 629–635. Bernheim, P.J. and Cox, J.N. (1960). Heat stroke and amphetamine intoxication in a sportsman. Schweizerische Medizinische Wochenschrift 90: 322–331. Besenhofer, L.M., Adegboyega, P.A., Berlels, M. et al. (2010). Inhibition of metabolism of diethylene glycol prevents target organ toxicity in rats. Toxicological Science 117 (1): 25–35. Besset, A., Tafti, M., Villemin, E. et al. (1995). Effects of zolpidem on the architecture and cyclical structure of sleep in poor sleepers. Drugs in Experimental Clinical Research 21 (4): 161–169. Beuck, S., Schänzer, W. and Thevis, M. (2011). Investigation of the in vitro metabolism of the emerging drug candidate S107 for doping-­ preventive purposes. Journal of Mass Spectrometry 46: 112–130. BGA (Bundesgesundheitsamt) (1992). Amalgame-­Nebenwirkungen und Bewertung der Toxizität. Zahnärzt Mitteilungen 19: 10/92. Bidlingmaier, M. and Strasburger, C.J. (2010). Growth hormone. Handbook of Experimental Pharmacology 195: 187–200. Bismuth, C., Inns, R.H. and Marrs, T.C. (1992). Efficacy, toxicology and clinical use of oximes. In: B. Ballantyne and T.C. Marrs (eds.), Clinical and Experimental Toxicology of Organophosphates and Carbamates, pp. 555–577. Oxford: Butterworth-­Heinemann. Blain, P.G. (2011). Organophosphorus poisoning (acute). Clinical Evidence (Online) 2102: 1–17. Bonte, W. (2000). Congener analysis. In: J.A. Siegel, P.J. Saukko, and G.C.  Knupfer, (eds.), Encyclopedia of Forensic Sciences, pp. 93–102. London: Academic Press.

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52.3 New Psychoactive Drugs Olaf H. Drummer

52.3.1 Introduction The term new or novel psychoactive drugs (NPSs) refer to new psychoactive drugs that largely mimic the effects of existing controlled drugs such as the amphetamines, benzodiazepines, cannabis, hallucinogens, opioids, etc. Initially, many of these substances were traded as legal replacements to the existing illicit drugs that required the establishment of various national and international regulatory mechanisms to control their sale

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52.3.2  Classes of Drugs Stimulants and hallucinogens

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Stimulants occupy a large class of the NPS, many of which also possess hallucinogenic properties, and indeed some are mainly hallucinogens, hence the reason to combine these drugs into one sub-­section (Table 52.3.1). Many of these substances possess activity to enhance the effects of endogenous neurotransmitters, particularly noradrenaline (norepinephrine) as well as dopamine and serotonin (5HT). They can do this by direct action on the receptor as partial or full agonists, by indirectly acting as substrates for reuptake into the nerve terminal and competing with endogenous transmitters, by inhibiting vesicular transporter, or by inhibiting the respective monoamine transporters (Iversen et al. 2014). A typical feature of hallucinogens is their serotonin agonist activity, particularly the 5HT2 subtype. The central serotonin receptors are key to regulating mood such that medicinal antidepressants will all have significant activity to enhance serotonin-­ mediated functions in the brain. The phenethylamine backbone is common to many of these substances. Most of these NPSs have substantial stimulant activity akin to amphetamine, an alpha-­ methyl substituted phenethylamine. Indeed, many of these substances are beta-­keto (bk) derivatives of this typical amphetamine structure, typified by cathinone, a naturally occurring alkaloid in the middle eastern plant, Khat (Catha edulis). Cathinones are the second most common NPS seen over the last several years with over 130 reported to the European Union drug monitoring network (EMCDDA) (Michou 2019). The

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and use. Today, many of these drugs are now controlled by most jurisdictions although some persist and each year new drugs appear in various parts of the world. The United Nations Office of Drug Control (UNODC) recorded the first NPS about 20 years ago (Fedotov 2019). As of the end of 2018, almost 900 of these substances were identified around the world, with one or more appearing in most countries of the world. The European Monitoring Centre for Drugs and Drug Addiction (EMCDDA) recorded to the end of 2018 over 730 of these drugs in 35 European countries (Michou 2019). Not every country will have seen most of these NPSs; their presence in a jurisdiction will vary with time depending on demand and evolution of legal control mechanisms. These NPSs continue to evolve largely dependent on the creativity of clandestine synthetic chemists. The UNODC Early Warning Advisory showed that synthetic opioids (34%) were the frequent new substances reported in 2019  in the 120  member countries and territories, followed by stimulants (29%) and synthetic cannabinoids (24%) (UNODC 2020). Previously emerging new synthetic cannabinoids and cathinones dominated the evolving NPS scene. The general adverse effects of these substances on users are summarized in Figure 52.3.1. Size of boxes refers to approximate scale of use in community There are also numerous plant-­based psychoactive materials that include kratom (Mitragyna speciosa), khat (Catha edulis), Salvia spp., and others (Fedotov 2019). This chapter provides an overview of a selection of these substances, their detection in biological material, known pharmacological and toxicological properties, and their relevance to forensic medicine.

Synthetic cannabinoids

Stimulants: agitation, anxiety, elevated

Anxiety, cognitive and psychomotor

FO

R

C

O

N

Stimulants and hallucinogens

blood pressure and heart rate,

impairment, seizures, heart failure,

restlessness, aggression, rebound

and sudden death

fatigue, and psychoses (paranoia). Hallucinogens: also visual and/or

Benzodiazepines

auditory hallucinations, and serotonin

Sedation, confusion, amnesia,

toxicity

dizziness, respiratory depression,

Opioids

parasomnias, and seizures

Sedation, respiratory depression,

Herbal substances

miosis, and sudden death

Stimulants and hallucinogens

Figure 52.3.1  Diagram illustrating a selection of common adverse effects of the respective NPS.

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Table 52.3.1  Examples of new psychoactive drugs by classes Key Examples

Synthetic Cannabinoids

5F-­ADB, 5F-­AMB, AB-­CHMINACA, AM-­2201, ADB-­FUBINACA, FUB-­AMB, MDMB-­CHMICA, MDMD-­ FUNINACA, AMB-­FUBINACA, JWH-­018, CUMYL-­PegACLONE, UR-­144, YLR-­11, 5F-­MDMB-­PINACA, 5F-­PB22, 5F-­APINACA

Stimulants and Hallucinogens

Cathinones: 4-­CEC, 4-­methylmethcathinone (4-­MMC, mephedrone)1, 3-­MMC, buphedrone, butylone, clephedrone, dibutylone, N-­ethylhexedrone, ephylone, ethylone, methedrone, methylone, N-­ethylpentylone, pentedrone, pentylone. Pyrovalerones: a-­pyrrolidinophenone, a-­pyrrolidinohexiophenone (α-­PHP), a-­pyrrolidinovalerophenone (a-­PVP), 3,4-­methylenedioxypyrovalerone (MDPV), 3,4-­methylenedioxy-­a-­pyrrolidinohexanophenone (3,4-­MDPHP), and a-­pyrrolidinopentiothiophenone (a-­PVT). 2C series: 2C-­E, 2C-­P, 2C-­bromo-­dragonfly, and 3C-­bromo-­dragonfly. NBOMe series: 25H-­NBOME, 25C-­NBOMe, 25B-­NBOMe, 25E-­NBOMe, and 25-­I-­NBOMe. Piperazines: Benzylpiperazine (BZP), 3,4-­methylenedioxybenzylpiperazine (MDBP), 1-­(3-­ trifluoromethylphenyl) piperazine (TFMPP), 1-­(3-­chlorophenyl)piperazine (m-­CPP), and 1-­(4-­ methoxyphenyl)piperazine (MeOPP). Tryptamines: a-­methyltryptamine (AMT), N,N-­dimethyltryptamine (DMT), di-­isopropyltryptamine (DiPT), 5-­methoxy-­diisopropyltryptamine (Foxy), and 4-­hydroxy-­dimethyltryptamine (psilocin). Others: 5,6-­methylenedioxy-­2-­aminoindane (MDAI), 6-­(2-­aminopropyl)-­benzofuran (benzofury or 6-­APB), 5-­APB, 2,3-­dihydro-­APB, N-­methyl-­APB, and N-­ethyl-­APB.

Benzodiazepines

Adinazolam, bromazolam, clonazolam, diclazepam, etizolam, flualprazolam, flubromazepam, flubromazolam, fluclotizolam, meclonazepam, nifoxipam, norfludiazepam, phenazepam, and pyrazolam.

Opioids

acetylfentanyl, acrylfentanyl, butyrylfentanyl, carfentanil, cyclopropylfentanyl, despropionylfentanyl, 2-­& 4-­fluorofentanyl, 4-­fluorobutyrfentanyl, furanylfentanyl, methoxyacetylfentanyl, 3-­methylfentanyl, ohmefentanyl, ocfentanil, thiofentanyl, isotonitazene, AH-­7921, U47700, and MT-­45.

Dissociatives

Methoxetamine, diphenidine, methoxyphenidine, 3-­MeO-­PCP, and 4-­MeO-­PCP.

Herbal NPS1

Areca catechu (areoline), Catha edulis (cathinone), Mitragyna speciosa (mitragynine), Piper methysticum (kawain), Psilocybe spp (psilocin), Salvia divinorum (salvinorin A), and Tabernanthe iboga (ibogaine).

TR IB

U

TO

R

U

SE

O

N

LY

Drug Classes

 most active substance in brackets

1

FO

R

C

O

N

most common recent examples include mephedrone (4-­methyl methcathinone, 4-­MMC), 3-­MMC, clephedrone (4-chloro-methcathinone), N-­ ethylhexedrone, ephylone (N-ethylpentylone), flephedrone (4-­fluoro-­methcathinone), and dibutylone (beta-­keto-­ dimethylbenzodioxolylbutanamine, bk-DMBDB). Another variation of the cathinone structure is the a-pyrrolidinophenones typified by 3,4-­methylenedioxypyrovalerone (MDPV) in which the nitrogen forms part of a pyrrolidino ring and the adjacent carbon is substituted with an alkyl chain. Analogs that include a-pyrrolidinovalerophenone (a-­ PVP) and a-pyrrolidinopentiothiophenone (a-­PVT), among others, and have become one of the more toxic cathinones. These analogs are potent norepinephrine and dopamine reuptake inhibitors without being transporter substrates themselves, similar to cocaine (Karila et al. 2018). Substitution on the phenyl ring, such as 4-­ methoxy (p-­ methoxy-amphetamine, PMA, and the N-­ methyl analog, methedrone) or 3-­4-­methylenedioxy (MDA and MDMA for the N-­methyl variant), has substantial serotonin reuptake inhibitory actions and associated empathogenic/entactogenic activities, whereas other variants with substitution around the aliphatic

nitrogen tend to have stronger activity on the noradrenaline and dopamine receptors (Figure 52.3.2). The piperazine stimulants are variants of the phenethylamines in which the nitrogen forms part of a piperazine ring. The earliest example was the weak stimulant 1-­ benzylpiperazine or BZP. Analogs with substitution on the phenyl ring include meta-­ chlorophenylpiperazine (m-­CPP), 3,4-­methylenedioxybenzylpip erazine (MDBP), and trifluoromethylphenylpiperazine (TFMPP) possess mostly serotonin agonist activity and are also more toxic (Arbo et al. 2012). Substitution of phenethylamine on the phenyl ring with 2-­and 5-­methoxy groups gave rise to the “2C” series of hallucinogens. These include 2C-­B (4-­bromo-­), 2C-­I (2-­iodo-­), and 2C-­E (4-­ ethyl) analogs. The naturally occurring mescaline isolated from some cactus species is the 3,4,5-­trimethoxy variant. When this 2C structure is further substituted on the nitrogen with the N-­benzylmethoxy moiety, a series of highly potent hallucinogens of the NBOMe series are produced. These include the 25C-­, 25B-­, 25H-­, and 25I-­NBOMe series of NPSs in which chlorine, bromine, hydrogen, and iodine are substituted at the 4-­position of

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CHAPTER 52   Toxicology of Specific Substances

R4 NR1R2

R5

R3

R1

R2

R3

R4

R5

H

H

CH3

H

H

Methamphetamine CH3

H

CH3

H

H

MDMA

CH3

H

CH3

H

3,4-

Mescaline

H

H

H

6-APB

H

H

CH3

Dibutylone

CH3

CH3

Ethyl

Ephylone

H

CH3

Propyl

Flephedrone

H

CH3

Methcathinone

CH3

Methedrone

H

Diphenidine

N-piperidine

LY

methylenedioxy 3,4,5-OCH3

H

6-Furan (3,4-

O

N

H

fused) 3,4-

SE

=O

methylenedioxy

R

U

=O

3,4methylenedioxy

=O

4-fluoro

H

CH3

=O

H

CH3

CH3

=O

4-methoxy

Phenyl

H

H

U

TO

CH3

TR IB

Amphetamine

Figure 52.3.2  Selected substituted phenethylamines and indole-­ethylamines. Source: Based on Roque Bravo et al. (2019).

FO

R

C

O

N

the 2,5-­dimethoxy substituted phenyl ring. All of these are potent hallucinogens with activity on one or more of the 5HT2 receptors, similar to LSD. A further variation of the 2C series occurs when the phenyl ring is substituted with two hydrogenated furan rings producing the “FLY” series of potent hallucinogens. Furan analogs are known as “Dragonfly” and include 2C-­and 3C-­bromo-­dragonfly, which are potent 5HT2 agonists. 3C-­bromo-­dragonfly also inhibits monoamine oxidase A, further enhancing its serotonergic effects and activity at sub-­ milligram doses. These, like the NBOMe series, can be impregnated on blotter papers. A noteworthy event was a mass poisoning of 29 persons in Germany involving a combination of 2C-­ E and bromo-­ dragonfly. All affected individuals survived although many had significant toxicity (respiratory distress and coma), with hallucinations and seizures also reported (Iwersen-­Bergmann et al. 2019). Tryptamines are largely hallucinogenic drugs based on the indole-­ ethylamine backbone that characterizes serotonin (5-­hydroxy-­tryptamine) and are mostly 5HT2 agonists. Ring substituted variants include alpha-­methyl-­(AMT), N,N-­dimethyl-­ (DMT), and di-­isopropyl-­tryptamines (DiPT). DMT is an active

principal of ayahuasca, a traditional spiritual medicine from South America. The active substance in the hallucinogenic psilocybin mushroom is the related 4-­hydroxy-­DMT or psilocin. A number of other chemical variations of these NPS stimulants include the aminoindanes and benzofurans. Aminoindanes have a cyclized form involving the alpha-­methyl group and the alkylamino side chain to form an indane ring system and include MDAI (5,6-­methylenedioxy-­2-­aminoindane), which is an inhibitor of the serotonin transporter, while MDAT is the tetralin analog. Benzofurans are mono-­substituted furan rings on the phenyl (as distinct from the di-­substituted FLY series) and include 5-­and 6-­APB, commonly known as benzofury. These two NPSs are potent inhibitors of all three monoamine transporters. 6-­APB is a potent 5HT2B agonist, while 5-­APB has activity on all three 5HT2 receptor subtypes. Other benzofurans include the 2,3-­dihydro (5-­ &  6-­APDB), N-­methyl (5-­ &  6-­MAPB), and N-­ethyl variants (5-­&  6-­EAPB). Since the first report in 2010, these benzofurans act much like MDMA but are more potent. Numerous presentations to hospitals and deaths have been reported (Roque Bravo et  al 2019). Selected examples of these stimulant and hallucinogenic NPSs are shown in Figure 52.3.2.

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TOXICOLOGY

MDPV

Bromo-dragonfly (DOB-DFLY)

O

N

LY

LSD

U

SE

5-MeO-DiPT (Foxy)

TO

R

25B-NBOMe

TR IB

U

BZP

m-CPP

O

N

Figure 52.3.2  (Continued)

FO

R

C

The stimulant and/or mixed hallucinogenic drugs that have activity as agonists on the 5HT2B subtype of serotonin receptor have been linked to the development of cardiac toxicity (Berger et al. 2009). These drugs include MDMA, MDA, and 5-­& 6-­APB (Iversen et al. 2014). The drugs that act primarily as stimulants have similar effects as for amphetamine and methamphetamine and tend to be more associated with behavioral aggression, nervousness, and anxiety. Interpretation of blood and tissues concentrations is even more complex with this class of drugs. Most often, use can lead to behavioral toxicity such as outbursts of violence and aggression or hallucinations. They have been the cause of death due to cardiovascular toxicity in numerous cases, and often they are associated with other CNS active drugs (Zaami et  al. 2018; Kraemer et al. 2019). Due to the serotonin activity of many of these drugs, they have the potential to interact adversely with therapeutic drugs that also have serotonin enhancing activity, such as the

antidepressants, tramadol, anti-­migraine drugs, and some opioids (see Chapter 47). Blood concentrations vary widely and depend not only on the drug potency but also on how they are used. As for all unknown cases, highly sensitive detection systems are required. Methods published include GC-­MS (Mercieca et al. 2018), tandem LC-­MS (Ambach et al. 2015), and high resolution LC-­MS (Mollerup et al. 2017).

Synthetic cannabinoids Cannabis (marijuana) is the most common illicit drug used in many countries and is usually smoked as a joint or blunt, or through a water pipe (bong). The most psychoactive component of cannabis THC released in the smoke is inhaled and is rapidly absorbed in the lungs reaching the brain (where it primarily acts) within seconds of inhalation. In forensic medicine, it is most

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CHAPTER 52   Toxicology of Specific Substances

I N OH

H

U

TR IB

N

O

C

R

FO

N

O

N

H

HN

O

O

N

O

THC

AM-1220

APICA

O O O

O

O

N

N

N

N

N

O

F

F

AB-FUBINACA O O

O

N

JWH-030

O

JWH-210

N

JWH-250

R

JWH-018

N

N H

N N

N

AB-PINACA

SE

O

O O O

N H

F

5F-ADB

U

N

O O O

N H

F

5F-PB22

AM-679

LY

N

Figure 52.3.3  Selected synthetic cannabinoids with THC as comparison

TO

often associated with cases of suspected impaired driving but has also been implicated as a possible contributor to some deaths in persons with significant heart disease (Drummer et al. 2019). Drugs designed to mimic the effects of cannabis have been one of the more dominant NPSs that have appeared since their first detection in 2008 and represent some 30% of all new drugs over the last several years. The usual practice has been to apply some of the drug powder to dry plant matter and smoke the burning blend similar to a joint, although vaping is now also popular (Blundell et al. 2018). Brand names such as Spice, K2, and Kronic have been used, among many others. While THC is a partial agonist, most of the synthetic cannabinoids are full agonists with many much more potent than THC itself on the CB1 receptor. Most of them also have substantial activity on the CB2 receptor, and in contrast to THC, many show selectivity for the CB1 receptor. The G-­protein coupled CB1 receptor is largely associated with the CNS although it has functions in the peripheral nervous system as well, while the CB2 receptor is largely associated with regulation of immune function. There are many seemingly diverse structures that include aminoalkylindoles (AM-­1220, WIN-­55,225), naphthoylindoles (JWH-­ 018, JWH-­ 210, AM-­ 2201), naphthoylpyrroles (JWH-­ 030, JWH-­145), phenylacetylindoles (JWH-­250, JWH-­203), benzoylindoles (AM-­679, AM-­694), alicyclic and polycyclic 3-­acylindoles (XLR-­12, UR-­144), adamantoylindoles (5F-­AKB-­48, APICA), indazole carboxamides (AB-­PINACA, AB-­FUBINACA), quinoyl esters (PB-­22, 5F PB22), and others. Many of the fluorinated analogs, such as 5F-­PB22, 5F-­AMB, and 5F-­ADB, are highly potent agonists on the CB1 receptor and feature in many of the intoxications. Selected examples are shown in Figure 52.3.3. In the past, the names of these synthetic cannabinoids have derived from companies or individual researchers publishing their scientific research and/or lodging patent applications, hence prefixes such as AM, JWH. The diverse structures and the sheer volume of synthetic cannabinoids have led to a naming convention in which a short-­hand systematic nomenclature is derived from names made sequentially from codes for a linked (or linking) group, followed by a tail group, core group, and linker. As with cannabis, the synthetic cannabinoids produce euphoria and a sense of relaxation associated with reduced vigilance and cognitive function and higher states of anxiety. However, acute intoxications are common and have involved over 50 different synthetic cannabinoids. Symptoms include agitation, nausea, and psychosis. Seizures and cardiac toxicity are more serious and can lead to sudden death (Worob and Wenthur 2019). Deaths have been reported from at least 35 synthetic cannabinoids including 5F-­AMB (N-­[[1-­(5-­fluoropentyl)-­1H-­indazol-­ 3-­ yl]carbonyl]-­ L-­ valine methyl ester) (Shanks and Behonick 2016), 5F-­ADB (Angerer et al. 2017; Ivanov et al. 2019; Kraemer et al. 2019; Hvozdovich et al. 2020), FUB-­AMB (Hvozdovich et al. 2020), XLR-­11 (3-­(tetramethyl-­cyclopropyl-­methanoyl)indole) (Shanks et  al. 2015), PB-­22 (1-­pentyl-­8-­quinolinyl ester-­1H-­ indole-­3-­carboxylic acid) (Gerostamoulos et  al. 2015), 5F-­PB22

(Angerer et  al. 2017), AB-­CHMINACA (Angerer et  al. 2017; Maeda et  al. 2018), MDMB-­CHMICA (Adamowicz 2016), UR-­ 144 (Paul et al. 2018), AM-­2201 (Labay et al. 2016), and JWH-­018 (Labay et al. 2016). As with most fatal intoxications, deaths often involve other drugs including other synthetic cannabinoids as well as associations with co-­morbidities such as various forms of heart disease. Fatalities and other deaths in which these drugs were detected have been reviewed in recent publications (Kraemer et al. 2019; Giorgetti et al. 2020). The UNODC report of new NPS reported in 2019 indicated that 5F-­MDMB-­PICA and AMB-­FUBINACA (FUB-­AMB) predominated with cases also of 5F-­MDMB-­PINACA (5F-­ADB) and ADB-­FUBINACA (UNODC 2020). There is no straightforward interpretation of blood or tissue concentrations given their diverse structures and potencies, and other competing drugs and co-­ morbidities in many cases. Redistributive processes are very likely further thwarting an understanding of toxic blood concentrations. Pathology findings are often non-­specific such as aspiration, coma, blood pressure, and breathing abnormalities. Co-­existing cardiovascular disease increases the risk of a fatal outcome, although it is likely that some members of this class of drugs will cause other adverse outcomes depending on their actions on other biological systems. Since these drugs adversely affect cognitive and psychomotor performance as well as vigilance, persons using these drugs will be adversely affected when driving motorized vehicles (Kaneko 2017; McCain et al. 2018). Blood concentrations are often below

O

N

TR IB

LY

N

U

TO

Benzodiazepines are one of the most widely used therapeutic class of drugs with one or more members of this class of drugs used as hypnotics, muscle relaxants, minor tranquilizers, or anti-­ convulsants. Benzodiazepines act as a positive allosteric modulator on the one or more of the subunits of the GABAA receptor, which is a ligand-­gated chloride-­selective ion channel that acts as a major inhibitory neurotransmitter in the brain (Sigel and Steinmann 2012). Not all benzodiazepines have the same net pharmacological effect; this depends on the affinity of the drug to one or more of the five subunits that make up the receptor. For example, benzodiazepines that have affinity for the α1βγ2 subtype have sedative activity, while effects on anxiety are linked more to the α2βγ2 subtype, although it is likely that these drugs interact with one or more of these subunits with differing affinity (Balkhi et al. 2020). In addition, their pharmacokinetic half-­lives are variable, and as for legal benzodiazepines, this also influences their use (hypnotic, sedative, etc.) and duration of action. Phenazepam and etizolam first appeared as novel benzodiazepines in Western Europe, although there were available legally in Russia and Japan, respectively (Michou 2019). Since 2012, over 20 novel benzodiazepine NPSs have appeared around the world. These include adinazolam, bromazolam, clonazolam, diclazepam, flualprazolam, flubromazepam, flubromazolam, fluclotizolam, meclonazepam, nifoxipam, and pyrazolam (Zawilska and Wojcieszak 2019). Of these, clonazolam and flualprazolam appear to be the most potent clinically (Balkhi et al. 2020). Structurally, all of these novel benzodiazepines are based on the original 1,4-­benzodiazepine nucleus or the triazolo or thieno-­triazolo variations (Zawilska and Wojcieszak 2019). In 2019, the UNODC reported that flualprazolam, ­flubromazepam,

O

Benzodiazepines

clonazolam, ­diclazepam, and etizolam were the most common benzodiazepines in cases of suspected driving under the influence (UNODC 2020) (Figure 52.3.4). Most appear to be in the form of solid dosage such as a tablet or capsule, but nasal insufflation and sublingual consumption from impregnated blotting paper and from smoking are known (Balkhi et al. 2020). Hospitalizations involving use and misuse of these novel benzodiazepines have been reported, particularly those involving flubromazolam (Backberg et  al. 2019) and clonazolam (Carpenter et al. 2019), although etizolam and phenazepam have also been frequently detected drugs. Clonazolam and flubromazolam have also appeared in one candy-­like pill involving a poisoning (Pope et al. 2018). Adverse symptoms range widely and include amnesia, ataxia, disinhibition, nausea, significant sedation to deep sleep, and respiratory depression. Illustrative of the dangers of potent novel benzodiazepines is a case report involving the ingestion of 3-­mg flubromazolam in a young male who was found some time later in a deep coma with unreactive pupils, hypotension, tachycardia, and slow respiration that was responsive to the antagonist flumazenil. His serum and urine taken 19 h after intoxication gave levels of 59 and 105 ng/ mL (Łukasik-­Głębocka et al. 2016). In another case report involving a middle-­aged male, persistent bradycardia was the dominant clinical feature with mild hypotension following the use of 3 mg of flubromazolam (Bohnenberger and Liu 2019). This long acting NPS has potent amnesic and hypnotic properties and is also subject to enterohepatic recycling (Huppertz et  al. 2018). Another case report of a young male using etizolam and diclazepam who had developed significant withdrawal symptoms (anxiety, sensitivity to temperature changes, light, and seizures) requiring treatment (Shapiro et al. 2019). Deaths caused from the toxic effects of these novel benzodiazepines have been occurred but have been invariably in the presence of another toxic drug, such as alcohol, common opioid or

SE

1 ng/mL; hence, sensitive detection systems are required, Tandem LC-­MS methods included (Tynon et al. 2017; Ong et al. 2019).

TOXICOLOGY

U

PART VII  

R

1230

O

H N

R

C

O

FO

Cl

O

H N

Adinazolam

Clonazolam

N Cl N

Flualprazolam

N N

Etizolam

F

F

F

N

N

N

F Cl

H N

Cl

Cl

Cl

Diazepam

O

H N

N

N N

Flubromazepam

Cl

S

N

Fluclotizolam

N N

Cl

S

N

N N

Phenazepam

Figure 52.3.4  Selected benzodiazepines with diazepam as comparison. Source: Based on UNODC (2020).

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CHAPTER 52   Toxicology of Specific Substances

FO

LY

N

O

SE

R

C

O

N

TR IB

U

TO

There have been numerous synthetic drugs released over the last several years that mimic the narcotic effects of heroin. Most of these are more potent than heroin and are even more potent than fentanyl itself. Indeed, most are based on the fentanyl structure. Arguably, the most potent NPS is carfentanil, which is some 100 times more potent than fentanyl. Fatalities have been reported from at least 16 analogs including acetylfentanyl, butyrylfentanyl, carfentanil, furanylfentanyl, methylfentanyls, 4-­methoxyfentanyl, ocfentanil, tetrahydrofuranylfentanyl, various fluorofentanyls, and some other structural variations with opioid activity known as AH-­7921, U-­47700, and MT-­45 (Zawilska 2017; Drummer 2019; Kraemer et al. 2019). Other novel opioids reported to cause respiratory depression chloroisobutryfentanyl, requiring hospitalization include 4-­ cyclopentylfentanyl, cyclopropylfentanyl, methoxyacetylfentanyl, and tetrahydrofuranfentanyl (Zawilska 2017; Kraemer et al. 2019) (Figure 52.3.5). A report from the UK Advisory Council on the Misuse of Drugs on fentanyls indicated that the following NPS opioids had been detected in the 2015–17 period, in decreasing order of fluorobutyrfentanyl, butyrfentanyl, detection, carfentanil, para-­

U

Opioids

acetylfentanyl, 2-­ fluorofentanyl, despropionylfentanyl, furanylfentanyl, 4-­ fluorofentanyl, cyclopropylfentanyl, methoxyacetylfentanyl, ocfentanil, and an unknown fluorofentanyl (Bowden-­Jones and Thomas 2020). The UNODC reported that acetylfentanyl, butyrfentanyl, and cyclopropylfentanyl were the most common novel fentanyls instrumental in causing drug-­ related fatalities in 2019 (UNODC 2020). More recently, non-­fentanyl and highly potent opioid NPSs have appeared. MT-­45, a 1-­substituted-­4-­(1,2-­diphenylethyl)piperazine first appeared in around 2012, followed by AH-­7921, a cyclohexylmethylbenzamide, and the more potent structural isomer U-­47700 (Figure 52.3.6). These have also caused numerous deaths, particularly U-­ 47700  in Europe and North America (Concheiro et al. 2018; Fels et al. 2019). In 2019, isotonitazene, a benzimidazole, which is as potent as fentanyl on the m-­opioid receptor with full agonist activity, was first detected in Canada, then Belgium, and later in the year the United States (Krotulski et al. 2020). It has appeared in at least several drug-­caused deaths nanogram per milliliter concentrations in blood and in sub-­ belongs to a number of related opioids, namely, clonitazene, etonitazene, and metonitazene. Since these drugs are potent m-­opioid receptor agonists, they will produce respiratory depression when used in sufficient doses relevant to any developed tolerance to the opioids. Given their mechanism of action and uncontrolled recreational use, and as with heroin users, many will also co-­consume other dangerous drugs and hence are more likely to present to emergency physicians and come under investigation by forensic medical practitioners. In some cases, heroin has been laced with a potent novel opioid such as fentanyl creating extreme danger to the user (Rodda et al. 2017). Because of issues of tolerance and use of other drugs, there will be no defined minimum fatal dose or concentration. Carfentanil,

R

stimulant, or another NPS. Polydrug use substantially reduces the ability to interpret the effect of one drug over another, let alone the relevance of a drug concentration. Developed tolerance and dependence will further exacerbate any straightforward interpretation of tissue levels. There are numerous analytical methods published for benzodiazepines; however, given the thermal instability of some members, LC-­MS methods are preferred (Pettersson Bergstrand et al. 2016).

Fentanyl

Acetylfentanyl

Cyclopropylfentanyl Ocfentanil

Butyrfentanyl

Carfentanil

Furanylfentanyl

4-Fluorofentanyl

Figure 52.3.5  Selected opioid NPS with fentanyl comparison. Sources: Adapted from Kraemer et al. (2019); Zawilska (2017).

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PART VII  

U-47700

TOXICOLOGY

MT-45

Isotonitazene

Figure 52.3.6  Selected non-­fentanyl novel opioid NPSs.

R

U

SE

O

N

LY

Examples of other dissociative NPSs that have appeared over the last several years include diphenidine, methoxyphenidine, 2-­oxo-­PCE (N-­ethyldeschloroketamine), and 3-­ & 4-­methoxy-­PCP (Morris and Wallach 2014; Backberg et al. 2015; Helander et al. 2015; Theofel et  al. 2019) (Figure  52.3.7). More recently, 2-­ fluorodeschloroketamine (2F-­ DCK) has emerged (Davidsen et al. 2020; Tang et al. 2020). As with other drugs of abuse intoxications leading to emergency management in hospitals and fatalities are rarely mono-­intoxications. Often other drugs are involved. An  excellent review on these dissociatives is by Morris and Wallach (2014).

Other Substances

TO

for example, will need detection in blood down to at least 0.1 ng/ mL; hence, the most sensitive analytical methods are required to detect many of these novel opioids. Drugs of most concern that are co-­used with opioids include other depressants of the central nervous system, such as alcohol and the benzodiazepines. The principal finding at autopsy is similar to that seen with heroin deaths and is associated with hypoxia, pulmonary edema, and features of congestion in other organs. Some of the novel opioids have been linked to kidney dysfunction, hearing loss (MT-­ 45), hemoptysis, acute lung injury, and diffuse alveolar hemorrhage (butyrfentanyl) (Cole et al. 2015; Mallappallil et al. 2017; Frisoni et al. 2018). Some immunoassays targeted toward fentanyl can detect novel opioids that are metabolized to norfentanyl (Helander et al. 2018; Guerrieri et al. 2019); however, immunoassays will not be able to detect all analogs. Hence, their detection in biological specimens requires the use of sensitive mass spectral detection procedures that are either based on tandem mass spectrometry or high-­ resolution mass spectrometry (Sofalvi et  al. 2017; Gerace et  al. 2018; Moody et al. 2018; Klingberg et al. 2019).

TR IB

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Mitragynine and the metabolite, 7-­hydroxy-­mitragynine, are alkaloids found in the southeast Asian plant, Mitragyna speciosa, commonly known as Kratom. These are potent partial m-­opioid receptor agonists with antagonist activity at k-­and d-­opioid receptors and also possess stimulant activity at lower doses. The plant material is often consumed in the form of dry leaves either directly or by infusion as a tea. It can cause significant euphoria, agitation, hallucinations, delusions, and vomiting and will cause respiratory depression, and if used repeatedly, drug dependence. A number of deaths have been reported with this substance with mitragynine and 7-­hydroxy-­mitragynine concentrations ranging from a few ng/mL in mono-­intoxications, but with mean blood concentrations of 0.4 and 0.7 mg/L, respectively (Corkery et al. 2019). Salvia divinorum is also a psychoactive plant that contains an opioid-­like compound. It contains as the most active constituent Salvinorin A that is a potent k-­opioid receptor agonist (Figure 52.3.7.1). It acts primarily as a hallucinogen, although it does not act on the 5HT2A receptor that is responsible for the hallucinogenic activity of LSD and other substances discussed earlier. The fresh or dried leaves can be chewed or made into a tea and even smoked. Another herbal NPS is khat grown on the Arabian Peninsula and northeast Africa. This contains cathinone (the basis for the cathinone stimulants) as well as cathine and other related alkaloids. This stimulant herb has been used locally for centuries, but its use has spread to many other countries. There are other herbal substances of interest. These are covered in detail in Lo Faro et al. (2020).

Dissociatives

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These are a class of hallucinogenic drugs that produce feelings of detachment and distort perceptions of sight and sound. Most of these dissociative drugs can also cause cognitive impairment and depress the CNS, among other adverse effects. Their principal mode of action is as non-­competitive inhibitors of the N-­methyl-­ D-­aspartate (NMDA) receptor. Ketamine and tiletamine are the most well-­known drugs of this type used in medicine for dissociative anesthesia, and phencyclidine (PCP) has been an illicit drug for many years. The antitussive dextromethorphan, when used in much larger doses than as an antitussive, also produces dissociative effects and continues to be abused in some parts of the world (Morris and Wallach 2014). Methoxetamine is an arylcyclohexylamine related to ketamine with actions as both an NMDA receptor antagonist and an inhibitor of serotonin reuptake that has led to numerous acute poisonings as well as fatalities, particularly in Europe and the United States. Clinical features include a dissociative/catatonic state with features resembling the use of a strong stimulant. Blood concentrations in intoxications tend to be around 0.1 mg/L and higher (Wood et al. 2012; Chiappini et al. 2015).

1233

Methoxetamine (MXE)

Diphenidine

Methoxyphenidine

3-MeO-PCP

2’-oxo-PCE

N

Ketamine

LY

CHAPTER 52   Toxicology of Specific Substances

7-Hydroxy-mitragynine

O

N

TR IB

Mitragynine

U

TO

R

U

SE

O

Figure 52.3.7  Selected dissociative drugs. Sources: Adapted from Morris and Wallach (2014); Helander et al. (2015); Backberg et al. (2015); Theofel et al. (2019).

C

Salvinorin A

Cathinone

FO

R

Figure 52.3.7.1  Examples of active constituents of Kratom, Salvia divinorum (salvinorin A), and Catha edulis (cathinone).

Methods of Analysis Analytical techniques to detect these NPSs let alone other more common drugs that may be present in a case involves the use of much more sophisticated methods than use of a series of immunoassays and/or basic mass spectrometric (MS) methods. Some immunoassays will detect one or more of a class of NPSs, but these will not be able to detect all members of a class, given their variable structures and likely differing immunoreactivities. The structural diversity, and in many cases high potency with consequent low concentrations in blood and other

s­pecimens, requires the use of tandem MS methods using drug-­independent acquisition mode, or better, high-­resolution MS (HR-­MS) covering the whole range of NPS that has been validated for the specimen of interest. Targeted methods will also work but are more dependent on obtaining analytical reference material for identification and targeting. New evolving NPS may not be detected if targeted methods are used, although even with HR-­MS analytical reference material will be required for formal quantification in submitted specimens. A crowd-­sourced online database is available (Mardal et  al. 2019).

FO

R

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O

N

TR IB

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N

O

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Adamowicz, P. (2016). Fatal intoxication with synthetic cannabinoid MDMB-­CHMICA. Forensic Science International 261: e5–e10. Ambach, L., Redondo, A.H., Konig, S. et al. (2015). Detection and quantification of 56 new psychoactive substances in whole blood and urine by LC-­MS/MS. Bioanalysis 7 (9): 1119–1136. Angerer, V., Jacobi, S., Franz, F. et al. (2017). Three fatalities associated with the synthetic cannabinoids 5F-­ADB, 5F-­PB-­22, and AB-­CHMINACA. Forensic Science International 281: e9–e15. Arbo, M.D., Bastos, M.L. and Carmo, H.F. (2012). Piperazine compounds as drugs of abuse. Drug and Alcohol Dependence 122 (3): 174–185. Backberg, M., Beck, O. and Helander, A. (2015). Phencyclidine analog use in Sweden-­-­intoxication cases involving 3-­MeO-­PCP and 4-­MeO-­PCP from the STRIDA project. Clinical Toxicology (Philadelphia) 53 (9): 856–864. Backberg, M., Pettersson Bergstrand, M., Beck, O. and Helander, A. (2019). Occurrence and time course of NPS benzodiazepines in Sweden  – results from intoxication cases in the STRIDA project. Clinical Toxicology (Philadelphia) 57 (3): 203–212. Balkhi, S.E., Monchaud, C., Herault, F. et al. (2020). Designer benzodiazepines’ pharmacological effects and potencies: How to find the information. Journal of Psychopharmacology 269881119901096. Berger, M., Gray, J.A. and Roth, B.L. (2009). The expanded biology of serotonin. Annual Review of Medicine 60: 355–366. Blundell, M., Dargan, P. and Wood, D. (2018). A cloud on the horizon-­a survey into the use of electronic vaping devices for recreational drug and new psychoactive substance (NPS) administration. QJM 111 (1): 9–14. Bohnenberger, K. and Liu, M.T. (2019). Flubromazolam overdose: A review of a new designer benzodiazepine and the role of flumazenil. Mental Health Clinician 9 (3): 133–137. Bowden-­Jones, O. and Thomas, S. (2020). Misuse of Fentanyl and Fentanyl Analogues. London, UK, Advisory Council on the Misuse of Drugs. Carpenter, J.E., Murray, B.P., Dunkley, C. et al. (2019). Designer benzodiazepines: A report of exposures recorded in the National Poison Data System, 2014-­2017. Clinical Toxicology (Philadelphia) 57 (4): 282–286. Chiappini, S., Claridge, H., Corkery, J.M. et al. (2015). Methoxetamine-­ related deaths in the UK: An overview. Human Psychopharmacology 30 (4): 244–248. Cole, J.B., Dunbar, J.F., McIntire, S.A. et al. (2015). Butyrfentanyl overdose resulting in diffuse alveolar hemorrhage. Pediatrics 135 (3): e740–e743. Concheiro, M., Chesser, R., Pardi, J. and Cooper, G. (2018). Postmortem toxicology of new synthetic opioids. Frontiers in Pharmacology 9: 1210. Corkery, J.M., Streete, P., Claridge, H. et  al. (2019). Characteristics of deaths associated with kratom use. Journal of Psychopharmacology 33 (9): 1102–1123. Davidsen, A.B., Mardal, M., Holm, N.B. et  al. (2020). Ketamine analogues: Comparative toxicokinetic in vitro-­in vivo extrapolation and quantification of 2-­fluorodeschloroketamine in forensic blood and hair samples. Journal of Pharmaceutical and Biomedical Analysis 180: 113049.

Drummer, O.H. (2019). Fatalities caused by novel opioids: A review. Forensic Sciences Research 4 (2): 95–110. Drummer, O.H., Gerostamoulos, D. and Woodford, N.W. (2019). Cannabis as a cause of death: A review. Forensic Science International 298: 298–306. Fedotov, Y. (2019). Global review of drug demand and supply. World Drug Report 2019. Vienna, Austria, United Nations. Fels, H., Lottner-­Nau, S., Sax, T. et  al. (2019). Postmortem concentrations of the synthetic opioid U-­47700 in 26 fatalities associated with the drug. Forensic Science International 301: e20–e28. Frisoni, P., Bacchio, E., Bilel, S. et al. (2018). Novel synthetic opioids: The pathologist’s point of view. Brain Sciences 8 (9): 170. Gerace, E., Salomone, A. and Vincenti, M. (2018). Analytical approaches in fatal intoxication cases involving new synthetic opioids. Current Pharmaceutical Biotechnology 19 (2): 113–123. Gerostamoulos, D., Drummer, O.H. and Woodford, N.W. (2015). Deaths linked to synthetic cannabinoids. Forensic Science, Medicine and Pathology 11 (3): 478. Giorgetti, A., Busardo, F.P., Tittarelli, R. et al. (2020). Post-­mortem toxicology: A systematic review of death cases involving synthetic cannabinoid receptor agonists. Frontiers in Psychiatry 11: 464. Guerrieri, D., Kjellqvist, F., Kronstrand, R. and Green, H. (2019). Validation and cross-­reactivity data for fentanyl analogs with the immunalysis fentanyl ELISA. Journal of Analytical Toxicology 43 (1): 18–24. Helander, A., Beck, O. and Backberg, M. (2015). Intoxications by the dissociative new psychoactive substances diphenidine and methoxphenidine. Clinical Toxicology (Philadelphia) 53 (5): 446–453. Helander, A., Stojanovic, K., Villen, T. and Beck, O. (2018). Detectability of fentanyl and designer fentanyls in urine by 3 commercial fentanyl immunoassays. Drug Testing and Analysis 10 (8): 1297–1304. Huppertz, L.M., Moosmann, B. and Auwarter, V. (2018). Flubromazolam -­ Basic pharmacokinetic evaluation of a highly potent designer benzodiazepine. Drug Testing and Analysis 10 (1): 206–211. Hvozdovich, J.A., Chronister, C.W., Logan, B.K. and Goldberger, B. (2020). Case report: Synthetic cannabinoid deaths in State of Florida prisoners. Journal of Analytical Toxicology e-­pub, 1–3. Ivanov, I.D., Stoykova, S., Ivanova, E. et al. (2019). A case of 5F-­ADB / AMB abuse: Drug-­ induced or drug-­ related death? Forensic FUB-­ Science International 297: 372–377. Iversen, L., White, M. and Treble, R. (2014). Designer psychostimulants: Pharmacology and differences. Neuropharmacology 87: 59–65. Iwersen-­Bergmann, S., Lehmann, S., Heinemann, A. et al. (2019). Mass poisoning with NPS: 2C-­ E and Bromo-­ Dragon Fly. International Journal of Legal Medicine 133 (1): 123–129. Kaneko, S. (2017). Motor vehicle collisions caused by the ’super-­strength’ synthetic cannabinoids, MAM-­2201, 5F-­PB-­22, 5F-­AB-­PINACA, 5F-­ AMB and 5F-­ADB in Japan experienced from 2012 to 2014. Forensic Toxicology 35 (2): 244–251. Karila, L., Lafaye, G., Scocard, A. et al. (2018). MDPV and alpha-­PVP use in humans: The twisted sisters. Neuropharmacology 134 (Pt A): 65–72. Klingberg, J., Cawley, A., Shimmon, R. and Fu, S. (2019). Collision-­ induced dissociation studies of synthetic opioids for non-­targeted analysis. Frontiers in Chemistry 7: 331.

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References

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Pettersson Bergstrand, M., Helander, A. and Beck, O. (2016). Development and application of a multi-­component LC-­MS/MS method for determination of designer benzodiazepines in urine. Journal of Chromatography B 1035: 104–110. Pope, J.D., Weng Choy, K., Drummer, O.H. and Schneider, H.G. (2018). Novel benzodiazepines (clonazolam and flubromazolam) identified in candy-­like pills. The Journal of Applied Laboratory Medicine 3 (1): 48–55. Rodda, L.N., Pilgrim, J.L., Di Rago, M. et al. (2017). A cluster of fentanyl-­ laced heroin deaths in 2015  in Melbourne, Australia. Journal of Analytical Toxicology 1–7. Roque Bravo, R., Carmo, H., Carvalho, F. et  al. (2019). Benzo fury: A new trend in the drug misuse scene. Journal of Applied Toxicology 39 (8): 1083–1095. Shanks, K.G. and Behonick, G.S. (2016). Death after use of the synthetic cannabinoid 5F-­AMB. Forensic Science International 262: e21–e24. Shanks, K.G., Winston, D., Heidingsfelder, J. and Behonick, G. (2015). Case reports of synthetic cannabinoid XLR-­11 associated fatalities. Forensic Science International 252: e6–e9. Shapiro, A.P., Krew, T.S., Vazirian, M. et al. (2019). Novel ways to acquire designer benzodiazepines: A case report and discussion of the changing role of the Internet. Psychosomatics 60 (6): 625–629. Sigel, E. and Steinmann, M.E. (2012). Structure, function, and modulation of GABA(A) receptors. Journal of Biological Chemistry 287 (48): 40224–40231. Sofalvi, S., Schueler, H.E., Lavins, E.S. et al. (2017). An LC-­MS-­MS Method for the Analysis of Carfentanil, 3-­Methylfentanyl, 2-­Furanyl Fentanyl, Acetyl Fentanyl, Fentanyl and Norfentanyl in Postmortem and Impaired-­ Driving Cases. Journal of Analytical Toxicology 41 (6): 473–483. Tang, M.H.Y., Li, T.C., Lai, C.K. et al. (2020). Emergence of new psychoactive substance 2-­fluorodeschloroketamine: Toxicology and urinary analysis in a cluster of patients exposed to ketamine and multiple analogues. Forensic Science International 312: 110327. Theofel, N., Moller, P., Vejmelka, E. et al. (2019). A Fatal Case Involving N-­Ethyldeschloroketamine (2-­Oxo-­PCE) and Venlafaxine. Journal of Analytical Toxicology 43(2): e2–e6. Tynon, M., Homan, J., Kacinko, S. et  al. (2017). Rapid and sensitive screening and confirmation of thirty-­four aminocarbonyl/carboxamide (NACA) and arylindole synthetic cannabinoid drugs in human whole blood. Drug Testing and Analysis 9 (6): 924–934. UNODC (2020). Current NPS Threats, Volume II. UNODC. Vienna, Austria, UNODC. Wood, D.M., Davies, S., Puchnarewicz, M. et  al. (2012). Acute toxicity associated with the recreational use of the ketamine derivative methoxetamine. European Journal of Clinical Pharmacology 68 (5): 853–856. Worob, A. and Wenthur, C. (2019). DARK classics in chemical neuroscience: Synthetic cannabinoids (Spice/K2). ACS Chemical Neuroscience 11 (23): 3881–3892. Zaami, S., Giorgetti, R., Pichini, S. et  al. (2018). Synthetic cathinones related fatalities: An update. European Review for Medical and Pharmacological Sciences 22 (1): 268–274. Zawilska, J.B. (2017). An expanding world of novel psychoactive substances: Opioids. Front Psychiatry 8: 110. Zawilska, J.B. and Wojcieszak, J. (2019). An expanding world of new ­psychoactive substances-­designer benzodiazepines. Neurotoxicology 73: 8–16.

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Kraemer, M., Boehmer, A., Madea, B. and Maas, A. (2019). Death cases involving certain new psychoactive substances: A review of the literature. Forensic Science International 298: 186–267. Kraemer, M., Fels, H., Dame, T. et al. (2019). Mono-­/polyintoxication with 5F-­ADB: A case series. Forensic Science International 301: e29–e37. Krotulski, A.J., Papsun, D.M., Kacinko, S.L. and Logan, B.K. (2020). Isotonitazene Quantitation and Metabolite Discovery in Authentic Forensic Casework. Journal of Analytical Toxicology 44 (6): 521–530. Labay, L.M., Caruso, J.L., Gilson, T.P. et al. (2016). Synthetic cannabinoid drug use as a cause or contributory cause of death. Forensic Science International 260: 31–39. Lo Faro, A.F., Di Trana, A., La Maida, N. et al. (2020). Biomedical analysis of New Psychoactive Substances (NPS) of natural origin. Journal of Pharmaceutical and Biomedical Analysis 179: 112945. Łukasik-­ Głębocka, M., Sommerfeld, K., Teżyk, A. et  al. (2016). Flubromazolam  – A new life-­threatening designer benzodiazepine. Clinical Toxicology (Phila) 54 (1): 66–68. Maeda, H., Kikura-­ Hanajiri, R., Kawamura, M. et  al. (2018). AB-­ CHMINACA-­induced sudden death from non-­cardiogenic pulmonary edema. Clinical Toxicology (Philadelphia) 56 (2): 143–145. Mallappallil, M., Sabu, J., Friedman, E.A. and Salifu, M. (2017). What do we know about opioids and the kidney? International Journal of Molecular Sciences 18 (1): 223. Mardal, M., Andreasen, M.F., Mollerup, C.B. et al. (2019). HighResNPS. com: An online crowd-­sourced HR-­MS database for suspect and non-­ targeted screening of new psychoactive substances. Journal of Analytical Toxicology 43 (7): 520–527. McCain, K.R., Jones, J.O., Chilbert, K.T. et al. (2018). Impaired Driving Associated with the Synthetic Cannabinoid 5f-­Adb. Journal of Forensic Science & Criminology 6 (1). Mercieca, G., Odoardi, S., Cassar, M. and Strano Rossi, S. (2018). Rapid and simple procedure for the determination of cathinones, amphetamine-­ like stimulants and other new psychoactive substances in blood and urine by GC-­MS. Journal of Pharmaceutical and Biomedical Analysis 149: 494–501. Michou, P. (2019). EU Drug Markets Report. The Hague, Netherlands, European monitoring centre for drugs and drug addiction. Mollerup, C.B., Dalsgaard, P.W., Mardal, M. and Linnet, K. (2017). Targeted and non-­targeted drug screening in whole blood by UHPLC-­ TOF-­MS with data-­independent acquisition. Drug Testing and Analysis 9 (7): 1052–1061. Moody, M.T., Diaz, S., Shah, P. et al. (2018). Analysis of fentanyl analogs and novel synthetic opioids in blood, serum/plasma, and urine in forensic casework. Drug Testing and Analysis 10 (9): 1358–1367. Morris, H. and Wallach, J. (2014). From PCP to MXE: A comprehensive review of the non-­medical use of dissociative drugs. Drug Testing and Analysis 6 (7–8): 614–632. Ong, R.S., Kappatos, D.C., Russell, S.G.G. et  al. (2020). Simultaneous analysis of 29 synthetic cannabinoids and metabolites, amphetamines, and cannabinoids in human whole blood by liquid chromatography-­ tandem mass spectrometry  – A New Zealand perspective of use in 2018. Drug Testing and Analysis 12 (2): 195–214. Paul, A.B.M., Simms, L., Amini, S. and Paul, A.E. (2018). Teens and spice: A review of adolescent fatalities associated with synthetic cannabinoid use. Journal of Forensic Sciences 63 (4): 1321–1324.

52.4.1 Introduction

Pharmacokinetics

Most sedative-­hypnotics are rapidly absorbed via the gastrointestinal (GI) tract. Some of them are also used intravenously for rapid onset such as thiopental or midazolam. When taken orally, barbiturates and benzodiazepines are primarily absorbed in the small intestine. Because they act at receptors in the brain, they must cross the blood–brain barrier. For that purpose, they must be lipophilic, which in turn means that they usually have to be metabolized to be ready for excretion. Being that lipophilic, they are redistributed to other (fatty) body tissues explaining the rapid loss of clinical effects, e.g., of ultra-­short acting barbiturates or intravenously used benzodiazepines such as midazolam. Another consequence of their lipophilicity is that they are often highly protein-­bound. Thus, being not filtered by the kidneys, they have to be eliminated via metabolism (Lee and Ferguson 2011). There are also exceptions such as the more water-­soluble drugs chloral hydrate or meprobamate and the longer-­acting barbiturates. In Table 52.4.1, some pharmacokinetic parameters for sedative-­hypnotics discussed in this chapter are given (taken from Ratz et al. 1999; Aktories et al. 2004; Baselt 2008; Sweetman 2009; Baselt 2017; Landry et al. 2020).

Pharmacodynamics

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Sedative-­hypnotics are drugs that lead to sedation by limiting excitability, and/or induce drowsiness and sleep (hypnosis). All sedative-­hypnotics induce central nervous system (CNS) depression. They are, e.g., agonists at the GABA A receptor, thus enhancing the function of GABA-­mediated chloride channels that are the primary mediators of inhibitory neurotransmission in the brain (Lee and Ferguson 2011). These receptors are members of the Cys-­loop pentameric ligand-­gated ion channel (LGIC) superfamily with gamma-­aminobutyric acid (GABA) as an endogenous ligand. The active site of the GABA A receptor is the binding site for GABA. The activity of the receptor can also be modulated via other allosteric binding sites at the protein (Barnard et  al. 1998; Olsen and Sieghart 2008). They are therefore targets of sedative-­hypnotic drugs such as barbiturates, benzodiazepines, benzodiazepine-­ related drugs, or piperidinedione derivatives. Benzodiazepines also interact with specific benzodiazepine receptors that are not associated with the GABA receptor. These receptors had been labelled omega receptors (ω-­receptors), a term that is nowadays obsolete. Instead, it has been recommended to use the benzodiazepine omega site (BZ/ω site). In fact, the heteromeric LGICs offer much greater heterogeneity than other known receptor subtypes. More details can be found in Barnard et al. 1998 and Olsen and Sieghart 2008. Some sedative-­ hypnotics also act by inhibiting excitatory neurotransmission via glutamate mediated N-­methyl-­d-­aspartate (NMDA) receptors. Newer sleep aids, such as melatonin or its analogs, are agonists at melatonin receptor subtypes MT-­1 and MT-­2 in the suprachiasmatic nucleus of the brain. Dexmedetomidine, used by intensive care units and anesthetists, is relatively unique in its ability to

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Sedatives and hypnotics form a heterogeneous group of substances. According to the WHO Anatomical Therapeutic Chemical (ATC) index, they can be divided in the following subgroups: barbiturates, aldehydes and derivatives, benzodiazepine derivatives, piperidinedione derivatives, benzodiazepine-­related drugs, melatonin receptor agonists, and other hypnotics and sedatives (WHO Collaborating Centre for Drug Statistics ­ Methodology 2019). In this ATC classification system, the active substances are divided into different groups according to the organ or system on which they act and their therapeutic, pharmacological, and chemical properties. This system was therefore adopted for this chapter.

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Thomas Krämer and Andrea E. Steuer

­ rovide sedation by acting as an agonist of α2-­adrenergic recepp tors similar to clonidine. Suvorexant is the first approved drug belonging to the group of dual orexin receptor antagonists (DORA). It is inducing sleep through the antagonism at excitatory orexin receptors that are involved in the regulation of wakefulness, for example, through activation of tubero-­mammillary histamine neurons that secrete wake-­maintaining histamine. In the meantime, the FDA has approved another DORA (i.e. lemborexant) for the treatment of insomnia in adults. Suvorexant and lemborexant bind to both OX1R and OX2R as reversible competitive antagonists. As lemborexant shows a stronger inhibition effect on OX2R than QX1R, it is expected to increase the non-­ REM sleep (Kishi et al. 2020).

N

52.4  Sedatives, Hypnotics

TOXICOLOGY

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Poisoning With most of them being centrally depressing agents, overdoses with sedative-­hypnotics lead to typical symptoms such as altered level of consciousness, difficulties in thinking, drowsiness, faulty judgment, incoordination, shallow breathing, slurred speech, sluggishness, staggering, or at larger doses even coma. Not in all cases, CNS depression parallels respiratory depression. Having similar mechanisms of action (e.g. prolongation of the opening of the chloride channel of the GABA receptor by barbiturates and increasing of the ionophore opening by benzodiazepines), the combination of different sedative-­hypnotics leads to synergistic effects aggravating the intoxication. Especially, the combination of ethanol with sedative-­hypnotics is a typical complication (Buckley et al. 1995; Verstraete and Buylaert 1995; Wilkinson 1995; Treweek et al. 2010). Only the new DORA suvorexant and lemborexant do not show interaction with ethanol (Hoyer et al. 2020).

LY

Bioavailability (%)

Volume of distribution (L/kg)

Protein binding (%)

Clearance (mL/min)

Alprazolam

90

0.8

70

60

hepatic

20–24

40

8–12

7–15

0.3

97

30

Choral hydrate

12–(70)

0.6

35

90

1.4

90

Clomethiazole

90

7.9±1.1

60–70

TO

Clobazam

0.33 (V/F)

Dexmedetomidine

3–7

94

98

100

1.1

Diphenhydramine

55

5

80

Doxylamine

2.7

Eszopiclone

1.1–1.7

Ethchlorvynol

2.4–3.2 -­

1

Flunitrazepam

85

3 (V/F)

Glutethimide

2.7

O

Ketazolam

49–76

Melatonin

R

Meprobamate

Methohexital Methyprylon Midazolam

FO

Methaqualone 25

45

1.0

4 min (6–10 TCE)

50

18 (50)

40±10

2.3–5

130 (CL/F)

2.0 (70)

1.0

1.0

3–10 30

40 (70)

1.0

800

6

0.9

174–240

7–13

52–59

4–9

62

19–32

50

1200

0.9

1.0

80

200 (CL/F)

16

1.0

54

5–22

93 94

1.5 90

1–2 (40–100)

1.8–2.5

61–78

40–50 min

0.7

20

6–17

6

80

2

80

0.6–1.5

60

1

95

C

Lemborexant Medazepam

N

Flumazenil

TR IB

Diazepam

U

Clorazepate

SE

100

0.9

strong

100

Carbromal Chlordiazepoxide

14

1.5–3

R

Camazepam

Extrarenal fraction, Q0

8–12

U

Bromisoval

Half-­life, t1/2 (h)

O

Substance

N

Table 52.4.1  Some pharmacokinetic parameters of sedative-­hypnotics

20–60 800

6 min (2)

1.0

7–11 400

2 (1)

1.0 (Continued)

LY N O SE

Table 52.4.1  (Continued) Volume of distribution (L/kg)

Protein binding (%)

Nitrazepam

80

2

90

Nordazepam

Ca 100

1

98

Oxazepam

>90

0.6

95

0.9

Half-­life, t1/2 (h)

Extrarenal fraction, Q0

60

26

1.0

10

70 (8)

1.0 (1.0)

70

8

1.0

hepatic

3–10

R

Paraldehyde

Clearance (mL/min)

U

Bioavailability (%)

TO

Substance

Pentobarbital

80

0.6

50

Phenobarbital

ca 100

0.54

50

Ramelteon

1.0–1.1

82

0.5–2.4

Suvorexant

0.5–0.9

>99

7–15

Tasimelteon

0.8–1.8 >80

1.4

Thiopental

-­

2.5

Triazolam

45

1

70

Zopiclone

80

U 20

0.7

1–5

97

100’/60

12/17

1.0

85

240

11 (22)

1.0

90

400

2.5

1.0

7–11 (trichloroethanol)

60 92

300

2.5

1.0

45

230

5

0.95

Sources: Adapted from Ratz et al. (1999); Aktories et al. (2004); Baselt R.C. (2008); Baselt R.C. (2017).

R

1.0

100

0.5 1.3

FO

30

1.0–1.5

C

Zolpidem

O

Zaleplon

N

Triclofos

TR IB

Temazepam

4.4

0.9–1.2

1239

CHAPTER 52   Toxicology of Specific Substances

R

C

O

N

TR IB

U

Drug tolerance occurs with sedative-­hypnotics and is defined as a progressive diminution of the susceptibility to the effects of a drug, resulting from its continued administration. Higher doses of the drug are necessary to achieve the same effect. Tolerance happens when adaptive neural and receptor changes (plasticity) occur after repeated exposures. These changes can be of different kind: downregulation may lead to a decrease in the number of receptors, receptor desensitization may lead to a reduction of firing, or receptor shifting may lead to changes in the receptor. Another possibility is the reduction of coupling of sedative-­ hypnotics from their respective GABA A receptor site (Lee and Ferguson 2011). It is also known that, e.g., a chronic use of ­benzodiazepines decreases the activity of the benzodiazepine site on the GABA receptor and at the same time decreases the binding affinity of the barbiturate sites. This phenomenon is called cross-­tolerance. The WHO defines dependence syndrome as a cluster of behavioral, cognitive, and physiological phenomena that develop after repeated substance use and that typically include a strong desire to take the drug, difficulties in controlling its use, persisting in its use despite harmful consequences, a higher priority given to drug use than to other activities and obligations, increased tolerance, and sometimes a physical withdrawal state (WHO, Management of Substance Abuse). The dependence syndrome may be present for a specific psychoactive substance (e.g. tobacco, alcohol, or diazepam), for a class of substances (e.g. opioid drugs or the sedative-­hypnotics), or for a wider range of pharmacologically different psychoactive substances.

FO

SE

O

N

LY

Barbiturates were already introduced in the beginning of the 20th century. This class of medications prevailed the sedative-­hypnotic market for more than 50 years. Having a narrow therapeutic-­to-­ toxic ratio and as given above a high potential for abuse, barbiturates became a health problem in many countries. This was a major cause for the efforts of pharmaceutical companies to find less toxic alternatives. Barbiturates were the number one suicide pills for several decades (Sandberg 1953; Locket 1954; Long 1960; Barraclough et al. 1971). Barbiturate mortality was even used as an index for its use (Richman and Orlaw 1965), and the addition of emetics had been proposed to prevent suicides (Sandberg 1953). Nowadays, barbiturates have again gained questionable fame as the drug of choice for assisted suicide endorsed by right-­ to-­die societies (Flanagan and Rooney 2002; Bosshard et al. 2003; Nordentoft et al. 2007; Ogden et al. 2010). Barbiturates are still used in therapy, but with decreasing tendency. Phenobarbital and its precursor primidone are still used as anticonvulsants for which drug monitoring is necessary. Thiopental is widely used as short-­ time intravenous anesthetic. Besides other central depressing substances, thiopental and its metabolite pentobarbital are often to be monitored for decision of brain death. Barbiturates are 5,5-­disubstituted barbituric acid derivatives which have no CNS depressing activity (Figure 52.4.1). In contrast to barbituric acid, which is five times more acidic (pKa 4.0) than acetic acid (pKa 4.75), the 5,5-­disubstituted derivatives are only weak acids (pKa about 8). This is of importance for their biological effects and for their analytical behavior. Under physiological conditions, barbituric acid is deprotonated, while 5,5-­disubstituted analogs are mainly unionized and can cross the blood–brain barrier to develop their central activity. The addition of various side chains influences pharmacologic properties of the barbiturates. Lipophilicity increases with longer side chains leading to easier crossing of the blood–brain barrier, higher potency, and slower rates of elimination. However, the duration of action of barbiturates does not correlate well with their biological half-­ lives because pharmacokinetic processes such as absorption, (re) distribution, and metabolism to active metabolites also influence their clinical effects. When given orally, barbiturates are readily absorbed from the small intestine. Elimination takes place via liver and kidneys. Barbiturates with higher lipophilicity are more bound and are mainly metabolized in the liver. protein-­ Phenobarbital as one of the few barbiturates that are still in use is also excreted into urine as an unchanged drug. Urinary excretion can be significantly increased by urine alkalinization that is a

TO

Tolerance, dependence, and withdrawal

52.4.2 Barbiturates

R

Sedative-­hypnotics are often the cause of unconsciousness in medical emergencies and should therefore be included in typical toxicological screening analyses (i.e. general unknown or systematic toxicological analysis [STA]) (Sauvage et  al. 2006; Maurer 2007; Maurer 2009; Peters et al. 2009; Sauvage et al. 2009; Sturm et  al. 2010; Johnson and Botch 2011). An additional field for determination of sedative-­hypnotics is the diagnosis of brain death, i.e. the exclusion of effective plasma concentrations of drugs that might mimic a flat-­line EEG. A minimum consensus for toxicological analysis in this context includes relevant analytes (thiopental, pentobarbital, methohexital, phenobarbital, diazepam, nordazepam, midazolam, and propofol) (Peters et al. 2005a; Peters et al. 2005b). The determination of sedative-­hypnotics is often an issue in forensics, e.g., the question of penal responsibility of a criminal after ingestion of sedative-­hypnotics must be answered. They may also reduce the driving ability or the ability to work at machines (Kraemer and Maurer 2007). Furthermore, in cases of drug-­ facilitated sexual assault, determination of sedative-­hypnotics might be necessary (Elliott 2008; Butler and Welch 2009; Shbair et al. 2010a; Shbair et al. 2010b; Shbair and Lhermitte 2010; Drummer et al. 2019).

The withdrawal state is defined as a group of symptoms of variable clustering and severity occurring on absolute or relative withdrawal of a psychoactive substance after persistent use of that substance. It may be complicated by convulsions. Some of the sedative-­hypnotics, classically the barbiturates, benzodiazepines, and ethanol, are associated with characteristic potentially life-­ threatening withdrawal syndromes.

U

Clinical and forensic toxicological aspects

1240

TOXICOLOGY

N

LY

PART VII  

O

Figure 52.4.1  Common chemical structure of barbiturates

U

SE

cerebral edema, and multi-­organ system failure as a result of prolonged cardiorespiratory depression.

R

52.4.3  Aldehydes and derivatives Aldehydes, such as chloral hydrate or paraldehyde, were introduced as sedative-­hypnotic drugs into therapeutics more than 120 years ago.

Chloral hydrate Chloral hydrate is the oldest known synthetic sedative-­hypnotic drug. However, it is still used today, especially in pediatrics. However, adverse effects are more and more discussed (Grissinger 2019). Chemically, it is the hydrate of trichloroacetaldehyde. Formulations contain either chloral hydrate alone or combinations with acetylglycinamide or antipyrine, named as acetylglycinamide chloral hydrate or dichloralphenazone, respectively. Trichloroethanol, its glucuronide, dichloroacetic acid, and trichloroacetic acid have been identified as chloral hydrate metabolites as shown in Figure  52.4.2. Trichloroethanol as an

FO

R

C

O

N

TR IB

U

TO

treatment regimen that should increase poison elimination by the administration of intravenous sodium bicarbonate to produce urine with a pH > or = 7.5. The term urine alkalinization emphasizes that urine pH manipulation rather than a diuresis is the prime objective of treatment. Therefore, the use of the terms forced alkaline diuresis and alkaline diuresis is obsolete. It has to be stated that nowadays urine alkalinization cannot be recommended as first-­line treatment in cases of phenobarbital poisoning, as multiple-­dose activated charcoal is superior (Proudfoot et al. 2004). Barbiturates are known to be inducers of cytochrome P450 (CYP) enzymes in the liver. CYP2B6, CYP2C9, and CYPs 3A4, 5, 7 are affected (Flockhart 2010). Thus, they can accelerate their own metabolism and that of other medicaments such as anticoagulants, corticoids, lamotrigine, doxycycline, chloramphenicol, antimycotics of the azole-­type, and oral contraceptives, leading to partial or complete loss of therapeutic effects. As explained in the introduction, overdoses with sedative-­ hypnotics lead to typical symptoms such as altered level of consciousness, difficulties in thinking, drowsiness, faulty judgment, incoordination, shallow breathing, slurred speech, sluggishness, staggering, or at larger doses even coma. Despite of being named after barbiturates, the so-­called “barb blisters,” cutaneous bullae, are also present in intoxications with other sedative-­hypnotics. Therefore, the name “coma blister” is more common today. Coma blisters are lesions that occur in the setting of a variety of neurological diseases. They develop 48–72 h after the onset of unconsciousness. Their etiology is complex and cannot simply be explained by pressure effects (Kato et al. 1996; Maguiness et al. 2002; Waring and Sandilands 2007; Basu et al. 2009; Rocha et al. 2009). Another common sign of sedative-­hypnotics overdose is hypothermia, but together with the coma blisters, these signs may be more pronounced with barbiturates. In severe intoxication cases with barbiturates, death is caused by respiratory arrest and cardiovascular collapse. If the patient survives the initial overdose, death may result from other complications such as acute renal failure, pneumonia, acute lung injury,

Figure 52.4.2  Metabolic pathway of chloral hydrate to trichloroacetic acid and the active metabolite trichloroethanol

1241

CHAPTER 52   Toxicology of Specific Substances

R

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O

N

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benzodiazepines are used as tranquilizers, hypnotics, anesthetics, anticonvulsants, or muscle relaxants and belong to the most frequently prescribed drugs. In recent years, in parallel with the massive increase of new psychoactive substances (NPS), also new (designer) benzodiazepines entered the illegal drug market. The first of its kind was phenazepam, first identified and described in 2007. The majority of these compounds never underwent clinical testing usually required prior to official drug approval by the authorities. The actual potency and pharmacokinetic behavior of individual compounds therefore remain largely unknown. Nevertheless, it can be assumed that designer benzodiazepines, in principle, act similar to classic prescription ones with similar side effects and toxicity in overdoses (Manchester et al. 2018). In less than 1% of the patients, paradoxical reactions to benzodiazepines may occur, characterized by increased talkativeness, emotional release, excitement, and excessive movement. The exact mechanism of paradoxical reactions still remains unclear (Gutierrez et  al. 2001; Robin and Trieger 2002; Mancuso et  al. 2004; McKenzie and Rosenberg 2010; Landolt and Dijk 2019). aryl-­ 1,4-­ diazepine The classical benzodiazepines have a 5-­ structure, characterized by a benzene ring condensed to the 10-­ and 11-­positions of the 1,4-­diazepine ring. The aryl substituent at position 5 is usually phenyl or a 2-­ halogenated phenyl ring (Figure 52.4.3). Annulation of an imidazole ring in 1,2-­position leads to imidazo-­or diazolo-­benzodiazepines such as midazolam, alprazolam, or triazolam. The primary benzene ring can be replaced with a thienyl ring to give compounds such as brotizolam, clotiazepam, and bentazepam. There are also some 1,5-­benzodiazepines such as clobazam. Flumazenil is a benzodiazepine receptor antagonist, which is structurally related to the benzodiazepines (Figure 52.4.3). Most benzodiazepines are lipophilic and therefore highly protein-­bound, which prevents renal excretion. This means that they must first be metabolized in the liver before they can be eliminated from the body. During phase I metabolism, benzodiazepines are N-­dealkylated, hydroxylated at the phenyl ring in C-­5, and/or hydroxylated at C-­3. Metabolism can lead to common metabolites, i.e. oxazepam is a common metabolite in urine for many 1,4-­benzodiazepines, such as diazepam, nordazepam, temazepam, clorazepate, chlordiazepoxide, ketazolam, camazepam, or medazepam (Figure 52.4.4a). Other 1,4-­benzodiazepines have a nitro-­ substituent at C-­ 7 (flunitrazepam, clonazepam, nitrazepam, etc.). They are metabolized to 7-­amino metabolites. Alprazolam, midazolam, and triazolam can additionally be hydroxylated at the methyl group of the annulated imidazole ring (Figure 52.4.4b). Only phase II metabolism (mainly glucuronidation) leads to inactive metabolites. However, the resulting glucuronides may undergo an enterohepatic circulation that may be responsible for the second peak of diazepam plasma concentration (Ma and Sun 1993). Again, because of the individual pharmacokinetics of sedative-­hypnotics and the production of active metabolites, there is often little correlation between the therapeutic and biological half-­lives.

TR IB

U

TO

active metabolite also contributes to the pharmacologically active principle of chloral hydrate therapy. Acute toxicity includes liver disease, gastric irritation, cardiac irritability, and CNS depression. Physical dependence and tolerance as well as withdrawal symptoms have been observed after chronic ingestion. Deaths due to chloral hydrate overdose were reported, and the cause of death was mainly attributed to cardiac arrhythmias (Steinberg 1993). Application of flumazenil was shown as a suitable therapy for chloral hydrate poisoning in some cases, resulting in improved consciousness and increased respiratory rate and blood pressure (Donovan and Fisher 1989). Possible carcinogenic properties of chloral hydrate are the subject of many discussions. Trichloroethylene, formerly used as a general anesthetic and disinfectant, is known to form chloral hydrate as its main metabolite. It was removed from the drug market due to its genotoxic properties observed in rodent studies. Very high doses of chloral hydrate were also demonstrated to alter the number of chromosomes. However, the impact of such studies on genotoxicity of chloral hydrate in humans is still controversial (Steinberg 1993; Salmon et al. 1995). Coadministration of chloral hydrate and ethanol leads to a significant potentiation of the hypnotic effect (Gessner and Cabana 1970; Owen and Taberner 1980). Various mechanisms have been discussed aiming to explain this phenomenon: a general additive effect due to the central depressant actions of both drugs, induction, and inhibition of their metabolism, as well as coupling of oxidation and reduction of ethanol and chloral hydrate, respectively (Owen and Taberner 1980). This synergism provoked particular interest, partly as a result of the efficacy of a “mickey finn” (Inciardi 1977). Even today, chloral hydrate should still be considered as a possible ingredient of knockout drops, e.g., in the context of drug-­facilitated crimes (Ashok et al. 2016).

N

Paraldehyde

FO

R

C

O

Paraldehyde is the cyclic trimer of acetaldehyde and was used as a sedative-­hypnotic drug and in therapy of delirium tremens in the first half of the 20th century. Nowadays, it is replaced by newer sedative-­hypnotic drugs, such as benzodiazepines or z-­drugs, but is still used as a reserve drug in epileptic therapy in pediatrics (Gessner and Shakarjian 1985; Ahmad et  al. 2006). One major drawback was its offensive taste and odor resulting from its degradation to acetic acid. Despite of the impression gained from smelling the breath of paraldehyde-­treated patients, the drug is mostly metabolized, not excreted by the lungs (Lasagna 1972). Generally, paraldehyde was considered as a rather safe hypnotic.

52.4.4 Benzodiazepine derivatives The benzodiazepines were discovered in the late fifties of the last century. Leo Sternbach had found that the new compounds had hypnotic, anxiolytic, and muscle relaxant properties. In the early 1960s, chlordiazepoxide (Librium®) followed by diazepam (Valium®) had then been introduced. Today, dozens of different

1242

TOXICOLOGY

U

SE

O

N

LY

PART VII  

O N

CI

H N

CI

OH

N

CI

R Chlorazepate

Alprazolam/triazolam/midazolam

O

N

FO

Temazepam

N

R

R

N

Chlordiazepoxide

CI

N

O

C

OH CI

N+ O–

N

(b)

O

O

O

N

CI

H N

N

Diazepam

N

U

N

H N

TR IB

(a)

TO

R

Figure 52.4.3  Structures of classic 1,4-­benzodiazepines (a), triazolo-­(b)-­, and imidazo-­(c) benzodiazepines, clobazam as 1,5 benzodiazepine (d), and flumazenil (e) as benzodiazepine receptor antagonists.

CI

Nordazepam

H N N

N

HO

O

N

N

R

N

R OH

CI

N R

CI

N R

Oxazepam

Glucuronidation

Glucuronidation

Glucuronidation

Glucuronidation

Figure 52.4.4  Main metabolic steps of classic 1,4-­benzodiazepines to common metabolites (a) and of imidazo-­/triazolo-­benzodiazepines (b)

1243

CHAPTER 52   Toxicology of Specific Substances

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TR IB

N

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C

R

FO

52.4.5  Piperidinedione derivatives Piperidinedione derivatives as glutethimide, methyprylon, and pyrithyldione (Figure  52.4.5) were first marketed as sedative-­hypnotic drugs in the 1950s. Initially developed as non-­barbiturate drugs, aimed to produce less undesired effects like respiratory depression and hypotension, they are rarely prescribed today. Some derivatives were even withdrawn from

O

O HN

O NH

O

O Glutethimide

Methyprylon

O

N H Pyrithyldion

Figure 52.4.5  Chemical structures of piperidinediones

R

U

SE

O

N

LY

the market in different countries. Chemically, they have structural resemblance to barbitals, being slightly acidic compounds, and also act through interaction with GABA A receptors. As such, they also lead to addiction and withdrawal symptoms (Tyrer 1993). Because of their low solubility in aqueous fluids, absorption from the gastrointestinal tract is retarded, which should be considered in cases of overdoses (Maher et al. 1962). Piperidinedione derivatives are extensively metabolized to ­various partly active metabolites. Main metabolites of glutethimide are, for example, 1’-­hydroxyglutethimide, 4-­hydroxyglutethimide, 2-­phenylglutarimide, 2-­ethyl-­2-­phenyl-­glutaconimide, and alpha-­ phenyl-­gammabutyrolactone. At least for 4-hydroxy­glu­tethimide, 2-­phenylglutarimide, and alpha-­phenyl-­gammabutyrolactone, pharmacological effects were shown (Baselt 2008). The same is methylpyrithyldione (methylpersedon), the active true for 5-­ metabolite of methyprylon. Pharmacokinetic interactions of glutethimide with codeine were described, resulting in increased morphine plasma concentrations and therefore potentiation of the pharmacological codeine/morphine effects. Abuse of this combination, so-­called “loads,” is described to produce euphoric effects similar to heroin (Khajawall et al. 1982; Popa et al. 2003) but also increase the risk of central depression and addiction. Part of the underlying interaction mechanism was assumed to be a competitive inhibition of morphine glucuronidation by glutethimide or one of its metabolites. Furthermore, glutethimide showed induction of various enzymes; however, systematic investigations of drug metabolizing enzymes are not available (Popa et al. 2003). Typical symptoms of glutethimide overdoses were tachycardia, coma, hypothermia in early stages switching to fever in later stages, infrequent respiratory depression, sudden apnea, hypotension, and anticholinergic effects. Compared to barbiturates, piperidinedione derivatives were described to have a lower risk for respiratory depression, but a higher incidence of shock. The cause of death in cases of lethal overdoses was attributable to irreversible shock, cardiac arrest, and sudden apnea. A typical characteristic of piperidinedione overdoses is often long-­lasting comas, persisting in some cases for up to seven days. Cyclically varying degrees of CNS depression over this time were observed (Hansen et al. 1975). Accumulation of reactive metabolites has been postulated as the main reason for the described symptoms (Kennedy and Fischer 1979). Due to poor water-­solubility, gastric lavage, which is usually performed with aqueous liquids, is not effective in the treatment of glutethimide poisoning. Usefulness of hemodialysis is controversially discussed (Maher et al. 1962; Baselt 2008).

TO

Benzodiazepines are considered to be safe drugs. Even after large overdoses, fatalities after mono-­ intoxications with them are rare. Typically, patients with overdoses of benzodiazepines only need supportive care. Most often deaths occur with combinations of benzodiazepines with other centrally depressing substances with the most common one being ethanol (Karavokiros and Tsipis 1990; Koski et al. 2002). Even the long-­term use of benzodiazepines is usually not associated with specific organ toxicity or any other systemic injury. The biggest problem associated with the use of benzodiazepines is their distinctive ability of producing tolerance and dependence in patients. Psychological and physical dependence can develop within a few weeks or years of regular or repeated use (Ashton 2005; Authier et  al. 2009; Hidalgo and Sheehan 2009; Liebrenz et  al. 2010). Discontinuation after long-­term use often leads to severe withdrawal symptoms such as perceptual distortions, paresthesia, feelings of unreality or depersonalization, pain, visual disturbances, depression, paranoid thoughts and feelings of persecution, gastrointestinal symptoms, increased sensitivity to light, noise, taste and smell, anxiety, tension, agitation, restlessness, and sleep disturbance (Authier et al. 2009). Switching a chronic user of a particular benzodiazepine to another benzodiazepine with different receptor activities may also lead to withdrawal. Even fatal outcomes of withdrawal are reported (Lann and Molina 2009; Votaw et al. 2019). Benzodiazepines are commonly administered to medical ICU patients. Propylene glycol (1,2-­propanediol) is the solvent used to deliver lorazepam and diazepam intravenously. After prolonged doses of these benzodiazepines, the diluent may accumulate and may also be associated with toxicity such as metabolic acidosis and hyperosmolar state (Wilson et al. 2005). Drug–drug interactions may also occur with benzodiazepines and may prolong the half-­life of them or change their potency or duration of action. It is well known that CYP3A4 inhibitors such as ketoconazole, ritonavir, and others can significantly inhibit midazolam metabolism, leading to a several hundred times longer half-­life (Ahonen et  al. 1999; Hamaoka et al. 1999; Oda et al. 1999; Wang et al. 1999; Wang et  al. 2000; Kanazu et  al. 2005; Vossen et  al. 2007; Sugiyama individual expression of et  al. 2011). The inter-­and intra-­ CYP3A4 is highly variable (40fold!) in humans. CYP3A4 overexpression might be the reason for high metabolite rates after midazolam therapy in some patients.

LY

N

O

52.4.7  Melatonin receptor agonists Melatonin (N-­acetyl-­5-­methoxytryptamine) is a physiological substance produced in the pineal gland. As a hormone, its physiological roles in humans are thought to embrace diurnal rhythm, sleep, mood, immunity, reproduction, intestinal motility, and metabolism mediated by three specific melatonin receptor subtypes, namely, MT1, MT2, and the putative MT3 binding site (Ma et  al. 2005). Melatonin secretion is associated with darkness, resulting in higher levels and light in lower levels. Therefore, increased melatonin concentrations, usually present in the evening, are responsible for inducing sleep and regulation of the sleep-­ wake cycle. When MT1 is occupied by a ligand, neuronal firing is restrained, and hence, sleep is induced (Mets et al. 2010; Landolt and Dijk 2019). Lacking interactions with GABA receptors like typical sedative-­hypnotic drugs, melatonin was thought as a versatile alternative sleeping pill without the typical side effects. Melatonin is available as dietary supplement in North America but was also marketed as a therapeutic drug in Europe in 2007. However, the efficacy of melatonin is inconclusive, and a recent meta-­analysis

FO

R

C

O

N

TR IB

U

TO

Zopiclone, zolpidem, zaleplon, and eszopiclone are hypnotic agents that act at the benzodiazepine recognition site of GABA A receptors, namely, the BZ/ω 1 site. They are often summarized as z-­drugs. Unlike benzodiazepines, they have weak myorelaxant and anticonvulsant effects, which are explained by their receptor selectivity. This receptor selectivity was also claimed to be the key to the alleged absence of potential for dependence. They belong to the most commonly prescribed hypnotics (Joester et al. 2010; Rust et al. 2012). They have rapid onset of action and short elimination half-­life (zolpidem 2h, (es)zopiclone 5–6h, and zaleplon 1–2h). Z-­drugs have little effect on sleep architecture and do not shorten REM sleep stages (Besset et  al. 1995; Hemmeter et  al. 2000; Uchimura et al. 2006). Zolpidem (Figure 52.4.6) is metabolized mainly by CYP3A4 to inactive metabolites via oxidation of each of the methyl groups on the phenyl moieties and via hydroxylation of the imidazopyridine moiety (Pichard et  al. 1995; von Moltke et  al. 2002). Zaleplon (Figure  52.4.6) is extensively metabolized also via CYP3A4 to inactive metabolites, mainly via oxidation to 5-­oxo-­zaleplon with less than 1% of dose excreted unchanged in urine (Renwick et al. 1998). Zopiclone (Figure  52.4.6) is extensively metabolized by CYP3A4 and CYP2C8 via three major pathways: decarboxylation, oxidation, and demethylation (Becquemont et al. 1999). As eszopiclone is claimed to be the active enantiomer, dosage is lower (1–3 mg) than that for the racemate zopiclone. Various pharmacokinetic interactions of z-­drugs with other xenobiotics have been reported. They seem to be less clinically important than those of the conventional benzodiazepines. An explanation may be a difference in CYP metabolism. While benzodiazepines such as triazolam and midazolam are biotransformed almost entirely via CYP3A4, the newer z-­ drugs are biotransformed by several CYP isoenzymes in addition to CYP3A4, resulting in CYP3A4  inhibitors and inducers having less effect on their biotransformation. Mono-­intoxications with z-­drugs lead to drowsiness and CNS depression, but prolonged coma with respiratory depression are markedly rare. Severe intoxications or even fatalities are described, but in most (not all) cases there had been concomitant intake of

other drugs (Garnier et al. 1994). The benzodiazepine antagonist flumazenil might also work for the z-­drugs. Even if the z-­drugs were thought to be a less dangerous group of drugs than the benzodiazepines, it has turned out that tolerance, dependence, and withdrawal are also common with them (Baruch et  al. 2007; Cubala and Landowski 2007; Huang et al. 2007; Sharan et al. 2007; Cubala et al. 2008; Jana et  al. 2008; Spyridi et  al. 2009; Aggarwal and Sharma 2010; Hsieh et al. 2011; Mattoo et al. 2011; Wang et al. 2011; Schifano et al. 2019). The WHO Expert Committee on Drug Dependence has already taken a close look on z-­drugs. For zopiclone, they have stated that it has some abuse potential together with the capacity to produce withdrawal syndrome upon its discontinuation. In terms of the number of adverse drug reaction reports related to abuse received by the international drug monitoring program, zopiclone ranks higher than nitrazepam and temazepam.

SE

52.4.6 Benzodiazepine-­related drugs

TOXICOLOGY

U

PART VII  

R

1244

Figure 52.4.6  Chemical structures of the z-­drugs zaleplon, zolpidem, zopiclone, and eszopiclone.

1245

CHAPTER 52   Toxicology of Specific Substances

O

O

N H

O

H H N

H

O

N H

O

O

N H melatonin

ramelteon

tasimelteon

Figure 52.4.7  Structures of melatonin receptor agonists

R

U

SE

O

N

LY

The oral bioavailability for ramelteon is less than 2% due to extensive first-­pass metabolism, which includes oxidation of the furan ring, side-­ chain hydroxylation, and ketone formation (Obach and Ryder 2010). Only the side-­chain hydroxyl metabolite showed slight pharmacological activity, but the impact is still unclear. It is discussed that because of its longer half-­life and accumulation in plasma, contribution to the overall effects is possible. Tasimelteon provides an overall higher absolute bioavailability of approximately 38%, although it is heavily metabolized as well. The four formed hydroxyl metabolites also exhibited melatonin receptor affinity, but about one-­tenth lower than the parent (Ogilvie et  al. 2015; Atkin et  al. 2018). CYP1A2 and CYP3A4 were the main isoenzymes involved in the biotransformation of both melatonin analogs. In  vivo, fluvoxamine was found to raise serum levels of ramelteon and tasimelteon by ­70-­fold and 6.5-­fold, respectively. Cigarette smoking lowered tasimelteon’s exposure by approximately 40% (Obach and Ryder 2010; Ogilvie et al. 2015).

FO

R

C

O

N

TR IB

U

TO

failed to show a significant effect on sleep onset latency (Mets et al. 2010). Melatonin is mainly metabolized by hydroxylation to form 6-­hydroxymelatonin and to a minor extent O-­desmethylmelatonin. They are excreted mainly as the respective sulfates or glucuronides. CYP1A2 represents the major CYP enzyme catalyzing the initial metabolic steps, with minor contribution of CYP1A1, 1B1, and 2C19 (Ma et al. 2005). As a potential sedative-­hypnotic drug, several attempted suicides were reported, but no severe clinical outcomes were observed (Chung 2001). However, the use of melatonin as a drug is limited by its short biological half-­life, poor bioavailability, and the fact that it has numerous other actions. Therefore, chemical modifications were performed in order to synthesize derivatives of melatonin with more receptor selectivity and better pharmacokinetic properties (Miyamoto 2009). Chemical structures of melatonin and its analogs ramelteon and tasimelteon are given in Figure 52.4.7. Ramelteon, first marketed in 2005, was synthesized as part of a program aimed at limiting the conformational flexibility of the methoxy group of melatonin, whose orientation is important for optimal binding to the melatonin MT1 receptor (Uchikawa et al. 2002). Compared to melatonin, ramelteon has a six-­fold higher binding potency for the human MT1 and a three-­fold higher affinity for the human MT2, acting via shortening the latency to sleep onset and by maintaining sleep. However, no impact on sleep quality was observed. Furthermore, reduction in the pharmacological effects was seen after 6 months of chronic ingestion (Mets et al. 2010). Tasimelteon, approved by the FDA in 2014, provides similar potencies as ramelteon with regard to MT1 and 2.1–4.4 times greater affinity for MT2 over MT1 (Atkin et  al. 2018). So far, tasimelteon has been only used in the treatment of Non-­24-­Hour Sleep-­Wake Disorder in blind people, while other indications are under investigation. Both drugs were well tolerated and showed only mild side effects. No severe intoxications or overdoses with significant toxicity have been described. Ramelteon, for instance, showed no impact on breathing, neither apnea nor other depressant effects on the CNS were observed. Its abuse potential is low and no next-­day residual effects have been demonstrated. Adverse side effects observed ranged from nausea to dizziness and somnolence. The intake of tasimelteon may produce headaches, elevation of serum hepatic enzymes, and unusual dreams (Mets et  al. 2010; Torres et  al. 2015; Baselt 2017).

52.4.8 Other hypnotics and sedatives Several other substances belonging to a large diversity of different chemical drug classes were or are still used as sedatives and hypnotics. Some chemical structures are given in Figure  52.4.8. Although meprobamate and the antihistaminic drugs diphenhydramine and doxylamine are not enclosed in the ATC classification system, they still represent important sedative-­ hypnotic drugs and are therefore discussed under this subchapter.

Methaqualone Methaqualone (Figure 52.4.8) is a powerful sedative, which was widely used. It has a high potential for addiction and misuse and was therefore scheduled. In the United States, methaqualone was removed from the market in 1984, but it has been available for much longer in European countries in combination with diphenhydramine (trade name: Mandrax). In South Africa, it is still a threat with a lot of DFSA cases involving methaqualone (Tiemensma and Davies 2018). However, since modern sedative-­ hypnotics show much less toxicity, its use in pharmacotherapy has decreased. Common symptoms of methaqualone overdose include depressed levels of consciousness, tachycardia, dysarthria, hyporeflexia, and mydriasis (Bailey 1981).

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N N

N

O

clomethiazole

methaqualon

F

Cl

S

N

N

N O

O

NH

LY

N

N O

N

diphenhydramine

doxylamine

O

Br

O Cl

N H

H2N

N H

O Cl

Br

carbromal

bromisoval

O O

N H

NH2

Cl ethchlorvynol

R Cl

dexmedetomidine

TR IB

meprobamate

H

TO

N

U

O

HO

triclofos

NH2 O

Cl

P OH OH

SE

H2N

O

U

O

O

N

niaprazine

N

N N O

N

O

N

N

suvorexant

Figure 52.4.8  Chemical structures of other sedative-­hypnotic drugs

N

Clomethiazole

FO

R

C

O

Clomethiazole (Figure  52.4.8) has sedative, hypnotic, and anti-­ convulsive effects. It is used in the treatment of the alcohol-­ withdrawal syndrome including delirium tremens (Sychla et  al. 2017). Fatalities due to its intake are described (Klug and Schneider 1984). Compared to clomethiazole, combined carbamazepine and tiapride alcohol detoxification treatment shows a lower level of sedation and lack of addictive potential (Diehl et al. 2007).

Niaprazine

Niaprazine is a phenylpiperazine derivative (Figure 52.4.8), mainly prescribed to adolescents (Mancini et al. 2006). Pharmacological binding affinity to dopaminergic, noradrenergic, histaminergic, and muscarinic cholinergic receptors was described. The main metabolite of niaprazine, p-­fluoro-­phenylpiperazine, additionally exhibits affinity to serotonin receptors (Scherman et  al. 1988). Development of adverse side effects has not been observed (Ottaviano et al. 1991), and no literature is available on overdoses concerning the ingestion of niaprazine.

H1-­antihistaminics H1-­antihistaminics like diphenhydramine and doxylamine were initially used in the therapy of hypersensitivity reactions. However, their central depressant side effects changed their field of application to more sedative-­ hypnotic indications. Furthermore, diphenhydramine is therapeutically used for its antiemetic effects. Diphenhydramine and doxylamine are non-­ prescription drugs but can still lead to severe poisonings and have also been abused in drug-­facilitated crimes (Pragst et al. 2006). Overdoses of antihistamines lead to anticholinergic effects and also to blockade of fast sodium channels, which can result in cardiac conduction problems including sinus tachycardia, ventricular tachycardia, and torsades de pointes (Jeffery and Lytle-­Saddler 2008). Rhabdomyolysis and prolonged QT intervals have been described as further, rare complications (Ramachandran and Sirop  2008). Already three years after its introduction, the first fatality due to diphenhydramine toxicity was reported in 1949. The most common symptoms for most fatalities in literature were cardiac dysrhythmias, seizure activity, and/or sympathetic pupil responses (Nine and Rund 2006). Sodium channel blockade

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Dexmedetomidine

O

N

LY

Dexmedetomidine is a central α2 adrenergic agonist that decreases central presynaptic catecholamine release. It was approved by the FDA in 1999 for short-­term use in the critical care setting. Dexmedetomidine offers a unique ability of providing both sedation and moderate analgesia with only minimal respiratory depression. It has a wide safety margin, excellent sedative capacity, and moderate analgesic properties. Although its main use is currently in patients of surgical and non-­surgical intensive care units, other promising applications in neuroprotection, cardioprotection, and renoprotection are under investigation (Afonso and Reis 2012; Lee 2019; Zhao et al. 2020). Some case reports describe the promising and successful application of dexmedetomidine in the treatment of different toxidromes, such as serotonin syndrome (Rushton and Charlton 2014) and anticholinergic syndrome caused by a diphenhydramine overdose (Walker et al. 2014). The antioxidant effect of dexmedetomidine was also shown to improve of hepatotoxicity and nephrotoxicity caused by acetaminophen overdoses (Tas et al. 2019). Generally considered as a safe drug, nausea, dry mouth, bradycardia, and varying effects on blood pressure (usually, hypertension followed by hypotension) are described as the most common adverse effects. Although the incidence of severe bradycardia is low, it should be carefully monitored under dexmedetomidine treatment/medication, as few cases with cardiac arrest have been described (Lee 2019; Zhao et al. 2020)

R

Bromisoval also known as bromvalerylurea is a sedative-­hypnotic drug first marketed in 1908. The chronic use of bromisoval has been associated with bromine poisoning. The same applies for carbromal (Maes et  al. 1985). Several acute intoxications with patients being comatose have been described. Bromide salts, particularly potassium bromide, were popular sedatives in the 19th and early 20th centuries. Due to their chronic toxicity, they were removed from the market. Acute bromide poisoning mainly lead to gastrointestinal symptoms, with nausea, vomiting, and diarrhea. Chronic intoxications are difficult to diagnose and to differentiate from other CNS diseases as only more or less unspecific symptoms like fatigue, loss of appetite, stomach pain, rapid respiration, stupor, and delirium with hallucinations occur (Baselt 2008). Intoxications may still happen these days, and the clinical and forensic toxicologist must be aware of it (Thornton and Haws 2020).

SE

Bromides and bromine-­containing sedative-­hypnotics

Maximal serum concentrations are attained within 60–90  minutes. Only 10% of ECV is excreted unchanged via the urine, feces, or as the volatile alcohol through the lungs (Yell 1990). Ethchlorvynol produces psychological dependence, tolerance, and physical dependence. Blood levels greater than 38 mg/L are associated with coma, hypotension, areflexia, and respiratory depression requiring intubation (Yell 1990). Hemoperfusion has been proposed for treatment of severe cases (Tozer et al. 1974).

U

should be recognized as a complication of pediatric diphenhydramine overdose, and hypertonic sodium bicarbonate has been proposed as treatment (Cole et al. 2011; Palmer et al. 2020).

TO

Meprobamate

FO

R

C

O

N

TR IB

U

Meprobamate is a mild tranquilizer, which has been used since the early 1950s. It is also a metabolite of the centrally acting muscle relaxant carisoprodol. Today, it has lost its significance since more modern medicaments such as benzodiazepines or benzodiazepine-­related drugs (z-­drugs) show less side effects. Nevertheless, it is still in use and intoxications still occur (Charron et  al. 2005; Daval et  al. 2006). Typical symptoms of overdose include CNS depression, weakness, clonus and hyperreflexia, tachycardia, hypotension, and respiratory depression. Serious and sometimes fatal developments are known, resulting from hemodynamic disturbance and circulatory collapse, secondary to severe acute cardiac failure (Buire et al. 2009). The benzodiazepine antagonist flumazenil might also work for meprobamate overdoses (Roberge et al. 2000). Withdrawal from meprobamate can lead to severe autonomic instability and death (Zink 2020).

Triclofos

Triclofos is a prodrug, which is metabolized in the liver to trichloroethanol, hence displaying similar effects as chloral hydrate (see Chapter 52.43).

Ethchlorvynol Ethchlorvynol was first introduced to the pharmaceutical market in 1955. Intoxication leads to effects similar to those of barbiturates, mainly characterized by a prolonged deep coma (Gustafsson et al. 1989; Baselt 2008). When taken orally at therapeutic doses, it is rapidly absorbed with a distribution half-­life of 1–3 hours.

Dual Orexin Receptor Antagonists (DORA): Suvorexant and lemborexant Suvorexant is a new sedative drug, first approved in 2015, and belongs to the new compound class of DORA. In 2019, another DORA lemborexant has also been approved by the US FDA. It has similar indications and properties as suvorexant, but because lemborexant shows a stronger inhibition effect on OX2R than QX1R, it is expected to increase the non-­REM sleep (Kishi et al. 2020). Orexins are peptides initially discovered in 1998 that are mainly synthesized in the lateral hypothalamus from where they are transported along neurons throughout the brain. Orexins act as excitatory neurotransmitters via two kinds of receptors. Their primary physiological role is the regulation of wakefulness, for example, through activation of tubero-­mammillary histamine neurons that secrete wake-­ maintaining histamine. Antagonism of the orexin system has been associated with diseases such as narcolepsy and catalepsy. With suvorexant, the first p ­ harmacological

dependence, and withdrawal are also quite common with sedative-­hypnotics and must be considered. The new DORA seem to have less side effects, but the future will tell, if this will remain so.

References

SE

O

N

LY

Afonso, J. and Reis, F. (2012). Dexmedetomidine: Current role in anesthesia and intensive care. Revista Brasileira de Anestesiologia 62 (1): 118–133. Aggarwal, A. and Sharma, D.D. (2010). Zolpidem withdrawal delirium: A case report. Journal of Neuropsychiatry and Clinical Neurosciences 22 (4): 451-­o e27–e28. Ahmad, S., Ellis, J.C., Kamwendo, H. and Molyneux, E. (2006). Efficacy and safety of intranasal lorazepam versus intramuscular paraldehyde for protracted convulsions in children: an open randomised trial. The Lancet 367 (9522): 1591–1597. Ahonen, J., Olkkola, K.T., Takala, A. and Neuvonen, P.J. (1999). Interaction between fluconazole and midazolam in intensive care patients. Acta Anaesthesiologica Scandinavica 43 (5): 509–514. Aktories, K., Förstermann, U., Hofmann, F. and Starke, K. (2004). Allgemeine und spezielle Pharmakologie und Toxikologie. München: Urban & Fischer. Ashok, J., Nair, M. and Friedman, R. (2016). Drug-­facilitated sexual assaults. In: A. Phenix and H. Hoberman (eds.), Sexual Offending. New York: Springer. Ashton, H. (2005). The diagnosis and management of benzodiazepine dependence. Current Opinion in Psychiatry 18 (3): 249–255. Atkin, T., Comai, S. and Gobbi, G. (2018). Drugs for insomnia beyond benzodiazepines: Pharmacology, clinical applications, and discovery. Pharmacological Reviews 70 (2): 197–245. Authier, N., Balayssac, D., Sautereau, M. et  al. (2009). Benzodiazepine dependence: focus on withdrawal syndrome. Annales Pharmaceutiques Françaises 67 (6): 408–413. Bailey, D.N. (1981). Methaqualone ingestion: Evaluation of present status. Journal of Analytical Toxicology 5 (6): 279–282. Barnard, E.A., Skolnick, P., Olsen, R.W. et al. (1998). International Union of Pharmacology. XV. Subtypes of gamma-­aminobutyric acidA receptors: Classification on the basis of subunit structure and receptor function. Pharmacological Reviews 50 (2): 291–313. Barraclough, B.M., Nelson, B., Bunch, J. and Sainsbury, P. (1971). Suicide and barbiturate prescribing. Journal of the Royal College of General Practitioners 21 (112): 645–653. Baruch, E., Vernon, L.F. and Hasbun, R.J. (2007). Intractable nausea caused by zolpidem withdrawal: a case report. Journal of Addiction Medicine 1 (1): 48–50. Baselt, R.C. (2008). Disposition of Toxic Drugs and Chemicals in Man. Foster City: Chemical Toxicology Institute. Baselt, R.C. (2017). Disposition of Toxic Drugs and Chemicals in Man. Foster City: Chemical Toxicology Institute. Basu, A., Brown, S., Kirkham, N. et al. (2009). Coma blisters in 2 children on anticonvulsant medication. Journal of Child Neurology 24 (8): 1021–1025.

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antagonist of the orexin receptor has been developed to suppress wakefulness, thereby consequently inducing sleep (Kripke 2015; Keks et al. 2017; Landolt and Dijk 2019). Depending on the dose, positive effects on both sleep onset and sleep maintenance were observed. Optimal dosing in terms of pharmacological effects and acceptable side effects are still critically discussed. Doses of 30–40  mg from initial clinical trials exhibited more compelling drug effects in contrast to the recommended oral doses of only 10–20 mg (lemborexant 5–10 mg). From the point of safety, it is strongly recommended not to exceed doses of 20  mg (Kripke 2015; Keks et  al. 2017; Landolt and Dijk 2019) (Scott 2020). Reported side effects include somnolence, muscle weakness, abnormal dreams, and headache. Next-­ day drowsiness was reported as low to moderate with no differences between older or younger people. Therefore, little effect on next-­morning driving performance was assumed. However, it should be mentioned that the development of another DORA, almorexant, was stopped in 2011 after concerns about its side effects (Keks et al. 2017; Waters et al. 2018; Landolt and Dijk 2019). Oral lemborexant was generally well tolerated in adults with insomnia disorder with a tolerability profile similar to that with placebo treatment (Scott 2020). In contrast to classic benzodiazepine sedatives, suvorexant is meant for long-­term therapy. Clinical trials so far have been promising and showed pharmacological effects for up to a year without significant rebound insomnia. No or minimal withdrawal effects were observed, which leads to the conclusion that the overall dependence potential appears low. However, this is controversially discussed as another study demonstrated that suvorexant had a similar ‘liking’ response to zolpidem which point to a certain abuse potential (Keks et al. 2017). Suvorexant is extensively metabolized by either aliphatic or aromatic primary hydroxylation followed by oxidation to the respective carboxylic acid and glucuronidation to pharmacologically inactive metabolites. Mainly CYP3A4 and to lesser extent CYP2C19 and CYP2D6 were shown to be involved in the initial metabolic steps (Cui et  al. 2016; Baselt 2017; Skillman and Kerrigan 2020). So far, no data on toxic or fatal overdoses with suvorexant or lemborexant have been described. Waters et al. detected suvorexant in three postmortem cases, but never as the (primary) cause of death (Waters et al. 2018).

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52.4.9 Summary

Sedatives and hypnotics form a heterogeneous group of xenobiotics with varying mechanisms of action. They are drugs that lead to sedation by limiting excitability, and/or induce drowsiness and sleep (hypnosis). All sedative-­hypnotics induce central nervous system (CNS) depression and can be the cause of unconsciousness in medical emergencies. Besides their importance in clinical toxicology, they are also targets for the forensic toxicologist, e.g., for the question of penal responsibility and for driving under the influence of drugs (DUID) or drug-­ facilitated sexual assault cases, as well as for investigations about cause of death. Tolerance,

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Kato, N., Ueno, H. and Mimura, M. (1996). Histopathology of cutaneous changes in non-­ drug-­ induced coma. American Journal of Dermatopathology 18 (4): 344–350. Keks, N.A., Hope, J. and Keogh, S. (2017). Suvorexant: Scientifically interesting, utility uncertain. Australas Psychiatry 25 (6): 622–624. Kennedy, K.A. and Fischer, L.J. (1979). Quantitative and stereochemical aspects of glutethimide metabolism in humans. Drug Metabolism and Disposition: The Biological Fate of Chemicals 7 (5): 319–324. Khajawall, A.M., Sramek, J.J., Jr. and Simpson, G.M. (1982). ’Loads’ alert. Western Journal of Medicine 137 (2): 166–168. Kishi, T., Nomura, I., Matsuda, Y. et al. (2020). Lemborexant vs suvorexant for insomnia: A systematic review and network meta-­analysis. Journal of Psychiatric Research 128: 68–74. Klug, E. and Schneider, V. (1984). [Clomethiazole poisoning]. Zeitschrift für Rechtsmedizin. Journal of Legal Medicine 93 (2): 89–94. Koski, A., Ojanpera, I. and Vuori, E. (2002). Alcohol and benzodiazepines in fatal poisonings. Alcoholism, Clinical and Experimental Research 26 (7): 956–959. Kraemer, T. and Maurer, H.H. (2007). Forensic toxicology: Sedatives and hypnotics. In: M. Bogusz (ed.), Handbook of analytical separation sciences: Forensic Sciences, 2nd ed. Amsterdam: Elsevier Science. Kripke, D.F. (2015). Is suvorexant a better choice than alternative hypnotics? F1000Research 4: 456. Landolt, H.-­ P. and Dijk, D.-­ J. (2019). Handbook of Experimental Pharmacology. Zurich, Switzerland and Guildford, UK: Springer. Landry, I., Nakai, K., Ferry, J. et al. (2020). Pharmacokinetics, pharmacodynamics, and safety of the dual orexin receptor antagonist lemborexant: Findings from single-­dose and multiple-­ascending-­dose phase 1 studies in healthy adults. Clinical Pharmacology in Drug Development 10 (2): 153–165. Lann, M.A. and Molina, D.K. (2009). A fatal case of benzodiazepine withdrawal. American Journal of Forensic Medicine and Pathology 30 (2): 177–179. Lasagna, L. (1972). Hypnotic drugs. New England Journal of Medicine 287 (23): 1182–1184. Lee, D.C. and Ferguson, K.L. (2011). Sedative-­Hypnotics. In: L.S. Nelson, N.A. Lewin, M.A. Howland, R.S. Hoffmann, L.R. Goldfrank and N.E. Flomenbaum (eds.), Goldfrank’s Toxicologic Emergencies. New  York: McGraw-­Hill Medical. Lee, S. (2019). Dexmedetomidine: present and future directions. Korean Journal of Anesthesiology 72 (4): 323–330. Liebrenz, M., Boesch, L., Stohler, R. and Caflisch, C. (2010). Benzodiazepine dependence: when abstinence is not an option. Addiction 105 (11): 1877–1878. Locket, S. (1954). Acute barbiturate poisoning and suicide. Medicine Illustrated 8 (4): 244–253. Long, R.H. (1960). Barbiturates, automatism and suicide. Postgraduate Medicine 28: A-­56–72. Ma, X., Idle, J.R., Krausz, K.W. and Gonzalez, F.J. (2005). Metabolism of melatonin by human cytochromes p450. Drug Metabolism and Disposition: The Biological Fate of Chemicals 33 (4): 489–494. Ma, Y.M. and Sun, R.Y. (1993). Second peak of plasma diazepam concentration and enterogastric circulation. Zhongguo Yao Li Xue Bao. Acta Pharmacologica Sinica 14 (3): 218–221.

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Rocha, J., Pereira, T., Ventura, F. et al. (2009). Coma blisters. Case Reports in Dermatology 1 (1): 66–70. Rushton, W.F. and Charlton, N.P. (2014). Dexmedetomidine in the treatment of serotonin syndrome. Annals of Pharmacotherapy 48 (12): 1651–1654. Rust, K.Y., Baumgartner, M.R., Meggiolaro, N. and Kraemer, T. (2012). Detection and validated quantification of 21 benzodiazepines and 3 “z-­drugs” in human hair by LC-­MS/MS. Forensic Science International 215 (1–3): 64–72. Salmon, A.G., Kizer, K.W., Zeise, L. et al. (1995). Potential carcinogenicity of chloral hydrate  – a review. Journal of Toxicology: Clinical Toxicology 33 (2): 115–121. Sandberg, F. (1953). The combination of barbiturates with an emetic as a method for the prevention of suicide. Acta Physiologica Scandinavica 28 (2–3): 272–278. Marcoux, F. et  al. (2009). General Sauvage, F.L., Picard, N., Saint-­ unknown screening procedure for the characterization of human drug metabolites in forensic toxicology: applications and constraints. Journal of Separation Science 32 (18): 3074–3083. Sauvage, F.L., Saint-­Marcoux, F., Duretz, B. et  al. (2006). Screening of drugs and toxic compounds with liquid chromatography-­linear ion trap tandem mass spectrometry. Clinical Chemistry 52 (9): 1735–1742. Scherman, D., Hamon, M., Gozlan, H. et al. (1988). Molecular pharmacology of niaprazine. Progress in Neuro-­ Psychopharmacology and Biological Psychiatry 12 (6): 989–1001. Schifano, F., Chiappini, S., Corkery, J.M. and Guirguis, A. (2019). An insight into Z-­drug abuse and dependence: An examination of reports to the European Medicines Agency database of suspected adverse drug reactions. International Journal of Neuropsychopharmacology 22 (4): 270–277. Scott, L.J. (2020). Lemborexant: First approval. Drugs 80 (4): 425–432. Sharan, P., Bharadwaj, R., Grover, S. et al. (2007). Dependence syndrome and intoxication delirium associated with zolpidem. National Medical Journal of India 20 (4): 180–181. Shbair, M.K., Eljabour, S., Bassyoni, I. and Lhermitte, M. (2010a). Drugs involved in drug-­facilitated crimes – part II: Drugs of abuse, prescripthe-­ counter medications. A review. Annales tion and over-­ Pharmaceutiques Françaises 68 (6): 319–331. Shbair, M.K., Eljabour, S. and Lhermitte, M. (2010b). Drugs involved in facilitated crimes: part I: alcohol, sedative-­ hypnotic drugs, drug-­ hydroxybutyrate and ketamine. A review. Annales gamma-­ Pharmaceutiques Françaises 68 (5): 275–825. Shbair, M.K. and Lhermitte, M. (2010). Drug-­ facilitated crimes: Definitions, prevalence, difficulties and recommendations. A review. Annales Pharmaceutiques Françaises 68 (3): 136–147. Skillman, B. and Kerrigan, S. (2020). CYP450-­mediated metabolism of suvorexant and investigation of metabolites in forensic case specimens. Forensic Science International 312: 110307. Spyridi, S., Diakogiannis, I., Nimatoudis, J. et  al. (2009). Zolpidem dependence in a geriatric patient: a case report. Journal of the American Geriatrics Society 57 (10): 1962–1963. Steinberg, A.D. (1993). Should chloral hydrate be banned? Pediatrics 92 (3): 442–446. Sturm, S., Hammann, F., Drewe, J. et al. (2010). An automated screening method for drugs and toxic compounds in human serum and urine

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Walker, A., Delle Donne, A., Douglas, E. et al. (2014). Novel use of dexmedetomidine for the treatment of anticholinergic toxidrome. Journal of Medical Toxicology 10 (4): 406–410. Wang, J.S., Backman, J.T., Kivisto, K.T. and Neuvonen, P.J. (2000). Effects of metronidazole on midazolam metabolism in  vitro and in  vivo. European Journal of Clinical Pharmacology 56 (8): 555–559. Wang, J.S., Wen, X., Backman, J.T. et  al. (1999). Midazolam alpha-­ hydroxylation by human liver microsomes in vitro: inhibition by calcium channel blockers, itraconazole and ketoconazole. Pharmacology and Toxicology 85 (4): 157–161. Wang, L.J., Ree, S.C., Chu, C.L. and Juang, Y.Y. (2011). Zolpidem dependence and withdrawal seizure  – report of two cases. Psychiatria Danubina 23 (1): 76–78. Waring, W.S. and Sandilands, E.A. (2007). Coma blisters. Clinical Toxicology (Philadelphia, Pa.) 45 (7): 808–809. Waters, B., Hara, K., Ikematsu, N. et  al. (2018). Tissue distribution of suvorexant in three forensic autopsy cases. Journal of Analytical Toxicology 42 (4): 276–283. WHO Management of Substance Abuse https://www.who.int/substance_ abuse/terminology/definition1/en/. WHO Collaborating Centre for Drug Statistics Methodology (2019). ATC/DDD Index. https://www.whocc.no/atc_ddd_index/?code=N05C. Wilkinson, C.J. (1995). The acute effects of zolpidem, administered alone and with alcohol, on cognitive and psychomotor function. Journal of Clinical Psychiatry 56 (7): 309–318. Wilson, K.C., Reardon, C., Theodore, A.C. and Farber, H.W. (2005). Propylene glycol toxicity: a severe iatrogenic illness in ICU patients receiving IV benzodiazepines: A case series and prospective, observational pilot study. Chest 128 (3): 1674–1681. Yell, R.P. (1990). Ethchlorvynol overdose. American Journal of Emergency Medicine 8 (3): 246–250. Zhao, Y., He, J., Yu, N. et al. (2020). Mechanisms of dexmedetomidine in neuropathic pain. Frontiers in Neuroscience 14: 330. Zink, T.M. (2020) A phantom of the past: Withdrawal from meprobamate presenting with focal seizures. Journal of Maine Medical Center 2(1, article 11): 1–4.

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using liquid chromatography-­tandem mass spectrometry. Journal of Chromatography. B: Analytical Technologies in the Biomedical and Life Sciences 878 (28): 2726–2732. Sugiyama, M., Fujita, K., Murayama, N. et al. (2011). Sorafenib and sunitinib, two anticancer drugs, inhibit CYP3A4-­mediated and activate CY3A5-­mediated midazolam 1’-­hydroxylation. Drug Metabolism and Disposition: The Biological Fate of Chemicals 39 (5): 757–762. Sweetman, S.C. (2009). Martindale: The Complete Drug Reference. London, Chicago: Pharmaceutical Press. Sychla, H., Grunder, G. and Lammertz, S.E. (2017). Comparison of clomethiazole and diazepam in the treatment of alcohol withdrawal syndrome in clinical practice. European Addiction Research 23 (4): 211–218. Tas, N., Altinbas, A., Noyan, T. et  al. (2019). Acute acetaminophene-­ induced hepatotoxicity and nephrotoxicity; therapeutic effect of dexmedetomidine. Bratislavske Lekarske Listy 120 (4): 270–276. Thornton, C.S. and Haws, J.T. (2020). Bromism in the Modern Day: Case Report and Canadian Review of Bromide Intoxication. Journal of General Internal Medicine 35 (8): 2459–2461. Tiemensma, M. and Davies, B. (2018). Investigating drug-­facilitated sexual assault at a dedicated forensic centre in Cape Town, South Africa. Forensic Science International 288: 115–122. Torres, R., Dressman, M.A., Kramer, W.G. and Baroldi, P. (2015). Absolute bioavailability of tasimelteon. American Journal of Therapeutics 22 (5): 355–360. Tozer, T.N., Witt, L.D., Gee, L. et al. (1974). Evaluation of hemodialysis for ethchlorvynol (Placidyl) overdose. American Journal of Hospital Pharmacy 31 (10): 986–989. Treweek, J.B., Roberts, A.J. and Janda, K.D. (2010). Superadditive effects of ethanol and flunitrazepam: Implications of using immunopharmacotherapy as a therapeutic. Molecular Pharmaceutics 7 (6): 2056–2068. Tyrer, P. (1993). ABC of sleep disorders. Withdrawal from hypnotic drugs. British Medical Journal 306 (6879): 706–708. Uchikawa, O., Fukatsu, K., Tokunoh, R. et al. (2002). Synthesis of a novel series of tricyclic indan derivatives as melatonin receptor agonists. Journal of Medicinal Chemistry 45 (19): 4222–4239. Uchimura, N., Nakajima, T., Hayash, K. et al. (2006) Effect of zolpidem on sleep architecture and its next-­morning residual effect in insomniac patients: A randomized crossover comparative study with brotizolam. Progress in Neuro-­ Psychopharmacology and Biological Psychiatry 30 (1): 22–29. Verstraete, A.G. and Buylaert, W.A. (1995). Survey of patients with acute poisoning seen in the Emergency Department of the University Hospital of Gent between 1983 and 1990. European Journal of Emergency Medicine 2 (4): 217–223. von Moltke, L.L., Weemhoff, J.L., Perloff, M.D. et al. (2002). Effect of zolpidem on human cytochrome P450 activity, and on transport mediated by P-­glycoprotein. Biopharmaceutics and Drug Disposition 23 (9): 361–367. Vossen, M., Sevestre, M., Niederalt, C. et al. (2007). Dynamically simulating the interaction of midazolam and the CYP3A4 inhibitor itraconazole using individual coupled whole-­body physiologically-­based pharmacokinetic (WB-­PBPK) models. Theoretical Biology & Medical Modelling 4: 13. Votaw, V.R., Geyer, R., Rieselbach, M.M. and McHugh, R.K. (2019). The epidemiology of benzodiazepine misuse: A systematic review. Drug and Alcohol Dependence 200: 95–114.

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52.5 Narcotics and Other Analgesics Dimitri Gerostamoulos and Olaf H. Drummer Narcotic drugs have traditionally derived from natural and synthetic morphine derivatives possessing morphine like actions. The most common of these include drugs such as morphine, heroin, codeine and are collectively known as opioids or narcotic analgesics. The global consumption of opioids more than tripled during the period 1991–2010 and ranks third in terms of community prevalence and use behind cannabis and stimulants. Legitimate opioids, such as morphine, codeine, oxycodone, hydrocodone and hydromorphone, are being consumed at an ever-­ increasing rate. According to the International Narcotic Control Board, approximately 50 countries have increased their consumption of opioid analgesics by more than 100% during the

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to relieve severe pain, while mild-­to-­moderate pain can be treated with codeine, dihydrocodeine and dextropropoxyphene. Some opioids, such as fentanyl and the fentanyl analogues alfentanil, remifentanil, and so on, can be used to induce or supplement anaesthesia. Opioids such as codeine, dihydrocodeine and, to a lesser extent, pholcodine can also be used as cough suppressants (antitussives). Synthetic opioids such as methadone, buprenorphine, naltrexone and naloxone can be used for the treatment of addiction to opioids. Combinations of pharmaceuticals, which include opioid and non-­opioid drugs, can be used to enhance analgesia (e.g. analgesic-­antipyretic preparations and use with some phenothiazines). There are substantial region-­to-­ region differences in the availability and usage of opioids. For example, the United States accounts for more than 99% of the global consumption of hydrocodone.

52.5.2  Sources of opioids

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Morphine and codeine are naturally occurring alkaloids that are extracted from the milky juice and stalks of the opium poppy, Papaver somniferum. The content of morphine in these plants can vary from as little as 5% to as high as 25%. Codeine is usually found in much smaller amounts (c. 0.2%) and the content of other naturally occurring opioids, such as thebaine (c. 0.1–0.3%), narcotine (c. 0.3%), narceine (c. 4–10%) and papaverine (c. 1%), can also vary depending upon the quality and origin of the plant (UNODC 2012). Many semisynthetic opioid derivatives are made by relatively simple modifications of the morphine or thebaine molecule, for example, heroin (diacetylmorphine) is a semisynthetic opioid produced from the acetylation of morphine. Other semisynthetic drugs produced from morphine or thebaine molecules include hydromorphone, oxycodone and naloxone (Figure 52.5.1).

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last decade (INCB 2012). The increase is particularly evident in Europe and North America. Other narcotics such as tilidine, diphenoxylate and fentanyl have all increased substantially over the last few years, while conversely dextropropoxyphene and pethidine have declined in use (Soumerai et al. 1987). The estimated annual prevalence of opioids in 2010 was 0.6– 0.8% of the population aged 15–64 years (between 26 million and 36  million opioid users worldwide), nearly half of whom used opioids, particularly heroin (UNODC 2012). In many countries, the majority of heavy drug users seeking treatment are primarily addicted to heroin; however, it is of increasing concern that legitimate opioids, such as oxycodone and methadone, are being diverted for illegal use. According to the 2010 (USA) National Survey on Drug Use and Health, there were 140,000 people aged 12 or older who had used heroin for the first time within the past 12 months (Substance Abuse and Mental Health Services 2011). This estimate was similar to the estimate in 2009 (180 000) and to estimates during 2002–2008 (ranging from 91 000 to 118 000 per year). Of concern was that the average age at first use among recent initiates aged 12–49 was 21.3 years, significantly lower than the 2009 estimate (25.5 years). Scientific reports published by the United Nations Drug Control Program (UNODC 2012) have showed that the world’s consumption of heroin has stabilised following increases in the 1980s and 1990s. Consumption has however increased significantly in Afghanistan and a number of countries involved in the transit of heroin. An indicator of the stabilisation of heroin consumption in Europe and other traditional markets such as Australia is the number of heroin-­related drug deaths as those deaths are predominantly linked to heroin use. Deaths due to heroin rose strongly in the 1980s and the 1990s and then declined slightly in the 2000s. Improvements in measures aimed at reducing the negative outcomes have also resulted in fewer deaths due to heroin. However, world events including the political changes in Afghanistan, the world’s largest producer of heroin, led to significant decline in the supply and subsequent demand for the drug. The use of heroin continues to cause widespread health and social problems in many countries especially in Europe and Asia where heroin injectors who regularly consume large amounts of different drugs face a risk of death, which may be 20–30 times higher than non-­drug users in the same age range. However, more recently reports from European countries such as Estonia and Finland suggest that fentanyl and buprenorphine may have displaced heroin use, while reports from Russia suggest that opimorphine) have oids such as desomorphine (dihydrodesoxy-­ been in high demand (UNODC 2012).

52.5.1 Use of narcotic drugs The primary use of legal opioids is for pain relief. This depends on the severity of pain as different opioids can be used to diminish or relieve pain. Opioids such as fentanyl, hydromorphone, methadone, morphine, oxycodone, tramadol, and pethidine can be used

52.5.3 Opioid pharmacology Opioids bind to opioid receptors throughout the central and peripheral nervous system to exert their diverse effects. Analgesia or the reduction in pain is the primary reason for prescribing opioids; they provide relief as the sensation of pain is reduced and the stress becomes less intense. In therapeutic doses, opioids relieve pain without causing general CNS depression as compared with general anaesthetics. In larger doses, opioids are more general depressants, and all are subject to misuse and addiction. Opioids also exhibit a range of unwanted side effects, which include nausea, vomiting and drowsiness. People prescribed opioids may experience euphoria, and as doses increase, side effects become more pronounced along with sedation, muscle rigidity and respiratory depression. As opioid doses increase, respiratory rates fall and progressively become shallower and irregular. This is due to the reduced sensitivity of the brain’s medullary centres to CO2. The respiratory depression and CO2 retention have important consequences; in high doses,

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breathing may fall to 3 or 4 breaths per minute instead of 12–16. Reduction in breathing can lead to hypoxia, coma and death. Heart rate can also be slowed by large doses of opioids. Opioids also depress the cough reflex by exerting a direct effect on the medulla. There seems to be no relationship between the analgesic, respiratory depressant action of opioids, and cough suppression; however, some opioids are more effective in depressing the cough reflex than others. Codeine and, to a greater extent, pholcodine are more effective than morphine in depressing the cough reflex. Another important effect of opioids is the varying effects on the gastrointestinal tract. Morphine causes a marked increase in tone and reduced motility, resulting in constipation. Constipation is a frequent side effect of repeated doses, and diarrhoea can effectively be treated with opioids. The smooth muscle of the biliary tract including the sphincter of Oddi is stimulated following administration of opioids. Other pharmacological effects of opioids include a decrease in pupillary size, often demonstrated in those using opioids.

Morphine

Codeine

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Thebaine

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6-Acetylmorphine

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52.5.4  Tolerance and dependence

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Tolerance is a condition where after repeated administration, a given dose of a drug produces a decreased effect and increasingly larger doses must be taken to obtain the pharmacological effects observed with the original dose. Tolerance to opioids develops much more quickly and to a much greater degree than tolerance to other drugs subject to abuse. Heroin addicts who have access to repeated and progressively larger doses are soon able to tolerate many multiples of the usual lethal doses. Tolerance is thus defined as a state where larger doses are required to elicit the same effect or when a fixed dose exerts a decreasing effect. The rate at which tolerance develops depends upon several factors, including especially the interval between doses and the amount of drug provided. The development of tolerance in heroin addicts is limited by the difficulty and expense involved in obtaining the drug, and many are able to continue more or less indefinitely on a fixed dose, continuing to experience the rush at the time of injection but showing little depression later. In the therapeutic situation, if constant and severe pain forces the administration of potent narcotics at 4–6-­hour intervals, the analgesic effect may decrease within a few days. If the interval between therapeutic doses can be lengthened well beyond the duration of action of the drug, i.e. given two times per day, tolerance may not be apparent after several weeks. In humans, for example, an initial dose of 100–200 mg of morphine would be sufficient to cause profound sedation, respiratory depression, anoxia and death; however, tolerant subjects can handle this and much more. Tolerance diminishes rapidly (few days) after withdrawal so that a previously tolerated dose may prove fatal if a period of abstinence of some days (or longer) occurs. Tolerance to other opioids can also occur depending on the potency of the drug.

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Diacetylmorphine (heroin)

N

Hydrocodone

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C

O

Oxycodone

Buprenorphine

Methadone

Figure 52.5.1  Chemical structures of some common opioids.

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due to incomplete absorption and metabolism of the drug as it passes through the liver and leads to a significant reduction in available drug (first pass metabolism) at approximately 25–35% of the same dose if given intravenously. Other opioids have higher bioavailability when given orally (Table 51.9). Some opioids are also available as sustained release preparations (i.e. slow or controlled release such as some oxycodone and morphine preparations). This means that a patient suffering from chronic pain may only have to take a tablet once or twice per day, which results in stable blood concentrations as a consequence of the drug’s more predictable pharmacokinetics. Methadone and oxycodone rectal suppositories are useful alternatives to oral sustained release morphine preparations. Opioids undergo extensive metabolism in humans. There are three main biotransformation pathways that have been established for the metabolism of opioids: hydrolysis, glucuronidation and oxidation. For example, following intravenous injection, heroin is rapidly converted (within seconds) to 6-­acetylmorphine ­(6-­AM), which is subsequently hydrolysed (within minutes) to morphine. The conversion of heroin to 6-­AM occurs as both a result of enzymes and spontaneous hydrolysis. The majority of opioid metabolism in humans occurs in the liver through a process called glucuronidation; this occurs mainly in the liver and to a lesser extent in the intestine and kidneys (Regnard and Twycross 1984). Morphine is conjugated primarily to morphine-­ 3-­ glucuronide (M3G) and to a lesser extent the biologically active morphine-­6-­glucuronide (M6G). There are also opioids considered to be prodrugs; these are drugs that are administered in a biologically inactive form and are biotransformed into an active metabolite. Hydrocodone and tramadol are prodrugs that are converted to active forms by CYP450 isoenzymes (Overholser and Foster 2011). Most opioids are excreted in the urine usually within the first 24 hours following administration (Table 52.5.1). The concentrations of conjugated metabolites usually exceed those of the parent drug and can be present for longer periods of time

TO

Parallel to the development of tolerance and independent of any psychoactive dependence, people addicted to opioids develop a need for the continued administration of opioids to prevent what is commonly known as withdrawal syndrome. Abrupt discontinuation of opioids in people physically dependent on opioids precipitates a withdrawal syndrome, the severity of which depends on the individual, the drug used, the size and frequency of the dose and the duration of drug use. Opioid analgesics with some antagonist activity, such as buprenorphine, butorphanol or pentazocine, may also precipitate withdrawal symptoms in patients who are dependent on opioid narcotics. The onset and duration of withdrawal symptoms also vary according to the duration of action of the specific drug. Withdrawal symptoms may be terminated by a suitable dose of morphine or another opioid. Methadone and buprenorphine are currently the most widely used pharmacotherapeutic agents for maintenance treatment of heroin addicts (Christrup  1997). Methadone and buprenorphine are effective in the suppression of withdrawal symptoms and in the reduction or elimination of an addict’s compulsion to take heroin. The major objective of an opioid maintenance programme is to achieve long-­lasting stabilisation of the user’s drug dependence by providing the drug indefinitely in doses large enough to produce a level of cross-­ tolerance that is sufficient to diminish the effects of ordinary doses of heroin.

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52.5.5 Absorption, metabolism and excretion

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The extent of absorption depends on the type of opioid and the route of administration. All opioids are well absorbed when given intravenously; however, the bioavailability of opioids when given orally varies between different compounds. For example, morphine has a greatly delayed onset of action following oral administration and must be several times the intravenous dose. This is

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Table 52.5.1  Elimination half-­life, volume of distribution, typical dose, duration of action and action as a drug receptor for selected opioids.

Vd (L/kg)

Dose (mg)

Duration of action (h)

Receptor action

2–3

3.2

5–20

4–5

Agonist

2–4

2.6–5

30–60

2–4

Agonist

Heroin

0.03–0.05

-­

5–10

-­

Agonist

Hydromorphone

1.5–4

3

2–4

1.5–4

Agonist

Oxycodone

3–6

1.8–3.7

10

4–5

Agonist

Buprenorphine

2–4

1.4–6.2

0.05–0.2

0.3–0.6

Partial agonist

Fentanyl

3–12

3–8

0.05–0.2

0.5–1

Agonist

Methadone

15–55

4–7

5–100

6–8

Agonist

T1/2 (h)

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Opioid

Naturally occurring Codeine

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Morphine

Semisynthetic

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T1/2, half-­life; Vd, volume of distribution.

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Opioids exhibit a remarkable absence of direct toxic effects. Much of the toxicity to opioids is typically combined with the use of other drugs including alcohol and/or benzodiazepines. Other factors that contribute to the overall poor health of opioid addicted people include crime, unsterile drug administration leading to disease such as the suppression of immune systems, family disruption, undernourishment, vomiting, diarrhoea leading to marked weight loss, unsafe sex practices, dehydration, ketosis and disturbance in acid–base balance. The manifestation of infectious diseases in intravenous drug users plays a critical role in life-­threatening complications for addicts. Needle sharing, unsafe sexual practices, poor hygiene, devastation of skin barrier and the injection of unfamiliar substances into the body all increase the risk of human immunodeficiency virus (HIV) transmission, viral hepatitis, pneumonia and tuberculosis. Infectious complications as a result of all these factors include endocarditis and septicaemia, viral hepatitis, liver cirrhosis, meningitis and tetanus (Hanks et  al. 1988). Many opioid-­dependent women who are pregnant also carry additional risks that may include abortion, low birth weight infants with intrauterine growth retardation and high rates of preterm birth. Infants of addicted women are often preterm and suffer from significant jaundice and neonatal opioid withdrawal. Signs of withdrawal in infants include irritability, rhythmic tremors, shrill and high pitch crying, and poor feeding. Much more important is the growing number of people addicted to prescription opioids. The worldwide epidemic of oxycodone-­related deaths accounts for a growing number of fatalities every year (Okie 2010). From 1999 to 2007, the number of US poisoning deaths involving any opioid analgesic (e.g. oxycodone, methadone or hydrocodone) more than tripled, from 4041 to 14 459, or 36% of the 40 059 total poisoning deaths in 2007 (Warner and Makuc 2009). Toxic effects of opioids can be exacerbated with the combined use of other drugs. Drugs such as alcohol and benzodiazepines enhance the depressant effects of opioids on the CNS (Moffat 2004). Opioid users frequently take benzodiazepines (e.g. diazepam, nitrazepam) to reduce anxiety and to minimise the unpleasantness of any withdrawal symptoms. Drug combinations are therefore potentially serious complications in opioid users, particularly for heroin users. Data from a review of deaths in Victoria,

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52.5.6  Toxic effects of opioids

Australia (Gerostamoulos et al. 2001), show that benzodiazepines were the most prevalent drug group (c. 55%) and alcohol was present in c. 30% of all heroin-­related deaths. Drugs such as the sedating antihistamines can prolong morphine metabolism, leading to increased respiratory depression. One of the problems confounding the forensic toxicologist and pathologist is the relevance and interpretation of drug concentrations in different specimens following the use of opioids. The mechanism of death in opioid users is often uncertain and is often due to many factors. For example, the use of heroin results in a wide range of adverse effects due to a variety of pharmacological and physiological responses to heroin, hypersensitivity reactions to the cutting agents or contaminants, and diseases associated with intravenous use. The largest numbers of deaths have been attributed to an acute reaction, whereby death occurs shortly after injection (Spitz et  al. 2006). Three overdose syndromes are recognised: death from profound respiratory depression, death from arrhythmia and cardiac arrest and death as a consequence of severe pulmonary oedema (Robbins et al. 2010). Death may also occur indirectly as a complication of unconsciousness. This is caused by a non-­fatal dose, leading to airways’ obstruction in a setting of diminished respiratory function. Respiratory disease can also reduce the ability of person to tolerate a dose of opioid. Fentanyl, methadone, oxycodone, ­ hydrocodone and morphine are the most common legal opioids known to cause death if misused.

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(Christrup  1997). As with most pharmacokinetic parameters, there is great individual variation in the elimination half-­life. This value is greatly prolonged in renal failure resulting in drug accumulation. Some opioids with long half-­lives such as methadone can accumulate in tissues. Such accumulation can be toxic or even fatal if doses are not appropriately administered and the drug not given enough time to be cleared by the body. This can lead to prolonged respiratory depression and pronounced enterohepatic recirculation (excretion into the gastrointestinal system through bile followed by reabsorption) (Hanks et al. 1988).

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52.5.7  Postmortem toxicology: Interpretation of opioid concentrations Understanding postmortem toxicology results is difficult and requires knowledge of postmortem processes that can affect interpretation (see also Chapter 49). Consideration must be given to the regularity of opioid use (i.e. acute vs chronic). Concentrations of drug may be typically higher in those who are regular users and hence may not be toxic compared to naive users. The route of administration is also important as oral ingestion of opioids means that the liver removes some of the efficacy of the dose before the drug enters the blood stream (first pass metabolism). Other factors include the presence of natural disease, which can impact metabolism, and excretion of opioids and metabolites as well as contribute to compromised respiratory function caused by the opioid. The accumulation of opioids may result in adverse effects and even death. Concurrent natural disease may predispose individuals to cardiovascular collapse and even unpredictable responses such as convulsions (Goodman et al. 2001). Drugs that interfere with the pharmacokinetics of opioids generally do so by altering their elimination and hence the presence of other drugs, the possibility of antagonism for one drug by another and more often the additive or synergistic effects produced by the interaction of two or more depressant drugs must all be considered (Cone et  al. 2004). The consideration of drug

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−

Toxic Deleterious Mn+ Possible death

Figure 52.5.2  Dependence of the biological response (i.e. effect) on the concentration of an essential trace element (solid line) and of a non-­essential element (broken line). Reproduced with permission from Seiler et al. (1994), © Springer; for the original figure, see Drasch (2003, p. 199).

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i­ nteractions involving opioid prodrugs is important because they can be clinically manifested in the opposite manner from an active parent drug. For example, the decreased metabolism of a prodrug would result in a decreased analgesic effect and potential treatment failure, whereas the decreased metabolism of an active parent drug would enhance an analgesic effect and potentially lead to opioid toxicity (Overholser and Foster 2011). Tissue distribution and redistribution of opioids and its metabolites are also important in assessing postmortem contributions of opioids to adverse effects and death, as blood concentrations increase after death for the more lipid soluble opioids, even when blood is taken from a peripheral source. As tolerance develops with opioids, there are enormous differences in the doses administered between those receiving acute postoperative pain relief and those in palliative care. Studies reporting the findings in living and deceased heroin addicts illustrate the extent of overlap between concentrations measured in opioid-­related and opioid-­caused deaths (Kennedy 2010) (Figure 52.5.2).

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Deficient

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Marginal

Hanks, G.W., Hoskin, P.J., Aherne, G.W., Chapman, D., Turner, P., and Poulain, P. (1988). Enterohepatic circulation of morphine. Lancet 1 (8583): 469. INCB (2012). Report of the International Narcotics Control Board for 2011. United Nations: New York. Kennedy, M.C. (2010). Post-­mortem drug concentrations. Intern Med J 40 (3): 183–7. Moffat, A.C. (2004). Clarke’s Analysis of Drugs and Poisons, 3rd edition. Pharmaceutical Press. National Survey on Drug Use and Health (2011). Results from the 2010 National Survey on Drug Use and Health: Summary of National Findings. Substance Abuse and Mental Health Services Administration: Rockville, MD. Okie, S. (2010). A flood of opioids, a rising tide of deaths. N Engl J Med 363 (21): 1981–5. Overholser, B.R. and Foster, D.R. (2011). Opioid pharmacokinetic drug-­ drug interactions. Am J Manag Care 17 (11): S276–87. Regnard, C.F. and Twycross, R.G. (1984). Metabolism of narcotics. Br Med J (Clin Res Ed) 288 (6420): 860. doi: https://doi.org/10.1136/ bmj.288.6420.860 Robbins, S.L., Kumar, V., and Cotran, R.S. (2010). Robbins and Cotran Pathologic Basis of Disease, 8th edition. Philadelphia, PA: Saunders/ Elsevier. xiv, 1450 p. https://doi.org/10.1002/path.1711600125. Soumerai, S.B., Avorn, J., Gortmaker, S., and Hawley, S. (1987). Effect of government and commercial warnings on reducing prescription misuse: The case of propoxyphene. Am J Public Health 77 (12): 1518–23. Spitz, W.U., Spitz, D.J., and Fisher, R.S. (2006). Spitz and Fisher’s Medicolegal Investigation of Death: Guidelines for the Application of Pathology to Crime Investigation, 4th edition. Springfield, Ill.: Charles C. Thomas. 1325 p.UNODC (2012). World Drug Report. United Nations: Vienna, Austria. Warner C.L. and Makuc, D.M. (2009). Increase in Fatal Poisonings Involving Opioid Analgesics in the United States, 1999-­2006., n. NCHS data brief, Editor, National Center for Health Statistics.: Hyattsville, MD.

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Optimal

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Biological response

Essential Mn+

FO

References

Christrup, L.L. (1997). Morphine metabolites. Acta Anaesthesiol Scand 41 (1 Pt 2): 116–22. Cone, E.J., Fant, R.V., Rohay, J.M., Caplan, Y.H., Ballina, M., Reder, R.F., and Haddox, J.D. (2004). Oxycodone involvement in drug abuse deaths. II. Evidence for toxic multiple drug-­drug interactions. J Anal Toxicol 28 (4): 217–25. Gerostamoulos, J., Staikos, V., and Drummer, O.H. (2001). Heroin-­ related deaths in Victoria: a review of cases for 1997 and 1998. Drug Alcohol Depend 61 (2): 123–7. Goodman, L.S., Hardman, J.G., Limbird, L.E., and Gilman, A.G. (2001). Goodman & Gilman’s the Pharmacological Basis of Therapeutics, 10th edition. New York: McGraw-­Hill. xxvii, 2148 p.

52.6  Toxic Elements Gustav Drasch Chemical elements and their ions can interact with a living organism in different ways: as electrolytes (like sodium or calcium) and essential trace elements (like cobalt or nickel) or in a toxic way. In principle, all elements are toxic: it is only a question of the dose. Nevertheless, the term ‘toxic elements’, often used synonymously with ‘toxic heavy metals’, is frequently used. Which elements are described in these terms depends mainly on the individual toxicity of an element and how often toxic concentrations of this element occur in the environment (in addition, in forensic toxicology, how often the element is used for suicide or homicide). To complicate the situation, some essential trace elements like selenium may also be toxic in higher concentrations. Figure 52.6.1 shows these correlations schematically. Typically, lead (Pb), cadmium (Cd) and mercury (Hg) are indicated as toxic heavy metals. Arsenic (As) is often included, even though it is a

ESSENTIAL

DEFICIENT CONCENTRATION OF METAL ION –

TOXIC DELETERIOUS Mn+ POSSIBLE DEATH

Figure 52.6.1  Dependence of the biological response (i.e. effect) on the concentration of an essential trace element (solid line) and of a non-­essential element (broken line), schematically illustrated (according to Seiler et al. 1994).

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transition element. The concentrations of essential trace elements are regulated in the body within narrow limits by homoeostasis. In contrast, non-­essential elements are not subject to such regulation. Therefore, their concentrations in tissues and body fluids differ much more intraindividually and are often caused by the body mistaking toxic with essential elements (e.g. Pb2+ with Ca2+, Tl+ with K+ or AsO43− with PO43−). With recent progress in trace analysis (speciation), the special importance of the binding form of an element (species) for its toxicity has been recently recognised. In the last few decades, the toxicological importance of elements has changed from classic acute and occupational intoxications to chronic exposure. Nevertheless, serious acute intoxications, such as accidental, iatrogenic, suicidal or homicidal, still occur worldwide. Moreover, the question of third-­party foul play rises much more often in the case of a severe intoxication with a toxic heavy metal than in the case of intoxication with pharmaceutical or addictive drugs. Therefore, it is still indispensable for the forensic toxicologist to be able to recognise such intoxications. A special problem in forensic toxicology is the interpretation of human tissue concentrations of toxic heavy metals. Recently, numerous data on background concentrations of elements have been published by environmental medicine specialists, but almost exclusively in so-­called biomonitors like blood, serum, urine or hair (i.e. analytes which can easily be obtained from a living individual). In contrast, reference values for human tissues are rare to find in recent publications. The use of tissue concentrations from older publications is critical for the following reasons. First, for many toxic metals, the background pollution has changed in the last decades. In contrast to received opinion, most of them have not increased but decreased. A further large problem is the accuracy of older reference values. For a valid comparison, it should be aimed to take only reference values measured under strict quality control with matrix-­matched reference samples. Moreover, in older publications, tissue concentrations are often referred to as dry-­weight or ash-­weight, while more recently wet-­weight concentrations have been preferred. A conversion is critical due to individual variations like different water contents.

LY

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BIOLOGICAL RESPONSE

EXCESS MARGINAL

Several threshold values have been proposed for toxic element concentrations not only in water, soil, air and foodstuff, but also in human biomonitors. The strict consideration of the definition of such a limit and the application of its last revised version is crucial for the correct interpretation of contaminant concentrations. In Germany, for toxic element concentrations in biomonitors, different approaches exist: reference values, human biomonitoring (HBM) values and biological threshold values (the Biologische Grenzwerte or BGW in German). German ‘reference values’ were derived from the upper 95% confidence interval of the 95 percentile of large cohort studies with some 1000 volunteers. These reference values are not toxicologically defined but give the upper limits of the actual background pollution. In contrast, HBM values are toxicologically defined. The HBM-­I value is defined as a ‘check value’, while HBM-­II is an ‘intervention value’. This means that with an excess of HBM-­I, no adverse effect is to be expected, but the source of the increased burden should be detected and, as far as possible, eliminated. At values above HBM-­II, adverse health effects cannot be excluded. A medical investigation for such effects and, if necessary, a medical treatment should be performed (HBM Commission 2007a, 2007b; Schulz et  al. 2011). Similarly defined, but internationally more common are the terms no observed adverse effect level (NOAEL) and lowest observed adverse effect level (LOAEL). Whereas the LOAEL can be calculated statistically from epidemiological studies on burdened populations, the NOAEL must be extrapolated from the LOAEL with a safety factor. From these limits, valid for the total population, limits for an occupational pollution (BGW limits) must be strictly separated. BGW limits are only valid for healthy adult workers under occupational medical supervision. Besides the well-­known Biologischer Arbeitsstoff-­Toleranzwert (BAT) values, for carcinogenic agents, the so-­called Biologischer Leitwert (BLW) values are given; furthermore, a Biologischer Arbeitsstoff Referenzwert (BAR) value is given, which is not toxicologically defined (DFG 2012). Other countries have and use other limits and other definitions, for example, the Reference Dose (RfD) given by the Environmental Protection Agency of the United States represents toxicologically defined limits. The interpretation of metal concentrations determined in autolytically changed tissues raises special problems in forensic toxicology. Postmortem element concentrations in tissues do not decrease by putrefaction similar to organic compounds like drugs. But for some relevant toxic metals, the living organism sustains an extreme gradient between different compartments. For example, the cadmium concentration in the kidney cortex of a merely background polluted person can exceed 100  mg/kg, while the cadmium blood concentration of the same person will just reach 1 μg/L. This is a gradient of five orders of magnitude, which causes a postmortem redistribution and therefore an increase in cadmium in postmortem blood up to values that can be misinterpreted as an acute intoxication (Götz et  al. 1993). Similar problems arise with histologically fixed materials. Moreover, some fixation fluids still contain mercuric salts for

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conservation, what may result in totally unrealistic high mercury concentrations in tissues, embedded in such fluids. Caused by spectacular cases in the past, a recurring question is the exclusion of arsenic intoxication in an exhumed corpse or even urn ash. For this, some details are given in Drasch (2003). Due to their specific relevance in forensic toxicology, the following is limited mainly to acute intoxications with lead, cadmium, mercury, thallium (T1) and arsenic.

Toxicity

52.6.1 Lead

Even for well soluble inorganic lead compounds like lead acetate, lethal doses for an adult are relatively high, approximately 20–50 g. Typical symptoms of an acute intoxication with inorganic lead compounds are heavy stomach colics (lead colic) with nausea and vomiting, constipation and diarrhoea. More characteristic is a sweet metallic taste in the mouth. Damage to liver and kidneys and hypertension may occur. Adverse effects on the CNS follow; at the end, lead encephalopathy develops. In the case of lead intoxicated children, often the CNS symptoms dominate. The target tissue for intoxication with organolead compounds is also the brain with hallucinations and hyperreflexia. Should the ­victim survive the acute phase, in many cases a chronic intoxication develops.

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In most industrial countries, background pollution with lead has been decreasing for decades due to the phasing out of lead in many products like wall paint or water pipes (both prohibited in Germany for decades, but still a problem in old houses in other countries like the United States), tin-­lead solders or car petrol. Regionally, there are some hotspots of lead in drinking water, especially from domestic wells with strong acidic water. Normally, approximately 80% of total lead intake originates from food, especially vegetables. Most lead in (or better: on) vegetables is superficial and can be removed almost completely by peeling or washing. For toddlers, the oral intake of lead-­contaminated soil via hand-­to-­mouth activities may contribute more to their total lead contamination than food. A special problem in forensic toxicology was the recent occurrence of severe lead-­adulterated marijuana in Germany (Busse et al. 2008). Further unexpected—­and therefore often misdiagnosed—­ lead intoxications have been reported with traditional Chinese medicine (TCM) or Ayurveda drugs, lead-­glazed and too-­low fired ceramics, especially from markets in the developing world, and lead shots or bullets that were not removed from the body (Seiler et al. 1994; Drasch 2003; Meißner et al. 2011).

Acute intoxication

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Exposure

As a typical heavy metal, lead interacts with sulphhydryl groups of many enzymes by blocking them. Moreover, lead affects calcium-­ dependent physiological functions. In bone, lead is incorporated in the form of hardly soluble lead phosphate (which does not harm the bone).

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Approximately 10% of soluble lead compounds are absorbed in the adult gastrointestinal tract, but up to 50% in the infant. In the lungs, lead containing particles brain) usually remains constant. In the case of an intoxication with MeHg, mercury in the brain may be as high as in the liver or kidneys (Drasch 2003; Baselt 2011; Arndt 2012b). Total mercury in hair correlates relatively well with the total mercury in the brain. In Germany, median values of c. 250 ng/g in hair were found. Normally, total mercury in hair is below 1000 ng/g. Toxic effects from MeHg in children were reported if total mercury in hair exceeded 5000 ng/g (HBM Commission 2009b).

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Reference concentrations

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52.6.4 Thallium

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Thallium is a rare element and of no ecotoxicological relevance. Previously, higher exposures have been reported in the vicinity of cement plants and lead, zinc and cadmium smelters. Thallium-­I-­ sulphate was the active ingredient of the common rodenticide Zelio, which has not been available for some decades. Nevertheless, some old packages may still be found stored and may therefore cause further acute intoxications, if ingested orally.

Reference values for thallium in urine of 0.5 μg/L (adults) and 0.6 μg/L (children) have been published by the German Environmental Agency. An HBM-­I of 5 μg/L is proposed (HBM Commission 2011b). In the case of severe thallium intoxications, peak concentrations of 25 mg Tl/L of urine were found, declining to 1–10 mg/L at the onset of alopecia (Drasch 2003; Baselt 2011; Klemm and Meißner 2012). In blood, similar background concentrations (1  mg/L) can be found. However, thallium levels in blood declines much quicker than those in urine. Typical background concentrations of thallium in tissues are in a range of 1  ng/g in kidneys and in bones and teeth up to 10 ng/g (Drasch 2003). In the case of a lethal thallium intoxication, tissue concentrations are in the range of 10–100 μg/g; after a longer time of survival, 1–10 μg/g will be found (Baselt 2011). In hair, normal thallium concentrations are in the range of 5–15 ng/g. Acute thallium intoxication may increase the thallium concentration in hair to more than 1 μg/g. The interpretation of thallium concentrations in hair, which was lost due to the intoxication, can be problematic. The supply of this in hair may already be altered days before. If any storage of thallium can be expected at all, it is in the roots (Drasch 2003).

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Soluble thallium compounds are quickly and almost completely absorbed in the gastrointestinal tract. The toxicokinetics of thallium are triphasic: In the first 4 hours, a quick distribution from the blood into a central distribution compartment (liver, muscles, kidneys), followed by a second phase with redistribution to peripheral compartments like the brain (4–24 hours after application). In a third phase, a further redistribution to the bones, teeth and skin occurs. Thallium is eliminated renally with a half-­ life of some days. Biliary excreted thallium is partially subject to an enterohepatic circulation (Seiler et  al. 1994; Drasch 2003; Klemm and Meißner 2012).

Toxicity

52.6.5 Arsenic

Acute intoxication

Exposure

Even after the intake of a lethal dose of thallium-­I-­sulphate (0.5–3 g), the first 1 or 2 days are usually free of symptoms. Not until 3–5 days after intake will the following symptoms develop slowly, reaching their maximum not before the second or third week: abdominal and especially retrosternal pain and polyneuropathy, ascending to the extremities. Often, a psycho-­organic syndrome (POS) develops. Other massive toxic damage of the central and peripheral nervous system, the optic nerves, the cardiac muscle, the lungs, the liver and the kidneys may follow. Some days

Arsenic is a typical accompanying element in many ores like copper, gold, zinc or lead. Increased arsenic concentrations may be found in special geogenic hotspots in water and soil, but also in the vicinity of smelters and waste dumps. Al-­Ga-­arse-­nide is used in the semiconductor industry. Most former applications of arsenic as in paints, pesticides, rodenticides or pharmaceuticals have been phased out due to its toxic and carcinogenic potentials. Fish is the highest arsenic burdened food. But this organic ‘fish-­arsenic’ (arsenic-­betaine, -­choline and -­lecithin) is much less toxic than

LY

N

Chronic intoxication

Several cases from the past have shown that chronic arsenic intoxication is even more often misdiagnosed than its acute form. The symptoms may be similar to an acute intoxication, but their onset is quite gradual and their sequence may be changed. A chronic arsenic intoxication may last for months or even years until death occurs by common cachexia (Drasch 2003).

TO

Soluble arsenic compounds like As2O3 or KAsO2 will almost be quantitatively absorbed in the gastrointestinal tract. The absorption rate after inhalation ranges widely according to the particle diameters. Absorbed As2+ is partially metabolised to the more toxic and carcinogenic As3+. This is slowly detoxified in the human body by methylation to monomethylarsonic acid (MMAA) and dimethylarsinic acid (DMAA). Approximately 35–75% of absorbed arsenic is eliminated predominantly renally within the first 1–2  days. In a second phase with a half-­life of 8–10 days, increasingly MMAA and DMAA are eliminated, the rest very slowly renally or incorporated in the skin, hair and nails. For organic arsenic compounds from fish, a biphasic elimination is reported: a first phase with a half-­life of 7–11 hours and a terminal one with approximately 75 hours duration. Arsenic crosses the placenta (Drasch 2003; Baselt 2011).

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Toxicokinetics

affected too. At 2–4 weeks after ingestion, a relatively characteristic massive loss of hair begins (very similar to a thallium intoxication). One month after ingestion, pale transverse stripes usually appear on the nails, again similar to thallium intoxication. Later, changes of the skin follow such as hyperkeratosis, melanosis and basal cell carcinoma (Drasch 2003). Approximately 10 ppm of arsine (AsH3) inhaled for 1 minute is markedly toxic; 250 ppm for 1 hour is lethal. Typical of arsine intoxication is a latency period of up to 24 hours. Predominant is a massive intravascular haemolysis. Death occurs within a few hours after the onset of symptoms by internal asphyxiation, caused by a lack of erythrocytes. 52.6.5.2 Arsenic halides like the chemical warfare agent lewisite are even more toxic than AsH3 and react strongly corrosively on skin and mucous membranes. A toxic oedema of the lung develops rapidly. Inhaled in high doses, death occurs within minutes by suffocation caused by mucus in the bronchi (Croddy and Wirtz 2005).

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inorganic arsenic species. Until the First World War, arsenic was undoubtedly the classic homicide poison because its symptoms were easily mistaken for food poisoning. Nevertheless, acute homicidal and suicidal intoxications with arsenic compounds have been described in the recent past, almost all with arsenic oxide (As2O3) or potassium arsenite (KAsO2). Of special interest in forensic toxicology are arsenic containing chemical warfare agents (alkyl-­arsines like lewisite) (Croddy and Wirtz 2005).

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The toxicity of arsenic compounds depends strongly on their binding forms. Inorganic arsenic compounds can be put in the following ascending order of toxicity: metallic arsenic > As5+ compounds > slightly soluble As3+ compounds > readily soluble As3+ compounds > arsine (AsH3). Some organoarsenic compounds like ‘fish-­arsenic’ have relatively lower levels of toxicity, while alkyl-­arsines like 2-­chlorovinyl-­dichloroarsine (lewisite) are extremely toxic and, therefore, used as chemical weapons.

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The acute lethal oral dose of As2O3 or soluble arsenites is in the order of 60–300 mg for an adult. Normally, after oral ingestion, the symptoms start rapidly with vomiting increasing until the onset of systemic effects. In some cases, a latency of several hours between first vomiting and systemic effects has been reported. These are abdominal pain and colic, mostly massive, and bloody diarrhoea, thirst, an inflammation of the throat, cyanosis, convulsions of the extremities, renal failure and blood circulation failure. As a rule, death occurs within 4–24 hours after ingestion. In some cases, the gastrointestinal symptoms have only been transient and the nervous system symptoms like confusion, cramps, somnolence and coma have begun immediately. If the acute intoxication is survived, long and painful aftereffects of the peripheral nervous system like polyneuropathies, motoric paralysis and cramps in the extremities follow. Sometimes the CNS is

A reference value for arsenic in urine of 15 μg/L was given for people without fish consumption within the last 2 days prior to sample collection (Wilhelm et  al. 2004). In some cases, highly organoarsenic burdened seafood may increase arsenic in urine for a short term up to 200 μg/L. For a differentiation between such a harmless increase in arsenic in urine by organoarsenic from seafood and the suspicion of an inorganic arsenic intoxication, a second urine sample, taken 3 or more days later after seafood abstinence, should be analysed. In the first days after a severe intoxication, arsenic concentrations in urine are often in the mg/L range, independent from the ingested arsenic species. Usually, it remains increased for at least 14  days (Drasch 2003; Baselt 2011). The determination of arsenic in the blood is less suitable to assess an arsenic intoxication than arsenic in urine. Arsenic is distributed quite equally between plasma and the corpuscular parts of the blood. In most cases, arsenic levels in the blood are somewhat lower than in urine. After an acute ingestion of toxic quantities of arsenic compounds, the arsenic level in the blood may be in the range of 100–1000 μg/L, declining rapidly within a few days (Drasch 2003; Baselt 2011). In hair, background levels of 10–100 ng As/g are usually found. Normal upper limits are up to 1 μg/g. In the case of a suspected intoxication, a sectioned investigation of the hair should be performed, to obtain information not only on the date and duration

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Busse, F., Omidi, L., Leichtle, A. et  al. (2008). Lead poisoning due to adulterated Marijuana. New England Journal of Medicine 358: 1641–1642. Croddy, E. and Wirtz, J.J. (2005). Weapons of Mass Destruction, Vol. I: Chemical and Biological Weapons. Santa Barbara, California: ABC-­ CLIO Inc. DFG (ed.) (2012). MAK-­ und BAT-­Werte-­Liste 2012. Weinheim, Germany: Wiley-­VCH-­Verlag. Drasch, G., Wanghofer, E. and Roider, G. (1997). Are blood, urine, hair and muscle valid biomonitors for the internal burden of men with the heavy metals mercury, lead and cadmium? Trace Elem. Electrolytes 14: 116–123. Drasch, G. (2003). Metalle und Verbindungen. In: Madea, B. & Brinkmann, B. (eds) Handbuch Gerichtliche Medizin Band 2, pp. 198– 238.) Berlin: Springer. Götz, D., Baudisch, H. and Köppel, C. (1993). Post mortem changes in cadmium blood concentration in patients with cardiovascular disease. Proceedings of the 8th International Symposium on Trace Elements in Man and Animals, Dresden. HBM (Human Biomonitoring) Commission of UBA (2002). Addendum zur “Stoffmonographie Blei – Referenz-­und Human-­Biomonitoring-­ Werte”. Bundesgesundheitsbl 45: 752–753. HBM (Human Biomonitoring) Commission of UBA (2007a). Ableitung von Human-­Biomonitoring-­(HBM-­)Werten auf der Basis tolerabler Aufnahmemengen  – Teil I: Einführung. Bundesgesundheitsbl 50: 249–250. HBM (Human Biomonitoring) Commission of UBA (2007b). Ableitung von Human-­Biomonitoring-­(HBM-­)Werten auf der Basis tolerabler Aufnahmemengen  – Teil II: Grundlagen und Ableitungsweg. Bundesgesundheitsbl 50: 251–254. HBM (Human Biomonitoring) Commission of UBA (2009a). 2. Addendum zur Stoffmonographie Blei  – Referenz-­und Human-­ Biomonitoring-­Werte. Bundesgesundheitsbl 52: 983–986. HBM (Human Biomonitoring) Commission of UBA (2009b). Addendum zur Stoffmonographie Quecksilber  – Referenz-­ und Human-­ Biomonitoring-­Werte. Bundesgesundheitsbl 52: 1228–1234. HBM (Human Biomonitoring) Commission of UBA (2009c). Neue und aktualisierte Referenzwerte für Antimon, Arsen und Metalle (Blei, Cadmium, Nickel, Quecksilber, Thallium und Uran) im Urin und im Blut von Kindern in Deutschland. Bundesgesundheitsbl 59: 77–982. HBM (Human Biomonitoring) Commission of UBA (2011a). Aktualisierung der Stoffmonographie Cadmium – Referenz-­und Human-­Biomonitoring (HBM)-­Werte. Bundesgesundheitsbl 54: 981–996. HBM (Human Biomonitoring) Commission of UBA (2011b). Stoffmonographie Thallium − Referenz-­und Human-­Biomonitoring-­ (HBM)-­Werte für Thallium im Urin. Bundesgesundheitsbl 54: 516–524. Kijewski, H (1993). Die forensische Bedeutung der Mineralgehalte in menschlichen Kopfhaaren. Lübeck: Schmidt-­Römhild. Klemm, M. and Meißner, D. (2012). Problematik, Klinik und Beispiele der Spurenelementvergiftung  – Thallium. Toxichem Krimtech 79: 19–22. Meißner, D., Klemm, M. and Zogbaum, M. (2011). Problematik, Klinik und Beispiele der Spurenelementvergiftung – Blei. Toxichem Krimtech 78: 453–464.

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of the burden but also on the individual background level of unburdened sections. For the reliable diagnosis of an arsenic intoxication, arsenic in hair must markedly exceed 1 μg/g. Principally, nails store arsenic as good as hair. But in the case of a suspected survived arsenic intoxication, the investigation of nails is difficult because from a living person only the foremost part of the nail can be cut. But arsenic is like other substances incorporated in the nailbeds and grows slowly forward with the nails (c. 5mm/month). Background levels of arsenic in internal tissues are normally low, approximately 10–100 μg/kg wet-­weight. In any case of lethal arsenic intoxication, the arsenic concentrations in the internal tissues are in the mg/kg range. Besides the stomach, after an acute intoxication, the highest concentrations can be found in the liver, spleen and kidneys. Brain concentrations are usually somewhat lower (Drasch 2003; Baselt 2011). As arsenic is radiopaque, a conspicuous radiograph of the stomach region, taken early in the hospital, is often of crucial diagnostic importance to detect an unknown arsenic intoxication. The interpretation of arsenic concentrations in exhumed corpses raises special problems. In principle, the following shifts may occur: leaching of arsenic out of the corpse into the environment (e.g. into the pall under the corpse); or an immigration of arsenic from outside into the corpse. The latter is especially suspect if the coffin has already collapsed and is filled with soil. Even unburdened soil can contain up to 20 mg As/kg dry matter (i.e. a concentration 2–3 orders of magnitude higher than in human tissues). The determination of the arsenic concentrations in different soil samples, collected according to the scheme of Specht and Katte (1954) from around and in the coffin is irremissible in such situations. A further common possibility for arsenic contamination is grave goods like crosses made from metal alloys, etc. Mould and keratin of hair and nails may enrich arsenic from groundwater. Arsenic intoxication is one of the few intoxications that can be proven even after cremation by the analysis of urn ash. Indispensable in such a case is the investigation of several other ash samples from corpses, incinerated in the same crematorium under the same conditions (i.e. temperature, heating material, etc.) for comparison. Moreover, arsenic contamination from coffin nails and so on must be considered (Drasch 2003).

References and further reading Arndt, T. (2012a). Problematik, Klinik und Beispiele der Spurenelementvergiftung – Cadmium. Toxichem Krimtech 79: 127–136. Arndt, T. (2012b). Problematik, Klinik und Beispiele der Spurenelementvergiftung – Quecksilber. Toxichem Krimtech 79: 51–60. Baselt, R.C. (2011). Disposition of Toxic Drugs and Chemicals in Men, 9th edn. Seal Beach, CA: Biomedical Publication. BGA (Bundesgesundheitsamt) (1992). Amalgame-­Nebenwirkungen und Bewertung der Toxizität. Zahnärztl. Mitteilungen 19: 10/92.

PART VII  

1986; Flanagan et  al. 2007). Various spectrophotometric or gas chromatography methods are described in the literature (Goldbaum et al. 1986; Van Dam and Daenens 1994; Widdop 2002; Boumba and Vougiouklakis 2005; Flanagan et al. 2007). Carbon monoxide affinity to haemoglobin is in the same molar ratio as oxygen but its binding is approximately 220 times stronger. Even traces of carbon monoxide in the air can be bound to haemoglobin gradually causing potentially toxic blood level of COHb after some time. The degree of intoxication of an individual correlates with the time of exposure and physical activity. COHb levels attained rapidly are reported to result in less dangerous effects than the same levels attained gradually (Baselt 2011). Dissolved carbon monoxide in plasma enters into cells and inhibits the function of heme proteins such as cytochromes and myoglobin with adverse impacts on the heart and brain function. The prolonged enzyme inhibition has more severe toxic impacts including neuropsychological disorders (e.g. memory defects, parkinsonism) (Neuhaus 1986; Lüllmann et al. 2002; Sevela et al. 2002). Carbon monoxide is oxidised to carbon dioxide to less than 1% with the major part eliminated unchanged by pulmonary excretion. After interruption of carbon monoxide exposure, COHb levels drop by about 30–50% per hour (Neuhaus 1986). The elimination can be accelerated by administration of pure oxygen achieving a half-­life of 1.5 h approximately (Lüllmann et  al. 2002). The replacement of bonded CO by oxygen and its elimination can be accelerated by administration of hyperbaric oxygen (Sevela et al. 2002; Patocka et al. 2004; Baselt 2011). The symptoms of intoxication at levels of COHb 20–40% are manifested by headache, dizziness, disorientation, vomiting and nausea. In subjects with some physiological dysfunction, dysrhythmias can also appear, and children and elderly people are at higher risk. COHb is a cherry-­red pigment that is apparent in the skin colour of intoxicated subjects. COHb blood levels higher than 50% are associated with deep coma, cramps, metabolic acidosis, cardiovascular failure with fatal lung and brain oedema, circulation failure and heart arrest. Bleeding into the brainstem cannot be excluded due to necrosis in the CNS generated during hypoxia (Neuhaus 1986; Niesink et al. 1996; Sevela et al. 2002).

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Acute or chronic poisonings caused by some gases or solvents occur with various incidences worldwide and can be associated with environmental or occupational pollution, or industrial accidents. They can cause serious intoxications in the criminal or suicidal context, and they can be misused by mistake. Those substances likely to be seen in forensic toxicology are discussed in this chapter.

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Marie A. Balíková

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52.7  Solvents and Gaseous Poisons

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Schulz, C., Wilhelm, M., Heudorf, U. and Kolossa-­Gehring, M. (2011). Update of reference and HBM values by the German Human Biomonitoring Commission. International Journal of Hygiene and Environmental Health 215: 26–35. Seiler, H.G., Sigel, A. and Sigel, H. (eds) (1994). Handbook on Metals in Clinical and Analytical Chemistry. New York: Dekker. Specht, W. and Katte W. (1954) Giftverdacht? Hamburg, Germany: Kriminalistik-­Verlag. UNEP (United Nations Environemt Programme) (ed.) (2013). The negotiating process. http://www.unep.org/hazardoussubstances/ Mercury/Negotiations/tabid/3320/Default.aspx Wilhelm, M., Ewers, U. and Schulz, C. (2004). Revised and new reference values for some trace elements in blood and urine for human Biomonitoring in environmental medicine. International Journal of Hygiene and Environmental Health 207: 69–73.

TOXICOLOGY

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52.7.1 Gases Carbon monoxide

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Carbon monoxide is an odourless, colourless and combustible gas, which does not irritate respiratory ways. Its density is approximately of the same value as air. It is produced by incomplete combustion of various organic materials. Its common sources are from cigarette smoke, fire and automobile exhaust. Surprisingly, it can be generated in the biotransformation of dichloromethane. Carbon monoxide (CO) is generated in small amounts endogenously by the catabolism of heme and is capable of establishing background carboxyhaemoglobin (COHb) saturation levels up to c. 0.5%. In ill patients with blood disorders, COHb level can rise up to 3% (Neuhaus 1986). Smokers can achieve a saturation level of up to 10%. Levels exceeding 50% saturation are considered life threatening (Neuhaus 1986; Baselt 2011). In forensic situations, the level of COHb can indicate whether the deceased was alive and died in the fire or died before the fire. In deceased subjects without spontaneous respiration, practically no carbon monoxide resorption and elimination occurs; therefore, the level of COHb can be determined for a long time after the death. However, the analysis should be performed without delay after blood sampling because COHb level tends to decrease with time (Neuhaus

Carbon dioxide Carbon dioxide is a colourless, odourless and non-­combustible gas, with a faint acid taste, with density higher than air c. 1.5 times. It is a part of the respiration process of plants and animals, and it is the product of biotransformation of various compounds. Expired air in humans contains c. 4% CO2. It is a constituent of volcanic gases, mineral waters and can accumulate at the bottom of caves and other closed spaces. As a product of burning an organic material and a waste industrial product, it is of public concern with respect to air pollution and potential climatic adverse impacts. Carbon dioxide is usually marketed under pressure as liquid or in solid form as dry ice. Its toxicity is generally low. The environmental or workplace threshold limit is close to the physiological value of expired air. At higher concentrations, respiratory stimulation and hyperventilation take place, with

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CHAPTER 52   Toxicology of Specific Substances

Nitrogen oxides

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Ammonia is a colourless gas, lighter than air with an unpleasant pungent odour. Threshold workplace concentration in air is 25–50 ppm; brief exposures to higher concentrations up to 300– 500 ppm can be tolerated (Moeschlin 1986). It is soluble in water forming ammonium hydroxide. Usually, it is marketed under pressure in steel cylinders or as ammonia water. Vapours have irritating effects on the eyes and respiratory tract. Inhalation of vapours is associated with alkali caustic effects on the respiratory tract. In massive acute exposures, asphyxia and lung oedema can occur and hepatoxicity has also been reported (Moeschlin 1986).

Phosgene

Phosgene, or carbonyl dichloride, is a colourless highly toxic gas with a suffocating odour. It has a boiling point of 8.2°C. Its mean lethal concentration is 3.2 mg/min/L (Patocka et al. 2004). It is slowly hydrolysed in water forming HCl. It can be used in chemical synthesis and was misused as a weapon of war in the First World War. Chlorinated hydrocarbons (chloroform, tetrachloromethane, trichloroethylene) form toxic phosgene when burnt in the open space. It is lipophilic, interfering with enzyme functions in the respiratory tract, and other biological membranes causing severe pulmonary oedema or pneumonia. Symptoms are not apparent immediately, even when fatal concentrations are inhaled. Initial symptoms of poisoning are only mild irritation in the respiratory tracts, headache, constricted feeling in the chest and vomiting. Full development of lung oedema can appear 6–12 hours after the dose, and death after 24–48 hours. Inhalation of sublethal doses causes choking, constricted feeling in the chest, coughing, impaired and painful breathing, headache and lassitude and bloody sputum. Vapours strongly irritate the eyes. (Moeschlin 1986; Merck Index 1996; Patocka et al. 2004).

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Dinitrogen oxide (N2O), or nitrous oxide, is a non-­toxic colourless gas that is heavier than air. Its content in atmosphere is of ecological interest since it belongs to the greenhouse gases. It is produced naturally in the soil during microbial processes. When inhaled, it has analgesic effects and it has application in medicine as a mild general anaesthetic, mainly in paediatrics. Its initial and transient excitation effects can be the reason for recreational use to achieve euphoria and slight hallucination. Nitric oxide (NO) is a colourless gas that reacts with oxygen to form nitrogen dioxide (a brown gas). It is used in the manufacture of nitric acid and as a bleaching or stabilising agent in industry. On contact with air, it is converted immediately to the more toxic nitrogen dioxide. Nitric oxide and nitrogen dioxide occur together as air polluting gases. Nitrogen dioxide (NO2) is a reddish-­brown gas with an irritating odour, perceptible at 0.11 ppm in air, and is poisonous. Nitrogen dioxide forms nitric acid upon contact with water, it is the intermediate in nitric acid manufacture, and it can be used as a nitration agent in the industrial production of various organic compounds. Acute toxic effects may appear with some time delay causing inflammation of the lungs leading to oedema several days later that may be fatal (Merck Index 1996; TOXNET NLM HSDB 2013). Nitric oxide in variable ratios with nitrogen dioxide is an irritating and polluting gas in the atmosphere, and its contents are of environmental concern. Exposure to these oxides results in pulmonary function impairment and possibly to development of methaemoglobinaemia and other adverse impacts.

Ammonia

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Ozone or triatomic oxygen is found in the atmosphere in various concentrations. It belongs to the group of photooxidants that are formed either naturally or by electrical discharge from lightning and from excessive smog at lower atmospheric layers (photo smog). The normal concentrations in air are reported within the range 0.02–0.04  mg/m3. In laboratory lamps, it is produced by ultraviolet (UV) radiation. It is a bluish explosive gas with a characteristic odour. Its usage is as a disinfectant due to its oxidative power. Concentrations higher than 0.2 mg/m3 may cause irritation of the eyes, skin and respiratory tract, causing bronchitis or asthma (Wagner 1984; Moeschlin 1986).

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Ozone

­ roduced by volcanic eruptions. Sulphur dioxide is an intermedip ate in the production of sulphuric acid. In burning sulphur containing coal or oil, sulphur dioxide emissions are a precursor of acid rain and contribute to air pollution with impact on human health. Sulphur dioxide is toxic in large amounts with long-­term exposure by inhalation associated with irritation of the eyes, skin and bronchoconstriction. The workplace threshold air concentration is 2 ppm (Moeschlin 1986). To date, its biological role in mammals has not been understood fully.

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headache, faintness, cramps and circulation failure. Humans cannot breathe air containing more than 10% carbon dioxide without adverse symptoms and at higher concentrations above 20% asphyxia and death occur (Moeschlin 1986).

Sulphur dioxide Sulphur dioxide is a colourless gas with a typical suffocating odour. It has antimicrobial properties, and it is used as a preservative in vegetables and fruits as a disinfectant and antioxidant in the food industry and as a bleaching compound. It is also

Phosphine Phosphine or hydrogen phosphide is a colourless poisonous gas, heavier than air with a typical garlic or decaying fish odour. It is spontaneously flammable in air. Toxicity is extremely high, with threshold workplace air concentration at only 0.3 ppm. Inhalation of 50 ppm can be immediately dangerous to life (CDC NIOSH 2011). Phosphine can be formed in traces by putrefaction of organic material containing phosphorus. Phosphine is also

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Fluorine is a diatomic pale-­yellow gas. Fluorine is the most electronegative element and the most reactive non-­metal. In nature, it occurs chemically bound in various compounds. Hydrogen fluoride or hydrofluoric acid gas is colourless, fuming in air, at a boiling point of 19.5°C. It is very soluble in water. Its salts, such as fluorides, are of low toxicity. Sodium fluoride is used in the fluoridation of drinking water, as a preservative or disinfectant, and as an insecticide. Acute poisoning via hydrogen fluoride inhalation is manifested with burns and necroses in the respiratory tract as for chlorine. Hydrogen fluoride is very aggressive to skin and eyes, causing severe burns, which may not be painful or visible for several hours (Moeschlin 1986; Merck Index 1996).

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Fluorine

fire smoke from burning nitrogen-­containing plastics. Its other potential sources are from biotransformation of organic compounds as nitriles (e.g. acetonitrile, acrylonitrile) or glycoside amygdaline and the antihypertensive drug nitroprusside. Poisoning occurs from inhalation of gaseous hydrogen cyanide or due to ingestion of various compounds producing hydrogen cyanide in the body. Hydrogen cyanide is formed from ingestion of amygdaline, nitriles or cyanides bound in various complex metal compounds (e.g. cyanocuprate, cyanonickelate). The hydrogen cyanide formation from these chemicals is slower than from the alkali metal salts; hence, toxic symptoms can be gradual or can be delayed. After inhalation of hydrogen cyanide, the absorption is very quick. At an air concentration of 300 ppm, death can occur within 1 minute, while at 100 ppm, death occurs within 1 hour (Sevela et al. 2002). The workplace threshold limit value for gaseous hydrogen cyanide is 10 ppm (11 mg/m3) over 8 hours, and 110 ppm value can be fatal after 1 hour (Moeschlin 1986; Baselt 2008). The toxic symptoms after ingestion of potassium or sodium salts are almost as fast as gas inhalation due to their fast hydrolysis in the gastric environment. Lethal doses for an adult ingesting potassium cyanide are 200–300  mg (Sevela et al. 2002; Patocka et al. 2004); however, doses of 600 mg potassium cyanide can lead to recovery with supportive therapy (Baselt 2011). The minimum lethal dose of sodium or potassium cyanide is estimated to be 3 mg/kg of body weight (Sevela et al. 2002). The reference plasma cyanide values in healthy subjects were reported at 0.004  mg/L in non-­ smokers and 0.006  mg/L in smokers, whereas in another group whole blood levels in non-­smokers were 0.016 mg/L and 0.041 mg/L in smokers. The average blood values of 0.18  mg/L (non-­smokers) and 0.56  mg/L (smokers) were reported in chronically cyanide-­exposed workers (Baselt 2011). Comatose or fatal plasma or serum values range from 1 to 3 mg/L (Schulz and Schmoldt 2003). Clinical features of poisoning include coma, respiratory arrest and cardiovascular collapse. The major part of the cyanide dose is metabolised in the liver to thiocyanates that are excreted in urine. The main toxic effect is inhibition of various enzymes including ferricytochromoxidase. Therapy for cyanide poisoning must start as soon as possible and includes high flow of oxygen and cardiopulmonary resuscitation. Administration of sodium thiosulphate supports the conversion to non-­toxic thiocyanates. Cobalt ethylenediaminetetraacetic acid (EDTA) or hydroxocobalamin binds cyanides into complexes and acts fast and efficiently, whereas the action of sodium thiosulphate is rather slow. Amyl nitrite, sodium nitrite or 4-­dimethylaminophenol is used as an antidote to form methaemoglobin and binds cyanides into the complex cyanomethemoglobin, thus restoring the ferricytochromoxidase function. However, not only the efficiency of antidotes but also their safety has been under debate (Sevela et al. 2002; Megarbane et al. 2003; Hall et al. 2007). Laboratory confirmation of acute cyanide poisoning is not usually necessary since poisoning occurs so rapidly. Cyanides are present largely in erythrocytes; therefore, whole blood is preferable to assay. After death, the concentration of cyanide is not stable in cadavers nor in stored tissue samples. Both decrease and

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formed by decomposition of metal phosphides in water or acid media. It is a component in acetylene manufacture from calcium carbide, which is attained by reduction in natural carbonate. During this reduction process, traces of phosphates present in raw material are reduced to phosphides. Phosphine is toxic to the respiratory tract, nervous system, liver and heart. Initial acute symptoms of intoxication resemble Salmonella or meat poisoning. Rapid coma and death may occur at high exposure associated with respiratory failure, heart arrest and lung oedema. Poisoned subjects who overcome the most critical stage can be afflicted by hepatoxicity and nephrotoxicity with a latency of 24–48 hours. In chronic exposure, cases of cardiomyopathy can develop (Moeschlin 1986).

TOXICOLOGY

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Chlorine

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Chlorine is a diatomic greenish-­yellow gas with a sharp suffocating odour. It is marketed in the form of compressed gas in steel cylinders. It is used in the industrial production of various materials, in the manufacture of chlorinated lime, as a bleaching and disinfecting agent and can be used for purifying water. Chlorine is a powerful irritant and can cause fatal pulmonary oedema. However, its typical sharp odour is apparent even in great dilution in air and this property usually prevents serious poisoning. Threshold odour detection is reported as 0.02– 0.04 ppm (Merck Index 1996). In contact with moisture such as wet skin or the respiratory tract, it is transformed into caustic hydrochloric acid causing burns.

Cyanides Hydrogen cyanide is a very toxic colourless gas or liquid (hydrocyanic acid) of a bitter almond-­like odour, with a boiling point of 26°C. Hydrocyanic acid and its various salts are used in industry for various purposes, such as electroplating solutions, metal polishes, fungicides and insecticides. The sodium or potassium salts of hydrogen cyanide are unstable in acidic media and under air access are gradually converted to carbonates. Hydrogen cyanide is present in exhaust gases, in tobacco and wood smoke, and in

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CHAPTER 52   Toxicology of Specific Substances

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The toxicity of aliphatic hydrocarbons is higher with more volatile methane, propane and butane. The major intake is via the lungs; absorption through the skin is negligible. They are excreted mainly as unchanged compounds via the lungs associated with a typical odour in expired air, with only a minor proportion excreted in urine. Volatile hydrocarbons can produce dose-­ related CNS depressant effects, with low doses producing euphoria or hallucinations. Higher doses may be life threatening causing asphyxia, convulsions and coma. The direct cause of death is cardiotoxicity or CNS toxicity. Inhalation of n-­butane may be associated with hypotonia, ventricular fibrillation, encephalopathy and myocardial infarction (Baselt 2008). Death may occur also indirectly by aspiration of gastric contents. Oral consumption of gasoline causes gastrointestinal mucosal membrane injury and affects respiration through aspiration and local toxicity in the lungs. The risk of aspiration is associated with viscosity, surface tension and volatility of hydrocarbons. Hydrocarbons with low viscosity and high volatility are more likely to cause aspiration pneumonia among other symptoms (Moeschlin 1986).

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Hydrogen sulphide is a colourless, flammable, poisonous gas that is heavier than air, with a typical odour of rotten eggs. It is formed during putrefaction of sulphur containing materials and can be produced as a by-­product of industrial processes. It also occurs naturally in mines, volcanic gases and hot sulphur springs. The workplace threshold concentration is 10 ppm (Moeschlin 1986). It is an insidious poison since the sense of smell may fail to give a warning at high concentrations. In humans, it is extremely hazardous. Low concentrations cause irritation of the conjunctival and mucosal membranes and cause headache, dizziness, nausea and lassitude (Merck Index 1996). It inhibits cytochromoxidases causing cellular asphyxia, leading to coma and death from respiratory failure within minutes of exposure (Table 52.7.1).

Aliphatic hydrocarbons

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Hydrogen sulphide

usually associated with inhalation of their vapours deliberately (glue or sniffing cigarette lighter refills) or from occupational or environmental exposure. Oral consumption is rare.

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increase in cyanide levels have been observed. The mechanisms include loss by evaporation, thiocyanate formation and reaction with tissues. On the contrary, under different conditions, the conversion of thiocyanate to cyanide has also been reported. The addition of sodium fluoride to samples to diminish postsampling changes is recommended (Flanagan et al. 2007; McAllister et al. 2008; Baselt 2011).

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52.7.2 Solvents

Aromatic hydrocarbons

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Crude oil, petroleum and gasoline are rich sources of hydrocarbons: both aliphatic and aromatic with various molecular structures and with a broad range of volatility. In crude oil, the predominating ones are paraffins (alkanes), naphthenes (cycloalkanes) and aromatics in variable abundance depending on the origin of the oil. The content of aromatics is limited in final gasoline products. Natural gas or fuel gas is the source of light hydrocarbons. Hydrocarbons are parts of various industrial and household solvents and are used commonly in laboratories and in industry. Hydrocarbons are strongly lipophilic substances capable of depressing the CNS. Poisoning with volatile substances is

Table 52.7.1  Toxicity of hydrogen sulphide (H2S). H2S concentration in air (ppm)

Biological effects

0.03

Smell threshold

0.05–30

Rotted eggs smell

50–100

Mild toxicity, respiratory tract irritation

>150

Smell sense paralysis

>500

Loss of consciousness, circulation collapse, respiratory arrest, death within 60 min

>1000

Immediate death within 1 min

Source: From TOXNET NLM HSDB (2013), © US National Library of Medicine.

Aromatic hydrocarbons have a high potential for depression of CNS due to their solubility in fat. They have lesser potential for cardiac disturbances and lesser risk of aspiration compared to aliphatic hydrocarbons. Nevertheless, the acute toxic effects of benzene are also due to myocardial sensitisation. The aromatic hydrocarbons have significant systemic toxicity. The health risk of chronic exposure to benzene includes haematological disturbances and carcinogenicity. Benzene is oxidised to phenol and other minor products. The majority of phenol is excreted by urine in conjugated form. Toluene vapour exposure is associated more with CNS depressant effects rather than organ toxicity, even if long-­term abuse can lead to encephalopathy, neurotoxicity and hepatorenal damage. Toluene is oxidised to benzoic acid, followed by conjugation with glycine or glucuronic acid. The glycine conjugate (hippuric acid) represents the major excretory product in urine. Xylene occurs in three structural isomeric forms. Xylenes are components of solvents used in paints, cleaning agents, pesticides and gasoline. Their toxicity is very similar to toluene (Moeschlin 1986; Sevela et al. 2002).

Chlorinated hydrocarbons Chlorinated hydrocarbons are used widely in industry and occur as components in various commercial cleaning products which are also available in households. Poisoning may occur by inhalation; intentional or accidental ingestions are rare. These compounds are highly lipid soluble with significant depressive effects

PART VII  

on CNS and with repeated doses tend to accumulate in adipose tissues (Moeschlin 1986). Tetrachloromethane, chloroform, tri-­ or tetrachloroethylene can induce hepatorenal toxicity, and tetrachloromethane is a potential carcinogen in humans. Chloroform had been used as an anaesthetic agent in surgery, but was replaced many years ago by less toxic compounds (halothane, 2-­bromo-­2-­ chloro-­ 1,1,1-­ trifluoroethane). Dichloromethane is considered less hepatotoxic than other chlorine substituted methanes. However, it is converted in  vivo to carbon monoxide. Trichloroethylene is extensively metabolised in humans, producing chloralhydrate as a transient metabolite with hypnotic activity. The major urinary metabolites are trichloroethanol and trichloroacetic acid, excreted partly as glucuronide conjugates. In an open flame, many chlorinated hydrocarbons are oxidised to highly toxic phosgene.

TOXICOLOGY

HO-CH2-CH2-OH

Ethylene glycol

HO-CH2-C

O

HO-CH2-C

H

Glycolic aldehyde

O

OH

O C-C

H

OH

Glyoxylic acid

O

OH

O

Glycols and their derivatives

O

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Formic acid

O C-C

HO

OH

Oxalic acid

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Figure 52.7.1  Biotransformation scheme of ethylene glycol.

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which is manifested by a pH 2 kDa

 6

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Initial testing procedure, MS-­based

N

#

Differentiation of endogenous / synthetic anabolic steroids

Urine

GC/C/IRMS*

 7

Proteases

Urine

SDS-­PAGE*, LC-­MS

 8

Small interfering RNA (siRNA)

Urine

SDS-­PAGE, LC-­MS

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O

LC-­MS

 9

Artificial oxygen carriers

Serum

LC-­MS

10

Xenon

Urine, serum

Headspace-­ GC-­MS

#

Initial testing procedure, not MS-­based

Matrix

Approach

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 1

Human chorionic gonadotropine (hCG)

Urine

ECLIA*

 2

Erythropoiesis-­stimulating agents (ESAs)

Urine, Serum

IEF* & WB, SDS-­/SAR*PAGE & WB*

 3

Haematological parameters

Blood

Flow cytometry

 4

Human growth hormone (hGH)

Serum

ILMA*

 5

Homologous blood transfusion

Blood

Flow cytometry

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52.10.2  Analytical aspects

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In doping controls, the preferred analytical techniques are mass spectrometry-­based methods due to its inherent identification power for most prohibited substances. Merely, the analysis of higher molecular mass analytes such as proteins (e.g. erythropoietin, growth hormone) or the determination of classical blood parameters (haematocrit, reticulocytes etc.) are realized by other complementary analytical approaches. A summary of the used methods and their application is shown in Table 52.10.3.

Mass spectrometric analysis Depending on the physical and chemical properties of the target analytes, mass spectrometry is commonly coupled to gas chromatography (volatile compounds) or liquid chromatography (non-­volatile compounds) in order to separate the complex sample matrix from the target analytes. While single quadrupole systems prevailed in the early days of systematic doping

LC-­MS

Urine, serum

TO

for both the athlete and the anti-­doping organization (ADO), because the whereabouts of all athletes must be known at all times. This has been realized and standardized worldwide by the introduction of the Anti-­ Doping Administration and Management System (ADAMS) of WADA in 2005. This system records and stores whereabout data, any therapeutic use exemptions (TUE), and laboratory test results for all athletes. The access rights are regulated accordingly, so that, for example, an athlete can only enter and change his/her whereabouts but has no access to other data. Extensive OOC testing currently enables a potentially complete monitoring of the athlete, although this of course considerably affects for the athlete’s privacy. Not all samples (urine or blood) are subjected to the analysis of the complete portfolio of prohibited substances, e.g. archery athletes are not tested for EPO or growth hormone, while 60% of all samples from cyclists are tested for EPO and 30% of the powerlifting samples are subjected to growth hormone analysis. Here, the technical document for sport specific analysis specifies the respective numbers of recommended analysis for each kind of sport and the respective federations were prompted to fulfil the criteria of this document (WADA 2020a). Sampling is executed and supervised by trained and authorized doping control personnel, which ensures the reliability of the complete sampling process. This is required due to the possible manipulation of the samples by addition tampering agents (e.g. proteases) or urine substitution. The sample collection always includes the splitting of the obtained volume into A-­and B-­sample, thus, in case of an adverse analytical finding, the athlete has the right to request the B-­sample analysis. The most crucial requirement of doping control sampling and analysis is the independence of all participating stakeholders. Thus, it must be ensured that the samples are anonymised, and information provided with each sample is limited to matrix, test type (IC/OOC), gender, discipline and medication taking during the last 7 days.

TOXICOLOGY

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 Abbreviations: GC-­MS: gas chromatography mass spectrometry, NPD: nitrogen-­phosphorus detector, LC-­MS: liquid chromatography mass spectrometry, IRMS: isotope-­ratio mass spectrometry, SDS-­PAGE: sodium dodecyl sulphate-­polyacrylamide gel electrophoresis, ECLIA: electrochemiluminescence immunoassay, IEF: isoelectric focusing, WB: western blotting, SAR: sarcosyl, ILMA: immunoluminometric assay.

*

controls, more recently tandem mass spectrometry (MS/MS) and/or high-­resolution mass spectrometry (HRMS) systems are utilized due to their superior performance. The unambiguous identification of the prohibited substance is enabled with minimum mass spectrometric and chromatographic criteria, which are defined in WADA’s technical document TDIDCR2021 (WADA 2020c). The analytical data of a suspicious sample are directly compared to the results of a blank sample and a blank

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CHAPTER 52   Toxicology of Specific Substances

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the retrospective evaluation of individual steroid profiles from different samples from the same athlete by independent experts enable the uncovering on an AAS misuse much more efficient than the T/EpiT alone. Valid evidence concerning the endogenous or exogenous origin of steroids is provided by IRMS. This technology enables the determination of isotopic features of steroids by directly measuring (stable) isotopes of an element. In sports drug testing, usually carbon isotope ratios (CIRs) are measured. Stable isotopes of carbon are 12C and 13C (one additional neutron) with a natural distribution of 98.9% 12C and 1.1% 13C. By determination of the δ13C values of target compounds such as T, EpiT, androsterone and etiocholanolone, the abundance of 13C of the respective substance in part-­per-­thousand (per-­mille, ‰) in relation to a fixed r­ eference standard is calculated. Significantly relevant differences in the abundance of 13C are expressed and indicate the exogenous origin of the administered steroids or its metabolites in ­comparison to an endogenous unaffected precursor (most often, pregnanediol). The analysis of CIRs requires the complete combustion of the molecule to CO2 and detection in a magnetic sector mass analyser. Such analysers detect ions with a mass at m/z 44, 45 and 46 by means of three Faraday cups. Therefore, the detection of these ions is very accurate and reliable. This is, in combination with optimal chromatographic conditions and resulting well separated and pure peaks, the basis for valid IRMS analysis. The ratio of m/z 45 to m/z 44 is monitored for the target steroids, which (in the case of endogenously produced steroids) reflects the individual carbon isotopic signature (mostly influenced by the nutrition). In contrast, synthetically produced steroids originate from other natural sources (e.g. phytosterols from soya). Here, the 13C-­content is significantly reduced, which can be distinguished from endogenously produced steroidal compounds accordingly. Thus, samples showing conspicuous or abnormal steroid profiles in the initial testing procedure are subjected to subsequent IRMS analysis to further confirm the presence of exogenous steroids. Noteworthy, IRMS confirmations can also potentially be applied to other endogenous, prohibited substances such as 5-­aminoimidazole-­4-­carboxamide ribonucleotide (AICAR) or corticoids (Piper et al. 2014).

TO

sample fortified with the suspected target analyte (positive control), all of which are tested in the same analytical batch. The minimum criteria for the chromatography define (besides others) a maximum permissible retention time difference (resp. relative retention time shift, if an internal standard is used) between the sample and the positive control of 90

30–50

21

Dihydropyridine calcium channel blocker

Aprindine

95–98

24–48

3

Class Ib antiarrhythmic agent

Atenolol

5–10

6–9

1

β-­adrenergic antagonist, ISA (−), MSA (-­)

Benazepril

94

0.6

0.7

Angiotensin converting enzyme inhibitor

Betaxolol

55

14–22

4.9–9.8

β-­adrenergic antagonist, ISA (−), MSA (low)

Bisoprolol

30

9–12

3

β-­adrenergic antagonist, ISA (−), MSA (-­)

Captopril

30

1–6

0.7

Angiotensin converting enzyme inhibitor

Carteolol

20–30

8–12

4

β-­adrenergic antagonist, ISA (+), MSA (-­)

FO

Drugs

(Continued)

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TOXICOLOGY

Protein binding (%)

Half-­life (hr)

Vd (l/kg)

Note

Carvedilol

25–35

6–8

1–2

β-­adrenergic antagonist, ISA (−), MSA (+)

Digitoxin

97

96–240

0.7–0.9

Digitalis

Digoxin

20

30–45

5.1–7.4

Digitalis

Diltiazem

80

2–5

3

Benzothiazepine calcium channel blocker

Disopyramide

20-­75

5–9

0.6

Class Ia antiarrhythmic agent

Enalapril

50-­60

2

1.7

Angiotensin converting enzyme inhibitor

Felodipine

> 99

11–16

10

Dihydropyridine calcium channel blocker

Flecainide

60

11–15

7–10

Class Ic antiarrhythmic agent

Isosorbide dinitrate

28

0.4–0.8

6.3–8.9

Organic nitrate

Isosorbide mononitrate

95

8

Nifedipine

99

2

Nisoldipine

99

7–12

Nitrendipine

98

10–22

Nitroglycerin

60

0.12

Olmesartan

99

Phenytoin

87–93

Pilsicainide

35

Pindolol

50

Procainamide

15

Propafenone Propranolol Quinapril Quinidine

N

Class Ib antiarrhythmic agent

R

U

SE

Angiotensin converting enzyme inhibitor

0.7

Dihydropyridine calcium channel blocker

1

Dihydropyridine calcium channel blocker

4–5

Dihydropyridine calcium channel blocker

4–8

Dihydropyridine calcium channel blocker

TO

U

LY

Drugs

O

Table 52.11.2  (Continued)

Organic nitrate

0.2–0.3

Angiotensin II receptor blocker

8–60

0.5–0.8

Class Ib antiarrhythmic agent

4–5

1.5

Class Ic antiarrhythmic agent

3–4

2

β-­adrenergic antagonist, ISA (+), MSA (+)

2–3

1.7–2.4

Class Ia antiarrhythmic agent

75–88

3–5

3.7

Class Ic antiarrhythmic agent

90

3–5

4

β-­adrenergic antagonist, ISA (−), MSA (+)

97

1

0.3

Angiotensin converting enzyme inhibitor

80–90

6–8

3

Class Ia antiarrhythmic agent

10

7–11

1.2–2.4

Class III antiarrhythmic agent

>99

18–30

6.6–7.6

Angiotensin II receptor blocker

80

0.6–0.8

0.2–0.3

Angiotensin converting enzyme inhibitor

Valsartan

95

4–12

0.14– 0.34

Angiotensin II receptor blocker

Verapamil

90

4.5–12

5

Phenylalkylamine calcium channel blocker

Telmisartan Trandolapril

ISA: intrinsic sympathomimetic activity MSA: membrane stabilizing activity Vd: volume of distribution

N O

C

R FO

Sotalol

TR IB

2.6

6–15

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CHAPTER 52   Toxicology of Specific Substances

affected by various factors such as sample condition, redistribution, and drug interactions, we have to consider these factors for proper interpretation (Pelissier-­Alicot et  al. 2003; Ferner 2008; Skopp 2010).

Table 52.11.3  Drugs that increase blood concentration of antiarrhythmic drugs Antiarrhythmic drug

Concomitant drug or substance

Amiodarone

Grapefruit juice

Aprindine

Amiodarone

52.11.2  Antiarrhythmic agents

Diltiazem Verapamil Disopyramide

U

TR IB

N

O

C

R

FO

Clarithromycin Erythromycin Indinavir

LY

Nelfinavir Ritonavir

O

Amiodarone

R

U

SE

Procainamide

N

Saquinavir

Flecainide

Propafenone

TO

An arrhythmia is defined as not sinus rhythm. Antiarrhythmic agents are used for the treatment of various kinds of arrhythmias. However, since antiarrhythmic agents themselves have arrhythmogenic properties, overdoses of those drugs may cause life-­threatening outcomes. Antiarrhythmic agents have been historically classified by the mechanism of action of each drug (Vaughan Williams classification) into four groups, class I (sodium (Na+) channel blockers), class II (β-­adrenergic antagonists), class III (potassium (K+) channel blockers), and class IV (calcium channel blockers). In this section, mainly class I and class III agents are described, and since there is a high incidence of poisoning reported by class II (β-­adrenergic antagonists) and class IV (calcium channel blockers) agents, they will be described in a separate section. All class I agents depress the voltage-­gated fast sodium channel. They are divided into three subgroups, class Ia (disopyramide, procainamide, quinidine), class Ib (lidocaine, mexiletine, phenytoin), and class Ic (flecainide, pilsicainide, propafenone: also block the potassium channel). The class I agents depress myocardial contractility, excitability, and conduction velocity, and the QRS or QT interval is prolonged. Sinus bradycardia, asystole, and polymorphous ventricular tachycardia are observed with overdose. The class III drugs (amiodarone, sotalol) prolong the action potential and effective refractory period. This results in QT interval prolongation with therapeutic doses by blocking the potassium channel. Since most of these antiarrhythmic drugs have a narrow therapeutic index, toxic symptoms such as hypotension, bradyarrhythmia, and ventricular arrhythmia due to QRS and QTc prolongation are observed with overdose. And we also consider pharmacokinetic drug interaction with other drugs, which increase the blood concentration of antiarrhythmic drugs (Table 52.11.3) (Anderson and Nawarskas 2001; Aonuma et al. 2017). The diagnosis of poisoning is based on the history of use of an antiarrhythmic agent and electrocardiographic and cardiac findings. In fatal cases, the postmortem drug concentration is one of the important factors for making the diagnosis. However, postmortem changes of blood concentrations of antiarrhythmic drugs depend on the sampling site (central or peripheral) and time. Concentrations of some kinds of drugs in postmortem samples were higher than those of antemortem serum (O’Sullivan et  al. 1995). This would be due to postmortem redistribution by ­various factors such as lipophilicity of the drug, distribution volume, and the state of ionization by pH changes (Pelissier-­Alicot et al. 2003; Ferner 2008; Skopp 2010; Yoshitome et al. 2008, 2010). These should be considered when interpreting concentrations in postmortem samples.

Quinidine

Cimetidine Amiodarone Cimetidine Levofloxacin Ofloxacin Trimethoprim Amiodarone Cimetidine Quinidine Amiodarone Cimetidine Erythromycin Itraconazole Ketoconazole

Sources: Adapted from Anderson and Nawarskas (2001); Aonuma et al. (2017).

52.11.3 β-­adrenergic antagonists (β-­blockers) The β-­adrenergic antagonists are used for the treatment of various cardiovascular diseases such as coronary heart diseases, hypertension, and tachyarrhythmia. The β-­adrenoceptor has three subtypes (β1, β2, and β3) and β1 and β2-­adrenoceptors increase cardiac performance acutely. The β1 and β2-­ adrenoceptors couple to the Gs-­protein-­adenylyl cyclase pathway, and its stimulation increases cyclic AMP (cAMP). The activation of β1-­adrenoceptors increases the opening of L-­type calcium channels via cAMP in cardiomyocytes, and it is a powerful physiologic mechanism to acutely increase cardiac performance (Brodde and Michel 1999; Brodde et  al. 2006). The β-­adrenergic antagonists cause negative inotropic and chronotropic responses by reducing the facilitation of calcium entry into cardiomyocytes.

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“intake during playing” and “medication errors” were noted in children (Christensen et al. 2018). The calcium channel blockers are divided into three groups by their chemical structure: (1) dihydropyridines (nifedipine, nicardipine, amlodipine); (2) benzothiazepines (diltiazem); and (3) phenylalkylamines (verapamil). Calcium channel blockers inhibit L-­type voltage-­sensitive calcium channels. The affinities of each group are different. At therapeutic doses, the dihydropyridines primarily affect smooth muscle in peripheral vessels, and they act as potent vasodilators, with little effect on the heart. On the other hand, benzothiazepines and phenylalkylamines affect both peripheral vessels and the heart. Since the tissue selectivity would be attenuated with increasing doses, this selectivity may be lost in overdose cases (DeWitt and Waksman 2004; Arroyo and Kao 2009; DeRoos 2011; Christensen et al. 2018). In overdose situations, calcium channel blockers decrease the force of contraction, decrease AV nodal conduction, and depress sinus node activity. They also cause peripheral vasodilation. Severe hypotension and bradycardia are observed. In addition to the above vital sign findings, the diagnosis is made based on the various findings, such as nausea and vomiting, metabolic acidosis, and hyperglycemia, with electrocardiographic findings such as sinus arrest or AV block without QRS prolongation. Hyperglycemia is caused by blockade of insulin release, which is dependent on the calcium influx via L-­type calcium channels (DeWitt and Waksman 2004; Arroyo and Kao 2009; DeRoos 2011). Measurement of postmortem drug concentrations confirms the diagnosis of the presence of a parent substance (the cause of poisoning) with its pharmacologically active metabolite (Kinoshita et al. 2003). The calcium channel blockers are well absorbed following ingestion, and they are metabolized in the liver. However, since the absorption of sustained-­release preparations is delayed, the toxic symptoms are delayed and continue for a long time (Ashraf et al. 1995). It is important to collect more detailed information concerning the preparation of the suspected drug (DeWitt and Waksman 2004; Arroyo and Kao 2009; DeRoos 2011). We also consider drug-­drug interactions with other cardiovascular drugs and substances. Co-­ingestion of verapamil and a β-­adrenergic antagonist may cause severe hypotension or bradyarrhythmia (Kinoshita et al. 2003). Since most calcium channel blockers are metabolized by CYP 3A4, grapefruit juice may increase the blood concentration of dihydropyridines such as nifedipine and nisoldipine (Bailey et al. 1998; Anderson and Nawarskas 2001).

52.11.4 Calcium channel blockers (Calcium channel antagonists)

Digitalis is one kind of cardiac glycoside present in the foxglove plant (Digitalis purpurea), which has been extracted from the plant leaf since ancient times. However, today it is produced in pharmaceutical plants and widely prescribed for the treatment of congestive heart failure or supraventricular arrhythmias. Digoxin and digitoxin, a derivative of digoxin, are commonly used. Although the differences in the chemical structure of each substance are small, the pharmacokinetic parameters, such as volume of distribution, biological half-­life, and protein binding rate,

FO

R

C

O

N

TR IB

U

TO

R

U

SE

O

N

LY

The β-­adrenergic antagonists have different pharmacological properties, such as receptor selectivity, intrinsic sympathomimetic activity (ISA), lipid solubility, and MSA, and they are used in their classification (DeWitt and Waksman 2004). All β-­adrenergic antagonists have an antagonistic action to the β1-­adrenergic receptor. Selectivity for the β1-­adrenergic receptor was observed with atenolol and metoprolol. Since β2-­adrenergic receptors mediate bronchodilation and regulation of insulin secretion, selective β1-­adrenergic receptor antagonists are safer for patients with asthma or diabetes mellitus than non-­selective β-­ adrenergic receptor antagonists such as propranolol or carteolol. However, this selectivity for the β-­adrenergic receptor is lost in overdose cases. ISA means the pharmacological action of a partial agonist at β-­adrenergic receptors. Since several β-­receptor antagonists such as pindolol and carteolol have ISA, they may avoid the decreased heart rate and may be safer in overdose situations. Seizure and coma were observed in cases of overdose by β-­ adrenergic receptor antagonists with high lipid solubility such as propranolol. The lipid soluble drugs are absorbed rapidly from the gastrointestinal tract following ingestion, and they easily enter the central nervous system. On the other hand, water soluble drugs tend to have slow absorption and are eliminated by the kidneys. Several β-­adrenergic receptor antagonists inhibit fast sodium channels, similar to class I antiarrhythmic agents, such as propranolol, acebutolol, and labetalol. This pharmacological action is called MSA. It depresses myocardial contractility and conduction velocity, and QRS widening is observed, and this modifies the toxicity in overdose situations. It is associated with the fatal outcome of poisoning by β-­adrenergic receptor antagonists. The β-­adrenergic receptor antagonists impair cardiac function by the competitive antagonism of the action of catecholamines at receptors. They cause depressed myocardial contractility and conduction, bradycardia, and hypotension (Brodde et  al. 2006). Refractory bradycardia and hypotension may be observed in overdose situations, with fatal outcomes in severe cases. In cases of co-­ingestion with other drugs such as calcium channel blockers, the toxicity of those drugs may be increased due to the synergistic effects (Kinoshita et al. 2003). The diagnosis of overdose is based on manifestations such as bradycardia and hypotension and a history of ingestion. Most of the symptoms occur within 2 hours after an acute overdose, and a small number of cases involving sustained-­release drugs showed delayed onset of symptoms and prolonged toxicity. In fatal cases, measurement of the postmortem drug concentrations confirms its diagnosis.

The calcium channel blockers are widely used for the treatment of hypertension, angina, arrhythmias, migraine, and subarachnoid hemorrhage. Since the calcium channel antagonists have a small therapeutic index, their overdose is frequently ­life-­threatening. It has been reported that “suicidal poisoning” and “medication errors” were frequent as a cause of poisoning in adult cases, and

52.11.5 Digitalis

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CHAPTER 52   Toxicology of Specific Substances

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I is converted to angiotensin II, a potent vasopressor peptide, by the angiotensin converting enzymes (ACEs) in pulmonary endothelial cells. Since ACEIs inhibit ACEs and reduce the formation of angiotensin II, they are potent vasodilators and widely used for the treatment of hypertension and heart failure. Since aldosterone synthesis is regulated by angiotensin II, ACEIs also decrease blood pressure (Flaten and Monte 2017; Laurent 2017). Hypotension was observed in overdose cases, and fatal cases were rare (Waeber et al. 1984; Park et al. 1990; Lip and Ferner 1995). Dry cough and angioedema have been reported as side effects. They are thought to be due to the increases of bradykinin concentration and other peptides by the inhibition of ACEs (Flaten and Monte 2017; Laurent 2017). Although angioedema occurs in 0.1–0.7% of patients taking ACEIs, it may cause life-­threatening airway obstruction with the development of edema in the upper respiratory tract (Inomata 2012).

Angiotensin II receptor blockers (ARBs)

R

U

Most vasoconstrictive effects of angiotensin II are mediated via the type I angiotensin II receptor. ARBs antagonize the pharmacological action of angiotensin II and reduce blood pressure (Flaten and Monte 2017; Laurent 2017). ARBs are also used for the pharmacotherapy of hypertension. Since ARBs do not interfere with bradykinin metabolism, the risk of angioedema is less. However, cases of ARB-­ associated angioedema have been reported (Malde et al. 2007; Brown et al. 2017).

FO

R

C

O

N

TR IB

U

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are markedly different. The pharmacological mechanism of digitalis is due to the inhibition of sodium-­potassium adenosine triphosphatase (Na+-­K+-­ATPase) on the surface of cardiomyocytes. This results in the elevation of intracellular calcium concentrations and increases the cardiac contractile force, which appears as a positive inotropic effect. Digitalis also increases vagal tone at a high level. It decreases the conduction velocity of the sinus and of atrioventricular node conduction. The automaticity of Purkinje fibers is also increased by the intracellular calcium accumulation. As a result of these pathophysiological actions, prolongation of the PR interval and shortening of the QT interval are observed on electrocardiographic findings. Most cases of acute poisoning by digitalis occur as accidental ingestion, attempted suicide, and errors (Burchell 1983; Kinoshita et al. 2009). Since the therapeutic range of digitalis concentration is narrow, monitoring of the serum concentration is important to prevent poisoning. Since patients with cardiovascular diseases are prescribed many kinds of drugs, such as for hypertension or arrhythmia, especially elderly patients, we have to consider drug interactions. Digoxin is eliminated through the kidneys, mediated by P-­glycoprotein (P-­gp), an ATP-­binding cassette (ABC) transporter. Verapamil and diltiazem increase the blood digoxin concentration via inhibition of P-­gp transport in epithelial cell membranes of the kidney. Amiodarone and quinidine inhibit the renal and nonrenal clearance of digoxin, which increases the toxicity of digoxin (Hauptman and Kelly 1999; Anderson and Nawarskas 2001; Takara et al. 2002; Ledwitch and Roberts 2017). In cases of acute poisoning, hyperkalemia due to the inhibition of Na+-­K+-­ATPase, sinus arrhythmia, secondary hypotension, and sinoatrial arrest due to inhibition of its automaticity can occur. Various kinds of arrhythmias such as premature ventricular contractions and ventricular tachycardia with/without second-­degree or third-­degree AV block were observed due to the direct effect of increasing the automaticity of cardiomyocytes. Death may result from refractory ventricular fibrillation or asystole. Symptoms such as nausea, vomiting, anorexia, abdominal pain, weakness, confusion, delirium, psychosis, and color vision disturbance have been reported with digitalis poisoning. Diagnosis of poisoning is based on the history of prescription, various symptoms, and characteristic arrhythmias such as bidirectional tachycardia and junctional rhythm. Hyperkalemia (serum K+ > 5.5 mmol/L) may be a marker of severe acute toxicity. The concentration of digitalis is an aid for the diagnosis. However, since the postmortem blood concentration of digitalis is affected by postmortem redistribution from surrounding tissues, we should be careful when interpreting the postmortem concentration (O’Sullivan et al. 1995; Ferner 2008).

52.11.6 Other cardiovascular drugs Angiotensin converting enzyme inhibitors (ACEIs) The renin-­ angiotensin-­ aldosterone system regulates blood pressure. The renin, secreted from juxtaglomerular cells in the kidney, forms angiotensin I from angiotensinogen. Angiotensin

Organic nitrates Organic nitrates, such as nitroglycerin, isosorbide dinitrate, and isosorbide mononitrate, are used as vasodilators for the pharmacotherapy of ischemic heart diseases including anginal episodes. Those compounds are sources of nitric oxide. The activation of guanylate cyclase by nitric oxide increases intracellular cyclic guanosine monophosphate (cGMP). The cGMP causes the relaxation of smooth muscle cells and dilation of the coronary arteries and peripheral vessels (Michel 2006). Nitrates cause hypotension by peripheral vasodilation. As side effects of these compounds, headache, transient dizziness, and weakness have been reported. Nitrates may be converted to nitrites in the gastrointestinal tract. Since nitrites are oxidizing agents, methemoglobinemia may occur through the oxidation of hemoglobin, especially in infants. Because infants have lower enzyme activity of NADH-­ cytochrome-­b5 reductase, an enzyme that converts methemoglobin to hemoglobin, infants have a risk for methemoglobinemia (Johnson 2019). We also have to consider drug interactions with phosphodiesterase-­5 (PDE5) inhibitors, such as sildenafil, tadalafil, and vardenafil. Those PDE5 inhibitors are prescribed for the treatment of erectile dysfunction. The combined use of a PDE5  inhibitor and nitrates may cause severe hypotension through enhancement of vasodilation (Michel 2006; Doumas et al. 2015).

52.11.7 Conclusions

Doumas, M., Lazaridis, A., Katsiki, N. et  al. (2015). PDE-­5  inhibitors: Clinical points. Current Drug Targets 16: 420–426. Drummer, O.H. (2008). Pharmacokinetics and metabolism. In: S. Jickells and A. Negrusz (eds.), Clarke’s Analytical Forensic Toxicology, pp. 13– 42, London: Pharmaceutical Press. Ferner, R.E. (2008). Post-­mortem clinical pharmacology. British Journal of Clinical Pharmacology 66: 430–443. Flaten, H.K. and Monte, A.A. (2017). The pharmacogenomic and metabolomic predictors of ACE inhibitor and angiotensin II receptor blocker effectiveness and safety. Cardiovascular Drugs and Therapy 31: 471–482. Gummin, D.D., Mowry, J.B., Spyker, D.A. et  al. (2019). 2018 annual report of the American Association of Poison Control Center’s National Poison Data System (NPDS): 36th annual report. Clinical Toxicology 57: 1220–1413. Hauptman, P.J. and Kelly, R.A. (1999). Digitalis. Circulation 99: 1265–1270. Inomata, N. (2012). Recent advances in drug-­induced angioedema. Allergology International 61: 545–557. Johnson, S.F. (2019). Methemoglobinemia: Infants at risk. Current Problems in Pediatric and Adolescent Health Care 49: 57–67. Kennedy, M.C. (2010). Post-­ mortem drug concentrations. Internal Medical Journal 40: 183–187. Kinoshita, H., Taniguchi, T., Nishiguchi, M. et  al. (2003). An autopsy case of combined drug intoxication involving verapamil, metoprolol and digoxin. Forensic Science International 133: 107–112. Kinoshita, H., Fuke, C., Nishiguchi, M. et al. (2009). A case of digoxin poisoning. Current Study of Environmental and Medical Sciences 2: 17–18. Laurent, S. (2017). Antihypertensive drugs. Pharmacological Research 124: 116–125. Ledwitch, K.V. and Ronerts, A.G. (2017). Cardiovascular ion channel inhibitor drug-­drug interactions with p-­glycoprotein. AAPS Journal 19: 409–420. Lip, G.Y. and Ferner, R.E. (1995). Poisoning with anti-­hypertensive drugs: Angiotensin converting enzyme inhibitors. Journal of Human Hypertension 9: 711–715. Lynch, T. and Price, A. (2007). The effect of cytochrome P450 metabolism on drug response, interactions, and adverse effects. American Family Physician 76: 391–396. Malde, B., Regalado, J., and Greenberger, P.A. (2007). Investigation of angioedema associated with the use of angiotensin-­converting enzyme inhibitors and angiotensin receptor blockers. Annals of Allergy, Asthma & Immunology 98: 57–63. Michel, T. (2006). Treatment of myocardial ischemia. In: L.L. Brunton, J.S. Lazo, and K.L. Parker (eds.), Goodman & Gilman’s The Pharmacological Basis of Therapeutics, 11th edn, pp. 823–844, New York: McGraw Hill. Moffat, A.C., Osselton, M.D., Widdop, B. (eds.) (2004). Clarke’s Analysis of Drug and Poisons in Pharmaceuticals, Body Fluids and Postmortem Materials, 3rd edn. London, Pharmaceutical Press. Musshoff, F., Padosch, S., Steinborm, S. et al. (2004). Fatal blood and tissue concentrations of more than 200 drugs. Forensic Science International 142: 161–210. O’Sullivan, J.J., McCarthy, P.T., and Wren, C. (1995). Differences in ­amiodarone, digoxin, flecainide and sotalol concentrations between

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Anderson, A.R. and Nawarskas, J.J. (2001). Cardiovascular drug-­drug interactions. Cardiology Clinics 19: 215–234. Aonuma, K., Shiga, T., and Atarashi, H., et  al. (2017). Guidelines for therapeutic drug monitoring of cardiovascular drugs clinical use of blood drug concentration monitoring (JCS2015) -­ digest version-­ . Circulation Journal 81: 581–612. Arroyo, A.M. and Kao, L.W. (2009). Calcium channel blocker toxicity. Pediatric Emergency Care 25: 532–541. Ashraf, M., Chaudhary, K., Nelson, J. et al. (1995). Massive overdose of sustained-­release verapamil: a case report and review of literature. American Journal of the Medical Sciences 310: 258–263. Bailey, D.G., Arnold, M.O., and Spence, J.D. (1998). Grapefruit juice-­drug interaction. British Journal of Clinical Pharmacology 46: 101–110. Baselt, R.C. (2017). Disposition of Toxic Drugs and Chemicals in Man, 11th edn. Seal Beach, CA: Biochemical Publications. Brodde, O-­E. and Michel, M.C. (1999). Adrenergic and muscarinic receptors in the human heart. Pharmacol Rev 51, 651–690. Brodde, O-­E., Bruck, H., and Leineweber, K. (2006). Cardiac adrenoceptors: physiological and pathophysiological relevance. Journal of Pharmacological Sciences 100: 323–337. Brown, T., Gonzalez, J., and Monteleone, C. (2017). Angiotensin-­ converting enzyme inhibitor-­induced angioedema: A review of the literature. Journal of Clinical Hypertension 19: 1377–1382. Burchell, H.B. (1983). Digitalis poisoning: Historical and forensic aspects. Journals of the American College of Cardiology 1: 506–516. Christensen, M.B., Petersen, K.M., Bøgevig, S. et al. (2018). Outcomes following calcium channel blocker exposures reported to a poison information center. BMC Pharmacology and Toxicology 19: 78. DeRoos, F.J. (2011). Calcium channel blockers. In: L.S. Nelson, N.A. Lewin, and M.A. Howland, et  al. (eds.), Goldfrank’s Toxicologic Emergencies, 9th edn, pp. 884–892, New York: McGraw Hill. DeWitt, C.R. and Waksman, J.C. (2004). Pharmacology, pathophysiology and management of calcium channel blocker and ß-­blocker toxicity. Toxicological Reviews 23: 223–238.

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References

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Cardiovascular drugs are a heterogeneous group with a variety of mechanisms of action. They cause various manifestations such as arrhythmias and hypotension in overdose cases, and they sometimes cause a fatal outcome. Their toxicity is immediately life-­ threatening, and the diagnosis of poisoning depends on the concentration in the blood. However, we have to consider various factors, such as drug-­drug interactions for antemortem specimens and sampling site and postmortem redistribution for postmortem specimens for the proper interpretation of drug concentrations. Diagnosis of poisoning is based on not only postmortem blood concentrations, but also the integration of information about other clinical conditions and circumstances prior to death (Kennedy 2010). Detailed investigation with collaboration between the pathologist and toxicologist is essential for proper diagnosis.

TOXICOLOGY

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encountered in emergency medical room and forensic practices. Thus, its wide availability and severe toxicities have placed APAP overdose as the leading cause for calls to Poison Control Centers in the United States (>100 000/year). Moreover, paracetamol/ acetaminophen overdose accounts for more than 56 000 emergency room visits, 2600 hospitalizations, and an estimated 458 deaths due to acute liver failure each year in the United States alone (Lee 2004). At the Japan Poisoning Center, about 6% of reports on drug poisoning over the past five years were related to paracetamol/acetaminophen poisoning, and in 2012, out of a total of 10 480 cases, 628  were inquiries about paracetamol/ acetaminophen. The most common paracetamol/acetaminophen content per tablet of the common cold medicines sold in Japan was as small as 100 mg or 200 mg. However, in recent years, drugs containing 300 mg or more of paracetamol/acetaminophen per tablet have begun to be launched in Japan. Furthermore, in the United States, drugs containing paracetamol/acetaminophen in large volumes of 500  mg or more per tablet are sold as over-­the-­ counter drugs, and with the spread of the Internet, they are easily available in Japan. Under such circumstances, it is easily expected that the number of cases of poisoning due to large doses of acetaminophen will increase regardless of suicide or accident.

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antemortem serum and femoral postmortem blood. Human and Experimental Toxicology 14: 605–608. Park, H., Purnell, G.V., and Mirchandani, H.G. (1990). Suicide by captopril overdose. Journal of Clinical Toxicology 28: 379–382. Pelissier-­ Alicot, A-­ L., Gaulier, J-­ M., Champsaur, P. et  al. (2003). Mechanisms underlying postmortem redistribution of drugs: A review. Journal of Analytical Toxicology 27: 533–544. Scheen, A.J. (2011). Cytochrome P450-­mediated cardiovascular drug interactions. Expert Opinion on Drug Metabolism & Toxicology 7: 1065–1082. Schulz, M., Schmoldt, A., Andresen-­Streichert, H. et al. (2020). Revisited: Therapeutic and toxic blood concentrations of more than 1100 drugs and other xenobiotics. Critical Care 24: 195. Skopp, G. (2010). Postmortem toxicology. Forensic Science, Medicine, and Pathology 6: 314–325. Takara, K., Kakumoto. M., Tanigawara, Y. et  al. (2002). Interaction of digoxin with antihypertensive drugs via MDR1. Life Sciences 70: 1491–1500. Truitt, C.A., Brooks, D.E., Dommer, P. et al. (2012). Outcomes of unintentional beta-­blocker or calcium channel blocker overdoses: a retrospective review of poison center data. Journal of Medical Toxicology 8: 135–139. Vucinić, S., Vicinić, Z., Segrt Z. et al. (2003). Acute poisoning with cardiovascular agents. Vojnosanitetski Pregled 60: 691–696. Waeber, B., Nussberger, J., and Brunner, H.R. (1984). Self poisoning with enalapril. British Medical Journal 288: 287–288. Yoshitome, K., Miyaishi, S., Yamamoto, Y. et  al. (2008). Postmortem increase of flecainide level in cardiac blood. Journal of Analytical Toxicology 32: 451–453. Yoshitome, K., Ishizu, H., and Miyaishi, S. (2010). Postmortem acidification of blood/organs induces an increase in flecainide concentration in cardiac blood and the concentration of the lung to this increase. Journal of Analytical Toxicology 34: 26–31. Zanger, U.M. and Schwab, M. (2013). Cytochrome P450 enzymes in drug metabolism: regulation of gene expression, enzyme activities, and impact of genetic variation. Pharmacology & Therapeutics 138: 103–141.

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52.12 Paracetamol/Acetaminophen

52.12.1 Blood concentrations The average of plasma paracetamol/acetaminophen concentrations was 4.2 mg/L (range, 2.4-­6.4) at 6 hours when 24 healthy males with oral single ingestion of 324 mg paracetamol (Thomas et  al. 1972). An oral administration of 1000  mg to six healthy men resulted in the serum concentration of 9 mg/L in average at 1, 2, and 3 hours (Weikel, 1958). Thirty minutes after the ingestion of 1300  mg, scrum concentrations reached at from 4.8 to 13 mg/L in five healthy adults (Fletterick et al. 1979). One hour after an oral administration of 1800 mg to eight healthy adults, plasma concentrations averaged 26  mg/L (range, 5.6–52) (Prescott et al. 1968).

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Toshikazu Kondo

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Paracetamol is also known as acetaminophen, which is widely used as an active ingredient in over-­the-­counter antipyretic and antipyretic analgesics. In general, it is mainly given orally or transrectally, and intravenous injection is also available. Paracetamol/ acetaminophen is given in order to relieve mild-­to-­moderate pain and is effective for fever reduction in children. Moreover, the combination of paracetamol and opioid drugs is applied to control cancer pain and surgery-­ induced pain (Kaplowitz 2004; Whitcomb 1994). Currently, there are many types of antipyretic analgesics and common cold drugs containing paracetamol/acetaminophen, and, in principle, those drugs are easily available without a prescription at drug stores. As a result, paracetamol/acetaminophen poisoning due to suicide attempts or incorrect doses is often

52.12.2  Metabolism and toxicity In therapeutic usage, paracetamol/acetaminophen is excreted largely in the urine as various conjugates: 45–55% as a glucuronide conjugate, 20–30% as a sulfate, 15–55% as cysteine and mercapturic acid conjugates, and 4–5% as methoxy acetaminophen. Approximately 2% of a dose is excreted unchanged in the urine (McGill and Jaeschke 2013) (Figure  52.12.1). Paracetamol/­ acetaminophen does not have carcinogenic effects, but its hepatotoxicity is much higher than that of the same antipyretic analgesics, aminopyrin, and phenacetin. Paracetamol/acetaminophen itself is not toxic, and most of acetaminophen is excreted in the urine as several conjugates such as glucuronide, sulfate, cysteine, and mercapturic acid (James et al. 2003). Paracetamol/­

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TOXICOLOGY

NHCOCH3

NHCOCH3

Paracetamol /acetaminophen Glucuronide or sulfate conjugates Glucuronide or sulfate

OH

CYP1A2 CYP2E1 CYP3A4

NCOCH3

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Covalent binding Hepatocyte necrosis

N

NAPQI

O

O

NHCOCH3

OH

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Glutathione

U

Mercapturic acid conjugates

TO

NHCOCH3

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Glutathione conjugates

OH

SCH2CHCOOH NHCOCH3

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Figure 52.12.1  Approximately 2% of a dose is excreted unchanged in the urine. Source: Based on McGill and Jaeschke (2013).

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a­cetaminophen-­induced toxicity is mediated by a toxic metabolite, N-­acetyl-­p-­benzoquinone imine (NAPQI) generated from paracetamol/acetaminophen. Conversion of paracetamol/acetaminophen into NAPQI is catalyzed by hepatic cytochrome P450 enzymes, particularly, CYP1A2, CYP2E1, and CYP3A4 (Brackett and Bloch 2000; Dahlin et al. 1984; Patten et al. 1993; Zaher et al. 1998). At therapeutic doses, NAPQI is usually detoxified by glutathione in hepatocytes followed by the excretion in urine. However, when acetaminophen is taken in large amounts, a proportionally large amount of NAPQI is produced, thus resulting in the depletion of glutathione for NAPQI detoxification. When the amount of glutathione falls below 20–30% of normal, NAPQI covalently binds to high-­molecular-­weight substances of cell proteins, eventually causing hepatocyte necrosis (James 2003). This is the basic mechanism of liver damage caused by high doses of acetaminophen (Figure 52.12.1). Hepatotoxicity of paracetamol/acetaminophen has occasionally been reported at lower-­than-­expected doses. In particular, paracetamol-­ induced hepatotoxicity is dependently on the amount of NAPQI, indicating that conditions enhancing CYP expression can exaggerate hepatotoxicity because of over-­ generation of CYP-­dependent NAPQI. The intrahepatic expression of CYPs is closely associated with hepatotoxicity of paracetamol/acetaminophen. The intake of paracetamol/­

Table 52.12.1  Representative drugs of CYPs CYP type

Inducer

CYP1A2

Omeprazole, dioxin, polycyclic aromatic hydrocarbon, carbamazepine, phenytoin

CYP2E1

Ethanol

CYP3A4

Phenobarbital, rifampicin, ritonavir, efavirenz, carbamazepine, phenytoin

acetaminophen with drugs that induce CYPs such as CYP1A2, CYP2E1, and CYP3A4 unexpectedly cause severe hepatic ­dysfunction. For examples, the intake of paracetamol with ethanol or phenobarbital would increase the risk of hepatic injury than expected (Table 52.12.1). Moreover, a recent study demonstrates that the herbal compound can also exacerbate paracetamol/acetaminophen-­induced liver failure (Britza et al. 2020). In addition to drugs, circadian rhythms have effects on intrahepatic CYP expression. Thus, when an identical dose of paracetamol is given, the difference of hepatotoxicity would be observed between day and night time. When paracetamol/acetaminophen is detected in patients or cadavers, forensic pathologists should consider the influence of intake time and other drugs including ethanol.

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CYPs

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Kupffer cell activation

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Glutathion depletion Covalent binding

NAPQI-SG

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Glutathione conjugate

Colorimetric methods are widely used in clinical laboratories (Liu and Oka, 1980). Gas chromatography or liquid chromatography is more available (Jeevanandam et  al. 1980; Feng et  al. 2009). In particular, liquid chromatography has the advantage for the measurement of the conjugated metabolites of acetaminophen in plasma, urine, and tissue homogenates (Jung and Zafar 1985; Colin et al. 1987). Paracetamol/acetaminophen is stable in plasma for 6 hours at room temperature and 2 weeks at -­20 °C (Feng et al. 2009).

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Paracetamol

N

Sterile inflammation

C

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Cytokines, chemokines, adhesion molecules

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Leukocyte accumulation

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52.12.5 Analyses

Hepatocyte

NAPQI

Currently, N-­acetylcysteine (NAC) is used as the only therapeutic agent in clinical practice. However, the efficacy of NAC is strongly associated with the passage of time after taking paracetamol/ acetaminophen. In other words, it is important to administer as early as possible (within 8 hours) after taking the drug, and the incidence of liver damage increases after 8 hours or more, and the efficacy is poor after 16 hours or more. However, in the field of emergency medical care, even if drug poisoning is suspected, the causative drug poison is unknown at the time of transportation, and it is not uncommon for it to take time to identify it. Therefore, in addition to NAC, effective treatment for paracetamol/acetaminophen poisoning is expected to be established even after a relatively long period of time.

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Paracetamol/acetaminophen hepatotoxicity is influenced further by several factors such as oxidative stress, inflammatory responses, cytokine production, and hepatocyte apoptosis (Ishida et al. 2002, 2004, 2006; Ishibe et al. 2009). These factors contribute synergistically to damage. Indeed, inflammatory cytokines, chemokines, and leukocytes have some detrimental roles in the pathogenesis of paracetamol/acetaminophen-­ induced liver injury. In paracetamol/acetaminophen-­induced liver injury, hepatocyte necrosis is induced by the large production of the degradation product NAPQI, and the subsequent inflammatory reaction is thought to exacerbate the liver injury. Chemical mediators such as cytokines and chemokines have a role as exacerbating factors or protecting factors in the inflammatory process (Figure 52.12.2).

52.12.4  Therapy for hepatotoxicity

N

52.12.3 Paracetamol/acetaminophen liver damage and cytokines/chemokines

iNOS, ROS

Liver damage

Figure 52.12.2  Chemical mediators such as cytokines and chemokines have a role as exacerbating factors or protecting factors in the inflammatory process.

References Brackett, C.C. and Bloch, J.D. (2000). Phenytoin as a possible cause of acetaminophen hepatotoxicity: case report and review of the literature. Pharmacotherapy 20 (2): 229–233. Britza, S.M., Musgrave, I.F., and Byard, R.W. (2020). Paracetamol (acetaminophen) hepatotoxicity increases in the presence of an added herbal compound. Legal Medicine (Tokyo, Japan) 47: 101740. https:// doi.org/10.1016/j.legalmed.2020.101740 Colin, P., Sirois, G., and Chakrabarti, S. (1987). Rapid high-­performance liquid chromatographic assay of acetaminophen in serum and tissue homogenates. Journal of Chromatography 413: 151–160. Dahlin, D.C., Miwa, G.T., Lu, A.Y., and Nelson, S.D. (1984). N-­acetyl-­p-­ benzoquinone imine: a cytochrome P-­450-­mediated oxidation product of acetaminophen. Proceedings of the National Academy of Sciences of the United States of America 81 (5): 1327–1331. Feng, S., Tian, Y., Zhang, Z. et al. (2009). Rapid simultaneous determination of paracetamol, amantadine hydrochloride, caffeine and chlorpheniramine maleate in human plasma by liquid chromatography/tandem mass spectrometry. Arzneimittel-­Forschung 59 (2): 86–95. Fletterick, C.G., Grove, T.H., and Hohnadel, D.C. (1979). Liquid-­ chromatographic determination of acetaminophen in serum. Clinical Chemistry 25 (3): 409–412.

52.13 Interpretation of Postmortem Forensic Toxicological Results Henrik Druid and Gisela Skopp

52.13.1 General considerations

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Postmortem toxicological analyses are today extensively performed on autopsy samples collected by forensic pathologists. The continuous increase in the availability and use of pharmaceuticals and illicit drugs, and the improved sensitivity of the analytical methods imply that it is likely that more cases will be tested, and that more drugs will be detected in each case. The rationale for postmortem drug testing is primarily to confirm a suspicion of fatal intoxication or to rule out intoxication or influence of drugs, even though the result may not always be conclusive. Sometimes, the question may be if a particular compound is present or not, or if the drug is present in a concentration that reasonably matches a prescribed dose regimen or a certain alleged intake. The forensic toxicology laboratories have the task to identify and quantify drugs in the submitted samples, but some forensic toxicologists also provide an interpretation of the results. However, most commonly the forensic pathologist has the responsibility to consider all findings of the postmortem examination, including the toxicological results, in order to determine the cause and manner of death. It is often stated that fatal intoxication is a diagnosis of exclusion. This designation has probably begun and maintained to be used by pathologists who are trained to rely on morphological changes in the tissues. In most intoxications, the morphological changes are minimal and unspecific, or absent. However, in many cases of intoxications, there are several observations and findings which together form a line of evidence. In the death investigation of possible intoxications, the toxicologist has a key role by having a profound expertise in pharmacology and drug toxicity, and in particular, a good knowledge of the drug panorama observed at the laboratory. The pathologist, on the other hand, will have all details of the case, including the circumstances surrounding death as reported by the police, autopsy findings (at a growing number of departments CT-­scans and even MRI images may be available), microscopy findings, and sometimes results of supplementary clinical chemistry analyses. Therefore, many forensic pathologists and forensic toxicologists today have established a more active dialogue regarding the interpretation of the toxicology results. Forensic toxicology laboratories worldwide typically participate, and perform well, in several quality assurance programs, where their performance is examined. Although methodological protocols and instrumentation vary between laboratories, most forensic toxicological analytical results will provide reliable information of the content of the sample in the test tube. The possible errors are rather dependent on changes in the body before the sampling, and sample types and sampling techniques, i.e. pre-­ analytical factors. Linnet et al. (2008) compared results obtained

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Ishida, Y., Kondo, T., Ohshima, T. et al. (2002). A pivotal involvement of IFN-­gamma in the pathogenesis of acetaminophen-­induced acute liver injury. FASEB Journal 16 (10): 1227–1236. Ishida, Y., Kondo, T., Tsuneyama, K. et al. (2004). The pathogenic roles of tumor necrosis factor receptor p55  in acetaminophen-­induced liver injury in mice. Journal of Leukocyte Biology 75 (1): 59–67. Ishida, Y., Kondo, T., Kimura, A. et al. (2006). Opposite roles of neutrophils and macrophages in the pathogenesis of acetaminophen-­induced acute liver injury. European Journal of Immunology 36 (4): 1028–1038. Ishibe, T., Kimura, A., Ishida, Y. et al. (2009). Reduced acetaminophen-­ induced liver injury in mice by genetic disruption of IL-­1 receptor antagonist. Laboratory Investigation 89 (1): 68–79. James, L.P., Mayeux, P.R., and Hinson, J.A. (2003). Acetaminophen-­ induced hepatotoxicity. Drug Metabolism and Disposition 31 (12): 1499–1506. Jeevanandam, M., Novic, B., Savich, R., and Wagman, E. (1980). Serum acetaminophen assay using activated charcoal adsorption and gas chromatography without derivatization. Journal of Analytical Toxicology 4 (3): 124–126. Jung, D. and Zafar, N.U. (1985). Micro high-­performance liquid chromatographic assay of acetaminophen and its major metabolites in plasma and urine. Journal of Chromatography 339 (1): 198–202. Kaplowitz, N. (2004). Acetaminophen hepatoxicity: What do we know, what don’t we know, and what do we do next? Hepatology  40 (1): 23–26. Lee, W.M. (2004). Acetaminophen and the U.S. Acute Liver Failure Study Group: Lowering the risks of hepatic failure. Hepatology 40 (1): 6–9. Liu, T.Z. and Oka, K.H. (1980). Spectrophotometric screening method for acetaminophen in serum and plasma. Clinical Chemistry 26 (1): 69–71. McGill, M.R. and Jaeschke, H. (2013). Metabolism and disposition of acetaminophen: recent advances in relation to hepatotoxicity and diagnosis. Pharmaceutical Research 30 (9): 2174–2187 Patten, C.J., Thomas, P.E., Guy, R.L. et  al. (1993). Cytochrome P450 enzymes involved in acetaminophen activation by rat and human liver microsomes and their kinetics. Chemical Research in Toxicology 6 (4): 511–518. Prescott, L.F., Sansur, M., Levin, W., and Conney, A.H. (1968). The comparative metabolism of phenacetin and N-­acetyl-­p-­aminophenol in man, with particular reference to effects on the kidney. Clinical Pharmacology and Therapeutics 9 (5): 605–614. Thomas, B.H., Coldwell, B.B., Zeitz, W., and Solomonraj, G. (1972). Effect of aspirin, caffeine, and codeine on the metabolism of phenacetin and acetaminophen. Clinical Pharmacology and Therapeutics 13 (6): 906–910. Weikel, J.H., Jr. (1958). A comparison of human serum levels of acetylsalicylic acid, salicylamide and N-­acetyl-­p-­aminophenol following oral administration. Journal of the American Pharmaceutical Association. American Pharmaceutical Association 47 (7): 477–479. Whitcomb, D.C. (1994). Acetaminophen poisoning and liver function. The New England Journal of Medicine 331 (19): 1311–1312. Zaher, H., Buters, J.T., Ward, J.M. et al. (1998). Protection against acetaminophen toxicity in CYP1A2 and CYP2E1 double-­ null mice. Toxicology and Applied Pharmacology, 152 (1): 193–199.

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The pharmacology and toxicology, including pharmacokinetics, pharmacodynamics, and toxic effects, of select drugs and drug groups are treated in Chapter  51. Although virtually any substance may be harmful at very high concentrations, in practice there is quite a difference in toxicity between drugs; for instance, antibiotics are rarely found to be causing death in contrast to fentanyl derivatives, where the mere presence in postmortem blood is highly associated with a fatal poisoning. The frequency of a particular drug encountered in fatal intoxications does not necessarily reflect the toxicity of the drug, since the availability and “popularity” as an intoxicant will also affect its occurrence in postmortem samples. Moreover, certain drugs are used extensively in the population and are frequently detected postmortem, but predominantly at non-­toxic concentrations. This means that care must be taken to identify the particular drug(s) responsible for the intoxication. The toxicity of a drug may be measured by its median lethal dose 50 (LD50). Even if this information is available for most substances encountered in the postmortem setting, its translation into fatal postmortem blood concentration is not straightforward. On the other hand, information about the therapeutic window and therapeutic index of a drug provided by the manufacturer and national drug control agencies may give a clue as to which drugs may be more harmful than others. This information is classically obtained from standardized animal experiments, but even more helpful for the appreciation of a drug´s toxicity is certainty safety factor [TD1/ED99], i.e. the toxic dose that 1% of the animals display and the effective dose that 99% of the animals show, respectively. The drawback with this and similar measures is that the toxic effect might not relate to the fatality risk, which may be rather caused by a different mechanism. So, in practice, forensic pathologists and toxicologists learn from each other primarily from reading case reports describing certified fatal intoxications by a new drug, and later on usually case series. On the other hand, uncertainties may remain for quite some time regarding drugs that we do detect, but about which there are no reported fatal poisonings. If and when such a fatality is reported, then in retrospect we may realize that this particular drug may have caused or contributed to death in one or more cases previously examined. The therapeutic window, or safety, of a drug may be different for various drug groups, i.e. that the serious side effects and risk

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52.13.2  Drug toxicity

for toxicity of a drug may be accepted for a drug that may be important to treat critical conditions, but not accepted for other drugs, particularly not for over-­the-­counter drugs. Propoxyphene was a widely prescribed painkiller, approved by the FDA already in 1957, but which became increasingly more popular during the 1980s and 1990s. Because of abuse, governmental drug safety authorities in several countries decided to direct warnings to the doctors for prescribing propoxyphene to known or suspect abusers, and eventually the manufacturers stopped the production. The latter decision was mainly based on the observation that a remarkably high proportion of the fatal poisonings were accidental, suggesting a too low therapeutic window. Similarly, remoxipride, which was introduced as a promising new antipsychotic by AstraZeneca 1990, was withdrawn from the market in 1993 due to reported blood dyscrasias. But while it was used, forensic pathologists observed that its mere presence in blood was associated with death by suicide. Hypothetically, this might be analogous to the disinhibition conveyed by tricyclic antidepressants before their antidepressant effect appear, so even if most of these suicides were not caused by a fatal intoxication by the drug per se, remoxipride was suspected to indirectly be responsible for some of these deaths. Drug groups which are commonly involved in fatal intoxications are mainly psychoactive drugs and heart medications. Hence analgesics, sedatives, hypnotics, antidepressants, antipsychotics, beta-­blockers, and calcium channel blockers dominate the pharmaceutical drugs involved in fatal intoxications. During the last two decades, fatalities due to drugs of abuse (which may include classified pharmaceutical drugs) have steadily increased whereas intoxications by non-­abused pharmaceutical drugs have stabilized or even decreased. As mentioned above, propoxyphene was removed from the market, and then the question was what drugs should take its place. The same gap between regular painkillers and stronger opioids occurred when pentazocine (although being a weak opioid) was withdrawn from the market. The death statistics support an overall reduction of pharmaceutical drug intoxications, whereas there has been a gradual increase in the number of fatal intoxications with opioids, although most of these opioid deaths are seemingly caused by non-­authorized accessed opioid drugs. Some drugs frequently detected in postmortem blood may not be particularly toxic, but rather represent incidental findings. For instance, cannabis is considered to be quite non-­toxic, but synthetic cannabinoids (so-­called spice) have been reported to cause deaths. Further, both cannabis and spice smoking occasionally results in impulsivity and indirectly leads to death e.g. by suicidal or accidental jump from a height. Having said that, low levels of tetrahydrocannabinol, caffeine, theophylline, paracetamol, acetyl salicylic acid, ibuprofen, diclofenac, and carbamazepine are rarely of any significance in a suspected poisoning. Regarding carbamazepine, low levels may on the contrary be a result of drug non-­ adherence, and might be an explanation for a sudden unexpected death in epilepsy (SUDEP). The same is true for very low levels of anti-­hypertensive drugs and anti-­diabetics if a fatal brain hemorrhage and hyperglycemia is found. Finally, some drugs may not

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by separate analysis of methadone in samples from the right and left femoral vein. They found that the total measurement uncertainty exceeded the analytical uncertainty severalfold due to pre-­ analytical factors. Hence, the forensic pathologists and forensic toxicologists will have to consider a number of factors when reviewing the toxicological results and making the diagnosis fatal intoxication, or excluding this alternative. Below, the principles for interpretation of postmortem toxicological results and various factors influencing the interpretation are described in some detail.

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The interpretation of postmortem toxicological results remains an important, but often difficult, task for the forensic pathologists and forensic toxicologists. In order to determine if a particular drug may have caused or contributed to the death, information about the general properties of the drug, in particular the toxicity (see section  52.13.2), should be gathered and reviewed if not well known. The second issue is the blood concentration, is it high? To assess this, reference data are necessary. Baselt´s “Disposition of toxic drugs and chemicals in man” has now appeared in its 12th edition (2020), and over the years the various editions of this book have been extensively used by forensic toxicologists since it provides literature information about drug properties, and in particular therapeutic plasma or serum concentrations, as well as reported toxic and fatal blood concentrations, if available, for most drugs that may occur in the postmortem setting. Another guide with exhaustive information about drug concentrations that can assist in the interpretation of toxicological analytical results is the publications by Schulz and Schmoldt and co-­workers, recently updated (Schulz et al. 2020). Long before that, a similar compilation by Stead and Moffat (1983) was extensively used by forensic toxicologists. Whereas such compilations are very valuable for clinicians in suspected or obvious intoxications, most of the data are based on analysis of samples collected from living subjects and there are no details regarding how the reports of postmortem fatal levels, if available, were selected and assessed. As will be discussed in some detail, in section 52.13.7 there are a number of factors that make postmortem drug concentrations different from those in living subjects. Hence, a more relevant reference information is the concentrations found in deceased subjects where the key substance is detected but the person did not die of an intoxication. Such data cannot be generated to any larger extent unless toxicology is liberally performed in non-­ intoxication cases examined at forensic pathology departments. The other requirement is that both forensic pathology data, including cause of death, can be merged with the toxicological results. In order to generate data for a sufficient number of cases without substantial manual efforts, computerized systems are necessary. In Sweden, all these requirements were met when data management systems for forensic pathology and toxicology were developed (Druid et al. 1996) and implemented at the same time as all routine case work in forensic medicine, toxicology, genetics, and diagnostic psychiatry were merged into a national authority in 1991. The data management systems were designed to work as real-­time databases that could cross-­talk, allowing for automated selection and compilation of combined forensic pathology and

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52.13.3 Interpretation of postmortem drug concentrations

toxicology data for all drugs of interest. Hence, a compilation of postmortem femoral blood concentrations of 83 drugs based on such data was first published (Druid and Holmgren 1997). Since then, additional compilations, focusing on certain drug groups, have also been published using the same strategy (Reis et al. 2007; Jonsson et al. 2014; Söderberg et al. 2016). The basic procedure for the selection, assessment, and compilation of the data for intoxications and postmortem control cases of a particular drug is displayed in Figure 52.13.1. The strategy divides fatal intoxications into cases where the particular drug was considered to solely have caused the death (A-­cases) and cases where the drug may have contributed to a fatal intoxication (B-­cases). The B-­cases might either be considered to have acted in concert with another drug, but a case may alternatively be a B-­case because of its presence in high concentration along with another drug in high concentration, regardless of whether the mechanism of toxicity is similar or different. The postmortem control cases are perhaps the most important cases since they will tell you at what concentrations the particular drug may occur in postmortem femoral blood without causing an incapacitation of the subject immediately before death. The suicidal hangings and accidental deaths of motor vehicle drivers are some examples of such cases. However, a back-­seat passenger dying in a car crash or a person found drowned in a bathtub is not suitable as controls since you cannot tell if they had been incapacitated by the drug detected in the postmortem blood prior to the incident. The same is true for victims with severe blunt trauma to the chest or abdomen if there are injuries to the esophagus, the ventricle or the liver since transfer to the still circulating blood of high concentrations of the drug may have occurred before complete cardiac arrest. When compiling the results, there will always be some overlap in the concentrations of intoxication cases and controls, but their distributions are almost always significantly different (Figure 52.13.2). This strategy for collecting and analyzing peripheral blood samples from postmortem control cases for comparing their

SE

cause death immediately, but by chronic exposure, e.g. carcinogens, and such compounds are typically not included in the toxicological analysis, and the association between such exposures and death are rarely a matter for the death investigation.

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1304

Rough selection – ICD–9 codes

Manual assessment C-cases

DUI levels TDM levels

Manual assessment A, B cases Comparison observers

Check select cases – database

Check select cases – original files Figure 52.13.1  Strategy for semi-­automated selection and manual perusal and assessment of toxicological and pathological findings in fatal intoxications and postmortem control cases. DUI = driving under the influence cases; TDM = therapeutic drug monitoring cases.

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CHAPTER 52   Toxicology of Specific Substances

Amitriptyline

Citalopram

***

Clomipramine

***

40

80

30

60

20 7

40 25

30 20

***

20

4

5

20

μg/g

μg/g

2

10

1

***

15

1 0

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0

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Figure 52.13.2  Postmortem femoral blood concentrations of three antidepressant drugs. A = single drug intoxication, B = intoxication with two or more drugs, C = postmortem control cases, T = therapeutic drug monitoring cases (through values in steady state). The median concentrations in the intoxication groups are significantly higher than the controls (not indicated). The concentration distribution patterns are fairly representative for most psychoactive drugs.

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l­evels with those found in suspected intoxication cases have in recent years been adopted by several more investigators (e.g. Skov et al. 2015; Alvarez et al. 2020; Lefrancois et al. 2021) so over time there will be much more postmortem reference levels for peripheral blood available to be used in the routine casework, reducing the limitations when translating serum or plasma concentrations in samples from living subjects. Meanwhile, compilation and analysis of large datasets of postmortem blood concentrations may be valuable as a reference, particularly when combined with information about the particular drugs´ propensity for postmortem redistribution (Launiainen and Ojanpera 2014; Ketola and Ojanpera 2019). The limitation is that the toxicity of the drug must be considered, since even if the concentration you observe in your blood sample is very high, it might still not be significant if the drug has a low toxicity. It should be noted that the reference concentrations discussed here refer to postmortem femoral blood concentrations, and that these may be applicable to other peripheral blood samples, but can be different from those found in central blood, i.e. cardiac blood, or blood collected from the large vessels. If femoral blood is lacking due to blood loss or decomposition, the pathologist may only obtain central blood or soft tissues for analysis. In such situations, postmortem drug redistribution should be considered. In their pioneering work, Prouty and Anderson (1990) compared the concentrations of a number of drugs in central and peripheral drugs in a large series of cases. Later on, Pounder and co-­ workers carried out several in-­ depth studies (Jones and Pounder, 1987; Pounder et al. 1990, 1996) and named the phenomenon of postmortem redistribution “a toxicological night-

mare” (Pounder and Jones, 1990). Experimental studies have also been conducted to assess the propensity of drugs for postmortem redistribution and such results have been compared to postmortem drug redistribution in postmortem human samples (Hilberg et al. 1999). Further on, additional studies have been published which also provides suggested factors that may be used in order to translate central blood and tissue concentrations into what could be expected in peripheral blood, and which can be used for guidance. The reason for using blood for analysis is that this is typically used in the clinical settling, since samples from soft tissues cannot be obtained. Hence, it might seem strange that samples from the target organs where the particular drug exerts its effect, such as the heart or the brain, are not analyzed, since these tissues are accessible postmortem. There are actually some studies that have addressed this issue (see e.g. by Mikkelsen et al. 2016, 2018; Skov et al. 2015; and Nedahl et al. 2018), but the problem is to build up a sufficiently large body of reference data for such specimens. Furthermore, such an approach requires careful sampling since drug concentrations may vary between different regions of a tissue. In Figure 52.13.3, an example of postmortem femoral blood concentrations in an acute suicidal intoxication are shown. This middle-­aged man was found dead in his apartment, lying on the floor in a prone position. Compared to postmortem femoral blood reference concentrations reported in several publications, the concentrations of zolpidem and tramadol are much higher than those seen in postmortem control cases, and match levels seen in fatal intoxications with each drug. In the apartment, empty blister packs of zopiclone and tramadol were found, as well

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Figure 52.13.3  Toxicological findings in femoral blood from a middle-­aged man who committed suicide by ingestion of a large amount of pills. For details, see text.

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as a suicide note. There were no major signs of diseases observed at autopsy. Most forensic pathologists and toxicologist will likely consider the cause of death to be a combined intoxication with zolpidem and tramadol. While this may be true, it cannot be excluded that the death was caused only by one of the drugs, whereas the other has not yet reached the fatal concentration for this individual. In this case, however, both drugs are central depressants, hence it is likely that they have produced the toxicity in concert (see section 52.13.5 for further discussion). The other drugs should not be considered as contributory, since they are present in concentrations that typically are seen in subjects who have not been incapacitated by drugs immediately before their death by other causes. The ethanol level is not impressive, but it is possible that alcohol may have contributed to the toxicity, particularly if the man did not drink much alcohol regularly. For the sake of clarity, the drug levels shown here are in μg/g, which is because the sample weight has been recorded instead of the volume. This is practiced by some toxicology laboratories to avoid problems with inhomogeneous samples, whereas others measure the volume. However, when compensating for the density of blood, the differences between the units are non-­significant. Sometimes exceptionally high concentrations are found in cases, where the cause of death is obviously not due to an intoxication. The pathologist is recommended to talk to the responsible forensic toxicologist, who may check the chromatogram and other documents, and in some cases, a re-­analysis might be justified. However, if everything has been done according to the book, we have to accept that certain subjects obviously can tolerate high drug levels without becoming incapacitated. For instance, it is fairly well known among experienced anesthesiologists that red-­ haired individuals may require much higher doses to induce sleep, but there are only a few controlled studies on this (Liem et al. 2004). Conversely, in clear-­cut cases, the pathologist should not shrink from making the diagnosis of fatal intoxication if the blood drug levels are low, but in such instances, a careful review of all possible competing causes of death is important. Reference drug levels are rather intended to assist when the circumstances

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and findings are not clear-­cut. Finally, in certain cases, in which there is a strong suspicion of a fatal intoxication, the toxicology comes out negative. Then the forensic pathologist and toxicologist need to discuss in-­depth. The drug screens used at most laboratories are today quite comprehensive, but certain drugs may go undetected. It therefore may be necessary to run a number of additional toxicological analysis in order to find a drug that could be responsible for the death. However, despite such efforts, the supplementary analyses may also be negative, and if there are no other significant findings at autopsy or during histopathological examination, the pathologist may eventually state that the cause of death could not be certified. In this section, the focus has been on intoxication with pharmaceutical drugs. Regarding opioids, please see section  52.13.6. Concerning other illicit drugs, psychostimulants such as cocaine and amphetamines are also abused, and in many countries, psychostimulants predominate over opioids in terms of popularity among drug addicts. However, psychostimulants cause death much more infrequently. Cocaine is considered more toxic than methamphetamine and definitely more toxic than amphetamine, and may cause a rapid death if high amounts are consumed, although the mechanisms are not well understood. If the person is drinking substantial amounts of alcohol together with cocaine, cocaethylene, which has severalfold higher toxicity than cocaine, will be formed and may reach fatal concentrations. High cocaine intake may also increase the risk for myocardial infarction, or generalized myocardial ischemia when the myocardial demand exceeds the blood supply of oxygen and nutrients. Common for all psychostimulants is their potential to produce very high blood pressure that can cause intracerebral hemorrhage, so if this is found during the postmortem examination of a young person, toxicological analysis of psychostimulants should be performed if not included in the regular toxicology panel. Finally, cocaine and some amphetamine derivatives may occasionally cause excited delirium, which may develop rapidly, and since it, in addition to hallucinations and paranoia, also typically produces aggressiveness and combativeness, these persons may end up in a fight with strangers, guards, paramedics, and the police. Such deaths almost always cause headlines, and an exhaustive death investigation is warranted. The pathobiology behind this condition is considered to be a failure to upregulate the dopamine transport protein. Neuropharmacological methods are available to examine this, and in addition, upregulation of heat shock protein 70 in the temporal lobe cortex can give support for significant hyperthermia that most subjects develop and which sometimes may be the immediate cause of death (Mash et al. 2009).

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52.13.4 Metabolites The postmortem reference levels discussed in section 52.13.3 are based on the parent compound. This is partly explained by the absence of methods to analytically detect many metabolites in the past, and partly because fewer active metabolites were known. Today, more active metabolites can be detected with improved ana-

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N-­acetyl-­p-­benzoquinone imine (NAPQI). However, at autopsy, these cases usually do not cause any trouble to diagnose given the striking structural changes in the liver. Rather, the question may concern if paracetamol has been involved in a death that occurred shortly after a multiple drug intake, when the metabolite had not yet been formed in any significant amounts. Even in cases where the postmortem blood concentrations of paracetamol exceed 100 μg/g, it is difficult to prove that paracetamol contributed to death if this had a rapid course since the parent drug has not been shown to produce severe acute toxicity. However, high levels of paracetamol in the blood might be an indicator of an acute overdose, and suggest that the concentrations of other, more toxic drugs found also represent a high acute intake. Gamma-­hydroxy-­butyrate is a small compound which share many effects with alcohol, but with a short half-­life. For the user, this means that he or she has to take the drug repeatedly, every other hour or so, in order to keep the desired euphoric effects. Typically, the GHB is prepared as a transparent liquid which is consumed by filling the cap of a PET bottle and drinking such a small amount, but frequently. The concentration of GHB in the liquid may vary, and hence some users can become severely intoxicated if the preparation was strong, and may even end up at an emergency room, or die (Zvosec et al. 2011). At the emergency room, the condition may be critical for a while, but those who survive will recover very quickly due to the rapid elimination. Since GHB is a classified substance in most countries, some abusers may instead use the precursors gamma-­butyrolactone (GBL) or butane-­diol (BD), both of which are transformed to GHB in the body. These compounds are difficult to classify since they are used extensively in industrial production of various chemicals, and in this way the abuser can avoid being charged with possession. The drawback is, however, that the user will have even more difficulties with the dosing. In the postmortem setting, the problem for the forensic pathologist and toxicologist is that we all have an endogenous production of GHB, and even if the levelsnever can reach those that kill, they may make it difficult to determine if GHB (or its precursors) had been consumed by the individual. Efforts to determine the compound that was ingested deduced from identification of metabolites is more common in clinical toxicological practice, e.g. when analyzing urine or hair samples from individuals suspected of petty drug offense (for review, see Welter-­Ludeke and Maurer 2016). This line of research is of course important when the parent compound is absent in blood, whereas it is less relevant in postmortem toxicology where the focus rather is on the compounds actually found in the blood. Still, this research has certainly contributed to our general understanding of metabolic pathways, particularly for new psychoactive substances (NPS), and given that some of them have active metabolites, it may be important to include these in the analytical methods. During the past decades, genetic factors impacting the effects of drugs have been increasingly studied, particularly in the development of personalized medicine. In forensic toxicology research, pharmacogenomic studies has evolved since the 1990s (Agrawal and Rennert 2012; Sajantila et al. 2010; Musshoff et al. 2010). At first, investigators were hoping that this line of research could

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lytical methods, and several of these may be of important for the toxicity. Having said that, for most drugs, the parent compound is still mainly responsible for fatal intoxications, and in order to obtain support for an acute high intake, the parent drug to metabolite concentration (P:M ratio), as suggested by Apple (1989) many years ago, is still useful. The steady state P:M ratio is however variable for different drugs; some being metabolized rapidly and others slowly before being eliminated by the kidneys. Hence, some drugs may in the steady state have a low metabolite concentration whereas the metabolite concentration may exceed the parent compound concentration during steady state. This pharmacokinetic information is fortunately well known for most drugs. The benzodiazepines that generates a 7-­ amino-­ metabolite should be treated differently. For nitrazepam, flunitrazepam, and clonazepam, the postmortem blood will typically contain much more of the 7-­amino-­metabolite due to rapid postmortem conversion; hence, to estimate the antemortem levels, the sum of the parent drug and its 7-­amino-­metabolite should be used to better estimate the antemortem level. The same is true for the hypnotic drug propiomazine and its main metabolite dihydropropiomazine. For drugs with an active metabolite, the interpretation is more difficult, and more studies focusing on the combined toxicity are needed to provide guidance. Such studies are difficult to carry out properly as long as both the parent compound and metabolite will appear together when the parent compound is administered. Hence, there is a need for evaluation of the toxicity of the metabolite with the absence of the parent compound. Even if this is done, the combined effect will still be difficult to assess since both the parent compound and metabolite may be binding to the same receptor, and the net effect of this is difficult to predict given that both compounds may saturate a larger amount of receptors, but also compete at the binding sites. Hence, if their efficacy upon receptor binding is significantly different, then the proportion of receptors occupied by each of them will determine the toxicological effects. In many situations, the intake of a particular drug detected in postmortem blood was neither an isolated acute intake, nor a long-­ term chronic intake, but the result of an irregular use, and this might result in intermediate P:M ratios, that may make the interpretation difficult. Even worse, a chronic intake may mask an acute ­ pioid mainintake of a high dose. Individuals participating in an o tenance program with methadone will typically show concentrations in postmortem femoral blood ranging 0­ .1-­0.3  μg/g, and if such subjects show levels exceeding 0.7 g/g or more, it usually means a high intake, even when differences in opioid tolerance is considered. The use of the P:M ratio of methadone and the metabolite 2-­ethylidene-­1,5-­dimethyl-­3,3-­diphenylpyrrolidine (EDDP) may be helpful to provide support for an acute intoxication, and the (R)/(S) enantiomer ratio can provide information as to if the raceenantiomer was consumed (Jantos and mate or only the (R)-­ Skopp 2013). Paracetamol (acetaminophen) is found very frequently in postmortem blood samples, but is rarely involved in acute intoxications. However, when taken in very large amounts liver damage may ensue, often several days later, caused by the metabolite

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example, half a pill of 1 g of flunitrazepam may induce sleep rapidly in aged subjects with insomnia problems, whereas an experienced abuser may ingest 10 tablets and get a high, often in combination with excitation and agitation. In contrast, the same dose to a non-­experienced user may result in a fatal intoxication (Druid et al. 2001). The paradoxical effect of flunitrazepam at high doses has not yet been clarified; one possibility is that the subjects reacting with hyperactivity might have in part conserved the neonatal excitatory response of the GABA-­receptor, but activation of other receptor systems at high doses can also be considered. The common notion that the addition of one drug will potentiate the toxicity caused by another drug is not always true. For instance, if naloxone is co-­ingested with morphine, it is likely to rather reduce the toxic effect of morphine. Further, suppose there is a “fairly” high concentration of both propranolol and zopiclone in postmortem blood, it is tempting to state that both drugs did not match the levels seen in fatal cases, but that they might have acted in concert. However, if propranolol causes death, it will be by causing a lethal arrhythmia, whereas zopiclone is a central depressant, acting on the brain. It is thus important to consider the pharmacological and toxicological properties of the drugs involved before reaching a conclusion. Even then, and a combined effect of the same or similar effector systems are logical for two drugs, such as alprazolam and zopiclone, there is not much support for combined toxicity in the literature. This means that caution is needed when making the diagnosis multiple drug overdose in an obscure case in which there is no drug that is present in a reasonably high concentration. Some experimental studies have however been conducted to address the effect of addition of one drug on the toxicity of another. Borron et al. (2002) pretreated groups of rats that were then given morphine, methadone, or buprenorphine, and found that flunitrazepam increased the mortality among rats given methadone and buprenorphine, but not morphine. The same research group and a few others have published a limited number of papers in similar settings, but more surprises may show up when such studies are performed on various other drug groups, so it is advisable to use a humble attitude when dealing with cases in which there are several possible candidate drugs to explain a possible fatal intoxication. Finally, when a fatal intoxication is suspected but the blood concentrations of one or more drugs are only moderately elevated as compared to levels in postmortem controls, it may seem reasonable to look at different pathologies that could have made the person more sensitive to the drugs detected. For instance, moderately elevated levels of a calcium channel blocker, a beta blocker or a tricyclic antidepressant, intuitively could be expected to induce a lethal arrythmia in an enlarged heart with patchy or diffuse fibrosis than in a heart that is normal. However, there are no postmortem studies that support the notion that those with such heart conditions die of arrhythmogenic drugs at lower concentrations than individuals with a normal heart. If the examination of the heart reveals substantial pathologies, it might be rather more likely that an acute complication to the heart condition that is the cause of death. Hence, again, it is advisable to be cautious when the ­circumstantial

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explain for unexpected high or low postmortem blood drug concentrations, e.g. that the deceased person was a poor metabolizer of the responsible enzyme, and hence reached a high concentration of the parent drug, despite “normal” intake over a period of time, during which the drug could have accumulated. Conversely, subjects with an ultrahigh genetic profile might produce much more of a metabolite that is toxic. Even though there are quite a number of such cases reported in the literature, they seemingly make up only a very small proportion of the deaths examined at forensic pathology departments. Furthermore, there is not much evidence to support the possibility that subjects with a “defect” metabolic profile will die at a different blood drug concentration than those who are genetically wildtype. However, the determination of the genotype corresponding to relevant metabolic enzymes may be important for the interpretation of the manner of death, i.e. if the levels observed in postmortem blood had been achieved by a repeated intake by a poor metabolizer, supporting an unintentional intoxication, or if they have resulted from an intentional overdose as a means of suicide. Whereas genetic polymorphism of drug metabolizing enzymes may result in higher or lower blood drug concentrations, this may partly be true also for certain transporters, such as P-­glycoprotein (P-­gp). The effect of the various functions of this transporter is somewhat difficult to assess since it is involved in both the absorption of many drugs from the intestines, and also a key player in the transport of drugs across the blood-­brain barrier. For other transporters, the effect of such polymorphisms may impact mainly on drug concentrations in the areas surrounding synapses and might not affect the blood concentrations. In a wider sense, the effect, if any, may be considered to be pharmacodynamic. To better understand the interindividual variation of blood drug concentrations in fatal intoxications (and variation in control subjects), estimation of the function of the target receptor system is desired. However, even if many receptor polymorphisms of functional relevance have been identified for a number of receptor types, this field has not as yet been much studied in postmortem toxicology.

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52.13.5  Contributory drugs and diseases

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Opioids remain indispensable in the treatment of patients with severe pain due to their strong analgesic effect. However, they all have an abuse potential since they share euphoric effects, and they can cause respiratory depression that may be fatal. These effects are mediated by mu-­opioid receptors. Several opioids also act on kappa and delta opioid receptors, but these effects are subordinate to the effects conveyed by the mu-­opioid receptors. When the Swedish investigators pursued the evaluation of postmortem toxicological results in large series of cases, substance by substance, they experienced great problems when trying to apply the strategy on illicit drugs, particularly opioids. It was found that the morphine concentrations seen in postmortem controls, and in postmortem cases treated at a burn center overlapped the levels found in obvious heroin and morphine intoxications, see Figure 52.13.4. The explanation for the very high levels in patients with burns is that they require very high doses of morphine (or similar opioids) to relieve the pains. All those with a very high concentration were ventilated in a respirator, lending support for the notion that most opioid deaths are caused by respiratory depression. Most publications regarding heroin overdose deaths report mean postmortem blood concentrations in the region of 0.2-­0.3

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μg/g, whereas those who commit suicide with morphine pills typically have postmortem blood concentrations of morphine exceeding 1 μg/g. There are two possible explanations for this difference: due to higher lipophilicity, heroin will cross the blood-­ brain barrier and reach the respiratory control center more rapidly than morphine, and 6-­acetylmorphine may in practice exert effect on the mu-­opioid receptors. The latter is supported by the finding of higher 6-­acetylmorphine concentrations in the brain during the first 30  minutes after heroin administration, matching the effects on the rats treated (Morland et al. 2013). In addition, heroin is often administered intravenously, snorted or smoked, and these modes of administration will result in a more rapid distribution of the drug to the brain than oral ingestion of morphine. It is also likely that a sudden, extensive occupation of any receptor system will cause a strong response as compared to a gradual saturation that allows the cells to react with defense mechanisms such as receptor internalization, uncoupling, and production of modulatory microRNAs. As pointed out above, all opioids share euphoric, analgesic, and respiratory depressant effects, but they also produce a significant tolerance development upon repeated exposures, which in the clinical setting is a great problem since the dose has to be gradually increased to achieve pain relief. In the postmortem situation, the problem is that the opioid blood concentrations that are found in fatal intoxications may vary tenfold or more. Unfortunately, there is no reliable biomarker that can be used to estimate the degree of opioid tolerance. As a proxy, segmental hair analysis may be performed in order to obtain an idea of the recent use of opioids (Druid et al. 2007). Actually, most individuals who die has no opioids in the most recent segment of their hair (Figure 52.13.5), suggesting that the death was caused by an acute intake after a period of abstinence, that may have been due

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Figure 52.13.5  Heroin metabolites in femoral blood and hair collected from a fatal heroin intoxication. The hair segments are 5 mm each, corresponding to a growth period of approximately 2 weeks. The results give support for a loss of tolerance after previous heroin use.

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able in urine. After yet some time, only morphine is detected, but then the glucuronides are usually seen in higher amounts. Whereas there is some support for the contribution of 6-­acetylmorphine to the toxicity of heroin, it is still a matter of debate to what extent morphine-­6-­glucuronide contributes. If it does, it is probably more significant in delayed deaths. Morphine-­ 3-­glucuronide is formed in larger amounts, but not considered toxic. A clue as to if the detected morpine is due to heroin or morphine intake is the finding of acetylcodeine, a byproduct of heroin synthesis. Fentanyl is an extremely potent analgesic that is also abused. During a period of several years, fentanyl analogs were produced and sold, mainly over the Internet, to abusers in many countries, contributing to what has been named the Opioid crisis. The producers frequently modified the structure of the molecule to avoid classification. Many of these drugs were distributed as powders, and intermediate vendors then dissolved them in liquid and sold them in nasal sprays. This resulted in a large number of deaths due to the difficulties in dosing. The epidemic peaked in 2017, and the number of deaths rapidly dropped because the production was effectively stopped. Some of the fentanyl analogs were so potent that their mere detection in postmortem blood usually implied that it was the cause of death. Like heroin toxicity deaths, most of these victims presented with foam in the airways and significant lung edema. The blood concentrations were quite variable just like in other opioid deaths. Buprenorphine is a partial mu-­receptor agonist with potent analgesic properties, and is both used for pain relief, but also in opioid replacement programs. Like all opioids, the postmortem concentrations are variable, but surprisingly, when evaluating the concentrations in buprenorphine toxicity deaths and in postmortem cases with a different cause of death, Selden et  al. (2012) paradoxically observed that the median buprenorphine blood drug concentration was lower in the former group (0.8 vs. 2.7 μg/g). The explanation was that the majority of those who died of other causes had been using buprenorphine for a longer period of time and thus developed substantial tolerance. Hence the median

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to visit to an addiction center, hospitalization, other sickness, or inability to obtain drugs. The term “overdose” is extensively used, but in most cases, the victims have actually taken a regular dose (or even a lower dose) but miscalculated his/her tolerance. It has been suggested that the development and loss of tolerance to the analgesic, respiratory depressant and euphoric effect of opioids may follow different time courses, and hence a dose that produces euphoria might exceed the dose that kill (White and Irvine 1999). It is generally believed that opioids cause death by respiratory depression via overstimulation of mu-­ opioid receptors on interneurons in the respiratory control centers, particularly the pre-­Bötzinger complex, which in the human brain has been identified to be localized in the anterolateral part of medulla oblongata (Schwarzacher et al. 2011). However, most opioid overdose deaths present with lung edema and froth in the airways (Figure 52.13.6) and the edema might in many cases be so massive that it may be considered the cause of death. The reason for the development of this phenomenon remains obscure. Interestingly, intensive care doctors rarely find lung edema in opioid intoxication cases, so most likely it develops rapidly and those who contract it might not survive during transport to the hospital. Opioid deaths may thus have a rapid course, but a proportion of these deaths are delayed, i.e. occur one hour a more after intake. A frequent report by witnesses is that the person had felt dizzy after snorting, smoking, or injection, then laid down on a couch, snoring for some time, and eventually to be found lifeless and impossible to resuscitate. Several reviews of opioid toxicity deaths have reported that 40–50% of the deaths are delayed. In addition to circumstantial information, delayed deaths might also be identified by looking at the metabolic pattern in postmortem blood and tissues (Thaulow et  al. 2018). Heroin is rapidly degraded to 6-­acetylmorphine, and then to morphine. Morphine in turn is metabolized to morphine-­3-­glucuronide and morphine-­ 6-­glucuronide. In rapid deaths, 6-­acetyl-­morphine may be seen in postmortem blood, whereas the glucuronides may be present in very low concentrations or absent. At a little longer interval after intake, 6-­acetylmorphine may be absent in blood but detect-

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Figure 52.13.6  a–c Froth in the larynx and nostrils, and massive lung edema in three cases of acute opioid intoxication. N.B. that in some cases, froth may have been wiped off by first-­attenders or lost during intubation attempts by staff at the emergency department.

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Druid, H. and Holmgren, P. (1998). Compilations of therapeutic, toxic, and fatal concentrations of drugs. Journal of Toxicology: Clinical Toxicology 36 (1–2): 133–134 author reply 135–136. Druid, H., Holmgren, P., and Ahlner, J. (2001). Flunitrazepam: an evaluation of use, abuse and toxicity. Forensic Science International 122 (2– 3): 136–141. Druid, H., Holmgren, P., and Löwenhielm, P. (1996). Computer-­assisted systems for forensic pathology and forensic toxicology. Journal of Forensic Sciences 41 (5): 830–836. Druid, H., Strandberg, J.J., Alkass, K. et al. (2007). Evaluation of the role of abstinence in heroin overdose deaths using segmental hair analysis. Forensic Science International 168 (2–3): 223–226. Drummer, O.H. (2007). Post-­ mortem toxicology. Forensic Science International 165 (2–3): 199–203. Geile, J., Maas, A., Kraemer, M. et al. (2019). Fatal misuse of transdermal fentanyl patches. Forensic Science International 302: 109858. Gerostamoulos, D., Beyer, J., Staikos, V. et al. (2012). The effect of the postmortem interval on the redistribution of drugs: A comparison of mortuary admission and autopsy blood specimens. Forensic Science, Medicine, and Pathology 8 (4): 373–379. Hilberg, T., Rogde, S., and Morland, J. (1999). Postmortem drug redistribution-­-­human cases related to results in experimental animals. Journal of Forensic Sciences 44 (1): 3–9. Ingelman-­Sundberg, M., Sim, S.C., Gomez, A., and Rodriguez-­Antona, C. (2007). Influence of cytochrome P450 polymorphisms on drug therapies: Pharmacogenetic, pharmacoepigenetic and clinical aspects. Pharmacology & Therapeutics 116 (3): 496–526. Jantos, R. and Skopp, G. (2013). Postmortem blood and tissue concentrations of R-­and S-­enantiomers of methadone and its metabolite EDDP. Forensic Science International 226 (1–3): 254–260. Jones, G.R. and Pounder, D.J. (1987). Site dependence of drug concentrations in postmortem blood-­ -­ a case study. Journal of Analytical Toxicology 11 (5): 186–190. Jonsson, A.K., Soderberg, C., Espnes, K.A. et  al. (2014). Sedative and hypnotic drugs-­-­fatal and non-­fatal reference blood concentrations. Forensic Science International 236: 138–145. Ketola, R.A. and Ojanpera, I. (2019). Summary statistics for drug concentrations in post-­mortem femoral blood representing all causes of death. Drug Testing and Analysis 11 (9): 1326–1337. Kronstrand, R., Guerrieri, D., Vikingsson, S. et al. (2018). Fatal poisonings associated with new psychoactive substances. Handbook of Experimental Pharmacology 252: 495–541. Kuepper, U., Musshoff, F., and Madea, B. (2011). Applicability of succinylmonocholine as a marker for succinylcholine administration  – comparative analysis of samples from a fatal succinylcholine-­intoxication versus postmortem control specimens. Forensic Science International 207 (1–3): 84–90. Langford, A.M. and Pounder, D.J. (1997). Possible markers for postmortem drug redistribution. Journal of Forensic Sciences 42 (1): 88–92. Launiainen, T. and Ojanpera, I. (2014). Drug concentrations in post-­ mortem femoral blood compared with therapeutic concentrations in plasma. Drug Testing and Analysis 6 (4): 308–316. Lefrancois, E., Reymond, N., Thomas, A. et al. (2021). Summary statistics for drugs and alcohol concentration recovered in post-­mortem

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Agrawal, Y.P. and Rennert, H. (2012). Pharmacogenomics and the future of toxicology testing. Clinics in Laboratory Medicine 32 (3): 509–523. Alvarez, J.C., Mayer-­Duverneuil, C., Cappy, J. et al. (2020). Postmortem fatal and non-­fatal concentrations of amlodipine. Forensic Science International 316: 110555. Apple, F.S. (1989). Postmortem tricyclic antidepressant concentrations: Assessing cause of death using parent drug to metabolite ratio. Journal of Analytical Toxicology 13 (4): 197–198. Baselt, R.C. (2020). Disposition of Toxic Drugs and Chemicals in Man, 12th edn. Seal Beach, CA: Biomedical Publications. Bergh, M.S., Bogen, I.L., Wilson, S.R., and Oiestad, A.M.L. (2018). Addressing the fentanyl analogue epidemic by multiplex UHPLC-­MS/ MS analysis of whole blood. Therapeutic Drug Monitoring 40 (6): 738–748. Borron, S.W., Monier, C., Risede, P. and Baud, F.J. (2002). Flunitrazepam variably alters morphine, buprenorphine, and methadone lethality in the rat. Human and Experimental Toxicology 21 (11): 599–605. Brøsen, K. (2004). Some aspects of genetic polymorphism in the biotransformation of antidepressants. Therapie 59 (1): 5–12. Burns, G., DeRienz, R.T., Baker, D.D. et  al. (2016). Could chest wall rigidity be a factor in rapid death from illicit fentanyl abuse? Clinical toxicology (Philadelphia) 54 (5): 420–423. Chevillard, L., Decleves, X., Baud, F.J. et al. (2013). Respiratory effects of diazepam/methadone combination in rats: A study based on concentration/effect relationships. Drug and Alcohol Dependence 131 (3): 298–307. Darke, S., Duflou, J., and Torok, M. (2009). Drugs and violent death: comparative toxicology of homicide and non-­substance toxicity suicide victims. Addiction 104 (6): 1000–1005. Dickson, E.W., Bird, S.B., Gaspari, R.J. et al. (2003). Diazepam inhibits organophosphate-­induced central respiratory depression. Academic Emergency Medicine 10 (12): 1303–1306. Druid, H. and Holmgren, P. (1997). A compilation of fatal and control concentrations of drugs in postmortem femoral blood. Journal of Forensic Sciences 42 (1): 79–87.

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ratio of norbuprenorphine/buprenorphine was higher in the intoxication group than in the postmortem controls (0.6 vs. 1.9) and (02 vs. 2.9) in blood and urine, respectively. Lastly, there are several weak opioids that are frequently encountered in postmortem blood, e.g. codeine. These show a postmortem blood concentration pattern that is similar to “regular” pharmaceutical drugs, i.e. with significantly higher median concentrations in certified intoxications compared to postmortem controls. The small amounts of morphine that may be found in codeine intoxications are typically low, and even though morphine is definitely more potent, the morphine concentrations are far much lower than those seen in pure morphine toxicity deaths. This suggests that codeine per se, and not morphine, produces the toxic effects in acute intoxications, but this warrants further studies.

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Pragst, F. and Balikova, M.A. (2006). State of the art in hair analysis for detection of drug and alcohol abuse. Clinica Chimica Acta 370 (1–2): 17–49. Prouty, R.W. and Anderson, W.H. (1990). The forensic science implications of site and temporal influences on postmortem blood-­drug concentrations. Journal of Forensic Sciences 35 (2): 243–270. Reis, M., Aamo, T., Ahlner, J., and Druid, H. (2007). Reference concentrations of antidepressants. A compilation of postmortem and therapeutic levels. Journal of Analytical Toxicology 31 (5): 254–264. Sajantila, A., Palo, J.U., Ojanpera, I. et al. (2010). Pharmacogenetics in medico-­legal context. Forensic Science International 203 (1–3): 44–52. Schulz, M., Iwersen-­Bergmann, S., Andresen, H., and Schmoldt, A. (2012). Therapeutic and toxic blood concentrations of nearly 1,000 drugs and other xenobiotics. Critical Care 16 (4): R136. Schulz, M., Schmoldt, A., Andresen-­ Streichert, H., and Iwersen-­ Bergmann, S. (2020). Revisited: Therapeutic and toxic blood concentrations of more than 1100 drugs and other xenobiotics. Critical Care 24 (1): 195. Schwarzacher, S.W., Rub, U., and Deller, T. (2011). Neuroanatomical characteristics of the human pre-­Botzinger complex and its involvement in neurodegenerative brainstem diseases. Brain 134 (Pt 1): 24–35. Selden, T., Ahlner, J., Druid, H., and Kronstrand, R. (2012). Toxicological and pathological findings in a series of buprenorphine related deaths. Possible risk factors for fatal outcome. Forensic Science International 220 (1–3): 284–290. Skov, L., Johansen, S.S., and Linnet, K. (2015). Postmortem femoral blood reference concentrations of aripiprazole, chlorprothixene, and quetiapine. Journal of Analytical Toxicology 39 (1): 41–44. Soderberg, C., Tillmar, A., Johansson, A. et al. (2020). The importance of sample size with regard to the robustness of postmortem reference values. Forensic Science International 311: 110292. Soderberg, C., Wernvik, E., Tillmar, A. et  al. (2016). Antipsychotics  – Postmortem fatal and non-­fatal reference concentrations. Forensic Science International 266: 91–101. Stead, A.H. and Moffat, A.C. (1983). A collection of therapeutic, toxic and fatal blood drug concentrations in man. Human Toxicology 2 (3): 437–464. Thaulow, C.H., Oiestad, A.M.L., Rogde, S. et  al (2018). Can measurements of heroin metabolites in post-­mortem matrices other than peripheral blood indicate if death was rapid or delayed? Forensic Science International 290: 121–128. Welter-­Luedeke, J. and Maurer, H.H. (2016). New psychoactive substances: Chemistry, pharmacology, metabolism, and detectability of amphetamine derivatives with modified ring systems. Therapeutic Drug Monitoring 38 (1): 4–11. White, J.M. and Irvine, R.J. (1999). Mechanisms of fatal opioid overdose. Addiction 94 (7): 961–972. Zvosec, D.L., Smith, S.W., Porrata, T. et  al. (2011). Case series of 226  gamma-­hydroxybutyrate-­associated deaths: lethal toxicity and trauma. American Journal of Emergency Medicine 29 (3): 319–332.

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femoral blood in Western Switzerland. Forensic Science International 325:110883. Linnet, K. (2012). Postmortem drug concentration intervals for the non-­ intoxicated state  – A review. Journal of Forensic and Legal Medicine 19 (5): 245–249. Linnet, K., Johansen, S.S., Buchard, A. et al. (2008). Dominance of pre-­ analytical over analytical variation for measurement of methadone and its main metabolite in postmortem femoral blood. Forensic Science International 179 (1): 78–82. Maas, A., Madea, B., and Hess, C. (2018). Confirmation of recent heroin abuse: Accepting the challenge. Drug Testing and Analysis 10 (1): 54–71. Mash, D.C., Duque, L., Pablo, J. et al. (2009). Brain biomarkers for identifying excited delirium as a cause of sudden death. Forensic Science International 190 (1–3): e13–19. Maurer, H.H. (2018). Impact of Pharmacogenomic variations and/or drug/drug(food) interactions on interpretation of clinical and forensic toxicology cases with CNS acting drugs. TIAFT Bulletin 48 (1): 27–30. Mikkelsen, C.R., Jornil, J.R., Andersen, L.V. et al. (2016). Quantification of 16 QT-­prolonging Drugs and Metabolites in Human Postmortem Blood and Cardiac Tissue Using UPLC-­MS-­MS. Journal of Analytical Toxicology 40 (4): 286–293. Mikkelsen, C.R., Jornil, J.R., Andersen, L.V. et al. (2018). Distribution of Eight QT-­prolonging drugs and their main metabolites between postmortem cardiac tissue and blood reveals potential pitfalls in toxicological interpretation. Journal of Analytical Toxicology 42 (6): 375–383. Musshoff, F., Stamer, U.M., and Madea, B. (2010). Pharmacogenetics and forensic toxicology. Forensic Science International 203 (1–3): 53–62. Nedahl, M., Johansen, S.S., and Linnet, K. (2018). Reference brain/blood concentrations of citalopram, duloxetine, mirtazapine and sertraline. Journal of Analytical Toxicology 42 (3): 149–156. Nedahl, M., Johansen, S.S., and Linnet, K. (2019). Brain-­blood ratio of morphine in heroin and morphine autopsy cases. Forensic Science International 301: 388–393. Pilgrim, J.L., Gerostamoulos, D., and Drummer, O.H. (2012). The role of toxicology interpretations in sudden death. Forensic Science, Medicine, and Pathology 8 (3): 263–269. Pilgrim, J.L., Gerostamoulos, D., and Drummer, O.H. (2010). Review: Pharmacogenetic aspects of the effect of cytochrome P450 polymorphisms on serotonergic drug metabolism, response, interactions, and adverse effects. Forensic Science, Medicine, and Pathology 7 (2): 162–184. Pounder, D.J. (1993). The nightmare of postmortem drug changes. Legal Medicine 163–191. Pounder, D.J., Adams, E., Fuke, C., and Langford, A.M. (1996). Site to site variability of postmortem drug concentrations in liver and lung. Journal of Forensic Sciences 41 (6): 927–932. Pounder, D.J. and Jones, G.R. (1990). Post-­mortem drug redistribution-­ -­ a toxicological nightmare. Forensic Science International 45 (3): 253–263.

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Traffic Medicine

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53 | Driving Aptitude and Fitness to Drive 54 | Effects of Cardiovascular Disease on Fitness to Drive 55 | Effects of Vision and Visual Fields on Fitness to Drive 56 | Effects of Epilepsy on Fitness to Drive 57 | Effects of Diabetes on Fitness to Drive 58 | Epidemiology and Causal Factors in Fitness to Drive 59 | Driving Under the Influence of Alcohol 60 | Effects of Illegal Drugs on Fitness to Drive 61 | Effects of Medicinal Drugs on Fitness to Drive 62 | Toxicological Markers of Chronic Alcohol Abuse 63 | Traffic Accidents Handbook of Forensic Medicine, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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53

Driving Aptitude and Fitness to Drive

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Frederick Vinckenbosch, Annemiek Vermeeren, Eef L. Theunissen and Johannes G. Ramaekers

domain of traffic medicine, a scientific discipline that aims to enhance human safety in traffic through research, and the practical application of medical findings and experiences. Provided the significant burden of road traffic accidents and the increasing number of motor vehicles on the roads, traffic medicine will remain an important discipline until technological advances eliminate the need for a human operator, which is conceivably still many years away.

53.1.1  Relevance of traffic medicine

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The number of annual road traffic deaths is steadily increasing. While the World Health Organization reported an annual road traffic death toll of 1.25  million in 2015 (World Health Organization, 2015), which seemed to have leveled off since 2007, in 2018 an annual road traffic death toll of 1.35  million was reported (World Health Organization,  2018). The increase in road traffic deaths is partly attributable to the growing number of motor vehicles that frequent the roads. Corrected for this growing fleet of motor vehicles, an encouraging steady decrease of the relative death toll can be observed. Nevertheless, each fatality is a tragedy for those involved and their social network, especially considering it is the leading cause of death in 5–29 year olds. In addition to fatalities, around 50 million people get injured in road traffic accidents each year (World Health Organization,  2018). Taken together, road traffic injuries and deaths account for about one-­third of the disability adjusted life years due to accidental injuries, and therefore put a significant burden on society as a whole (Haagsma et al., 2016). Curbing these numbers starts with eliminating the causal factors of road traffic accidents. Human error lies at the basis of the vast majority of cases (Singh, Kushwaha, Agarwal, & Sandhu, 2016). To minimize the likelihood of human error, it is necessary to identify causal factors during driving and to be able to make informed decisions about an individual drivers’ ability to adequately and responsibly operate a motor vehicle. This is the

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53.1 Introduction

Handbook of Forensic Medicine, Volume 3, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

53.1.2  Traffic medicine and forensic medicine Traffic medicine and forensic medicine are closely related disciplines that overlap a great deal. Traffic medicine is mainly concerned with driving capability, the ability to safely operate a motor vehicle. Driving capability can be thought of as the sum of driving aptitude, fitness to drive, and driving skills (Berghaus & Schnabel, 2014; Staak, Hobi, & Berghaus, 1988). Driving aptitude is determined by physical and psychological traits and functions that are stable over time. It is dependent on driver dispositions such as personality traits and psychomotor functioning and can be affected by chronic medical conditions such as epilepsy, sleep apnea, narcolepsy, uncorrectable visual impairments, certain personality disorders, and many more. In traffic medicine, the assessment of driving aptitude is crucial for advising patients on whether to drive or determine circumstances under which ­driving is possible or discouraged. Unlike driving aptitude, fitness to drive is not stable over time. It is defined as the situation-­and

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method for identifying factors associated with increased risk of traffic accident involvement has been the study of differential accident involvement (Ranney,  1994). This approach investigates which characteristics and factors are more prevalent or pronounced in drivers involved in a road traffic accident. Because of obvious ethical objections, the study of differential accident involvement cannot rely on experimental study designs. The approach is therefore limited to epidemiological research methods. The most widely adopted study design for investigating contributing factors to road traffic accidents and at-­risk drivers is the case–control study. Case–control studies start with identifying cases, i.e. drivers who have been involved in a traffic accident, and controls, e.g. a random sample of drivers recruited at roadside checkpoints (Houwing et  al.  2009; Vandenbroucke and Pearce  2012). It is then determined whether the odds of being exposed to a certain external factor (e.g. alcohol consumption) prior to the accident or the odds of having an inherent quality (e.g. impulsivity) are higher for the group of drivers involved in a road traffic accident. Over the years, case–control studies have helped identify countless factors and characteristics that are associated with road traffic accidents, ranging from stable traits, such as demographics (e.g. age, gender, etc.) and personality characteristics (e.g. neuroticism, aggression, etc.) (Dumais et al., 2005; Mullin, Jackson, Langley, & Norton, 2000; Wang et al., 2012), to more transient states such as drug intoxications (Gjerde et al. 2011; Ravera et al. 2011). One of the most noteworthy results of case–control research has been providing the rationale for per se limits of alcohol in drivers. The first case–control study investigating the association between blood alcohol concentrations (BACs) and road traffic accidents is known as the Grand Rapids study (Borkenstein, Crowther, & Shumate, 1974; Hurst, Harte, & Frith, 1994). This study reported an exponential increase in crash risk from a BAC > 0.4 g/L onward. Together with more recent findings confirming that relative crash risk increase at a BAC > 0.4–0.5 g/L, the Grand Rapids study laid the foundation for the per se BAC limit of 0.5 g/L adhered to in many countries worldwide (Jones, 2010). Another common type of epidemiological study is the retrospective cohort study. This type of study is a useful alternative to the case–control study when the factor or characteristic under investigation is relatively rare in the general population, e.g. suffering from narcolepsy (Tzeng et  al.,  2019). In retrospective cohort studies, participants who are known to present with a specific feature of interest are grouped and compared to a control group regarding the prior incidence of motor vehicle accidents. Alternatively, in a prospective cohort study, groups of presumed at-­risk drivers and controls are compared regarding the incidence of motor vehicle accidents during a predetermined period following study enrollment, e.g. Nabi et al. (2006). Although the epidemiological approach has proven valuable for identifying at-­risk groups and framing legislation, these types of studies are subject to some practical and theoretical limitations. A major practical concern of epidemiological research is the general requirement for large sample sizes. Especially retro-­ and prospective cohort studies face this obstacle because of the

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time-­dependent ability to drive a vehicle. Fitness to drive can be influenced by transient factors such as fatigue or the occasional use of psychoactive substances. For traffic medicine, fitness to drive is vital to consider for advising patients on when to avoid operating a vehicle and for how long, e.g. when prescribing sedating medication. Lastly, driving skills are the operations required to operate a motor vehicle successfully and can be improved through training and experience. Whereas traffic medicine is mainly concerned with driving capability, forensic medicine is concerned with driver culpability. It builds on the methods and insights of traffic medicine regarding driving capability to help frame legislation and devise methods that aid in enforcing this legislation. An obvious example is the determination of accident culpability of drivers who tested positive for alcohol or other psychoactive substances in biological samples (breath, urine, saliva, or blood). In addition, forensic medicine experts often serve as expert witnesses in court to help answer the question of culpability and risk of recidivism, which is important for determining penalties. In this capacity, the forensic expert is faced with questions regarding driver aptitude, e.g. advising on whether to suspend a driver’s license of an older driver, and fitness to drive, e.g. determining the likelihood that a driver was significantly intoxicated at a given time. Thus, the forensic expert will translate the findings of traffic medicine into practice where the circumstances often permit little room for error. Therefore, it is important that the forensic expert is familiar with the insights and methods of traffic medicine in order to formulate appropriate advice.

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53.1.3  Outline and purpose of the chapter

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The purpose of this chapter is to familiarize the reader with different empirical methods for the identification of groups of drivers who have a chronically (driving aptitude) or transiently (driver fitness) increased risk of being involved in a road traffic accident. To this end, epidemiological methods for studying differential accident involvement and experimental methods for the assessment of driving performance are discussed. Next, focus is shifted from the group level to the individual level. General considerations regarding the assessment of individual driving aptitude and fitness to drive are discussed.

53.2 Empirical methods for identifying risk factors for road traffic accidents 53.2.1  Epidemiological research: differential accident involvement In practical terms, traffic and forensic medicine are mainly concerned with identifying drivers at increased risk of being involved in traffic accidents. During most of the previous century, the main

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ing performance assessment, ecological validity is the extent to which test outcomes translate to actual “real-­world” driving performance. Hence, a test that closely resembles actual driving has high ecological validity. There is no established approach for determining ecological validity other than the careful consideration of the characteristics of the respective test and construct under investigation (Schmuckler, 2001). Lastly, for a test to have construct validity, it should be reliable. Reliability refers to the repeatability of a test outcome when administered under similar conditions (Golafshani, 2003). Criterion validity captures how well performance on a specific task (e.g. reaction times in a visual search task) predicts a certain outcome (e.g. pass/fail during a driving examination or future accident risk). This form of validity is more related to performance thresholds than the test itself. Performance thresholds are necessary for making unequivocal decisions regarding fitness to drive and driving aptitude on both the group and individual levels. Hence, if a performance threshold is defined for a particular test, it should be supported by proof of criterion validity in order to be useful for making decisions regarding driver (un)fitness. Criterion validity can be demonstrated by comparing a tests’ outcome to that of a test with high ecological validity, such as on-­ road driving tests (see section on “On-­road driving tests” below). Lastly, content validity is the extent to which a test (battery) assesses all relevant functions that underlie (a specific aspect of) driving performance. As a heuristic, the hierarchical models of the driving task by Michon (1985) and Keskinen (1994) can be applied. The hierarchical model of the driving task proposed by Michon (1985) organized the driving task into three hierarchical levels. The operational level (1) at the bottom of the hierarchy consists of highly automated tasks required for basic vehicle control, such as road tracking and braking. Directly above the operational level is the tactical level (2). This level describes operations that allow the driver to negotiate the traffic environment. For example, overtaking and turning on an intersection depends on actions at the maneuvering level. The highest level of the hierarchy is the strategic level (3). On this level, route planning and a priori risk assessment (e.g. assessing the risk of going for a drive in bad weather) takes place. It is important to note that the different levels are interdependent, or as Michon (1985) puts it, exist in a “nested hierarchy.” It is clear that the goals set at the strategic level, e.g. arriving at work on time, can only be accomplished if the necessary actions on the maneuvering level are performed, e.g. overtaking slower traffic or driving faster. In turn, the actions planned on the maneuvering level require the execution of the corresponding action patterns on the operational level to steer the vehicle along the envisioned course. Keskinen (1994) extended Michon’s (1985) model with a fourth hierarchical level (Keskinen 1994; Hatakka et al. 2002). The three lowest levels, i.e. (1) vehicle maneuvering, (2) mastering traffic situations, and (3) goals and context of driving, correspond to Michon’s (1985) operational, tactical, and strategic levels, respectively. The fourth level, labeled goals for life and skills for living, can be understood as reflecting a driver’s personal attitudes relevant to driving, such as risk-­taking behavior. The fourth level is

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relatively low frequency of traffic accidents. Case–control studies start with identifying cases of drivers involved in traffic accidents and therefore circumvent the methodological problem of infrequent accident occurrences. However, like all epidemiological study designs, case–control studies face the problem that traffic accidents happen because of a multitude of reasons. This dilutes the relative prevalence of the characteristic or factor of interest. Again, this results in the need for large sample sizes. The need for large samples is not an insurmountable obstacle as evidenced by the many epidemiological studies that have been conducted successfully. However, this type of research is limited in a more fundamental way. Epidemiological research methods are associative in nature. Because of the lack of randomization and manipulation, epidemiological studies are prone to confounding. A straightforward way of dealing with confounders is to measure or document likely confounders and then statistically correct for them. However, the many possible confounders make it challenging and laborious to deal with this issue. Furthermore, adding explanatory variables to statistical analyses generally decreases the statistical power to detect a true relation. In conclusion, epidemiological research designs are not optimal for demonstrating causality. In order to do that, epidemiological findings require support from experimental research. For further reading on types, strengths, and weaknesses of epidemiological study designs, the reader is referred to DiPietro (2010).

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53.2.2  Experimental research: assessment of driving performance

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In contrast to epidemiological studies, experimental studies are the preferred research method for studying mechanisms and demonstrating causality. However, in the field of traffic medicine, the benefit of demonstrating causality comes at a great cost. The actual outcome of interest, i.e. traffic accidents, cannot be studied directly. Instead, experimental researchers have to resort to the assessment of driving performance. It is presumed that traffic accidents are preceded by chronic or acute impairments in driving performance, and hence, that the assessment of driving performance can predict the likelihood of a road traffic accident occurring. A central question for the assessment of driving performance is the validity of the applied assessment techniques. Three types of validity should be considered when selecting tests or test batteries to assess driving performance, i.e. construct validity, criterion validity, and content validity (Crossley, Humphris, & Jolly, 2002; Cureton, 1951; Downing, 2003). Construct validity is the extent to which a specific test (battery) assesses the construct of interest, i.e. (a specific aspect of) driving performance. In practice, the demonstration of construct validity relies on showing an association between test performances on different tasks hypothesized to assess the same underlying construct, a.k.a. convergent validity, which can be considered to be a part of construct validity (Carlson & Herdman, 2012). Another aspect of construct validity is ecological validity. In terms of driv-

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Table 53.1  Example of items and weights (obtainable scores) of the Test Ride for Investigating Practical (TRIP) fitness to drive. Score range

Lateral position on the road at speed below 50 km/h

2–8

Lateral position on the road at speed above 50 km/h

2–8

Mechanical operations

3–12

Speed adaptations at speed below 50 km/h

2–8

Speed adaptations at speed above 50 km/h

2–8

Gap distance at speed below 50 km/h

2–8

Gap distance at speed above 50 km/h

2–8

Lane position change

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Visual behavior and communication

8–32

Understanding, insight, and quality of traffic perception

2–8

Turning left

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Anticipation and perception of traffic light and signs

Joining the traffic stream

6–24

Total score:

49–196

Source: Adapted from Devos et al. (2013).

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an important addition because personal attitudes can potentially compensate or undermine functioning at the lower levels. A comprehensive assessment of driving performance preferably covers all levels described in Keskinen’s (1994) model. However, there is arguably no single assessment method that adequately covers all the hierarchical levels of the driving task. Also, which functional levels need to be assessed depends on the suspected functional impairments. For example, elderly drivers are arguably more likely to be impaired on the lower hierarchical levels i.e. operational functions/vehicle maneuvering and tactical functions/mastering of traffic situations, than younger drivers who are more likely to exhibit more risk-­taking behavior (Ball et  al.  1998; McGwin Jr. and Brown  1999; Karthaus and Falkenstein 2016). Hence, in the context of driving performance, content validity is the extent to which all aspects of driving performance relevant to the specific population and research question at hand are assessed. It follows that the assessment of driving performance requires the careful assembly of a test battery consisting of on-­road and/or simulated driving tasks combined with neurocognitive and psychological testing, depending on the driving behaviors or functional domains of interest. The rest of the section provides a general overview of the methods for the assessment of driving performance in experimental research.

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introduces safety concerns for the participant, examiner, and other road users. Second, a glance at the example checklist in Table 53.1 shows that on-­road driving tests mainly assess operations at the two lower levels of Michon’s (1985) and Keskinen’s (1996) hierarchical models, i.e. skills directly related to operating a vehicle in traffic. The higher levels, where, for instance, risk-­ taking behavior and route planning reside, are not assessed despite their likely considerable impact on lower levels of functioning. Lastly, no two drives are the same because of the ever-­ changing traffic environment. Therefore, the applied checklists for rating driving performance must be limited to skills and maneuvers that can be assessed during every drive. Hence, reactions to unexpected and/or near-­crash events, arguably better predictors of crash risk, cannot be assessed.

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On-­road driving tests are often considered as the most ecologically valid assessments of driving performance. The tests are similar to a classic driving examination. The participant drives a predetermined route through actual traffic while a licensed instructor rates performance on items listed on a standardized checklist (for an example, see Table 53.1). Several on-­road driving test procedures have been developed over the years. A recent systematic review by Sawada et al. (2019) investigated the validity and reliability of commonly used driving tests and concluded that the Washington University Road Test (Hunt et  al.,  1997), the Rhode Island Road Test (Brown et al., 2005), the Test Ride for Investigating Practical (TRIP) fitness to drive (Devos et al., 2007; Tant, Brouwer, Cornelissen, & Kooijman, 2002), and the Performance Analysis for Driving Ability (Patomella, Caneman, Kottorp, & Tham, 2004) were highly reliable assessment methods. Proof of criterion or construct validity was gathered by comparing the test outcomes to those of other on-­road or neurocognitive tests, respectively. It should be noted that these tests were specifically designed to assess the driving performance of rehabilitation patients. Nevertheless, the listed tests are good examples of high-­quality on-­road driving tests. On-­road driving tests are considered the “golden standard” (Sawada et al., 2019) because the tests’ high degree of ecological validity cannot be matched by any other assessment technique. These tests take place in actual traffic and usually cover a wide range of traffic environments (city centers, motorways, various types of intersections, etc.). However, several conceivable shortcomings deserve consideration. First, exposing a potentially impaired driver to the complexities of real-­world traffic situations

The standardized on-­the-­road highway driving test The standardized on-­the-­road highway driving test is a special case of on-­ road driving tests (Ramaekers,  2017; Verster & Roth, 2011). The test requires participants to complete a 100 km course on a public highway. The driver is instructed to maintain a steady lateral position in the right traffic lane and to keep a steady velocity of 95 km/h. It is allowed to ignore these instructions to overtake slower traffic, leaving the highway halfway to turn back, or because of immediate safety concerns. A licensed driving instructor who has access to dual controls, i.e. brake, accelerator, and clutch, accompanies the driver. Both driver and instructor can decide to abort the test if safety concerns arise. During the drive, an optic device mounted on top of the test vehicle continuously monitors the vehicle’s lateral position relative to the traffic lane

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The standardized on-­the-­road highway driving test is subject to the same limitations as other on-­road driving tests. Even more, it arguably only assesses functioning on the lowest level of Michon’s (1985) and Keskinen’s (1996) hierarchical models of driving as road tracking is a highly automated function in licensed/experienced drivers. However, because of the model’s hierarchical nature, it can be reasoned that if performance is impaired at the lowest level, functioning is impaired on all levels. The standardized on-­road highway driving test is a valuable tool for the assessment of sustained attention, because of its rather long duration (approximately 60 minutes) and the monotonous nature, in an ecologically valid setting. This makes the test a superior method for the assessment of the impairing effects of sedative and dissociative drugs (Leufkens, Ramaekers, de Weerd, Riedel, & Vermeeren, 2009; Ramaekers, Robbe, & O’Hanlon, 2000; Theunissen, Vermeeren, & Ramaekers, 2006), as well as sleep and attentional disorders (Jongen, Perrier, Vuurman, Ramaekers, & Vermeeren, 2015; Verster et al., 2008).

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demarcation on the left, which is documented on the on-­board computer drive. Also, velocity is continuously monitored. The data are later edited offline to mark events such as overtake maneuvers. The remaining data are then used to determine the mean speed, mean lateral position, standard deviation of speed, and the standard deviation of the lateral position (SDLP). The SDLP is the primary outcome measure of the standardized on-­road highway driving test. It is essentially a quantification of road-­tracking performance or lane weaving, with higher values reflecting poorer performance. The measure has proven to be a very sensitive parameter for detecting the impairing effects of licit and illicit sedating intoxicants such as benzodiazepine receptor agonists, antihistamines, tri-­and tetracyclic antidepressants, cannabis, alcohol, but also sleepiness and fatigue (Ramaekers et  al.  2000; Vermeeren  2004; Theunissen et  al.  2006; Bosker et al. 2012; Jongen et al. 2015; van der Sluiszen et al. 2016; Jongen et  al.  2017). Randomized, placebo-­controlled, crossover trials have reliably demonstrated a 2.4 cm increase in SDLP at a blood– alcohol concentration of 0.5 g/L (Jongen et al., 2017). Considering the finding of the aforementioned Grand Rapids study that demonstrated an exponentially increased accident risk at this BAC (Borkenstein et al., 1974; Hurst et al., 1994), a 2.4 cm increase in SDLP serves as the benchmark for clinically or functionally relevant impairment and is considered to reflect the threshold for increased accident risk (Ramaekers, 2017). See Figure 53.1 for an illustration of the standardized on the road driving test (Ramaekers, 2017). 

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Lateral position

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Sedating drug (SDLP increases) 100 km Figure 53.1  Illustration of the standardized on-­road highway driving test and its main outcome measure, the standard deviation of lateral position (SDLP). Source: After Ramaekers (2017).

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for investigating driving behavior under specific (rare or dangerous) circumstances. Also, by manipulating the simulated environment, tests of executive functioning (e.g. working memory, selective attention, etc.) can be embedded in the scenarios (e.g. counting the number of red vehicles encountered) (Fisher et al., 2011). Lastly, an advantage of driving simulator testing is the abundance of outcome measures that can be readily documented, ranging from parameters of direct vehicle control (e.g. steering wheel angle, braking force, accelerator release, etc.) to more dynamic outcome measures (e.g. distance keeping, lateral position, velocity, accelerator release in response to an event, etc.) (Fisher et al., 2011). However, driving simulators have their own unique limitations. The most prominent one is the possibility of simulator sickness (Helland et  al.,  2016; Kennedy, Lane, Berbaum, & Lilienthal, 1993). Simulator sickness is similar to motion sickness and likely arises from the mismatch between visual, vestibular, and proprioceptive input (Groen & Bos, 2008). Common complaints are oculomotor strain, feelings of disorientation, and nausea (Kennedy et al., 1993). It is clear that these complaints have the potential to distort the measurements during simulator testing (Helland et al., 2016). Even if there are no apparent signs of simulator sickness, driving performance might still be influenced

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advanced driving simulator is the National Advanced Driving Simulator 1 (NADS-­1) at the University of Iowa (Chen, Papelis, Waston, & Solis, 2001) (Figure 53.2, top right and left). Despite the superior ecological validity, there are few simulators of the latter type, likely because of the high costs associated with purchase, construction, maintenance, and operation. More commonly used are simpler setups consisting of a driver seat, pedals (accelerator, brake, and clutch), steering wheel, and a screen displaying the traffic environment and dashboard, all mounted on a fixed base (Cuenen et al., 2019; Mets et al., 2011). Other, more advanced fixed-­base setups involve the body of an actual vehicle mounted in front of a large projection screen (Figure 53.2, bottom) (Hussain, Pirdavani, Ariën, Brijs, & Alhajyaseen, 2018). Driving simulator testing circumvents many problems associated with on-­road driving tests. First, there are minimal, if any, safety concerns associated with simulator testing. It is therefore possible to assess driving performance of impaired drivers without introducing potential hazards. Driving simulators often come equipped with standard driving scenario’s, but most simulation software also allows for the “de novo” design or tweaking of existing scenario’s (Fisher, Rizzo, Caird, & Lee, 2011). As such, virtually every aspect of the traffic environment can be manipulated by the experimenter. This makes driving simulators a potent tool

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Figure 53.2  Photographs taken from the inside (top left) and outside (top right) the National Advanced Driving Simulator (NADS-­1), a highly advanced moving base driving simulator at the University of Iowa, USA. The bottom image displays a more advanced fixed-­base simulator that features the cabin of an actual vehicle, as setup at the Institute for Mobility (IMOB) at Hasselt University, Belgium.

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Table 53.2  Overview of the five functional domains proposed by the FDA (Food and Drug Administration, 2017) which need to be assessed at a minimum for the investigation of driver fitness, complemented with a sixth domain comprising personal attitudes and characteristics, with examples of relevant functions and applicable tests.

Alertness and arousal

Alerting, orienting, sustained attention

ANTa, UFOVb, PVTc

Attention and processing speed

Stimulus response matching, divided attention, selective attention, top-­down controlled visual search

ANTa, SARTd, DSSTe, TMTf, Stroop testg, DATh

Reaction time and psychomotor functioning

Fine motor skills, reaction speed

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Executive functioning

Personal attitudes and characteristics

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CTTi, PRTj, FTTk, PVTc

Visual acuity, contrast sensitivity, movement perception, object recognition

ATTPTl, UFOVb, CCFm, motion perceptionn, dynamic visual acuityo

mapping dynamic traffic scene, evaluation of performance, planning, working memory

Spatial working memory testp, WCSTq, Tower of Londonr, n-­back tasks

Impulsivity, risk taking, aggression, respecting traffic rules

RT-­18t, BISu, DBQv

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In addition to on-­road and driving simulator testing, neurocognitive and psychological test batteries are frequently used to related functioning. The US Food and Drug assess driving-­ Administration (FDA) provided guidelines for evaluating drug effects on fitness to drive in which they list the minimal required functional domains to be assessed for evaluating driving-­related functioning, i.e. (1) alertness and arousal, (2) attention and processing speed, (3) reaction time and psychomotor functioning, (4) sensory perceptual functioning, and (5) executive functioning (Food and Drug Administration,  2017). These functions are rather broad and require deconstruction into subfunctions in order to be assessable. Table 53.2 shows examples of subfunctions and applicable neurocognitive and psychomotor tests to assess the functional domains listed by the FDA. These functional domains cover functioning/operating at the first three levels of Michon’s (1985) and Keskinen’s (1994) models, but disregard the highest level of Keskinen’s (1994) model which covers relevant personal attitudes and characteristics such as risk taking, impulsivity, aggression, and conscientiousness. Hence, a sixth functional domain with examples of relevant tests was added to Table 53.2. The list in Table 53.2 only serves to present examples of relevant functions and tests and is by no means intended to define the necessary features of a comprehensive and universal test battery. Neurocognitive and psychological test batteries are often assembled with specific driver groups in mind, e.g. stroke rehabilitation or traumatic brain injury patients (McKenna, Jefferies, Dobson, & Frude, 2004; Nouri & Lincoln,  1993), and validated through comparison with on-­road driving test results (pass/fail) with participants from the same population. Not only test batteries but also individual tests are often investigated for their suitability for the assessment of driving performance under specific circumstances, e.g. impairing effects of CNS depressants (Jongen, Vuurman, Ramaekers, & Vermeeren, 2016) or sleep deprivation (Jongen et al., 2015). Thus, when assessing driving performance, it should be considered whether the selected test (battery) is suitable for the demonstration of impairment in the population or context under investigation.

Examples of relevant functions

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Functional domain

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by the absence of vestibular and proprioceptive input that plays an important role in perceiving acceleration, deceleration, and centrifugal forces during turns (Reymond, Kemeny, Droulez, & Berthoz, 2001). In order to prevent these limitations, highly advanced moving base simulators can be used to mimic actual vehicle movements during driving. However, as stated above, advanced moving base simulators like the NADS-­1 are rare. Lastly, risk-­taking behavior is not easily assessed in a naturalistic way during driving simulator testing because of the driver’s awareness of the artificial nature of the simulated drive, which might prompt more risk taking because of the absence of real danger. Conversely, the driver might be inclined drive more responsibly because of test instruction or the social desire to appear as a capable driver to the experimenter.

 Attention Network Test (Fan, McCandliss, Sommer, Raz, & Posner, 2002);  Useful Field of View test (Ball & Owsley, 1993); c  Psychomotor Vigilance Task (Dinges & Powell, 1985); d  Sustained Attention to Response Test (Robertson, Manly, Andrade, Baddeley, & Yiend, 1997); e  Digit-­Symbol Substitution Test (McLeod, Griffiths, Bigelow, & Yingling, 1982); f  Trailmaking Test (Tombaugh, 2004); g  Stroop Color and Word Test (Stroop, 1935); h  Divided Attention Test (Moskowitz, 1973); i  Critical Tracking Task (Jex, McDonnell, & Phatak, 1966); j  Pursuit Rotor Task (Ammons, 1947); k  Finger Tapping Test (Morrison, Gregory, & Paul, 1979); l  Adaptive Tachistoscopic Traffic Perception Test (Schuhfried, 2009); m  Critical Flicker Fusion Threshold (Landis, 1954); n  (Lacherez, Au, & Wood, 2014; Wood, 2002); o  (Herdman et al., 1998; Vital et al., 2010); p  (Owen, Downes, Sahakian, Polkey, & Robbins, 1990); q  Wisconsin Card Sorting Test (Berg, 1948); r  (Krikorian, Bartok, & Gay, 1994); s  (Owen, McMillan, Laird, & Bullmore, 2005); t  (De Haan et al., 2011); u  Barratt Impulsiveness Scale (Patton, Stanford, & Barratt, 1995; Stanford et al., 2009); v  Driver Behavior Questionnaire (Martinussen, Hakamies-­Blomqvist, Møller, Özkan, & Lajunen, 2013). a

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53.2.3 Observational research of driving performance

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As described in Section 53.1.2, fitness to drive is defined as the situation-­and time-­dependent ability to drive a vehicle. Clinicians most often deal with the need for anticipation of unfitness to drive, e.g. impairing effects of prescribed medications, while forensic experts will generally deal with determining (un)fitness to drive retrospectively based on evidence collected at the time of suspicion. Examples of situations where fitness to drive can be impaired are acute drug intoxications, certain medical conditions, such as a transient ischemic attack, temporary mood, or emotional states such as depression or a fit of anger, and many more. These conditions’ transient nature introduces the necessity to determine fitness to drive soon after a suspicion of unfitness to drive arises. However, the resources for assessing driving performance and driving-­related functioning are usually not available when a suspicion of unfitness to drive presents itself. Therefore, the temporary (in)ability to adequately operate a motor vehicle should be predicted or evaluated after the fact based on available knowledge and evidence. Hence, the forensic expert will usually provide an expert opinion regarding fitness to drive in the context of a police investigation. The police investigation starts when suspicion of unfitness to drive occurs. This suspicion usually results from the observation of deviating driving behavior by either police officials or eyewitnesses. Examples of relevant observations are handling of the car, behavior of driver and passengers, weaving, driving in the middle of the road, maneuvering (skidding on a curve or curb), speed (too fast and too slow), tailgating, reactions (slow and belated), starting or breaking abruptly or belatedly, and crashes or near-­crashes (Berghaus & Schnabel, 2014). Especially when the police were the observers of the suspicious driving behavior, the observation itself already provides valuable information about the driver’s potential functional impairment at the time of the observation. However, it is not sufficient for the determination of (un)fitness to drive. It is crucial to establish whether the deviating driving behavior persists for some time, as every driver occasionally commits a driving error. Continued observation of driving behavior can be performed under circumstances where there is no immediate danger to the suspected driver or other road users. However, for safety reasons, the further collection of information by the police is usually carried out at the roadside after the driver has been stopped. The main source of information that might help identify drivers who are unfit to drive at roadside is the driver’s behavior and appearance. Examples of relevant observations are behavior and mood as well as changes in behavior and mood (tiredness, aggressiveness, etc.), physical abnormalities (speech, walking, agitation, tremor, etc.), orientation, responsiveness, reaction, logical reasoning, abnormalities concerning clothes or smell (in the car and in the respiratory air), and pupils (extension and reaction of the pupil) (Berghaus & Schnabel, 2014). In response to observations of driving behavior and appearance, the police can choose to collect evidence of acute intoxications affecting fitness to drive.

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Besides epidemiological and experimental research designs, there is also a third option, which is a hybrid form of the former study types. Observational research of driving performance selects pre-­ existing groups, e.g. long-­term hypnotic or anxiolytic users (van der Sluiszen et  al.,  2019), and applies testing procedures commonly used in experimental studies for the assessment of driving performance. The performance of the group of interest is then compared to that of a control group. These between-­subject research designs suffer from the same limitations regarding demonstrating causality as epidemiological studies due to the lack of randomization. They also lack the potential to demonstrate predictive validity because of the inability to directly assess actual road traffic accidents as an outcome. Nevertheless, observational studies can contribute to proof of convergent validity together with epidemiological findings whenever experimental manipulation is not possible, e.g. diseases and prolonged or irreversible treatments.

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The main advantage of neurocognitive and psychological testing is that the tests can be administered in laboratory or clinic settings, which is convenient, affordable, and safe. In addition, the approach allows for the detailed assessment of functioning relevant at all levels of Michon’s and Keskinen’s hierarchical models (Keskinen, 1994; Michon, 1985). Unlike the aforementioned testing methods, neurocognitive and psychological test batteries can be constructed to thoroughly assess higher order functions (executive functioning), such as route planning and risk assessment, and personal attitudes, such as consciousness and risk-­taking behavior. The clear disadvantage of neurocognitive and psychological testing is the lack of ecological validity. Unless supported by proof of convergent validity from on-­road and simulator testing, it is uncertain what role specific neurocognitive functions or ­ p sychological characteristics play in overall driving performance.

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53.3 Individual assessment of driving aptitude and fitness to drive Epidemiological and experimental research attempts to uncover risk factors for road traffic accidents. Although these empirical methods allow for identifying groups of drivers who have an increased risk of being involved in a road traffic accident, the findings cannot be extrapolated directly to individual drivers. Also, not all individual cases permit a thorough assessment of driving performance because of circumstantial or practical constraints. The current section discusses some important considerations in the individual assessment of fitness to drive and driving aptitude.

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(WAT) test. The HGN test requires the driver to follow an object (e.g. a pen or fingertip) with the eyes. The OLS assesses whether the driver is capable of standing on one leg for 30 seconds with the other foot approximately 15 cm above the ground. Lastly, the WAT consists of nine heel-­to-­toe steps along a straight line, after which the driver must turn back on one foot and take another nine heel-­ to-­toe steps along the straight line. The SFST was developed for the detection of alcohol-­induced impairment but has also been demonstrated to pick up stimulant and CNS depressant induced impairment (Porath-­ Waller & Beirness,  2014). However, it is unclear to what extent impaired performance during the SFTS is indicative of actual driving impairment. Also, the sensitivity of the SFST might be suboptimal. For instance, a study of the effects of THC (dronabinol) on driving performance demonstrated impaired performance in the on-­the-­road highway driving test in both occasional and heavy cannabis users, while the SFST was not able to discriminate between placebo and dronabinol conditions (Bosker et  al.,  2012). Nevertheless, it appears highly likely that severe impairment during the SFST is indicative of actual driving impairment. In conclusion, the transient nature of (un)fitness to drive makes a thorough assessment of driving performance and driving-­related functioning practically impossible. Hence, the forensic expert is required to carefully consider all the pieces of information that are available, i.e. police reports of observed driving behavior or traffic accident circumstances, behavior and appearance of the driver as observed by police, results of toxicological screening for the presence of driving impairing drugs, and information yielded during potential systematic assessments such as the SFST, in order to formulate proper advice.

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Depending on the legal possibilities within a given jurisdiction, common test procedures involve detecting or measuring drug concentrations in biological samples, such as breath (alcohol breathalyzer), saliva, urine, or blood plasma, or behavioral testing, such as the standardized field sobriety test (SFST). A few points regarding the collection of biological samples to detect substances known to impair driving deserve consideration. First, it is important to consider that detecting a drug known to impair driving does not necessarily indicate functional impairment. For instance, two studies on the effects of smoked cannabis on psychomotor and neurocognitive functioning in heavy cannabis users failed to demonstrate significant functional impairment, despite the fact that the tetrahydrocannabinol (THC) concentrations in blood plasma corresponded to oral fluid THC concentrations that would provide a positive result on some roadside saliva drug tests (Huestis and Cone  2004; Ramaekers et al. 2009; Wille et al. 2010; Ramaekers et al. 2011; Dobri et al. 2019). Roadside saliva and urine drug tests usually provide a binary outcome, i.e. positive or negative result for the presence of the drug (or metabolite) of interest. However, depending on the legislation (some jurisdiction apply per se limits while others have a zero tolerance policy), or specific circumstances (e.g. investigating whether a driver who tested positive for ketamine ran over his neighbor by accident or not), it might be necessary to follow up a positive result by the determination of actual drug concentrations to establish whether the drug was likely to cause the presumed driving impairment (Raes & Verstraete,  2005). Drug concentrations are usually determined in blood plasma samples. Most often, the collection of a blood plasma sample takes place some time after the positive drug test. Hence, the outcome of interest for the forensic expert is the estimated drug concentration at that time that aberrant driving was observed or a road traffic accident occurred. The estimated drug plasma concentration can then be compared to findings from epidemiological and experimental research so that the plausibility of the drug being a causal factor for driving impairment can be considered. However, in the absence of information on drug dosing and time or route of administration, it is not possible to retrospectively determine blood plasma concentrations with certainty. Furthermore, findings from epidemiological and experimental studies indicate drug concentrations at which an increased crash risk or impairment of driving (related functions) becomes apparent at the group level. However, group-­level findings from empirical research cannot be extrapolated to the individual without further consideration. Whether a given person is functionally impaired at a certain drug concentration arguably depends on many factors such as tolerance, concomitant drug use, or personality (e.g. tendency for aggression or impulsivity). Hence, the police’s observations regarding driving and driver behavior are valuable pieces of information that can help determine whether a driver was likely impaired due to drug use at a specific time in the past. In addition to general observations, an SFST (Burns & Moskowitz, 1977) is used in some jurisdictions, e.g. USA and Canada. The SFST consists of three parts, i.e. horizontal ­ alk-­and-­turn gaze nystagmus (HGN), one leg stand (OLS), and w

53.3.2  Assessment of driving aptitude After a driver has been judged to be unfit to drive, an assessment of driving aptitude may be necessary. Also, when there is doubt regarding the (un)fitness to drive at some point in the past, it might be advisable to assess driving aptitude. Examples of other reasons for ordering the assessment of driving aptitude can be traffic-­related criminal offenses or criminal offenses with a high aggression potential, subsequent to a medical assessment if deemed necessary, renewal of the driving license after license withdrawal due to penalty points, renewal of driving license at a certain age (required for elderly drivers in some jurisdictions), and repeated or severe offenses (Berghaus and Schnabel 2014). The methods for assessing driving aptitude consist largely of the same techniques used in experimental research (see Section 53.2.2). As discussed, there is a wide variety of on-­road driving, simulated driving, and neurocognitive and psychological tests available to assess driving performance and driving-­related functioning. The selected assessment methods must specifically consider the concrete information leading to the doubts about the driver’s aptitude. Often validated test batteries have been constructed for the assessment of driving aptitude in specific

References and further reading

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Ammons, R.B. (1947). Acquisition of motor skill: II. Rotary pursuit performance with continuous practice before and after a single rest. Journal of Experimental Psychology 37 (5): 393. Ball, K. and Owsley, C. (1993). The useful field of view test: A new technique for evaluating age-­related declines in visual function. Journal of the American Optometric Association 64 (1): 71–79. Ball, K., Owsley, C., Stalvey, B. et al. (1998). Driving avoidance and functional impairment in older drivers. Accident Analysis & Prevention 30 (3): 313–322. Berg, E.A. (1948). A simple objective technique for measuring flexibility in thinking. The Journal of General Psychology 39 (1): 15–22. Berghaus, G., and Schnabel, E. (2014). Driving aptitude and fitness to drive. Handbook of Forensic Medicine 995–1010. Borkenstein, R.F., Crowther, R.F., and Shumate, R.P. (1974). The role of the drinking driver in traffic accidents (the Grand Rapids study). Blutalkohol, 11 (Suppl): 1–131. Bosker, W., Kuypers, K., Theunissen, E. et  al. (2012). Medicinal THC (dronabinol) impairs on-­the-­road driving performance of occasional and heavy cannabis users but is not detected in Standardized Field Sobriety Tests. Addiction 107 (10): 1837–1844. Brown, L.B., Ott, B.R., Papandonatos, G.D. et  al. (2005). Prediction of on-­road driving performance in patients with early Alzheimer’s disease. Journal of the American Geriatrics Society 53 (1): 94–98. Burns, M. and Moskowitz, H. (1977). Psychophysical Tests for DWI Arrest. Retrieved from https://rosap.ntl.bts.gov/view/dot/1186 Carlson, K.D. and Herdman, A.O. (2012). Understanding the impact of convergent validity on research results. Organizational Research Methods 15 (1): 17–32. Chen, L., Papelis, Y., Waston, G., and Solis, D. (2001). NADS at the University of IOWA: A Tool for Driving Safety Research. Paper presented at the Proceedings of the 1st human-­centered transportation simulation conference. Crossley, J., Humphris, G., and Jolly, B. (2002). Assessing health professionals. Medical Education 36 (9): 800–804. Cuenen, A., Jongen, E. M., Brijs, T. et al. (2019). The effect of a simulator based training on specific measures of driving ability in older drivers. Transportation Research Part F: Traffic Psychology and Behaviour 64: 38–46. Cureton, E. (1951). Educational measurement. American Council on Education. De Haan, L., Kuipers, E., Kuerten, Y. et  al. (2011). The RT-­18: A new screening tool to assess young adult risk-­taking behavior. International Journal of General Medicine 4: 575. Devos, H., Nieuwboer, A., Vandenberghe, W. et al. (2013). Validation of driving simulation to assess on-­road performance in Huntington disease. Proceedings of the Seventh International Driving Symposium on Human Factors in Driver Assessment, Training and Vehicle Design, June 17–20, 2013, Bolton Landing, New  York. Iowa City, IA: Public Policy Center, University of Iowa, pp. 241–247. Devos, H., Vandenberghe, W., Nieuwboer, A. et al. (2007). Predictors of fitness to drive in people with Parkinson disease. Neurology 69 (14): 1434–1441.

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populations, e.g. patients with dementia or mild cognitive impairment (Fuermaier et al., 2017; Piersma et al., 2018; Piersma et al., 2016), or traumatic brain injury (McKenna et al., 2004). For certain medical conditions, the experimental methods discussed in Section 53.2.2 are inadequate to assess driving aptitude. Driving inaptitude can be conceptualized as a constant inability to safely operate a motor vehicle in traffic (e.g. patients with traumatic brain injury with lasting functional consequences) or as recurring bouts of unfitness to drive (e.g. epilepsy patients). Especially, medical conditions that cause driving inaptitude in the latter sense are not easily assessed with the methods described in Section  53.2.2. Instead, a thorough medical examination should be conducted, which also considers general physical and psychological condition, controllability of the disease by therapy (including medication), side effects of the medicines used, duration, intensity and severity of the disease, a combination of diseases, kind of participation in traffic of the driver (personal activities, commercial vehicles, and professional drivers), and the possibility to compensate for deficits (Berghaus and Schnabel  2014). An example of an alternative method for determining driver aptitude in epilepsy patients was proposed by Somerville et al. (2019), who introduced a decision tree incorporating the recent medical history of the patient. The individual assessment of driving aptitude has to be coherent and reviewable. This means that the inducing facts and resulting questions must be included, i.e. the applied assessment methods, and, if necessary, an explanation and discussion of the findings and their meaning. The report has to clearly distinguish between the previous history and the current findings. Additionally, the assessment report must be comprehensible, so that the assessed person and the driving license agency, court, and other relevant parties are able to comprehend the report. If the assessment leads to a negative result concerning the driver’s aptitude, recommendations should be given on how the preconditions for meeting the requirements can be improved (e.g. by substance abuse counseling or a therapeutic measure). A positive assessment may be made possible by naming impositions (e.g. wearing glasses) or restrictions (e.g. speed limitations).

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53.4 Conclusion

The current chapter aimed to provide a general overview of the techniques for identifying risk factors for road traffic accidents and the assessment of driving performance and related functioning, as well as their strengths and weaknesses. In addition, the available techniques and difficulties in assessing individual ­fitness to drive and driving aptitude were considered. It is clear that the selected sources of knowledge and methods of assessment greatly depend on the question at hand. The following chapters will elaborate in more detail about the findings and relevant aspects of different influential factors of driver fitness and driver aptitude.

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for drivers with dementia of the Alzheimer type. Archives of Neurology 54 (6): 707–712. Hurst, P.M., Harte, D., and Frith, W.J. (1994). The grand rapids dip revisited. Accident Analysis & Prevention 26 (5): 647–654. Hussain, Q., Pirdavani, A., Ariën, C. et al. (2018). The impact of perceptual countermeasures on driving behavior in rural-­urban transition road segments: A driving simulator study. Advances in Transportation Studies 46. Jex, H.R., McDonnell, J.D., and Phatak, A.V. (1966). A “critical” tracking task for manual control research. IEEE Transactions on Human Factors in Electronics 7 (4): 138–145. Jones, A.W. (2010). The relationship between blood alcohol concentration (BAC) and breath alcohol concentration (BrAC): A review of the evidence. Road Safety Web Publication 15: 1–43. Jongen, S., Perrier, J., Vuurman, E. et al. (2015). Sensitivity and validity of psychometric tests for assessing driving impairment: Effects of sleep deprivation. PLoS one 10 (2): e0117045. Jongen, S., Vermeeren, A., van der Sluiszen, N. et al. (2017). A pooled analysis of on-­the-­road highway driving studies in actual traffic measuring standard deviation of lateral position (ie,“weaving”) while driving at a blood alcohol concentration of 0.5 g/L. Psychopharmacology 234 (5): 837–844. Jongen, S., Vuurman, E., Ramaekers, J., and Vermeeren, A. (2016). The sensitivity of laboratory tests assessing driving related skills to dose-­ related impairment of alcohol: A literature review. Accident Analysis & Prevention 89: 31–48. Karthaus, M. and Falkenstein, M. (2016). Functional changes and driving performance in older drivers: assessment and interventions. Geriatrics 1 (2): 12. Kennedy, R.S., Lane, N.E., Berbaum, K.S., and Lilienthal, M.G. (1993). Simulator sickness questionnaire: An enhanced method for quantifying simulator sickness. The International Journal of Aviation Psychology 3 (3): 203–220. Keskinen, E. (1994). Why do young drivers have more accidents? Junge Fahrer und Fahrerinnen. Referate der Ersten Interdiziplinaren Fachkonferenz, December 12–14, 1994 in Koln. Krikorian, R., Bartok, J., and Gay, N. (1994). Tower of London procedure: a standard method and developmental data. Journal of clinical and Experimental Neuropsychology 16 (6): 840–850. Lacherez, P., Au, S., and Wood, J.M. (2014). Visual motion perception predicts driving hazard perception ability. Acta Ophthalmologica 92(1): 88–93. Landis, C. (1954). Determinants of the critical flicker-­fusion threshold. Physiological Reviews 34 (2): 259–286. Leufkens, T., Ramaekers, J., de Weerd, A. et al. (2009). On-­the-­road driving performance and driving related skills in untreated insomnia patients and chronic users of hypnotics. Hypnotics and Anxiolytics 97. Martinussen, L.M., Hakamies-­Blomqvist, L., Møller, M. et  al. (2013). Age, gender, mileage and the DBQ: The validity of the Driver Behavior Questionnaire in different driver groups. Accident Analysis & Prevention 52: 228–236. McGwin, G., Jr. and Brown, D.B. (1999). Characteristics of traffic crashes among young, middle-­aged, and older drivers. Accident Analysis & Prevention 31 (3): 181–198. McKenna, P., Jefferies, L., Dobson, A., and Frude, N. (2004). The use of a cognitive battery to predict who will fail an on-­road driving test. British Journal of Clinical Psychology 43 (3): 325–336.

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McLeod, D.R., Griffiths, R.R., Bigelow, G.E., and Yingling, J. (1982). An automated version of the digit symbol substitution test (DSST). Behavior Research Methods & Instrumentation 14 (5): 463–466. Mets, M.A., Kuipers, E., de Senerpont Domis, L.M. et al. (2011). Effects of alcohol on highway driving in the STISIM driving simulator. Human Psychopharmacology: Clinical and Experimental 26 (6): 434–439. Michon, J.A. (1985). A critical view of driver behavior models: What do we know, what should we do? Human Behavior and Traffic Safety (pp. 485–524), Boston, MA: Springer. Morrison, M.W., Gregory, R.J., and Paul, J.J. (1979). Reliability of the Finger Tapping Test and a note on sex differences. Perceptual and Motor Skills 48 (1): 139–142. Moskowitz, H. (1973). Laboratory studies of the effects of alcohol on some variables related to driving. Journal of Safety Research 5 (3): 185–199. Mullin, B., Jackson, R., Langley, J., and Norton, R. (2000). Increasing age and experience: are both protective against motorcycle injury? A case– control study. Injury Prevention 6 (1): 32–35. Nabi, H., Guéguen, A., Chiron, M. et  al. (2006). Awareness of driving while sleepy and road traffic accidents: prospective study in GAZEL cohort. BMJ: British Medical Journal 333 (7558): 75. Nouri, F.M., and Lincoln, N.B. (1993). Predicting driving performance after stroke. BMJ: British Medical Journal 307 (6902): 482. Owen, A.M., Downes, J.J., Sahakian, B.J. et al. (1990). Planning and spatial working memory following frontal lobe lesions in man. Neuropsychologia 28 (10): 1021–1034. Owen, A.M., McMillan, K.M., Laird, A.R., and Bullmore, E. (2005). N-­back working memory paradigm: A meta-­analysis of normative functional neuroimaging studies. Human Brain Mapping 25 (1): 46–59. Patomella, A.-­H., Caneman, G., Kottorp, A., and Tham, K. (2004). Identifying scale and person response validity of a new assessment of driving ability. Scandinavian Journal of Occupational Therapy 11 (2): 70–77. Patton, J.H., Stanford, M.S., and Barratt, E.S. (1995). Factor structure of the Barratt impulsiveness scale. Journal of Clinical Psychology, 51 (6): 768–774. Piersma, D., Fuermaier, A.B., De Waard, D. et al. (2018). Assessing fitness to drive in patients with different types of dementia. Alzheimer Disease and Associated Disorders 32 (1): 70. Piersma, D., Fuermaier, A.B., De Waard, D. et al. (2016). Prediction of fitness to drive in patients with Alzheimer’s dementia. PLoS one 11 (2): e0149566. Porath-­Waller, A.J. and Beirness, D.J. (2014). An examination of the validity of the standardized field sobriety test in detecting drug impairment using data from the drug evaluation and classification program. Traffic Injury Prevention 15 (2): 125–131. Raes, E. and Verstraete, A.G. (2005). Usefulness of roadside urine drug screening in drivers suspected of driving under the influence of drugs (DUID). Journal of Analytical Toxicology 29 (7): 632–636. Ramaekers, J.G. (2017). Drugs and driving research in medicinal drug development. Trends in Pharmacological Sciences 38 (4): 319–321. Ramaekers, J.G., Kauert, G., Theunissen, E. et al. (2009). Neurocognitive performance during acute THC intoxication in heavy and occasional cannabis users. Journal of Psychopharmacology 23 (3): 266–277.

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van der Sluiszen, N.N., Vermeeren, A., Jongen, S. et al. (2016). On-­the-­ road driving performance after use of the antihistamines mequitazine and l-­ mequitazine, alone and with alcohol. Psychopharmacology 233 (18): 3461–3469. van der Sluiszen, N.N., Vermeeren, A., Verster, J.C. et al. (2019). Driving performance and neurocognitive skills of long-­term users of benzodiazepine anxiolytics and hypnotics. Human Psychopharmacology: Clinical and Experimental 34 (6): e2715. Vandenbroucke, J.P. and Pearce, N. (2012). Case–control studies: Basic concepts. International Journal of Epidemiology 41 (5): 1480–1489. Vermeeren, A. (2004). Residual effects of hypnotics. CNS Drugs 18 (5): 297–328. Verster, J.C., Bekker, E.M., de Roos, M. et  al. (2008). Methylphenidate significantly improves driving performance of adults with attention-­ deficit hyperactivity disorder: A randomized crossover trial. Journal of Psychopharmacology 22 (3): 230–237. Verster, J.C. and Roth, T. (2011). Standard operation procedures for conducting the on-­the-­road driving test, and measurement of the standard deviation of lateral position (SDLP). International Journal of General Medicine 4: 359.

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Effects of Cardiovascular Disease on Fitness to Drive

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safety risks associated with cardiovascular diseases from a current medical perspective and to formulate appropriate guidelines. All EU Member States were invited to nominate an expert from their respective country, and the working group was formally established in June 2010. The Working Group comprised 12 experts from ten Member States, including Norway and Switzerland. The experts were coming from the field of cardiovascular disorders (representing the European Society of Cardiology) and from the licensing regulatory authorities. In addition, the Working Group also consulted with external experts. The Report of the Expert Group for Driving and Cardiovascular Diseases was adopted by the European Commission and came into force on 7 July 2016 in the Official Journal of the European Union (Directive 2016/1106 of 7  July 2016 amending Directive 2006/126/EC of the European Parliament and of the Council on driving licences). According to article 2 of the European Directive 2016/1106, all European countries had to issue and publish legal and administrative regulations latest by 1 January 2018. The general rules for participation in road traffic are specified in the German Driving Licence Regulations (FeV). The assessment of fitness to drive motor vehicles is, in addition to Annex 4 of the FeV, governed by the guidelines of the German Federal Highway Research Institute, which implements the requirements of the European Union in Germany. By anchoring the assessment guidelines on fitness to drive in the FeV (Annex 4) and the publication in the Traffic Journal, the guidelines have  a normative character. On December 31, 2019, the 14th

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54.1.1  New standards for driving with cardiovascular diseases: European perspective and current guidelines of the German Federal Highway Research Institute

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54.1 Introduction

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The recommendations in this chapter are guidelines for the minimum standards required for fitness to drive in applicants or drivers with cardiovascular disease(s). The recommendations for driving are primarily based on the Report of the Expert Group for Driving and Cardiovascular Diseases that formed the basis for suggested amendments to the Annex III to Directive 2006/126/ EC on driving licences (Directive 2006/126/EC of the European Parliament and of the Council of 20 December 2006 on driving licences). The provisions on cardiovascular diseases in the current Annex III, which lays down minimum standards of physical and mental fitness for driving, was last revised in 1991 and are generic in nature. A medical opinion delivered by the European Society of Cardiology and the European Heart Rhythm Association to the Commission emphasised the need to update the provisions to scientific progress and new forms of treatment, like the use of implantable cardioverter defibrillators. Against this background, the Driving Licence Committee agreed in February 2010 to establish a Working Group on Driving and Cardiovascular Diseases. The Working Group’s objective was to assess the road

Handbook of Forensic Medicine, Volume 3, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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There are no prospective, controlled studies where patients have been randomly assigned to permit or proscribe the driving privilege, or where patients have been randomly assigned to receive or not receive a physician’s advice not to drive. Furthermore, the defined standard of risk used in this chapter, while sensibly derived, is arbitrary and was not based on any evidence other than what had been acceptable historically. Given that all recommendations for driving eligibility are based on a comparison with this arbitrary standard, they are based on expert opinion only (Jung 2012). Wherever possible, best evidence was used to calculate the risks of driving, but it should be noted that the evidence itself does not support or deny driving licence restrictions for cardiac patients or the mandatory reporting of such patients by their physicians (Simpson et al. 2004). In this chapter, an effort has been made to consider the inherently subjective nature of society’s tolerance for risk, while also applying a scientifically based risk assessment mechanism in an effort to make the recommendations not just acceptable to society, but also consistent and justifiable.

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Directive on driving licences (Directive 91/439/EEC of 29 July 1991) further harmonised the standards at EU level. In its Annex III, the Directive distinguished between two groups of drivers (Report of the Expert Group for Driving and Cardiovascular Diseases 2013): • Group 1 is formed by drivers of vehicles of categories A, A1, A2, AM, B, B1 and BE. This comprises drivers of, e.g. motor cycles, passenger cars and other small vehicles with or without a trailer. • Group 2 is formed by drivers of vehicles of categories C, CE, C1, C1E, D, DE, D1 and D1E. This includes drivers of, e.g. vehicles over 3.5 tonnes or vehicles designed for the carriage of more than nine passengers with including the driver. This chapter uses the definitions used in the Driving licence Directive to distinguish between professional driving (group 2) and private driving (group 1). The experts in the Working Group strongly believe that, regardless of groups, particular consideration must be given to certain drivers such as drivers of taxis, ambulances and other professional drivers who spend many hours per day behind the wheel or carry passengers most of the time as they should be considered at higher risk. Under this perspective, the recommendations made for group 2 could also be applied to these drivers. Clinical judgement should prevail in borderline cases, for example for drivers of a small truck, where driving does not constitute the drivers’ main activity or where it is for leisure activities.

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a­ mendment of the FeV and other road traffic regulations came into force with the newly revised Chapter 3.4 on “Cardiovascular diseases” (Jung et  al. 2018; Gräcmann and Albrecht  2019; Cooper et al. 2020).

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54.1.5  Medical aspects of fitness to drive

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54.1.3  Psychosocial issues of driving restriction and adherence to recommendations

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The negative effects of driving restrictions have been of concern when outlining recommendations for driving for patients with cardiovascular diseases. Additional burden on recipients and their family’s needs to be avoided. At the same time, adherence to advice given by healthcare professionals needs to be maximised. As the driving restrictions can make the life situation of the patient and their families more difficult, this may affect adherence to the recommendations. Several studies (Jung and Lüderitz 1996a; Conti et al. 1997; Mylotte et al. 2013) point in the direction of low adherence amongst recipients to the driving ban advised by healthcare professionals. As there seems to be a gap between recommendations and patient adherence to these recommendations, an adequate education and follow-­up of patients and family is pivotal.

54.1.4  Definition of private drivers and professional drivers In the EU, the First Directive on driving licences (Directive 80/1263/EEC of 4 December 1980) proposed the establishment of a common European driving licence. The Second

Driving is a universal activity in all developed nations. It has been estimated that ordinary drivers of private vehicles, depending on age and occupation, may spend an average of 250 hours a year at the wheel. For professional or vocational drivers, the figure is many times higher (Petch 1998). In modern societies, the motor car has become an essential part of daily living. People who live in rural communities rely on the car for such things as getting to work, shopping and visiting. In all countries, some regulation of driving activity is usual with insistence on a level of competence and medical fitness. To deny an individual an ordinary driving licence may seriously restrict their lifestyle and should not be undertaken lightly. Regulations therefore have to strike a balance between the liberty of the individual and the threat that individual might pose to others by virtue of being a potential cause of a road traffic accident (Jung and Lüderitz 1996b). Road traffic accidents are the commonest cause of death in young people and account for a significant minority of deaths each year in most developed countries, for example in 1995 approximately 40 000 in the USA, 10 000 in Germany and 4000  in the UK. According to the German Federal Highway Research Institute, a significant decline of lethal traffic accidents has been observed. In 2019, 3046 fatal road accidents occurred in Germany. There is, in addition, a substantial morbidity and a significant economic impact, both of which are very difficult to estimate (Petch 1998).

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injury-­producing accident (Ac). Expressing this statement as a formula:

RH TD x V x SCI x Ac

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Fewer than 2% of reported incidents of driver sudden death or loss of consciousness have resulted in injury or death to other road users or bystanders. In the formula, therefore, Ac = 0.02 for all drivers. There is evidence that loss of control of a heavy truck or passenger-­carrying vehicle results in a more devastating accident than loss of control of a private automobile. Truckers are involved in only about 2% of all road accidents but in approximately 7.2% of all fatal accidents. In the formula, if V = 1 for a commercial driver, then V = 0.28 for a private driver. There is no published standard or definition of what level of risk is considered acceptable to society even through this is crucial in the formulation of guidelines based on the probability of some event occurring in a defined time period. It was necessary, therefore, to develop such a standard. For several years, the guidelines of the CCS, the Canadian Medical Association and the Canadian Council of Motor Transport Administrators, have permitted the driver of a heavy truck to return to that occupation following an acute myocardial infarction (MI) provided that he or she is functional class I with a negative exercise stress test at seven metabolic equivalents, has no disqualifying ventricular arrhythmias and is at least 3 months post infarct. On the basis of available data, however, such a person cannot be assigned a risk lower than 1% of cardiac death in the next year. The risk of sudden death would be lower than this but is at least partially offset by the risk of other suddenly disabling events such as syncope or stroke. For such a person, SCI is estimated to be equal to 0.01 in the formula. It may be assumed that the average commercial driver spends 25% of his or her time behind the wheel. Thus, in the formula, TD = 0.25. As indicated above, V may be assigned a value of 1 for commercial drivers, with an Ac of 0.02 for all drivers. Substituting into the formula:

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European data suggest that approximately 1.5–3.4% of road accidents are attributed to sudden driver incapacity, with only a minority being arrhythmia-­related (Jung et  al. 1997). Based on the Canadian experience, only 5% of fatal road accidents were due to an unexpected medical condition. Fatigue, alcohol and drugs accounted for a large number of accidents, with medical causes being infrequent. Age is another factor to consider; an increased risk of death amongst the young and elderly has already been accepted by society for years. It is of note that falling asleep, fatigue and alcohol represent a much greater risk for death and injury at the wheel than sudden incapacitation caused by ventricular tachyarrhythmias (Jung and Lüderitz 1996c). Given the inherent difficulty of determining an arrhythmic cause for accidents, currently available data can only be considered a rough estimate. These data do not convincingly show that sudden cardiac death (SCD) while driving is a major public safety issue, but rather they show it is a rare event and that fewer than 2% of sudden driver incapacitations result in death or injury to other road users or innocent bystanders (Jung et  al. 1997). Whether high-­ risk populations (e.g. implantable cardioverter-­ defibrillator (ICD) patients) are responsible for a higher frequency of arrhythmia-­related motor vehicle accidents has not yet been proven (Lüderitz and Jung  1996). In order to establish driving regulations for ICD patients, it seems reasonable that these recommendations should be based on an actuarial approach, such as that pioneered in the field of aviation medicine.

• The probability that such an event will result in a fatal or

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54.2 Derivation of the risk of harm formula

RH TD V SCI Ac 0.25 1 0.01 0.02 0.00005

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In 1992, the Canadian Cardiovascular Society (CCS) consensus conference document ‘Assessment of the cardiac patient for fitness to drive’ was published (Brennan et al. 1992). Four years later, as a result of significant advances in the investigation and management of both arrhythmias and syncope, an update was deemed necessary by the CCS Task Force that penned the original document. This document has served as the standard of assessment since that time (Brennan et  al. 1996). In 2003, the membership perceived that a further update Fitness to Drive and Fly was required, since significant developments had again occurred in the evaluation and treatment of cardiac disorders, rendering some of the recommendations obsolete (Simpson et al. 2004). In 1992, the CCS consensus conference developed a risk of harm (RH) formula to quantify the level of risk to other road users posed by a driver with heart disease (Brennan et al. 1992). This RH is assumed to be directly proportional to the following (Simpson et al. 2004): • Time spent behind the wheel or distance driven in a given time period (TD). • Type of vehicle driven (V). • Risk of sudden cardiac incapacitation (SCI).

Allowing such a driver on the road is associated with an annual risk of death or injury to others of approximately 1 in 20 000 (0.00005). This level of risk appears to be generally acceptable to society. A similar standard may be applied to the driver of a private automobile. The average private driver spends approximately 4% of his or her time behind the wheel (TD = 0.04). As indicated above, for such a driver, V = 0.28 and Ac = 0.02. The acceptable yearly risk of sudden death or cardiac incapacitation for such a person would be calculated as follows:



RH TD x V x SCI x Ac 0.00005 0.04 x 0.28 x SCI x 0.02 SCI 0.223

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54.3.1  Hospital mortality and prognosis Registry data consistently show that NSTE-­ACS is more frequent than STE-­ACS (Hamm et al. 2011). The annual incidence is about three per 1000  inhabitants, but this varies between countries. Hospital mortality in the first 15 days is higher in patients with STEMI than amongst those with NSTE-­ ACS (7% vs. 3–5%, respectively), but then a steadily decrease occurs (Fox et al. 2006). At 1 month, the mortality rates are 9% for STEMI patients and approximately 6% for NSTEMI patients. At 6 months, the mortality rates are very similar in both conditions (12% and 13%, respectively). After the 15th day following MI, mortality is less than 22% over a calculated 1-­year period, thus the yearly risk for sudden cardiac incapacitation is less than the 22% accepted risk boundary for a private driver. Long-­term follow-­up showed that death rates were higher amongst patients with NSTE-­ACS than with STE-­ACS, with a twofold difference at 4 years. This difference in mid-­and long-­term evolution may be due to different patient profiles since NSTE-­ACS patients tend to be older, with more co-­morbidities, especially diabetes and renal failure. In the CAD population, SCD is a well-­recognised phenomenon. While death is termed ‘sudden’ in epidemiological literature, it may not be instantaneous; there may be preceding symptoms. Clearly, the patient who experiences chest pain during driving does not pose the same risk as the driver who has a sudden ventricular arrhythmia while behind the wheel. However, ventricular arrhythmias may be more likely to occur during the first 24 hours of an ACS. But, it is neither reasonable nor practical to impose a driving restriction on all people at risk for developing a coronary syndrome, given its high prevalence in our society and given our inability to predict the timing of an occurrence with any degree of accuracy (CCS 2003). Finally, most acute coronary syndromes are not associated with sudden cardiac incapacitation. In the Randomised Intervention Trial of unstable Angina 3 (RITA) study, patients were randomly assigned to coronary angiography and appropriate intervention (n = 895) or to a symptom-­ guided conservative strategy (n = 915) (Poole-­Wilson et al. 2006). The death rate declined over time. In both groups combined, the death rate in months 1, 2–4 and 5–12  was 0.48, 0.15 and 0.07 deaths/1000  days of follow-­up, respectively. In the first week, patients in both groups were at highest risk of death, MI or

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Cardiovascular diseases are currently the leading cause of death in industrialised countries and are expected to become so in emerging countries by 2020 (Table 54.1). Amongst these, coronary artery disease (CAD) is the most prevalent manifestation and is associated with high mortality and morbidity. The clinical presentation of CAD includes stable angina pectoris and the acute coronary syndrome that covers the spectrum of unstable angina, non-­ST-­elevation MI (NSTEMI) and ST-­elevation MI (STEMI) (Hamm et al. 2011). MI is characterised by well-­known clinical, electrocardiographic (ECG), biochemical and pathological characteristics. • Patients with acute chest pain and persistent (>20 minutes) ST segment elevation. This is termed ST-­elevation acute coronary syndrome (STE-­ACS) and generally reflects an acute total coronary occlusion. Most of these patients will ultimately develop a STEMI. The therapeutic objective is to achieve rapid,

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c­ omplete and sustained reperfusion by primary angioplasty or fibrinolytic therapy. • Patients with acute chest pain but without persistent ST segment elevation. These patients have rather persistent or transient ST segment depression or T-­wave inversion, flat T waves, pseudo-­ normalisation of T waves or no ECG changes at presentation. The initial strategy in these patients is to alleviate ischaemia and symptoms, to monitor the patient with serial ECGs and to repeat measurements of markers of myocardial necrosis. The working diagnosis of non-­ ST-­ elevation ACS (NSTE-­ ACS), based on the measurement of troponins, will be further qualified as NSTEMI or unstable angina.

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Thus, the private automobile driver with a 22% risk of sustaining a sudden cardiac incapacitation in the next year poses no greater threat to public safety than the heavy truck driver with a 1% risk. Finally, for the commercial driver who drives a light vehicle, such as a taxicab or delivery truck, V = 0.28 and­ TD = 0.25, placing them at a risk between that of the private driver and the heavy truck driver.

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Table 54.1  Coronary heart disease (CAD) Group 1

Group 2

Acute coronary syndrome (ACS)

Driving may be allowed provided free of symptoms. In case of significant myocardial damage driving is allowed after four weeks

Driving may be allowed six weeks after the acute event provided free of symptoms; and exercise or other functional test requirements can be met.

Percutaneous coronary intervention (PCI)

Driving may be allowed after elective PCI, if good clinical outcome

Driving may be allowed four weeks after elective PCI if good clinical outcome provided the applicant or driver is free of symptoms, and that the functional test requirements are met.

Coronary artery bypass graft (CABG)

Driving may be allowed after sufficient wound healing and clinical recovery.

Driving may be allowed after sufficient wound healing, clinical recovery and functional test requirements are met.

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endothelial healing, strut fracture, late acquired malapposition and neoatherosclerosis are proposed pathophysiological mechanisms associated with drug-­eluting stent (DES) thrombosis. The relative safety of DES and bare metal stents, especially with respect to ST, continues to be debated. Better selection of stent size, higher insufflation pressures, more frequent non-­compliant balloon post-­dilatation, and intravascular ultrasound are currently used for correct stent expansion. Data from a comprehensive network meta-­ analysis including 49 trials with 50 844 patients randomly assigned to treatment groups showed that 1-­year definite ST was significantly lower with cobalt–chromium everolimus-­ eluting stents (CoCr-­ EESs) than with bare metal stents (odds ratio 0.23). The significant difference in ST between CoCr-­EESs and bare metal stents was evident as early as 30 days (odds ratio 0.21) and was also significant between 31 days and 1 year (odds ratio 0.27) (Palmerini et al. 2012a). EESs compared with other DESs reduced the relative risk of early ST (within 30 days), late ST (31 days to 1 year), cumulative 1-­year ST and very late ST (1–2 years) (Palmerini et al. 2012b). At 30-­day follow-­up, there were 15 episodes (0.2%) of early definite ST amongst the 9294 patients treated with an EES and 42 (0.6%) amongst the 6857 patients treated with a paclitaxel-­eluting stent (PES), sirolimus-­eluting stent (SES) or zotarolimus-­eluting stent (ZES). Use of an EES was associated with a significant reduction in early definite ST (relative risk (RR) 0.28) compared with pooled PES, SES and ZES stents. There were 10 episodes (0.1%) of late definite ST amongst the 8515 patients treated with EESs and 24 episodes (0.4%) of late definite ST amongst the 6111 patients treated with PES, SES and ZES stents. At 1-­year follow-­up, there were 25 episodes (0.3%) of definite ST amongst the 9512 patients treated with EESs and 66 (0.9%) amongst the 6963 patients treated with PES, SES or ZES stents. At 2-­year follow-­up, there were 32 episodes (0.5%) of definite ST amongst the 6825 patients treated with EESs and 68 (1.3%) amongst the 5109 patients treated with PES, SES or ZES stents. EES use compared with a pooled group of PES, SES and ZES use is associated with a significant reduction of definite ST, an effect that appears early and increases in magnitude through at least 2 years. Earlier data from a large, two institution cohort study (Daemen et al. 2007) showed that late ST occurred steadily at a constant rate of 0.6% per year up to 3 years after stent implantation. The incidence of early ST was similar for SES (1.1%) and PES (1.3%) stents.

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r­ efractory angina (incidence rate 40 times higher than in months 5–12 of follow-­up). Within 28  days, 24 patients died in both groups, of which SCD accounted for only 3 of 15 deaths in the interventional group and 3 of 9 deaths in the conservative group, respectively. The risk of sudden death from cardiac causes is increased amongst survivors of acute MI with reduced left ventricular systolic function. In the Valsartan in Acute Myocardial Infarction Trial (VALIANT) study (Solomon et al. 2005), this risk was highest in the first 30 days after MI (1.4% per month) and decreased to 0.14% per month after 2 years. Patients with a left ventricular ejection fraction of ≤30% were at highest risk in this early period (2.3% per month). Nineteen per cent of all sudden deaths or episodes of cardiac arrest with resuscitation occurred within the first 30 days after MI, and 83% of all patients who died suddenly in the first 30 days did so after hospital discharge. Each decrease of 5% points in the left ventricular ejection fraction was associated with a 21% adjusted increase in the risk of sudden death or cardiac arrest with resuscitation in the first 30 days. The risk of SCD following MI in community practice has declined significantly over the past 30 years. In the Olmsted County study (Adabag et al. 2008), SCD was independently associated with heart failure but not with recurrent ischaemia. The 30-­ day cumulative incidence of SCD was 1.2%, which was fourfold higher than expected. Thereafter, the rate of SCD was constant at 1.2% per year, yielding a 5-­year cumulative incidence of 6.9%. In the current therapeutic era, SCD accounts for about one-­third of cardiovascular deaths after NSTE ACS. Risk stratification can be performed with good accuracy using commonly collected clinical variables. Clinical events occurring after the index hospitalisation are underappreciated but important risk factors (Hess et al. 2016).

Stable coronary artery disease

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In the Prevention of Events with Angiotensin Converting Enzyme Inhibition (PEACE) trial (Hsia et  al. 2008), 8290 patients with stable CAD and preserved left ventricular systolic function ≥40% were randomised to trandolapril or placebo. The overall incidence of SCD was 0.32% per year, contrasting with 0.70% per year in the Heart Outcomes Protection Evaluation trial (HOPE) study (Teo et al. 2004) and 0.67% per year in the study published by Ikeda et al. (2006).

Percutaneous coronary interventions: in-­ stent restenosis and stent thrombosis In-­stent restenosis and stent thrombosis remain the major limitations of percutaneous coronary intervention. In-­stent restenosis reflects a complex underlying pathophysiology that involves various combinations of residual coronary stenosis, recoil and neointimal proliferation. Symptom status is an unreliable predictor of restenosis since many patients complain of non-­cardiac pain after angioplasty. Stent thrombosis (ST) is usually associated with symptoms. Persistent inflammation, hypersensitivity reactions, delayed

54.4 Arrhythmias 54.4.1  Bradycardia with and without cardiac pacing The safety of a patient driving with the diagnosis of a bradyarrhythmia is dependent on the presence of syncope before ­placement of the pacemaker, the reliability of the pacing system and the underlying cardiac pathology. Bradyarrhythmias can be

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Patients with supraventricular tachycardias that appear to be successfully ablated may drive after recovery from the procedure because the risk of arrhythmia recurrence and risk of injury from recurrence are low.

54.4.3 Ventricular tachycardia

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There is no reason a priori to make different recommendations for patients with ventricular tachycardia (VT) or ventricular fibrillation (VF) whose primary treatment is anti-­arrhythmic drug therapy rather than ICD therapy (Epstein et  al. 1996). Patients with sustained VT or VF who are treated with anti-­ arrhythmic drugs should be prohibited from non-­commercial driving for the first 6 months after initiation of therapy to document an arrhythmia-­free interval before they are allowed to resume driving. If a second episode of ventricular arrhythmia occurs after resuming driving, then another 6-­month period of abstinence from driving should be recommended to provide sufficient time to judge whether the changes in medical therapy have adequately suppressed arrhythmia recurrences. As with ICD-­ treated patients, it is recommended that all commercial driving be permanently prohibited in patients with VT or VF who are treated primarily with anti-­arrhythmic drugs. It is probably reasonable to make exceptions to the above recommendations for patients who have idiopathic VT and who do not have symptoms of impaired consciousness with their presenting arrhythmia. Such patients should be shown by extensive evaluation to have normal ventricular function, normal coronary arteries, no evidence of hypertrophic cardiomyopathy and no evidence of right ventricular dysplasia. Because such patients do not have obstructive CAD, they are unlikely to develop enough myocardial ischaemia during their arrhythmia to render the tachycardia electrically unstable; the degeneration of VT to VF in such patients is rare (Epstein et al. 1996). Accordingly, it is likely that if the patient has not had symptoms of impaired consciousness with the presenting arrhythmia, he or she will tolerate future episodes equally well. Thus, in selected individuals, a shorter period of driving restriction may be appropriate once medical therapy has been initiated.

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divided into those with and those without syncope or presyncope. Patients with symptoms require cardiac pacing to drive. Those without symptoms do not need a pacemaker and can drive for as long as they are symptom free. Patients who have cardiac pacemakers are unlikely to have further symptomatic bradycardia, and if symptoms recur, they are unlikely to be due to pacemaker malfunction. The risk of further driving is dependent on the underlying cardiac pathology and is more likely to be related to tachyarrhythmia than bradyarrhythmia (Epstein et al. 1996). Pacemakers implanted for neurally mediated syncope may fail to prevent recurrent syncope if there is a prominent vasodepressor component. Frequently, pacemakers are implanted to increase cardiac output above resting levels but not to prevent syncope. It is inappropriate to restrict the driving of persons with a pacemaker; indirect support for this position lies in the absence of problems observed. Indeed, when pacemakers were first used, concerns about driving were raised but disappeared when no problem was seen. For patients who have lost consciousness due to a bradyarrhythmia, a period of time should pass to ensure stable lead function before they return to driving; this should be 1  week for non-­commercial drivers and 4  weeks for commercial drivers because commercial drivers drive larger vehicles for a longer time duration and they may also have greater physical demands (e.g. loading and unloading a truck). Pacemaker patients followed according to the published follow-­up guidelines with pacemakers programmed to an appropriate pacing safety margin provide adequate assurance that the pacemaker will protect the patient from syncope due to bradycardia.

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54.4.2 Supraventricular tachycardia

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The supraventricular tachycardias comprise the atrial tachycardias, including focal atrial tachycardias, atrial fibrillation, atrial flutter, atrioventricular re-­entry and atrioventricular nodal re-­ entry. Although the general prognosis in patients with the Wolff– Parkinson–White syndrome and other supraventricular tachycardias is generally quite favourable, at least a single episode of syncope is reported in approximately 25% of patients referred to electrophysiology laboratories for assessment (Epstein et  al. 1996). Sudden death is rare as the first manifestation of the Wolff–Parkinson–White syndrome and is seen only sporadically excitation. in asymptomatic individuals with ventricular pre-­ Atrial tachyarrhythmias in young patients following the repair of congenital heart disease can be associated with severe symptoms. Atrial fibrillation is the most common supraventricular arrhythmia. Syncope related to atrial fibrillation is readily recognised in most instances as resulting from intermittent bradycardia or, conversely, a rapid ventricular rate. There are no data documenting the frequency with which ­syncope related to supraventricular tachycardia causes motor vehicle accidents, but it is probably rare. A wide range of catheter ablative, operative and medical therapies offer definitive cure or excellent control of symptoms with supraventricular t­ achycardias.

54.4.4  Implantable cardioverter defibrillators The guidelines for ICD implantation and the management of patients with ventricular arrhythmias and the prevention of SCD have been revised in recent years (Jung  1995; Jung et  al. 2006; Priori et  al. 2015). In 1997, the Working Groups on Cardiac Pacing and Arrhythmias of the European Society of Cardiology published recommendations on driving by patients with ICDs (Jung et  al. 1997). In 1996, in the USA, the American Heart Association developed a scientific statement entitled ‘Personal and public safety issues related to arrhythmias that may affect consciousness: implications for regulation and physician

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Group 2

ICD implantation for secondary prevention

Driving must cease for 3 months

Permanent driving ban

ICD implantation for primary prevention

Driving must cease for 2 weeks

Permanent driving ban

ICD therapy: appropriate

Driving must cease for 3 months

Not applicable (permanent driving ban)

ICD therapy: inappropriate

Driving must cease until measures are taken to prevent subsequent inappropriate therapy

Not applicable (permanent driving ban)

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r­ecommendations’ (Epstein et  al. 1996). Both reports mainly focused on patients with secondary prevention and recommended a driving restriction of 6 months after ICD implantation for this patient population (Table 54.2). Since the original publication of these medical/scientific statements, multiple trials have established the role of ICDs for the primary prevention of SCD in patients at risk for life-­threatening ventricular arrhythmias who have never had sustained VT or VF. In 2007, an addendum was developed in the United States to extend the original 1996 recommendations and to provide updated guidelines regarding individuals with ICDs implanted for primary prevention who may undertake activities, specifically driving, that may put themselves or others at risk if consciousness were to be impaired by a cardiac arrhythmia. Furthermore, in 2009 a consensus statement of the European Heart Rhythm Association presenting updated recommendations for driving by patients with ICDs was published (Vijgen et al. 2009). Factors that determine the risk of harm are as follows: • The frequency and the time course of arrhythmia recurrence. • The likelihood that such recurrences are associated with impaired consciousness. • The risk that such an event will cause an accident. • The probability that such an accident will result in death or injury to the patient and other road users, innocent bystanders or passengers.

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Group 1

predictor of an increased risk of first arrhythmia occurrence is a severely depressed left ventricular ejection fraction, leading to an actuarial incidence of the first ICD therapy at least two to three times greater than that of patients with well-­preserved left ventricular function. The absence of ICD therapy during the first year of ICD implantation has been associated with a lower risk of arrhythmia recurrence, ranging from approximately 10% to 20% annually. In contrast, patients who experience ICD therapy are at increased risk of receiving further ICD discharges during the subsequent years, with an estimated actuarial incidence in the order of 20–40% in the next 3 years. Published data suggest that the occurrence of a first shock did not predict a subsequent period of freedom from a second shock. Furthermore, the second shock-­free survival curve indicated that the majority of patients who will experience a second shock did so within 6 months or less of the first shock. The results of another study supported the findings that subsequent ICD discharges occurred earlier than the first ICD therapy and were not predictable by any clinical variable (Jung et al. 1997). Unlike the occurrence of first shock therapies, no clinical variable identifies a group at lower risk for subsequent ICD therapies. Data from the Polish registry of 2162 patients implanted for secondary prevention of SCD showed that the probability of ICD intervention for VF or fast VT during 10 years of follow-­up was 52.3% (Vijgen et  al. 2009). The mean time to first intervention was 344 ± 416 days. Fifty per cent of patients had an appropriate ICD intervention during the first 194  days after implantation. The probability of arrhythmic episodes was 1.9% in the first month, 3.3% in the second month and 3.7% in the third month. In the 3 months thereafter, the added probability remained below 2% per month. Although guidelines recommend driving restrictions for 3–6 months after appropriate ICD shocks, contemporary data to support these recommendations are lacking. To define the time course of subsequent shocks after an initial ICD discharge, a retrospective analysis of a nationwide cohort of 14 230 ICD recipients enrolled in a remote monitoring program was undertaken. In this large cohort of ICD recipients for primary and secondary prevention, the incidence of a second shock after an initial ICD discharge was lower than previously reported and is dependent on several programmed ICD variables, including the number of therapy zones and lowest heart rate for which therapy is delivered (Merchant et al. 2016).

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Table 54.2  Implantable cardioverter-­defibrillator (ICD).

Risk of recurrence of arrhythmia in patients implanted for secondary prevention The 5-­year actuarial incidence of appropriate ICD shocks varies somewhat from study to study, ranging from 55% to 70%, and may also be influenced by the study population receiving the ICD. Several studies have shown an incidence of ICD therapy delivery of approximately 50% within the first year of ICD implantation, after which hazard rates calculated for consecutive 12-­month intervals decline markedly (Epstein et  al. 1996; Jung et al. 1997). The initial 6 months are the period of highest risk for appropriate as well as inappropriate shock therapy. A powerful

Risk of syncope in patients for secondary prevention

implanted

Although actuarial data for the incidence of ICD therapy delivery are available, this does not necessarily equate with the incidence of incapacitation. It is important to stress that most patients with sustained ventricular arrhythmias experience only minimal or no symptoms prior to ICD discharge (Jung et al. 1997). The likelihood that the treatment of a ventricular tachyarrhythmia in an ICD patient will be associated with disabling symptoms or even syncope has been addressed in a study by Kou et  al. (1991) in

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DiCarlo et al. (1992) reviewed driving restrictions in the mid-­ west USA, and compatibility with state laws. Fifty-­three per cent of responding cardiologists only advised those ICD patients who had arrhythmia-­induced presyncope or physical collapse to cease driving. The remainder advised all implanted patients to stop driving. Most cardiologists recommended a temporary driving abstinence for a period of 2–12 months (6 ± 3 months). Minimal legal requirements in the mid-­west states were variable. Conti et al. (1997) surveyed 82 patients who were followed for 6 ± 1.3 years and drove 33 ± 43.5  km a day (20.5 ± 27  miles/day). All patients in this group had defibrillator shocks. Ninety per cent of the 52 patients resumed driving, and none experienced device discharge while driving during the follow-­up time period. Curtis et al. (1995) surveyed 742 US physicians who followed defibrillator patients: 452 physicians responded, and a total of 30 motor vehicle accidents related to shocks from implanted defibrillators were reported by 25 physicians over a 12-­year period. The estimated fatality rate for patients with a defibrillator was 7.5 per 100 000 patient-­years, significantly lower than for the general population (18.4 per 100 000 patient-­years). The injury rate for ICD patients was also very substantially less than for the general public, 17.6 versus 2224 per 100 000 patient-­years. Of 286 defibrillator discharges documented while driving, 10.5% resulted in an accident. Trappe et al. (1998) examined the driving behaviour of 291 ICD patients. Fifty-­nine per cent of 241 patients continued driving post implant and were followed for 38 ± 26 months. No patients died while driving; there were 11 accidents, but only 1 caused by the driver with an ICD, and none were related to syncopal symptoms or ICD therapy. Five per cent of all patients over this time received ICD therapy while driving; 74% of these occurred more than 2 years post implant. No patient had syncope or an accident with this event. Larsen et al. (1994) followed 511 patients with an implanted defibrillator for a mean of 26 months. The 1-­year event rate for all patients was 17%. The monthly hazard rate for defibrillator discharge was 4.22% per month in the first month, declining to 1.81% per month at months 2–7 and subsequently to 0.63% per month. This risk of defibrillator discharge per month was only slightly higher than the risk of any traffic accident involving death, injury or major damage amongst all licenced Oregon drivers (0.4% per month) or drivers aged 16–19 (0.9% per month). Since only 8% of the patients in this study were treated with an ICD, these data predominantly refer to patients receiving anti-­arrhythmic drug treatment. Akiyama et al. (2001) administered questionnaires regarding driving to 909 patients in the AVID study. Of the 758 patients who responded (83% of the total), 627 drove in the year prior to their index episode of ventricular arrhythmia. Fifty-­seven per cent of these drivers resumed driving within 3 months after randomisation in the AVID trial, 78% within 6  months and 88% within 12 months. Two per cent of patients during follow-­up had a syncopal episode while driving, and 11% had dizziness or palpitations that required stopping the vehicle. Eight per cent of the patients with an ICD received a shock while driving. Of the 55 accidents during 1619 patient-­years of follow-­up after resumption of driving, 11% were preceded by any symptom of possible

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180 ICD recipients. The investigators showed that only 16 of 106 patients (15%) who received shock therapy from their ICD had syncope. Unfortunately, no clinical variables, including age, sex, history of syncope, left ventricular function, type of underlying heart disease, electrophysiological findings, rate of VT, anti-­ arrhythmic medications and type of pulse generator implanted, were found to be predictors of syncope. Kou and co-­workers showed that two-­thirds of the patients who experienced syncope in association with subsequent ICD shocks did not experience syncope during their first shock. Based on their data, they concluded that an absence of syncope during the first shock did not predict freedom from syncope during subsequent shocks. Freedberg et  al. (2001) followed 125 ICD patients for 408 ± 321 days. Sustained monomorphic VT and low ejection fraction predicted a higher risk of future ICD therapy. Patients with minimally symptomatic tachycardia at first occurrence had a high likelihood of having asymptomatic subsequent symptoms and were felt to be at very low risk for syncopal ICD therapy if the first event was asymptomatic or minimally symptomatic. Bansch et al. (1998) retrospectively analysed data on 421 patients with an ICD followed for 26 ± 18 months. Of these patients, 229 (54.4%) had recurrent VT/VF, and 62 (14.7%) had syncope. Low baseline left ventricular ejection fraction, induction of fast VT (cycle length 35%

NYHA II

No restriction

No restriction, if LVEF >35%

NYHA III

No restriction

Disqualified

NYHA IV

Disqualified

Disqualified

NYHA: New York Heart Classification.

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54.6.5  Heart failure mortality and prognosis Mortality rates amongst groups of HF patients are highly variable and range from 5% to 75% per year. The results of both the Framingham Heart Study (Levy et al. 2002) and a population-­ based study in Olmsted County, Minnesota (Roger et al. 2004), suggested decreases of age-­adjusted mortality rates in patients after the onset of HF in the last decades. However, 5-­year age-­ adjusted mortality rates after onset of HF remained high in these two studies, with higher rates in men (50% in men vs. 46% in women for the Olmsted County population-­based study). The vast majority of patients with HF die from cardiovascular causes; estimates vary from 50% to 90%, depending on the HF population studied. Furthermore, the mode of death is also divergent, where some patients die suddenly (many of ventricular arrhythmia) and others die of progressive failure of cardiac function (pump failure). Amongst cardiovascular causes of death, SCD poses a major threat, with up to 50% of HF patients dying of SCD. Importantly, the relative contribution of sudden death to total death rate decreases when the clinical severity of HF increases. It is at its maximum (50%) in patients with low left ventricular ejection fraction (LVEF) and in NYHA classes I and II, and lowest in patients with advanced HF where patients die mostly from pump failure. This is an important element influencing the relative benefit, and, therefore, the indications for ICDs. Prediction of the likely mode of death in an individual HF patient, and of the relative and absolute risks of the different modes of death amongst different HF patients, might allow more rational or cost-­ effective use of specific HF medications or

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Heart failure is associated with ischaemic heart disease (46–68%), arterial hypertension (53–66%), diabetes (27–38%), arrhythmia, especially atrial fibrillation (21–42%) and renal insufficiency (17–53%). HF is associated with serious morbidities, such as renal failure, cancer, cirrhosis, hepatic insufficiency or chronic obstructive pulmonary disease (Zannad et al. 2009).

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A common description of HF distinguishes different types of the syndrome according to its mechanism (systolic vs. diastolic), its aetiology (ischaemic vs. non-­ischaemic) or its clinical presentation (acute vs. chronic) (Zannad et al. 2009).

54.6.4  Causes and co-­morbidity of heart failure

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Mild, moderate or severe heart failure (HF) is used as a clinical symptomatic description, where mild is used for patients who can move around with no important limitations of dyspnoea or fatigue, severe for patients who are markedly symptomatic and need frequent medical attention, and moderate for the remaining patient cohort (Table 54.4). Two classifications of the severity of HF are commonly employed. One is based on symptoms and exercise capacity  – the New  York Heart Association (NYHA) functional classification. The NYHA functional classification has proved to be clinically useful, and it is employed routinely in most randomised clinical trials. The other classification describes HF in stages based on structural changes and symptoms. All patients with overt HF are in stages C and D.

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trends were reported in the Rotterdam study (Mosterd et  al. 1999): from 1% in persons aged 55–64 years to 13% in those aged 75–84 years. The rise in the incidence and prevalence of HF globally is the result of improved care of acute MI combined with the ageing of the population and the emerging pandemic of cardiovascular disease in developing countries. HF hospitalisation represents 1–2% of all hospital admissions, which makes it the leading cause of hospitalisation for patients older than 65 years. HF has a huge impact on health-­related quality of life and appears as an economic burden nowadays. It is estimated that 1–2% of all healthcare expenditure is devoted to HF in developed countries (Zannad et al. 2009).

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54.6.6  Driving restrictions Heart failure with NYHA I

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NYHA class II patients are at proportionally higher risk of sudden death and less risk of progressive HF. Based on the β-­blocker and angiotensin-­converting enzyme inhibitor trials, the annual risk of death in the treatment arm was 7.2–10.4% and the annual risk of sudden death in a NYHA class II patient was approximately 6%. These risks are within the risk of annual sudden cardiac incapacitation of 22% and, as such, patients who are NYHA class II are fit for driving a private motor vehicle. However, again, this risk would be too high to allow commercial driving where the LVEF is below 35%. For those patients with a more preserved LVEF of >35%, permission for commercial driving has to be decided individually.

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devices. Prior reports have investigated the relationship of single risk factors with mode of death in HF, but prediction models that combine the information from multiple risk factors may more optimally capture overall risk (Mozaffarian et  al. 2007). The Seattle heart failure model (SHFM) is a validated prediction model that estimates total mortality in patients with HF by using commonly obtained clinical, laboratory, medication and device variables (Levy et al. 2006). Optimal HF therapy as tolerated is appropriate across the whole spectrum of the SHFM score. The proportion of deaths that resulted from sudden death was greater in patients with lower SHFM scores because the incidence of pump failure death increased faster than sudden death with higher SHFM scores (Mozaffarian et al. 2007). In the Metoprolol CR/XL Randomised Intervention Trial in Congestive Heart Failure (MERIT-­ HF) (MERIT-­HF Study Group  1999), the proportion of deaths that resulted from sudden death in NYHA class II, III and IV patients was 64%, 57% and 33%, respectively, whereas the 1-­year absolute risk (cumulative incidence) of sudden death was 4%, 6% and 6%, respectively (only 145 patients were NYHA class IV). In the SHFM analysis, the proportion of deaths that resulted from sudden death in NYHA class II, III and IV patients was 65%, 51% and 45%, respectively, whereas the 1-­year absolute risk was 4%, 7% and 13%, respectively. In comparison, across increasing SHFM scores (0–4), the proportion of deaths that resulted from sudden death was 79%, 57%, 43%, 40% and 28%, whereas the absolute risk at 1 year was 4%, 6%, 10%, 23% and 25%, respectively (Mozaffarian et al. 2007). These findings suggest that the SHFM more finely discriminates between HF severity and related risk than does NYHA class. To determine potential optimal treatments, both absolute rates of different modes of death and proportions of deaths (competing risks) from different modes of death might be important. NYHA class is commonly used as a proxy for HF severity and for the definition of patient sub-­groups to evaluate or form recommendations with regard to specific treatments. The SHFM and NYHA class systems were compared for predicting absolute rates and proportions of different modes of death. Across categories of NYHA class, both absolute rates and proportional risks were relatively similar for patients with the same SHFM score. For example, amongst patients with an SHFM score of 2, rates of sudden death were 8, 10 and 11 per 100 person-­years for NYHA classes II, III and IV, respectively, whereas proportions of sudden deaths were 39%, 40% and 45% for NYHA classes II, III and IV, respectively. In contrast, within each category of NYHA class, the SHFM strongly predicted absolute rates and proportional risks of sudden death and pump failure (P for trend 11.1 mmol/L) to about 100 mg/dL (c. 5.6 mmol/L) often causes symptoms of hypoglycaemia. People whose treatment includes insulin, sulfonylurea and glinides are at high risk for hypoglycaemia, for example in situations of exercise or

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Hypertension is the main risk factor for cerebrovascular disease. People with diabetes have an increased prevalence and incidence for stroke. A tight control of glucose, blood pressure and lipidaemia is needed. Ultrasonic evaluation of the cerebral arteries, the carotid intima media thickness and the degree of stenosis of the carotids may lead to the diagnosis (Hanefeld et al. 2000). Diabetes-­ related peripheral vascular disease affects the arteries of the lower limbs and can cause intermittent claudication and foot problems.

Hypertension High blood pressure is twofold increased in diabetics compared to non-­diabetics. Hypertension is a major part of the metabolic syndrome. It consists of diabetes or insulin resistance, hypertension, dyslipoproteinaemia and obesity, resulting in a 3–4 time increase in the risk for CVD. Hypertension is defined as systolic ≥140 and/or diastolic ≥90 mmHg blood pressure. Optimal treatment of hypertension mostly needs a combination therapy to reach the goal of 120–129/80–84 mmHg of blood pressure (recommendation of the European Society of Cardiology and European Society of Hypertension; Williams et  al. 2018). Self-­ monitoring of blood pressure by the patient is standard. Medial sclerosis, where there is an abnormal increased stiffness of the peripheral arteries, will result in incorrect, too high reading of the blood pressure.

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Microangiopathic disorders in diabetes are nephropathy and retinopathy. Diabetic nephropathy results from long-­term hyperglycaemia. Early stages are classified by the amount of urinary loss of albumins. To prevent progression and end-­stage renal disease (ESRD), a tight glucose, lipid and hypertension management is needed. Without therapy, a loss of renal function will lead to dialysis. In Europe and the USA, diabetic nephropathy is the leading cause of ESRD. Serum creatinine levels higher than 2.0 mg/dL (>180 μmol/L) usually require treatment with insulin. Because of the lower degradation rate of insulin, smaller doses of insulin are recommended in ESRD to prevent hypoglycaemia. Diabetic retinopathy is the major eye complication in patients with diabetes. It is now the leading cause of legal blindness. Further diabetes-­related eye complications are cataracts and glaucoma. Neovascularisations with retinal bleeding characterise the proliferative form of retinopathy. To classify and prevent all this, a minimum of one annual screening and ophthalmoscopic examination by an ophthalmologist are recommended. Perimetrically visual field testing can detect scotomas. Treatment options are laser coagulations and vitrectomy. In general, early treatment of diabetes and hypertension can prevent eye complications. To avoid retinal bleeding after long-­term hyperglycaemia, blood glucose must be lowered slowly. Rapid changes of blood glucose concentrations lead to refraction disorders (blurred vision). This can happen by turgor change of the eye lens (osmotic effects) when a better diabetes therapy causes a significant improvement of the HbA1c-­ level in a short time. Impaired vision is one of the key points that can make driving impossible. Therefore, after every adjustment of diabetes-medication, driving ability may be temporary not given, until the metabolic situation is stable again.

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Diabetes is a chronic disease with a progressive course. Sometimes chronic diabetic complications can be found long before diabetes manifestation. They are relevant for driving because of their psychological and physical burden. Hence, it is important to get an expert opinion before driving (Ausschuss Soziales der DDG 2004).

The macrovascular disease of diabetes is mostly located in the coronary arteries. In contrast to non-­diabetics, cardiovascular mortality is threefold higher in people suffering from diabetes and metabolic syndrome. After 8–15 years of having diabetes without knowledge of cardiovascular disease (CVD), diabetics will have the same risk for myocardial infarction or stroke as non-­ diabetics after one myocardial infarction (Haffner et al. 1998).

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decline of the BG to 70 mg/dL (3.9 mmol/L) can cause neurohypoglycaemic reactions, such as misinterpretation of car speed, distance and risk. Sometimes this comes together with aggressive behaviour. Diabetics unknowingly suffering from hypoglycaemia are a danger to others and should not actively take part in traffic. The DCCT Research Group has shown that high HbA1c levels do not guarantee the absence of hypoglycaemic events (DCCT 1991). Also, short-­acting hypoglycaemias can lead to cerebral dysfunctions and persistent hypoglycaemias via fatal arrhythmias or accidents to death. Adequate awareness of hypoglycaemia can be defined as whether the license holder/applicant is capable of bringing their vehicle to a safe controlled stop (GOV.UK 2016).

Neuropathy Neuropathy in diabetes can affect the peripheral and the autonomic nerve system. In type 1 diabetes, neuropathy will occur after years of poor metabolic control. In type 2 diabetes, neuropathy can often be seen before diagnosis of metabolic disorder. Metabolic effects and vascular factors are thought to be the main causes of neuropathy. About 60–70% of people with diabetes will develop a neuropathy during their lifetime. Most common is a diffuse neuropathy affecting the feet and ankles (distal symmetrical sensorimotor polyneuropathy (DSSP)). Most people describe the feeling as pins and needles or like the movement of insects on the skin. In some cases, severe pain is reported. Typically, discomfort increases during the night, especially when the feet get warmer in bed. Peripheral neuropathy can be screened by a sensory test with a calibrated tuning fork

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57.3  Driving and treatment of diabetes 57.3.1  General treatment and education

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General treatment of diabetes utilises medical nutrition therapy, exercise, pharmacological therapies and insulin. Good diabetes lessons by certified diabetes educators and BG self-­control are necessary to manage the chronic disease over decades. Education is given for inpatients or outpatients by practice and clinical specialists. Supportive too is the organisation of people affected in self-­help groups. Type 1 diabetics need lifelong supplementation with insulin. Type 2 diabetics need a staged therapy starting with healthy food and exercise. Later they need the help of oral agents, and as soon as the endogenous insulin decreases, exogenous insulin application is necessary. Especially in the obese, incretin mimetics and SGLT-­2  inhibitors are helpful in glucose control and weight loss. BG control facilitates self-­management of diabetes. A tight blood pressure control and lipid control additionally helps to prevent organ failure. In 2015, the American Diabetes Association published revisions of “Standards of Medical Care in Diabetes” (American Diabetes Association 2015). In addition to other general treatment, i.e. physical activity, all individuals should be encouraged to limit the amount of time they spend sitting and smoking cessation must also include the use of e-­cigarettes.

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Lesions of the lower limb and foot are caused by angiopathy, neuropathy or both. The lesions are mostly graded according to Wagner’s classification. The treatment of diabetic foot ulcers is very complex and time-­consuming, and most patients are hospitalised. After accurate diagnosis (using wound examination and X-­ray, Doppler or duplex examination to quantify foot pulse status and pressure) and surgical debridement, an immobilisation will follow with pressure relief of the wound. In the case of infection, effective antibiotic combinations are needed. Often, skin dressings must be renewed daily. Nearly half of all foot ulcers are caused by inadequate shoes (Greitemann 1998). To prevent future lesions, adequate footwear should be built by an interdisciplinary specialist team (physician, surgeon, chiropodist, podiatrist and wound nurse) (Morbach et  al. 2004). Charcot neuropathy (diabetic neuropathic osteoarthropathy, DNOAP) mostly affects the forefoot and midfoot, limiting joint mobility and causing microfractures. The foot looks swollen and is unstably deformed. To reach stabilisation, casts and arthrodesis are used to prevent weight bearing for some months. Patients with severe neuropathy of the lower limbs and foot ulcers, especially when they wear casts or undergo arthrodesis, are unable to drive a car. In the case of amputations of the foot, lower limb or total limb, mobility is ensured by prostheses and/or wheelchairs. But the proper operation of the car’s foot controls may be impaired. Motor vehicles must have suitable adaptations according to the driver’s needs. Sometimes the use of a car with an automatic transmission will be adequate, and in other cases break and accelerator pedals must change to hand operation. The ability to safely control a vehicle at all times is the essential requirement (GOV. UK 2016).

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and monofilament. Additionally, test batteries can be used. Diabetics with severely impaired feeling of the feet may be unable to drive a car if they cannot adequately work the accelerator and brake pedals. Different organ systems can be affected by autonomic neuropathy, such as the heart and cardiovascular system, the stomach (gastroparesis), the gastrointestinal tract (diarrhoea) and the penis with genitourinary tract (erectile dysfunction). Autonomic neuropathy is not as common as diffuse peripheral neuropathy. Autonomic neuropathy of the heart can lead to a ‘silent infarction’ by affecting the pain fibres. Erectile dysfunction correlates to CVD and is mostly hidden by affected men but is always incriminating. The gastroparesis is important for traffic medicine. It can lead to severe hypoglycaemia through retarded gastric emptying. In persons using insulin or insulinotropic agents with pre-­meal administration, there can be a fatal mismatch between acting insulin and the post-­meal glucose rise. Patients with gastroparesis are characterised by an instable metabolic situation. Diagnostic procedures are sonography, breath test, gastroscopy and scintigraphy of the gastrointestinal tract.

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57.3.2 Oral agents Classic oral agents for the therapy of type 2 diabetes are metformin and the sulfonylureas. The guidelines of national diabetes organisations recommend these drugs in an early stage of diabetes (Matthaei et al. 2009). The biguanide metformin suppresses excess glucose production in the liver by inhibiting gluconeogenesis and glycogenolysis. Metformin also improves glucose sensitivity in muscle and fat tissues. It also shows a low anorexic effect, which helps weight loss. In elderly patients and those with impaired kidney function, metformin may cause kidney failure because of its primarily renal clearance. Acarbose and miglitol are α-­ glucosidase inhibitors. They inhibit competitively α-­ glucosidase enzymes in the small intestine, so carbohydrate cannot be broken down to simple sugars. Taken with a meal this will lead to a delay in glucose absorption. An intestinal side effect is flatulence. The group of thiazolidinediones is insulin sensitisers, improving tissue glucose uptake and suppressing hepatic glucose output. In some countries, only pioglitazone is still in use. Positive effects on the cardiovascular system are in discussion. Pioglitazone is used in several combinations with other oral agents and also with insulin. The most common oral agents are the second-­generation sulfonylureas such as glibenclamide (glyburide in the USA), glipizide and the third-­ generation glimepiride. They directly stimulate the pancreatic β-­cells to secrete insulin. This effect can cause severe and ongoing hypoglycaemias (Holstein et al. 2001, 2003). Repaglinide and nateglinide are insulinotropic oral agents

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TissueGlucose

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CarbohydratIntake GLP-1 α-GlucosidaseInhibitors

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Metformin Insulins Insulins Blood Glucose

Insulin

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Figure 57.2  Common insulins and antidiabetic agents in their main relations to human target organs (schematic).

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(in the group of meglitinides) that act as prandial glucose regulators. The effect of meglitinides on the β-­cell is less, and they are shorter acting than sulfonylureas. Meglitinides are administered before each meal (‘one meal one pill’). They are an alternative to sulfonylureas with less risk for severe hypoglycaemias. Metformin, α-­ glucosidase inhibitors, thiazolidinediones, DPP-­IV inhibitors and SGLT 2 inhibitors are oral agents with low or no hypoglycaemic potency, which makes them preferable for diabetics who wish to drive. Insulinotropic agents such as sulfonylureas and meglitinides bear a higher risk for severe hypoglycaemias (Figure 57.2).

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57.3.3 Insulin treatment There are three forms of insulin therapy. A conventional therapy uses mixed insulin injections twice daily; this form of therapy is suitable for people with stable mealtimes. The injections cover the basal and prandial insulin requirements. People with greater variability during the daytime, changing carbohydrate intake and different activities should have an intensified therapy using basal/ bolus insulin (MIT). This better imitates a physiological insulin secretion than conventional therapy does. The basal insulins are commonly the intermediate-­acting neutral protamine Hagedorn (NPH) insulins (average action time 8–12 hours), long-­acting analogues (insulin detemir and glargine, 18–26 hours) or very long-­ acting insulin analogues (insulin degludec, >42 hours). Bolus insulins are regular and rapid-acting insulins. The duration

of ­regular insulin is dose dependent, acting for about 4–8 hours, so it covers a meal and a following smaller meal (e.g. first and second breakfast). Skipping the second meal can lead to hypoglycaemia. Rapid-­acting analogues such as aspart, glulisine or lispro have shorter reaction times (3–4 hours) and cover only one meal. Therefore, they give more flexibility to the user and severe hypoglycaemic reactions occur 25% less often (Hirsch 2005). The most technical insulin delivery system is the CSII. Insulin infusion pumps are constructed to deliver regular or rapid-­acting insulin from a small tank via a catheter in the subcutaneous tissue (Henrichs et  al. 2009). A programmed basal rate provides the body for 24 hours with the basic needs of insulin (overnight and fasting). During exercise and higher activity, the basal rate can be temporarily lowered. Meal-­time insulin will be delivered as a pre-­ prandial bolus. Most pumps can provide several different pre-­ meal boluses. A small snack needs a short bolus, a large pizza a dual bolus. All types of diabetic treatment need SMBG for adequate dosing (e.g. intensive insulin injection and pump therapy). A good knowledge of nutrition therapy is also necessary. Inadequate insulin dosing can lead to hyper-­or hypoglycaemia. CGM has been available since 1999 and helps to evaluate the metabolic situation from time to time. Wearing such a system continuously, individually programmed alarms can prevent from hypo-­and hyperglycaemic situations. Actual tissue glucose values and trend analysis can be seen on the monitor display. One type of CGM is compatible with an insulin infusion pump (sensor-­ augmented pump). CGM data and glucose curves are then shown on the pump’s display. In a case of severe hypoglycaemia and

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tablets were approved in the European Union in 2012. The new inhibitor lowers BG by removing excess glucose in the urine. All the new drugs are potentially able to lower the BG without causing a significant risk of hypoglycaemia. Besides metformin, incretin mimetics, DPP-­4 inhibitors and SGLT2 inhibitors seem to have a useful future in the treatment of drivers suffering from type 2 diabetes. Caution is needed with new combinations in one pill or syringe. Those fix combinations may contain DPP-­4 inhibitors and metformin, DPP-­4 inhibitors and SGLT2 inhibitors, and SGLT2 inhibitors and metformin which seem to be save concerning hypoglycaemias. In contrast, a combination of GLP-­1 agonists and basal insulins may cause hypoglycaemia.

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57.4  Driving performance and diabetes Diabetes is an endocrine disorder that is characterised primarily by hyperglycaemia and acute and chronic metabolic complications. Untreated or inadequate therapy will result in persistent hyperglycaemia, which constitutes a serious impairment for the sufferer with a direct influence on the driving ability of those affected. Treatment with insulin or oral insulinotropic agents bears a danger of sudden hypoglycaemia. Diabetes-­related complications such as micro-­and macroangiopathy and neuropathy with their endpoints of kidney failure, blindness, myocardial infarction, cerebral stroke and amputation will restrict active participation in road traffic. Because the disease is progressive, it can be present in patients with type 2 diabetes for a long asymptomatic time. Therefore, the detection of diabetic organ complications may be earlier than the diagnosis of diabetes itself. In addition, there will be a significant co-­morbidity in older patients. However, a prospective follow-­up study in Norway showed no increased risk of being involved in a road traffic accident for drivers taking oral glucose lowering agents. A slightly increased risk was observed for the group using insulin, with the highest risk for people aged 18–34 years (Skurveit et al. 2009). Driving by people with diabetes is impaired by the factors such as hyperglycaemia, hypoglycaemia and diabetic complications (Stork et al. 2006). A 2-­hour continuous driving training on a driving training course was performed to investigate the effect of prolonged acute mental stress. In a post-­prandial state of insulin-­treated people with type 1 and type 2 diabetes, blood glucose, systolic and diastolic blood pressure, heart rate, salivary cortisol concentrations and subjective stress perception were analysed on a control and a stress day. The average blood glucose at the beginning was 136.8 mg/dL (7.6 mmol/L). The peak of glucose concentrations was 90 minutes after intake of a liquid standard meal and glucose rested elevated for 4 hours on both days. This was to prevent serious hypoglycaemia during driving. All other physical parameters and the stress perception showed significantly higher values on the driving training day which begun immediately after start of the practice round. The findings hypothesise that post-­prandial glucose control is not significantly altered in insulin-­treated peo-

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unconsciousness, the sensor itself stops the basal insulin infusion of the pump for 2 hours. Then, the pump starts insulin perfusion again, to prevent ketoacidosis. During the 2 hours of basal rate inactivity, blood glucose will increase about 40 mg/dL (2.2 mmol/L) per hour (in total 80 mg/dL or 4.4 mmol/L), leading to consciousness. New kinds of insulin delivering systems are micro or patch pumps (without catheters). Some of them will soon be available in combination with a matching glucose sensor system. All the pumps on the market are external pumps; implantable pumps are not yet available. All users of insulin pump systems need special training to ensure successful therapy. The permanent use of a glucose management system with a pump is recommended, which may help to prevent users from overdosing insulin and overcorrections (shortly after a meal). Both the latter can lead to severe hypoglycaemia. The most often reported reasons from a patient’s point of view for a change from MIT to CSII are unpredictable metabolic variations, severe and stressing hypoglycaemias as well as nocturnal hypoglycaemic events. Immediately after start of pump therapy, they notice an increase in working ability and physical and mental performance (Quester et al. 2014). Overdosage or fatal complications of human insulin and its synthetic analogues can now be analysed by immunoaffinity and liquid chromatography-­mass spectrometry (Hess et al. 2012a, b). This is important for clinic as well as for forensic toxicology.

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57.3.4 New agents

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New agents for the treatment of diabetes are the incretin mimetics (GLP-­1  mimetics) and the DPP-­4  inhibitors (Figure  57.2). Glucagon-­like peptide 1 (GLP-­1) is a peptide hormone deriving from the small intestine. The half-­life is only 1 minute and it is degraded by an enzyme called dipeptidyl dipeptidase-­4 (DPP-­4). The first GLP-­1  mimetic was exendin-­4 from the saliva of the Gila monster (Heloderma suspectum). The synthetic hormone exenatide cannot be converted by DPP-­4 enzymes. Exenatide is injected subcutaneously 60 minutes before a meal in the morning and in the evening. It enhances insulin secretion, slows absorption of glucose from the gastrointestinal tract and suppresses glucagon production (Mazze et al. 2007). Because it imitates the appetite suppression mechanisms of incretins, exenatide not only leads to a blood glucose control but also to a significant weight loss. Therefore, it is recommended to overweight patients with type 2 diabetes. The most advanced galenic of exenatide is injected once weekly. Liraglutide and Lixisenatid, other synthetic incretin mimetics, must be injected once daily. The approximate glucose lowering potential of incretin mimetics is about 30 mg/ dL (1.7  mmol/L), meaning a 1% HbAlc reduction. Incretin mimetics show a low hypoglycaemic risk. DPP-­4 enzymes perform a rapid degradation of human GLP-­1. The new class of gliptines, the DPP-­4  inhibitors (sitagliptin, vildagliptin, saxagliptin and linagliptin), enhance the endogenous GLP-­1  levels. A new group of selective inhibitors of the sodium-­dependent glucose co-­transporter 2 (SGLT2), working independently from insulin, can be used for the treatment of type 2 diabetes. Dapagliflozin

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licences, even those issued before the date of application of Directive 2006/126/EC (Commission Decision (EU) 2016/1945 of 14 October 2016 on equivalences between categories of driving licences, notified under document C (2016) 6517). Usually, there are no restrictions for diabetics to get a licence to drive vehicles from group 1, classes A, A1, B, BE, M, L and T (smaller and less powerful motorcycles and private cars with no more than eight seats). There is a discussion in Germany and elsewhere about whether to ask those seeking a group 1 driver’s licence about their diabetes disease. There are progressive restrictions excluding insulin-­treated diabetics from driving group 2, class C1 and D1 vehicles (i.e. heavy trucks, cars for passenger transport and buses). Directive 91/439 of the European Union is interpreted differently throughout the European authorities (DVLA 2011). Many US states also have a restrictive licensing programme for diabetic drivers. Usually, insulin-­treated diabetics are denied an interstate commercial driving licence. However, there is no general reporting system for unexpected loss of consciousness due to hypoglycaemia (Stork et al. 2006). The findings of a study from the UK on road traffic accidents and diabetes suggest no increased risk to road safety for the group of insulin-­treated patients. Therefore, the authors proposed an individualised risk-­based assessment when considering driving restrictions (Lonnen et al. 2008). In Germany and other states, authorities can force drivers, after traffic mishaps due to diabetes problems, to consult a diabetes physician with special traffic experience for a medical examination. This is reported to the authorities (§11 Fahrerlaubnis-­Verordnung). The reporting specialist must be a different physician from their normally consulted healthcare professional. The driver may lose his/her driving permission for a defined period. A Blood Glucose Awareness Training (BGAT) can be enforced by law. This psycho-­educational training program is known to help the affected to better understand, recognise, prevent and counteract hypoglycaemic episodes. After optimising diabetes medication and patient behaviours to restore driving ability, a new medical evaluation takes place. The license can be retained or granted with or without restrictions. Evaluation guidelines for driving ability (Begutachtungsleitlinien zur Kraftfahreignung  2019) are now reconsidered in Germany. All physical and/or mental handicaps that make people unable to drive or limit their driving ability are listed in this reference book. Together with the S2e-­guideline “Diabetes and Road Traffic” of the German Diabetes Association DDG (Ebert et al. 2018), they are used by healthcare professionals for a case-­by-­case evaluation of driving ability. The guidelines give advice regarding the evaluation instructions, ways to compensate handicaps and regulate the legal position of the evaluating physicians (e.g. board approved specialists for diabetology). Needed for secure driving is a stable metabolic situation for at least 3  month and an unimpaired awareness of hypoglycaemias. In Australia, people with private or commercial driving licences treated with glucose-­lowering agents or insulin must notify their Driver Licensing Authority (DLA). They are not regarded as fit to hold an unconditional licence. Private licence holders need a five-­yearly review for licensing by their treating

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ple with T1D or T2D despite a driving-­ associated stress (Truninger et al. 2013). Under continuous glucose monitoring, a small group of 10 drivers with well-­balanced T1D was accompanied while steering a car under real-­life conditions. About 2 hours before the road trip in the city started, they had breakfast and their normal appropriate insulin injection. The average interstitial glucose was 168.5 mg/dL respective 9.36 mmol/L. After the first 35 minutes driving time, the CGM values showed a small peak (+7.2 mg/dL or +0.401  mmol/L) that was explained by the authors as stress associated. After 2 hours of driving, the CGM values were lower compared to the start (−15.8 mg/dL or −0.88 mmol/L). This may be due to the increasing glucose requirements after the initial stress situation. The following day and at the same time, without driving, this phenomenon did not happen. Hypoglycaemic reactions did not occur during the total time of the study. In conclusion, short driving times seem to be without risk of hypoglycaemia in people with well-­controlled type 1 diabetes (Schmied and Zulewski 2019). Meta-­analyses and meta-­regressions of 28 studies were performed to prove whether diabetes is associated with an increased collision risk and to show its effects of gender and age (Hostiuc et al. 2016). Overall, the authors found no statistically significant difference for unfavourable traffic events between drivers with and without diabetes. Neither gender nor age of diabetic patients had significant influence on traffic accidents. Only elderly and insulin-­ dependent people tended to have a higher risk. Interestingly, European drivers with diabetes showed a significantly lower collision risk compared with their North American counterparts. This depended, obviously, on the general risk for road fatalities in both continents.

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57.4.1  Drivers’ licences and diabetes

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For social participation and inclusion of people with chronic diseases, a major premise may be private and occupational mobility (Ebert et al. 2019). Restrictions regarding driving, without consideration of individual skills and physical abilities, may be regarded as discrimination. Worldwide there are a great variety of licensing requirements applied by both state and federal jurisdictions for people with diabetes who wish to operate motor vehicles (ADA 2012). In general, it is accepted that metabolic, well-­ adjusted and adequately educated people with diabetes can drive motor vehicles safely. Therapy concepts, use of vehicles and an undisturbed hypoglycaemia awareness must be considered by the medical assessment (evaluation guidelines for driving ability; Begutachtungsleitlinien zur Kraftfahreignung  2019). Diabetes alone is not sufficient to make any judgements about individual driver capacity (ADA 2014). It differs from country to country whether student drivers with diabetes must self-­identify before the start of the license application process or not. In Europe, there are drivers’ licences in two groups and several classes, covering all vehicles in road traffic. All member states of the European Union should mutually recognise their driving

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generation BGMS did not meet accuracy criteria based on latest EN ISO 15197:2015 (Pleus et al. 2020).

57.4.2  Driving and hypoglycaemia

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Nowadays, the treatment of people suffering from diabetes aims towards the goal of near normoglycaemia to prevent chronic diabetic complications. This is only possible if no severe hypoglycaemic reactions occur. Secure and effective diabetic therapies are obviously preferable for drivers. Severe hypoglycaemia can not only lead to sudden loss of consciousness, some hypoglycaemias start with constraints of cognitive functions. Reduced alertness, awareness and ability to respond on street or traffic conditions, impaired visual acuity, memory decline and mental disorders can happen (Table  57.1). The combination of hypoglycaemia and drunk driving amplifies those effects as alcohol consumption impairs the hepatic glucose contra-­regulation. Drivers of motorcycles are particularly endangered by even light hypoglycaemia because of the early dysfunction of their sense of balance (Finck 2009). As shown above, disrupted driving performance will be detected in people with a BG of less than 70 mg/dL (3.9 mmol/L) (Cox et al. 1993, 2000). Driving simulator studies during moderate hypoglycaemia showed deviations from the road, too fast driving and compensatory slowing of speed. In another experimental setting, the decision not to drive during hypoglycaemia was tested. Type 1 diabetics unaware of their hypoglycaemia frequently decided to drive in a hypoglycaemic situation. Those who were aware of their hypoglycaemia made safer decisions. Potentially dangerous decisions were made by people with type 2

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doctor, when on an insulin biennial review. Commercial drivers only on metformin will be reviewed annually by their treating physician, and all others on glucose-­lowering agents or insulin must have annual reviews for licensing by an endocrinologist or consultant physician specialising in diabetes. Patients and health professionals have the legal obligation to notify the DLA when driving is likely to be affected. The DLA assesses each medical report considering the nature of the driving task (farmers vs. interstate vehicle drivers), particularly when granting a conditional licence (National Transport Commission 2016). In addition, driving skills may be assessed by the authorities or an occupational therapist. In the UK, interstitial glucose-­monitoring systems may be used by drivers of group 1. Additional finger prick capillary glucose testing is required for adjusting the system and when symptoms of hypoglycaemia or low readings occur (glucose level ≤4.0 mmol/L or ≤70 mg/dL). Group 2 drivers are not permitted to use FGM or rtCGM devices. Drivers of group 1 vehicles and motorcycles must test capillary glucose within 2 hours before start of journey and every 2 hours after driving has started. Only a maximum of 2 hours is allowed between pre-­driving check and first glucose test while driving. In addition, group 2 bus and lorry drivers must perform at least 2 regular blood glucose self-­tests, even on days when not driving (GOV.UK 2016). One of the main problems of diabetes care is the continued poor performance and accuracy of some systems for self-­ monitoring of capillary blood glucose. They are used widespread without fulfilling legal accuracy criteria for autonomous control of the metabolism, to make therapy decisions and adjust CGM systems (Baumstark et  al. 2019). More than 20% of current-­

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Table 57.1  Blood glucose levels and their main relations to symptoms and human behaviour.

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4.2 3.9 3.3

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Normal reaction counterregulatory hormones Autonomous symptomes sweating, tremor, fear, palpitation, hunger Early neurohypoglycemic symptoms cognitive dysfunktion, odd behavior (aggressiveness) Late neurohypoglycemic symptoms drowsiness, confusion, disturbance of speech Severe neurohypoglycemic symptoms unconsciousness, convulsion

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As expected, the drivers at low risk and routine care (LR-­RC) had the lowest risk of 0.75 driving mishaps in 12 months. The studies demonstrate the fortune of the RADD questionnaire (now on the ADA website) in identifying drivers with type 1 diabetes at high risk and the beneficial help of the Internet intervention to prevent from traffic mishaps (DiabetesDriving.com). Interesting for people treated with insulin or insulinotropic oral agents is the continuous monitoring of interstitial fluid glucose with alarm functions, warning from high or low glucose levels (real-­time continuous glucose-­monitoring systems, rtCGM) or flash glucose-­monitoring systems (FGM, or intermittently scanned continuous glucose monitoring, iscCGM). A comparison regarding glycaemic variability (GV) and hypoglycaemic excursions with these two different systems in adults with highest risk type 1 diabetes showed a slight improvement of GV measures with rtCGM (Avari et al. 2019). In a head-­to head pilot study, the impact of CGM and FGM was assessed in people with type 1 diabetes and impaired awareness of hypoglycaemia (Reddy et  al. 2018). In both groups, glucose monitoring over 8 weeks improved HbA1c and percentage time spent in glucose target. CGM more effectively reduced time spent in hypoglycaemia at all thresholds and also hypoglycaemia fear. Limitations to the study include the small number of participants and the short follow-­up period; the self-­reported awareness of hypoglycaemia remained unchanged with both groups. For people with impaired awareness of hypoglycaemias, a blood glucose awareness training could be effective in reducing traffic violations and accidents (Stork et al. 2006). In special cases, the daily use of an insulin pump connected with rtCGM and an automatic stop of basal rate in case of hypoglycaemia may be possible (sensor-­augmented pump). A last generation of insulin pumps even automatically adjusts every 5 minutes the basal rate based on real-­time sensor glucose values (Cohen 2019). The users of this approved system must count and program only the ingested amount of carbohydrates. First, studies show a minimum time in the range of 70% and only short times of 2.4% below 70 mg/dL (3.9 mmol/L). Extremely critical is the use of patient created “artificial pancreas systems” using a commercial insulin pump and an rtCGM connected by a personal programmed software. In the Internet, such software is shared. Those systems are not officially approved. In the case of misfunction and/or injuries, there will be no insurance coverage. Healthcare professionals are not allowed to support users of such systems. Glucose meters must be certified, plasma calibrated and should have a memory to cover at least 3 month of readings, and/ or the user must use a diabetes diary for regular reviews with a physician providing diabetes care. In the UK, drivers who experience an episode of severe hypoglycaemia whilst driving, having more than one severe hypoglycaemia in the last 12  month or impaired awareness of hypoglycaemia must not drive and must notify the Driver and Vehicle Licensing Agency (DVLA). In some states of the USA, healthcare professionals have to report this to the authorities. In Germany, physicians must inform their patients not to drive. Only in severe cases when patients who must refrain

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diabetes, particularly when treated with oral hypoglycaemic agents (Stork et al. 2007). Some diabetes medications are combinations of non-­ hypoglycaemic agents and sulfonylureas, also seen in Traditional Chinese Medicine (TCM) or related drugs. Special information must be given to people with diabetes, using those drug combinations. Unacceptable are over-the -counter liquid antidiabetic drugs containing herb extracts because they contain up to 70% alcohol (i.e. Syzygium cumini). About 25% of patients with type 1 diabetes are unaware of their hypoglycaemia (Stork et al. 2006). People with this complication should not take part in road traffic. Strict avoidance of low glucose levels, multiple SMBG, no skipping of meals and less corrections of hyperglycaemic values may ease the problem. It is recommended to start driving only on blood glucose levels higher than 90 mg/dL (>5 mmol/L), otherwise the intake of carbohydrates is needed. SMBGs should be performed every 2–3 hours during the road trip. In the case of first signs of a hypoglycaemic episode, the driver must immediately stop the vehicle for a break. Blood glucose levels lower than 70 mg/dL (250 mg/dL or 13.9 mmol/L) were reported with an average number of driving mishaps/per year/per driver from 0.25 in the group with low risk to 0.61 in the group of high risk. Post hoc tests showed no significant differences between the intervention groups with Internet education of five units and with Internet education plus motivational telephone interviewing (0.54 vs. 0.59 driving mishaps/year/ driver). It may still be difficult to make an acute or chronic hyperglycaemia or coma responsible for a fatal accident. Post-­mortem glucose is rapidly metabolised into lactate. HbA1c or fructosamin values are long-­term indicators and do not reflect the actual metabolism. Therefore, a sum formula of Traub and the decreased concentration of 1,5-­anhydroglucocitol, the 1-­deoxy-­form of glucose, may give a hint to a severe hyperglycaemic situation (Hess et al. 2012a, b).

All diabetics treated with insulin or insulinotropic substances are in danger from hypoglycaemia. This risk can be minimised by special diabetes training, careful metabolic control and frequent BG self-­controls and/or sensor systems. Education must include the knowledge of potential hazards associated with travel on the road. This should be documented by the attending physician to avoid running the risk of any blame in the event of an accident involving his or her patient. There are restrictions for drivers with diabetes worldwide. In some countries, they range from more frequent than usual medical examinations to denial of driving privileges for certain groups, for example patients with hypoglycaemia unawareness (Stork et al. 2006). According to the German road traffic licensing regulation, StVZO section  2, people suffering from illness, including diabetes, are required to assess their driving skills before every departure as a driver, sometimes with the help of a physician. Therefore, from a legal perspective, it is important to provide people with diabetes with structured education covering all aspects of driving safely with the illness. The training should mandatorily include the following advice: • Start and continue driving only when feeling well. • Exclude a metabolic imbalance (hypoglycaemia or hyperglycaemia) by SMBG before departure. • Keep a diabetes diary. • Carry all utensils for SMBG (e.g. lancing device, meter and test strips). • Carry all utensils to guide your therapy (e.g. insulin, syringes and pump catheters). • Do not overcorrect with insulin or glucose lowering agents. • Carry rapid-­acting carbohydrate (e.g. glucose, apple juice and lemonade). • While driving make regular and sufficient breaks and perform metabolic control. • Stop driving immediately if there are signs of hypoglycaemia or discomfort. • Consume carbohydrates before testing blood glucose for hypoglycaemia. • Consume carbohydrates if there are low blood glucose values as soon as possible. • Consume adequate amounts of carbohydrates. • Find out the cause of hypoglycaemia. • Take precautions to avoid a second hypoglycaemia. • Continue driving only after all symptoms of hypoglycaemia have completely disappeared. Adherence to these precautions is necessary to ensure the safe driving performance of those with diabetes (Quester and Tschöpe 2007). A new idea to make driving safer is to use a pre-­driving checklist just as pilots do with their pre-­flight checklist (Cox et  al. 2017). After a pre-­driving SMBG, a traffic light symbol reminds drivers of benchmarks. Green colour indicates they

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57.4.3  Driving and hyperglycaemia

57.5  Therapeutic aspects for driving safely with diabetes

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range must start higher, 100–180  mg/dL (5.6–10  mmol/L). A good and stable time in range must be 70% or more. To show hypoglycaemias, a time below range (TBR, ≤70  mg/dL or ≤3.9  mmol/L) is calculated. Interesting, too, is the time above range (TAR, >180 mg/dL or >10 mmol/L). A new simple composite metric for the assessment of glycaemic status was introduced in 2017 (Hirsch et al. 2017). It is called Personal Glycaemic State (PGS). The data derived were equated solely from an rtCGM device composed by the items mean glucose, glycaemic variability, time in range and frequency and severity of hypoglycaemias. Hypoglycaemias were counted by the distinct thresholds 54  mg/dL (3.0  mmol/L) and 70  mg/L (3.9  mmol/L). Severe hypoglycaemias with neurocognitive impairment, unconsciousness and the possible need of assistance may occur below the lower threshold. In the second range 55–70  mg/dL (3.1–3.9  mmol/L), the adequate reaction is the intake of fast-­acting carbohydrates. After a waiting time of about 20  minutes, the glucose level will rise higher than 90  mg/dL (5.0 mmol/L). Only when the driver has fully recovered, the trip can be continued. It means that potentially clinically serious hypoglycaemias and less serious incidents of biochemical hypoglycaemias were separated. The values of the PGS are classified in quartiles (excellent, good, moderate and poor) to allow users and healthcare professionals a quick decision about the quality of glycaemic control and enable them to optimise further treatment. In the future also, other rtCGM or iscCGM may use such a composite index. Reflecting the circumstance that only about 65% of insulin using drivers in the UK consider blood glucose testing before driving, the use of sensor-­based glucose information has the potential to make driving more secure (Rayman et al. 2017). Only 71.6% of women and 77.8% of men on insulin pumps recorded by the German Insulin Pump Registry constantly used diabetes diaries (manually or electronically) (Quester et al. 2014). This is the most educated and best prepared group of insulin-­dependent diabetics.

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could start the road trip (BG > 90 mg/dL or > 5 mmol/L). Yellow means caution of hypoglycaemia during the ride and the need of prophylactic intake of carbohydrates (BG between 70 and 90 mg/ dL or 3.9 and 5  mmol/L). The red light (BG < 70  mg/dL or 3.9  mmol/L) indicates danger, and the driving must be interrupted. The immediate consumption of fast-­acting carbohydrates and long-­acting carbohydrates is recommended. Every driver should have easily accessible the following items in the car: a BG meter, and fast-­acting (dextrose tablets) and long-­ acting carbohydrates (cheese crackers). During the pre-­driving procedure, the driver must consider the potential risk of hypoglycaemias during the road trip. It is still not clear whether diabetics more frequently cause traffic accidents than non-­diabetics. As shown, the published studies provide some conflicting evidence or are methodologically vulnerable. Often no information is given on the driving performance of the comparison groups. In Germany, diabetic patients had no higher risk for traffic accidents compared with nondiabetics (Ausschuss Soziales der DDG 1999). A case–control study on diabetes and car accidents regarding over 65 year olds showed no significant difference between groups in terms of accident causation (McGwin et al. 1999). Part of the European Union project IMMORTAL was a literature search to assess the relationship between accident risk and age and various diseases (Vaa 2003); 25 reports of diabetes were evaluated. The relative risk for being involved in a traffic accident was given for diabetes as 1.56 (highly significant). To judge the driving ability of people with diabetes, all facets of the disease, including co-­morbidities, are assessed individually. The severity and stage of the disease, its predominant symptoms, the complications that occurred and its functional effects in interactions with other diseases all need to be considered (Ausschuss Soziales der DDG 2004). Furthermore, critical evaluation is needed concerning the quality of treatment, possible alternative treatment options and the scope for compensation of fitness for relevant functional or performance constraints. The metabolic situation must not be judged only by HbA1c-­ values as practiced in the past. Better parameters to assess for glycaemic variability will be the mean amplitude of glycaemic excursions (MAGE), the average daily risk range (ADRR) and the relative time lower or higher the normal glucose range (Time-­In-­ Range, TIR). Large MAGE values are associated with unstable diabetes (Service et al. 1970; Yu et al. 2018). The ADRR is also calculated from routine self-­monitored blood glucose data. The ADRR formula reflects the average of the risk range per day and shows a good balance of sensitivity in predicting future hypoglycaemia and hyperglycaemia (Kovatchev et al. 2006). By these calculations based on conventional SMBG (from diabetes diaries), the risk of hypoglycaemic episodes may be better estimated than by HbA1c-­values alone. Especially, the TIR shows the benefits of rtCGM but depends on the target range. By definition, the time in range is counted between 70 and 180  mg/dL (3.9–10  mmol/L) (Thomas 2019). Individually, a tighter glucose control can be set to 70–140 mg/dL (3.9–7.8 mmol/L). In the case of sudden hypoglycaemias or hypoglycaemia unawareness, the accepted target

57.6  People with diabetes as victims of traffic injuries When a traffic accident occurs, special care must be taken by rescue workers when victims are people with diabetes. Some of them may not only suffer from a trauma but additionally from diabetes problems. As shown, unconsciousness can be caused by a severe hypoglycaemia or hyperglycaemia, so the glycaemic status must be tested immediately. Also signs of paralyses, strokes and seizures can be caused by hypoglycaemia. Staged correction of the metabolic situation with glucose infusions are needed. Sulfonylureas especially when overdosed can cause series of hypoglycaemias within the next 2 days. During an accident, the catheter of an insulin pump may be disconnected or blocked, and blood glucose increases rapidly towards a coma. In other cases, during a hypoglycaemia caused absence, the insulin pump is still delivering insulin. So, first aiders must look for hidden devices

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American Diabetes Association (2014). Diabetes and driving. Diabetes Care 37 (Suppl. 1): 97–103. American Diabetes Association (2015). Summary of revisions of standards of medical care in diabetes. Diabetes Care 38 (Suppl. 1): 54. Ausschuss Soziales der DDG (Deutschen Diabetes-­Gesellschaft) (1999). Die Fahrerlaubnis beim Diabetes mellitus. Diabetes und Stoffwechsel 8: 35–38. Ausschuss Soziales der DDG (Deutschen Diabetes-­Gesellschaft) (2004). Information, Ausschuss Soziales der Deutschen Diabetes-­Gesellschaft. Empfehlungen zur Beurteilung beruflicher Möglichkeiten von Personen mit Diabetes. www.deutsche-­diabetes-­gesellschaft.de (last accessed 1 October 2013). Avari, P., Moscardo, V., Jugnee, N. et al. (2019). Glycemic variability and hypoglycemic excursions with continuous glucose monitoring compared to intermittently scanned continuous glucose monitoring in adults with highest risk type 1 diabetes. Journal of Diabetes Science and Technology Aug 2: 1932296819867688. DOI: 10.1177/ 1932296819867688 (last assessed 18 February 2020). Baumstark, A., Jendrike, N., Pleus, S. et al. (2019). Evaluation of system accuracy of four systems for Self-­Monitoring of Blood Glucose purchased in discounters and drugstores in the course of healthcare research adapted from ISO 15197. Diabetologie und Stoffwechsel 14: 456–461. Begutachtungsleitlinien zur Kraftfahreignung (2019). Bundesanstalt für Straßenwesen, Bergisch Gladbach, Stand 31. Dezember 2019, 32–36. Belchetz, P. and Hammond, P. (2003). Mosby’s Color Atlas and Text of Diabetes and Endocrinology. Amsterdam: Elsevier Science. Cohen, O. (2019). MiniMed™ 670G System: Practical Guidance to Onboard Your Patients Successfully. Oral presentation. Medtronic Symposium 20.020.2019, ATTD Congress Berlin 2019. Cox, D.J., Gonder-­Frederick, L. and Clarke, W. (1993). Driving decrements in type 1 diabetes during moderate hypoglycaemia. Diabetes 42: 239–243. Cox, D.J., Gonder-­ Frederick, L.A., Kovatchev, B.P. et  al. (2000). Progressive hypoglycemia’s impact on driving simulation performance. Diabetes Care 23 (2): 163–170. Cox, D.J., Gonder-­Frederick, L.A., Singh, H. et al. (2017). Predicting and reducing driving mishaps among drivers with type 1 diabetes. Diabetes Care 40: 742–750. DCCT (Diabetes Control and Complications Trial) Research Group (1991). Epidemiology of severe hypoglycemia in the diabetes control and complications trial. American Journal of Medicine 90: 450–459. DCCT (Diabetes Control and Complications Trial) Research Group (1993). The effect of intensive treatment of diabetes on the development and progression of long-­term complications in insulin-­dependent diabetes mellitus. New England Journal of Medicine 329: 977–986. DVLA (Drivers and Vehicle Licensing Agency) (2011). At a Glance. Guide to the Current Medical Standards of Fitness to Drive. Swansea: Drivers Medical Group DVLA. Ebert, O., Bohn, B., Bertram, B. et al. (2018). S2e-­Leitlinie Diabetes und Straßenverkehr. Deutsche Diabetes Gesellschaft (DDG). Diabetologie und Stoffwechsel 13: 54–97. Ebert, O., Bohn, B., Bertram, B. et al. (2019). Diabetes and road traffic. Experimental and Clinical Endocrinology & Diabetes 127 (Suppl. 1): 114–123.

57.7 Conclusions

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Diabetology is one of the most developed disciplines in medicine. On the other hand, we recognise an increase of people with diabetes worldwide, an epidemic. Today, patients can use the newest medications and insulins. Electronic devices such as sensors and pumps for insulin delivery are common. In the not so distant future, a complete artificial pancreas will further ease the situation. The interdisciplinary collaboration of specialists in diabetes, endocrinology, internal medicine, cardiology, angiology, neurology, ophthalmology, surgery, podiatry, obstetrics, pharmacology, psychology and diabetes education provides a great chance for people with diabetes to manage their disease favourably. Even commercial pilots who lost their professional licenses (CPL) were treated by a personalised type 2 diabetes management called “De-­ Escalation Treatment” (DET). It successfully preserved beta-­cell function up to several years, and they regained certificates (Pfützner and Rose 2019). All the tremendous efforts in diabetes treatment of the last years, together with the great improvements in diabetes technology, will make driving for those affected safer despite the knowledge that most traffic fatalities are caused by human error or the lack of attention.

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such as pumps or CGMS. The simplest way to stop insulin delivery in an emergency is to tear out the catheter tip or needle. Catheters are generally inserted at the abdomen, waist, hips or thighs. Small children can wear the catheter at the buttock under the diaper. Patch pumps can be detached completely from the skin and are usually found on the abdomen and arms. Pumps may be damaged or show malfunction due to accident or safety seat belt. All this must be documented in time to inform paramedics and the emergency room (ER) or intensive care unit (ICU) staff. The devices should be secured and given to the hospital. Pump and CGM internal memories can be read out later and show helpful data. Long-­lasting insulins are more secure than NPH insulins regarding hypoglycaemias in normal life. But they can be active far more than 24 hours, so after an accident the metabolic situation must be monitored. The readings of internal memories of systems for self-­ monitoring of blood glucose and rtCGM or iscCGM are used to have a look at the time before accident. This helps to understand the drivers mental and physical situation during the driving mishap. Other efforts were put to reconstruct the metabolism by biochemical methods from blood and urine.

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References and further reading American Diabetes Association. (2005). Defining and reporting hypoglycaemia in diabetes: A report from the American Diabetes Association Workgroup on Hypoglycemia. Diabetes Care 28: 1245–1249.

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Kovatchev, B.P., Gonder-­Frederick, L., Otto, E. et al. (2006). Evaluation of a new measure of blood glucose variability in diabetes. Diabetes Care 29 (11): 2433–2438. Lonnen, K.F., Powell, R.J., Taylor, D. et al. (2008). Road traffic accidents and diabetes: Insulin use does not determine risk. Diabetic Medicine 25 (5): 578–584. Matthaei, S., Bierwirth, R., Fritsche, A. et  al. (2009). Medikamentöse antihyperglykämische Therapie des Diabetes mellitus Typ 2. Update der Evidenzbasierten Leitlinie der Deutschen Diabetes-­Gesellschaft. Diabetologie 4: 32–64. Mazze, R.S., Strock, E., Simonson, G. and Bergenstal, R. (2007). Staged Diabetes Management: A Systematic Approach, revised 2nd edn. Chichester: John Wiley & Sons. McGwin, G., Pulley, L.V., Sims, R.V. and Roseman, J.M. (1999). Diabetes and automobile crashes in the elderly. Diabetes Care 22 (2): 220–227. Morbach, S., Müller, E., Reihe, H. et  al. (2004). Diagnostik, Therapie, Verlaufskontrolle und Prävention des diabetischen Fußsyndroms. In: W.A. Scherbaum and Th. Haak (eds.), Evidenzbasierte Leitlinien. Diabetes und Stoffwechsel, 2nd edn, pp. 1–84. www.deutsche-­diabetes-­ gesellschaft.de/ (last accessed 15 April 2013). National Transport Commission Australia (2017). Assessing Fitness to Drive 2016. For Commercial and Private Vehicle Drivers. Austroroads Publication Number: AP-­G 56-­17, 59–66. Nguyen, T., Dao, V.V., Timberman, J. et al. (2019). The effects of acute hyperglycemia on driving performance in people with T2DM. Diabetes 2019 Jun; 68 (Supplement 1): https://doi.org/10.2337/db19-­ PUB (last assessed 18 February 2020). Wieland, D., Müller, U.A. et  al. (2019a). Petersmann, A. Müller-­ Definition, Klassifikation und Diagnostik des Diabetes mellitus. Diabetologie 14: 111–118. Petersmann, A. Müller-­Wieland, D., Müller, U.A. et al. (2019b). Definition, classification and diagnosis of diabetes mellitus. Experimental and Clinical Endocrinology & Diabetes 127 (Suppl. 1): 1–7 Pfeiffer, A.F.H. and Freckmann, G. (2004). Kontinuierliche Glucosemessung und künstliches Pankreas. In: H. Schatz (ed.), Diabetologie kompakt, pp. 102–104. Stuttgart: Thieme. Pfützner, A. and Rose, D.-­M. (2019). Erfolgreiches personalisiertes Typ-­ 2-­ Diabetes-­ Management bei Berufspiloten durch β-­ zellschonende Deeskalationstherapie (DET)  – Hintergrund und Fallbeispiele. Diabetes, Stoffwechsel und Herz 28: 181–186. Pleus, S., Baumstark, A., Jendrike, N. et al. (2020). System accuracy evaluation of 18 CE-­marked current-­generation blood glucose monitoring systems based on EN ISO 15197:2015. BMJ Open Diabetes Research 2019-­ 001067 (last & Care 2020; 8: e001067. doi: 10.1136/bmjdrc-­ assessed 21 January 2020) Quester, W. and Tschöpe, D. (2007). Diabetes mellitus. In: B. Madea, F. Mußhoff, and G. Berghaus (eds.), Verkehrsmedizin. Fahreignung, Fahrsicherheit, Unfallrekonstruktion, 2nd edn, pp. 345–367. Köln: Deutscher Ärzte-­Verlag. Quester, W., Reichel, A. and Henrichs, H.R. (2014). Ergebnisse eines Registers zur Insulinpumpentherapie in Deutschland. Der Diabetologe 10: 43–47. Rayman, G., Kröger, J. and Bolinder J. on behalf of Diabetes UK (2018). Short report: Care delivery. could FreeStyle Libre™ sensor glucose data support decisions for safe driving? Diabetic Medicine 35: 491–494.

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Fanelli, C.G. (2004). Insulin therapy and hypoglycaemia: The size of the problem. Diabetes–Metabolism Research and Reviews 20 (Suppl. 2): 32–42. Finck, H. (2009). Sozialmedizinische Aspekte des Diabetes mellitus. Der Diabetologe 5: 371–385. Frier, B.M. (2008). How hypoglycemia can affect the life of a person with diabetes. Diabetes–Metabolism Research and Reviews 24 (2): 87–92. GOV.UK (2016). Diabetes Mellitus: Assessing Fitness to Drive. Last updated 14 February 2019 https://www.gov.uk/guidance/diabetes-­ mellitus-­assessing-­fitness-­to-­drive (last assessed 28.1.2020) Greitemann, B. (1998). Der diabetische Fuß: Ursachen – Prophylaxe – konservative und operative Therapie. In: H. Reike (ed.), Diabetisches Fuß-­Syndrom, pp. 166–182. Berlin: de Gruyter. Haffner, S.M., Letho, S., Rönnemaa, T. et al. (1998). Mortality from coronary heart disease in subjects with type 2 diabetes and in nondiabetic subjects with and without prior myocardial infarction. New England Journal of Medicine 339: 229–234. Hanefeld, M., Koehler, C., Henkel, E. et al. (2000). Post-­challenge hyperglycaemia relates more strongly than fasting hyperglycaemia with carotid intima-­media thickness: the RIAD Study. Diabetic Medicine 17: 835–840. Henrichs, H.R., Liebl, A., Reichel, A. et  al. (2009). Experimentelle Untersuchungen und klinische Evidenz der Insulinpumpentherapie (CSII). Diabetologie 4: 390–397. Hess, C., Thomas, A., Thevis, M. et al. (2012a). Simultaneous determination and validated quantification of human insulin and its synthetic analogues in human blood serum by immunoaffinity purification and mass spectrometry. Analytical and liquid chromatography-­ Bioanalytical Chemistry 404: 1813–1822. Hess, C., Stratmann, B., Quester, W. et al. (2012b). Clinical and forensic examinations of glycemic marker 1,5-­anhydroglucitol by means of high performance liquid chromatography tandem mass spectrometry. Forensic Science International 222: 132–136. Hirsch, I.B. (2005). Insulin analogues. New England Journal of Medicine 352 (2): 174–183. Hirsch, I.B., Balo, A.K., Sayer, K. et al. (2017). A simple composite metric for the assessment of glycemic status from continuous glucose monitoring data: Implications for clinical practice and the artificial pancreas. Diabetes Technology & Therapeutics 19 (Suppl. 3): 38–48. Holstein, A., Plaschke, A. and Egberts, E.H. (2001). Lower incidence of severe hypoglycemia in patients with type 2 diabetes treated with glimepiride versus glibenclamide. Diabetes–Metabolism Research and Reviews 17: 467–473. Holstein, A., Plaschke, A., Hammer, C. and Egberts, E.H. (2003). Characteristics and time course of severe glimepiride-­versus glibenclamide-­induced hypoglycemia. European Journal of Clinical Pharmacology 59: 91–97. Hostiuc, S., Negoi, I. and Hostiuc, M. (2016). Diabetes and collision risk. A meta-­analysis and meta-­regression. International Journal of Clinical Practice 70 (7): 554–568. International Diabetes Federation. IDF Diabetes Atlas, 9th edn. Brussels, Belgium: 2019. Available at: https://www.diabetesatlas.org (last accessed 29 January 2020). Klinkert, C. and Quester, W. (2004). Insulinpumpentherapie bei Typ1-­ Diabetes mellitus. Deutsche Medizinische Wochenschrift 129: 1149–1154.

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Stork, A., Van Haeften, T.W. and Veneman, T. (2007). The decision not to drive during hypoglycemia in patients with type 1 and type 2 diabetes according to hypoglycemia awareness. Diabetes Care 30 (11): 2822–2826. Thomas, A. (2019). “Time in Range” – ein neuer “Goldstandard” für die Beurteilung der Glykämie? Oder gar der neue Goldstandard? Diabetes Stoffwechsel und Herz 28 (Diabetes, Metabolism, and the Heart 28): 146–149. Truninger, R., Uthoff, H., Capraro, J. et al. (2013). Glucose control during a driving training in patients with type 1 and type 2 diabetes mellitus – a randomised, controlled trial. Experimental and Clinical Endocrinology & Diabetes 121: 420–424. Vaa, T. (2003). Impairments, Diseases, Age and Their Relative Risks of Accident Involvement: Results from Meta-­ Analysis, IMMORTAL. http://www.immortal.or.at (last accessed 29 January 2013). Williams, B., Mancia, G., Spiering, W. et  al. (2018). 2018 ESC/ESH Guidelines for the management of arterial hypertension: The Task Force for the management of arterial hypertension of the European Society of Cardiology (ESC) and the European Society of Hypertension. European Heart Journal 39 (33): 3021–3104. Yu, X., Lin L., Shen J., Chen Z. et al. (2018). Calculating the mean amplitude of glycemic excursions from continuous glucose data using an open-­code programmable algorithm based on the integer nonlinear method. Hindawi Computational and Mathematical Methods in Medicine 2018, Article ID 6286893, 9 pages.

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Reddy, M., Jungee, N., El Laboudi, A. et al. (2018). A randomized controlled pilot study of continuous glucose monitoring and flash glucose monitoring in people with Type 1 diabetes and impaired awareness of hypoglycaemia. Diabetic Medicine 35: 483–490 Ridel, D., Borowsky, A., Parmet, Y. et al. (2019). Acute Hyperglycemia may impair driving skills of young T1DM patients. Proceedings of the Human Factors and Ergonomics Society 2019 Annual Meeting. Riehl, J., Biermann, E. and Standl, E. (2002). Insulinresistenz und Typ-­2-­diabetes. Die IRIS-­Studie. Diabetes und Stoffwechsel 11: 150–158. Schäfer-­Graf, U., Laubner, K., Hummel, S. et al. (2019). Gestationsdiabetes mellitus (GDM), Diagnostik, Therapie und Nachsorge. Diabetologie 14 (Suppl. 2): 196–206. Schmied, L.S. and Zulewski, H. (2019). Glucose variations during driving in people with type 1 diabetes using a continuous glucose monitoring system. Diabetes Care 42: 1340–1343. Service, F.J., Molnar, G.D., Rosevear, J.W. et al. (1970). Mean amplitude of glycemic excursions, a measure of diabetic instability. Diabetes 1970 Sep; 19 (9): 644–655. Skurveit, S., Strøm, H., Skrivarhaug, T. et al. (2009). Road traffic accident risk in patients with diabetes mellitus receiving blood glucose-­ lowering drugs. Diabetic Medicine 26 (4): 404–408. Stork, A., Van Haeften, T.W. and Veneman, T. (2006). Diabetes and driving. Desired data, research methods and their pitfalls, current knowledge, and future research. Diabetes Care 29 (8): 1942–1949.

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Epidemiology and Causal Factors in Fitness to Drive

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analyse factors influencing occurrence rates (analytical epidemiology). For a sound scientific description of epidemiological study types, compare Kleinbaum et  al. (1982), Howe and Choi (1983) and Shadis et al. (2002).

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Epidemiology in traffic medicine needs to detect and to quantify the risks for traffic safety caused by drivers and other road users. The aim of epidemiological research is to quantify and rank causal factors that may emerge through experience, physiological knowledge or descriptive or experimental studies. Epidemiology, unlike experimental research, is primarily aimed at detecting real risk rather than potential risk. A particular illegal drug, for example, may have negative effects on fitness to drive, but if nobody uses the drug or drives a car when under the influence of this drug, the drug is of no relevance for road traffic safety. Hence, epidemiology makes it possible to rank the real risks (e.g. ‘crash’ as a statistical unit) created by causal factors. Section 58.2 looks at the essential methods of epidemiological research as well as their appropriate quality criteria based on publications by Berghaus and Krüger (2007), Berghaus et  al. (2007), Berghaus and Hilgers (2009), Drummer (2009) and Houwing et al. (2009). In Section 58.3, the full scope of causal factors is weighted and the risk of the influencing factors on road traffic safety outlined in Chapter 52 is discussed, providing some examples.

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58.1 Introduction

58.2  Epidemiological methods Epidemiological methods can be roughly divided in two groups: (i) those methods that provide frequency measures of safety relevant events (descriptive epidemiology) and (ii) methods that

Handbook of Forensic Medicine, Volume 3, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

58.2.1  Descriptive epidemiology

Prevalence and incidence are the central terms in descriptive epidemiology. Prevalence gives an estimate of the event probability in a defined population. It is given for a single time point (point prevalence) or is related to a time period (period prevalence). Prevalence related, for example, to events over the past year and recent events is used for the estimate of crash frequency over 1 year. In contrast, incidence relates only to newly observed crashes within a small space of time. It can be used, for example, to show the increase in the frequency of crashes per time unit. It seems trivial, but when reading articles in newspapers or listening to statistical information on road traffic by politicians, it is important to remember that an accurate interpretation of prevalence and incidence is only possible given clear definitions of the event (variable and its classification) and, especially, of the reference population. Hence, there are considerable differences between the statements: ‘20% of drivers had cannabis in their blood’, ‘20% of drivers with psychoactive substances in their blood had cannabis in blood’, ‘20% of drivers involved in a crash operated the car under the influence of cannabis’ and ‘20% of crashes are caused by cannabis’. Information concerning safety-­relevant events can be collected from the following sources:

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driving fitness if it can be compared with other figures, such as the frequency of use of a drug in the given region and time period. Roadside surveys are cross-­sectional studies. In contrast to crash surveys, roadside surveys are intended to provide an overview of the frequency of a defined causal factor in a well-­defined time period and region in crash-­free drivers in normal traffic. Two requirements are of special interest concerning the quality of a roadside survey study. Since it is impossible to include all drivers, a representative sample of the population of all drivers has to be used in the survey. Therefore, the random sample has to be adjusted for essential characteristics (age, sex of driver, region, time of day, type of road, etc.) compared with total drivers. The number of non-­responders (i.e. the number of drivers who refuse to participate in the survey) should be as small as possible. The smaller the number of non-­responders, the more reliable the estimate of prevalence. At first glance it seems meaningful to compare the results of a crash survey with the findings of a roadside survey with respect to a defined causal factor, using the figures of a roadside survey as a control for the results of a crash survey. But the outcome of such a comparison can only be a first hint for an increase of risk. In general, roadside surveys and crash surveys disagree in many variables of the study designs and hence are not comparable. A substantial disadvantage of crash surveys and roadside surveys is the fact that it is, for legal reasons, impossible to take specimens from drivers without their consent in many countries. Frequently, it is those drivers who have just used drugs before driving who will refuse to give a specimen. As a result, crash surveys and roadside surveys are seldom performed. It is only in recent years that some countries have started to regulate by law taking blood of drivers, depending on the seriousness of the injuries of the persons involved in a crash. The reader is referred to Assum et al. (2007), Pil et al. (2007) and Berghaus and Hilgers (2009) for further information on crash surveys and roadside surveys. Interviewing road traffic participants is a methodological approach with a lot of potential. Even very small populations (users of unusual medicines, rare diseases, younger people with diseases, etc.) or special groups of traffic participants (e.g. younger or older people) can be analysed as well as problems that cannot be solved by an experimental or epidemiological approach. For example, the question about the motivation of driving under the influence of a drug is an essential one in promoting traffic safety. In addition to core questions concerning the kind, frequency and time of participating in road traffic, with or without the preceding use of drugs, questions about near accidents and about the subjective rating of driving aptitude and driver fitness, as well as many others, can be asked. Besides general quality control, additional quality aspects are necessary when performing interviews with road users  – for example clear-­cut standardised questions concentrating on essential topics, the use of empirically validated scales, attempting to control the validity of answers and using a trained interviewer. Computer meta-­analyses are aimed at gathering and meaningfully analysing the results from as many publications on a defined

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Official statistics. Crash surveys or hospital studies. Roadside surveys. Questionnaires and interviews with road users. Meta-­analyses. In developed countries, official statistics are published in statistical annals. In addition to data on the number and kind of cars, licences, the road system, traffic volume, etc., official statistics may include (depending on the country) the number of road traffic crashes distinguished according to crash location (in town, out of town, highway, etc.), crash severity (with or without property damage, with injured and with killed persons), cause of crash (speed, right of way, car distance, driver fitness, etc.), time of crash, age and gender of involved drivers and other criteria. Normally, official statistics describe an entity that covers the road traffic crashes of a whole country, a whole state or a special region. Absolute risks can be calculated by relating crash data to data relevant to road traffic, for example, miles driven, number of vehicles, number of trips, population, number of licensed drivers, etc. This allows a comparison of absolute numbers between cities or states within countries. In addition, time series of crashes or absolute risks within a country can be established, giving a good view on the developing nature of crashes. In this connection, it is essential to look at the definitions and categorisations of variables because from time to time definitions are changed so that new data are no longer comparable with older data. In theory it should be possible to provide a valuable information basis for transport policy comparing the official statistics of different countries. But there are two obstacles to this: (i) there are important differences concerning cultural and legal backgrounds, mobility, road systems, traffic rules (e.g. speed limits), authority of the police and many more influencing factors on crash detection and frequency; and (ii) the different definitions and categorisations of variables. So, to give only one example, the definition of a ‘crash with killed persons’ is different between European countries depending on the period of time that has passed between the crash and death. The period ranges between ‘killed immediately at the scene of the crash’ and ‘died within a (country-­specific) defined period of time after the crash’. Hence, it is difficult to categorise crash figures between countries in terms of ‘better’ or ‘worse’. Crash surveys, also referred to as hospital studies, are performed with the objective of estimating the frequency of a defined causal factor in crash scenes in a well-­defined time period and region. The sample can consist of all crashes or a representative part of all crashes. Concerning the causal factor ‘effects of drugs’, for example, the sample could consist of all drivers in a region who are injured in a crash and who are brought to a hospital or who are killed using blood or other specimens taken from each driver independent of culpability for the crash and screened chemically/toxicologically for a defined number of drugs. Crash surveys provide no evidence of whether the detected influencing factor really was the cause of the crash. The frequency of drugs detected only gives a first hint about the influence of drugs on

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Odds ratio OR of being injured after substance use C :C : F : F

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Methodological approaches of analytical epidemiology (analysing factors determining occurrence rates) comprise the following: • Case–control studies. • Culpability (responsibility) studies. • Pharmaco-­epidemiological studies. Case–control studies provide an optimal method of estimating the risk of being involved, injured or killed in a crash under the effect of a defined causal factor (disease, drug, etc.). The principle of a case–control study consists in contrasting a risk factor, for example, the effect of a drug, with an outcome measure, for example, an injury resulting from a crash. The risk can be calculated if the exposure is measured for a random sample of cases as well as for a comparable sample of controls. In contrast to comparing the results between independently performed roadside surveys and crash surveys, the data of a case–control study are comparable a priori due to the identical epidemiological design. The procedure and calculation of risk factors for the example ‘driver injured in a crash’ are as follows: • Target population: car drivers. • Cases: car drivers injured in a crash. • Controls: comparable car drivers without a crash. • Risk factor: defined psychoactive substance in the blood, with a defined cut-­off value.

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58.2.2  Analytical epidemiology

threefold as likely in the population of crash drivers. Besides the OR itself, the study should state if the OR is statistically significant and/or the confidence interval (CI) should be calculated using the data. If the CI includes 1, the OR is not significant, which means that the causal factor has no influence on the crash scene. Some other aspects of case–control studies have to be mentioned. With respect to the target population, one has to differentiate between drivers or passengers in a crash (not injured, injured and killed) and between drivers culpable or not culpable for the crash. The OR provides information on the involvement risk (the risk of being involved in a drug crash and being either not injured, injured or killed) and on the causation risk (the risk of causing a crash under the effect of the drug and being either not injured, injured or killed). up of the control group, Depending on the make-­ population-­based case–control studies have to be differentiated from matched case–control studies. A population-­based case–control study would be where a roadside survey and at the same time a crash survey are performed in the same region. Since the data of the control group have to be comparable to the data of the case group, the data have to be adjusted before analysis with respect to confounding variables such as age, gender of drivers, time and location of crashes, etc. Therefore, it useful to perform a matched case–control study. Typical characteristics of all crashes of a region are determined (location, time, etc.). If every case is related to a corresponding control (matching), the distribution of confounding variables is the same in the two groups. Assuming that the exposure of the risk factor and the sample size are high enough and the risk factor (e.g. alcohol) can be categorised in classes, ORs can be calculated for the individual classes (e.g. concentration-­dependent ORs for alcohol). Hence, the risk potential can be illustrated as a hazard curve. Apart from difficulties concerning the selection of cases and controls, and the non-­responder rate, the need for ethical approval can complicate the realisation and interpretation of a case–control study. For further information on such studies, including a formula for calculating a relative risk (RR = (C+: all positives):(C−: all negatives)), compare Schlesselmann (1982), Howe and Choi (1983) and Houwing et al. (2009). The culpability analysis (or responsibility analysis) is a modification of the case–control method. It is used for analysing the influence of drugs on road traffic safety and is more economical than case–control studies. Instead of selecting the control sample from crash-­free road traffic, specimens (usually blood) are taken from all drivers in a region who are involved in a crash. In addition, it is determined without knowledge of drug influence if a driver is responsible or not for the crash. The drug-­free drivers are used as ‘controls’. They provide the baseline culpability rate ((F+:F−), where + means culpable) which is compared with the culpability rate of the drug-­positive group (C+:C−):

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topic (e.g. experimental or epidemiological studies on alcohol, illegal drugs and medicines) as possible. The first thing is to have an acceptable number of publications so a qualitative screening can be undertaken. The relevant information from the remaining studies can be stored in standardised mode in a computer for statistical analysis. In contrast to traditional reviews based on only a few publications, meta-­analyses allow information from many studies to be condensed and the analysis of target variables dependent on many influencing factors.

where Positive cases=car drivers injured in a crash with the substance (C+) Negative cases=car drivers injured in a crash without the ­substance (C+) Positive controls=crash-free drivers with the substance (F+) Negative controls=crash-free drivers without the substance (F–) An OR of 1 indicates that the use of the substance is equally likely in crash drivers and crash-­free drivers. An OR of 1 the use of the substance is more likely in crash drivers. An OR of, for example, 3 for a defined drug means that the use of the drug versus the non-­use of the drug is



OR

C :C

: F :F



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In multi-­ centre epidemiological studies, it is particularly important to ensure that the same procedures are used in all individual centres. However, this can be difficult to achieve, especially for centres in different countries, as there are some factors that cannot be standardised. Extensive studies should be preceded by a pilot study to check quality assurance and economics. Quality aspects cannot be covered in enough depth here, so the reader is directed to the citations in this chapter and to Ferrara (1992) and Simpson und Vingilis (1992).

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58.3  Examples of epidemiological studies

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58.3.1  Preliminary remarks: dependence of results on the individual design of a study

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In the preceding section, the concepts of individual methodological approaches were introduced without going into details of the methods and how they can influence the results. Before reporting concrete examples of epidemiological studies, it is important to demonstrate the degree of alteration of study results through apparently small modifications of the design or calculation procedures. As an example we can look at the influence of adjustment variables on the findings of five case–control studies calculating ORs for the effects of Δ-­9-­tetrahydrocannabinol (THC) (Table 58.1). From the multitude of different design aspects, only the country and the effects of different adjustment variables are shown in the table. Besides the considerable variation of the unadjusted ORs (the real differences between the ORs are also affected by the differences in study design), Table  58.1 illustrates how the adjustment procedure alters the ORs. In particular the studies of Haworth et al. (1997) and Blows et al. (2004) indicate how different the ORs are after different adjustments even if other aspects of the design remain the same. A statistically significant increase can change to a non-­significant result. If a single calculation procedure within a study can make such an explicit difference in the ORs, one can imagine the degree of variability if, in addition, other parts of the design are different. The number of factors that impact the target variable is great. Based on an inspection of published epidemiological studies, Table  58.2 lists some of the multitude of possible variations between study designs in case–control studies. A glance at this list immediately indicates the extent of variation in study designs. Even the definition of individual characteristics may be different, influencing the result of a study. How is the degree of injury defined? For example, for ‘moderately injured’, is this only at the first aid point? Or is hospitalisation up to a defined time period? Or is it only treated at the scene? This means that when giving examples of different methodological approaches in the following discussion I primarily focus only on results as it is not possible to consider all the influencing

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The division into ‘culpable’ or ‘not culpable’ is of fundamental importance. There are a lot of different procedures. Drummer (2009) categorises the types of culpability determination from the simplest methods to sophisticated ones into ‘at fault assessment’, ‘graduated culpability rate’ and ‘semiquantitative assessment of culpability’. Even for the culpability analysis, statistical adjustment is necessary to account for variations (e.g. age, gender, time of day and type of crash). Compare Robertson and Drummer (1994) and Drummer (2009) for further information on culpability analysis. Due to the required large random sample sizes, case–control studies and culpability studies are only meaningful for those causal factors where the exposure is high enough, especially for alcohol. Concerning medicaments, for example, these studies can only be used for the whole groups of drugs (e.g. benzodiazepines) and not for individual substances. Therefore, pharmaco-­ epidemiological studies have been introduced into the epidemiological scope of these methods. These studies are based on two data sets: (i) patient-­based prescription data from defined medicines in a defined time period and a defined region and (ii) data of drivers involved in a traffic accident at the same time and in the same region as the prescriptions (using crash registers and/or emergency and hospital data). The OR can be calculated if one compares the data sets and if, additionally, a representative control group without medicine use is established. Apart from the fact that the four groups necessary to calculate the OR may result from four different populations (which intensifies the problem of bias), pharmaco-­epidemiological studies include the disadvantage that one does not know if patients really use their medicines or if other factors contribute to a crash (e.g. the use of additional psychoactive substances).

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58.2.3  Quality assurance

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To finish this section on epidemiological methods, it is very important to look at quality assurance. It should be obvious that a study only can be interpreted adequately if a lot of quality criteria are fulfilled. Such quality criteria pertain to all aspects of epidemiological studies, for example: • Sample: unambiguous definition of the population, sample size, representativeness, selection of undesirable determinants, as few non-­responders as possible, etc. • Design: region, time of day and year, lighting, road design features, kind of specimen, number of drugs included in the chemical analysis, cut-­offs, detection methods, etc. • Data collection: relevance, information content, unambiguous definitions, completeness, homogeneity, etc. • Preparation and analysis of data: adjusting to confounding variables, weighting, homogeneity, adequate statistical procedures, calculation of CI (probability at most 95%), etc. • Interpretation: adequate concerning sample, data compilation and controls, analysis of selection effects, etc. • Publication: including all important information, even negative aspects, etc.

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Table 58.1  Odds ratios of THC from various case–control studies: dependence of the results on adjustment variables. ORs (95% CI) unadjusted

ORs adjusteda

Country

Reference

38.2 (13.8–105.8)

6.4–35.6

Australia

Haworth et al. 1997

2.0 (1.4–2.9)

1.6

Canada

Brault et al. 2004

1.45 (0.64–3.29)

1.29

Netherlands

Mathijssen and Houwing 2005

11.4 (3.6–35.4)

0.8–6.0

New Zealand

Blows et al. 2004

0.19–51.02

0.29–28.88

Nine European countries

Hels et al. 2011

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CI, confidence interval; OR, odds ratio; THC, Δ-­9-­tetrahydrocannabinol. a  Adjusted for different groups out of the following confounding variables: blood alcohol concentration, age, sex, hour, day, year, quarter of the year, ethnicity, driving exposure, vehicle age, seatbelt use and speed.

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Table 58.2  Selection of determinants affecting the results of an epidemiological study. Characteristics

Country

Culture, law, traffic rules, road system

Study design

City, rural region, mixed duration of study

Sample size (random error)

Small to large

Cases (crashes)

Kind of crash: single vehicle, multiple vehicle; crashes with fatally, seriously, moderately, not injured participants; cars, vans, other vehicles Data: only one hospital, multiple hospitals, hospital and forensic science institute Driver, passenger/s Driver characteristics: age, sex, profession, etc. of the driver, annual mileage, risky behaviour tendency Diseases: severity of diseases, duration of treatment, with and without other influencing factors (diseases, co-­morbidities, concomitant drugs) Number and kind of drugs

Controls (adjustment/ confounding variables)

Alcohol-­free, illegal drugs-­free, medicines-­free, free of all drugs Traffic vs. non-­traffic Matched or population based criteria: vehicle type (cars, vans and other vehicles); season, weather conditions, time of month, week and day, lighting, road type; age, sex and other characteristics of the driver (ethnicity, driving exposure, seatbelt use, speed and profession)

Selection of unsuitable cases and controls

Sleep disorders, pregnancy, diseases, etc.

Taking and measuring of the causal factors (cases/ controls)

Drugs: Matrix: blood (whole blood, serum), urine, sweat, saliva Interview (subjective) Taking technique of the specimen Conditions of transportation and storage Kind and number of substances searched for chemical–toxicological detection methods Cut-­offs and threshold value Estimation and estimation method of concentrations for time of crash Diseases: Method of diagnosis Phase and severity

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Determinant

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factors that would be necessary to evaluate the results. For an explicit description, it is essential to look at the original publication because even in traditional reviews it is impossible to summarise all aspects of the individual design. Many sources of original studies are listed in the book edited by Verster et  al. (2009) and in the epidemiological part of the DRUID (Driving under the Influence of Drugs, Alcohol and Medicines) project, especially Hels et al. (2011).

58.3.2 Overall spectrum of causal factors To start with, this section gives an overview of those causal factors that are the most common ones with respect to traffic safety. More than 90% of all road traffic crashes in developed countries are caused by human factors, about 5% by environmental conditions, 1% by technical deficits and about 1% by other reasons (GIDAS database, quoted in Golz et al. (2010)).

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Table 58.3  Number of crashes with injured persons as a result of human factors in 2003. Based on statistics from the Federal Office of Statistics, Germany.

78 728

Turn/turn-­over, driving backwards, driving in and out of running traffic

65 558

Right of way

63 896

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Speed

Distance

50 771

Road use

32 030

Fitness to drive Affected by alcohol

28 959 22 674 17 961

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17 677

Other mistakes by drivers

87 713

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Overall

443 293

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Source: After Hasse and Schepers (2007).

Table 58.4  Types of ‘driver unfitness’ (see Table 58.3). Cause of crash

Absolute number of crashes

Percentage

Effects of alcohol

22 674

78

Overtiredness

2034

7

Illegal drugs and medicines

1341

5

Other physical and mental disorders

2910

10

Overall

28 959

100

Source: After Albrecht (2007), © Ärzte-­Verlag, Köln.

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Number of crashes

The majority of epidemiological studies have studied the effect of drugs on fitness to drive, especially alcohol. Before summarising findings on individual substances, their ranking should be discussed as well as the importance of simultaneous use of ­ ­different drugs. A large multicentre roadside survey was performed in the context of the DRUID project, with 13 European countries participating. The data were gathered and prepared according to a generally uniform design (Houwing et al. 2011). In total, almost 50  000 randomly selected drivers participated between January 2007 and July 2009 in this study. Twenty-­three important psychoactive substances were included in the core substance list. The main results of this roadside survey are summarised as follows (Houwing et al. 2011, pp. 4–5): • Alcohol is still by far the number one psychoactive substance on European roads, followed by illicit drugs and medicinal drugs. • On a European level, alcohol is estimated to be used by 3.48% of the drivers, illicit drugs by 1.90% of the drivers, medicinal drugs by 1.36% of the drivers, drug–drug combinations by 0.39% of the drivers and alcohol–drug combinations by 0.37% of the drivers. • For illicit drugs, THC is the most frequently detected drug in traffic followed by cocaine. • Amphetamines and illicit opiates were less frequently detected. • Illicit drugs were in general mainly detected among young male drivers during all times of the day but mainly in the weekend. • Medicinal drugs were in general mainly detected among older female drivers during daytime hours. • Benzodiazepines were the most prevalent medicinal drug in traffic, and Z-­drugs were less prevalent. However, considerable differences between countries were present. The use of substances among drivers in the general driving population in Europe (prevalence) varies very much per country, but general patterns can be distinguished on the level of European regions: • The medicinal drugs, Z-­drugs and medicinal opiates and opioids were in general relatively frequently detected in Northern European countries. • Illicit drugs, alcohol and benzodiazepines are relatively frequently detected in Southern European countries. • In Eastern Europe, the prevalence of alcohol and drugs was relatively low compared to the other European regions. • In Western Europe, drug use is more or less on the European average. On the basis of crash studies, Isalberti et  al. (2011) reported within the same DRUID project on the prevalence of alcohol and other psychoactive substances in injured and killed drivers. Specimens of seriously injured drivers were sampled in a uniform design between October 2007 and May 2010 (n = 2492 car and van drivers in six European countries) and from killed drivers sampled between January 2006 and December 2009 (n = 1118 car and van drivers in four European countries). The non-­ responder rates

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58.3.3 Overall spectrum of drugs

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To differentiate the human factors, the official statistics of Germany are used as an example. Table  58.3 lists the human errors of drivers involved in crashes with injured persons, according to the frequency of occurrence, with speed being the most frequent. Lack of fitness to drive covers 6.5% of crashes and ranks only sixth. This causal factor includes effects of alcohol, illegal drugs and medicaments, overtiredness and other physical and mental deficits. The distribution of the 28 959 cases in the category ‘fitness to drive’ are shown in more detail in Table 58.4. This demonstrates the predominance of alcohol effects: about 80% of mistakes happened under the effect of alcohol compared with only 20% due to other causal factors. However, there are considerable shortcomings in official statistics, for example, it is almost impossible to identify the real cause of a crash. Concerning ‘overtiredness’, it is difficult to observe this causal factor after an individual crash. Even a ‘physical or mental disorder’ will only be considered if the drivers themselves mention the disease. For alcohol, illegal drugs and medicines, the police in Germany are only allowed to enforce a test if they identify abnormal behaviour by the driver.

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Table 58.6  Relative risk level of getting seriously injured or killed (based on ORs of nine European countries) when taking different substances and driving. Substance group

Risk

Risk level

1–3

Slightly increased

Benzoylecgonine Cocaine Illicit opiates Benzodiazepines and Z-­drugs Medicinal opioids

2–10

Mediumly increased

Alcohol (all concentrations) Amphetamines Multiple drugs

5–30

Highly increased

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20–200

Extremely increased

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Hels et al. (2011) point out that the risk estimates for cannabis and amphetamines (very different single country estimates) and for benzoylecgonine, cocaine and illicit opiates (few positive cases and controls) must be treated with caution. Source: After Hels et al. (2011, Table 33, p. 42).

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Hels et al. (2011) combined the roadside surveys (Houwing et al. 2011) and the case studies (Isalberti et  al. 2011) into a case–­ control study. In total, 2490 seriously injured drivers (in Denmark, Finland, Lithuania, Italy, Belgium and the Netherlands) and 1112  killed drivers (in Finland, Norway, Sweden and Portugal) were included, and 15  832 drivers participated in the control sample of the seriously injured drivers and 21 917 drivers in the control samples of killed drivers. Based on the ORs of the individual European countries, the authors weighted the data for the national distribution of traffic in each of eight time periods of the week and ranked the different substances according to their relative risks (adjusted for age and gender) (Table 58.6). The results of culpability studies in samples of injured drivers demonstrated, depending on the individual design of the study, increased risks for alcohol, THC, benzodiazepines, stimulants and opioids with considerably increased ORs for combinations of psychoactive drugs (e.g. Soderstrom et  al. 1990, 2005; Terhune et  al. 1992; Benzodiazepine/Driving Collaborative Group  1993; Longo et  al. 2000b; Lowenstein and Koziol-­ McLain  2001; Drummer et al. 2004; for more citations see Drummer 2009). In these studies, the ORs are generally ‘means’ because the prevalence of almost all the agents was so small that concentration-­ dependent calculations of ORs could not be carried out. For those few substances where the concentration-­dependent calculations were possible (mainly for alcohol, and sometimes for THC or benzodiazepines), the risk of participating in a crash as an injured or killed person or even causing a crash is considerably higher with raised concentrations of an agent in the blood (Box 58.1).

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­ iffered between 0% and 8.5% for injured drivers and between d 5.7% and 41% for killed drivers. Drug levels were conducted on blood samples. The percentage of subjects testing positive for at least one psychoactive substance ranged between 28% and 53% for seriously injured drivers and between 31% and 48% for killed drivers. For both groups, the majority of drugs appeared to be used in combination with other psychoactive substances. The percentage distribution of drug-­positive cases is shown in Table 58.5. Alcohol was the most common finding, both in seriously injured and in killed drivers. Among all seriously injured drivers, THC and benzodiazepines were the next most common. Benzodiazepines, amphetamines and THC took places 2–4 in the killed drivers. The prevalence ranges mentioned in these studies represent in essence the international findings for these study types, as can be shown with two examples. Maes et  al. (1999) summarised the results of crash surveys performed in Spain, Norway and Belgium. Alcohol was detected in 27–63% of the blood specimens, and illegal drugs or medicaments in 14–24%. Cannabis and the benzodiazepine group were leading substances within the groups of illegal drugs and medicaments. From 2500  injured drivers in Australia, about one-­quarter had at least one psychoactive agent in blood. Alcohol (8.6%) was the most frequently detected followed by THC (2.8%) and benzodiazepines (1.8%) and stimulating agents (0.8%) (Longo et al. 2000a). The considerable differences between the results of the roadside studies and the crash/hospital studies raised the suspicion of the risk potential of the various substances and substance groups.

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Table 58.5  Prevalence of use of substance groups in drug-­positive drivers. Percentage of drivers positive Seriously injured drivers

Killed drivers

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Substance

19.0–44.9

16.1–38.2

16.3–35.1

0.1–2.6

0.0–7.4

Benzoylecgonine but negative for cocaine

0.0–2.8

0.0–0.7

Cocaine

17.7–42.5

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Alcohol (≥0.1 g/L) Alcohol (≥0.5 g/L)

0.0–2.7

0.0–0.7

Cocaine and/or benzoylecgonine

0.0–5.4

0.0–1.4

THC–COOH but negative for THC

0.0–5.3

0.0–5.2

THC

0.5–7.6

0.0–6.1

THC and/or THC––COOH

1.6–9.9

1.3–6.1

Illicit opiates

0.0–2.1

0.0

Benzodiazepines

0.0–10.2

1.8–13.9

Z-­drugs (zopiclone, zolpidem and zaleplon

0.0–3.8

0.0–4.4

Medicinal opioids

0.5–7.8

1.7–2.1

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Amphetamines

THC, Δ-­9-­tetrahydrocannabinol; THC-­COOH, 11-­nor-­9-­carboxy-­THC. Source: Summarised from Isalberti et al. (2011, p. 4).

Alcohol Due to the high prevalence of alcohol involvement in crashes and roadside surveys, many concentration-­dependent studies could be performed. The most famous one, well known as the

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Box 58.1  Rank order of drugs within the causal factor ‘drug use’.

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• Roadside surveys: Alcohol is by far the number one drug taken by on road users, followed by illegal drugs and medicines. • Crash studies: Alcohol is the most dangerous agent in crashes with seriously injured or killed persons, followed by illicit drugs (THC, amphetamines) and medicines (benzodiazepines); simultaneous use of several psychoactive drugs plays a prominent role in crashes. • Case–control/culpability studies: The risk level concerning crashes with seriously injured or killed persons is extremely increased by alcohol and simultaneous drug use. • For all drugs: There is a positive correlation between the concentration of a substance in the blood and the level of risk, with increasing concentration of the agent the risk increases dramatically.

represents a maximum of crashes caused by medicaments. Diazepam was most frequently detected followed by other benzodiazepines. One of the few studies that differentiated according to the level of concentration of the medicament (Longo et al. 2000b) showed for benzodiazepines an OR of 1.3 for sub-­therapeutic levels, of 3.3 for therapeutic concentrations and of 3.6 for over-­therapeutic/ toxic levels. Hence, even for medicaments there is a positive correlation between risk and concentration. Owing to the frequently detected, relatively high concentrations, in addition to the simultaneous verification of other drugs, it seemed not to be the ‘normal’ patient using a medicament over a short period that is at risk of a crash but more those dependent on medicaments or illegal drugs with a parallel use of other drugs. Since the prevalence of most of the medicines was very small, a pharmaco-­epidemiological approach was tried to close the knowledge gap. A very detailed study was performed by Ravera and de Gier (2010) in the Netherlands. On the basis of pharmacy prescription data, police traffic accident data and driving licence data for a total study population of 4784, ORs were calculated, using an alcohol-­free population of 3963 cases (adults who had a traffic accident between 2000 and 2007 and were driving and received medical assistance) and 18 828 controls (adults who had a driving licence but had no traffic accident during the study period; controls were matched for sex, age, zip code and data of the accident). Various variables, such as age, sex, medicine half-­life, mono and combination therapy, alcohol use and categorisation of the medicines were considered for the analysis. Crude ORs showed a positive association between the risk of having a traffic accident and the exposure to at least one psychotropic medication (opioids, antipsychotics, anxiolytics, hypnotics and sedatives, antidepressants and antihistamines for systemic use) (OR 1.28; 95% CI 1.12– 1.46). The association was found to be higher in combination therapy users, new users, intermediate and long half-­life benzodiazepine users, female users and young/middle-­ aged users. Concerning the different psychotropic medicine group users, the crude ORs ranged between 1.01 and 1.59 with statistical significance for anxiolytics, hypnotics and antidepressants. For further details, see the original study (Ravera and de Gier 2010). Earlier pharmaco-­epidemiological studies (Neutel 1995, 1998; Hemmelgarn et  al. 1997; Barbone et  al. 1998) in part showed higher ORs. The following ORs were ascertained: benzodiazepines 0.9–5.2, opioid analgesics 1.8, antidepressants 0.9–2.3, antihistamines 0.7–1.8, asthma agents 2.9 and diuretics 2.9. The risks were higher when starting a therapy with medicaments than later in the therapy period. Patients using for the first time a medicament and long half-­life benzodiazepines showed an elevated risk.

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≥0.1 to 80 years) before re-­licencing, including tests of their eyesight and physical and mental health as well as prohibiting the use of handheld cellular phones while driving (Shiner  2007; Jones et al. 2019). The dangers of drinking alcohol before driving an ‘automobile wagon’ were recognised early in the history of transportation as evidenced by an editorial appearing in the 1907 issue of Quarterly Journal of Inebriety (Crothers 1904). Despite these early warning signs, it took several decades before laws were passed that regulated the use of alcohol before driving (Jones et al. 2019). Norway was the first country to enact a statutory blood-­alcohol concentration (BAC) limit for driving in 1936 followed by Sweden in 1941. However, there was lack of agreement as to what limit should be enforced, and Norway opted for 0.50 g/L and Sweden set a limit of 0.80 g/L, whereas many states of America enforced a BAC limit of 1.5 g/L as prima facie evidence of being drunk at the wheel (Lerner 2011). Even today, statutory BAC limits for driving differ between countries from 0.20 g/L (Norway, Poland, Sweden and China) to 0.80 g/L (USA, Canada, England and Wales). This fourfold difference probably reflects local politics rather than results of traffic-­safety research and the risk of causing a crash

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According to a report from the World Health Organization (WHO), as many as 1.34 million people were killed in road-­traffic crashes in 2016 (WHO  2018). These fatalities were the leading cause of death in children and young adults aged between 5 and 29 years. In addition, the WHO report also mentioned that between 20 and 50  million people suffered non-­fatal injuries, which often needed emergency medical treatment. Heavy drinking and drunkenness, especially in connection with driving, is a global public health problem (Hanson and Li  2003; Room et al. 2005). The underlying cause or causes of road-­traffic crashes are complex and involve an interaction between three main factors: the driver, the vehicle and the environment (Rolison et al. 2018). Traffic medicine research is a multi-­disciplinary topic dealing with epidemiology of road-­ traffic crashes (Christophersen et al. 2016), ways to prevent these from happening (Voas 2020), culpability for the crash (Poulsen et  al.  2014), and the long  term  consequences for public health and longevity (Gopalakrishnan 2012). Traffic safety is a top priority for many government agencies as well as for the manufacturers of automobiles, and efforts to produce safer vehicles are high on the agenda. This might involve stronger construction aimed at reducing the harm to the driver and passengers if a crash occurs and development of computer software to pre-­warn of potential dangers in the traffic flow, such as speeding, braking, lane positioning as well as traditional safety features, such as seat belts and air bags, both of which have saved countless lives (Voas 2020). The notion of driverless vehicles is no

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59.1 Introduction

Handbook of Forensic Medicine, Volume 3, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

Environment

Age and gender Driving experience Personality Fatigue/sleep loss Using a cellular phone Alcohol ingestion Drugs, medication State of health

Weather (rain, snow) Road conditions Day or night time Traffic intensity Passengers in vehicle

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Figure 59.1  Examples of factors that might contribute to a road-­traffic crash including aspects of the vehicle, the driver and the environment.

59.2  Properties of alcohol Alcohol (ethanol) is a legal drug, and light-­to-­moderate drinking should not pose a danger for health and longevity. However, heavy drinking and drunkenness are major public health problems worldwide (Probst et al. 2020). The principal psychoactive substance in all alcoholic beverages is ethanol (CH3CH2OH), which acts as a depressant of the central nervous system (Goldstein 1983). The mechanism of action of ethanol on the central nervous system (brain and spinal cord) resembles the effects produced by barbiturate and benzodiazepine drugs and also the general anaesthetic gases (Wallner and Olsen 2008). However, because of ethanol’s high solubility in water and low solubility in lipids, larger quantities must be ingested to cause impairment of normal body functioning. The clinical manifestations of ethanol intoxication depend on many factors, such as the type of drink consumed (beer, wine or liquor), and the dose ingested and speed of drinking (Jones  2019a). The consumption of ethanol on an empty stomach leads to a higher and earlier occurring peak BAC compared with the same quantity ingested after or together with a meal. Another factor to consider is habituation, and therefore previous experience with alcohol consumption and the development of acute and chronic tolerance to effects on the brain (Kalant 1998; Cromer et al. 2010).

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after drinking alcohol (Jones 2011a). One way to prevent drunken driving is to fit vehicles with an ignition interlock device, which would necessitate that a driver provides a negative breath-­alcohol test result before being able to start the vehicle (Fell et al. 2009). The factors that might contribute to a road-­traffic crash are summarised in Figure 59.1 including aspects related to the vehicle (brakes, speed, tyres and steering), the environment (weather, traffic density and road conditions) and the driver (age, distraction, medical and mental health, in-­vehicle distraction (cellular phone, text messaging, etc.), sleep loss and fatigue). One preventable cause of road-­traffic crashes is drinking alcohol or using other psychoactive drugs (Jones et  al.  2019). Investigations of drivers killed in road-­traffic crashes showed that between 20 and 50% of the victims had a BAC above the legal limit for driving (Legrand et al. 2014). The effects of alcohol on skills necessary for safe driving have been investigated extensively since the 1940s, both in laboratory experiments, which focus on recording behavioural changes and diminished cognitive and psychomotor performance (Drew et  al.  1958; Mitchell  1985) and actual on-­the-­road driving tests (Jongen et  al.  2017). The latter approach is more realistic and mimics real-­word conditions, such as using closed circuit roads or in specially designed dual-­control vehicles in the traffic flow (Laurell 1977; Kenntner-­Mabiala et al. 2015). By means of laboratory tests and controlled drinking studies, a dose–response relationship has been established between a person’s BAC and the impairment of the skills necessary for safe driving (Drew et al. 1959; Jongen et al. 2016). This type of research also verifies that a person’s BAC can be determined much more reliably than performance decrement an can the impairment of body functioning. The main reason for this is that people develop a tolerance to the effects of alcohol, and the signs and symptoms of drunkenness might not always be obvious even to trained observers

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(Brick and Erickson 2009; Olson et al. 2013). This supports the use of measuring ethanol concentrations in blood or other biological fluid as evidence for the prosecution of traffic offenders. Enforcement of drink-­driving laws would be much more difficult if the evidence was based only on the results from a clinical examination of suspects and signs and symptoms of drunkenness. Measuring the concentration of alcohol in samples of blood or breath furnishes a much more pragmatic way to enforce drink-­ driving legislation (Jones 2011a; DuPont et al. 2012). This chapter is an update of the information contained in the first edition of Handbook of Forensic Medicine that dealt with the effects of alcohol on fitness to drive. Over the past 10 years, not much has changed regarding the prevalence of traffic crimes and the demographics of offenders. Males still dominate among traffic offenders (80–90%) compared with females (10–20%). Their average age is between 35 and 45 years, and many had previous arrests for driving under the influence of alcohol (DUI). Furthermore, the average DUI offender had consumed massive amounts of alcohol, because their mean BAC when arrested was between 1.5 and 1.8 g/L.

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59.2.1  Relationship between blood alcohol and impairment The signs and symptoms of alcohol influence depend on many factors and there are large inter-­individual variations after the

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The impairment effects of ethanol are more pronounced on the rising limb close to the time of reaching the peak BAC. Several hours later, when the post-­absorptive phase is well established, the intoxicating effects of alcohol subside or disappear completely (Alha 1951). Highly sensitive tests are necessary to demonstrate impairment in the post-­absorptive declining phase of the BAC curve, which might still exceed the statutory limit for driving in some countries. Nevertheless, the risk of involvement in a road-­ traffic crash might still be appreciable, because after drinking alcohol people feel sleepy and are anxious owing to the after-­ effects of recent drinking. Alcohol has been called the Jekyll and Hyde of the drug world because moderate drinking is harmless, whereas over-­ consumption and abuse wrecks people’s lives, strains family relationships and damages body organs and tissues, especially the liver (Gibbons 1992). In a UK survey of the dangerousness of recreational drugs in society, (physical harm, social harm and risk of dependence) alcohol topped the list, even being ahead of heroin and crack cocaine (Nutt et al. 2010).

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59.3 Medicolegal alcohol determinations The concentration of ethanol determined in a person’s blood or breath is widely accepted as the most objective evidence and proof of over-­consumption of alcohol and unfitness to drive (Moriya  2005; Jones  2020b). The concentration of ethanol in a specimen of blood or exhaled breath can be determined with a high degree of accuracy, precision and specificity much more so than clinical signs and symptoms of drunkenness. Many clinical tests of impairment suffer from a lack of sensitivity and a lot depends on the skill and training of the person that administers them (Penttila and Tenhu 1976). The evidence that a person was in breach of the drink-­driving law depended on the concentration of ethanol determined in a sample of blood or urine taken from a suspect 1–2 hours after driving (Jones 2020a). However, in parallel with developments in electronics and microprocessor technology, instruments for breath-­alcohol analysis are increasingly being used for legal purposes. The sampling and analysis of ethanol in breath is less intrusive compared with sampling blood or urine. Two types of breath-­ alcohol analyser are currently available, (i) handheld instruments mainly intended as a roadside screening test for alcohol influences and (ii) evidential quality instruments located and used at police stations. The acceptance of evidential breath-­ alcohol instruments led to a change in the law and introduction of statutory limits of breath-­alcohol concentration (BrAC). These were derived from the existing statutory BAC limits and assuming a population average blood–breath ratio (BBR) of alcohol. However, different countries opted for different BBRs, ranging from 2000:1 to 2400:1 as shown in Table 59.2, which summarises the concentration units used to report BAC and BrAC for legal purposes.

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same dose and BAC reached (Paton  2005; Roberts and Dollard  2010). Drinking small amounts of alcohol makes people feel relaxed, they become more talkative and experience a mild euphoria (BAC < 0.5 g/L). As larger amounts are consumed and the BAC reaches 0.8–1.0 g/L, people tend to be more daring and have poor judgement and self-­ control. Consumption of alcohol to reach BAC exceeding 1.0 g/L makes people more inclined to take risks, which increases the probability of causing a crash. As drinking continues and BAC increases further, the impairment of sensory and motor functioning becomes more obvious. Among other things, there is marked deterioration in fine muscular control, as reflected in staggering gait and inability to walk without help. Important for traffic safety are ethanol’s effects on reaction time, tracking and divided attention tasks, the latter being particularly sensitive to elevated BAC (Liu and Ho 2010). Continuing to drink to reach even higher BAC (e.g. 1.5 g/L or more), causes confusion and many people are now emotionally unstable, and some are hostile and aggressive. Many medical books and review articles dealing with alcohol-­ use disorders and addiction provide information about the relationship between BAC and the signs and symptoms of drunkenness. The BAC is displayed in one column and alongside another column lists the typically expected impairment effects on the individual. A widely used compilation was constructed by Dr Kurt Dubowski (1921–2017), and his ‘Dubowski table’ has received a lot of attention in both addiction medicine and in law enforcement (Dubowski 2012). Table 59.1 shows measured BAC (left column) expressed as g% (g/100 mL), which is the concentration units used in USA, mg% (mg/100 mL) used in UK and g/L (mg/mL) used in many EU nations. The various stages of alcohol influence are designated, as are the expected clinical effects on the drinker. The signs and symptoms of alcohol influence are generally more pronounced on the rising phase of the BAC before reaching the maximum concentration in blood. Table  59.1 shows that there are six different stages of alcohol influence ranging from none observable (sobriety) to gross intoxication unconsciousness and death. The left column shows overlapping BAC ranges from 5.0 g/L. The right column lists the various clinical signs and symptoms that manifest when people consume alcohol to reach these BAC levels. The overlapping BAC ranges signifies the existence of large inter-­subject variations in the impairment effects of alcohol despite the same BAC. Furthermore, the information in Table  59.1 applies to social drinkers and might not necessarily be the same in heavy drinkers and alcoholics, who develop a tolerance to many impairing effects of ethanol. The information contained in Table 59.1 is intended to provide a general guideline whenever questions arise about interpreting BAC for legal purposes in relation to the degree of alcohol influence and expect clinical effects. Temporal variations also exist between impairment and BAC, depending on whether the tests or observations were made on the rising or declining limbs of the BAC curve (Schweizer et al. 2006).

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Table 59.1  Examples of six stages of alcohol influence in relation to blood-­alcohol concentration (BAC) and clinical signs and symptoms of  intoxication. Note that the BAC is reported here in three different concentration units (g/L, g% and mg%). Stage of alcohol influenceb

Clinical signs and/or symptoms associated with BAC rangesc

0.1–0.5 g/L 0.01–0.05 g% 10–50 mg%

Subclinical

Behaviour nearly normal by ordinary observation. Influence/effects usually not apparent or obvious. Impairment detectable by special tests.

0.3–1.2 g/L 0.03–0.12 g% 30–120 mg%

Euphoria

Mild euphoria, sociability, talkativeness. Increased self-­confidence, decreased inhibitions. Diminished attention, judgement and control. Some sensory-­motor impairment. Slowed information processing. Loss of efficiency in critical performance tests.

0.9–2.5 g/L 0.09–0.25 g% 90–250 mg%

Excitement

Emotional instability, loss of critical judgement. Impairment of perception, memory and comprehension. Decreased sensatory response, increased reaction time. Reduced visual acuity and peripheral vision and slower glare recovery. Sensory-­motor in-­coordination, impaired balance, slurred speech. Vomiting, drowsiness.

1.8–3.0 g/L 0.18–0.30 g% 180–300 mg%

Confusion

Disorientation, mental confusion, vertigo, dysphoria. Exaggerated emotional states (fear, rage, grief, etc.). Disturbances of vision (diplopia, etc.) and of perception of colour, form, motion, dimensions. Increased pain threshold. Increased muscular in-­coordination, staggering gait, ataxia. Memory loss. Apathy with progressive lethargy.

2.5–4.0 g/L 0.25–0.40 g% 250–400 mg%

Stupor

3.5–5.0 g/L 0.35–0.50 g% 350–500 mg%

Coma

>3.6 g/L (median) >0.36 g% (median) >360 mg% (median)

Death

General inertia, approaching loss of motor functions. Markedly decreased response to stimuli. Marked muscular in-­coordination, inability to stand or walk. Vomiting, incontinence of urine and faeces. Impaired consciousness, sleep or stupor, deep snoring. Complete unconsciousness, coma, anaesthesia. Depressed or abolished reflexes. Subnormal temperature. Impairment/irregularities of circulation and respiration. Grave risk of death. Death from respiratory failure and/or cardiac arrest.

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 Blood-­alcohol concentrations (BAC) for legal purposes are reported in different ways depending on the countries concerned, exemplified by g/L, g% and mg%.  Ethanol-­induced impairment is more pronounced on the rising part of the BAC curve close to Cmax compared with the same BAC on the declining phase of the curve after absorption and distribution are complete. This indicates the development of acute tolerance to ethanol’s effects. c  Large inter-­subject variations exist within the same BAC interval depending on age, previous drinking experience, speed of drinking and central nervous system tolerance. a

59.3.1  Blood-­alcohol analysis Methods for the analysis of ethanol in blood have been available for more than a century (Jones 2020a). A major advance occurred in 1922 when a micro-­diffusion method and chemical oxidation procedure was published (Widmark  1922), which because the gold standard in several countries. By the 1950s, a further analytical development saw the introduction of enzymatic oxidation,

which was more specific for ethanol analysis compared with chemical oxidation (Bonnichsen and Theorell  1951). In the 1960s, the method of gas-­liquid chromatography (GLC) was adopted in forensic laboratories, and this proved to be ideal for the determination of volatile substances in blood and other biological specimens (Curry et al. 1966). GLC methods later became the ‘gold standard’ for quantitative and qualitative analysis of ethanol in blood, and other volatile substances, such as acetone,

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Table 59.2  Examples of the concentration units used to report blood-­alcohol concentration (BAC) and breath-­alcohol concentration (BrAC) in different countries for legal purposes. Countries where these units are currently used

Concentration unit for reporting BrAC

Countries where these units are currently used

mg/g (g/kg)

Sweden, Denmark, Norway, Finland, Germany

mg/L

Austria, Sweden, Denmark, Norway, Finland, Germany, Spain and other European countries

mg/mL (g/L)

Austria, France, Holland, Spain, Belgium

μg/L

Holland

mg/100 mL (mg%)

UK, Ireland, Canada, New Zealand

μg/100 mL

UK, Ireland, New Zealand

g/100 mL (g%)

USA, Australia

g/210 L

USA

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Table 59.3  Chronological development in analytical methods used for qualitative and quantitative analyses of ethanol in blood for legal purposes. Method of analysis

Brief details of the analytical procedure

1920s

Chemical oxidation

Separation of ethanol from blood by diffusion or distillation followed by oxidation with an excess of potassium dichromate in sulphuric acid. The amount of chromic acid remaining is then determined by volumetric analysis and iodometric titration.

1950s

Enzymatic oxidation

Precipitation of blood proteins with perchloric acid, and the analysis of the supernatant after adding buffer to give pH of 9.4. Ethanol is oxidised with alcohol dehydrogenase (ADH) and the formation of reduced coenzyme NADH is monitored at a UV wavelength of 340 nm.

1960s

Gas chromatography (GC)

Dilution of an aliquot of the blood specimen (10 μL) 10 times with aqueous n-­propanol or t-­butanol as internal standard. Then, injecting 1 μL of the mixture into a GC instrument fitted with flame ionisation detector.

1970s

Headspace gas chromatography (HS-­GC)

1990s

HS-­GC with mass spectrometric detector

An aliquot of the blood specimen (10 μL) is diluted 10 times with aqueous n-­propanol or t-­butanol as internal standard. The mixture is allowed equilibrated at 50–60 °C in a glass vial made airtight with rubber septum and crimped-­on aluminium cap. After waiting 30 minutes to reach equilibrium, a sample of the headspace vapour is removed and injected into the GC instrument for analysis with flame ionisation detector. The GC analysis is accomplished as above but the effluent from the column is directed to an electron impact mass spectrometric detector. Ethanol shows prominent mass fragments at m/z 31 (base peak), m/z 46 (molecular ion) and m/z 45.

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ether, acetaldehyde and methanol could also be determined in the same specimen (Machata 1975). The positive identification of ethanol in blood was achieved by measuring its retention time (RT), that is, the time after injecting the sample until the peak response appears on the gas chromatogram. An aliquot of blood is first diluted with an internal standard (e.g. n-­propanol), and the relative retention times (RRTs) of these two substances are measured and used for identification. The amount of substance in the sample (quantitative analysis) was measured with a flame ionisation detector (FID) by integration of the area under the peak response on the chromatogram (Tagliaro et al. 1992; Seto 1994). Some forensic laboratories have now began using a mass s­ pectrometric (MS)

detector to identify ethanol in blood in drink-­driving casework (Tiscione et al. 2011). Table 59.3 presents a historical review of major developments of the methods used for determination of ethanol in blood for legal purposes (Jones 2020a). A robust and well-­proven gas chromatographic method of analysis entails diluting an aliquot of the blood specimen (100 μL) with 1  mL of an aqueous solution of an internal standard (n-­propanol or t-­butanol) and then injecting 1–2 μL of the mixture into the heated inlet port of the GLC instrument (Curry et al. 1966). The volatiles in the sample vapourise in the injection port, mix with the flowing carrier gas (N2 or He) and are transported through the GLC column where a partition between the

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As mentioned above, the method of HS-­GLC-­FID is the ‘gold standard’ for determination of ethanol in blood and other biological specimens for legal purposes worldwide. However, there is a trend towards obtaining higher analytical specificity by use of an MS detector in addition to the FID detector (Tiscione et al. 2011). With the MS detector, besides use of RT for the identification of ethanol, its mass fragments m/z 31 (base peak), m/z 46 (molecular ion) and a prominent ion at m/z 45 are also used. However, no agreement has been reached so far among forensic practitioners as to whether or not the more sophisticated technology of HS-­GLC-­MS is necessary when ethanol is determined in biological samples. This follows because the concentrations of ethanol in the blood of apprehended drivers are about 1000 times higher than most other psychoactive substances, being in the g/L range and not mg/L (Jones  2019b). Some laboratories prefer to determine ethanol in blood by two independent analytical methods, such as HS-­GLC-­FID and an enzymatic oxidation method (Krause  2007). This means that a potential interfering substance would need to have the same RT as ethanol by single-­column GLC analysis and also be oxidisable by alcohol dehydrogenase.

59.3.2  Breath-­alcohol analysis The first instruments designed for breath-­ alcohol analysis appeared in the USA in the 1940s, although these were rather primitive and were often challenged in court as being unreliable (Jones  2020a). The Breathalyzer instrument was developed in 1954, and by the 1960s was used in most states of the USA and in Canada for legal purposes (Borkenstein and Smith 1961). By the 1980s, the technology for breath-­alcohol analysis was greatly improved and this prompted EU nations to consider this analytical method both as a roadside test of driver sobriety (Zuba 2008) and also for evidential quality results and the prosecution of traffic offenders (Schechtman and Shinar 2011). The main advantage of breath analysis over blood analysis is the non-­invasive nature of the sampling, and the fact that the result of a test is obtained within a few minutes after sampling (Jones 2020b).

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moving gas phase and stationary liquid phase takes place. Volatile substances in the blood, such as ethanol and the internal standard, are separated from one another before they pass through the column and reach the detector for quantitative analysis. A modification of the GLC-­FID method of analysis entailed sampling the headspace vapour in equilibrium with the liquid blood sample. This had the advantage that the GLC column did not get contaminated with non-­volatile components of blood, such as protein and lipids. An aliquot of the blood sample (100 μL) is first diluted with 1 mL of internal standard in a glass vial, which is made airtight with a rubber stopper and a crimped-­on aluminium cap (Machata 1975; O’Neal et al. 1996). The vial is then heated to about 50 or 60 °C until an equilibrium is reached between volatiles in the blood and the air phase. Thereafter, a portion of the headspace vapour (1 mL) is removed with a gas-­tight syringe or some automated sampling system and injected into the GLC-­FID instrument for analysis (Seto 1994). The principles and practice of headspace GLC-­FID analysis are described in more detail elsewhere (Hachenberg and Schmidt 1977). All forensic blood sample should be analysed in duplicate, and the two determinations should be performed on two chromatographic systems that yield different RTs for ethanol. Finding ­co-­incident RTs on both GLC systems for ethanol and some unknown volatile substance that might have been present in blood is highly unlikely. Few chemical or toxicological substances can be determined with such a high accuracy, precision and specificity as blood-­ ethanol concentration. Table  59.4 shows the results of an inter-­ laboratory proficiency trial of blood-­ ethanol determination at specialist forensic laboratories in the Nordic countries. The blood specimens were taken from apprehended drivers in the usual way be venepuncture and split into 1 mL portions and shipped for analysis to the participating laboratories as a declared proficiency trial. The mean BAC reported by the various laboratories ranged from 0.47 to 2.88 g/L for the six specimens, and inter-­laboratory coefficients of variation (CV%) were only 2–3% indicating good interlaboratory precision. The precision within laboratories was even better (CV < 1%) as determined by replicate determinations on each blood sample (data not shown). The results of an inter-­laboratory proficiency trial of blood-­ethanol determinations at laboratories in Italy gave less acceptable results (Zamengo et al. 2019).

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Table 59.4  Results of an inter-­laboratory proficiency test of blood-­ethanol analysis at specialist forensic toxicology laboratories in the Nordic countries. Ethanol concentrations in blood are reported in per mille (g/kg); SD: standard deviation; CV: coefficient of variation. Blood sample

Denmark

Finland

Iceland

Norway

Sweden

Mean ± SD

CV%

1

0.49

0.46

0.47

0.48

0.47

0.474 ± 0.011

2.3

2

1.77

1.63

1.71

1.78

1.75

1.728 ± 0.061

3.5

3

2.69

2.58

2.68

2.76

2.70

2.682 ± 0.065

2.4

4

0.72

0.71

0.70

0.76

0.74

0.726 ± 0.024

3.3

5

1.12

1.09

1.10

1.13

1.12

1.112 ± 0.016

1.4

6

2.98

2.87

2.99

3.01

2.88

2.946 ± 0.066

2.2

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ples, such as infrared spectrometry at 9.5 μm and electrochemical oxidation for analysis of ethanol. This makes it virtually impossible that there could have been an interfering volatile substance emitted in human breath that caused a false positive analytical result (Caldwell and Kim 1997). Defence arguments maintaining that the result of an evidential breath-­alcohol test was caused by some volatile substance other than ethanol are simply not credible (Jones and Andersson 2008). Refusal to provide the required sample of breath or not being able to make a continuous exhalation for at least 6 seconds, because of some medical condition, means that a specimen of blood will be taken instead. The pros and cons of forensic blood-­ alcohol analysis as opposed to breath testing in law enforcement have been reviewed elsewhere (Jones  2000). Breath sampling is non-­intrusive which makes it more acceptable to the suspect, and whether the statutory limit is exceeded becomes known immediately, whereas the results of blood-­alcohol analysis take about one week to be finalised. However, compared with blood analysis, breath testing is more prone to physiological variations, depending on subject demographics, lung capacity and pattern of breathing prior to making a prolonged exhalation (Hlastala and Anderson 2016; Anderson and Hlastala 2019). Examples of some modern evidential breath-­alcohol instruments currently used in law enforcement are presented in Table 59.5. Experience has shown that many elderly female drivers, e.g. those with short stature and who are also smokers, have difficulty in providing two consecutive end-­exhaled breath samples, which is a minimum requirement in most countries for legal purposes (Jones and Andersson 1996). Whenever this happens, it is customary to require a sample of blood for the analysis at an accredited forensic laboratory. Because of analytical and biological variations and the existence of a razor-­sharp difference in penalty for those close to a statutory alcohol limit for driving, great care is needed when the analytical results are used in criminal prosecutions (Gullberg 2012; Vosk and Emery 2015). Uncertainty in the results can be allowed for by making a deduction from the mean BrAC (Searle 2015). After making this allowance, the final result used for prosecution can be stated as being not less than the true alcohol concentration with a high degree of statistical confidence, such as 99% or 99.9% (Jones  2003). Another approach is to adopt a safety margin or guard band, which means that if the statutory limit for driving is 35 μg/100  mL breath, a prosecution will not initiated until the lower of two tests is 40  μg/100  mL or higher, which gives an allowance of 14% and some benefit of doubt to a suspect in borderline cases (Gullberg 2020). If the observed impairment of a driver is caused by drugs other than alcohol, then it is necessary to obtained samples of blood for laboratory analysis (Jones et al. 2020). However, there is also on-­going research aimed at developing methods for analysis of non-­alcohol drugs in exhaled breath (Trefz et al. 2017). Measuring non-­alcohol drugs in expired air constitutes a novel approach for use in toxicology, and has already been used as proof of drug-­free status, but not yet in the enforcement of safety laws and impaired driving (Beck et  al.  2013; traffic-­

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Most of the instruments used as roadside screening tests of alcohol influence are handheld electronic devices that determine ethanol in the breath sample by electrochemical oxidation, which is a highly specific method of analysis (Zuba 2008). Some instruments report results simply as pass, warn or fail, whereas others give a digital display of the BrAC. In some of the clinical applications of breath-­alcohol analysis, the results are reported as the presumed BAC and this requires a pre-­calibration assuming a constant BBR of alcohol. Handheld breath-­alcohol analysers are an effective way to control sobriety of motorists and a driver can be tested sitting behind the steering wheel. Depending on the breath-­test results, the police then make a decision to arrest the suspect for further testing or allow the person to continue driving on the public roads. In conjunction with every evidential breath-­alcohol test, it is important to verify that the instrument was properly calibrated and thus giving accurate results. This is usually done by generating known strength concentrations of ethanol in air, such as by bath simulator device. Alternatively, known means of a wet-­ amounts of ethanol are available in pressurised cylinders mixed with argon or nitrogen gas (Dubowski and Essary  1996). Calibration control of the instrument at the time a suspect is tested represents one element of quality assurance, which is necessary when analytical results are used as evidence in criminal cases (Dubowski 1994; Gullberg 2020). Breath alcohol analysis should never be done within the first 15 minutes after the last drink, otherwise the result is likely to be falsely too high, owing to a mouth-­alcohol effect (Sterling 2012). Modern evidential breath-­alcohol instruments are fitted with so-­ called slope detectors that monitor a person’s BrAC during a prolonged exhalation. In this way, they can distinguish if the breath specimen is contaminated with excess alcohol from the oral cavity, such as after a recent drink or regurgitation of stomach contents. Depending on jurisdiction, each subject is expected to provide two breath samples at least 5 minutes but not more than 15  minutes apart, and the two results must agree within pre-­ defined tolerance limits for approval (Gullberg and Polissar 2011). On completion of the evidential testing, all analytical results are printed out in real time and stored on-­line for later retrieval and verification if this becomes necessary. The results of the two breath tests, the calibration control check and analysis of room air ‘blanks’ are all shown on a printed record made available as evidence. This print-­out should contain the name and date of birth of the suspect or other positive identification (e.g. passport or driving licence number) and then signed by the instrument operator to indicate that proper operating procedures were followed (Hodgson 2008). Evidential breath-­alcohol instruments that incorporate infrared absorption spectrometry determine ethanol from its molecular vibrations when exposed to wavelengths of 3.4 μm, which corresponds to C–H stretching, and 9.5 μm, which corresponds to C–O stretching vibrations. Some evidential instruments have multiple IR absorption wavelengths close to 3.4 μm for enhanced selectivity (Harding and Zettl  2008). A few evidential breath-­ alcohol analysers incorporate two independent analytical princi-

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Table 59.5  Examples of the development of instruments for the analysis of alcohol in breath for legal purposes, used both as a roadside screening test and for evidential purposes. The approximate time period when these instruments emerged and the operating principle for the determination of ethanol are shown. Instrument

Brief details of the analytical procedure

1950s

Breathalyzer

Chemical oxidation with potassium dichromate and sulphuric acid and end-­point determination by photometry.

1970s

GC Intoximeter Alco-­Analyzer

Small compact gas chromatographs designed to capture a sample of end-­exhaled breath. GC Intoximeter used a flame ionisation detector and AlcoAnalyzer used a thermal conductivity detector.

1970s

Intoxilyzer

Single wavelength (3.39 μm) infrared spectrometer measuring C–H stretching vibrations in ethanol molecule.

1975

Alcolmeter AlcoSensor

Small handheld instruments incorporating an electrochemical sensor for the oxidation of ethanol in a sample of end-­expired breath.

1980s

DataMaster Intoxilyzer 5000

Multiple infrared filters in the 3.3–3.5 μm range corresponding to C–H stretching in ethanol molecules allowed differentiating ethanol from acetone in breath.

1990s

Alcotest 7110 Intoximeter EC-­IR

Instruments that incorporate both an infrared and an electrochemical detector for the determination of ethanol.

2000s

DataMaster Intoxilyzer 8000

Instruments that incorporate infrared absorption at wavelengths of 3.4 μm (C–H stretch) and 9.5 μm (C–O) stretch in ethanol molecules.

2010s

Alcotest 9510 DataMaster DMT Evidenzer

More robust and portable instruments for conducting evidential testing at the roadside. Ethanol in breath is determined with multiple infrared filters or with a single filter (9.5 μm) in addition to an electrochemical detector.

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connected by the use of a population average BBR of alcohol, which varies between countries from 2000:1 to 2400:1. The BBR of alcohol depends to some extent on whether BAC was reported in mass/mass or mass/volume units. In Germany and

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Beck et al. 2019). The breath samples must be stored on special absorbent chemicals that are later shipped to a laboratory for analysis by LC-­MS methods (Ullah et al. 2018). Another problem with analysis of non-­alcohol drugs in breath is that there does not appear to be any association with the concentrations in samples of blood or serum, which limits practical usefulness of this methodology (Bakke et al. 2020). By contrast, many countries have enacted statutory concentration limits for selected psychoactive drugs in blood of drivers above which it is an offence to drive (Vindenes et al. 2012).

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59.4 Statutory alcohol limits for driving The enforcement of statutory BAC or BrAC limits for driving are well established in most countries worldwide (Voas and Fell 2011). However, these permissible alcohol limits vary fourfold, and China, Sweden, Norway and Poland enforce a limit of 0.2 g/L in blood or the equivalent concentration in breath. Most EU nations enforce a BAC limit of 0.5 g/L as does Scotland, whereas in England and Wales a limit of 0.8 g/L applies (Jones et al. 2019). In USA, the statutory limit for driving is 0.08 g% in blood or 0.08 g/210 L in breath, although the state of Utah lowered this to 0.05 g% (0.05 g/210 L breath) from January 2019, and there is strong support among traffic-­safety researchers that other states follow this lead (Fell and Scherer 2017). Table 59.6 compares statutory alcohol limits for driving in several European nations where both evidential blood-­and breath-­ alcohol testing is done. The alcohol limits in breath and blood are

Table 59.6  Statutory limits of blood-­alcohol concentration (BAC), breath-­alcohol concentration (BrAC) and the assumed blood–breath ratio (BBR) of ethanol in several European countries. Countrya

Statutory BAC

Statutory BrAC

BBR

Austria

0.50 g/L

0.25 mg/L

2000:1

Belgium

0.50 g/L

0.22 mg/L

2300:1

France

0.50 g/L

0.25 mg/L

2000:1

Greece

0.50 g/L

0.25 mg/L

2000:1

Italy

0.50 g/L

0.50 g/L

2000:1

Poland

0.20 g/L

0.10 mg/L

2000:1

Portugal

0.50 g/L

0.50 g/L

2300:1

Republic of Ireland

50 mg/100 mL

22 μg/100 mL

2300:1

Spain

0.50 g/L

0.25 mg/L

2000:1

The Netherlands

0.50 mg/mL

220 μg/L

2300:1

Scotland

50 mg/100 mL

22 μg/100 mL

2300:1

England and Wales

80 mg/100 mL

35 μg/100 mL

2300:1

a  In some countries, the BAC limits enforced are lower for learner drivers or those recently qualified (novice drivers), as well as for public transport and commercial vehicles.

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Table 59.7  Statutory limits of blood-­alcohol concentrations (BAC), breath-­alcohol concentration (BrAC) and blood–breath ratios (BBR) of alcohol adopted in Germany and Scandinavian countries where BAC is reported in mass/mass concentration units. Statutory BrAC (mg/L)

BBR

Denmark

0.50

0.25

2100:1

Finland

0.50

0.22

2400:1

Germany

0.50

0.25

2100:1

Norway

0.20

0.10

2100:1

Sweden

0.20

0.10

2100:1

Statutory BAC

Statutory BrAC

BBR

Australiaa

0.05 g/100 mL

0.05 g/100 mL

2100:1

Canadaa

80 mg/100 mL

80 mg/100 mL

2100:1

New Zealand

50 mg/100 mL

250 μg/L

2000:1

USAb

0.08 g/100 mL

0.08 g/210 L

2100:1

 In these countries, the result of the breath-­alcohol test is converted into the BAC using a population average BBR of 2100:1. b  Lower statutory limit (e.g. 0.02 g/100 mL) for people under 21 years. The BAC limit is also lower (e.g. 0.04 g/100 mL) for drivers of public transport and heavy commercial vehicles in some of the countries. a

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the Nordic countries BAC for legal purposes is reported in mass/ mass concentration units (mg/g or g/kg). Because the density of whole blood is 1.055 g/mL, a BAC of 1.0 g/kg is the same as 1.055 g/L. This 5.5% difference means that BBRs of alcohol are higher when mass/volume concentration units are used. Table 59.7 summarises the statutory alcohol limits for driving in various countries including the relationships between BAC and BrAC. The Breathalyzer instrument, which was invented in 1954, was widely used for legal purposes in Australia, Canada and the USA. This instrument incorporated a BBR of 2100:1 to convert the measured BrAC into the co-­ existing BAC (Borkenstein and Smith 1961). However, the results from many controlled drinking studies showed that the Breathalyzer instrument gave results that were systematically lower than the venous BAC by about 10% on average. A closer agreement was obtained using a 2300:1 BBR to calibrate the instrument. Indeed, a 2300:1 BBR was later adopted in some countries, such as Ireland, UK, Belgium and Netherlands, when their threshold BrAC limits were established. Research done in the 1970s showed the existence of large biological variations in the BBR of alcohol, and this became a major point of controversy when people were prosecuted. This led to a subsequent change in the law so that statutory alcohol limits for driving were defined as breath-­alcohol concentration (BrAC). This meant that in the USA a BAC of 0.08 g/100  mL was deemed equivalent to a BrAC of 0.08 g/210  L, thus assuming a BBR of 2100:1, which still exists today. Table  59.8 compares ­statutory

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Statutory BACa (g/kg)

Table 59.8  Statutory blood-­alcohol concentrations (BAC), breath-­ alcohol concentrations (BrAC) and the blood–breath ratio (BBR) of alcohol in the USA, Canada, Australia and New Zealand.

Drunken driving and road-­traffic crashes represent a major cause of morbidity and mortality worldwide. The age of offenders ranges from 14 to 90 years with a clear predominance of males in the age group of 35–45 years (Holubowycz et al. 1994). Descriptive statistics comparing age, gender and mean BAC for apprehended drivers in Sweden are shown in Table  59.9. The female traffic offenders were slightly older than the males by 2.8 years on average, and this difference was statistically significant and consistent over time. A relative frequency distribution of the age of male and female traffic offenders is shown in Figure 59.2 and it can be seen that the relative proportion of males was highest in the age group 15–30 years, whereas females where mostly aged 35–60 years when apprehended. These differences in propensity for drunken driving as a function of gender and age might reflect developments and changes in drinking habits (Holubowycz and McLean  1995). Young men seem more likely to engage in irre-

Table 59.9  Demographics of people arrested in Sweden for drunken driving in relation to their mean, median and highest blood-­alcohol concentration (BAC). Subjects

N (%)

Age year ± SD) (range of values)

Mean BAC ± SD) (g/L)

Median BAC (g/L)

Highest BAC (g/L)

Males

29 370a (89.5%)

39.0 ± 14.6b (14–90 yr)

1.73 ± 0.85c

1.77

5.02

Females

3444 (10.5%)

41.8 ± 13.6 (14–82 yr)

1.78 ± 0.87

1.83

5.18

Both gender

32 814 (100%)

39.3 ± 14.5 (14–90 yr)

1.74 ± 0.85

1.78

5.18

 Significantly higher proportion of males to females (p < 0.001).  Average age of men is less than women (p < 0.001). c  Mean blood-­ethanol concentration in men is lower than women (p < 0.01). a

b

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20 Women (N= 3444) mean age 42 yr 15

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*

*

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Men (N= 29370), mean age 39 yr

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Women (N= 3444) median blood ethanol 1.77 g/L

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sponsible drinking, they show poor judgement and self-­control resulting in them being arrested by the police for drunken driving. By contrast, women become more of a problem when they reach middle age, perhaps coinciding with other problems in life and heavy drinking in general (Skurtveit et al. 1995). Historically, over-­consumption of alcohol and drunken driving has been a male-­ dominated form of deviant behaviour. However, the number of females has increased appreciably over the past decades from being 1.6% of the total in 1967 to 6.2% in 1986, 10% in 2007 and 12% in 2011 based on studies in Sweden (Jones et al. 1989). In Norway, 7.6% of all arrested drivers were women in a study from 1992 to 1993 (Skurtveit et al. 1995). More and more women obtain a driving licence earlier in life and there is a definite change in drinking habits among women. This includes frequency of intake and total consumption, which probably explains the increasing prevalence of female traffic offenders in most nations (Romano et al. 2012).

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Figure 59.2  Relative frequency distribution of the age of people apprehended in Sweden for drunken driving showing a higher proportion of males to females at certain age groups as indicated with * sign.

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Blood Ethanol, g/L Figure 59.3  Relative frequency distribution of blood-­alcohol concentrations (BAC) in apprehended drivers in Sweden with no statistically significant differences between male and female traffic offenders. Source: Modified from Jones and Holmgren (2009).

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Most people arrested for drunken driving are hardly moderate drinkers, and their BAC ranges from below the statutory limits (0.2–0.8 g/L) to concentrations that are considered sufficient to cause death by acute alcohol poisoning >4 g/L (Jones  1999; Kriikku and Ojanpera  2020). Figure  59.3 shows a relative frequency distribution of the BAC determined in drunken drivers apprehended in Sweden for male and female offenders plotted separately (Jones and Holmgren 2009). The mean and distribution of BAC in these traffic offenders were remarkably similar for males and females, which suggests similar patterns of drinking in this population of traffic offenders. The results showed that 38% of men and 40% of women had a BAC exceeding 2.0 g/L, which is a strong indicator of binge drinking and most likely an alcohol-­use disorder (Schuckit 2009). In these high-­risk offenders (BAC > 2 g/L), the notion of some

type of treatment and rehabilitation for alcohol problems might be more worthwhile than conventional punishment, fines or imprisonment, for this traffic offence. The average BAC of 1.74 g/L (median 1.78 g/L) and many individuals exceeding 3.0 g/L, strongly suggests an alcohol-­abuse problem. The BAC can be translated into the amount of alcohol absorbed and distributed in all body fluids at the time of sampling blood using the well-­known Widmark equation (Widmark 1932,  1981). Accordingly, a male offender weighing 75  kg and with a BAC of 1.74 g/L assuming a distribution factor for ethanol of 0.70 will have 91 g of pure ethanol absorbed and distributed in all body fluids and tissues. This corresponds to consumption of at least 11 standard drinks or units of alcohol (one unit = 8 g). The total amount of alcohol consumed was even greater because of the amount eliminated by metabolism from the time of starting

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Table 59.10  Demographics of people arrested in Sweden for drunken driving between 2003 and 2007. Note that the mean age of offenders (both males and females) as well as their mean blood-­alcohol concentrations (BAC) remained remarkably constant over the 5-­year study period. Year

N a

Mean age ± SD (yr)

Mean BAC ± SD (g/L)

2003

4381

39 ± 14.4

2004

4084

2005

N (%)

Mean age ± SD (yr)

Mean BAC ± SD (g/L)

1.75± 0.84

Male Female

3950 (90.2%) 431 (9.8%)

39 ± 14.5 42 ± 13.5

1.75 ± 0.84 1.75 ± 0.85

39 ± 14.4

1.73± 0.83

Male Female

3663 (89.7%) 421 (10.3%)

39 ± 14.5 41 ± 13.5

1.73 ± 0.93 1.74 ± 0.83

3873

40 ± 14.7

1.75± 0.83

Male Female

3466 (89.5%) 407 (10.5%)

39 ± 14.8 43 ± 13.7

1.74 ± 0.83 1.81 ± 0.83

2006

3870

40 ± 14.9

1.73± 0.84

Male Female

3439 (88.9%) 431 (11.1%)

39 ± 14.9 42 ± 14.8

1.72 ± 0.83 1.82 ± 0.86

2007

3814

39 ± 14.9

1.72± 0.83

Male Female

3412 (89.5%) 402 (10.5%)

39 ± 14.7 41 ± 15.2

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 Apprehended drivers providing an evidential breath-­alcohol test are not included.

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Table 59.11  Demographics of people arrested once or several times for drunken driving in Sweden. Males dominated among first offenders and recidivists. Gender

N (%)

Age ± SD (yr)

Mean BAC ± SD (g/L)

One

Male Female

12 555 (89)b 1560 (11)

38.8 ± 14.9c 41.5 ± 13.5

1.69 ± 0.78 1.73 ± 0.79

Two or morea

Male Female

2672 (93)b 188 (7)

39.8 ± 13.1c 43.4 ± 10.2

1.85 ± 0.82d 1.69 ± 0.82

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to drink until the time of obtaining a blood sample (Gullberg and Jones 1994). The average rate of ethanol metabolism is 7–8 g (~1 unit) per hour.

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1.71 ± 0.82 1.80 ± 0.86

Arrests

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Repeat offending is a major problem for traffic-­safety organisations and government authorities (Cavaiola et al. 2007). A typical sanction for a drunken driving conviction is revocation of the driving permit for 12–24 months, although this does not seem to deter hard-­core offenders, many of whom still continue to drive even without a valid permit (Nochajski and Stasiewicz 2006). Re-­arrest rates differ between countries, and about 30% of people were re-­ arrested one or more times over a 7-­ year period (Christophersen et al. 2002). Table  59.11 compares the demographics of first-­time and repeat offenders in Sweden within 4 years of the first offence. Males were re-­arrested more frequently than females (93% vs. 7%) when compared with first-­time offenders (89% vs. 11%). Moreover, mean BAC was higher in male repeat offenders (1.86 g/L) compared with first-­time offenders (1.69 g/L). The average number of re-­arrests was 2.5 for men compared with 2.3 for women.

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The average age, the predominance of males to females and the consistency in the average BAC can be gleaned from Table 59.10, which shows these statistics between 2003 and 2007. These data make it perfectly clear that drunken drivers in Sweden are predominantly males (85–90%) with an average age of 39 years. The women are 2–3 years older on average, but they drink similar amounts of alcohol, because the mean and median BAC were about the same or slightly higher in females.

59.5.5  Amounts of alcohol consumed After absorption of alcohol from the stomach and intestines into the portal venous blood, the ethanol molecules are distributed throughout the total body water compartment, and there is no evidence of any binding to plasma proteins and solubility in fat and bone is negligible compared with water solubility (Kalant  1996; Jones 2019a). The forensic pharmacokinetics of ethanol in humans has been studied extensively since the 1930s (Widmark  1932). Such pharmacokinetic calculations are performed to verify the amounts of ethanol consumed and to back-­calculate the BAC at the time of sampling to the BAC at an earlier time, such as the time of driving (Jones 2011b). These calculations are mostly done using the Widmark equation below: A grams

BAC g / L

body weight kg

rho L / kg

TRAFFIC MEDICINE

59.6.1  Roadside surveys

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The most convincing evidence of a quantitative relationship between a driver’s BAC and the probability of involved in a crash comes from the results of roadside surveys, the most famous of which was done in the city of Grand Rapids, USA, in the 1960s (Borkenstein et al. 1974). This involved measuring the BrAC of drivers actually involved in traffic crashes (test group) and comparing this with the BrAC of a matched group of drivers not involved in crashes (control group). The accident and control groups of drivers were matched as closely as possible for time of day, day of week, direction of traffic flow and location of the crash. From the information collected, likelihood or odds ratios were calculated for a driver being involved in a crash in relation to his or her BrAC. The results of such studies are usually depicted as a relative risk curve as shown in Figure 59.4. The plot in Figure  59.4 presents the results from two such roadside surveys, one done in Florida (Fort Lauderdale) and the other in California (Long Beach) (Blomberg et al. 2009). The risk of a crash definitely begins to increase as the driver’s BAC passes 0.5 g/L, and the dotted lines in Figure 59.4 shows hypothetically what might be expected for people with different tolerance to ethanol’s impairment effects. The results shown in Figure 59.4 confirm and extend the findings from the original Grand Rapids relative risk study done in the 1960s (Borkenstein et al. 1974). Moreover, some of the deficiencies in the original study were corrected, such as by the use of more modern breath-­alcohol instruments to determine BrAC and also active pursuit of hit-­and-­run drivers, many of whom had

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59.6  Alcohol ingestion and crash risk

electronic devices, videos and DVDs, also increase the propensity for involvement in a crash.

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where A (g) is the amount of ethanol absorbed and distributed in all body fluids and tissues, BAC is the person’s blood-­alcohol concentration (g/L) and rho (L/kg) is a physiological factor characteristic for that individual. The rho-­factor shown in the equation is now more commonly referred to as the volume of distribution (Vd) of ethanol, which averages 0.70 L/kg for men and 0.60 L/kg for women, although the inter-­individual variability is about ±20%. This stems from biological factors, such as body composition, especially the proportion of fat to lead body mass (Watson et al. 1981; Maskell et al. 2020). Ethanol distributes into the water compartment of the body, and total body water in per cent of body weight is lower in women (~50%) compared with men (~60%) and also lower in obesity (body mass index (BMI) > 30) compared with lean individuals (BMI of 20–25) (Forrest  1986; Jones 2019a). In clinical medicine, risky drinking is often classified as the number of units of alcohol consumed per day or per week with one unit corresponding to 8 g ethanol in UK (Miller et al. 1991). Eight grams of ethanol is the amount contained in ½ pint (275 mL) medium strength beer (4% v/v), a small glass of table wine (100  mL) or one measure (25  mL) of spirits (40% v/v). Although opinions differ regarding what constitutes risky drinking, the consumption of three units of alcohol per day for a man and two units per day for a woman and elderly (>60 years) is generally not considered an immediate danger to health and longevity (Dawson 2011).

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Normal tolerance High alcohol tolerance Low alcohol tolerance

18 Relative Risk of a Crash

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The experimental evidence supporting the role of alcohol as a causative factor in traffic crashes is extensive and derives from various sources, including results of controlled laboratory studies aimed at measuring diminished performance in relation to BAC (Tiplady et al. 2004), impairment of cognitive and psychomotor functions, use of driving simulators and perhaps most importantly the results of case-­controlled roadside surveys (Blomberg et al. 2009). The skills necessary for safe driving include good visual search and recognition, vigilance, rapid information processing and decision-­making, often in choice situations, as well as adequate sensory and motor control necessary to perform the driving task (Ferrara et  al.  1994). A host of laboratory studies have investigated ethanol’s effects on eye movements, glare recovery, visual perception, auditory stimuli, reaction time in choice situations, speed in processing information and psychomotor functions. Divided attention tasks, such as when two or more sources of visual information must be processed simultaneously, are particularly sensitive to the effects of ethanol. Using a cellular phone when driving or reading or sending text messages has become a major traffic-­safety hazard because the person has to concentrate on several things at the same time (Strayer et al. 2006). Indeed, some nations have introduced legislation making it an offence to use a mobile phone or send text messages when driving. Other in-­vehicle distraction, such as

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1.6

Blood Alcohol Concentration, g/L Figure 59.4  Relationship between the relative risk of involvement in a road-­traffic crash and the driver’s blood-­alcohol concentration. The dashed lines are hypothetical and represent what might be expectations for drivers with different tolerance to alcohol’s impairment effects.

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Many types of skills are necessary for safe driving, especially sensory and motor functions, memory and attention to visual and auditory cues (Ferrara et  al.  1994). Although the results from controlled laboratory drinking studies are unlike real-­ world conditions, they nevertheless provide a useful starting point for the planning and execution of field studies (Drew et al. 1959; Grant et al. 2000). The various tests of performance decrement differ widely in terms of their sensitivity to alcohol, and the results from hundreds of laboratory studies have been extensively reviewed (Gjerde et  al.  2019; Moskowitz and Fiorentino 2000). A common experimental protocol is to administer a battery of sensory and/or psychomotor tasks and determine the person’s subjective feelings of intoxication and changes in mood after they receive known amounts of alcohol (Goldberg  1943). The various tests are done before drinking any alcohol, thus providing

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59.6.2 Laboratory drinking studies

baseline scores, and at repeated intervals for several hours after the end of drinking (Jones and Neri 1994). Examples of such laboratory tests include measuring standing steadiness (with open and closed eyes), hand-­steadiness, reaction time (simple and choice situations) and tracking performance (Jongen et al. 2016). The test results are then plotted as a function of time after drinking in relation to the prevailing BAC or BrAC, which serves as the independent variable. It is common practice to express the results as deviation from baseline scores, so both positive and negative deviations are possible. The latter indicates an improvement in test performance. The experiments should also be repeated after the same subjects receive a placebo drink instead of alcohol, and this ‘dry-­run’ can help to disclose whether learning effects need to be considered (Brumback et al. 2007). Vision and hearing are fairly resistant to the effects of alcohol and impairment of these functions first becomes obvious at fairly high BAC (>0.8 g/L). The person’s ability to process information deteriorates more rapidly, especially when different sources of information must be considered simultaneously. Complexity of the performance test is a critical element when attempting to measure the effects of alcohol on skills necessary for safe driving (Martin et al. 2013; Jongen et al. 2016). Table 59.12 gives examples of various methods used over the years to determine the effects of alcohol on a person’s performance and behaviour in relation to skills necessary for safe driving. When results of performance tests are interpreted, it is essential to know the concentration of alcohol in a sample of the ­person’s blood or breath at the same time the test was done. This

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been drinking alcohol (Blomberg et al. 2009). Neither the Grand Rapids study (Borkenstein et al. 1974) nor the more recent roadside survey (Bloomberg et  al. 2009) attempted to differentiate between drivers on the rising or declining limbs of the BAC curve. This information might have been gleaned from questioning the driver about their prior drinking pattern, amounts of alcohol consumed and the time of the last drink before involvement in a crash.

Types of evaluation made

Quantitative response factors studied

N

Perception and information processing

C

R

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Cognition

O

Ocular functions (vision) Psychomotor functioning

TR IB

Table 59.12  Examples of cognitive and psychomotor tests used to investigate the effects of alcohol on skills necessary for safe driving and on actual driving performance and the risk of being involved in a crash.

Visual search, auditory signal detection, pattern recognition, identification of traffic hazards, critical flicker fusion. Glare recovery, contrast sensitivity, visual acuity, depth perception, gaze nystagmus, positional nystagmus, roving ocular movements. Reaction time (simple and complex), finger tapping, body sway and balance disturbances, hand tremor, tracking tasks. Vigilance, divided attention, response latency, grammatical reasoning, mathematical processing, memory tests, digital symbol substitution tasks.

Laboratory studies of post-­drinking hangover effects

Sleepiness, anxiety and inability to concentrate, reduced working capacity and crash risk.

Driving performance

Computer-­aided tasks mimicking driving and the use of actual driving simulators of various dignity.

On-­the-­road driving

Closed-­track driving tasks, control of speed and emergency braking etc as well as driving on the highway with a dual-­controlled vehicle and monitoring distance from other traffic, weaving, lateral positioning, maintaining optimal speed, and over-­taking.

Roadside surveys and case-­controlled studies

The risk or probability of involvement in a traffic crash is determined by comparing BrAC of drivers actually involved in crashes with a matched control group of non-­crash drivers.

Autopsy studies of fatally injured drivers

Measuring the blood-­ethanol concentration of drivers killed in traffic crashes, determining their culpability for the crash and whether there are differences between single-­and multiple-­vehicle crashes.

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Clinical tests done before and after alcohol ingestion

6

• • • • • •

4

•

2

• • • •

0.75 g/kg N = 31 1.00 g/kg N = 30 1.25 g/kg N = 29

10 8

0

0

60

120

180

240

300

360

Pulse frequency Flushed face Walking (with open eyes) Walking (with closed eyes) Gait in turning Standing steadiness (open eyes) Standing steadiness (closed eyes) Finger-to-finger test Match test Speech and behaviour Errors in counting backwards

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Alcohol Influence (mean score)

12

Time After Start of Drinking, min

O

N

Figure 59.5  The time course of the ‘alcohol influence score’ according to the results of a clinical examination of heathy men after they drank a bolus dose of ethanol (0.75 g/kg, 1.0 g/kg or 1.25 g/kg) on an empty stomach. The various tests of alcohol influence (see bullet points) were administered at different times post-­dosing.

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their BAC at autopsy was above the legal limit for driving (Legrand et al. 2014; Ponce et al. 2019). The percentages of fatalities with elevated BAC was greater for single-­vehicle crashes, which strongly implicates the role of alcohol consumption and culpability on the part of the driver (Jones et al. 2009). International comparisons of driver killed in crashes are not easy to interpret because there is a fourfold difference in the statutory BAC limits for driving in different nations (Jones et al. 2019; Valen et al. 2019). This means that the proportion of victims considered above the legal limit is higher in those nations with the lowest statutory BAC limits (Lerner 2012). In Norway and Sweden, for example, a fatality injured driver with a BAC >0.2 g/L would be classified as exceeding the drink-­driving limit, whereas in USA and Canada a BAC of 0.8 g/L is needed to draw the same conclusion. When investigating driver fatalities, an autopsy BAC >0.1 g/L is mostly considered an alcohol-­positive case, although care is needed because under some circumstances alcohol can be generated in the body after death (Kugelberg and Jones  2007). This seems to be the case in traumatic deaths with open blood vessels, which tends to facilitate the synthesis of endogenous alcohol by the action of bacteria and yeasts (Jones 2016). Another problem with the statistics concerning alcohol-­related crashes is that not all driver fatalities undergo a forensic autopsy including comprehensive toxicological analysis. Much depends on survival time after the crash, the part of the country where this occurs and emergency hospital treatment of survivors. In a study from Norway, it was reported that information about alcohol and/or drug use by killed drivers was only available in 59% of fatalities (Gjerde et al. 2011). Figure 59.6 is a bar graph showing the percentages of drivers killed in crashes with BAC above the statutory limit for driving in Sweden of (0.2 g/L). Also shown on the graph is the average BAC in these victims, which was eight times (1.6 g/L) above the legal limit verifying binge drinking before the crash occurred (Jones et al. 2009). Over the 8 years’ of the study, about 22% of drivers killed were above the prescribed limit for driving, and their

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follows because of large inter-­individual variation in pharmacokinetics of ethanol, even when the same dose is consumed and when this is adjusted for gender and body weight. Moreover, the results of impairment tests need to be evaluated on the ascending and descending limbs of the BAC curve (see later under tolerance), which necessitates repetitive sampling of blood or breath during absorption, distribution and elimination processes (Martin and Earleywine 1990; Martin and Moss 1993). Experiments done by researchers in the Netherlands involved the use of dual-­controlled vehicles and the test subjects were allowed to drive on the highway after drinking known amounts of alcohol (Jongen et al. 2014). The most sensitive outcome measure was the standard deviation of lateral positioning of the vehicle after driving a given distance on the highway and maintaining a constant speed (Jongen et al. 2017). Examples of the time course of ethanol’s effects on cognitive and psychomotor functions are shown in Figure  59.5 in relation to the dose of ethanol administered (Alha 1951). The test subjects were healthy men, who consumed three different doses of alcohol on an empty stomach in 5 minutes. Thereafter, each subject was examined by a physician who recorded the results of clinical tests listed on the right side of the graph. The overall test results were then combined and plotted in relation to BAC as a function of time after drinking ended. The peak impairment score tended to occur before reaching the peak BAC, decreasing afterwards to reach baseline values (pre-­drinking) before BAC returned to zero. In fact, the mean impairment score in these clinical tests was not significantly different from pre-­drinking values although BAC was still above 0.5–0.8 g/L in some subjects.

59.6.3  Driver fatalities Statistical surveys of drivers killed on the roads verify that between 20–50% of victims had consumed alcohol before the crash, and

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1.0

20

0.5

0

2000 2001 2002 2003 2004 2005 2006 2007

0.0

Year Figure 59.6  Percentages of drivers killed in motor vehicle crashes in Sweden 2000–2007 with an autopsy blood-­alcohol concentration (BAC) above the statutory limit for driving (0.20 g/L). The mean BAC (± standard error) in each year is plotted above the bars for the 8-­year period (2000–2007).

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The development of tolerance to the effect of drugs is an important concept in pharmacology and is often depicted by a shift in the sigmoid dose–response curve to the right, which indicates a diminished response and higher doses are therefore needed to achieve the same effect as before (Roberts and Dollard 2010). It is common knowledge that people differ in their response to a given dose of alcohol, which is interpreted to mean that some individuals are more resistant to ethanol’s effects and signs and symptoms of drunkenness might not be evident (Brick and Erickson 2009). Some people need to drink more alcohol and reach higher BAC than others to have the same impairment effects on performance and behavior. This is one indication of the development of central nervous tolerance. Essentially, three types of tolerance to ethanol are recognised by pharmacologists: (i) dispositional or metabolic tolerance, (ii) acute or functional tolerance, which develops during a single exposure and (iii) chronic or cellular tolerance, which develops after a period of continuous heavy drinking lasting weeks or months (Kalant 1998). Dispositional tolerance is related to altered absorption, distribution, metabolism or elimination processes after a period of heavy drinking. Metabolic tolerance is reflected in a more rapid rate of ethanol elimination from the body, which is attributed to

5 Symptoms

Blood alcohol conc., g/L

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1.5 Alcohol Dose 0.75 g/kg mean curves, N = 31 subjects

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1.0 3

BAC curve

2 0.5 1 0.0

0

50

100

150

200

250

300

Clinical signs and symptoms

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59.7  Alcohol tolerance

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average BAC was consistently high (mean 1.6 g/L). These statistics are robust because about 95–97% of all drivers killed on the roads in Sweden underwent a forensic autopsy and a toxicological analysis was included. Research done in USA shows that young women are increasingly being involved in alcohol-­related crashes (Voas et al. 2012).

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40

N

1.5

O

60

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2.0

enzyme induction, whereby the microsomal enzyme (CYP2E1) increases its capacity to oxidise ethanol (Lieber 1997; Zanger and Schwab 2013). Faster elimination rates from blood were observed in some, but not all, alcoholics during detoxification for whom the average rate was 0.22 g/L/h compared with 0.15 g/L/h in moderate drinkers (Jones 2010). The development of acute tolerance occurs during a single exposure to ethanol and this phenomenon was first reported by the British pharmacologist Sir Edward Mellanby (1884–1955), based on experiments in dogs (Mellanby  1920; Holland and Ferner 2017). Acute tolerance to ethanol’s impairment effects is a robust observation and has been verified by different investigators in both humans and animals (Ginsburg et al. 2008). Subjective feelings of intoxication are more pronounced on the ascending limb of the BAC curve compared with the same BAC existing on the descending limb after passing the peak concentration (Martin and Moss 1993). Furthermore, decrement in cognitive and psychomotor functioning is also more pronounced on the ascending limb of the BAC curve compared with the post-­absorptive phase (Schweizer et al. 2006). Figure  59.7 shows the results of controlled drinking experiments in healthy men (N = 31), who ingested 0.75 g/kg body weight in the morning on an empty stomach. The battery of tests described in Figure 59.5 was administered before drinking and then at various times after drinking. The dose of ethanol was consumed in 5 min and tests were done on the rising and declining portions of the BAC curve (Alha  1951). By 120  minutes post-­ dosing, the clinical symptoms had more or less disappeared and were indistinguishable from pre-­drinking scores, despite the fact that BAC still remained elevated. However, this should not be interpreted to mean that these individuals were safe drivers, because many of the test subjects felt tired several hours after drinking alcohol, and fatigue is a well-known risk factor for traffic safety.

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Mean ± SE autopsy BAC g/L

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80

2.5

Blood alcohol conc., g/L

Per cent above BAC limit

100

0

Time After Start of Drinking, min Figure 59.7  Temporal variations in the symptoms of alcohol intoxication and the time course of blood-­alcohol concentration (BAC) after healthy volunteers drank 0.75 g/kg as a bolus dose on an empty stomach (mean curves for N = 31 subjects are plotted).

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Table 59.13  Relationship between clinical signs and symptoms of alcohol influence in relation to blood-­alcohol concentration (BAC). Apprehended drivers (N = 244) were examined by the same physician, who had to judge whether they were under the influence of alcohol. The degree of influence was graded as none, slight, moderate or severe. Judgement of alcohol influence None

Slight

Moderate

Severe

Total (%)

0.0–0.49

3

26

0

0

29 (12)

0.5–0.99

2

37

0

0

39 (16)

1.0–1.49

2

41

10

1

54 (22)

1.5–1.99

1

24

17

5

47 (19)

2.0–2.49

0

20

22

5

47 (19)

2.5–2.99

0

7

11

7

3.0–3.49

0

1

1

1

Totals

8 (3)

156 (64)

61 (25)

N

LY

BAC (g/L)

3 (1) 244 (100)

O

19 (8)

25 (10)

Source: Jones (2011a).

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Tenhu 1976). The police surgeon who administered the various clinical tests was not aware of the person’s BAC but knew they were suspected of drunken driving. After considering all the findings, the physician concluded whether the suspect was or was not impaired by alcohol and whether the level of impairment was slight, moderate or severe. Thereafter, a sample of blood was taken for the determination of ethanol. Table  59.13 shows considerable variation within the same BAC interval as to being judged as mild, moderate or severe intoxication. The results would probably have been worse if different physicians had examined the 244 suspects. Other reasons for the poor correlation between BAC and signs and symptoms of drunkenness are acute and chronic tolerance to ethanol’s effects and the fact that some people can pull themselves together in critical situations (Cherpitel et al. 2005).

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All-­in-­all, the results from laboratory studies suggest that risk of involvement in a traffic crash is greatest when the BAC curve is in the rising phase and close to the peak (Ginsburg et al. 2008). On reaching the declining limb of the BAC curve, the brain seemingly adapts to many of the behavioral and impairment effects of ethanol. A topic that has received rather little attention in traffic medicine is the after-­effects of an evening’s heavy drinking and performance decrement during a hangover (Swift and Davidson 1998). During a hangover, many people feel anxious, nervous, often suffer from nausea and many are sleepy, all of which have negative effects on performance of skilled tasks necessary for safe driving (Alford et al. 2020). Chronic tolerance develops more gradually and is reflected in a diminished response to the pharmacological effects of ethanol after multiple exposures over several weeks or months (Bennett et  al.  1993). Both acute and chronic tolerance coexist to some extent, which probably explains why many drunken drivers, when they are examined by a police surgeon, are judged as not being under the influence of alcohol despite them having an appreciable BAC. Apart from the smell of alcohol on breath, even trained physicians have difficulty in interpreting the results from a clinical examination of arrested drivers using simple cognitive and psychomotor tests of impairment as shown by the data in Table 59.13 (Jones 2011a). Table  59.13 presents results from a medical examination of 244 drivers arrested in Sweden who were suspected of being drunk at the wheel. Each person was examined by the same physician about 1–2 hours after making an arrest. The examination followed a standard protocol and consisted of asking various questions about recent consumption of alcohol and/or use of medication or recreational drugs. The examining physician made notes about the person’s general appearance, pulse rate, orientation in time and space, ataxia (Romberg’s test), speech, finger and hand movements, reactions of pupils to light as well as various cognitive and psychomotor tests (Penttila and

59.8  Concluding remarks Alcohol is a legal drug, and light-­to-­moderate drinking is a normal part of social life in most nations. The problem is that between 10 and 15% of those who begin as social drinkers, especially among men, increase their consumption pattern and they become regular heavy drinkers (Carvalho et al. 2019). In effect, they develop an alcohol-­use disorder and might be diagnosed as being alcohol dependent. Among the wide spectrum of pharmacological and behavioural effects of ethanol, people feel less inhibited and they experience a mild euphoria as BAC reaches 0.5 g/L. As the BAC increases further, many people become self-­ confident and more daring and they are also likely to take risks, including engaging in unsafe sex and drunken driving (Lane et al. 2004; Davis et al. 2009). Several studies of people arrested for drunken driving in Sweden show that this form of deviant behaviour is tightly linked to psychiatric co-­morbidity and criminality (Hubicka et al. 2007;

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Lowering the statutory blood-­alcohol limit from 0.8 to 0.5 g/L or even 0.2 g/L is not likely to deter hard-­core offenders because recidivism rates are high in these traffic delinquents (Schell et al. 2006; Williams et al. 2007). However, enforcing lower statutory alcohol limits for driving for newly qualified drivers shows promise in reducing the number of crashes caused by these individuals (Blackman et al. 2001). In the USA, the legal drinking age is 21 years, although a driving permit can be obtained at age 17 years, which speaks towards enforcing lower blood or breath-­ alcohol limits for novice drivers (0.2 g/L). Other traffic-­safety innovation for newly qualified drivers includes graduated licencing, with phased-­in privileges, such as carrying passengers in the vehicle, night-­time curfews and restrictions on the use of certain roads or highways (McKnight and Peck 2003). All these measures have proven effective in reducing fatalities in younger age groups of drivers (Fell, 2020). Laws that allow the traffic police to conduct random breath-­ alcohol testing of drivers have proven a very effective way to lower the incidence of drunken driving by acting as a deterrent (Homel  1983; Ferris et  al.  2013). Random breath testing is a needed and effective way to improve traffic safety, and represents only a minor intrusion of privacy considering the potential benefit this has on preventing alcohol-­impaired driving fatalities. A police officer in uniform should therefore have the power to stop a vehicle at random and perform a sobriety check on the driver with a handheld breath-­alcohol analyser. Such testing can be performed with the driver sitting behind the wheel so there is little inconvenience or time delay provided the result is negative. This seems a small price to pay for increasing the effectiveness of the police in detecting drunk driving offenders and improving traffic safety. The large scale use of random roadside breath-­alcohol testing makes motorists aware that they risk being stopped and breath tested without any suspicion of them having consumed alcohol, which should increase deterrence (Peek-­Asa  1999). In the USA, some state legislation permits targeted sobriety checkpoints on specific weekdays and times of day, such as weekend nights, when the prevalence of impaired driving is highest and this approach has proven effective in reducing road-­traffic deaths (Fell et al. 2004). Laws that permit impounding the vehicle owned by repeat offenders or removing the registration plates are also in existence in some nations. These hard-­core offenders don’t seem to care about driving when drunk, because many of them are criminal elements in society and have been convicted earlier for various non-­traffic related crimes (Cavaiola et al. 2007). There is increasing evidence that repeat offenders suffer from an alcohol-­use disorder and would benefit from some type of medical intervention (Sloan et al. 2014). The usual sanctions, such as monetary fines, revocation of the driving permit, or impounding the vehicle might be effective in the short term only, because the underlying problem is a substance abuse disorder (Snenghi et al. 2015). The mandatory fitting of an ignition interlock device to the vehicle owned by repeat offenders as one of the conditions for re-­licencing seems well motivated (Voas and Marques 2004). Experience shows that ignition interlock devices

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Hubicka et  al.  2008). There is a clear predominance of male offenders (80–90%) aged between 35 and 45 years with high mean BAC when arrested (1.5–1.7 g/L). Such a high BAC can only be reached after binge drinking, which is usually defined as a pattern of drinking necessary to reach 0.8 g/L or more within 2 hours of starting to drink. The number of standard drinks required to reach a BAC of 0.8 g/L depends on the person’s age and gender and equates to about five or more drinks for men and four drinks for women of average body weight (Courtney and Polich 2009). Many traffic offenders do not have a valid driving permit, probably because of a previous conviction for drunken driving (McCartt and Shabanova 2002). Recidivism rates are high and the best way to deal with hard-­core offenders is a major dilemma for traffic-­safety organizations and the legal authorities. The notion of sentencing repeat offenders to some form of treatment and/or rehabilitation programme for alcohol-­use disorder instead of conventional penalties for drunken driving (fines and short terms of incarceration) should be carefully considered in the future (DeYoung 1997; Fell 2020). Lowering statutory BAC or BrAC limits for driving does not seem to deter hard-­core traffic offenders, especially those suffering from a personality disorder or other mental health problem (Karjalainen et  al.  2013). There is a strong association between mental illness, alcohol and drugs abuse and criminal activity, including multiple arrests for drunken driving and premature death (LaBrie et  al.  2007; Karjalainen et  al.  2019). People who exhibit sensation seeking behaviour are over-­represented among those arrested for driving under the influence of alcohol and/or other drugs (LaPlante et al. 2008). A study of over 100 suspected DUI offenders by the Harvard Medical School showed that more than 45% of these who had been arrested multiple times suffered from a mental health disorder in addition to having problems with alcohol or drug abuse (Shaffer et al. 2007). Road-­safety campaigns should target these high-­risk groups, especially young males and first-­time offenders, before their alcohol-­use disorder becomes more difficult to treat. Drinking habits of women are becoming more like men, and this is also reflected in a higher prevalence of females among people arrested for drunken driving. Organisations in the USA that emerged in the 1980s, such as Mothers Against Drunk Driving (MADD) or Remove Intoxicated Drivers (RID), have done much to draw media attention to the hazards of drunken driving by highlighting family bereavement and the increasing number of people killed on the roads by an impaired driver (Fell and Voas 2006). A promising and recent enforcement strategy is the mandatory fitting of an ignition interlock device to the vehicles owned by first-­time offenders (Elder et al. 2011) and this has done much to prevent recidivism, at least during the time the interlock device is activated (Roth et al. 2007). To start the vehicle, a driver has to pass a breath-­alcohol test, which is set at a relatively low critical threshold, such as 0.2 g/L in blood. Use of other in-­vehicle safety technology might become available in the future which can hinder or prevent an alcohol or drug-­impaired person from starting their vehicle (Voas 2020).

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Borkenstein, R.F., Crowther, R.E., Shumate, R.P. et al. (1974). The role of the drinking driver in traffic accidents (the Grand Rapids study). Blutalkohol 11 (Suppl. 1): 1–132. Borkenstein, R.F. and Smith, H.W. (1961). The Breathalyzer and its applications. Medicine, Science and the Law 1: 13–23. Brick, J. and Erickson, C. K. (2009). Intoxication is not always visible: An unrecognized prevention challenge. Alcoholism, Clinical and Experimental Research 33: 1489–1507. Brumback, T., Cao, D. and King, A. (2007). Effects of alcohol on psychomotor performance and perceived impairment in heavy binge social drinkers. Drug and Alcohol Dependence 91: 10–17. Caldwell, J.P. and Kim, N.D. (1997). The response of the Intoxilyzer 5000 to five potential interfering substances. Journal of Forensic Sciences 42: 1080–1087. Carvalho, A.F., Heilig, M., Perez, A. et al. (2019). Alcohol use disorders. Lancet 394: 781–792. Cavaiola, A.A., Strohmetz, D.B. and Abreo, S.D. (2007). Characteristics of DUI recidivists: A 12-­year follow-­up study of first time DUI offenders. Addictive Behaviors 32: 855–861. Cherpitel, C., Bond, J., Ye, Y. et al. (2005). Clinical assessment compared with Breathalyser readings in the emergency room: Concordance of ICD-­10 Y90 and Y91 codes. Emergency Medicine Journal, 22: 689–695. Christophersen, A.S., Morland, J., Stewart, K. et al. (2016). International trends in alcohol and drug use among vehicle drivers. Forensic Science Review 28: 37–66. Christophersen, A.S., Skurtveit, S., Grung, M. et al. (2002). Rearrest rates among Norwegian drugged drivers compared with drunken drivers. Drug and Alcohol Dependence 66: 85–92. Courtney, K.E. and Polich, J. (2009). Binge drinking in young adults: Data, definitions, and determinants. Psychological Bulletin 135: 142–156. Cromer, J.R., Cromer, J.A., Maruff, P. et al. (2010). Perception of alcohol intoxication shows acute tolerance while executive functions remain impaired. Experimental and Clinical Psychopharmacology 18: 329–339. Crothers, T.D. (1904). Editorial. Quarterly Journal of Inebriety XXV1: 308–309. Curl, A. and Fitt, H. (2019). Will driverless cars be good for us? Now is the time for public health to act together with urban and transport planning. Journal of Global Health 9: 020303. Curry, A.S., Walker, G.W. and Simpson, G.S. (1966). Determination of ethanol in blood by gas chromatography. Analyst, 91: 742–744. Davis, K.C., George, W.H., Norris, J. et al. (2009). Effects of alcohol and blood alcohol concentration limb on sexual risk-­taking intentions. Journal of Studies on Alcohol and Drugs 70: 499–507. Dawson, D.A. (2011). Defining risk drinking. Alcohol Research and Health 34: 144–156. DeYoung, D.J. (1997). An evaluation of the effectiveness of alcohol treatment, driver license actions and jail terms in reducing drunk driving recidivism in California. Addiction 92: 989–997. Drew, G.C., Colquhoun, W.P. and Long, H.A. (1958). Effect of small doses of alcohol on a skill resembling driving. British Medical Journal 2: 993–999. Drew, G.C., Colquhoun, W.P. and Long, H.A. (1959). Effect of Small Doses of Alcohol on a Skill Resembling Driving, Vol. 38. London: HMSO.

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are very effective in preventing people from continuing to drive when drunk (Voas et al. 2016). The enforcement of a zero-­tolerance drink-­driving law is not feasible, because of the existence of small amounts of ethanol (1–3 mg/L) produced endogenously (Walker and Curry 1966). Furthermore, many household products and some pharmaceuticals contain alcohol, such as mouthwash, cough syrups and vitamin tonics, as well as perfumes and colognes, etc. Likewise, during the COVID-­19 pandemic, the widespread use of hand sanitisers containing alcohol is becoming a problem because some people are inclined to drink this product for intoxication purposes (Reynolds et al. 2006). For these reasons, enforcing a BAC of 0.20 g/L or a BrAC of 0.10 mg/L is probably as close as one can come to achieving a zero-­tolerance law for alcohol use before driving.

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Alford, C., Broom, C., Carver, H. et  al. (2020). The impact of alcohol hangover on simulated driving performance during a ‘Commute to Work’-­zero and residual alcohol effects compared. Journal of Clinical Medicine 9: 1435; doi:10.3390/jcm9051435. Alha, A.R. (1951). Blood alcohol and clinical inebriation in Finnish men: A medico-­legal study. Annales Academiæ Scientiarum Fennicæ 26: 1–92. Anderson, J.C. and Hlastala, M.P. (2019). The alcohol breath test in practice: Effects of exhaled volume. Journal of Applied Physiology 126: 1630–1635. Bakke, E., Hoiseth, G., Furuhaugen, H. et al. (2020). Oral fluid to blood concentration ratios of different psychoactive drugs in samples from suspected drugged drivers. Therapeutic Drug Monitoring 42: 795–800. Beck, O., Stephanson, N., Sandqvist, S. et al. (2013). Detection of drugs of abuse in exhaled breath using a device for rapid collection: Comparison with plasma, urine and self-­reporting in 47 drug users. Journal of Breath Research 7: 026006. Beck, O., Ullah, S. and Kronstrand, R. (2019). First evaluation of the possibility of testing for drugged driving using exhaled breath sampling. Traffic Injury Prevention 20: 238–243. Bennett, R.H., Cherek, D.R. and Spiga, R. (1993). Acute and chronic alcohol tolerance in humans: Effects of dose and consecutive days of exposure. Alcoholism, Clinical and Experimental Research 17: 740–745. Berghaus, G., Ramaekers, J.G. and Drummer, O.H. (2007). Demands on scientific studies in different fields of forensic medicine and forensic sciences. Traffic medicine – impaired driver: Alcohol, drugs, diseases. Forensic Science International 165: 233–237. Blackman, K., Voas, R.B. and Gullberg, R.G. (2001). Enforcement of zero tolerance in the state of Washington – evidence from breath test results Forensic Science Review 13: 77–86. Blomberg, R.D., Peck, R.C., Moskowitz, H. et al. (2009). The Long Beach/Fort Lauderdale relative risk study. Journal of Safety Research 40: 285–292. Bonnichsen, R.K. and Theorell, H. (1951). An enzymatic method for the microdetermination of ethanol. Scandinavian Journal of Clinical and Laboratory Investigation 3: 58–62.

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Jones, A.W. (2010). Evidence-­based survey of the elimination rates of ethanol from blood with applications in forensic casework. Forensic Science International 200: 1–20. Jones, A.W. (2011a). Driving under the influence of alcohol. In A.C. Moffat, M.D. Osselton, B. Widdop and J. Watts (eds.), Clarke’s Analysis of Drugs and Poisons, pp. 87–114. London: The Pharmaceutical Press. Jones, A.W. (2011b). Pharmacokinetics of ethanol  – issues of forensic importance. Forensic Science Review 23: 91–136. Jones, A.W. (2016). Impact of trauma, massive blood loss and administration of resuscitation fluids on a person’s blood-­alcohol concentration and rate of ethanol metabolism. Academic Forensic Pathology 6: 77–88. Jones, A.W. (2019a). Alcohol, its absorption, distribution, metabolism, and excretion in the body and pharmacokinetic calculations. WIRE Forensic Science 1: 1–26 https://doi.org/10.1002/wfs1002.1340. Jones, A.W. (2019b). Alcohol, its analysis in blood and breath for forensic purposes, impairment effects, and acute toxicity. WIRE Forensic Science 1: 1–24 https://doi.org/10.1002/wfs1002.1353. Jones, A.W. (2020a). The analysis of ethanol in blood and breath for legal purposes: A historical review. In A.W. Jones, J.G. Morland and R.H. Liu (eds.), Alcohol, Drugs and Impaired Driving: Forensic Science and Law Enforcement Issues (pp. 105–153). Boca Raton: CRC Press. Jones, A.W. (2020b). Use of punishable limits of blood and breath-­ alcohol concentration in traffic-­law enforcement: Some advantages and limitations. In A.W. Jones, J.G. Morland and R.H. Liu (eds.), Alcohol, Drugs and Impaired Driving: Forensic Science and Law Enforcement Issues (pp. 155–202). Boca Raton: CRC Press. Jones, A.W. and Andersson, L. (1996). Variability of the blood/breath alcohol ratio in drinking drivers. Journal of Forensic Sciences 41: 916–921. Jones, A.W. and Andersson, L. (2008). Determination of ethanol in breath for legal purposes using a five-­filter infrared analyzer: Studies on response to volatile interfering substances. Journal of Breath Research 2: 026006. Jones, A.W. and Holmgren, A. (2009). Age and gender differences in blood-­alcohol concentration in apprehended drivers in relation to the amounts of alcohol consumed. Forensic Science International 188: 40–45. Jones, A.W., Kugelberg, F.C., Holmgren, A. et al. (2009). Five-­year update on the occurrence of alcohol and other drugs in blood samples from drivers killed in road-­ traffic crashes in Sweden. Forensic Science International 186: 56–62. Jones, A.W., Morland, J.G. and Liu, R.H. (2019). Driving under the influence of psychoactive substances – a historical review. Forensic Science Review 31: 103–140. Jones, A.W., Morland, J.G. and Liu, R.H. (2020). Alcohol Drugs and Impaired Driving: Forensic Science and Law Enforcement Issues. Boca Raton: CRC Press. Jones, A.W. and Neri, A. (1994). Age-­related differences in the effects of ethanol on performance and behaviour in healthy men. Alcohol and Alcoholism 29: 171–179. Jones, A.W. (2000). Medicolegal alcohol determinations  – blood or breath alcohol concentration?. Forensic Science Review 12: 23–48. Jones, A.W. (2003). Dealing with uncertainty in chemical measurements. IACT Newsletter 14: 6–11.

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O’Neal, C.L., Wolf, C.E., 2nd, Levine, B. et  al. (1996). Gas chromatographic procedures for determination of ethanol in postmortem blood using t-­butanol and methyl ethyl ketone as internal standards. Forensic Science International 83: 31–38. Olson, K.N., Smith, S.W., Kloss, J.S. et al. (2013). Relationship between blood alcohol concentration and observable symptoms of intoxication in patients presenting to an emergency department. Alcohol and Alcoholism 48: 386–389. Paton, A. (2005). Alcohol in the body. British Medical Journal 330: 85–87. Peek-­Asa, C. (1999). The effect of random alcohol screening in reducing motor vehicle crash injuries. American Journal of Preventive Medicine 16: 57–67. Penttila, A. and Tenhu, M. (1976). Clinical examination as medicolegal proof of alcohol intoxication. Medicine, Science and the Law 16: 95–103. Ponce, J.C., Andreuccetti, G., Goncalves, R.E.M. et al. (2019). Comparison of traffic data and blood alcohol concentration among fatally injured drivers in Norway and Sao Paulo, Brazil, 2005-­2015. Traffic Injury Prevention 20: 673–678. Poulsen, H., Moar, R. and Pirie, R. (2014). The culpability of drivers killed in New Zealand road crashes and their use of alcohol and other drugs. Accident Analysis and Prevention 67: 119–128. Probst, C., Kilian, C., Sanchez, S. et al. (2020). The role of alcohol use and drinking patterns in socioeconomic inequalities in mortality: A systematic review. Lancet Public Health 5: e324–e332. Reynolds, S.A., Levy, F. and Walker, E.S. (2006). Hand sanitizer alert. Journal of Environmental Health 69: 48, 51. Roberts, J.R. and Dollard, D. (2010). Alcohol levels do not accurately predict physical or mental impairment in ethanol-­tolerant subjects: Relevance to emergency medicine and dram shop laws. Journal of Medical Toxicology 6: 438–442. Rolison, J.J., Regev, S., Moutari, S. et al. (2018). What are the factors that contribute to road accidents? An assessment of law enforcement views, ordinary drivers’ opinions, and road accident records. Accident Analysis and Prevention 115: 11–24. Romano, E.O., Peck, R.C. and Voas, R.B. (2012). Traffic environment and demographic factors affecting impaired driving and crashes. Journal of Safety Research 43: 75–82. Room, R., Babor, T. and Rehm, J. (2005). Alcohol and public health. Lancet 365: 519–530. Roth, R., Voas, R. and Marques, P. (2007). Mandating interlocks for fully revoked offenders: The New Mexico experience. Traffic Injury Prevention 8: 20–25. Schechtman, E. and Shinar, D. (2011). An analysis of alcohol breath tests results with portable and desktop breath testers as surrogates of blood alcohol levels. Accident Analysis and Prevention 43: 2188–2194. Schell, T.L., Chan, K.S. and Morral, A.R. (2006). Predicting DUI recidivism: Personality, attitudinal, and behavioral risk factors. Drug and Alcohol Dependence 82: 33–40. Schuckit, M.A. (2009). Alcohol-­use disorders. Lancet 373: 492–501. Schweizer, T.A., Vogel-­ Sprott, M., Danckert, J. et  al. (2006). Neuropsychological profile of acute alcohol intoxication during ascending and descending blood alcohol concentrations. Neuropsychopharmacology 31: 1301–1309.

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Legrand, S.A., Gjerde, H., Isalberti, C. et al. (2014). Prevalence of alcohol, illicit drugs and psychoactive medicines in killed drivers in four European countries. International Journal of Injury Control and Safety Promotion 21: 17–28. Lerner, B.H. (2012). Drunk driving across the globe: Let’s learn from one another. Lancet 379: 1870–1871. Lerner, B.H. (2011). One for the Road  – Drunk Driving since 1900. Baltimore: The Johns Hopkins University Press. Lieber, C.S. (1997). Cytochrome P4502E1 Its physiological and pathological role. Physiological Reviews 77: 517–544. Liu, Y.C. and Ho, C.H. (2010). Effects of different blood alcohol concentrations and post-­alcohol impairment on driving behavior and task performance. Traffic Injury Prevention 11: 334–341. Machata, G. (1975). The advantages of automated blood alcohol determination by head space analysis. Zeitschrift für Rechtsmedizin 75: 229–234. Martin, C.S. and Earleywine, M. (1990). Ascending and descending rates of change in blood alcohol concentrations and subjective intoxication ratings. Journal of Substance Abuse 2: 345–352. Martin, C.S. and Moss, H.B. (1993). Measurement of acute tolerance to alcohol in human subjects. Alcoholism, Clinical and Experimental Research 17: 211–216. Martin, T.L., Solbeck, P.A., Mayers, D.J. et  al. (2013). A review of alcohol-­impaired driving: The role of blood alcohol concentration and complexity of the driving task. Journal of Forensic Sciences 58: 1238–1250. Maskell, P.D., Jones, A.W., Heymsfield, S.B. et  al. (2020). Total body water is the preferred method to use in forensic blood-­alcohol calculations rather than ethanol’s volume of distribution. Forensic Science International 316: 110532. McCartt, A.T. and Shabanova, V.I. (2002). Effects of enhanced sanctions for high-­BAC DWI offenders on case dispositions and rates of recidivism. Annual Proceedings of the Association for the Advancement of Automotive Medicine 46: 193–209. McKnight, A.J. and Peck, R.C. (2003). Graduated driver licensing and safer driving. Journal of Safety Research 34: 85–89. Mellanby, E. (1920). Alcohol and alcoholic intoxication. British Journal of Inebriety 17: 157–178. Miller, W.R., Heather, N. and Hall, W. (1991). Calculating standard drink units: International comparisons. British Journal of Addiction 86: 43–47. Mitchell, M.C. (1985). Alcohol-­induced impairment of central nervous system function: Behavioral skills involved in driving. Journal of Studies on Alcohol. Supplement 10: 109–116. Moriya, F. (2005). Forensic sciences  – alcohol in body fluids. In P. Worsfold, A. Townshend and C. Poole (eds.), Encyclopedia of Analytical Sciences (pp. 358–365). London: Academic Press. Moskowitz, H. and Fiorentino, D. (2000). A Review of Literature on the Effects of Low Doses of Alcohol on Driving-­Related Skills. Washington, D.C.: US Department of Transportation National Highway Traffic Safety Administration DOT HS 809 028. Nochajski, T.H. and Stasiewicz, P.R. (2006). Relapse to driving under the influence (DUI): A review. Clinical Psychology Review 26: 179–195. Nutt, D.J., King, L.A. and Phillips, L.D. (2010). Drug harms in the UK: A multicriteria decision analysis. Lancet 376: 1558–1565.

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Voas, R.B. (2020). Vehicle safety features aimed at preventing alcohol-­ related crashes. Forensic Science Review 32: 55–81. Voas, R.B. and Fell, J.C. (2011). Preventing impaired driving opportunities and problems. Alcohol Research and Health 34: 225–235. Voas, R.B. and Marques, P.R. (2004). Emerging technological approaches for controlling the hard core DUI offender in the U.S. Traffic Injury Prevention 5: 309–316. Voas, R.B., Tippetts, A.S., Bergen, G. et al. (2016). Mandating treatment based on interlock performance: Evidence for effectiveness. Alcoholism, Clinical and Experimental Research 40: 1953–1960. Voas, R.B., Torres, P., Romano, E. et al. (2012). Alcohol-­related risk of driver fatalities: An update using 2007  data. Journal of Studies on Alcohol and Drugs 73: 341–350. Vosk, T. and Emery, A.F. (2015). Forensic Metrology: Scientific Measurement and Inference for Lawyers, Judges, and Criminalists. Boca Raton: Taylor & Francis. Walker, G.W. and Curry, A.S. (1966). ‘Endogenous’ alcohol in body fluids. Nature 210: 1368. Wallner, M. and Olsen, R.W. (2008). Physiology and pharmacology of alcohol: The imidazobenzodiazepine alcohol antagonist site on subtypes of GABAA receptors as an opportunity for drug development? British Journal of Pharmacology 154: 288–298. Watson, P.E., Watson, I.D. and Batt, R.D. (1981). Prediction of blood alcohol concentrations in human subjects. Updating the Widmark equation. Journal of Studies on Alcohol 42: 547–556. WHO. (2018). Global Status Report on Road Safety 2018, Geneva, World Health Organization, https://www.Who.Int/violence_injury_ prevention/road_safety_status/2018/gsrrs2018_summary_en.Pdf. Widmark, E.M.P. (1922). Eine Mikromethode zur Bestimmung von Äthylalkohol im Blut. Biochemische Zeitschrift 131: 473–484. Widmark, E.M.P. (1932). Die theoretischen Grundlagen und die praktische Verwendbarkeit der gerichtlich-­ medizinischen Alkoholbestimmung. Berlin: Urban & Schwarzenberg. Widmark, E.M.P. (1981). Principles and Applications of Medicolegal Alcohol Determinations. Davis: Biomedical Publications. Williams, A.F., McCartt, A.T. and Ferguson, S.A. (2007). Hardcore drinking drivers and other contributors to the alcohol-­impaired driving problem: Need for a comprehensive approach. Traffic Injury Prevention 8: 1–10. Zamengo, L., Tedeschi, G., Frison, G. et al. (2019). Inter-­laboratory proficiency results of blood alcohol determinations at clinical and forensic laboratories in Italy. Forensic Science International 295: 213–218. Zanger, U.M. and Schwab, M. (2013). Cytochrome p450 enzymes in drug metabolism: Regulation of gene expression, enzyme activities, and impact of genetic variation. Pharmacology and Therapeutics 138: 103–141. Zuba, D. (2008). Accuracy and reliability of breath alcohol testing by handheld electrochemical analysers. Forensic Science International 178: e29–33.

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Searle, J. (2015). Alcohol calculations and their uncertainty. Medicine, Science and the Law 55: 58–64. Seto, Y. (1994). Determination of volatile substances in biological samples by headspace gas chromatography. Journal of Chromatography A 674: 25–62. Shaffer, H.J., Nelson, S.E., LaPlante, D.A. et al. (2007). The epidemiology of psychiatric disorders among repeat DUI offenders accepting a treatment-­sentencing option. Journal of Consulting and Clinical Psychology 75: 795–804. Shiner, D. (2007). Traffic Safety and Human Behavior. Bingley: Emerald Group Publishing Limited. Skurtveit, S., Christophersen, A.S. and Morland, J. (1995). Female drivers suspected for drunken or drugged driving. Forensic Science International 75: 139–148. Sloan, F.A., Eldred, L.M. and Davis, D.V. (2014). Addiction, drinking behavior, and driving under the influence. Substance Use and Misuse 49: 661–676. Snenghi, R., Forza, G., Favretto, D. et al. (2015). Underlying substance abuse problems in drunk drivers. Traffic Injury Prevention 16: 435–439. Sterling, K. (2012). The rate of dissipation of mouth alcohol in alcohol positive subjects. Journal of Forensic Sciences 57: 802–805. Strayer, D.L., Drews, F.A. and Crouch, D.J. (2006). A comparison of the cell phone driver and the drunk driver. Human Factors 48: 381–391. Swift, R. and Davidson, D (1998). Alcohol hangover – mechanism and mediators. Alcohol, Health Research World 22: 54–60. Tagliaro, F., Lubli, G., Ghielmi, S. et al. (1992). Chromatographic methods for blood alcohol determination. Journal of Chromatography 580: 161–190. Tiplady, B., Franklin, N. and Scholey, A. (2004). Effect of ethanol on judgments of performance. British Journal of Psychology 95: 105–118. Tiscione, N.B., Alford, I., Yeatman, D.T. et al. (2011). Ethanol analysis by headspace gas chromatography with simultaneous flame-­ionization and mass spectrometry detection. Journal of Analytical Toxicology 35: 501–511. Trefz, P., Kamysek, S., Fuchs, P. et al. (2017). Drug detection in breath: Non-­invasive assessment of illicit or pharmaceutical drugs. Journal of Breath Research 11: 024001. Ullah, S., Sandqvist, S. and Beck, O. (2018). A liquid chromatography and tandem mass spectrometry method to determine 28 non-­volatile drugs of abuse in exhaled breath. Journal of Pharmaceutical and Biomedical Analysis 148: 251–258. Valen, A., Bogstrand, S.T., Vindenes, V. et al. (2019). Fatally injured drivers in Norway 2005-­2015 -­Trends in substance use and crash characteristics. Traffic Injury Prevention 20: 460–466. Vindenes, V., Jordbru, D., Knapskog, A.B. et  al. (2012). Impairment based legislative limits for driving under the influence of non-­alcohol drugs in Norway. Forensic Science International 219: 1–11.

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Effects of Illegal Drugs on Fitness to Drive

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Persons who are addicted to prescription medications or consume certain substances abusively or regularly are also not qualified to drive a vehicle since their mental and physical performance is low and their decision-­making ability can be impaired suddenly and unexpectedly by the drugs taken. Centrally acting substances influence inhibition control as well as intellectual and motor skills and are thus similar to alcohol regarding their individual effects. There are a number of different legal approaches. An approach focusing on impairment can show if the driver has clear symptoms of impairment, whether this is in personal behaviour or driving style, leading to prosecution. In some countries, there are so-­ called ‘per se limits’ for illicit drugs and if drugs are found in a driver’s body fluid (blood and in some countries oral fluid) above a defined cut-­off concentration, he or she will be prosecuted. In a few countries (e.g. in Germany), there is a two-­tier system that allows combining the advantages of two legal regulations  – for example combining a lighter sanction when drugs are present above the ‘per se limits’ and more severe sanctions when the driver was actually impaired. Civil and insurance legislation might be affected as well: • There may be joint liability of the insured person when an accident is caused by alcohol consumption or the use of other intoxicating substances. • The insurer can subrogate against the insured individual if an accident is alcohol-­or drug-­related. • Car insurance companies can refuse payment if an accident is related to alcohol or drugs.

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Driving in road traffic is subject to numerous legal regulations. Thus, a driving licence can only be obtained by a person with a physical and mental state that conforms to legal requirements and who has not previously violated either seriously or repeatedly traffic rules or legislation. Driving capability describes that part of driving that comprises training, exercise and experience in the sense of ‘skilfulness’ necessary for driving a vehicle safely. The term fitness to drive refers to a person’s condition to drive a ­vehicle in a certain situation at a particular time. This fitness can change quickly due to external factors and impairment of the driver (caused by alcohol, drugs, pharmaceuticals, tiredness, etc.). Driving ability means the permanent ability to drive a vehicle based on a driver’s personal character traits. Drinking alcohol or taking drugs and/or pharmaceuticals may cause reversible unfitness to drive if the driver’s abilities are reduced due to mental and physical deterioration of actual performance. Due to this deterioration of performance, the driver is actually unable to drive longer distances or to react appropriately to sudden changes in traffic. Central nervous system depressants alone and combined with other factors (e.g. tiredness and intake of pharmaceuticals) may be relevant in this context. A person endangering traffic safety due to physical, mental or personal problems is unable to drive a vehicle safely. Persons with pre-­existing diseases as well as persons consuming cannabis or other illegal drugs on a regular basis are unqualified to drive.

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Handbook of Forensic Medicine, Volume 3, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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Epidemiological data on the problem of driving under the influence of drugs (DUID) can be obtained by several procedures. According to Raes et  al. (2008), epidemiological studies can be classified into the following types: 1. Roadside surveys. 2. Prevalence studies in subsets of drivers: a. drivers injured in traffic accidents b. drivers killed in traffic accidents c. drivers involved in traffic accidents d. drivers suspected of driving under the influence of drugs e. drivers suspected of driving under the influence of alcohol. 3. Surveys on driving under the influence of drugs. 4. Accident risk studies. 5. Responsibility analysis studies. 6. Pharmaco-­epidemiological studies. Roadside surveys are used to determine the prevalence of drug use in the general driving population. In these surveys, drivers are randomly selected and tested for drugs and alcohol. In the European Union’s research project on Driving Under the Influence of Drugs, Alcohol and Medicines (DRUID project), roadside surveys were conducted in 13 countries across Europe. The analyses of blood or oral fluid samples from 50 000 drivers revealed that alcohol was present in 3.48%, illicit drugs in 1.90%, medicines in 1.36%, combinations of drugs or medicines in 0.39% and alcohol combined with drugs or medicines in 0.37% (EMCDDA 2012). Prevalence studies investigating drug use among drivers suspected of driving impaired usually show a higher prevalence of licit and illicit drugs, but the detection of these drivers is highly dependent on the officer’s subjective impressions. There are remarkable differences between countries, possibly because of varying national road traffic regulations and the amount of attention paid to the problem. Maes et  al. (1999) reported that there are also different methods of data collection and analysis, especially regarding individual attitudes towards consumption behaviour. Additionally, descriptive epidemiological studies only examine the prevalence of certain drugs in road traffic, but do not elucidate the role of drugs in crash responsibility. For this purpose, responsibility analysis studies might give answers. In the Roadside Testing Assessment (ROSITA) project, Maes et  al. (1999) reported a study by Drummer (1994) who evaluated data from 1045 drivers killed in motor vehicle accidents in Australia. Responsibility was determined using eight factors (without knowing the results of drug testing), and drivers were then classified into three groups: culpable, contributory and not culpable. The culpability ratio was calculated, and drivers with higher than therapeutic drug concentrations were either culpable for or contributory to the accident. Persons driving under the influence of more than one drug were found to be culpable for the accident in each case. For cannabis-­ positive drivers, lower culpability ratios were found. However, this study also included

drivers with only the inactive metabolite of Δ9-­ tetrahydrocannabinol (THC) 11-­ nor-­ Δ9-­tetrahydrocannabi nol-­9-­carboxylic acid (THC-­COOH) in their blood. In contrast, ­drivers who tested positive for THC only showed an increased crash risk (Drummer et al. 2004). Terhune et al. (1992) reported in a study of 1882 fatally injured drivers in the United States that the responsibility rates of drivers with alcohol alone and with alcohol–drug combinations were significantly higher than that of the drug-­free drivers. Drivers positive for THC only had a responsibility rate below that of the drug-­ free group. Conversely, the responsibility rate of the amphetamine-­only drivers was higher than the drug-­free rate. Drivers who tested positive for benzodiazepines only did not have an increased risk. In contrast, Drummer (1994) found higher culpability ratios for drivers with benzodiazepines. In both studies, however, significantly higher culpability ratios were observed when benzodiazepines were combined with alcohol. Moreover, Terhune et al. (1992) found that crash responsibility increased significantly with the number of drugs detected in a driver’s blood. Longo et al. (2000) reported that drivers who tested positive for alcohol only, benzodiazepines only, alcohol and THC, and alcohol and benzodiazepines were significantly more likely to be responsible for the car crash than those of the drug-­free group. For drivers with THC only (with increasing concentration), no increased crash risk was observed. This study again demonstrates that a combination of substances increases the accident risk. For medicines, pharmaco-­epidemiological studies can provide very useful information on the risk of crash involvement. Elderly drivers in particular have a relative risk of injurious crash involvement if they are under medication (benzodiazepines, antidepressants and others) which is markedly dose-­dependent (Maes et al. 1999). Raes et al. (2008) concluded that epidemiological studies are limited since there may be risk factors associated with drug use which do not emerge from the study results. Furthermore, it is not possible to distinguish ‘real’ risk factors from other factors that might be highly correlated with an actual risk factor (Berghaus  2007). Epidemiological studies are also difficult to compare because of differences in study design such as the following (summarised from Raes et al. 2008): • Different sample populations (age, gender, etc.). • Different times at which studies are performed, such as year and day of the week (e.g. studies conducted on weekend nights with higher percentages of drug-­positive drivers). • Biological samples are analysed for different types of psychoactive substances (prevalence results can depend on the number and types of drugs tested). • Different types of biological samples are used with varying windows of detection (e.g. cannabinoid metabolites can be detected in urine for a relatively long period and its presence in urine does not necessarily prove an acute effect). • Different analytical techniques are used to analyse samples with different limits of detection and quantification. • Different cut-­off levels are used in analyses. Considering all these factors, there is a potential for bias in epidemiological studies, making it almost impossible to compare

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60.2 Epidemiological data and its limitations

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60.4.1 Cannabis

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Cannabis represents the preparation of the Cannabis sativa plant. Used products are marihuana (dried parts of the plant), hashish (resin of the female flowering tops) and hashish oil (extract from the resin). Further data are summarised in Chapter 51. In general, cannabis consumption can lead to disturbances in time perception, changes in visual and auditory perception, decreased concentration and reduced ability to react quickly. Although several studies have been carried out, there is no consensus regarding the significance of the adverse effects on safe driving caused by cannabis abuse. Laboratory studies have mostly shown considerable negative effects on individual parameters. Driving simulator and on-­road studies regularly show slight adverse effects. Effects and side effects of cannabis consumption that may affect driving are as follows: sedation, intense tiredness, motor dysfunction, delayed reaction, concentration and attention problems, perception of irrelevant stimuli (causing subsequent disregard of traffic lights, etc.) and reduced ability to detect moving objects (e.g. pedestrians intending to cross the road and playing children). These effects may result in varying driving speeds, lane weaving and subsequent steering corrections. Especially in stressful and complex situations, prolonged reaction times, accumulation of wrong or inappropriate r­ eactions

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In general, drivers can be stopped by the police and tested for alcohol and drugs for several reasons, including, for example, general or random traffic controls, specific roadblocks, accidents or if the officer has a reasonable suspicion that someone is driving while impaired. This suspicion may be established by observing inappropriate driving behaviour including for example weaving, driving without headlights at night or speed and braking problems (Box 60.1). These observations of the vehicle in motion are considered as Phase 1 of the DUID detection process. In Phase 2, the officer will engage the driver in conversation and look for signs of impairment such as bloodshot eyes, slurred speech, slowed reaction time, sweating or odour of alcohol or marijuana. If the officer suspects that the driver is intoxicated, they will instruct the driver to exit the vehicle and observe the driver’s exit. In this phase, the officer can also perform preliminary tests (e.g. breath alcohol test and on-­site drug testing). During Phase 3, the officer having probable cause that the driver is impaired will arrest them and transport them to the police station for a medical examination and blood sampling. All observations made during these phases have to be carefully documented and are used along with the blood test result in court as evidence of impaired driving. In addition to breath alcohol tests, on-­site urine or saliva drug tests are increasingly used by the police to obtain sufficient evidence to justify a blood test. However, like on-­site breath alcohol tests, on-­ site drug tests are only preliminary tests based on immunochemical methods that still have substantial disadvantages (Table 60.1).

The next sections give an overview of drug effects on driving based on various sources: ADF (2020), Austrian Road Safety Board (2003), Couper et al. (2004), Madea et al. (2012), Maes et al. (1999), Musshoff (2011), Pennings et  al. (2002), Raes et  al. (2008) and Verster et  al. (2009). The following references are recommended for further reading: Jones (2007), Jones et  al. (2008), Logan (1996, 2005), Madea et al. (2011, 2012) and Musshoff (2011).

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different results. Consequently, there is obviously a need for methodological guidelines (Raes et al. 2008). Recently, a consensus report has been published defining guidelines, standards, core data variables and other controls that could be the basis for future research (Walsh et al. 2008).

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Table 60.1  Advantages and disadvantages of the different matrices for roadside drug testing. Advantages

Disadvantages

Urine

Availability of several reliable on-­site tests High concentrations of drugs/metabolites present Robust and well-­known technology

Delayed appearance of drugs in urine No correlation with impairment Sampling difficult at the roadside Risk of sample adulteration

Oral fluid (saliva)

Presence of parent drug Sampling can be performed without embarrassment Some correlation with impairment

No reliable on-­site test presently available Little and very viscous oral fluid after recent intake of drug Very low concentrations of THC and benzodiazepines Still much research necessary

Sweat

Presence of parent drug Sampling can be performed without embarrassment

No reliable on-­site test presently available No standardisation of sampling Delayed appearance of drugs in sweat Very low concentrations of THC and benzodiazepines Possibility of environmental contamination Still much research necessary

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Matrix

THC, Δ9-­tetrahydrocannabinol. Source: Summarised from Walsh et al. (2004).

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Box 60.1  DUID detection process.

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First phase: vehicle in traffic/traffic accident • Weaving from side to side, hugging the centre line and straddling a line • Inappropriate steering, skidding and inappropriate speed • Violation of right of way and endangering other road users • Nearly hitting objects on or beside the road • Driving on closed roads • Driving without headlights at night and incorrect signalling • Driving too close behind the car ahead and driving into oncoming or crossing traffic • Illegal or sudden turning, turning with a wide radius or hitting the kerb when turning right • Stopping for no apparent reason or stopping inappropriately • Slow response to traffic lights and sudden acceleration or braking for no apparent reason • Suspicious operation of the vehicle or suspicious state of the vehicle • Unusual behaviour of the passengers • Accidents: veering off the road, collisions with the kerb or collisions with oncoming or crossing vehicles, rear-­end collisions, accidents due to inappropriate speed and collisions while parking

Centrally depressant THC effects slow down the activity of the brain and other areas of the central nervous system (CNS), while minor hallucinogenic effects can distort a person’s perception of the environment. The effects of cannabis can be different for each person and are influenced by factors such as mentioned below: • Dose (dependent on the THC content of the preparation). • Ingestion route: When cannabis is smoked, the effects are experienced very quickly and may last up to 5–8 hours; after oral consumption, the onset of effects can be delayed by about 60–90 minutes and can last up to 24 hours. • Consumer’s psychological and physical attributes, and general factors such as mental or emotional state and physical health can also influence individual responses to drugs. Therefore, it is difficult to accurately predict in what way and for how long cannabis will affect a person’s ability to drive safely. As a general guideline, some of the effects of cannabis that can affect a person’s driving ability include the following: ○○ Decreased coordination and slower reaction times. ○○ Reduced information processing ability, confusion and impaired thinking. ○○ Changes in visual, auditory, time and space perception. In general, it is difficult to relate THC blood concentrations to performance impairment. Concentrations of THC and its metabolites are extremely dependent on dose and pattern of use. THC concentrations typically peak during smoking, decrease rapidly thereafter and are often 1.0 >1.0 >0.8 0.5–0.8   >0.5

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Lormetazepam

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Lemborexant Loprazolam

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Gaboxadol

N

Flurazepam

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Flunitrazepam

Reference

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Day and time of testing

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Dose regimen

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Hypnotic

A: afternoon; BAC, blood alcohol concentration; M: morning; MOTN: middle-­of-­the-­night administration; SUB: sub-­lingual. a  In a patient sample. b  In an elderly sample (65–80 years).

A 40 mg dose impaired driving in the morning after 1 and 8 days of dosing. All increases in SDLP were, however, small (Vermeeren et al. 2015). In an elderly population (65–76 years), evening doses of 15 and 30 mg of suvorexant did not impair driving performance in the morning after 1 or 8 days of treatment (Vermeeren et al. 2016).

Lemborexant also acts as an antagonist at orexin receptors (tmax = 1–2 hours). In a population of healthy adult and elderly volunteers, lemborexant in bedtime doses of 2.5, 5 and 10 mg did not impair driving performance after 1 or 8  days of dosing (Vermeeren et al. 2019).

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Table 61.2  Overview of the studies testing anxiolytics in the on-­the-­road driving test. The acute and sub-­chronic effects are indicated in terms of blood alcohol concentration (BAC) required to achieve the same impairment level. In bold are results on the driving test that are comparable or larger than the effect of a BAC 0.5 g/L. Significant impairment

Equivalent BAC (g/L)

Alprazolam   Alprazolam XR Buspirone

1 mg 1 mg 1 mg 5 mg tid 10 mg tid 5 mg tid 5 mg 5 mg 10 mg 10 mg 5 mg tid 5 mg tid 0.5 mg tid 1 mg tid 1.5 mg bid 10 mg 30 mg 10 mg tid 50 mg evening dose 5 mg bid 0.2 mg tid

Day 1 Day 1 Day 1 Day 1 Day 1 Day 1 Day 1 Day 1 Day 1 Day 1 Day 1 Day 8 Days 2, 9 Day 1 Day 7 Day 1 Day 1 Day 1 Day 2 (10 h) Day 2 (16 h) Day 7 Days 2, 9

Yes Yes Yes Yes Yes Yes No Yes Yes Yes Yes Yes Yes Yes Yes Yes Yes Yes Yes No No Yes

>1.5 >1.0 >0.5 ≈0.5 ≈0.5 ≈0.5   ≈0.5 ≈1.0 >0.5 >0.5 >1.0 >0.8 and >1.0 >1.5 >1.0 1.0 >0.5 >0.5

Ritanserin Suriclone

Verster et al. (2002b) Leufkens et al. (2007) Leufkens et al. (2007) Volkerts et al. (1987)

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Oxazepam

>1.0 and >0.5

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Lorazepam

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Clorazepate Diazepam

Reference

Brookhuis and Borgman (1988) O’Hanlon et al. (1982) Van Veggel and O’Hanlon (1993) O’Hanlon et al. (1982) Jongen et al. (2018) Volkerts et al. (1987) Van Veggel and O’Hanlon (1993) Uiterwijk and O’Hanlon (1994) Volkerts et al. (1987) van Laar et al. (2001) Jongen et al. (2018) Jongen et al. (2018) Brookhuis and Borgman (1988) Volkerts et al. (1992a) Volkerts et al. (1992a) van Laar et al. (2001) Uiterwijk and O’Hanlon (1994)

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Day of testing (time post-­dose)

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Dose regimen

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Anxiolytic

BAC, blood alcohol concentration; bid, twice a day; pd, post dose; tid, three times a day; XR: extended release.

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Because of their short half-­life, some hypnotics have a shorter duration of action and less severe residual effects. However, insomniacs also often take this medication in the middle of the night, so it is also important to measure the effects after MOTN administration. The short-­acting hypnotic zaleplon (Tmax  = 1 hour) had no significant residual effects on driving performance in doses of 10 and 20  mg, both after an evening and MOTN administration (Vermeeren et  al. 1998a;Vermeeren et  al. 2002b; Verster et  al. 2002a). For zolpidem (Tmax = 45 min), residual impairment was found on driving performance after MOTN treatment with 10 and 20  mg doses (Verster et  al. 2002a; Leufkens et  al. 2009b). Administration of zolpidem 10 mg in the evening did not impair driving performance in women complaining of chronic sleep disturbances (Vermeeren et  al. 1995). Buffered sub-­ lingual tablets of zolpidem (ZST) 3.5 mg reach Tmax rapidly (±38 min.). When administered in the middle of the night, ZST significantly impaired driving, although the SDLP increases were small (Vermeeren et al. 2014). For zopiclone 7.5  mg, evening doses were found to produce residual driving impairment in the morning in several studies (Vermeeren et al. 1998a; Vermeeren et al. 2002b; Leufkens et al. 2009a; Leufkens et al. 2009b; Mets et al. 2011; Ramaekers et al. 2011; Leufkens et  al. 2014b; Vermeeren et  al. 2014; Vermeeren

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et  al. 2015; Vermeeren et  al. 2016; Vermeeren et  al. 2019). As expected, the residual effects after MOTN administration were even more extensive (Vermeeren et al. 1998a). A pooled analysis confirmed that zopiclone 7.5  mg has significant and clinically relevant residual effects on driving in the morning (Leufkens and Vermeeren 2014). This analysis also showed that the effects were independent of sex or age, at least until the age of 75. Zopiclone 7.5 mg was found to also cause significant driving impairment in insomnia patients although to a lesser extent than in healthy controls (Leufkens et al. 2014b). Gaboxadol, a GABA-­a receptor agonist, has comparable effects on driving as the non-­benzodiazepines. Driving performance was unimpaired after an evening dose of 15  mg, whereas an MOTN dose did impair driving in the morning (Leufkens et al. 2009b). Esmirtazapine is the (S)-­(+)-­enantiomer of mirtazapine ­(anti-­depressant) and has similar overall pharmacology, including actions at H1, 5-­HT2 and α2-­adrenergic receptors. Esmirtazapine was under development for the treatment of insomnia. After single and repeated dosing with esmirtazapine 1.5  mg, driving performance was not impaired. A single dose of 4.5 mg did cause driving impairment (almost equivalent to a BAC of 0.5 g/L), but after repeated doses, this impairment was no longer present (Ramaekers et al. 2011). Ramelteon is a selective melatonin agonist with minimal affinity for GABA receptors. Therefore, it was expected to have less impairing effects compared with other hypnotics. However, a

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Short-­acting hypnotics (half-­life 0.8

84 mg IN 20 mg 50 mg 50 mg bid 10 mg tid

Day 1 (8 h) Days 1, 8, 22 Day 1 After 1 week Day 1

No No Yes No Yes

10 mg tid 15 mg days 1–7 30 mg days 8–15 30 mg 30 mg days 1–7 30 mg days 1–7 30 mg days 1–15   45 mg days 8–15 60 mg days 8–15

After 1 week Days 2, 8 Days 9, 16 Day 1 (8 h) Day 2 Day 2 Day 2 Day 16 Days 9, 16   Days 9, 16 Days 1, 8 Days 1, 8 Day 1 Day 8 Days 1, 8 Days 1, 7, 8, 15 Days 8, 15 Days 2, 16

Yes No No Yes Yes Yes Yes No No   Yes No No No Yes No No No No

Moclobemide Nefazodone

Paroxetine Venlafaxine

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Robbe and O’Hanlon (1995) Robbe and O’Hanlon (1995); Louwerens and Brookhuis (1984) Ramaekers et al. (1995) Ramaekers et al. (1995) Ramaekers et al. (1995) Louwerens and Brookhuis (1984); Schoenmakers et al. (1989); Ramaekers et al. (1994) Schoenmakers et al. (1989); Ramaekers et al. (1994) van de Loo et al. (2017) Ramaekers et al. (1995) van Laar et al. (1995) van Laar et al. (1995) Louwerens and Brookhuis (1984); Ramaekers et al. (1992a); O’Hanlon et al. (1998) O’Hanlon et al. (1998) Ramaekers et al. (1998) Ramaekers et al. (1998) van de Loo et al. (2017) Wingen et al. (2005) Ramaekers et al. (1998) Theunissen et al. (2013) Theunissen et al. (2013) Wingen et al. (2005)   Ramaekers et al. (1998) Ramaekers et al. (1992a) van Laar et al. (1995)

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0.5 ≈0.5 0.8

0.5 >0.5

Theunissen et al. (2004) Theunissen et al. (2004) Theunissen et al. (2006) van der Sluiszen et al. (2016) Theunissen et al. (2006) Theunissen et al. (2004) Vuurman et al. (1994) Vuurman et al. (1994) Vuurman et al. (1994)

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Table 61.4  (Continued) Day of testing (time post-­dose)

Gender

Significant impairment

Rupatadine Terfenadine

10 mg 60 mg od

Day 1 Day 1 (1 and 3 h)

Mixed Males, females

No No

60 mg bid

Day 1 (1 h)

Males

No

60 mg bid

Day 4 (1 and 2 h)

Males

No

120 mg od

Day 1 (1 and 3 h)

Females, males

No

180 mg od 120 mg bid 120 mg bid 120 mg bid

Day 1 (1 and 3 h) Day 1 (2 and 4 h) Day 4 (2 h) Day 4 (4 h)

Females Mixed Mixed Mixed

No No Yes No

Equivalent BAC (g/L)

0.8 and 1.0 g/L, respectively) and midazolam increased SDLP only on day 5 (equivalent to a BAC of 3 nights/week) (Leufkens et al. 2014a). Both groups of patients did not demonstrate impaired driving compared to healthy controls. A subsequent study compared the effects of zopiclone 7.5 mg in chronic and infrequent hypnotics users and confirmed driving impairment in both patient groups, although this was less pronounced than a healthy control group (Leufkens et al. 2014b). In a recent study, driving performance of long-­term users of benzodiazepine hypnotics was compared to a normative control group of healthy volunteers (van der Sluiszen et al. 2019). Driving performance was found to be impaired in patients treated with hypnotics compared to controls. This seemed mainly due to patients who had been using hypnotics for less than 3 years, as those who had received hypnotic treatment for more than 3 years did not show driving impairment (van der Sluiszen et al. 2019). Another concern is that gender differences have been found in some studies, showing that females are more susceptible to the impairing effects of benzodiazepine hypnotics and zolpidem (Verster et al. 2002a, 2004), possibly due to differences in weight and body fat distribution. Several other studies, including a pooled analysis of four studies with zopiclone, did not show differences between men and women (Leufkens and Vermeeren  2014). Because gender differences could not be

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BAC, blood alcohol concentration; bid, twice a day; CR, controlled release; od, once a day; pd, post dose; tid, three times a day; XR: extended release.

61.2.2 Anxiolytics Anxiolytic drugs are used for the treatment of anxiety. Most of the anxiolytic drugs on the market work via GABA, while a few act on serotonin. The studies described below mostly examined the acute effects of anxiolytics, but some also investigated the effects after sub-­chronic or repeated dose treatment. An overview of the results of anxiolytics on the on-­the-­road driving test is shown in Table 61.2.

Benzodiazepines The effects of alprazolam 1  mg on driving performance were quite severe; one study found an increase in SDLP comparable to

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driving performance than the serotonergic drugs that have a lower potential for impairment. The effects of anxiolytics are also found to be dose-­related. In addition to the studies with healthy volunteers described above, several researchers have looked at the effects of anxiolytics on driving performance in anxious patients (Brookhuis and Borgman  1988; van Laar et  al. 1992; Vermeeren et  al. 1994; O’Hanlon et  al. 1995). These studies demonstrated that the effects  of diazepam, lorazepam, buspirone, oxazepam and clorazepate in patients are comparable to the effects in healthy volunteers, and that improvement in their condition does not lead to an improvement in drug-­induced driving impairment. In one study, patients were treated with diazepam 5  mg tid for 4  weeks and it was established that tolerance to the effects of diazepam develops very slowly as the increase in SDLP was still larger than a BAC equivalent to 0.5 g/L in week 4. In a study with term daily users of benzodiazepine-­ related anxiolytics long-­ (including alprazolam, lorazepam and oxazepam), driving performance was found not to be impaired compared to a matched control group (van der Sluiszen et al. 2019).

61.2.3  Anti-­depressants There are several different classes of anti-­depressants. These classes are based on their mechanism of action. Because of this difference in mechanism, the effects of anti-­ depressants on driving performance are very diverse. An overview of the results of anti-­depressants on the on-­the-­road driving test is shown in Table 61.3.

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the effects of a BAC of 1.5 g/L (Verster et al. 2002b; Leufkens et al. 2007). When administered as an extended-­release formulation (in which the drug is slowly released over time), the effect of alprazolam 1  mg was much less, but still considerable (>BAC 0.5 g/L) (Leufkens et al. 2007). Lorazepam in doses of 0.5 mg three times a day (tid) and 1 mg tid caused severe acute driving impairment (Volkerts et al. 1987; Brookhuis and Borgman  1988). Treatment with lorazepam 0.5 mg tid still caused severe driving impairment on days 2 and 9 (Uiterwijk and O’Hanlon 1994). Repeated dose treatment with lorazepam 1.5  mg twice a day (bid) showed that impairment persisted for at least a week (van Laar et al. 2001). Diazepam’s effects on driving have been tested in doses of 5–15 mg a day. A 5 mg dose did not impair driving acutely in one study (O’Hanlon et al. 1982), but did cause mild impairment in another study (Van Veggel and O’Hanlon 1993). A three-­ times-­daily dosing with diazepam 5  mg (Volkerts et  al. 1987) and a 10  mg dose caused significant driving impairment (O’Hanlon et  al. 1982; Jongen 2018). Sub-­chronic effects of doses of 5 mg diazepam tid caused driving impairment larger than the effect of a BAC of 1.0 g/L (Van Veggel and O’Hanlon 1993). The active metabolite of diazepam, oxazepam, demonstrated impairing effects of driving performance after several dosing regimens. Single oral doses of oxazepam 10  mg caused mild driving impairment acutely, while a 30 mg dose severely impaired performance, comparable to a BAC > 1.0 g/L (Jongen et al. 2018). A 10  mg dose tid also resulted in acutely impaired driving performance (Brookhuis and Borgman  1988). Volkerts et  al. (1992) demonstrated that a 50  mg evening dose’s residual impairing effect was no longer present in the afternoon (Volkerts et al. 1992a). Suriclone 0.2  mg tid significantly impaired driving performance, both on the second day of treatment (BAC >1.0 g/L) and after 9  days of treatment (BAC >0.5  g/L) (Uiterwijk and O’Hanlon 1994; O’Hanlon et al. 1995). Clorazepate 5 mg tid had moderate impairing effects in healthy volunteers (Brookhuis and Borgman 1988).

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Buspirone is the only serotonergic anxiolytic currently on the market. Buspirone given in doses of 5 and 10 mg tid both impaired driving performance acutely to about the same extent as a BAC of 0.5 g/L (Volkerts et al. 1987). Ritanserin is another serotonergic anxiolytic, although the development of this drug has discontinued. One study looked at the sub-­chronic effects of ritanserin 5 mg bid and found that after 7 days of treatment ritanserin did not cause driving impairment (van Laar et al. 2001).

Discussion Most of the currently available anxiolytics act on the GABA system. Overall, the GABA agonists have a more severe effect on

Sedating anti-­depressants Tricyclic anti-­depressants (TCAs) are the oldest class of anti-­ depressants. They act on cholinergic, adrenergic and histaminergic receptors and are therefore related to cognitive impairment and sedation. TCAs have been found to result in serious driving impairment. TCAs such as doxepine (Louwerens and Brookhuis 1984; Schoenmakers et  al. 1989; Ramaekers et  al. 1994), imipramine (van Laar et al. 1995) and amitriptyline (Robbe and O’Hanlon 1995) cause a significant driving impairment with SDLP increases comparable to the effect of a BAC of 0.8 g/L at clinically effective doses. After 1 or 2 weeks of treatment, TCAs no longer impair driving performance as a result of tolerance (Louwerens and Brookhuis  1984; Schoenmakers et  al. 1989; Ramaekers et al. 1994; van Laar et al. 1995). A way to overcome TCAs’ acute impairing effects is to administer them as a nocturnal dose, so that only residual effects remain in the morning. A study with nocturnal dothiepin doses found no residual impairment in the morning after a single nocturnal dose (Ramaekers et al. 1995). Impairment was still absent after repeated nocturnal doses with dothiepin (Ramaekers et al. 1995). The noradrenergic and specific serotonergic anti-­depressants, such as mianserine and mirtazapine, significantly impair driving

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No driving impairment was found for the following: moclobemide (Ramaekers et  al. 1992a), a reversible inhibitor of monoamine oxidase A; selective serotonin reuptake inhibitors such as fluoxetine (Ramaekers et  al. 1995) and paroxetine (Robbe and O’Hanlon 1995; van Laar et al. 1995); and venlafaxine (O’Hanlon et  al. 1998), a serotonin–norepinephrine reuptake inhibitor. Nefazodone, a serotonin receptor antagonist and reuptake inhibitor, only caused minor driving impairment after repeated administration of a high dose (van Laar et  al. 1995). Also no driving impairment was found for the more recently developed atypical anti-­depressants, vortioxetine (Theunissen et  al. 2013), which act in a multimodal way as a serotonin agonist, antagonist and reuptake inhibitor while also affecting other neurotransmitter systems. An intranasal formulation of esketamine (van de Loo et  al. 2017), a rapid-­ acting, anti-­ depressant with NMDA antagonistic properties, was also found not to impair driving performance 8 hours after administration. Impairing effects were absent after single as well as repeated doses of these drugs. The only exception to this was a repeated and high dose of nefazodone, which did impair driving performance, but this impairment was less than the effect of 0.5 g/L BAC. Vortioxetine belongs to a new chemical class of psychotropics, the bis-­aryl-­sulfanyl amines, which possess unique properties and are structurally different from all currently known psychotropics. in  vitro studies indicate that vortioxetine is a 5-­HT3, 5-­HT7 and 5-­HT1D receptor antagonist, 5 HT1B receptor partial agonist, 5-­HT1A receptor agonist and inhibitor of the 5-­HT transporter. All of these activities are considered to be of potential clinical relevance and may be involved in the therapeutic mechanism of action of vortioxetine.

TCAs, mianserin and mirtazapine clearly impaired driving performance, while no driving impairment was found for the other classes of anti-­depressants. Nocturnal administration of sedating anti-­depressants is an effective method to reduce the impairing effects. Prolonged treatment with the sedating anti-­ depressants showed a decrease in impairment after 1 or 2 weeks, indicating the development of tolerance. In depressed patients, however, long-­ term anti-­ depressant treatment (including venlafaxine and paroxetine) resulted in poorer driving performance as compared to controls (Wingen et  al. 2006). Nevertheless, it has also been demonstrated that driving impairment is reduced in these long-­ term treated patients compared to untreated patients (van der Sluiszen et  al. 2017). Another study in patients using sedating anti-­ depressants (amitriptyline or mirtazapine) found no impaired driving compared to healthy controls, except for patients who had been using anti-­depressants for less than 3 years (van der Sluiszen et al. 2020). Special precautions have to be taken when co-­medication with anti-­depressants takes place. For instance, a study in depressed patients demonstrated that non-­sedating anti-­depressants can cause sedation when given in combination with benzodiazepines. Subjects received fluoxetine or moclobemide, while about 80% of the patients were chronically using benzodiazepines as co-­ medication. An increase in SDLP was demonstrated after the first week of treatment for a sub-­group of patients. These patients were using benzodiazepines that are usually metabolised by enzymes that were now being blocked by the anti-­depressants. It is, therefore, essential to make sure that patients’ co-­medication is compatible with their anti-­depressant. Variability in drug response might also be due to differences in metabolising rate. For instance, esmirtazapine is metabolised through CYP2D6, and its clearance is twice as low in poor metabolisers as compared with extensive metabolisers. A recent study showed a more pronounced driving impairment in poor metabolisers after treatment with esmirtazapine (Ramaekers et  al. 2011). This suggests that subjects with genetic CYP deviations (e.g. poor CYP2D6 metabolisers) may be at higher risk when exposed to higher levels of drugs or metabolites that have the potential to impair driving (Ramaekers et al. 2011).

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performance. In acute doses, mianserine 10  mg tid caused driving impairment comparable to the effects of a BAC of 1.0 g/L (Louwerens and Brookhuis 1984; Ramaekers et al. 1992a; O’Hanlon et  al. 1998). After 1 and 2  weeks of treatment, the driving impairment was less but still significantly worse than placebo and was comparable to a BAC of more than 0.5  g/L (O’Hanlon et  al. 1998). Nocturnal administration of 15  mg mianserine did not impair driving performance the next morning or after 7 days of treatment. Even after dose increase to 30  mg (days 8–15), impairment was absent the morning after the first dose and at steady state (Ramaekers et  al. 1998). Nocturnal dose studies with mirtazapine 30  mg showed significantly impaired driving performance after a single dose (Ramaekers et al. 1998; Wingen et al. 2005). After dose increase to 45 and 60 mg (days 8–15), only a high dose of mirtazapine still impaired driving performance, indicating that tolerance was not complete (Ramaekers et al. 1998). Recently, the effects of mirtazapine were replicated with 30  mg of mirtazapine causing significant impairment 8 hours after a morning dose (van de Loo et al. 2017) and on the morning after an evening dosing (Theunissen et  al. 2013), but not after 2  weeks of treatment (Theunissen et al. 2013).

61.2.4  Anti-­histamines The group of anti-­ histamines includes chemically different compounds that all share the pharmacological capacity of preventing histamine from binding to its receptor and diminishing allergies’ symptoms. The elimination half-­life of anti-­histamines ranges from a few hours to days (Simons et al. 1987). In general, driving tests have been performed at maximal concentration of the drug, both after acute and sub-­chronic dosing. Often a distinction is made between first-­and second-­ generation anti-­histamines. First-­generation anti-­histamines

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Hydroxizine 50  mg acutely impaired driving performance, with an SDLP increase larger than the effect of a BAC of 0.8  g/L (Vuurman et al. 2007). After 8 days of treatment with hydroxizine 50  mg, driving performance was still impaired, but to a lesser extent (Conen et al. 2011). In a study with female subjects, driving was impaired after diphenhydramine 50  mg treatment (Ramaekers and O’Hanlon 1994). Two other studies demonstrated a lower SDLP increase in a mixed gender population (Verster et al. 2003; Vuurman et al. 2004), but the effects were still larger than a BAC of 0.5 g/L. After 4 days of treatment, the effect of 50 mg diphenhydramine once daily (od) was still significantly different from placebo (Verster et al. 2003). Clemastine 2 mg was studied as a once-­a-­day dose (Vuurman et al. 1994) as well as a twice-­a-­day dose (Vermeeren et al. 1998b). The one-­time administration resulted in the highest increase in SDLP, whereas the twice-­a-­day regimen showed a smaller but still significant driving impairment. After 4  days of treatment with clemastine 2  mg bid, the SDLP increase was still significantly larger than placebo (Vermeeren et al. 1998b). Triprolidine caused the largest driving impairment when it was administered in a 5 mg twice-­a-­day regimen (Riedel et al. 1990). After 4  days of treatment with triprolidine 5  mg bid, impaired driving performance was still present (Riedel et al. 1990; Volkerts et  al. 1992b). When administered as a controlled release formulation, a 10 mg od dose still impaired driving performance in a male population, but this effect disappeared after 4 days of treatment (Robbe and O’ Hanlon 1990). Dexchlorpheniramine 6 mg in a controlled release formulation significantly impaired driving performance acutely in two studies (Theunissen et al. 2004; Theunissen et al. 2006). In one study, the effects were larger than the effect of a BAC of 0.5 g/L, while the effect was less in the other study. After 8  days of treatment, dexchlorpheniramine’s effects on driving no longer differed from placebo (Theunissen et al. 2006).

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both doses of emedastine still significantly increased the SDLP (Vermeeren et al. 2002a). Mizolastine was tested in four different doses (5, 10, 20 and 40 mg) (Vuurman et al. 1994). The two lowest doses had moderate, non-­significant effects on driving performance, whereas the 20 and 40  mg doses significantly increased SDLP. This study also demonstrated that females reached the sedative threshold after lower doses of mizolastine compared with males. Acrivastine 8 mg tid a day did not lead to significant increases in SDLP in male subjects (Robbe and O’Hanlon 1990). In female subjects (Ramaekers and O’Hanlon  1994), significant dose-­ related SDLP increments were demonstrated for once-­ a-­ day treatment with 8, 16 and 24 mg doses. In one study, mequitazine demonstrated dose-­related driving impairment, although the separate doses (5, 10 and 15 mg) did not reach significance (Theunissen et al. 2004). In another study, driving performance was acutely impaired after mequitazine 10  mg treatment (Theunissen et  al. 2006). This effect was no longer significantly different from placebo after 8  days of treatment with the drug (Theunissen et al. 2006). A more recent study did not demonstrate acute driving impairment with mequitazine 10 mg (van der Sluiszen et al. 2016). In this study, however, the driving test was performed 1–2 hours after Tmax, indicating that the mequitazine effect might be short-­lived. Inconsistent findings have been found for cetirizine. Two studies (Ramaekers et al. 1992b; Vermeeren et al. 2002a) found cetirizine (10  mg od) to cause a significant SDLP increase, although in the latter this was only for the female participants. This effect was persistent after 4 days of treatment for the females but not for the whole group. Three other studies (Theunissen et al. 2004; Theunissen et al. 2006; Volkerts et al. 1992b), in male subjects and in a mixed population, found no acute or sub-­ chronic effects of cetirizine. Three studies confirmed that single doses of loratadine 10 and 20 mg did not impair driving performance (Riedel et al. 1989; Riedel et al. 1990; Ramaekers et al. 1992b). A repeated administration of 10  mg loratadine for 4  days did not increase SDLP (Riedel et  al. 1989), whereas 4 days of treatment with a 20 mg od dose almost significantly increased SDLP (Riedel et al. 1990). Some recently developed anti-­histamines are levocetirizine, rupatadine, bilastine and levomequitazine. A mixed-­gender study showed that levocetirizine 5 mg had no acute or sub-­chronic effect on SDLP (Verster et  al. 2003). The effects of rupatadine in its therapeutic dose of 10  mg (Vuurman et  al. 2007) were also comparable to those of placebo treatment. Bilastine was tested in 20 and 40 mg doses, both after acute dosing and repeated doses, and was found not to cause driving impairment in any of these dosing conditions (Conen et al. 2011). Levomequitazine, on the other hand, was found to impair driving acutely after a high dose (10 mg), but not after low doses (2.5 and 5 mg) (van der Sluiszen et al. 2016). Instead of causing impairment, a few anti-­histamines showed a decrease in SDLP, indicating a mild improvement in driving performance. This was true for terfenadine 60, 120 and 180 mg (Riedel et  al. 1989; Riedel et  al. 1990; Volkerts et  al. 1992b;

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readily cross the blood–brain barrier and bind to H1 receptors in the central nervous system (Quach et  al. 1979; Nicholson et  al. 1991), causing sedation. Second-­generation anti-­histamines have a reduced ability to cross the blood–brain barrier and therefore cause less sedation. Although they are less sedating than first-­ generation anti-­histamines, patients occasionally report sedative side effects when treated with second-­generation anti-­histamines, especially when the therapeutic dose is exceeded (Mann et  al. 2000; Casale et al. 2003). Therefore, most driving studies have also investigated larger than therapeutic doses of anti-­histamines. An overview of the effects of anti-­histamines on the on-­the-­ road driving test is shown in Table 61.4.

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Second-­generation anti-­histamines Emedastine 2 and 4  mg bid caused significant acute driving impairment 3 hours after drug intake. After 4 days of treatment,

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Over the years, many anti-­histamines have been developed, and an obvious improvement in the safety profile can be seen with the development of second-­generation anti-­histamines. Although second-­generation anti-­histamines are often called ‘non-­sedating anti-­histamines’, it should be clear that they are not free of sedative effects. Some of the more recent developed anti-­ histamines demonstrated no impairment (e.g. fexofenadine, desloratadine and levocetirizine) and are therefore sometimes called ‘third-­ generation anti-­histamines’. histamines, driving impairment was only For several anti-­ demonstrated in an exclusively female population, and a few studies also demonstrated women’s greater sensitivity to the impairing effects of anti-­histamines. These findings suggest that women might be more sensitive to the effects of anti-­histamines. It has been hypothesised that this greater sensitivity of women is a consequence of the lower average body weight of participants in the respective studies, although this has never been demonstrated. When drug administration was continued over several days, anti-­histamines’ effects on driving performance diminished but remained significant in most cases. Only the effect of ebastine changed from acutely stimulating to sub-­chronically sedating, probably as a consequence of the accumulation of the drug in the brain.

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Methylphenidate (Ritalin), a psychostimulant drug used for treating attention deficit hyperactivity disorder (ADHD) and narcolepsy, was found to improve driving performance in adult ADHD patients (Verster and Cox 2008) and 3,4-­methylenedioxy methamphetamine (MDMA; e.g. ecstasy) users (Ramaekers and Kuypers 2004; Ramaekers et al. 2006), as evidenced by a decrease in SDLP compared to placebo. The opioids methadone (5 and 10  mg) and buprenorphine (0.2  and 0.4  mg), which are used as analgesics and in opioid maintenance treatment, have recently been tested in healthy volunteers (Strand et al. 2019). While methadone did not impair driving performance, buprenorphine produced mild impairing effects in the high dose condition. In recent years, cannabis and synthetic cannabinoids developed for medical use and have become more prevalent in many countries due to changes in legislation. The effects of cannabis have previously been studied in recreational users of cannabis (Robbe 1998; Ramaekers et al. 2000). Low (100 μg/kg of THC) and medium (200 μg/kg of THC) doses significantly impaired driving performance in a dose-­related manner, with both doses causing comparable effects to alcohol 0.5 g/L (Ramaekers et al. 2000). The effects of oral doses of dronabinol, a synthetic THC formulation approved for treating, i.e. nausea in chemotherapy, was studied in occasional and regular healthy cannabis users (Bosker et al. 2012). Both doses of 10 and 20 mg significantly impaired driving performance in a dose-­ related manner. Performance impaired both groups of users significantly, although to a lesser extent in regular users, possibly due to tolerance development.

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Ramaekers and O’Hanlon 1994), ebastine 10, 20 and 30 mg (De Vries et al. 1989; O’Hanlon and Ramaekers 1995), desloratadine 5  mg (Vuurman et  al. 2004) and fexofenadine 120  mg bid (Vermeeren and O’Hanlon 1998). When the administration was prolonged, terfenadine 120 mg bid did cause a significant rise in SDLP when driving was performed 2 hours after intake, but this effect disappeared again 2 hours later (Riedel et al. 1990). Also for ebastine 30 mg, an impairing effect was demonstrated after 5 days of treatment (O’Hanlon and Ramaekers 1995).

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The anti-­epileptic drugs carbamazepine and remacemide were compared in one driving study using healthy volunteers (Ramaekers et  al. 2002). The effects on driving performance were measured after 8, 10 and 12 days of a 12-­day incremental dosing regimen. Remacemide did not affect the subjects’ driving performance. Carbamazepine increased SDLP throughout treatment compared to those of drivers with BACs of 0.5 g/L. One study looked at the effects of mefloquine, an anti-­malarial drug, on actual driving performance (Vuurman et al. 1996). The medication was taken in a 250 mg dose during the evenings of days 1, 2, 3, 8, 15, 22 and 29. Testing was performed on days 4, 23 and 30, the latter after repeated doses of alcohol sufficient to sustain a blood concentration of about 0.35  g/L. Mefloquine caused no significant impairment in any test at any time relative to placebo. It significantly improved driving performance in the on-­the-­road driving test on day 4.

61.3 Conclusions This chapter gives an overview of the studies using the on-­the-­ road driving test for measuring the impairing effects of psychoactive prescription drugs on driving performance. Driving impairment has been demonstrated for several hypnotics, anxiolytics, anti-­depressants and anti-­histamines. Within each drug class, large differences in impairing potential have been shown. To ensure traffic safety, governments apply strict laws to driving under the influence of alcohol. Applying such laws for medicinal drugs is, however, not that simple because of various reasons. For example, a large proportion of the population would be excluded from driving. In addition, relatively reliable and straightforward breath analyser tests are available for estimating the amount of alcohol in the blood circulation, but such easy tests do not exist for measuring circulating levels of psychoactive substances. Therefore, it is essential that drugs are screened thoroughly on their potential to affect patients’ driving performance and that patients receive the most up-­ to-­ date information about the impairing potential of the drugs they are using. For this reason, the International Council on Alcohol, Drugs and Traffic Safety (ICADTS) has provided a categorisation

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Bosker, W., Kuypers, K., Theunissen, E. et  al. (2012). Medicinal THC (dronabinol) impairs on-­the-­road driving performance of occasional and heavy cannabis users but is not detected in Standardized Field Sobriety Tests. Addiction 107 (10): 1837–1844. Brookhuis, K.A. and Borgman, A.E. (1988). The effects of some anxiolytics on driving performance. Journal for Drugtherapy and Research 13: 228–231. Brookhuis, K.A., Volkerts, E.R. and O’Hanlon, J.F. (1990). Repeated dose effects of lormetazepam and flurazepam upon driving performance. European Journal of Clinical Pharmacology 39 (1): 83–87. Casale, T.B., Blaiss, M.S., Gelfand, E. et  al. (2003). First do no harm: managing antihistamine impairment in patients with allergic rhinitis. Journal of Allergy and Clinical Immunology 111 (5): S835–842. Conen, S., Theunissen, E.L., Oers, A.C.M. et  al. (2011). Acute and subchronic effects of bilastine (20 and 40  mg) and hydroxyzine (50 mg) on actual driving performance in healthy volunteers. Journal of psychopharmacology 25 (11): 1517–1523. De Vries, G., de Waard, D. and Brookhuis, K.A. (1989). A Double Blind Study to Compare the Acute and Subchronic Effects of Ebastine 10, 20, 30  mg o.d., Triprolidine 10  mg o.d. and Placebo on Car Driving Performance. Groningen Drake, C.L., Roehrs, T., Roth, T. et  al. (2003). Insomnia causes, consequences, and therapeutics: An overview. Depression & Anxiety 18 (4): 163–176. European Monitoring Centre for Drugs and Drug Addiction. (2012). Driving Under the Influence of Drugs, Alcohol and Medicines in Europe —­Findings from the DRUID Project. Luxembourg. European Monitoring Centre for Drugs and Drug Addiction. (2014). Drug Use, Impaired Driving and Traffic Accidents. P. O. o. t. E. Union: Luxembourg. Jongen, S., Vuurman, E., Ramaekers, J. and Vermeeren, A. (2018). Comparing the effects of oxazepam and diazepam in actual highway driving and neurocognitive test performance: a validation study. Psychopharmacology 235 (4): 1283–1294. Leufkens, T., Ramaekers, J.G., De Weerd, A. et al. (2014a). On-­the-­road driving performance and driving-­related skills in older untreated insomnia patients and chronic users of hypnotics. Psychopharmacology 231 (14): 2851–2865. Leufkens, T., Ramaekers, J.G., De Weerd, A. et  al. (2014b). Residual effects of zopiclone 7.5  mg on highway driving performance in insomnia patients and healthy controls: a placebo controlled crossover study. Psychopharmacology 231 (14): 2785–2798. Leufkens, T.R. and Vermeeren, A. (2014). Zopiclone’s residual effects on actual driving performance in a standardized test: A pooled analysis of

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age and sex effects in 4 placebo-­controlled studies. Clinical therapeutics 36 (1): 141–150. Leufkens, T.R., Vermeeren, A., Leufkens, T.R.M. and Vermeeren, A. (2009a). Highway driving in the elderly the morning after bedtime use of hypnotics: a comparison between temazepam 20  mg, zopiclone 7.5  mg, and placebo. Journal of Clinical Psychopharmacology 29 (5): 432–438. Leufkens, T.R.M., Lund, J.S. and Vermeeren, A. (2009b). Highway driving performance and cognitive functioning the morning after bedtime and middle-­of-­the-­night use of gaboxadol, zopiclone and zolpidem. Journal of Sleep Research 18 (4): 387–396. Leufkens, T.R.M., Vermeeren, A., Smink, B.E. et  al. (2007). Cognitive, psychomotor and actual driving performance in healthy volunteers after immediate and extended release formulations of alprazolam 1 mg. Psychopharmacology 191 (4): 951–959. Louwerens, J.W. and Brookhuis, K.A. (1984). The Effects of the Antidepressants Oxaprotiline, Mianserin, Amitryptiline and Doxepin upon Actual Driving Performance (Report VK 83-­05). Groningen. Louwerens, J.W., Gloerich, A.B.M., de Vries, G. et al. (1987). The Relationship between Drivers’ Blood Alcohol Concentration (BAC) and Actual Driving Performance during High Speed Travel. International Congres on Alcohol, Drugs and Traffic Safety, T86, Amsterdam, Exerpta Medica. Mann, R.D., Pearce, G.L., Dunn, N. and Shakir, S. (2000). Sedation with “non-­sedating” antihistamines: Four prescription-­event monitoring studies in general practice. British Medical Journal 320 (7243): 1184–1186. Mets, M.A., de Vries, J.M., de Senerpont Domis, L.M. et al. (2011). Next-­ day effects of ramelteon (8 mg), zopiclone (7.5 mg), and placebo on highway driving performance, memory functioning, psychomotor performance, and mood in healthy adult subjects. Sleep 34 (10): 1327–1334. Nicholson, A.N., Pascoe, P.A., Turner, C. et  al. (1991). Sedation and histamine H1-­ receptor antagonism: Studies in man with the enantiomers of chlorpheniramine and dimethindene. British Journal of Pharmacology 104 (1): 270–276. O’Hanlon, J.F. (1984). Driving performance under the influence of drugs: rationale for, and application of, a new test. British Journal of Clinical Pharmacology 18 (Suppl. 1): 121S–129S. O’Hanlon, J.F., Haak, T.W., Blaauw, G.J. and Riemersma, J.B. (1982). Diazepam impairs lateral position control in highway driving. Science 217 (4554): 79–81. O’Hanlon, J.F. and Ramaekers, J.G. (1995). Antihistamine effects on actual driving performance in a standard test: a summary of Dutch experience, 1989-­94. Allergy 50 (3): 234–342. O’Hanlon, J.F., Robbe, H.W., Vermeeren, A. et al. (1998). Venlafaxine’s effects on healthy volunteers’ driving, psychomotor, and vigilance performance during 15-­day fixed and incremental dosing regimens. Journal of Clinical Psychopharmacology 18 (3): 212–221. O’Hanlon, J.F., Vermeeren, A., Uiterwijk, M.M. et al. (1995). Anxiolytics’ effects on the actual driving performance of patients and healthy volunteers in a standardized test. An integration of three studies. Neuropsychobiology 31 (2): 81–88. O’Hanlon, J.F. and Volkerts, E.R. (1986). Hypnotics and actual driving performance. Acta Psychiatrica Scandinavica, Supplementum 332: 95–104.

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system for medicinal drugs affecting driving performance, in which categories I, II and III are used and are equivalent to a BAC of 0.8  g/L. Using this system, prescribing doctors and dispensing pharmacists can select the safest drug within a class and warn patients about the possible risks associated with the drug.

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Quach, T.T., Duchemin, A.M., Rose, C. and Schwartz, J.C. (1979). in vivo occupation of cerebral histamine H1-­receptors evaluated with 3H-­ mepyramine may predict sedative properties of psychotropic drugs. European Journal of Pharmacology 60 (4): 391–392. Ramaekers, G., Lamers, J., Verhey, F. et al. (2002). A comparative study of the effects of carbamazepine and the NMDA receptor antagonist following performance in remacemide on road tracking and car-­ actual traffic. Psychopharmacology (Berl) 159 (2): 203–210. Ramaekers, J.G. (2003). Antidepressants and driver impairment: the-­ road test. Journal of empirical evidence from a standard on-­ Clinical Psychiatry 64 (1): 20–29. Ramaekers, J.G., Conen, S., de Kam, P.J. et  al. (2011). Residual effects of  esmirtazapine on actual driving performance: Overall findings and  an exploratory analysis into the role of CYP2D6 phenotype. Psychopharmacology 215 (2): 321–332. Ramaekers, J.G., Kuypers, K.P. and Samyn, N. (2006). Stimulant effects methylenedioxymethamphetamine (MDMA) 75  mg and of 3,4-­ methylphenidate 20  mg on actual driving during intoxication and withdrawal. Addiction Abingdon, England 101 (11): 1614–1621. Ramaekers, J.G. and Kuypers, K.P.C. (2004). A Placebo Controlled Study on the Effects of 3,4-­Methylenedioxymethamphetamine (MDMA) 75mg and Methylphenidate 20mg on Actual Driving Performance, Visuospatial Attention and Memory during Intoxication and Withdrawal. Maastricht Ramaekers, J.G., Muntjewerff, N.D. and O’Hanlon, J.F. (1995). A comparative study of acute and subchronic effects of dothiepin, fluoxetine and placebo on psychomotor and actual driving performance. British Journal of Clinical Pharmacology 39 (4): 397–404. Ramaekers, J.G., Muntjewerff, N.D., Van Veggel, L.M.A. et  al. (1998). Effects of nocturnal doses of mirtazapine and mianserin on sleep and on daytime psychomotor and driving performance in young, healthy volunteers. Human Psychopharmacology Clinical and Experimental 13 (Suppl. 2): S87–S97. Ramaekers, J.G. and O’Hanlon, J.F. (1994). Acrivastine, terfenadine and diphenhydramine effects on driving performance as a function of dose and time after dosing. European Journal of Clinical Pharmacology 47 (3): 261–266. Ramaekers, J.G., Robbe, H.W.J. and O’Hanlon, J.F. (2000). Marijuana, alcohol and actual driving performance. Human Psychopharmacology: Clinical and Experimental 15 (7): 551–558. Ramaekers, J.G., Swijgman, H.F. and O’Hanlon, J.F. (1992a). Effects of  moclobemide and mianserin on highway driving, psychometric performance and subjective parameters, relative to placebo. Psychopharmacology 106 Suppl: S62–67. Ramaekers, J.G., Uiterwijk, M.M. and O’Hanlon, J.F. (1992b). Effects of loratadine and cetirizine on actual driving and psychometric test performance, and EEG during driving. European Journal of Clinical Pharmacology 42 (4): 363–369. Ramaekers, J.G., van Veggel, L.M. and O’Hanlon, J.F. (1994). A cross-­ study comparison of the effects of moclobemide and brofaromine on actual driving performance and estimated sleep. Clinical Neuropharmacology 17 (Suppl. 1): S9–18. Riedel, W.J., Quasten, R., Hausen, C. and O’Hanlon, J.F. (1988). A Study Comparing the Hypnotic Efficasies and Residual Effects on Actual Driving Performance of Midazolam 15 mg, Triazolam 0.5 mg, Temazepam 20 mg and Placebo in Shiftworkers on Night Duty. Maastricht.

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Vermeeren, A., Swijgman, H.F. and O’Hanlon, J.F. (1994). Acute and Subchronic Effects of Alpidem, Lorazepam, and Placebo on Anxiety, Psychometric Test Performance and Actual Driving in Anxious Patients. Maastricht. Vermeeren, A., Vets, E., Vuurman, E.F. et al. (2016). On-­the-­road driving performance the morning after bedtime use of suvorexant 15 and 30 mg in healthy elderly. Psychopharmacology 233 (18): 3341–3351. Vermeeren, A., Vuurman, E.F., Leufkens, T.R. et  al. (2014). Residual effects of low-­ dose sublingual zolpidem on highway driving performance the morning after middle-­of-­the-­night use. Sleep 37 (3): 489–496. Verster, J.C. and Cox, D.J. (2008). ADHD, methylphenidate and driving:  does some legislation endanger public health? Journal of Psychopharmacology 22 (3): 227–229. Verster, J.C., Veldhuijzen, D.S. and Volkerts, E.R. (2004). Residual effects of sleep medication on driving ability. Sleep Medicine Reviews 8 (4): 309–325. Verster, J.C., Volkerts, E.R., Schreuder, A.H.C.M.L. et al. (2002a). Residual effects of middle-­of-­the-­night administration of zaleplon and zolpidem on driving ability, memory functions, and psychomotor performance. Journal of Clinical Psychopharmacology 22 (6): 576–583. Verster, J.C., Volkerts, E.R., van Oosterwijck, A.W. et al. (2003). Acute and subchronic effects of levocetirizine and diphenhydramine on memory functioning, psychomotor performance, and mood. Journal of Allergy and Clinical Immunology 111 (3): 623–627. Verster, J.C., Volkerts, E.R. and Verbaten, M.N. (2002b). Effects of alprazolam on driving ability, memory functioning and psychomotor performance: A randomized, placebo-­ controlled study.″ Neuropsychopharmacology official publication of the American College of Neuropsychopharmacology 27 (2): 260–269. Vinckenbosch, F., Vermeeren, A., Verster, J. et al. (2020). Validating lane drifts as a predictive measure of drug or sleepiness induced driving impairment. Psychopharmacology 237 (3): 877–886. Volkerts, E.R., Brookhuis, K.A. and O’Hanlon, J.F. (1987). Comparison of the Effects of Buspirone 5 mg and 10 mg, Diazepam 5 mg, and Lorazepam 1 mg (t.i.d.) upon Actual Driving Performance (VK 87-­02). Haren. Volkerts, E.R., Louwerens, J.W., Gloerich, A.B.M. et al. (1984). Zopiclone’s residual effect upon actual driving performance versus those of nitrazepam and flunitrazepam. VSC, Report 84-­ 10. Groningen, TrafficResearchCentre. Volkerts, E.R., van Laar, M.W. and van Willigenburg, A.P.P. (1992a). A comparative study of on-­ the-­ road driving and simulated driving performance after nocturnal treatment with lormetazepam 1 mg and oxazepam 50  mg. Human Psychopharmacology: Clinical and Experimental 7: 297–309. Volkerts, E.R., Van Willigenburg, A.P., Van Laar, M.W. and Maes, R.A. (1992b). Does cetirizine belong to the new generation of antihistamines? An investigation into its acute and subchronic effects on highway driving, psychometric test performance and daytime sleepiness. Human Psychopharmacology Clinical and Experimental 7 (4): 227–238. Vuurman, E.F., Muntjewerff, N.D., Uiterwijk, M.M. et al. (1996). Effects of mefloquine alone and with alcohol on psychomotor and driving performance. European Journal of Clinical Pharmacology 50 (6): 475–482.

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sedating antidepressants. Human Psychopharmacology: Clinical and Experimental: e2762. van der Sluiszen, N.N., Vermeeren, A., Verster, J.C. et al. (2019). Driving performance and neurocognitive skills of long-­ term users of benzodiazepine anxiolytics and hypnotics. Human Psychopharmacology: Clinical and Experimental 34 (6): e2715. van der Sluiszen, N.N., Wingen, M., Vermeeren, A. et al. (2017). Driving performance of depressed patients who are untreated or receive long-­ term antidepressant (SSRI/SNRI) treatment. Pharmacopsychiatry 50 (05): 182–188. van Laar, M., Volkerts, E. and Verbaten, M. (2001). Subchronic effects of the GABA-­agonist lorazepam and the 5-­HT2A/2C antagonist ritanserin on driving performance, slow wave sleep and daytime sleepiness in healthy volunteers. Psychopharmacology 154 (2): 189–197. van Laar, M.W., van Willigenburg, A.P. and Volkerts, E.R. (1995). Acute and subchronic effects of nefazodone and imipramine on highway driving, cognitive functions, and daytime sleepiness in healthy adult and elderly subjects. Journal of Clinical Psychopharmacology 15 (1): 30–40. van Laar, M.W., Volkerts, E.R. and van Willigenburg, A.P. (1992). Therapeutic effects and effects on actual driving performance of chronically administered buspirone and diazepam in anxious outpatients. Journal of Clinical Psychopharmacology 12 (2): 86–95. Van Veggel, L.M. and O’Hanlon, J.F. (1993). Effects of Ondansetron and Diazepam on Driving and Psychometric Performance in Healthy Volunteers (IGVG-­P26). Maastricht. Vermeeren, A., Danjou, P.E. and O’Hanlon, J.F. (1998a). Residual effects of evening and middle-­of-­the-­night administration of zaleplon 10 and 20  mg on memory and actual driving performance. Human Psychopharmacology: Clinical and Experimental 13: S98–S107. Vermeeren, A., Jongen, S., Murphy, P. et al. (2019). On-­the-­road driving performance the morning after bedtime administration of lemborexant in healthy adult and elderly volunteers. Sleep 42 (4): zsy260. Vermeeren, A. and O’Hanlon, J.F. (1998). Fexofenadine’s effects, alone and with alcohol, on actual driving and psychomotor performance. Journal of Allergy and Clinical Immunology 101 (3): 306–311. Vermeeren, A., O’Hanlon, J.F., Declerck, A.C. and Kho, L. (1995). Acute effects of zolpidem and flunitrazepam on sleep, memory, and driving performance, compared to those of partial sleep deprivation and placebo. Acta Therapeutica 21: 47–64. Vermeeren, A., Ramaekers, J.G. and O’Hanlon, J.F. (2002a). Effects of emedastine and cetirizine, alone and with alcohol, on actual driving of males and females. Journal of Psychopharmacology 16 (1): 57–64. Vermeeren, A., Ramaekers, J.G., Van Leeuwen, C.J. and O’ Hanlon, J.F. (1998b). Residual effects on actual car driving of evening dosing of chlorpheniramine 8 and 12 mg when use with terfenadine 60 mg in the morning. Human Psychopharmacology Clinical and Experimental 13 (Suppl. 2): S79–S86. Vermeeren, A., Riedel, W.J., van Boxtel, M.P. et al. (2002b). Differential residual effects of zaleplon and zopiclone on actual driving: a comparison with a low dose of alcohol. Sleep 25 (2): 224–231. Vermeeren, A., Sun, H., Vuurman, E.F. et al. (2015). On-­the-­road driving performance the morning after bedtime use of suvorexant 20 and 40  mg: a study in non-­elderly healthy volunteers. Sleep 38 (11): 1803–1813.

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and psychomotor performance measurements. European Journal of Clinical Pharmacology 60 (5): 307–313. Wingen, M., Bothmer, J., Langer, S. et  al. (2005). Actual driving performance and psychomotor function in healthy subjects after acute and subchronic treatment with escitalopram, mirtazapine, and placebo: a crossover trial. Journal of Clinical Psychiatry 66 (4): 436–443. Wingen, M., Ramaekers, J.G., Schmitt, J.A. et  al. (2006). Driving impairment in depressed patients receiving long-­term antidepressant treatment. Psychopharmacology 188 (1): 84–91.

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Vuurman, E.F., Theunissen, E.L., Oers, A.C.M. et  al. (2007). Lack of effects between rupatadine 10mg and placebo on actual driving performance of healthy volunteers. Human Psychopharmacology 22: 289–297. Vuurman, E.F., Uiterwijk, M.M., Rosenzweig, P. and O’Hanlon, J.F. (1994). Effects of mizolastine and clemastine on actual driving and psychomotor performance in healthy volunteers. European Journal of Clinical Pharmacology 47 (3): 253–259. Vuurman, E.F.P.M., Rikken, G.H., Muntjewerff, N.D. et al. (2004). Effects of desloratadine, diphenhydramine, and placebo on driving performance

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Toxicological Markers of Chronic Alcohol Abuse

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Generally, there are two types of alcohol biomarkers: trait markers that characterise the genetic disposition and vulnerability to become addicted to alcohol, and state markers that characterise the real drinking behaviour of an individual.

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Alcohol is the most frequently misused substance worldwide, and alcohol dependence or excessive drinking without being addicted leads to immense harm in medical, social, legal and economic terms. From psychiatric point of view, alcohol use disorder (AUD) is a psychiatric disorder characterised by excessive and uncontrolled drinking that causes harm and distress, and has devastating consequences for men and women of all ages. It combines alcohol dependence and alcohol abuse. The diagnosis is based on criteria of the Diagnostic and Statistical Manual of Mental Disorders (DSM-­IV) of the American Psychiatric Association and on the International Classification of Diseases (ICD-­10) of the World Health Organization (Hasin 2003; Lago et al. 2016). Therefore, biomarkers for the detection of excessive alcohol consumption are a topic of ongoing research and have extensively been reviewed in several papers (Jones 2008; Nanau and Neumann 2015; Niemalä 2016; Andresen-­Streichert et al. 2018). Besides in traffic medicine, they are important in several other contexts of forensic medicine such as workplace testing, family law and child custody, probation conditions of offenders and decisions for liver transplantation. Furthermore, there are medical applications, e.g. in the control of alcohol withdrawal treatment or the detection of alcohol abuse during pregnancy. This chapter mainly addresses the use of alcohol markers related to drunk driving although the principles are the same in all other fields of application with only different assessment criteria.

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Handbook of Forensic Medicine, Volume 3, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

62.2  Trait markers and polygenic risk scores of alcohol use disorder

It follows from twin and adoption studies that alcohol use disorder is approximately 50% heritable with the other 50% caused by environmental factors (Verhulst et al. 2015). The immense progress in genotyping technologies and the formation of large genomic banks in the last years has enabled significant efforts to identify specific variations within the genome related to the aetiology of AUD. Using current methods, genome-­wide association studies (GWAS) can individually test for an association between a phenotype of interest and about 7 million variants (single nucleotide polymorphisms, SNPs) across the human genome. From such studies, it was demonstrated that not a limited number of specific polymorphisms but hundreds of variants across the genome, most of relatively small effect, contribute to the genetic aetiology of AUD (Deak et al. 2019). It means that AUD is a trait with a complex inheritance pattern and a high level of polygenicity including hundreds of variants across the genome with a minor effect size.

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dependence could also lead to discrimination and disadvantages in professional and private life. Therefore, ethical aspects will probably limit the benefits of a broad application of genetic alcohol trait markers.

62.3  State biomarkers of alcohol consumption

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62.3.1  Overview and characteristics of alcohol state markers

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An overview of state markers with a greater practical importance is given in Table 62.1. They can be classified with respect to their time window for the detection of acute or chronic consumption or with respect to their connection to ethanol for indirect or direct markers. Direct ethanol (referred to as alcohol from now on) markers contain the two carbon atoms of ethanol. Therefore, they can only originate from alcohol. These markers are ethanol itself and its minor metabolites ethyl glucuronide, ethyl sulphate, fatty acid ethyl esters, phosphatidyl ethanol and the combined marker of cocaine and alcohol use cocaethylene (structures shown in Figure 62.1). All other markers given in Table 62.1 are indirect markers that are based on biochemical or pathological changes caused by alcohol but are not structurally linked to ethanol. During the last decade, a shift from indirect markers to direct markers has occurred because of their higher specificity. Blood and urine as the traditional specimens and increasingly hair with its longer detection window are preferably used as sample materials for this purpose. The applicability of an alcohol consumption marker is characterised by the following criteria: • Biochemical mechanism: functional connection between alcohol consumption and the value of the parameter. • Practicability: availability of the test, difficulty and costs of performance. • Reference range: values for teetotallers, social drinkers and alcohol abuse. • Cut-­off values: decision limit for discrimination between the different consumer groups. • Threshold dose: tolerance of single drinking events. • Sensitivity (%): 100  minus the percentage of false-­negative results at a fixed cut-­off. • Specificity (%): 100  minus the percentage of false-­positive results at a fixed cut-­off. • Discrimination power: accuracy of discrimination between consumers groups. • Disturbances: sources of false-­ positive and false-­ negative results. Since these criteria depend to high degree on the investigated population, quite different values were obtained in the many studies described in the literature and only a qualitative rating can be given in Table  62.1. In the following sections, the most important biomarkers of alcohol consumption are described.

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Nevertheless, some specific gene loci associated with AUD were identified in such genome-­ wide studies by comparison between large numbers of individuals with alcohol dependence and controls (Hart and Kranzler 2015; Sanchez-­Roige et al. 2019; Walters et al. 2018; Thompson et al. 2020). This concerns between others, the ADH-­ALDH gene cluster on chromosome 4 that encodes the ethanol-­metabolising enzymes and the DRD2 locus on chromosome 11 that encodes the dopamine receptor 2 subtype, and is linked to several neurobiological processes, including functional activation of reward circuits. Furthermore, significant genetic correlations were observed with 17 phenotypes, including schizophrenia, attention deficit hyperactivity disorder (ADHD), depression, and the use of cigarettes and cannabis. A genetic distinction between pathological and non-­ pathological drinking behaviours was also seen. A more straightforward way to use genome-­wide association studies for the detection of alcohol use disorder is the development of polygenic risk scores (PRSs) as it is under investigation in other inherited complex disorders such as coronary artery disease, type 2 diabetes, Alzheimer’s disease or psychiatric disorders (Lewis and Vassos 2020). This score is statistically calculated for each individual as the weighted sum of the number of the relevant risk alleles which the individual carries. The outcome is a single score of each individual’s genetic loading for a disease or for a continuous trait. As an example, a Finnish group built an AUD polygenic risk score based on self-­reported alcohol consumption data, GGT-­ values and alcohol-­related morbidity and mortality of 39  695 participants including 4202  non-­ drinkers and the FinnGen genomic database (Kiiskinen et al. 2020). The number of SNPs in the final score was 1.1 million. The PRS was normalised and used as a continuous variable. It appeared to be strongly associated with alcohol consumption, and the increase in the PRS by 1 SD (standard deviation) was associated with the increase of the weekly alcohol consumption by 11.2 g. By comparing the lowest and highest quintiles of the score, a clear association was also seen for alcohol-­related morbidity events and all-­cause mortality. Its predictive capacity was studied in two cohorts of 5732 and 18 427 participants with detailed baseline data and ≥10 years of follow-­up time. It was shown to provide a genetic basis for potentially identifying a subset of high-­risk individuals even early on in life. From the present point of view, such polygenic risk scores seem to be the most suitable approach to alcohol trait markers in the future and superior to any other single parameter. However, until now no application in forensic medicine was described, and further research is necessary with larger cohorts of individuals with more reliable data about alcohol consumption and dependence as well as including other than European and North American populations. The early detection of a genetic vulnerability to alcohol use disorders can serve as a motivator for reducing drinking or adhering to abstinence and is a cost-­effective and efficient strategy to reduce alcohol-­related harms with clinical and forensic respect. However, the documented information about the genetic disposition to alcohol

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Table 62.1  Characteristics of alcohol biomarkers. Alcohol markera

Indication

Time window

Availabilityb

Thresholdc

Sensitivityd

Specificityd

EtOH in blood

Actual drinking

Hours to 1 d

A, C

0.1 g/kg

Not definede

100%

EtOH in breath

Actual drinking

Hours to 1 d

E

0.05 mg/L

Not defined

e

100%

EtOH in urine

Recent drinking

Hours to 1 d

A, C

0.1 g/kg

Not defined

e

EtOH in sweat

Abstinence control

Minutes to hours

F

Not defined

High

EtG in urine

Recent drinking

Up to 130 h

D

100 ng/mL

Not definede

High

EtS in urine

Recent drinking

Up to 80 h

D

Not defined

Not defined

High

5-­HTOL/5-­HIAA in urine

Recent drinking

6–20 h

C

15 nmol/μmol

Not definede

>90%

MeOH in blood

Chronic episodic drinking

Hours to 1 d

C

0.01 g/kg

Low

High

GGT in plasma, serum

Excessive drinking

2–6 wk

A

55–64 IU/L (male) 36–42 IU/L (female)

MCV in blood

Excessive drinking

Several months

A

98–100 fL

CDT in plasma, serum

Excessive drinking

3–4 wk

B

1.7-­2% of total transferrin

C D

EtG in hair

Excessive drinking

Several months

D

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2–3 wk Several months

200 ng/mL

Low-­moderate

Moderate

Low

Moderate

Moderate

High

High

High

0.35 ng/mg (0–3 cm)

High

High

30 pg/mg (0–3 cm)

High

High

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Excessive drinking Excessive drinking

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PEth 16:0/18:1 in blood EtPa in hair

100% 100%

N OH

O

CH3

CH2

R

O

CH2 CH3

C O CH2 CH3 O

H3C N

O

C O

H5C6

EtS

EtG

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O

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HOOC

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a  Abbreviations: CDT, carbohydrate deficient transferrin; EtG, ethyl glucuronide; EtOH, ethanol; EtS, ethyl sulphate; EtPa, ethyl palmitate; GGT, gamma-­ glutamyltransferase; 5-­HTOL/5-­HIAA, ratio of 5-­hydroxytryptophol and 5-­hydroxyindolylacetic acid; MCV, mean corpuscular volume of erythrocytes; MeOH, methanol; PEth, phosphatidylethanol. b  Test availability: A, low expenses, all clinical laboratories; B, increased expenses, central clinical laboratories; C, increased expenses, forensic alcohol laboratories; D, high expenses, specialised laboratories; E, on-­site testing, police, emergency units; F, pilot projects, individual application, continuous testing by wrist or ankle bracelet. c  Threshold values of some markers may depend on analytical method, age and gender, pregnancy or aim of diagnosis. d  Literature data about sensitivity and selectivity strongly depend on evaluated population and used decision criteria (cut-­offs). Therefore, only a qualitative assessment can be given. e  The sensitivity of detecting actual and recent alcohol drinking depends on alcohol dose and time after cessation of drinking and, therefore, cannot be defined. Source: Bortolotti et al. (2018), Brinkmann et al. (2000), Conigrave et al. (2003), Hannuksela et al. (2007), Niemelä (2016), Jones (2008), Liangpunsakul et al. (2010), Pirro et al. (2011), Pragst et al. (2017) and Ulwelling and Smith (2018).

Cocaethylene O O

CH2

OH

HO

CH3

O

FAEEs (example: ethyl palmitate)

N H

O O

O O

P

Serotonin degradation

O O

CH2 CH3

O

5-HIAA

NADH excess OH

HO

O

PhEt (example: PhEt 16:0/18:1)

N H

5-HTOL

Figure 62.1  Structure of some alcohol biomarkers: ethyl glucuronide (EtG), ethyl sulphate (EtS), cocaethylene (combined alcohol and cocaine metabolite), fatty acid ethyl esters (FAEEs, group of minor ethanol metabolites), phosphatidylethanol (PEth, group of minor ethanol metabolites), 5-­hydroxytryptophol (5-­HTOL) and 5-­hydroxyindolylacetic acid (5-­HIAA).

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Some indications of an alcohol problem are obtained from inexpensive clinical laboratory parameters that are routinely determined, for instance during a health check of a person. These are the liver enzymes γ-­glutamyltransferase (GGT), alanine transaminase (ALAT) and aspartate transaminase (ASAT) and the erythrocyte mean corpuscular volume (MCV) (Sillanaukee et al. 1998; Conigrave et  al.  2003; Hietala et  al.  2006; Liangpunsakul et al. 2010). They are indirect alcohol markers. Of these, GGT is the most frequently used parameter to get first information about heavy drinking as well as for monitoring changes in following up during withdrawal treatment. This liver enzyme catalyses the transfer of the γ-­glutamyl moiety from glutathione to various small proteins. Elevated values in serum occur after daily drinking of 60–80 g ethanol over at least 3 weeks by leakage from damaged liver cells or by enzyme induction; 4–5  weeks of abstinence are required before their return to normal values. False-­positive results are obtained by the presence of hepatitis, liver cirrhosis, liver carcinoma or several medicinal drugs. Reference ranges depend on age and gender and may be changed in cases of pregnancy, obesity or diabetes. Its diagnostic validity can be improved in combination with alcohol screening questionnaires such as AUDIT and CAGE (Choe et al. 2019). ALAT and ASAT are hepatic enzymes involved in the metabolism of the amino acids, alanine and aspartate. Their analysis is a part of a routine liver function test. Elevated values indicate liver dysfunction caused either by heavy drinking or by liver diseases. The sensitivity and specificity as biomarkers of heavy drinking are relatively low. In addition to the enzyme activities, a ratio of ASAT:ALAT of >2 can be used as additional evidence of liver dysfunction. The MCV of the erythrocytes is enlarged after continuous heavy drinking for 4–8 weeks. The specificity is higher than that of GGT, ALAT or ASAT but is limited also by non-­alcoholic liver diseases, coagulation disorders or folate deficiency. The sensitivity is very low. In a study with 6244 drivers involved in traffic accidents it failed to reveal individuals at risk who tend to drive in a condition of low-­ to-­moderate alcohol intoxication (Porpigli et al. 2019). Therefore, it should be used only in combination with other parameters. These traditional markers were generally found to have imperfect sensitivity and specificity. For instance, in a large, population-­based study of more than 8700 participants, these markers of heavy drinking, either alone or in combination, did not appear to be useful in identifying unhealthy drinking (Liangpunsakul et al. 2010). However, they have a clinical role in the detection of complications of drinking and co-­morbid conditions that may increase the risk of drinking.

­ rinkers, the tetrasialo isoform widely dominates. Continuous d consumption of more than 50–70 g alcohol per day for at least seven consecutive days leads to a disturbance of the hepatic glycosylation in the transferrin synthesis, and an increased portion of isoforms with only two or even without any sialo groups is formed. The ratio of this carbohydrate-­ deficient transferrin (CDT) to the total transferrin is used as a marker of alcohol abuse (Arndt et al. 2008; Helander et al. 2016; Bortolotti et al. 2018). The procedures for the determination of CDT are based on various analytical techniques (immunoassays, HPLC, capillary electrophoresis and nephelometry). They were at first differing in the definition of the analyte and used different reference intervals and cut-­off values. Therefore, in 2005 a scientific Working Group on Standardization of CDT (WG-­CDT) was founded by the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) with the aim of harmonising and standardising CDT measurement and interpretation. An HPLC candidate reference measurement procedure was validated for CDT (ratio of disialotransferrin to total transferrin) based on peak areas (Schellenberg et  al.  2017). The performance of this reference method with previously developed commutable calibrators allows the standardisation of the currently available commercial measurement procedures for CDT. Nevertheless, at present, the term CDT still includes two transferrin fractions, asialotransferrin and disialotransferrin, both of them causally and directly related to excessive alcohol consumption in a dose-­related manner, as reported in a review paper summarising the actions taken by this working group (Helander et al. 2016).

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62.3.3  Carbohydrate-­deficient transferrin in serum or plasma Transferrin is a transport glycoprotein for iron ions (Fe3+). It contains two carbohydrate side chains with 2–6 terminal sialic acid groups for binding the iron. In healthy non-­and moderate



CDT %

100 disialo asialo / total transferrin



The analytical determination of CDT is more expensive than the parameters described in Section  62.3.2. Depending on the studied population and the diagnostic requirements, 1.7– 2.0% CDT is used as a cut-­off for heavy drinking. In the case of pregnancy, the CDT reference values increase progressively from the first to the third trimester (Bortolotti et  al.  2020). For alcoholics, the CDT is usually below 20% but may range up to 40% in rare cases. Although being an indirect alcohol marker, CDT is much more specific than, for instance, GGT. False-­positive results are rare and are caused, for example, by unusual genetic variants of the transferrin or by a rare congenital disorder of glycosylation. Published data about the sensitivity varies strongly from 90% depending on the evaluated drinker groups with highest probability of 50–70%. The detection window is 2–3  weeks of abstinence. Besides in serum, CDT can be measured in dried blood spots (Bertaso et al. 2016). In post-­mortem serum, CDT can be used as marker of pre-­mortem alcohol misuse up to 76  hours or even 152 hours after death (Rainio et  al.  2008). However, with increasing post-­mortem interval, haemolysis and protein degradation can lead to falsely high or low results. CDT is frequently used in comparison and in combination with the traditional alcohol markers (Sillanaukee et  al.  1998;

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Phosphatidylethanol (PEth) is a group of minor non-­oxidative metabolites of alcohol formed after alcohol ingestion from phospholipids by enzymatic replacement of choline or ethanolamine by ethanol (Viel et al. 2012; Hahn et al. 2016). The structure is shown in Figure  62.1. The reaction with ethanol is catalysed by phospholipase D, an ubiquitary enzyme, which normally hydrolyses phospholipids to phosphatidic acid. PEth is located in all cell membranes, for instance in blood cells, brain, kidney, liver, skeletal muscle and heart. But, different from other tissues, erythrocytes are not able to efficiently degrade PEth, probably due to the absence of phosphatidylcholine phospholipase C activity. For this reason, PEth accumulates in the membranes of red blood cells and is generally determined in whole blood. At present, the analytical determination is generally performed by liquid chromatography/tandem mass spectrometry (LC-­MS/MS). Up to 48 homologues of this group have been identified by LC-­MS/MS (Gnann et  al.,  2010). They differ by the fatty acid CO and R2-­ CO. The single homologues are substituents R1-­ commonly indicated as ‘PEth A:B/C:D’, where A and C are the number of carbons in the carboxylic sidechains and B and D are the number of double bonds in each side chain. For instance, PEth 16:0/18:1 carries the palmitoyl and the oleoyl residues (Figure  62.1). The concentrations of the homologues were determined in many studies and appeared to be in a typical ratio. As an example, in a study with 47 heavy drinkers, the range of the total PEth concentration was 0.82–11.7 (mean 6.35, median 5.88) μmol/L. PEth 16:0/18:1, 16:0/18:2 and 16:0/20:4 were found to be the predominant molecular species and accounted for on average ~42%, ~26% and ~9%, respectively, of total PEth in these samples without significant differences between male and female (Helander et al. 2019). Therefore, the use of PEth for the detection of alcohol abuse was restricted from the total amount to usually PEth 16:0/18:1 alone as the most sensitive homologue. Meanwhile, the increase in sensitivity of the LC-­ MS/MS methods enables the determination of PEth for social drinkers and even after a single alcohol intake with a blood-­ alcohol concentration of 1 g/kg with a detection time of 3–12  days (Schröck et al. 2017). Therefore, PEth in blood has a potential in abstinence monitoring, similar to EtG in urine (see Section 62.3.6). The study of the pharmacokinetics of the three homologues PEth 16:0/18:1,16:0/18:2 and 16:0/20:4 after a single oral ethanol dose of 0.4 and 0.8 g/kg led to half-­lives of 7.8 ± 3.3, 6.4 ± 5.0 and 2.1 ± 3 days, respectively (Hill-­Kapturczak et al. 2018; Lopez-­Cruzan et al. 2018). Similar results were obtained by following up these three homologues in blood during alcohol detoxification of 47 heavy drinkers with an about threefold spread in half-­live values

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62.3.4  Phosphatidylethanol in blood

(Helander et al. 2019). From this, it follows a detection window of about 3 weeks depending on the level of initial alcohol consumption and on individual factors (Stenton et al., 2019). PEth 16:0/18:1 concentrations were stable (defined as 0.1 indicate that the drug was used regularly in combination with high alcohol levels whereas CE/COC in the lower range should show that alcohol was present only occasionally or in low concentrations during cocaine use. Differences to other alcohol markers can be explained by non-­concurrent use of cocaine and alcohol, positive cocaine result caused by external contamination or different effects of hair care and cosmetics. Furthermore, the occasional presence of cocethylene in illicit cocaine that originates from the use of ethanol for the purification of crude cocaine can lead to misinterpretation concerning alcohol consumption (Casale et al. 2008).



Sensitivity 1 false negatives / true negatives



Specificity 1 false positive / true positives

62.5  Control of abstinence Total abstinence, for example for 1 year, after withdrawal treatment from alcohol addiction can be required as a prerequisite of driving licence re-­granting or for other reasons such as obtaining or maintaining child custody, after domestic violence offences, staying in jobs with high endangering potential or allowing licensed professionals with an alcohol problem to continue practicing without jeopardising community safety. This must be controlled since the probability of relapse is very high. The following two approaches are practically applied: 1. Frequent and at random measurement of alcohol markers with a short time window. This concerns ethanol in the breath, blood, urine or saliva, or EtG in the urine with the time

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Figure 62.2  Evaluation of the discrimination power of alcohol biomarkers by receiver operation characteristic (ROC) analysis. (a) 125 subjects, area under curves (AUCs): ALAT 0.73, ASAT 0.79, GGT 0.79, CDT 0.84, EtG in hair 0.99. Source: Kharbouche et al. (2012). (b) 341 subjects, AUCs: MeOH 0.728, A+2P 0.878, GGT 0.903, CDT 0.963. Source: Brinkmann et al. (2000). (c) 175 subjects, AUCs: CDT 0.771, EtG in hair 0.982; not shown: GGT 0.834, MCV 0.730, ALAT 0.728, ASAT 0.612. Source: Pirro et al. (2011). (d) 261 subjects, AUC: EtG in hair 0.954; 229 subjects, AUC: FAEE in hair 0.955. Source: Suesse et al. (2012). Assessment of discrimination power of the tests (Greiner et al. 2000): AUC = 0.5, non-­informative; 0.5 < AUC ≤ 0.7, less accurate; 0.7 < AUC ≤ 0.9, moderate accurate; 0.9 < AUC < 1.0, highly accurate; AUC = 1.0, perfect. A + 2P, acetone + isopropanol; ALAT, alanine transaminase; ASAT, aspartate transaminase; CDT, carbohydrate deficient transferrin; EtG, ethyl glucuronide; FAEE, fatty acid ethyl ester; GGT, γ-­glutamyltransferase; MCV, mean corpuscular volume of erythrocytes; MeOH, methanol.

windows being up to 3  days longer than that of ethanol. In Germany, EtG in urine (cut-­off 100 ng/mL) as well as in hair (cut-­off 7 pg/mg) has become of broader importance for traffic cases (Schubert et al. 2013). The advantage of EtG in urine is a higher sensitivity to single drinking events if the time window is met. However, complete supervision is not possible and occasional drinking cannot be excluded. In future, PEth in blood could gain importance because of the improved detectability by LC-­MS/MS, its longer detection window and its lower vulnerability to manipulation as compared to urine (cf. Section 62.3.4). 2. Measurement of alcohol markers with a longer time window (such as GGT, CDT or particularly EtG in hair) at the

beginning, repeatedly within, and at the end of the control period. Termination of drinking is indicated by a corresponding change of the values. Since these markers are not able to detect single or even occasional drinking, this is a way to detect heavy and longer lasting relapses rather than a real abstinence control. Nevertheless, it has proved to be a more or less successful method since persons controlled for abstinence usually relapse with more than a single drink. In Germany, EtG in hair is regularly used for this purpose with the cut-­off 7 pg/mg. The superiority of hair as the sample material was shown in a study where only 92 out of 4248 urine samples (2%) were tested positive, whereas 81 out of 386 hair samples (21%) were positive

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Alladio, E., Giacomelli, L., Biosa, G. et al. (2018). Development and validation of a partial least squares-­ discriminant Analysis (PLS-­ DA) model based on the determination of ethyl glucuronide (EtG) and fatty acid ethyl esters (FAEEs) in hair for the diagnosis of chronic alcohol abuse. Forensic Science International 282: 221–30. Alonso-­Martín, C., Tejedor-­Tejada, J., Valenzuela, E.F. et  al. (2020). Utility of ethyl-­glucuronide for evaluation of abstinence in patients presenting for liver transplantation. Transplantation Proceedings 52: 1450–2. Ammann, D., Becker, R. Kohl, A., Hänisch, J., and Nehls, I. (2014). Degradation of the ethyl glucuronide content in hair by hydrogen peroxide and a non-­destructive assay for oxidative hair treatment using infra-­ red spectroscopy. Forensic Science International 244: 30–5. Andraus, M., Apppenzeller, B., Baumgartner, M. et  al. (2019). 2019 Consensus for the use of alcohol markers in hair for supporting the assessment of abstinence and chronic alcohol consumption. https:// soht.org/images/pdf/Revision_2019_Alcoholmarkers.pdf. Assessed 24.10.2020. Andresen-­Streichert, H., Müller, A., Glahn, A. et  al. (2018). Alcohol biomarkers in clinical and forensic contexts. Deutsches Ärzteblatt International 115: 309–15. Andresen-­Streichert, H., Beres, Y., Weinmann W. et al. (2017). Improved detection of alcohol consumption using the novel marker phosphatidylethanol in the transplant setting: results of a prospective study. Transplant International 2017; 30: 611–20. Arndt, T., Guessregen, B., Hallermann, D. et al. (2008). Forensic analysis of carbohydrate-­deficient transferrin (CDT) by HPLC – statistics and extreme CDT values. Forensic Science International 175: 27–30. Arndt, T., Grüner, J., Schröfel, S. and Stemmerich, K. (2012). False-­ positive ethyl glucuronide immunoassay screening caused by a propyl alcohol-­ based hand sanitizer. Forensic Science International 223: 359–63. Arndt, T., Schröfel, S. and Stemmerich, K. (2013). Ethyl glucuronide identified in commercial hair tonics. Forensic Science International 231: 195–8. Arndt, T., Schröfel, S., Güssregen, B. and Stemmerich, K. (2014). Inhalation but not transdermal resorption of hand sanitizer ethanol causes positive ethyl glucuronide findings in urine. Forensic Science International 237: 126–30. Aradottir, S., Seidl, S., Wurst, F.M., Jonsson, B.A.G. and Alling, C. (2004). Phosphatidylethanol in human organs and blood: a study on autopsy material and influences by storage conditions. Alcoholism: Clinical and Experimental Research, 28: 1718–23. Auwärter, V., Sporkert, F., Hartwig, S. et al. (2001). Fatty acid ethyl esters in hair as markers of alcohol consumption. Segmental hair analysis of alcoholics, social drinkers, and teetotallers. Clinical Chemistry 47: 2114–23. Becker, R., Lô, I., Sporkert, F. and Baumgartner, M. (2018). The determination of ethyl glucuronide in hair: Experiences from nine consecutive interlaboratory comparison rounds. Forensic Science International 288: 67–71. Bertaso, A., Sorio, D., Vandoros, A. et  al. (2016). Use of finger-­prick dried blood spots (fpDBS) and capillary electrophoresis for carbohydrate deficient transferrin (CDT) screening in forensic toxicology. Electrophoresis 37: 2867–74.

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in the same time period (Agius et al. 2012). In Switzerland, the cut-­off value of 30 pg/mg in hair is used in order to control imposed ‘zero tolerance when driving’ (ZTD) which means that a driver is not allowed to drink any alcohol before driving (0.0 mg/g in blood, usually 0.5  mg/g) and to consume alcohol above the level of social drinking at any other time (Muskovich and Haag-­Dawoud 2012). An approach to a real abstinence control is continuous transdermal alcohol monitoring, using ankle or wrist bracelets (Hawthorne and Wojcik  2006; Alessi et  al.  2017). The Secure Continuous Remote Alcohol Monitor (SCRAM™) relies on the excretion of alcohol through sweat (about 1% of the ingested ethanol), either as sensible perspiration (sweat in the liquid phase) or insensible perspiration (constant, unnoticeable sweat in the vapour phase). The device is a tamper-­and water-­resistant bracelet, containing an electrochemical sensor that is attached to the test person’s ankle using a durable strap. Every half hour, it draws a certain amount from an enclosed air volume above the test person’s skin into a chamber containing the electrochemical alcohol sensor and analyses it for ethanol. In addition, the temperature and a signal from an infrared obstruction sensor are measured as anti-­circumvention features. The results are stored on a flash memory stick and are transferred, for instance, every 24 hours wirelessly to a modem that is linked to the computer of the control agency. Studies with several hundred participants showed that this is a valid and reliable method of determining compliance with court-­ordered abstinence. Furthermore, it can be useful for obtaining more realistic consumption data in epidemiologic alcohol research (Greenfield et al. 2014). It can be expected that with further miniaturisation of the alcohol sensors this could be the preferred way of controlling absolute abstinence.

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Agius, R., Nadulski, T., Kahl, H.G. and Dufaux, B. (2012). Ethyl glucuronide in hair – A highly effective test for the monitoring of alcohol consumption. Forensic Science International 218: 10–14. Albermann, M.E., Musshoff, F. and Madea, B. (2011). Comparison of ethyl glucuronide (EtG) and fatty acid ethyl esters (FAEEs) concentrations in hair for testing abstinence. Analytical and Bioanalytical Chemistry 400: 175–81. Albermann, M.E., Musshoff, F., Aengenheister, L. and Madea B. (2012). Investigations on the influence of different grinding procedures on measured ethyl glucuronide concentrations in hair determined with an optimized and validated LC-­ MS/MS method. Analytical and Bioanalytical Chemistry 403: 769–76. Albermann, M.E., Madea, B. and Musshoff, F. (2014). A SPME-­GC/MS procedure for the determination of fatty acid ethyl esters in hair for confirmation of abstinence test results. Journal of Chromatographic Science 52: 955–60. Alessi, S.M., Barnett, N.P. and Petry, N.M. (2017). Experiences with SCRAMx alcohol monitoring technology in 100 alcohol treatment outpatients. Drug Alcohol Depend. 178: 417–24.

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misclassification of social drinkers. Alcoholism: Clinical and Experimental Research 40: 451–456. Fosen, J.T., Mørland, J. and Høiseth, G. (2020). The relation between ingested dose of ethanol and amount of ethyl glucuronide formed in blood. Journal of Analytical Toxicology. 31, 44: 861–3. Gareri, J., Appenzeller, B., Walasek, P. and Koren, G. (2011). Impact of hair-­care products on FAEE hair concentrations in substance abuse monitoring. Analytical and Bioanalytical Chemistry 400: 183–188. Gerbase, F.E., Tegner, M., Krutzmann, M.E. et  al. (2021). Blood phosphatidyl ethanol levels as a tool to detect alcohol misuse in trauma patients. Clinical Toxicology (Philadelphia), 59: 418–25. Gnann, H., Engelmann, C., Skopp G. et al. (2010). Identification of 48 ESI-­ MS/MS. homologues of phosphatidylethanol in blood by LC-­ Analytical and Bioanalytical Chemistry 396: 2415–2423. Greenfield, T.K., Jason Bond, J. and Kerr, W.C. (2014). Biomonitoring for improving alcohol consumption surveys  -­The new gold standard? Alcohol Research 36: 39–45. Greiner, M., Pfeiffer, D. and Schmidt, R.D. (2000). Principles, practical application of the receiver-­ operating characteristic analysis for diagnostic tests. Preventive Veterinary Medicine 45: 23–41. Gough, G., Heathers, L., Puckett, D. et al. (2015). The utility of commonly used laboratory tests to screen for excessive alcohol use in clinical practice. Alcoholism: Clinical and Experimental Research 39: 1493–1500. Haffner, H.Th., Banger, M., Graw, M. et al. (1997). The kinetics of methanol elimination in alcoholics and the influence of ethanol. Forensic Science International 89: 129–136. Hahn, J.A., Anton, R.F. and Javors, M.A. (2016). The formation, elimination, interpretation, and future research needs of phosphatidylethanol for research studies and clinical practice. Alcoholism: Clinical and Experimental Research 40: 2292–2295. Hannuksela, M.L., Liisanantti, M.K., Nissinen, A.E. and Savolainen, M.J. (2007). Biochemical markers of alcoholism. Clinical Chemistry and Laboratory Medicine 45: 953–961. Hart, A.B. and Kranzler, H.R. (2015). Alcohol dependence genetics: Lessons learned from genome-­wide association studies (GWAS) and post-­GWAS analyses. Alcoholism: Clinical and Experimental Research 39: 1312–1327. Hartwig, S., Auwärter, V. and Pragst, F. (2003a). Fatty Acid ethyl esters in scalp, pubic, axillary, beard and body hair as markers for alcohol misuse. Alcohol and Alcoholism 38: 163–167. Hartwig, S., Auwärter, V. and Pragst, F. (2003b). Effect of hair care and hair cosmetics on the consumption. Forensic Science International 131: 90–97. Hasin, D. (2003). Classification of alcohol use disorders. Alcohol Research & Health 27: 5–17. Hawthorne, J.S. and Wojcik, M.H. (2006). Transdermal alcohol measurement: a review of the literature. Journal of the Canadian Society of Forensic Science 39: 65–71. Helander, A., Beck, O. and Jones, A.W. (1996). Laboratory testing for recent alcohol consumption: comparison of ethanol, methanol, and 5-­hydroxytryptophol. Clinical Chemistry 42: 618–624. Helander, A. and Eriksson, C.J. (2002). Laboratory tests for acute alcohol consumption: results of the WHO/ISBRA study on state and trait markers of alcohol use and dependence. Alcoholism: Clinical and Experimental Research 26: 1070–1077.

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Biondi, A., Freni, F., Carelli, C., Moretti, M. and Morini, L (2019). Ethyl glucuronide hair testing: A review. Forensic Science International 300: 106–19. Boll, R., Johnson, T., Kaaks, R., Tilman Kühn, T. and Skopp, G. (2017). Is it possible to detect PEth 16:0/18:1 and PEth 18:1/18:1 in red blood cells after 20 years of storage in liquid nitrogen? International Journal of Legal Medicine 131: 1291–7. Bortolotti, F., Sorio, D., Bertaso, A. and Tagliaro, F. (2018). Analytical and diagnostic aspects of carbohydrate deficient transferrin (CDT): A critical review over years 2007-­2017. Journal of Pharmaceutical and Biomedical Analysis 147: 2–12. Bortolotti, F., Raffaelli, R., Di Simone, N., et al. (2020). CDT reference values for monitoring chronic alcohol abuse in pregnancy. Clinica Chimica Acta 507: 156–60. Borucki, K., Schreiner, R., Dierkes, J. et  al. (2005). Determination of recent ethanol intake with new markers: comparison of fatty acid ethyl esters in serum and of ethyl glucuronide and the ratio of 5-­ hydroxytryptophol to 5-­ hydroxy indole acetic acid in urine. Alcoholism: Clinical and Experimental Research 29: 781–7. Brzezinski, M.R., Abraham, T.L., Stone, C.L., Dean, R.A. and Bosron, W.F. (1994). Purification and characterization of a human liver cocaine carboxylesterase that catalyzes the production of benzoylecgonine and the formation of cocaethylene from alcohol and cocaine. Biochemical Pharmacology 48: 1747–55. Brinkmann, B., Köhler, H., Banaschak, S. et al. (2000). ROC analysis of alcoholism markers – 100% specificity. International Journal of Legal Medicine 113: 293–299. Buchholtz, U. (1993). Blood methanol as a marker of alcoholism. A diagnostic component within the scope of expert assessment of driving competence in alcoholic intoxication. Blutalkohol 30: 43–51. Casale, F.J., Boudreau, D.K. and Jones, L.M. (2008). Tropane ethyl esters in illicit cocaine: isolation, detection, and determination of new manufacturing by-­products from the clandestine purification of crude cocaine base with ethanol. Journal of Forensic Science 53: 661–667. Casati, S., Ravelli, A., Angeli, I., et al. (2021). PTCA (1-­H-­Pyrrole-­2,3,5-­tr icarboxylic acid) as a marker for oxidative hair treatment: Distribution, reports. Journal of gender aspects, correlation with EtG and self-­ Analytical Toxicology, 45: 513–20. Choe, Y.M., Lee, B,C., Choi, I.G. et al. (2019). Combination of the CAGE and serum gamma-­glutamyl transferase: an effective screening tool for alcohol use disorder and alcohol dependence. Neuropsychiatric Disease and Treatment 15: 1507–1515. Conigrave, K.M., Davies, P., Haber. P. and Whitfield, J.B. (2003). Traditional markers of excessive alcohol use. Addiction 98 (Suppl. 2): 31–43. Crunelle, C.L., Yegles, M., van Nuijs, A.L.N. et al. (2014). Hair ethyl glucuronide levels as a marker for alcohol use and abuse: a review of the current state of the art. Drug and Alcohol Dependence 134: 1–11. Crunelle, C.L., Yegles, M., De Doncker, M. et al. (2017). Hair ethyl glucuronide concentrations in teetotalers: should we re-­evaluate the lower cut-­off? Forensic Science International 274: 107–108. Deak, J.D., Miller, A.P. and Gizer, I.R. (2019). Genetics of alcohol use disorder: A review. Current Opinion in Psychology 27: 56–61. Fosen, J.T., Morini, L., Sempio, C. et al. (2016). Levels of hair ethyl glucuronide in patients with decreased kidney function: Possibility of

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Helander, A., Böttcher, M., Fehr, C. et  al. (2009). Detection times for urinary ethyl glucuronide and ethyl sulfate in heavy drinkers during alcohol detoxification. Alcohol and Alcoholism 44: 55–61. Helander, A., Wielders, J., Anton, R. et al. (2016). Standardisation and use of the alcohol biomarker carbohydrate-­ deficient transferrin (CDT). Clinica Chimica Acta 459: 19–24. Helander, A., Böttcher, M., Dahmen, N. and Beck, O. (2019). Elimination characteristics of the alcohol biomarker phosphatidylethanol (PEth) in blood during alcohol detoxification. Alcohol and Alcoholism 54: 251–257. Herbst, E.D., Harris, D.S., Everhart, E.T. et al. (2011). Cocaethylene formation following ethanol and cocaine administration by different routes. Experimental and Clinical Psychopharmacology 19: 95–104. Hietala, J., Koivistom, H., Anttila, P. and Niemelä, O. (2006). Comparison of the combined marker GGT-­CDT and the conventional laboratory markers of alcohol abuse in heavy drinkers, moderate drinkers and abstainers. Alcohol and Alcoholism 41: 528–533. Hill-­ Kapturczak, N., Dougherty, D.M., Roache, J.D. et  al. (2018). Differences in the synthesis and elimination of phosphatidylethanol 16:0/18:1 and 16:0/18:2 after acute doses of alcohol. Alcoholism: Clinical and Experimental Research 42: 851–860. Høiseth, G., Bernard, J.P., Stephanson, N. et  al. (2008). Comparison between the urinary alcohol markers EtG, EtS, and GTOL/5-­HIAA in a controlled drinking experiment. Alcohol and Alcoholism 43: 187–191. Høiseth, G., Morini, L., Polettini, A. et al. (2009). Blood kinetics of ethyl glucuronide and ethyl sulphate in heavy drinkers during alcohol detoxification. Forensic Science International 188: 52–56. Høiseth, G., Karinen, R., Christophersen, A. and Mørland, J. (2010). Practical use of ethyl glucuronide and ethyl sulfate in postmortem cases as markers of antemortem alcohol ingestion. International Journal of Legal Medicine 124: 143–148. Høiseth, G., Morini, L., Ganss, R. et al. (2013). Higher levels of hair ethyl glucuronide in patients with decreased kidney function. Alcoholism: Clinical and Experimental Research 37 (Suppl. 1): E14–16. Huppertz, L.M., Gunsilius, L., Lardi, C. et al. (2015). Influence of Gilbert’s syndrome on the formation of ethyl glucuronide. International Journal of Legal Medicine 129: 1005–1010. Iffland, R., Schmidt, V., Oehmichen, M. et  al. (1988). Aceton-­und 2-­Propanol-­Konzentration im Blut. Abhängigkeit vom akuten und chronischen Alkoholkonsum (Alkoholismus). Blutalkohol 25: 80–96. Isaksson, A., Walther, L., Hansson, T. et al. (2018). High-­throughput LC-­ MS/MS method for determination of the alcohol use biomarker phosphatidylethanol in clinical samples by use of a simple automated extraction procedure  – preanalytical and analytical conditions. The Journal of Applied Laboratory Medicine 2: 880–892. Jones, A.W. (2008). Biomarkers of acute and chronic alcohol ingestion. In: J.C. Garriott (ed.), Medicolegal Aspects of Alcohol, 5th edn, pp. 157– 203. Tucson, AZ: Lawyers and Judges Publishing Company. Kerekes, I. and Yegles, M. (2013). Coloring, bleaching, and perming: influence on EtG content in hair. Therapeutic Drug Monitoring 35:  527–529. Kiiskinen, T., Mars, N.J., Palviainen, T. et al. (2020). Genomic prediction of alcohol-­related morbidity and mortality. Translational Psychiatry 10: 23. doi.org/10.1038/s41398-­019-­0676-­2 Kharbouche, H., Faouzi, M., Sanchez, N. et al. (2012). Diagnostic performance of ethyl glucuronide in hair for the investigation of alcohol

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Pragst, F., Krumbiegel, F. and Thurmann, D. et  al. (2020). Positive findings of ethyl glucuronide in hair of young children from families with addiction background. International Journal of Legal Medicine 134: 523–532. Rainio, J., De Paoli, G., Druid, H. et al. (2008). Post-­mortem stability and redistribution of carbohydrate-­deficient transferrin (CDT). Forensic Science International 174: 161–165. Reisfield, G.M., Teitelbaum, S.A., Shannon, O. et al. (2020). The roles of phosphatidylethanol, ethyl glucuronide, and ethyl sulfate in identifying alcohol consumption among participants in professionals health programs. Drug Testing and Analysis 12: 1102–1108. Salomone, A., Pirro, V., Lombardo, T. et  al. (2014). Interpretation of group-­level factors from a large population dataset in the determination of ethyl glucuronide in hair. Drug Testing and Analysis 7: 407–413. Salomone, A., A Bozzo, A., Di Corcia, D. et al. (2018). Occupational exposure to alcohol-­based hand sanitizers: The diagnostic role of alcohol biomarkers in hair. Journal of Analytical Toxicology 42: 157–162. Sanchez-­Roige, S., Palmer, A.A., Fontanillas, P. et  al. (2019). Genome-­ wide association study meta-­analysis of the Alcohol Use Disorder Identification Test (AUDIT) in two population-­ based cohorts. American Journal of Psychiatry 176: 107–118. Schellenberg, F., Wielders, J., Anton, R. et  al. (2017). IFCC approved HPLC reference measurement procedure for the alcohol consumption biomarker carbohydrate-­deficient transferrin (CDT): Its validation and use. Clinica Chimica Acta 465: 91–100. Schubert, W., Dittmann, V. and Brenner-­Hartmann J. (2013). Urteilsbildung in der Fahreignungsbegutachtung – Beurteilungskriterien. Kirschbaum – Fachverlag für Verkehr und Technik, Bonn, Germany 2013. Schräder, J., Rothe, M. and Pragst, F. (2012). Ethyl glucuronide concentrations in beard hair after a single alcohol dose: evidence for incorporation in hair root. International Journal of Legal Medicine 126: 791–799. Schröck, A., Redondo, A.H., Fabritius, M.A. et  al. (2016). Phosphatidylethanol (PEth) in blood samples from “driving under the influence” cases as indicator for prolonged excessive alcohol consumption. International Journal of Legal Medicine 130: 393–400. Schröck, A., Thierauf-­Emberger, A., Schürch, S. and Weinmann, W. (2017). Phosphatidylethanol (PEth) detected in blood for 3 to 12 days after single consumption of alcohol-­ a drinking study with 16 volunteers. International Journal of Legal Medicine 131: 153–160. Sillanaukee, P., Aalto, M. and Seppä, K. (1998). Carbohydrate-­deficient transferrin and conventional alcohol markers as indicators for brief intervention among heavy drinkers in primary health care. Alcoholism: Clinical and Experimental Research 22: 892–896. Skråstad, R.B., Spigset, O., Aamo, T.O. and Andreassen, T.N. (2021). Stability of phosphatidylethanol 16:0/18:1 in freshly drawn, authentic samples from healthy volunteers. Journal of Analytical Toxicology, 45: 417–21. Sporkert, F., Kharbouche, H., Augsburger, M.P. et al. (2012). Positive EtG findings in hair as a result of a cosmetic treatment. Forensic Science International 218: 97–100. Stenton, J., Walther, S., Hansson, T. et al. (2019). Interindividual variation and factors regulating the formation of phosphatidylethanol. Alcoholism: Clinical and Experimental Research 43: 2322–2331. Stephanson, N., Helander, A. and Beck, O. (2007). Alcohol biomarker analysis: simultaneous determination of 5-­ hydroxytryptophol glucuronide and 5-­hydroxyindoleacetic acid by direct injection of

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Muskovich, M. and Haag-­Dawoud, M. (2012). Alcohol consumption among drivers subject to the Swiss license restriction of zero tolerance when driving. Traffic Injury Prevention 13: 537–543. Musshoff, F., Albermann, E. and Madea, B. (2010). Ethyl glucuronide and ethyl sulfate in urine after consumption of various beverages and foods  – misleading results? International Journal of Legal Medicine 124: 623–630. Nanau, R.M. and Neuman, G.M. (2015). Biomolecules and biomarkers used in diagnosis of alcohol drinking and in monitoring therapeutic interventions. Biomolecules 5: 1339–1385. Niemelä, O. (2016). Biomarker-­based approaches for assessing alcohol use disorders. International Journal of Environmental Research and Public Health 13: 166. Oppolzer, D., Barroso, M., Passarinha, L. and Gallardo, E. (2017). Determination of ethyl glucuronide and fatty acid ethyl esters in hair samples. Biomedical Chromatography 31 (4). doi: 10.1002/bmc.3858. Petzel-­Witt, S., Pogoda, W., Wunder, C. et  al. (2018a). Influence of bleaching and coloring on ethyl glucuronide content in human hair. Drug Testing and Analysis 10: 177–183. Petzel-­Witt, S., Meier, S.I., Schubert-­Zsilavecz, M. and Toennes, S.W. (2018b). PTCA (1H-­pyrrole-­2,3,5-­tricarboxylic acid) as a marker for oxidative hair treatment. Drug Testing and Analysis 10: 768–773. Pianta, A., Liniger, B. and Baumgartner, M.R. (2013). Ethyl glucuronide in scalp and non-­head hair: An intra-­individual comparison. Alcohol and Alcoholism 48: 295–302. Pirro, V., Valente, V., Oliveri, P. et al. (2011). Chemometric evaluation of nine alcohol biomarkers in a large population of clinically-­classified subjects: pre-­eminence of ethyl glucuronide concentration in hair for confirmatory classification. Analytical and Bioanalytical Chemistry 401: 2153–2164. Pirro, V., Di Corcia, D., Seganti, F. et al. (2013). Determination of ethyl glucuronide levels in hair for the assessment of alcohol abstinence. Forensic Science International 232: 229–236. Politi, L., Zucchella, A., Morini, L. et  al. (2007). Markers of chronic alcohol use in hair: comparison of ethyl glucuronide and cocaethylene in cocaine users. Forensic Science International 172: 23–27. Porpigli, N.M., Bortolotti, F., Romolo, M. et al. (2019). Critical evaluation of the association between elevated mean corpuscular volume and alcohol-­related traffic accidents: A retrospective study on 6244 car crash cases. Alcoholism: Clinical and Experimental Research 43: 1528–1532. Pragst, F., Auwaerter, V., Sporkert, F. and Spiegel, K. (2001). Analysis of fatty acid ethyl esters in hair as possible markers of chronically elevated alcohol consumption by headspace solid-­phase microextraction (HS-­ SPME) and gas chromatography-­ mass spectrometry (GC-­ MS). Forensic Science International 121: 76–88. Pragst, F., Rothe, M., Moench, B. et al. (2010). Combined use of fatty acid ethyl esters and ethyl glucuronide in hair for diagnosis of alcohol abuse: interpretation and advantages. Forensic Science International 196: 101–110. Pragst, F. (2015). Alcohol biomarkers in hair. In: P. Kintz, A. Salomone and M. Vincenti (eds.), Hair Analysis in Clinical and Forensic Toxicology, pp. 71–139. Amsterdam: Academic Press. Pragst, F., Suesse, S., Salomone, A. et al. (2017). Commentary on current changes of the SoHT 2016 consensus on alcohol markers in hair and further background information. Forensic Science International 278:   326–333.

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concentrations in clinical and post-­mortem urine. Drug Testing and Analysis 5: 439–445. Ulwelling, W. and Smith K (2018). The PEth blood test in the security environment: What it is; why it is important; and interpretative guidelines. Journal of Forensic Science 63: 1634–1640. Verhulst, B., Neale, M.C. and Kendler, K.S. (2015). The heritability of alcohol use disorders: a meta-­analysis of twin and adoption studies. Psychological Medicine 45: 1061–1072. Berto, R., Giovanni Cecchetto, G. et  al. (2012). Viel, G., Boscolo-­ Phosphatidylethanol in blood as a marker of chronic alcohol use: analysis. International Journal of A  systematic review and meta-­ Molecular Sciences 13: 14788–14812. Walters, R.K., Polimanti, R., Johnson, E.C. et al. (2018). Trans-­ancestral GWAS of alcohol dependence reveals common genetic underpinnings with psychiatric disorders. Nature Neuroscience 21: 1656–1669. Witt, S., Wunder, C., Paulke, A. et al. (2016). Detection of oxidative hair treatment using fluorescence microscopy. Drug Testing and Analysis 8: 826–831.

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urine using ultra-­performance liquid chromatography-­tandem mass spectrometry. Journal of Mass Spectrometry 42: 940–949. Suesse, S., Pragst, F., Mieczkowski T. et al. (2012). Practical experiences in application of hair fatty acid ethyl esters and ethyl glucuronide for detection of chronic alcohol abuse in forensic cases. Forensic Science International 218: 82–91. Sundström, M., Jones, A.W. and Ojanperä, I. (2016). Utility of urinary mortem toxicology when ethyl glucuronide analysis in post-­ investigating alcohol-­related deaths. Drug and Alcohol Dependence 159: 272–276. Thompson, A., Cook, J., Choquet, H. et al. (2020). Functional validity, role, and implications of heavy alcohol consumption genetic loci. Science Advances 6: eaay5034. doi: 10.1126/sciadv.aay5034 Tsanaclis, L., Bagley, K., Bevan, S. and Wicks J. (2020). The effect of prolonged storage time on the stability of fatty acid ethyl esters in hair samples. Journal of Analytical Toxicology, 44: 829–33. Turfus, S.C., Vo, T., Niehaus, N. et al. (2012). An evaluation of the DRI-­ ETG EIA method for the determination of ethyl glucuronide

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63.1.1  Car-­to-­pedestrian collision

the impacting front, it is also possible to be impacted towards the head end of one’s mass centre (Figure  63.1.2b, phase 1). Then, the pedestrian is prostrated (Figure  63.1.2b, phases 2 and 3). 2. Loading: The pedestrian is catapulted onto the hood of the car due to the rotation (lateral body axis or arrow axis). The head can reach the lower or even the upper beam of the windshield depending on the impact velocity (Figure  63.1.2a, phases 2–5). 3. Shuffling off: The pedestrian is shuffled off due to the car breaking and the kinetics follow the laws of lateral force impacts (Figure 63.1.2a, phases 5–7). In accordance with the laws of classical mechanics, the impact dynamics during the accident cause the reversible clear motion sequences of the casualty during the impact and afterwards. By identifying the trajectories – the routes of the impact of the casualty – the quality and quantity of the causal dynamic are determined. Distinctive support points of the trajectory of the accident casualty are his or her contact points or area with the impacting tool (here a car) or other limiting constraining forces (e.g. road surface, guarding rail, road signs and tree trunks). It is these constraining forces in particular that leave the impacting partners with a trace exchange (Figure  63.1.3) and mark the casualty with their characteristic morphology injuries. Therefore, from careful trace analysis and observation of the morphology of the injuries to the type of corresponding constraining forces conveying contact points or areas, the trajectory of the accident casualty and, from this, the dynamic of the impact can be worked out.

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From a mechanical point of view the collision of a pedestrian with a car resembles an inelastic impact in which the inelastic negligible mass of the human being in comparison with the car takes on the velocity of the impacting car within the shortest period of time, and the relative velocity between car and human being is reduced to zero. The velocity reduction leads to an energy absorption which equals that kinetic fall energy which is achieved with the impact of the collision velocity onto a hard base from a respective height. Such energy absorptions cause continuum destructions due to the deformation of the human tissue material. The sum of destruction is scalable and has been used to create the internationally acknowledged abbreviated injury scale (AIS), where the values correlate with the collision velocity (Figure 63.1.1). The injuries caused by the dynamics of the accident are mainly determined by the mass ratio of the impacting partner, the contours of the impacting car and the anatomy and current posture of the pedestrian (which are unfavourable for the pedestrian). The type of impact, the circumstance, whether it is a frontal accident with complete coverage or a streaking impact with a partial coverage, has a great influence on the kinetics and dynamics. In the case of a complete coverage, one usually faces the following kinetic sequences: 1. Impact: The pedestrian is usually impacted towards the foot end of his mass centre (Figure 63.1.2a, phase 1). Depending on

Handbook of Forensic Medicine, Volume 3, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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Figure 63.1.1  The abbreviated injury scale (AIS), showing on the right hand a rough relation between the AIS score and the collision velocity. The left-­hand side shows the relationship between the height of fall and the collision velocity.

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The investigation should be structured considering the typical accident phases (impact, loading and shuffling off). The observed injuries of the pedestrian are mainly blunt force trauma or ­sometimes a partially sharp trauma: haematomas, abrasions and separation of structures (ruptures, contusions to the crushing, contused lacerations with accompanying décollation, décollements and fractures). The impact of formed forces by, for e­ xample, especially structured car part can often be seen in typical skin desiccation patterns. In the case of a frontal collision with complete cover between the upright walking pedestrian and a car, the primary impact (Figure  63.1.2a, phase 1) is usually a blunt force trauma to the lower leg via the front spoiler transferred to the bumper. Since the pedestrian achieves a translational acceleration component due to the impact, the shoe soles glide on the road surface. This causes corresponding abrasion traces (Figure 63.1.4) – on two soles in a standing pedestrian and one sole in a walking pedestrian – that indicate the position of the pedestrian during the impact and the direction of impact. Since the standing surface (shoe sole), dependent on the weight of the pedestrian, has traction, a moment is exercised through the translational force of the impacting car contours (Figure 63.1.5a) which may cause flexion fractures close to the ankle with protrusion on the opposite side

of the impact (Figure  63.1.5b). In the mid-­tibial shaft, typical bending fractures take place with wedge-­shaped fractures (so-­ called Messerer’s fracture, Figure 63.1.5c–e). The wedge tip points in the direction of the impact so that the position of the pedestrian and, possibly, his or her walking direction at the time of impact can be determined. The impact to the lower leg causes, on the side close to the impact, abrasions and contusions with associated haemorrhages and also some wound pocket formations. The height above the shoe sole level of such injuries can be used to work out the height of the primary impacting car part. The breaking process and associated ‘nodding motion’ of the car may result in a lower impact to the pedestrian than in an impact with a stationary car. In the area where the impact occurred, the desiccations often provide information about the structure of the impacting car parts. Their manifestation is therefore of diagnostic importance. If the primary impact site (e.g. the hood edge) is at the height of the pelvis and is dorsal, vertebra and pelvic fractures and renal ruptures may occur (Figure 63.1.6). Impacts higher up on the side of the body can cause rip fractures and liver or spleen ruptures. The impact progresses to the loading phase (see Figure 63.1.2a, phases 2–4). During the loading process, the pedestrian is subject to a tangential movement towards the car hood edge or the

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Figure 63.1.2  (a) Phases of car to pedestrian collisions for a primary impact. (b) The mass centre of the pedestrian in a frontal collision.

Figure 63.1.3  A typical trace pattern on the surface of the car after a car-­to-­pedestrian collision.

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Figure 63.1.4  Abrasion of the shoe heel area after a car-­to-­ pedestrian frontal collision.

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engine hood which causes, respectively, textile abrasion and corresponding skin abrasions and scratches with fresh haemorrhaging into the tissue below (Figure  63.1.6). The main movement component, however, is a rotation around the vertical axis of the pedestrian. Simultaneously to this rotation, a rotation around the longitudinal axis occurs. Both rotations are ended with a blunt impact on the engine hood. Blunt injuries to the thorax (rib fractures and pneumohaemothorax, lung and cardiac contusion) and blunt abdominal traumas (contusions and ruptures of the major abdominal organs, haemorrhaging into the great net and mesenteric ruptures) are the consequence of this (Figure 63.1.7). Depending on the velocity, the head will impact on the hood on the lower bar of the windshield or the upper bar. Such impacts manifest themselves as contusive lacerations, facial fractures and skull cap fractures, which are usually located in the parietal bone region and continue into the base of the skull. The injuries leave traces (blood, tissue particles, hair, etc.) on the vehicle that can be morphologically and molecular biologically matched to the corpse. An impact to the windshield usually results in breaking and causing lacerations and a transfer of the glass splinters to the contact surface of the corpse of the victim (Figure 63.1.8). The velocity of the vehicle at the moment of collision might be extremely high so that if the induced rotation is maintained, the pedestrian may fly over the roof. However, in most cases after the loading, a breaking-­induced shuffling off with impact to the ground and subsequent sliding process takes place. Since shuffling off is subject to the law of slanted (here horizontal) shuffling off, the shuffling distance can be used to estimate the vehicle

(d) Figure 63.1.5  Lower leg injuries caused by impact with a vehicle: (a) the dynamics, (b) a flexion fracture close to the ankle with protrusion, (c) mid-­tibial shaft fracture (so-­called Messerer’s fracture), (d) typical lower leg injury caused by impact with a vehicle with a Messerer’s fracture and (e) typical Messerer’s fracture, original drawing from Messerer’s monograph 1880. Source: Messerer (1880).

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Figure 63.1.5  (Continued )

Figure 63.1.6  Injuries caused during the loading phase of a car to pedestrian collision: (a) abrasions and scratches, (b) fresh haemorrhaging into the tissue below, (c) destruction of a kidney and (d) tension scratches of the abdominal skin in a pedestrian who was hit by a car with a high velocity in the back (for injury mechanism see Figure 63.1.2a). (Continued on next page.)

velocity at the time of the shuffling off. The impact to the ground causes fall injuries that might be superimposed on the primary impact injuries. The sliding process finds its morphological equivalence in partially marked, extensive abrasions, avulsions, opening wound pocket formations and bone abrasions

(Figure  63.1.9). Particles of the ground at the shuffled off site might be identified in the wounds. If the pedestrian is not hit totally, but partially, the movement process is that of a streaked impact. The pedestrian suffers a rotation movement around his or her vertical axis which causes

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Figure 63.1.6  (Continued )

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Figure 63.1.7  (a) Blunt injuries of the thorax (rib fractures, pneumohaematothorax, lung and cardiac confusion). (b) Blunt injuries of the abdomen (contusions and ruptures of the major abdominal organs, here the liver).

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­ rimarily the head, but also other body parts, to impact the side p parts of the vehicle, mainly the A pillar of the vehicle. The expected injuries are therefore contusing lacerations in the area of the head and skull fractures, but also lacerations that are caused by prominent components. The recumbent pedestrian may also be over-­run. Often tyre tread marks can be seen within the skin (Figure 63.1.10a). Also, the clothes and the body are in contact with the materials adhering to the tyre tread (Figure 63.1.10b). Since the skin is stretched tangentially to the surface (Figure 63.1.11a), typical stretch marks occur (Figure 63.1.11b) which are accompanied by widespread avulsions (or even detachment). If a casualty is run over by a heavy vehicle, there may be burst fractures due to indirect impact in the area of the skull (Figure 63.1.11c) until the skull is crushed and there is an

explosive decerebration. In the area of the thorax, there may be rib fractures and fractures of the spine as well as organ ruptures and organ eversions due to extrusion. Finally, the recumbent pedestrian can also be dragged along behind or under the vehicle which will cause areal abrasions with consequent skin desiccations.

63.1.2  Car-­to-­car collision Box 63.1.1 outlines the frequency of the different types of car-­to-­ car collisions found. In a car-­to-­car collision, the collision-­related injury patterns to the passengers may be matched to clear trauma mechanical processes, and therefore the observed injury topography leads to an understanding of trauma mechanical processes.

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Figure 63.1.8  Face injuries caused by a frontal windshield contact (incisions by glass splinters, teeth fractures, contusions and lacerations).

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Figure 63.1.9  Extensive abrasions, opening wound pocket formations and bone abrasions after shuffling off. Particles of the ground can be seen in the wound.

The geometry and structure of the passenger cabin are clearly of major importance to the cause of injuries if one considers that the energy destructing the integrity of the human body is introduced through so-­called constraining forces in a mechanical sense (the

Figure 63.1.10  Characteristics of over-­run: (a) tyre tread marks and (b) tyre contact traces.

forces which inhibit the motion which is happening though limitations or rejections). The belt and airbag system are also part of such constraining forces because they limit the movement of the passengers, but are capable of reducing the injury potential of other constraining forces. Other important aspects are the so-­called initial conditions: the location of the passenger at the time of collision (sitting position and sitting posture) and the velocity of the passenger at the time of collision (equal to the collision velocity). From studies of front-­end, side and rear-­end collisions, the important elements for mixed form collisions can be calculated.

Front-­end collision The deceleration that occurs during the front-­ end collision induces a velocity and therefore a resulting movement of the driving passenger forwards (Figure 63.1.12a, phases 1–3). If the driver or passenger does not wear a seatbelt, he or she will touch the dashboard with the knees where a direct (usually an impact from a shaped object although not always) force with consecutive

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Figure 63.1.11  Over-­running the head: (a) dynamics of a bursting fracture, (b) typical stretch marks, (c) morphology of a bursting fracture.

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Box 63.1.1  Frequency of car-­to-­car collision types. • 45% of cases are a front-­end collision: ○○ In 25% there is complete coverage. ○○ In 25% there is 30–50% right-­side coverage. ○○ In 50% there is 30–50% left side coverage. • 20% of cases are side collisions: • 10% of cases are rear-­end collisions. • 10% involve a somersault. • 5% involve front-­end under-­driving.

c­ontusive lacerations occurs. The compression process to the thigh may cause bloody cartilage contusions of the femoral condyles with condylar spongiosa sintering, to intracondylar chisel fractures of the femur to supracondylar flexion fractures. Force introduction close to the knee may also cause an indirect bending of the femur with a bending fracture or even a posterior luxation fracture of the hip joint. Fractures close to joints are usually associated with destructions of the ligaments and the capsules. If the driver slams his feet on the footrest as a reflex, pilon fractures of the ankle joint may occur; in impacts with the car pedals (possibly due to the leverage effect to the foot), fractures of the metatarsal bone can be observed.

Figure 63.1.12  Phases of a front-­end collision: (a) driver without a seatbelt and (b) driver wearing a seatbelt.

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whether a seatbelt was worn or not at the time of collision is of interest to the criminal and civil law. Therefore, the examination of seatbelt injuries is cautiously performed in a positive way. Due to the mirror symmetry of the belt course, the driver and passenger can be distinguished with help of their seatbelt injuries. The casualty shown in Figure 63.1.13 (in a German vehicle) was the passenger. A lack of seatbelt injuries, however, does not exclude the use of a seatbelt. There are some injuries that are incompatible with wearing a seatbelt and prove that a seatbelt was not worn. In rare cases, the use of a seatbelt is of disadvantage to the occupant. The seatbelt might be too loose or the lower body of the occupant might slide underneath the seatbelt (submarining). Injuries to the inner organs (intestine and kidneys) can then occur. If the occupant cannot unbuckle in time, he or she might be subject to other damaging events after the primary collision (e.g. second collision and fire).

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In a side collision to the passenger cabin, torso injuries of the passenger close to the impact can cause spleen ruptures (side impacts from the left) and liver ruptures (side impacts from the right). The seatbelt provides little protection for the occupant close to the impact. Because of the law of inertia, the side impact causes an abrupt lateral inclination of the head, compressing the cervical spine close to the impact, with the tensile stress to the cervical spine on the opposite side of the impact. Compression and tension may be involved in such injuries. For the passenger on the opposite side of the impact, a seatbelt does provide protection because, through the fixation of the seatbelt, contacts with solid marginal structures within the passenger cabin are averted. In contrast to the seatbelt, a side airbag system definitely reduces injuries.

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The upper body and abdominal part of the driver who is not wearing a seatbelt reaches the steering wheel. The injuries ­commonly observed include rip fractures, cardiac and lung contusions, lung ruptures, aortic ruptures, liver rupture, spleen rupture, pancreatic contusion as well as intestinal and mesenterial haemorrhaging. Body fixation by the steering wheel causes a flexion (‘nodding’) of the head. If during this the face hits the steering wheel, facial injuries and fractures occur as discussed for the head impact injuries of the pedestrian. If the steering wheel stop the driver’s body, his or her face may hit the windshield. Typical glass splinter injuries are added to the blunt injuries. Finding the glass splinters is an important criterion in reconstructing the accident. The brace position with hands and arms causes extremity fracture (e.g. dislocation fractures within the thumb basal joint and compression fractures within the wrist joint). A successful brace position is not possible in crash velocities faster than 15 km/h. The injury possibilities for a driver are considerably lowered when wearing a seatbelt and using an airbag system (Figure 63.1.12b). If the seatbelt is properly applied, fatal injuries are rare in collision velocities of up to 40–45 km/h. However, seatbelt injuries can occur. The collision-­induced acceleration pressure causes haematomas in the form of the seatbelt and minor injuries to the upper skin layer which cause post-­mortal desiccations shaped by the seatbelt (Figure  63.1.13). High collision velocities can also cause rib fractures and injuries to the inner organs (liver, spleen and rarely the heart). Because of the fixation of the buckled-­in torso, collision-­related deceleration causes a forced ventral flexion of the head. Such a sudden bending of the spine causes on the concave side (ventrally) compressions and on the convex side (dorsally) distensions; these are part of the trauma mechanic injuries and lead to clinical complaints. Injuries caused by the seatbelt prove that a seatbelt was worn. The detection of

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Figure 63.1.13  A seatbelt-­shaped desiccation after a frontal impact.

Rear-­end collision In a rear-­end collision, the torso pushed forwards due to the back of the seat, in contrast to the head which is subject to the law of inertia. This process causes a shear pressure to the cervical vertebrae leading to a forced reclination of the head. This pathological movement process is called cervical vertebrae distortion. The underlying pathomechanical force is called cervical vertebrae trauma. There is pressure exercised onto the discs in the sense of a compressive load and neck muscles, vessels and ligaments (interior longitudinal ligament) are tensile strained. The injuries occur once the biomechanical tolerance limit is surpassed. The complaints that occur after such a cervical spine trauma are summed up under the definition ‘cervical spine syndrome after cervical spine whiplash’. It can be difficult to clinically differentiate such cervical spine traumas of another origin unless there is knowledge of whether a rear-­end collision occurred or not. In disagreements subject to civil law, in order to prove that a cervical spine syndrome was the result of a rear-­end accident, the velocity of the car driven into must be above a certain threshold. For a previously uninjured cervical spine, this threshold is approximately

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In order to adequately categorise the injury topography after a two-­wheeler-­to-­car collision, two different accident courses must be identified: • The vehicle hits the two-­wheeler in the side (Figure 63.1.14a). • The two-­wheeler hits the vehicle in the side (Figure 63.1.14b and c). During the collision phase, if a vehicle collides with its front with the side of a two-­wheeler (Figure 63.1.14a), the cyclist or motorcyclist who first hits the bumper or the hood, as in the case of the pedestrian, is mainly subject to a rotation movement. During the rotation movement, the cyclist is loaded or picked up and reaches, depending on the velocity, the hood with his head, or the windshield or the upper windshield frame. According to

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the primary contact site, there may be hip joint fractures, femoral neck fractures or femoral shaft fractures. During loading, injuries can be caused within the abdomen and chest organs, along with pelvis and rib fractures and, after the impact of the head, severe intracranial injuries. However, the intracranial injuries are not as common in the two-­wheeler cyclist as in a pedestrian. After the loading, there is shuffling off and injuries caused by being thrown off. If the two-­wheeler collides with the side of a car and if this collision is at the height of the passenger cabin (Figure 63.1.14b), there is a contact of the head/upper body part with the roof frame of the car in nearly all cases. This is particularly shape aggressive so that this type of collision is connected to very high mortality rates. Due to the unsuccessful brace position at the steering wheel, there occur metacarpal fractures, compression-­related dislocation fractures of the carpal bone and also compression-­related radius fractures in a typical location (loco typico). If the introduced force flow reaches the elbow, one can expect an extension fracture to the elbow close to the humerus. Since the two-­wheeler cyclist is moved forwards in a crouched position after the i­mpact-­related deceleration of the

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13 km/h. It is lower if there was previous damage to the cervical spine or if at the time of collision an out-­of-­position posture existed.

Figure 63.1.14  The most important types of two-­wheeler-­to-­car collisions: (a) vehicle hits a two-­wheeler in the side, (b) two-­wheeler hits a car at the cabin and (c) two-­wheeler hits a car at the bonnet.

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Madea, B. and Doberentz, E. (2019). Der Verkehrsunfall. In: Der Notfallsanitäter, Verkehrsmedizin, Luhri-­Verlag, 19: 20–56. Madea, B., Mußhoff, F. and Berghaus, G. (2012). Verkehrsmedizin. Fahreignung, Fahrsicherheit, Unfallrekonstruktion, 2. Aufl. Köln: Deutscher Ärzte-­Verlag. Maresch, W. and Maurer, H. (1985). Der Verkehrsunfall in gerichtsmedizinischer Sicht. Graz: Leykam. Mason, J.K. (1993). The Pathology of Trauma. London: Edward Arnold. Mattern, R. (2004). Verkehrsunfall. In: B. Brinkmann and B. Madea (eds.), Handbuch Gerichtliche Medizin, Band 1, pp. 1171–1214. Berlin Heidelberg New York: Springer. Ponsold, A. (1967). Lehrbuch der Gerichtlichen Medizin, 3. Aufl. Stuttgart: Thieme Verlag. Ropohl, D. (1990). Die rechtsmedizinische Rekonstruktion von Verkehrsunfällen. Stuttgart: Deutsche Automobil Treuhand. Schmitt, K.U., Niederer, P.F. and Walz, F. (2010). Trauma Biomechanics: Accidental Injury in Traffic and Sport. Berlin Heidelberg: Springer. Wagner, K. and Wagner, H.J. (Hrsg.) (1968). Handbuch der Verkehrsmedizin. Berlin Heidelberg New York: Springer. Wagner, H.J. (Hrsg.) (1984). Verkehrsmedizin. Berlin Heidelberg New York Tokyo: Springer. Wehner, H.D. (2012). Der Verkehrsunfall und seine Rekonstruktion. In: B. Madea, F. Mußhoff and G. Berghaus (Hrsg.), Verkehrsmedizin. Fahreignung, Fahrsicherheit, Unfallrekonstruktion. 2. Aufl., pp. 703–773. Köln: Deutscher Ärzte-­Verlag. Wehner, H.D. (2015). Der Verkehrsunfall. In: Madea B (Hrsg.), Rechtsmedizin. Befunderhebung, Rekonstruktion, Begutachtung. 3. Aufl., pp. 763–782. Berlin Heidelberg New York: Springer.

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two-­wheeler as dull mass, the knee contacts the vehicle and injuries occur. Within the frame of the movement process, the handlebar may be a constraining force introducing structures important to the injury of the femoral area. The collision phase is followed by a flying phase which entails certain impact-­related injuries (fractures of the upper extremities, intracranial injuries, rib and clavicle bone fractures and bending and compression injuries to the spine in force introduction through the head). In the subsequent sliding phase there are skin, muscle and bone abrasions that are often contaminated with dirt from the road. The flying phase as well as the sliding phase may be ended by an impact with a further obstacle (sidewalk, poles, guarding rail, etc.) which then introduces a blunt force to the injury morphology. If the brunt of the impact takes place on the side of the car at the height of the hood, the injury pattern is mainly marked by the flying phase (catapulting motion over the hood) (Figure 63.1.14c). In the context of the reconstruction of a two-­wheeler collision, a very important aspect is whether the cranial injury inflicted could have been prevented if a helmet had been worn. The helmet can fly off during the collision if not properly worn (poor fit, not adequate or missing chin strap closure). The damage to the helmet can be used to work out if it was incorrectly worn or not worn at all. The investigation also has to include whether the chin strap damage is due to the present accident or to past stretches. The forensic contribution to solving such a case is modest, but often decisive: injuries within the front and side neck areas with abrasions to the upper skin layer (and maybe the thereby caused post-­mortal desiccations) are a positive criterion. Since most two-­wheeler collision casualties present with head injuries, a lack of intracranial injuries imply that a helmet was worn.

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Adamec, J. and Graw, M. (2013). Rekonstruktion von Verkehrsunfällen. Fußgängerkollisionen. Rechtsmedizin 23: 423–438. Becker, K., Friedrich, K. and Rothschild, M.A. (2011). Rekonstruktion von Verkehrsunfällen. Sitzposition der Pkw-­Insassen. Rechtsmedizin 21: 561–573. Buhtz, G. (1938). Der Verkehrsunfall. Stuttgart: Ferdinand Enke Verlag. Donaldson, L. and Scally, G. (2009). Donaldson’s Essential Public Health, 3rd edn. Oxford New York: Radcliffe Publishing. Dürwald, W. (1966). Gerichtsmedizinische Untersuchungen bei Verkehrsunfällen. Leipzig: VEB Georg Thieme. Graw, M. and Adamec, J. (2017). Traffic injuries and deaths. In: M.M. Houck (ed.), Forensic Pathology. London: Elsevier Academic Press. Graw, M. and Koenig, H.G. (2002). Fatal pedestrian – bicycle collisions. Forensic Science International 126: 241–247. Laves, W., Bitzel, F. and Berger, E. (1956). Der Straßenverkehrsunfall. Ursachen Aufklärung Beurteilung. Stuttgart: Ferdinand Enke. Madea, B. (2015). Rechtsmedizin. Befunderhebung, Rekonstruktion, Begutachtung. 3. Aufl. Berlin Heidelberg New York: Springer.

63.2 Sudden Natural Death while Driving Guy Rutty

63.2.1 Introduction According to the World Health Organization, 1.35 million people, equating to 18.2/100 000 population, died worldwide in 2016 consequent to road-­traffic-­related incidents (WHO, 2018). Although the vast majority will die at the time from injuries sustained during the incident, or at a later date, of complications arising from the injuries sustained, a small proportion of all cases will die as a consequence of natural disease whilst in charge of transportation. As sudden death from natural causes may occur at any time and in connection with any occasion or any of someone’s daily tasks, then it is to be expected that, on occasion, sudden death will occur whilst a person is in charge of transportation. These deaths, depending upon the year and country of origin of the source publication, will make up between 3.5 and 25% of one’s road traffic autopsy-­related practice (West et al. 1968; Mant 1977; Halinen and Jaussi 1994; Lau 1996; Osawa et al. 1998; Tervo et al. 2013; Brodie et al. 2019). This is not unexpected as approximately 10% of all deaths are sudden,

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defined as ‘the person who was driving or propelling the vehicle when the fatal collapse occurred’ (Mant 1977). He goes on to inform us that ‘it is immaterial whether the driver has stopped the car on the highway or in a car park or collapsed by the side of the vehicle. A collapse indicates a collapse from which no recovery has occurred. It is not necessary for death to ensue immediately upon collapse. For example, a driver may collapse from a cerebrovascular accident and die several hours later in the hospital without regaining consciousness’. Although the vast majority of these deaths will be associated with domestic cars, they are also reported in lesser numbers in relation to commercial heavy vehicles, buses, minibuses, tractors, excavators as well as two-­wheeled vehicles such as motorbikes, mopeds and pedal cycles. Deaths in charge of electric milk floats and invalid carriages have been reported (Lau 1996, 1994; Cheng and Whittington 1998; Buettner et al. 1999). As the onset of a natural disease event is unpredictable, death may occur anywhere a vehicle can access. Unlike other fatal road traffic incidents, most cases associated with natural death occur during the daytime (Osawa et  al. 1998). There is no increase in incidence on any particular day of the week. One series reported a slight increase in natural deaths during the months of November and December (Oström and Erikkson 1987). If the driver is within the vehicle at the time and the vehicle is in motion, the vehicle itself often sustains little, if any damage, as the driver may have sufficient time to bring the vehicle to a halt prior to losing consciousness; hence, drivers are found dead in stationary vehicles (Baker and Spitz 1970; Antecol and Roberts 1990). Drivers may also be in the process of getting into or out of vehicles at the time of loss of consciousness; hence, they are found by the side of their vehicle. Deaths have also occurred whilst drivers have been pushing their vehicles, having arguments with other drivers and even whilst giving statements to the police (Bowen 1973). At other times vehicles are witnessed to veer suddenly from their direction of travel, cross intersections without stopping, plunge into rivers or impact head on with other vehicles (Levy 1963; West et al. 1968; Lau 1996; Sjogren et al. 1996; Osawa et al. 1998; Tervo et  al. 2013). LeCount and Ruckstinat (1929), in the first known published account of natural death at the wheel, describe two cases where the vehicles involved suddenly started to weave from side to side before leaving the road and overturning (Lecount 1929). Under these circumstances, the vehicle may collide with parked vehicles or road furniture resulting in minor damage to the vehicle (Oström and Erikkson 1987). In the case of head on impact, however, damage may be substantial, especially if it occurs at speed. Passengers, on seeing the driver collapse, may attempt to take over the controls of the vehicle to avert a collision.

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occurring within 1 hour of onset of symptoms. Thus, an incidence of 10% of sudden natural death amongst motorists is consistent with the incidence amongst the general population (Brodie et al. 2019). Although many of these deaths will gain little attention and may not be included in national road-­traffic-­related statistics, some incidents may gain special attention if, for example, heavy vehicles carrying dangerous goods become involved or others are injured or die as a consequence of the incident. The investigation of natural death whilst in charge of transportation poses a particular problem to pathologists. Although, as detailed in the following sections, there are consistent characteristics associated with these deaths which can be used to assist attributing the death to natural disease, where extensive traumatic injuries arise as a consequence of the incident; as elderly driver would be expected to have pre-­existing natural disease, it may be difficult for the pathologist to decided what role, if any, this natural disease had in the death. It is the classic problem of trying to distinguish between those dying off and dying with a natural disease. Thus, it should always be born in mind that just because a driver has sustained traumatic fatal injuries consequent to a road traffic incident that natural disease may be the initiating causative factor to the incident. In cases where there is obvious recent myocardial infarct, a ruptured Berry aneurysm or fresh massive pulmonary thromboembolism, the diagnostic process in an uninjured or only slightly injured person should not cause difficulties. However, where a person has pre-­ existing epileptic seizures or stenosing coronary atherosclerosis without thrombosis and myocardial infarct or the body is severely damaged by vehicle related fire, the pathological findings must be carefully assessed in the context of the entire case.

63.2.2 Demographics

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Most of the cases reported in the literature occur in males (Mant 1977; Oström and Erikkson 1987; Sjogren et  al. 1996; Buettner et  al. 1999). Although, as with the general population, sudden natural death can occur at any age in adulthood, in terms of natural death whilst in charge of transportation, most are reported to occur within the 50–60 age range, with a median age around 57 years common to many series (Schmidt et al. 1990; Cheng and Whittington 1998; Buettner et al. 1999). It is unusual for passengers, other road users or pedestrians to be killed in relation to the incident, although injuries may occur, especially if a head on impact occurs. An example where other vehicle occupants may die is provided by Levy who reported an incident where several bus passengers drowned after the driver of the bus suffered a cardiac event at the wheel, causing the bus to plunge into a river (Levy 1963).

63.2.3 Circumstances When considering the circumstances under which these deaths occur, we are assisted by Mant who provides a definition of what is meant by ‘in charge of transportation’. Mant suggests this can be

63.2.4  Cause of death In theory, any disease that can cause sudden unexpected death within the general population could have its onset whilst in charge of transportation. Having said that, the commonest forms of natural disease reported in relation to this type of road-­traffic-­ related death are associated with the cardiovascular and cerebrovascular systems as well as diabetes and epilepsy.

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63.2.5 Sleep-­related road traffic collisions

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It is estimated that between 10 and 30% of all fatal road traffic collisions are attributed to sleepiness at the wheel. Bioulac et al. (2017) define behavioural sleepiness as ‘difficulty in remaining awake even while carrying out activities’. They go on to define sleepiness at the wheel as ‘difficulty in remaining awake interfering with driving skills’ (Bioulac et al. 2017). These fatalities are different compared to those attributable to those caused by natural death at the wheel. First, the victims are often less than 30 years of age (Horne and Reyner 1999). The collision often happens at high speed, with no skid marks or hard breaking and with head on impacts or veering off the road to hit roadside furniture (Findley et al. 1991; Herman et al. 2014). In terms of road traffic collisions in general, commercial vehicles on non-­urban roads are more involved than domestic vehicles due to monotonous driving inducing sleepiness. It is suggested that in the USA, every lorry will be involved in one sleep-­related crash in its lifetime. The driver involved is usually healthy and often does not recall that they have fallen asleep. Risk factors for sleep-­related accidents include sleep deprivation due to having multiple jobs, working night shifts, unusual work schedules and less than 6 hours sleep in a 24-­hour period. The peak time is between 0200 and 0600 hours and 1400 and 1600 hours. The older driver is vulnerable to afternoon sleepiness. Sleep apnoea is another risk factor for both non-­ fatal and fatal collisions. Obstructive sleep apnoea is associated with obesity, particularly those with a fat neck. Narcolepsy may also be a risk factor. Circadian rhythms can affect the sleepiness effect of alcohol, prescription drugs and other substances. For example, alcohol consumed in the afternoon is about twice as potent in causing sleepiness than when taken in the early evening.

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About 85–95% of such deaths result from cardiovascular disease. Although a variety of myocardial, valvular, conducting system and aortic pathologies have all been described in association with death in charge of transportation, the commonest cause of death results from coronary artery disease with luminal stenosis over 75% (Grattan and Jeffcoate 1968; Baker and Spitz 1970; Mant 1977; James et al. 1980; Antecol and Roberts 1990; Schmidt et al. 1990; Buettner et al. 1999; Toit-­Prinsloo and Saayman 2016). They are usually associated with pre-­existing myocardial fibrosis from previous ischemic episodes as well as cardiac hypertrophy (Copeland 1987). Luminal occlusion by acute thrombosis is uncommon. Driving, especially under difficult conditions, is recognized to be associated with significant emotion-­induced stress upon the heart and circulatory system (Bellet et al. 1968). Urinary catecholamine excretion may increase by 100%, and several changes have been observed on electrocardiographic monitoring including ischaemia-­associated T wave change and ventricular ectopic beats (Schmid and Meythaler 1964). Heart rate may increase in excess of 140 beats per minute in non-­ professional drivers, particularly within urban rather than rural settings, and has been reported to exceed 200 beats per minute in professional racing drivers (Taggart et al. 1969). Blood pressure alters little in healthy drivers but increases in those with established coronary artery disease (Simonson et al. 1968). Increases in blood pressure and pulse do not appear to be related to driving at speed. Drivers suffering a cardiac ischaemic event may suffer prodromal symptoms; hence, they may have time to steer the vehicle to the side of the road and come to a stop before collapsing at the wheel. After cardiovascular causes, cerebrovascular causes are the next leading cause of death of which subarachnoid haemorrhage caused by ruptured aneurysm is the most frequently observed finding at autopsy. Unlike cardiovascular causes, the driver may get little if any warning prior to aneurysmal rupture which accounts for vehicles that have been observed to suddenly veer off the road or into the path of oncoming vehicles. No breaking or crash avoidance is observed in such circumstances and, unlike the more controlled stop scenario, severe traumatic injuries may be sustained. After cardiac and intracranial causes diabetics associated hypoglycaemia and epilepsy account for the majority of the remaining reported causes. Driving fatalities are rare in those with epilepsy, occurring in a younger age group. There is an increased risk in those who suffer from status epilepticus or neoplasm-­ associated epilepsy (Richards 2004; Sheth cerebral-­ et al. 2004; Krumholz 2009). Finally, a number of miscellaneous causes of death are reported such as pulmonary thromboembolism, pneumonia and chronic obstructive lung disease. Intoxication with alcohol is not a feature usually seen in these deaths. Deafness is also not thought to be a factor in road traffic accidents, but visual impairment in at least one eye, such that the driver is unable to see the other vehicle on the side of the defective vision, is a factor in fatal road traffic incidents (Grattan and Jeffcoate 1968). Alzheimer’s disease is the commonest cause of impaired cognitive function in the elderly. One study suggests that 47–53% of elderly drivers killed may have incipient Alzheimer’s disease and that these drivers had more accidents compared to age-­matched controls (Johansson et al. 1997).

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References and further reading Antecol, D.H. and Roberts, W.C. (1990). Sudden death behind the wheel from natural disease in drivers of four-­wheeled motorized vehicles. American Journal of Cardiology 66: 1329–1335. Baker, S.P. and Spitz, W.U. (1970). An evaluation of the hazard created by natural death at the wheel. New England Journal of Medicine 283: 405–409. Bellet, S., Roman, L., Kostis, J. and Slater, A. (1968). Continuous electrocardiographic monitoring during automobile driving. Studies in normal subjects and patients with coronary artery disease. American Journal of Cardiology 22: 856–862. Bioulac, S., Micoulaud-­Franchi, J.A., Arnaud, M. et  al. (2017). Risk of motor vehicle accidents related to sleepiness at the wheel: A systematic review and meta-­analysis. Sleep 4: zsx134. doi: 10.1093/sleep/zsx134. Bowen, D.A.L. (1973). Deaths of drivers of automobiles due to trauma and ischaemic heart disease: A survey and assessment. Forensic Science 2: 285–290. Brodie, L.R., Odell, M., Ranson, D. et al. (2019). Sudden natural death behind the wheel: Review of driver deaths and fitness to drive assessment history in Victoria, Australia 2012–2013. Journal of Forensic and Legal Medicine 63: 31–33.

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Schmidt, P., Haarhoff, K. and Bonte, W. (1990). Sudden natural death at the wheel  – a particular problem of the elderly? Forensic Science International 48: 155–162. Sheth, S.G., Krauss, G., Krumholtz, A. and Li, G. (2004). Mortality in epilepsy. Driving fatalities vs other causes of death in patients with epilepsy. Neurology 63: 1002–1007. Simonson, E., Baker, C., Burns, N. et  al. (1968). Cardiovascular stress (electocardiographic changes) produced by driving an automobile. American Heart Journal 75: 125–135. Taggart, P., Gibbons, D. and Somerville, W. (1969). Some effects of motor-­car driving on the normal and abnormal heart. British Medical Journal 4: 130–134. Tervo, T., Raty, E., Sulander, P. et al. (2013). Sudden death at the wheel due to a disease attack. Traffic Injury Prevention 14: 138–144. Toit-­Prinsloo, L. and Saayman, G. (2016). “Death at the wheel” due to tuberculosis of the myocardium: A case report. Cardiovascular Pathology 25: 271–274. West, I., Nielsen, G.L., Gilmore, A.E. and Ryan, J.R. (1968). Natural death at the wheel. JAMA 205: 68–73. World Health Organisations (WHO) (2018). Global Status Report on Road Safety 2018. https://www.who.int/publications/i/item/global­status-­report-­on-­road-­safety-­2018

63.3  Motor-­Vehicle-­Assisted Suicide Guy Rutty

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Buettner, A., Heimpel, M. and Eisenmenger, W. (1999). Sudden natural death ‘at the wheel’. A retrospective study over a 15–year time period (1982–1996). Forensic Science International 103: 101–112. Cheng, L.H.H. and Whittington, R.M. (1998). Natural deaths while driving: would screening for risk be ethically justified? Journal of Medical Ethics 24: 248–251. Copeland, A.R. (1987). Sudden natural death ‘at the wheel’ – revisited. Medicine, Science and the Law 27: 106–113. Findley, L.J., Weiss, J.W. and Jabour, E.R. (1991). Drivers with untreated sleep apnea. A cause of death and serious injury. Archives of Internal Medicine151: 1451–1452. Grattan, E. and Jeffcoate, G.O. (1968). Medical factors and road accidents. British Medical Journal 1: 75–79. Halinen, M.O. and Jaussi, A. (1994). Fatal road accidents caused by sudden death of the driver in Finland and Vaud, Switzerland. European Heart Journal 15: 888–894. Herman, J., Kafoa, B., Wainiqolo, I. et al. (2014). Driver sleepiness and rick of motor vehicle crash injuries: A population-­based case control study in Fiji (TRIP 12). International Journal of the Care of the Injured 45: 586–591. Horne, J. and Reyner, L. (1999). Vehicle accidents related to sleep: A review. Occupational and Environmental Medicine 56: 289–294. James, T.N., Pearce, W.N. Jr. and Givhan, E.G. Jr. (1980). Sudden death while driving. Role of sinus perinodal degeneration and cardiac neutral degeneration and ganglionitis. American Journal of Cardiology 45: 1095–1102. Johansson, K., Bogdanovic, N., Kalimo, H. et al. (1997). Alzheimer’s disease and apolipoprotein E e4 allele in older drivers who died in automobile accidents. Lancet 349: 1143–1144. Krumholz, A. (2009). Driving issues in epilepsy: Past, present, and future. Epilepsy Currents 9: 31–35. Lau, G. (1994). Ischaemic heart disease as a natural cause of death in motorists. Singapore Medical Journal 35: 467–470. Lau, G. (1996). Natural disease and alcohol intoxication amongst drivers of motor vehicle in Singapore from 1989 to 1993: A study of 140 necropsies. Annals of the Academy of Medicine of Singapore 25: 516–521. Levy, R.L. (1963). Heart disease in drivers of public motor vehicles as a cause of highway accidents. JAMA 184: 481–484. LeCount, E.R. and Ruckstinat, G.J. (1929). Sudden death from heart disease while motoring. JAMA 92: 1347–1348. Mant, A.K. (1977). Natural death while in charge of transportation. Legal Medicine Annual, pp. 111–126. Appleton-­Century-­Crofts, New York, 1969–1977. Osawa, M., Nagasawa, T., Yukawa, N. et al. (1998). Sudden natural death in driving: Case studies in the Western area of Kanagawa. Journal of Forensic and Legal Medicine 52: 315–318. Oström, M. and Erikkson, A. (1987). Natural death while driving. Journal of Forensic Science 32: 988–998. Richards, K.C. (2004). The risk of fatal car crashes in people with epilepsy. Neurology 63: E12. Sjogren, H., Eriksson, A. and Ostrom, M. (1996). Role of disease in initiating the crashes of fatally injured drivers. Accident Analysis & Prevention 28: 307–314. Schmid, E. and Meythaler, C. (1964). Appraisal of the sympathico-­ adrenal reaction during driving by dosage of vanilmandelic acide in urine. Klinische Wochenschrift 42: 139.

63.3.1 Introduction Suicide is the deliberate act of ending one’s life and is the second leading cause of death among people age 15–24 years (Rudy 2012). The choice of method will depend upon accessibility, availability, lethality, familiarity, frequency of use and the potential for concealing the suicide. Selzer and Payne inform us that ‘The automobile lends itself admirably to attempts at self-­ destruction because of the frequency of its use, the generally accepted inherent hazards of driving, and the fact that it offers the individual an opportunity to imperil or end his life without consciously confronting himself with his suicidal intent’ (Selzer and Payne 1962). They go on to inform the reader that ‘The automobile presents the depressed and frustrated individual with an opportunity to end his life in what he may perceive as a burst of glory. The automobile may also constitute a special enticement to the aggressive and vengeful feelings present in many would-­be suicides’. Both mobile and stationary two-­and four-­wheeled vehicles may be used to commit suicide. Although it is estimated that between 1.5% and 15% of single vehicle road traffic collision fatalities may be as a result of suicide, it is generally accepted that this is an underestimate and that the figure may be nearer to 30% (Hernetkoski Keskinen 1998; Alvestad and Haugen 1999; Ahlm et  al. 2001; Sarsone et  al. 2010; Henderson and Joseph 2012; Pompili et al. 2012; Straka et al. 2013; Razvodovsky 2017). Wyatt

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63.3.2 Categories of motor-­vehicle-­related suicides

Over 90% of cases are males, usually within the age range 25–34 years old. The deceased personal circumstances may include being unmarried, employed, although some suggest unemployed may be a feature, with intermediate education and higher socioeconomic skills. The vehicle is usually driven at speed to cause a head on impact, often into a heavy goods vehicle although other solid structures such as road furniture, walls or trees may also be targeted. No seatbelt may be worn at the time of impact. No evidence of skid marks, loss of control or braking is observed during scene examination. Typically, there is a history of alcohol/substance intoxication or abuse, bad driving, significant psychosocial stress, mental disorders such as depression and previous suicide attempts, recent negative life events and impulsivity and low distress tolerance personality trait. When examining these cases, it is worth considering that there is a reported association between latent Toxoplasma gondii infection and both an increased risk of road traffic collisions and suicide attempts in general (Sutterland et al. 2019). The blunt trauma injuries sustained are those expected for the type of collision that occurs. In the case of head-­on collisions, they are often extreme due to the effect of significant compartment intrusion resulting from the impact. The typical thoracic aortic injuries associated with rapid deacceleration may be present. The use of cross-­sectional imaging may identify occipital condyle, extensive multi-­level vertebrae, long bone, hand and feet bone as well as sacral and pelvic fractures that may be difficult to identify or visualise in detail through conventional invasive autopsy. The weight of the vehicle can be used to commit suicide by drowning (Byard and James 2001). Examples of this is when the vehicle is intentionally driven off mooring jetties either on beaches or large inland rivers, or off bridges or cliff edges into deep water. When combined with a remote location, then rescue is unlikely. Imajo (1983) described two cases where women used this method whilst their children were also in the vehicle. Although in one case the child survived, in the other the three children also drowned (Imago 1983). Vehicles driving off a height onto land may result in lethal blunt trauma injuries whereas falls into water often show no such injuries and death results from drowning (Hardwicke et al. 1985).

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et al. (2009) suggest that this is in part due to the use of the word ‘accident’ in relation to road traffic related deaths, as the word ‘accident’ lulls one into assuming that such deaths are unintentional (Hernetkoski et al. 2009). The principal problem in distinguishing between a suicide and a non-­suicide collision is that crash scene investigation on its own is often insufficient to distinguish between the two scenarios (Wyatt et al. 2009). A lack of skid or brake marks may assist the scene investigation, but it is often hard to prove intent, as up to 75% of cases are reported to lack a suicide note. There may also have been an intentional act to conceal the collision as a suicide, for example due to cultural norms or financial compensation reasons, or it could simply be an impulsive, unplanned act (Ohberg et al. 1997; Murray and de Leo 2007; Milner and De Leo 2012). Unlike other forms of suicide, those committing suicide with a road going vehicle may not consider the effect of their action upon other road users which may result in personal injury, death or post-­ traumatic distress (Pridmore et al. 2017). Radun et al. (2019) in their paper considering the views of Finnish heavy vehicle drivers towards suicidal crashes report that 20% of heavy vehicle drivers suspect they have been targeted in a suicide attempt and 80% viewed it as an occupational hazard.

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Motor-­vehicle-­assisted suicides can be considered in eight broad categories (Byard and James 2001): (1) speed and mass of their own vehicle to inflict lethal injury, (2) speed and mass of another vehicle to inflict lethal injury, (3) exiting a moving vehicle to inflict lethal injury, (4) using the vehicle to facilitate ligature-­ associated lethal neck injury, (5) toxic gases from or within a vehicle, (6) vehicle flammability to assist self-­immolation, (7) incised wounds inflicted inside vehicles and (8) vehicle-­associated firearm suicide.

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Speed and mass of their own vehicle to inflict lethal injury

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This is by far the commonest category of vehicle-­assisted suicide reported in the literature. Although it usually involves a single driver, driver and passenger suicides as well as murder–suicide using this method are reported (Atri et al. 1997; Pridmore et al. 2017). The latter is rare, an example of which is reported by Byard et al. (2018), who described a 39-­year old female who intentionally crashed her car, killing herself and her 11-­year-­old daughter at the same time. Although cars are the principal vehicle of choice, motorcycles riders may also choose this method to commit suicide (Milner and De Leo 2012). A number of risk factors are associated with these deaths (Selzer and Payne 1962; Hardwicke et al. 1985; Hernetkoski and Keskinen 1998; Murray and de Leo 2007; Bjornstig et  al. 2008; Hernetkoski et  al. 2009; Wyatt et  al. 2009; Sarsone et  al. 2010; Henderson and Joseph 2012; Gauthier et al. 2015; Bryard 2018).

Speed and mass of another vehicle to inflict lethal injury Three scenarios exist under this category. The first are pedestrians who intentionally step out or run out in front of cars or heavy vehicles both in urban and motorway environments (Hernetkoski and Keskinen 1998; Milner and De Leo 2012; Rudy 2012). They may also position themselves under vehicles or jump from bridges into the path of vehicles. Prior to impact, the pedestrian may hesitate, i.e. they may be seen or captured stepping out in front of a vehicle but then stepping back before impact only to step in front of another vehicle. The resulting blunt trauma injuries conform to the usual primary, secondary and tertiary inquiry patterns. When jumping from a height it may be possible, depending upon

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Each year a number of people will die as a result of jumping or falling from vehicles. The majority of these will be males aged between 10 and 24 years (Williams and Goins 1981). They fall or jump off the back of pickup trucks, often in rural environments, jump from the interior passenger compartment of moving vehicles or fall from opened doors. Occasionally, both drivers and passenger will alight a moving vehicle with the intention to end their life. In doing so, they will receive blunt trauma injuries. Maujean et al. (2016) provide a model to consider the trajectory of the body to assist with injury interpretation whilst Greenwood reports three patients who released their seatbelt restraints in the back of the ambulances in which they were travelling and consequently jumped from the rear door of the vehicles to their deaths (Greenwood 2006).

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Both hanging and ligature strangulation, with the associated extreme event of decapitation, have all been described in association with vehicle-­assisted suicide. Hanging can occur both inside and outside the vehicle. Pampin and Ridriguez (2001) describe two cases, both males found on the backseat of the vehicle. One had secured a belt between the window and the vehicle frame whereas the other had utilized the front driver seatbelt to take the weight of the body upon the neck. In the latter case, the images provided show the deceased nose and mouth pressed into the back of the driver’s seat, raising the possibility that the mechanism of smothering may have also contributed to the death. A rope attached to the front passenger door grab handle with the deceased lying across the front seat with his head below the front passenger seat was reported by Durso et al. (1995). The use of a seatbelt as the ligature was also reported by Chmura et al. (2018). In their case, the victim was sat outside the vehicle with their back towards an open door and the head on the threshold of the vehicle. Not only was there a ligature around the neck created by the seatbelt, but a second ligature in the form of a shoelace was present around the neck which was attached to the car’s steering wheel. Under such circumstances, the pathological findings are as would be expected for a partially ­suspended/ recumbent hanging. Ligature strangulation can be achieved by attaching the ligature around the neck with the other end attached to a solid external structure such as road furniture or another vehicle. The engine need not be on as releasing the handbrake to allow the weight of the vehicle to apply pressure to the neck is all that is required (Hardwicke et  al. 1985; Barranco et  al. 2018). If the engine is on and the vehicle is driven away, then decapitation may ensue. This has been described with the use of nylon, rope, metal and hemp ligatures attached to trees, fences, gates and streetlights (Zhao et al. 2008; Turk and Tsokos 2005; Hejna and Havel 2012;

Exiting a moving vehicle to inflict lethal injury

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Samberkar 2012; Marchard et al. 2019). Typically, the deceased is found within a heavily blood-­stained vehicle, which has its engine engaged, often on a sloop with the head within the vehicle but the ligature attached to a solid object some distance behind the vehicle. The neck wound is described as clean cut with haemorrhagic infiltration of the soft tissues and neck structures at the level of transection. The use of a tractor loader to enable suicidal decapitation of a person outside the vehicle, braced against a wall, has been described (Racette et  al. 2007). In this case, the head was within the loader whose engine was still running causing the tractor to bounce repeatedly off a silo wall against which the headless body was slumped.

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the extent of the injuries, to determine whether they landed first on their feet, back or head prior to vehicle impact. They may also show other forms of recent or historical self-­inflicted injury, for  example self-­ inflicted incised wounds to the forearms. Establishing intent in these circumstances can be assisted by the review of local closed-­circuit television or in vehicle dash-­cam footage. The second scenario is when a vehicle occupant intentionally leans out of a moving vehicle to impact upon another vehicle or road furniture. Boglioli et al. (1988) described a sole occupant of a moving vehicle who leaned out of the front-­passenger window, sustaining fatal injuries when their body struck a road sign. The final scenario is parking one’s vehicle on a railway line so that it can be struck by a train (Byard and James 2001). In such a scenario, if the vehicle is at right angles to the train, then the resulting blunt trauma injuries may be extensive and follow a side-­impact pattern of distribution.

Toxic gases from or within a vehicle Vehicular suicide involving toxic gases may be achieved by three methods. The first is the use of the vehicle engine to generate the fumes, the second is to run a generator within the vehicle compartment and the third method is to generate toxic gases inside a stationary sealed vehicle compartment through the mixing of chemicals. Inhalation of carbon monoxide from vehicle exhaust fumes was a common form of vehicular suicide until the introduction of catalytic converters when numbers associated with this method fell (Kendell 1998; Yoshioka et al. 2019). With the steady increase in electric motor vehicles, once all combustion engine vehicles no longer exist then this method of suicide will be confined to the history books. Until this happens, this remains a method available to a person wishing to take their own life. The victim is usually a male, aged 20–50 years. It is usual to find them between 0600 and 1200 hours, and suicide notes are reported to be present in approximately 42% of cases. Intoxication with drugs or alcohol can be a feature, and pets may also be found dead with the person (Thomsen and Gregersen 2006). Traditionally, a pipe is used to channel the exhaust fumes from the vehicle exhaust pipe into the car interior whose windows will be closed. This can lead to black deposits that have been observed to the hands of the deceased from attaching the pipe to the soiled exhaust pipe. Other times

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been used (McNew et al., 2011). When mixed, for example in a bucket in the footwell of the vehicle, the liberated hydrogen sulphide acts in a similar way to hydrogen cyanide, inhibiting mitochondrial enzymes by binding to the Fe3+ of cytochrome oxidase. The scene of these deaths are dangerous to emergency services and must be approached with appropriate health and safety advice and precautions. Luckily, the decedent will often alert responders to the danger by placing a warning sign in the window of the vehicle. Although the classic rotten egg smell of hydrogen sulphide can be detected as low as 0.3 ppm, levels of over 150 ppm will not have the smell due to olfactory nerve paralysis. Levels of 700–900 ppm cause central respiratory paralysis and beyond 1000  ppm near instantaneous respiratory paralysis and coma (McNew et  al., 2011). The autopsy findings are nonspecific although conjunctivitis and scleritis as well as evidence of respiratory irritation are described. Small punctate burns may be seen to the face and hands as a result of corrosive spatter. Greenish discoloration of the cerebral grey matter along with cherry-­red or pink lividity may occur.

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Occasionally, an individual will undertake self-­immolation whilst inside a motor vehicle, usually on their own and, in the authors’ experience, often in remote isolated locations (Shkrum and Jonston 1992; Byard and James 2001). Occasionally, fire can be used for murder–suicide as described by Bugelli et al. (2017). In their case, a 40-­year-­old female sedated her children at the home before latter setting fire to herself and the children inside a car. Subsequent toxicology undertaken upon larvae feeding on the remains of the children confirmed the presence of diazepam that had been given to the children’s in soft drinks. In these cases, there must be careful consideration of the scene to ensure one is not dealing with a homicide deposition site. Members of the public may alert police to the scene having misinterpreted the cracking of vehicle glass during the fire to the distant sound of gunshots. It is essential to try and identify the presence of an accelerant along with a method of conveying it to the vehicle. The scene may be complicated if the suicide has unusual elements, for example as described by Byard and James where, to prevent escape, the deceased had tied wire around one of his wrists and attached it to the steering wheel (Byard and James 2001), or in the case reported by Adair et al. (2003), the female had set fire to the vehicle whilst she was in the car trunk. Autopsy examination should identify the typical cherry pink change related to a raised carboxyhaemoglobin level along with soot and thermal injury to the respiratory tract although examination of the body may be hindered due to the effect of flame and heat. Toxicological examination is essential in these cases.

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the victim may be found under the exhaust pipe or the pipe may be connected to a plastic bag over the deceased head. The deceased may also be found within a sealed garage with the car engine running (Hardwicke et al. 1985). Although cars are traditionally used, cases are reported of motorbikes been used inside garages or the exhaust fumes been channelled from the bike by a pipe into a tent (Busuttil et  al. 1997; Martinez and Ballesteros 2006). The deceased will show the typical autopsy features of carbon monoxide poisoning. Superficial burns may be observed to exposed skin due to the action of hot fumes. Although catalytic converters reduce the carbon monoxide content of exhaust fumes to less than 0.1% in modern idle vehicles, it remains possible to commit suicide with a vehicle fitted with one. During a cold start, whilst the exhaust fumes remain below 250 °C, the catalytic converter does not function and thus for approximately the first 3 minutes high levels of carbon monoxide are produced. Depending upon the age of the vehicle, the material in the catalytic converter may have deteriorated. Under both conditions, suicide remains possible (Routley and Ozanne-­ Smith 1998; Thomsen and Gregersen 2006). When a vehicle with a catalytic converter is operated in a sealed environment, over time atmospheric oxygen is gradually reduced resulting in a decrease in engine efficiency and an increase in carbon monoxide production. Despite the presence of a catalytic converter, potentially lethal levels of carbon monoxide can be achieved (deRoux 2006; Hampson et al. 2017). During this time, the availability of oxygen is reduced and other gases such as carbon dioxide accumulate with resultant hypoxia. The formation of methemoglobinemia from the presence of oxides of nitrogen within the fumes may also be important in these cases (Vevelstad and Morild 2009). A vehicular suicide involving an electric car is reported by Bohnert and Zollinger (1994). The exhaust gas in this case was produced by a small motor generator used for charging the vehicles battery which had been placed into the loading space behind a seat in the vehicle. This case demonstrates that carbon monoxide suicide can occur in association with an electric car and emphasises the importance of a thorough scene examination to identify the source of the carbon monoxide in circumstances where carbon monoxide would not be immediately expected to be the cause of death. Toxic gases may be intentionally generated within a sealed vehicle compartment, the classic one has been hydrogen sulphide although hydrogen cyanide is another example. First reported in 2007  in Japan, thousands have died across the world by this method due to its dissemination via the Internet (Bott and Dodd 2013). The victim is often male, and the method chosen as it is reported to cause loss of consciousness and death in one breath, so-­called ‘knock down’. Hydrogen sulphide is generated by mixing two easily accessible common household chemicals: hydrochloric acid from products such as toilet or pool cleaners and lime sulphur (calcium polysulphide) from pesticides, tree spray or bath salts. Other chemicals such as ammonium hydroxide, aluminium sulphide, calcium hypochlorite, calcium sulphide, germanium oxide, hydrochloric acid, potassium ferrocyanide, sodium hypochlorite, sulphur, sulphuric acid and trichloroethylene have

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Incised wounds inflicted inside vehicles Self-­inflicted incised wounds inside vehicles are very uncommon. A non-­fatal case was reported in the Littlehampton Gazette, UK, in 2019. The driver of the car, having driven the vehicle into

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(2008) reports the most common method employed was by gunshot injury. Ninety-­six per cent of the victims were salaried, white males. Ortmann et al. (2016) describe a case of a 70-­year old male who shot himself to the left temple with a small calibre ‘bull-­dog’ revolver (Ortmann et al. 2016). This case is unusual as the gunshot injury was considered not to have been immediately fatal, but rather the authors suggest that these was a prolonged agony period during which he developed hypothermia.

References and further readings

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Adair, T., DeLong. L., Dobersen, M.J. et al. (2003). Suicide by fire in a car trunk: A case with potential pitfalls. Journal of Forensic Sciences 48: 1113–1116. Ahlm, K., Eriksson, A., Lekander, T. and Bjornstig, U. (2001). All traffic deaths are not “fatalities” – analysis of the official Swedish statistics of traffic accident fatalities in 1999. Lakartidningen 98: 2016–2022. Alvestad, M. and Haugen, O.A. (1999). Death behind the wheel. Tidsskr Nor Laegeforen 119: 966–968. Atri, A., Penttila, A. and Lonnqyist, J. (1997). Driver suicides. British Journal of Psychiatry 171: 468–472. Barranco, R., Caputo, F., Bonsignore, A. et  al. (2018). A rare vehicle-­ assisted ligature hanging: Suicide at the wheel. American Journal of Forensic Medicine and Pathology 39: 69–72. Bjornstig, U., Bjornstig, J. and Eriksson, A. (2008). Passenger car collision fatalities-­with special emphasis on collisions with heavy vehicles. Accident Analysis & Prevention 40: 158–166. Boglioli, L.R., Taff, M.L., Green, A.S. et al. (1988). A bizarre case of vehicular suicide. American Journal of Forensic Medicine and Pathology 9: 169–178. Bohnert, M. and Zollinger, U. (1994). Suicide carbon monoxide poisoning in an electric car. An unusual case report. Archiv für Kriminologie 194: 145–148. Bott, E. and Dodd, M. (2013). Suicide by hydrogen sulfide inhalation. American Journal of Forensic Medicine and Pathology 34: 23–25. Bryard, R.W. (2018). Vehicular suicides resulting in the deaths of others. Australasian Psychiatry 26: 225. Bugelli, V., Papi, L., Fornaro, S. et al. (2017). Entomotoxicology in burnt bodies: A case of maternal filicide-­suicide by fire. International Journal of Legal Medicine 131: 1299–1306. Busuttil A., Moody, G.H., Obafunwa, J.O. and Dewar, C. (1997). The skeletonized body: suicidal inhalation of motorbike exhaust. American Journal of Forensic Medicine and Pathology 18: 50–55. Byard, R.W. and James, R.A. (2001). Unusual motor vehicle suicides. Journal of Clinical Forensic Medicine 8: 1–4. Byard, R.W., O’Donovan, S., van den Huevel, C. and Baldock M. (2018). Familial vehicular murder-­suicide. Journal of Forensic Sciences 63: 1307–1308. Chmura, N., Borowska-­Solonynko, A., Brzozowska, M. et  al. (2018). Unusual case of suicide-­hanging in a car using two nooses. Archiwum Medycyny Sądowej i Kryminologii 68: 10–19. deRoux, S.J. (2006). Suicidal asphyxiation by inhalation of automobile emission without carbon monoxide poisoning. Journal of Forensic Sciences 51: 1158–1159.

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A variety of guns have been reported to have been used to commit suicide in stationary and moving vehicles. The use of moving vehicles and a projectile weapon allows these cases to be considered as so-­called ‘complex suicides’, i.e. where one or more mechanisms of death are used to ensure a fatal outcome. Murphy describes two such cases (Murphy 1997). The first case describes a male with a single gunshot injury to the right temple caused by a .22  long rifle semiautomatic pistol. As only minor collision damage was present to the vehicle, suggesting that the injury was inflicted whilst driving, the principal pathology was that of the gunshot injury to the head with the projectile recovered from the left temporalis muscle. The second case describes another male with an intraoral 12-­gauge shotgun injury to the head. The vehicle was found with the engine running at the bottom of a drainage ditch leading to an interpretation that the injury was inflicted whilst driving. Earlier Murphy had described a female who shot herself to the mid-­anterior chest with a .357 revolver whilst driving. In this case, the vehicle travelled approximately 200 feet along the roadside before becoming airborne and coming to rest on its roof. Thus, not only did the female sustain a gunshot injury to the central structures of the chest with the projectile recovered from the soft tissue posterior to the fifth thoracic vertebrae but she also sustained collision-­related injuries to her face, shoulders and upper extremities as a result of the rollover collision (Murphy 1989). In the cases above, no other vehicle was involved. Straka et al. (2013) describe a case which on the face of it is a typical single vehicle suicide in as much that a 20-­year old male’s car deviated suddenly into the opposite traffic to crash head on a truck with resultant external and internal blunt trauma impact injuries (Straka et  al. 2013). However, he was found to have a gunshot injury to his right temple caused by a .22 Long Rifle auto-­loading pistol. As with Murphy’s case, a deformed projectile was recovered from the left temporal region. Self-­inflicted gunshot injuries are also described in stationary vehicle. In a series of 217 work-­related parking lot suicides, Fayard

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several parked cars and damaged street furniture, was found by police, in the car, with two stab wounds to the chest. It is unclear from the limited information available within the report whether they were self-­inflicted or inflicted by another (https://www. littlehamptongazette.co.uk/news/driver-­found-­stab-­wounds-­ after-­lancing-­crash-­166958). A different scenario was reported by the Mail Tribune, USA, 2020. Here, after a high-­speed police chase ended, whilst the driver was removed from the vehicle by police, he was found to have several self-­inflicted stab wounds to the neck. Aid was rendered to him, but he died, and death has been attributed to the neck injuries (https://mailtribune.com/ news/top-­stories/highway-­99-­temporarily-­closed-­near-­talent-­ during-­ police-­ investigation). Although no further details are available, as it was reported that the vehicle stalled and thus no vehicle trauma has occurred, the findings at autopsy would be expected to be those diagnostics of self-­inflicted incised wounds.

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Murray, D. and de Leo, D. (2007). Suicidal behavior by motor vehicle collision. Traffic Injury Prevention 8: 244–247. Ohberg, A.A., Penttila, A. and Lonnqvist, J. (1997). Driver suicides. British Journal of Psychiatry 171: 468–472. Ortmann, J., Doberentz, E., Kernbach-­ Wigthon, G. and Madea, B. (2016). Lethal hypothermia after firing a suicidal shot to the head in a car. Archiv für Kriminologie 238: 188–197. Pampin, J.M.B. and Rodriguez, B.A.L. (2001). Suicidal hanging within an automobile. American Journal of Forensic Medicine and Pathology 22: 367–369. Pompili, M., Serafini, G., Innamorati, M. et al. (2012). Car accidents as a method of suicide: A comprehensive overview. Forensic Science International 223: 1–9. Pridmore, S., Vardanov. S. and Sale, I. (2017). Suicide and murder-­ suicide involving automobiles. Australian Psychiatry 25: 32–34. Racette, S., Vo, T.T. and Sauvageau, A. (2007). Suicidal decapitation using a tractor loader: A case report and review of the literature. Journal of Forensic Sciences 52: 192–194. Radun, I., Radun, J., Kaistinen, J. et al. (2019). Suicide by crashing into a heavy vehicle: Professional driver’s views. Traffic Injury Prevention 20: 826–831. Razvodovsky, Y. (2017). Suicides and road traffic deaths in Russia: A comparative analysis of trends. European Psychiatry 41: S891 Routley, V.H. and Ozanne-­Smith, J. (1998). The impact of catalytic converters on motor vehicle exhaust gas suicides. MJA 168: 65–67. Rudy, B.S. (2012). Suicide by pedestrian versus motor vehicle. A case report. American Journal of Forensic Medicine 33: 268–269. Samberkar, P.N. (2012). Motor vehicle-­assisted ligature strangulation causing complete decapitation. American Journal of Forensic Medicine and Pathology 33: 86–87. Sams, R.N., Carver, H.W., Catanese, C. and Gilson, T. (2013). Suicide with hydrogen sulfide. American Journal of Forensic Medicine and Pathology 34: 81–82. Sarsone, R.A., Lam, C. and Wiederman, M.W. (2010). History of attempted suicide and reckless driving: Cross-­sectional study in primary care. Primary Care Companion Journal of Clinical Psychiatry 12: PCC.09m00888. Selzer, M.L. and Payne, C.E. (1962). Automobile accidents, suicide and unconscious motivation. American Journal of Psychiatry 119: 237–240. Shkrum, M.J. and Jonston, K.A. (1992). Fire and suicide: A three-­year study of self-­ immolation deaths. Journal of Forensic Sciences 37: 208–221. Straka, L., Novomesky, F., Stuller, F. et  al. (2013). A planned complex suicide by gunshot and vehicular crash. Forensic Science International 228: e50–e53. Straka, L., Novomesky, F., Stuller, F. et  al. (2013). A planned complex suicide by gunshot and vehicular crash. Forensic Science International 228: e50–e53. Sutterland, A., Kuin, A., Kuiper, B. et  al. (2019). Driving us mad: the association of toxoplasma gondii with suicide attempts and traffic accidents  – a systematic review and meta-­ analysis. Psychological Medicine 49: 1608–1623. Thomsen, A.H. and Gregersen, M. (2006). Suicide by carbon monoxide from car exhaust-­ gas in Denmark 1995–1999. Forensic Science International 161: 41–46.

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Durso, S., Vecchio, S.D. and Ciallella, C. (1995). Hanging in an automobile: A report on a unique case history. American Journal of Forensic Medicine and Pathology 16: 352–354. Fayard, G.M. (2008). Work-­related fatal injuries in parking lots, 1993– 2002. Journal of Safety Research 39: 9–18. Gauthier, S., Reisch, T., Ajdacic-­Gross, V. and Bartsch, C. (2015). Road traffic suicide in Switzerland. Traffic Injury Prevention 16: 768–772. Greenwood, M.J. (2006). Self-­inflicted death during interfacility transfer. Annals of Emergency Medicine 47: 212. Hampson, M.B., Holm, J.R. and Courtney, T.G. (2017). Garage carbon monoxide levels from sources commonly used in intentional poisonings. UHM 44: 11–15. Hardwicke, W.B., Taff, M.L. and Spitz, W.U. (1985). A case of suicide hanging in an automobile. American Journal of Forensic Medicine and Pathology 6: 362–364. Hejna, P. and Havel, R. (2012). Vehicle-­assisted decapitation. American Journal of Forensic Medicine and Pathology 33: 73–74. Henderson, A. and Joseph, A.P. (2012). Motor vehicle accident or driver suicide? Identifying cases of failed driver suicide in the trauma setting. Injury International Journal of the Care of the Injured 43: 18–21. Hernetkoski, K. and Keskinen, E. (1998). Self-­destruction in Finnish motor traffic accidents in 1974–1992. Accident Analysis & Prevention 30: 697–704. Hernetkoski, K.M., Keskinen, E.O. and Parkkari, I.K. (2009). Driver ­suicides in Finland  – are they different in northern and southern Finland? International Journal of Circumpolar Health 68: 249–260. https://mailtribune.com/news/top-­stories/highway-­99-­temporarily-­ closed-­near-­talent-­during-­police-­investigation https://www. littlehamptongazette.co.uk/news/driver-­found-­stab-­wounds-­after­lancing-­crash-­166958 Imago, T. (1983). Suicide by motor vehicle. Journal of Forensic Sciences 28: 83–89. Kendell, R.E. (1998). Catalytic converters and prevention of suicide. Lancet 352: 1525. Marchard, E., Mesli, V., Le Garff, E. et al. (2019). Vehicle-­assisted ligature decapitation: A case report and a review of the literature. Journal of Forensic and Legal Medicine 65: 119–123. Martinez, M.A. and Ballesteros S. (2006). Suicidal inhalation of motorbike exhaust: adding new data to the literature about the contribution of gasoline in the cause of death. Journal of Analytical Toxicology 30: 697–702. Maujean, G., Guinet, T. and Malicier, D. (2016). Fall from a car driving at high speed: A case report. Forensic Science International 261: e11–e16. McNew, J.L., Rigouard, S.G., Welles, W.L. et al. (2011). Chemical Suicides in Automobiles  —­Six States, 2006–2010. Morbidity and Mortality Weekly Report. Centers for Disease Control and Prevention. https:// www.cdc.gov/mmwr/pdf/wk/mm6035.pdf Milner, A. and De Leo, D. (2012). Suicide by motor vehicle “accident” in Queensland. Traffic Injury Prevention 13: 342–347. Murphy, G.K. (1989). Suicide by gunshot while driving an automobile. American Journal of Forensic Medicine and Pathology 10: 285–288. Murphy, G.K. (1997). Suicide by gunshot while driving a motor vehicle: two additional cases. American Journal of Forensic Medicine and Pathology 18: 295–298.

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European Union (https://ec.europa.eu/eurostat/statistics-­explained/ index.php/Rail_accident_fatalities_in_the_EU#:~:text=In%20 2018%2C%20there%20were%201,and%20748%20persons%20 seriously%20injured), 1844  in 2019  in the USA (https://www.bts. gov/content/transportation-­fatalities-­mode) and 283 suicides and 23 non-­suicide deaths occurred in the United Kingdom 2019/2020 (https://www.networkrail.co.uk/running-­the-­railway/looking-­ after-­the-­railway/delays-­explained/fatalities/). Typically, these deaths will make up approximately 1% of a forensic pathologist’s workload with the majority been suicides (Driever et al. 2002). Due to the difference in mass between a human body and a train, large kinetic energy exchanges occur (Akkas et al. 2011). The body is thus often severely injured. Identification may thus play an important role in the investigation of these deaths. The possibility that the railway has been used to cover up a homicide must always be within the mind of the pathologist investigating the death. Acts of terrorism and multiple fatality train-­ related incidents are considered elsewhere.

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Turk, E.E. and Tsokos, M. (2005). Vehicle-­assisted suicide resulting in complete decapitation. American Journal of Forensic Medicine and Pathology 26: 292–293. Vevelstad, M. and Morild, I. (2009). Lethal methemoglobinemia and automobile exhaust inhalation. Forensic Science International 187: e1–e5. Williams, A.F. and Goins, S.E. (1981). Fatal falls and jumps from motor vehicles. AJPH 71: 275–279. Wyatt, J.P., Squires, T., Collis, S. and Broadley, R. (2009). Road traffic suicides. Journal of Forensic and Legal Medicine 16: 212–214. Yoshioka, E., Hanley, S.J.B. and Saijo, Y. (2019). Time trends in suicide rates by domestic gas or car exhaust gas inhalation in Japan, 1968– 1994. Epidemiology and Psychiatric Sciences 28: 644–654. Zhao, D., Ishikawa, T., Quan, L. et al. (2008). Suicidal vehicle-­assisted ligature strangulation resulting in complete decapitation: an autopsy report and review of the literature. Legal Medicine 10: 310–315.

63.4  Railroad-­Related Deaths

63.4.2 Demographics

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For railway-­related deaths, males outnumber females. There is a very wide age range reported, from infants as young as 3 weeks to adults in their 80s (Shapiro et al. 1994; Davis et al. 1997; Spaite et  al. 1988), with a peak range of 20–40 years often quoted for suicides (Mohanty et  al. 2007; Krysinska and De Leo 2008). Analysis of weekday and seasonal variations reports conflicting observations, although in general accidents tend to occur during commuter rush hours whereas suicides are reported more often towards the end of the week and in afternoons (Lerer and Matzopoulos 1997; Erazo et  al. 2004; Radbo et  al. 2005; Sousa et al. 2015). Intoxication with alcohol is a frequent finding in both suicidal and non-­suicidal railway-­related deaths (Matzopoulos et al. 2006; Silla and Luoma 2012). Unlike other forms of suicide attempt such as by hanging or tablet overdose where the individual may survive, railway-­ related suicide has up to a suggested 94% success rate (Krysinska and De Leo 2008; Taylor et al. 2016). Suicide is associated with psychopathology, especially depression and schizophrenia, a history of psychiatric hospitalisation, proximity of the mental health institution to the railway track, familiarity with the concept of train-­related suicide, dense population areas and ease of access to the rail track (De Leo and Krysinska 2008; Van Houwelingen et  al. 2010). Open track between stations is favoured over stations (De Leo and Krysinska 2008; Lukaschek et  al. 2011). Often the victim may exhibit unusual behaviour prior to the suicide attempt including choosing an eminent spot on the station platform, standing at the edge of the platform, deviant behaviour, dropping or leaving personal belongings, avoidance of eye contact, erratic gesture, mimic or movement, erratic communication, general confusion, wandering aimlessly, waiting or hiding to jump in front of train and wearing out of ordinary clothing (Lukaschek et  al. 2011; Dinkel et  al. 2011; Radbo and Andersson 2012).

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63.4.1 Introduction

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Murphy (1976) defined the term ‘railroad-­related death’ as ‘a death either occurring on railroad property, including railroad cars, tracks, and right of way, or in which the decedent’s body was first discovered either on or very near railroad property’. Using this definition, the first rail-­related death was reported in 1830 when Rt. Hon William Huskisson, member of Parliament, whilst attempting to shake hands with the Duke of Wellington at the opening of the Liverpool and Manchester Railway, failed to notice George Stephenson’s Rocket locomotive travelling down the opposite track. Attempting to swing into a carriage he fell onto the tracks. His leg was severely injured by the passage of the Rocket. He was taken, by train, to hospital, but subsequently died of his injuries (Creevey 2020). The first railway suicide in England was recorded in the 1856 Registrar General’s report. A table on page 170 of this report records a suicide connected to the railway of a male in 1852 (Suicides in England 1852–1856). The first railway-­related murder in England was in 1864. Thomas Briggs was assaulted by Franz Muller in a railway carriage and subsequently found unconscious with severe injuries between Hackney Wick and Bow stations. He died of his injuries the following night (https://www.btp.police.uk/about_us/our_history/ crime_history/the_first_railway_murder.aspx). Finally, the first act of railway terrorism in Britain was in 1880 when a bundle of explosives was placed on a track between Bushey and Watford (https://www.btp.police.uk/about_us/our_history/timeline/ timeline_1851-­1950.aspx). Today, single or multiple railway-­related deaths occur throughout the world, for example 853 fatalities occurred in 2018 across the

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63.4.3 Circumstances As far back as the 1870 edition of the Annual Report of the Registrar General, we see how railway-­related deaths can be categorised into a number of distinct themes. At this time, the themes were as follows: run over on the line, fall from a carriage or engine, killed by collisions, killed by crushing, killed by a fall of heavy substances and killed from burns (Thirty-­First Annual Report of the Registrar-­General of Births, Deaths and Marriages in England 1870). With refinement, taking into account of methods of modern-­day transportation, these deaths can be considered in six broad themes: persons on foot on the tracks, bodies found on or near railway property, riding on trains, falls from trains and platforms, vehicle–train collisions and miscellaneous causes.

Bodies found on or near railway property Bodies may be found on or in railway property without any witness to the death. These should be treated as suspicious until an explanation can be asserted. Those attempting suicide may jump from a height, for example from bridges or buildings, onto railway property to be found subsequently adjacent to the tracks. Under such circumstances, the injuries sustained are those from the fall from a height. Patterned injuries from railroad ballast may be present, although such injuries may also occur when bodies are struck and thrown by trains. People walking on tracks may collapse of natural disease. Under such circumstances, there is a paucity of traumatic injuries other than the typical injuries of an uncontrolled collapse to the ground. Drug users may overdose in railroad carriages and warehouses. Those sleeping rough may die of fire-­related mechanisms when personal property, such as sleeping bags, or railroad property catches fire through unguarded fires or natural flames such as by using candles. Bodies that have been shot, strangled or stabbed may be left on rail lines as a means of disposal; the intention has been that the disruption of the body by the impact of a train to coverup the trauma that caused the death (Davis et al. 1997). Bodies may also be disposed of through burial on railroad property. Radiology examination is recommended in all unwitnessed railway deaths, especially disrupted bodies, not only to aid identification but to consider the cause of death as gravel has a lower radiodensity to metal projectiles.

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Railroad tracks and property are private property. Thus, by definition, persons who are neither railroad employees nor passengers who are on the tracks are trespassing (Pelletier 1997). Each year a number of people die in any given country, whilst on foot on railroad tracks. Although some will be employees of the railroad working at the time, the vast majority are trespassers. Those that are struck by trains whilst on foot traditionally are undertaking one of three activities: walking on or by the track, sitting or kneeling on or by the track or platform edge and lying across or in between the tracks (Murphy 1976; Davis et al. 1997; Lerer and Matzopoulos 1997; De Leo and Krysinska 2008; Krysinska and De Leo 2008; Silla and Luoma 2012). Those kneeling by the track may be doing it in such a way with the intention of committing suicide by presenting their head to the oncoming train. When walking they may be struck by a projecting part of the train or carriage which may cause patterned marks to the body that can be identified at subsequent autopsy. Platform and trackside jumping into the path of the train occurs in those with suicidal intent. Employees working on the line can, as with the general population, die of natural, principally cardiac events (Davis et al. 1997). They can also die of trauma when trains back into them, crush them between vehicles or run them over. This is reported to occur more often in non-­metropolitan railway lines. As with the public, they may not see or hear the vehicle prior to impact, or their attention may be distracted from the impending danger at the time of the incident (Murphy 1976). Familiarity with the railroad environment may breed complacency. Pathologists investigating such deaths should seek out any evidence of visual or hearing impediment through review of the deceased medical records and consider undertaking an examination of the optical and auditory sensory organs and pathways by the most appropriate means available to them. In terms of the general public, in some parts of the world, the close proximity of low economic accommodation to the rail tracks increases the risk of residents been struck by a train (Lerer and Matzopoulos 1996). Deaths can be caused by people running across tracks to try and catch trains, avoid ticket controls or

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fleeing from violence for example criminal gangs praying on commuters. Children under the age of 3 years, unaware of the danger that the tracks present, may simply wander onto the line (Nixon et al. 1998).

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Riding on trains Riding within a train compartment is normally considered a relatively safe mode of transport although natural, accidental, suicidal and homicide deaths do occur both on stationary and mobile trains (Murphy 1976). Assaults onboard trains causing death have been reported since 1864. People can sustain injuries from objects thrown at trains. Murphy (1976) reports that both members of the public and rail employees have been killed by people shooting at trains. Leaning out of the window of a moving train can result in the individuals head striking fixed objects external to the train such as posts, trees or electric poles (Mohanty et al. 2007). Although often popularised in media such as film, electrical injury from third rails as found, for example, in association with subway systems results in few deaths. Unlike the high-­tension, 25  000 V alternating current cables above the train, third rails carry direct current of approximately 600 V. Although unintentional occupational or general public contact by hand, tools or falls may occur with or onto the rail, this usually does not cause

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down on the tracks. Vehicle drivers may not see warning signs (assuming they are operational at the time) or the approaching train or there may be wanton disregard for safety as the driver tries to cross despite warning signals (Murphy 1976). They may wait for the first train to pass only to be struck by the second that may come from the opposite direction.

Miscellaneous causes

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As the risk is the same as that of the general population, a number of people will die suddenly and unexpectedly of natural disease, principally of ischemic heart disease whilst travelling on the railway networks. Fire-­ related deaths, not associated with derailment, occur occasionally due to train–train or train–bus collisions (Evans 2011). An unusual case of self-­ immolation on the Japanese Shinkansen high-­speed railway is reported (Morita et al. 2016). In this case, an elderly male performed self-­immolation with oils at the front of the front passenger compartment. He died whilst a number of other passengers sitting in close proximity to him suffered tracheal thermal burns and carbon monoxide poisoning. Although not considered in detail in this chapter, head-­on train–train collisions can result in front carriage passengers becoming entrapped within the wreckage and succumbing to injuries sustained through mechanical trauma, burns and scalds or traumatic asphyxia (Hambeck and Pueschel 1981).

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fatalities. Although fatalities may occur due to electric-­induced cardiac arrest, the more common scenario is that they are struck by the oncoming subway train during the incident (Rabban et al. 1997). A case of an individual walking on the railroad with a fishing pole which made contact with the overhead line causing them to collapse to the ground only to be cut in half by a passing train illustrates the difficulty that may face investigators in reconstructing the events associated with the death (Preuss et al. 2020). On the other hand, deaths associated with electricity and railroads can occur due to riding on top of the train. Known as ‘train surfing’, where a person stands up or crouches on the roof of the train, they can die either from impact with oncoming overhead obstacles or due to contact with the overhead electric cables (Sternick et  al. 2000). Contact with the cable however need not occur. Due to the high-­tension voltage, the phenomenon of arcing can occur. Here, the victim is grounded by contact with the metal parts of the carriage which is in contact with the ground by the rails. Ionised particles occur between the cable and the victim. These will cross 2–3 cm for every 10 000 V. The arc generates a temperature of 3000–20 000 °C, igniting clothing, causing burns and throwing the victim downwards. The current is not thought to pass through the victim (Koller 1991). A reflection of modern culture, particularly reported in India, sees arc and contact-­induced fatalities associated with taking self-­portraits (‘selfies’) with smartphones whilst riding on top of trains (Cuculic and Sosa 2019).

Falls from trains and platforms

63.4.4  Cause of death

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Although falling from a stationary train does not usually result in fatal injuries in adults, getting on and off a train or falls from a moving train, for example by international jumping or due to overcrowding, can result in fatalities. Victims of rape and robbery may be thrown from moving trains (Lerer and Matzopoulos 1997). Metrorail systems see pedestrians interact with the trains only at stations/platforms, have slower train speeds and are associated with traumatic limb amputations following falls from the platform (Donnally et al. 2019). As the person gets on or off the train, they may fall between the train and the platform. This may cause them to go under the wheels of the train and sustain fatal injuries (Shapiro et al. 1994). They can also become caught by a component of the train or carriage and be dragged (Murphy 1976).

Vehicle–train collisions Kligman et  al. (1999) report that, in 1999, a collision occurred between a motor vehicle or a pedestrian and a railway train every 100  minutes in the USA and that a motorist is 40 times more likely to die following a collision with a train than with another motor vehicle. Motor vehicle–train collisions usually occur at railway crossings. Cars, bicycles, motorbikes and heavy vehicles may all be involved (Shapiro et al. 1994; Davis et al. 1997). The train may be moving or stationary at the time. Vehicles can be intentionally driven into the train or the vehicle may have stalled or broken

As highlighted already, railroad-­ related deaths often exhibit multisystem, severe traumatic injuries. Although it is all too often easy to attribute the cause of death to ‘polytrauma’ or ‘multiple injuries’, where possible the pathologist should strive to be more specific as to the most significant life-­threatening injury present. For example, decapitation is a non-­survivable injury whereas lower limb amputation, although life threatening, need not be fatal. A spectrum of injuries, ranging from burns to crushing, avulsions, amputations and whole-­body disruption, is reported in children and adults in a range of circumstances related to railroad-­related fatalities (Moore et  al. 1991; Shapiro et  al. 1994; Lerer and Matzopoulos 1997; de Giorgio et  al. 2006; Mohanty et  al. 2007; Akkas et al. 2011). An unusual case of suicidal longitudinal hemi-­ section is reported by Palanco et al. (1999). When there has been contact with the wheels, wheel tract marks along with dirt and grease contamination of the wounds are a feature. These observations may assist with confirming that the individual has been struck by the train. The body may be thrown a distance from the train during the impact and body parts may be spread over a wide area of the track as well as contaminating the engine or carriages themselves. Striking the track ballast can produce distinctive cranial bone injuries which may be mistaken as weapon injuries (Bowen 1966). The most detailed account of the morphological findings in railway-­related deaths to date is the paper of Driever et al. (2002).

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Detailing the typical blunt trauma injuries caused by aspects of the engine and carriages as well as the semi-­sharp force exerted by the wheels, the author categorises the injuries according to the speed and position of the body at the time of impact (Figure  63.4.1). The injuries associated with different collision speeds, according to Driever et  al. (2002), are shown in Table 63.4.1. A summary of the injuries sustained by impact body position is shown in Table 63.4.2.

head and trunk, are seen. The ribs and pelvis are most frequently injured, but the spine less so. The extremities are often injury free. Internally, brain contusions, liver and splenic rupture as well as avulsion of the kidney and ureters may occur. If the body is subsequently run over, then the injury pattern corresponds to those primarily over-­rolled in a lying position.

Collison with a person in an upright position

If the body is lying totally outside the track when struck, the injuries are caused by projective side elements of the train. Injuries under these circumstances are often seen to the head. There is usually a lack of extremity and extra-­cranial organ injury. Decapitation may occur when the neck is rested neck

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The body is usually thrown to the side or the track and onto the embankment. Unilateral patterned impact abrasions and localised impact fractures particularly to the axial skeleton, usually the

Collision with a person lying outside the track

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Figure 63.4.1  (a) Body of a male thrown on to the embankment after being over-­run by a train. (b) Complete amputation of both legs (thighs) due to a lying position on the track. (c) Partial amputation of the left lower leg with adherent axle grease and dirt. (d) Almost complete horizontal dismembering of a female (abdominal region and lying position).

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Other types of collisions

Train velocity

Injuries

80% of each other and can be deduced as two heterozygous genotypes; the higher peak height for allele 14 at vWA indicates allele sharing between the two mixture contributors. Mixture B has one locus with five alleles and must have at least three contributors. There is no deducible major or minor component, and this mixture can only be used for direct comparisons to a reference or other evidence. Mixture C has lower peak heights and allelic drop-­out, in this case two unlabeled peaks below the analytical threshold of 50 RFU. While major components for two contributor mixtures can be unambiguous, alleles of the minor component are often below the stochastic thresholds and can be masked by allele sharing or the presence of stutter. In order to not introduce any bias, it is important to finalize possible allele combinations (genotypes) for each mixture component blindly without first consulting reference profiles for suspected contributors. An exception to this notion is cases with intimate samples or other items where the victim’s DNA can be logically expected to be present; here, the use of a known contributor can help in resolving the foreign genotype present in the mixture (Butler 2015). Mixture interpretation becomes more difficult for degraded or low quantities of DNA. For these mixtures, one cannot always assume that all alleles belonging to the original DNA sources have been detected. Aside from unlabeled but visible peaks like in mixture C in Figure 64.4, there is also the possibility of complete allelic drop-­out with no residual signal. Comparing known references to mixtures with suspected allelic drop-­out can be difficult and may lead to a situation where the comparison is inconclusive. Probabilistic genotyping uses statistical models informed by research data on peak height variation, stutter rates, and drop-­out probabilities to infer genotypes and account for missing alleles in sample comparisons (Gill et al. 2015). See Section 64.1.5 on mixture statistics for more discussion. Mixtures also gain complexity with increasing number of contributors. Due to allele sharing, there will always be a level of uncertainty about how many individuals’ DNA are present in a mixture. By looking at the maximum allele count at any given loci, it is possible to state the minimum number of contributors, e.g. for up to four alleles at least two people must have contributed, and three for up to six alleles, but allele sharing will mask allele contributions for higher number of contributors, especially for less informative loci (J.S. Buckleton et al. 2007). Assumptions about the number of contributors are an important part of mixture interpretation and statistical evaluation. Several new computational, machine learning, or probabilistic approaches have been developed for this task. These software programs make better use of available information and have been shown to improve manual maximum allele count estimates (e.g. Swaminathan et al. 2015; Marciano and Adelman 2019).

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size in base pairs. Note that alleles for D22S1045 are 3 bp apart, for the other STRs the distance is 4 bp. The highly polymorphic SE33 locus is characterized by a large range of alleles (4.2–37) and several intermediate alleles (Butler et al. 2009). All genetic typing assays must be validated to ensure the reliability of the data. As discussed under PCR characteristics, laboratories are required to define the minimum RFU value above which peaks are identified as alleles, determine the RFU threshold for true homozygotes, and establish stutter percentages for each locus. Other scientific decisions to be made by the DNA laboratory include rules for the removal of artifacts caused by overamplified samples or poor spectral calibration. A laboratory’s STR interpretation protocol must be based on validation studies that explore the limitations of each multiplex assay and include a range of known samples representing typical casework specimen like compromised and mixed samples (SWGDAM 2015). Reporting analysts are trained to recognize artifacts, and classify and interpret complete, partial, or mixed DNA profiles in a consistent manner across a laboratory. Manual STR data interpretation strictly based on thresholds can become quite complex, and there has been a shift toward software to support an analyst’s interpretation and apply probabilistic principles to genotyping. Refer to Section 64.1.5 on mixture statistics for more information and to Butler and Willis (2020) for a list of open source or commercial software available for this purpose.

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DNA mixtures are common for intimate samples like vaginal swabs or contact traces collected from objects touched by multiple individuals. In recent years, forensic DNA casework has shifted from predominantly high-­quality DNA extracted from biological fluids like blood or semen to contact traces causing laboratories to encounter an increased number of complex mixtures. As discussed above, if one STR marker displays more than the expected two heterozygote alleles, the sample must be evaluated as a potential mixture. Examining all loci in a multiplex genotype helps to recognize stutter, unspecific electrophoresis artifacts or genetic anomalies such as a localized chromosomal rearrangement that could cause additional peaks (Butler  2015). The complexity of results for DNA mixtures varies immensely. A two-­contributor mixture with a clear major component can be very straightforward and will often allow for an unambiguous deduction of at least the major contributor genotype for database entry. The key to this type of mixture deduction is a consistent ratio between the two contributors, and the expectation that two heterozygote alleles from the same source should display similar signal strength. Under these assumptions, each locus can be evaluated for possible genotypes combinations. The use of quantitative peak height information for the deconvolution of mixtures has first been described in 1998 and has been incorporated in manual interpretation guidelines and probabilistic genotyping software (Clayton et al. 1998; Butler 2015). Figure 64.4 shows three different types of mixtures that illustrate the challenges of mixture interpretation. Mixture A has high peak heights and with three to four alleles per locus is consistent with two contributors. For locus D3, the

Allele-­specific mixture amplification A specialized approach to overcome problems with mixture interpretation, specifically the masking of and inability to detect minor components in imbalanced mixtures, is allele-­ specific amplification. Also deemed “compound biomarker profiling” the

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Figure 64.4  Examples for three different types of mixtures. Shown are two STR loci from the Globalfiler kit. Peaks are labeled with repeat number and peak height. Mixture A has high peak heights and is consistent with two contributors. Mixture B must have at least three contributors. Mixture C has lower peak heights and two unlabeled peaks not in a –­4 bp stutter position.

method takes advantage of insertion/deletion (INDEL, sometimes called deletion/insertion DIP) or SNP polymorphisms adjacent to STR loci to design primers that will only amplify the allele carrying a specific INDEL or SNP allele. A non-­STR compound biomarker type combines a deletion/insertion polymorphism with a binary SNP. Oldoni and Podini provide an overview on assay design and validation results for these DIP-­STR, DIP-­ SNP, or SNP-­STR systems (Oldoni and Podini  2019). All three assays can detect a minor component down to a ratio of 1:1000. This level of sensitivity could be a useful tool in selected cases, but

to target the minor component all possible mixture contributors must first be sequenced to identify suitable flanking INDELs or SNPs. The selected minor component INDEL or SNP allele must be unique to the mixture, which means the major contributor must be homozygote for the opposite allele. Multiple marker sets combining up to nine DIP-­STRs or eleven SNP-­STRs have been published and successfully tested on controlled mixtures and sexual assault casework samples. Another application for these markers is the detection of circulating fetal DNA in maternal plasma (Oldoni and Podini 2019).

IDENTIFICATION

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Allele frequencies are determined using unrelated test subjects from different ethnic groups. The requisite size of such a population frequency database depends on the number and distribution of alleles at a specific locus, but generally local DNA databases vary from 150 to 500 samples. Allele frequencies have been shown to differ between major ethnic groups, and separate databases are required (Butler 2015). While allele frequencies can be measured, genotype frequencies are calculated based on Mendelian inheritance rules and an expectation of random mating and Hardy Weinberg equilibrium. The concept of random mating does not realistically apply to human populations, and it is generally expected that local populations have population substructure, meaning that many individuals share some common ancestry, which will lead to an increased number of homozygote genotypes. This and other sampling factors have been evaluated to develop appropriate statistical adjustments (Balding  2005). A substructure correction factor of theta 0.1 for general populations and theta 0.3 for smaller, more isolated populations has been suggested by the US National Research Council and is widely accepted in the forensic community. This correction is

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If a suspect or other known reference has been positively associated with an unknown evidence profile, it is imperative to assign a statistical weight or significance to this association. Not doing so can be misleading to a judge or jury and fails to properly assess the risk of a false-­positive or fortuitous inclusion. The theoretical framework for the required calculations is based on the fact that DNA alleles have measurable allele frequencies and that their co-­ dominant genetic inheritance pattern leads to predictable genotype frequencies. More complex profiles like DNA mixtures are assessed using probabilistic reasoning.

i­ncorporated to correct the standard genotype calculation for heterozygote (2pq) and homozygote genotypes (p2) using an equation developed by Balding and Nichols (National Research Council  2009). Under the assumption of independence, the resulting genotype frequencies can be multiplied with each other to establish a cumulative genotype combination frequency or its inverse, the random match probability (RMP), as shown in Table 64.3. The table illustrates the components of the applicable statistical calculation for an example with eight STR loci with a final RMP value greater than 1  in 36 billion. This number increases if more loci are included. Results for a large multiplex with 20 or more loci were shown to range from 1 in 5.5 × 1023 to 1 in 6.3 × 1027 which would be in the sextillion to octillion range (Butler 2015). This calculation can only be applied to single source DNA or major component profiles and describes how often one would expect to see this particular allele combination across all loci in a population of unrelated individuals. If a case scenario involves relatives, a different value must be calculated adjusting for expected allele sharing (J.S. Buckleton, Bright et al. 2016a). The more STR loci are included and still match; the less frequent the combination across all loci is expected to occur and it becomes increasingly unlikely that a random unrelated person would also share this profile. This makes it more unlikely to obtain this DNA result if somebody else is the true source of the evidence. The random match probability is equivalent to a likelihood ratio expressing the probability of the evidence if the tested suspect is the source of the DNA versus the hypothesis/proposition that a different unrelated individual left the DNA:

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associations. As explained above, some mixtures allow for the unambiguous deduction of a major or minor component. In this case, it is possible to apply a random match probability (RMP) for the resulting DNA contributor genotype. But as soon as drop-­out, stutter, or allele sharing make the correct assignment of homozygote genotypes ambiguous, using the homozygote genotype frequency in the RMP calculation will unduly inflate the result and modified formulas are required. One approach is to expand RMP calculations to consider the presence of more than one possible genotype (J.S. Buckleton, Taylor et al. 2016). For mixtures with no deduced major or minor components, reference samples are compared to all detected alleles, and it can be difficult to determine if a specific reference is associated to a mixture or not. Only if the examiner can be sure that all possible alleles have been detected, anybody who has an allele that is not present in the mixture cannot have been a contributor and is excluded. For mixtures with high RFU values supporting the assumption that all alleles are present, it is possible to calculate a combined probability of exclusion, by combining all allele frequencies over all loci (Bieber et al. 2016). The resulting value is also called random man not excluded (RMNE) and applies to this evidence profile without making any assumptions about the underlying genotypes or number of contributors. While this can be a useful approach, it is only valid under a very defined set of circumstances, has been frequently misapplied to unsuitable samples, and does not make the best use of the data. Most types of mixtures are much better served if all profile comparisons are performed using probabilistic genotyping and likelihood ratios (Gill et al. 2006, 2012). Probabilistic genotyping programs such as EuroForMix, TrueAllele, or STRmix (more open source or commercial software options are listed by Butler and Willis (2020)) provide forensic laboratories with a ranked list of possible genotype combinations useful for database entry/searches and a likelihood ratio (LR) calculation tool for comparisons between the unknown mixture and one or more possible contributors. Earlier likelihood ratio assignment software (LRAS) programs such as LRmix or FST were “semi-­continuous” and only considered the presence or absence of allele peaks and could not infer probable genotypes. Refer to these two reviews for a brief history of probabilistic genotyping, challenges, and solutions (Gill et  al.  2015; Coble and Bright  2019). “Continuous” programs use peak height data to model possible alleles and genotype combinations by incorporating relevant parameters such as peak height variation, different types of stutter, drop-­out and drop-­in, and DNA degradation effects. Underlying model assumptions and calculations vary from program to program and can cause different results for the same mixture (Steele and Balding  2014; Taylor et  al.  2016). In addition to programming choices, the outcome of an LR calculation is affected by the choice of propositions, the local allele frequency database, and the use of population substructure corrections. With all these variables and due to the probabilistic nature of all models it has to be understood that there is no underlying “true” likelihood ratio that can be obtained (Gill et al. 2018). This concept has been a concern for court admissibility and triggered efforts to establish the reliability of LRAS.

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defense invokes an unrelated individual who happens to have the same allele combination. Numerically the prosecutor’s hypothesis is expressed a “1” reflecting the match between the evidence and the suspect at all loci. The probability of seeing the evidence if an unrelated person sharing the same genotype left the DNA is determined by establishing the appropriate profile frequency (simplified – Pr(E Hd) = 2pq × 2pq × q2 . . ..). The profile frequency determination is based on established genetic and statistical principles and will produce reasonable estimates for single DNA component profiles. To account for possible stain donors not necessarily sharing the suspect’s ethnic group, it is important to apply allele frequencies from all relevant populations and report results for more than one ethnic group (Butler 2015). Reporting the statistical weight may follow a frequentist approach and report the combined genotype frequencies as the random match probability. For example, using the result from Table 64.3, “The DNA profile of the blood stain matches the DNA profile of the defendant and this blood could have come him. If this blood did not come from the defendant, the DNA profile must match by chance. The odds of obtaining this profile from an unrelated individual of population A is approximately 1 in 36 billion.” The likelihood ratio based on the pair of propositions shown above would be reported as follows: “The DNA profile of the blood stain matches the DNA profile of the defendant and this blood could have come him. If this blood did not come from the defendant, the DNA profile must match by chance. The results of the DNA profile are approximately 36 billion times more likely if the sample originated from the defendant than if it came from a random unrelated individual of population A.” Both statements are modified from Goodwin et al. (2011). There are many equally valid statements to express the significance of a positive association, but there are also incorrect and possibly misleading expressions for the same value. A common problem is the transposition of the conditional such as stating “The probability that this evidence came from someone other than the defendant is 1  in 36 billion.” This is also called the prosecutor’s fallacy, and instead of assigning the statistical value to the evidence it now pertains to the hypothesis. Equally misleading is the assumption that if the DNA type of a crime scene stain has an estimated frequency of 1 in 3 million, then it is true that in a city of nine million, three people are expected to have this profile and could have left the stain so that the probability of the defendant having left the DNA is only one in three. This is called the defense fallacy. The error here is applying an equal opportunity of access to these theoretical three people that could be small children or may not even exist (Evett and Weir  1998). Presenting statistics in court is a complex issue and textbooks like Buckleton Bright, and Taylor (2016b) as well as the sources cited above are recommended for more detail.

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calculations for validation and casework purposes, and different approaches on how to offer this result as additional information in court have been discussed (Gill et al. 2018). Likelihood ratio calculations require a suspect as the comparison sample and a pair of competing, mutually exclusive propositions to be evaluated. A simple pair of propositions was presented for single source samples (see above). For DNA mixtures, the propositions now must address the prosecution’s explanation of the DNA result (Hp) versus the defense’s allegation (Hd). This assessment must include the number of contributors present in the mixture and can add “conditioning” DNA profiles of individuals expected to be present, e.g. a victim on a vaginal swab. The defense proposition will replace the suspect’s profile with an unknown individual and may disagree with the number of contributors and may disallow a known contributor. A typical pair of propositions for a two-­person mixture with one suspect (S) and either an unknown (U) or a known (K) contributor would be:

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H p : S + U and H d : U + U or H p : S + K and H d : U + K For more complex mixtures, the propositions add increasing number of contributors, e.g. H p : S + 2U and H d : 3U or H p : S + K + U and H d : K + 2U

It is important to note that in cases with two suspects linked to the same crime, combining both suspects in a single calculation, e.g. Hp: S1 + S2 + K and Hd: 2U + K, will yield a different LR than individual calculations and may add undue weight to one of the associations, especially if one of the suspects is a very minor component with missing alleles (Gill et al. 2018). The uncertainty surrounding the true number of contributors in a mixture causes some laboratories to offer more than one LR result for different assumptions. The defense can theoretically opt for a different number of contributors, but in general increasing the number of assumed contributors in the denominator will not be advantageous for the defendant (Gill et al. 2018). Separately optimizing the number of contributors for both competing propositions may be a good approach to resolve disagreements on this parameter (J.S. Buckleton et al. 2019). Forensic laboratories may not be aware of specific case circumstances and scenarios considered by the prosecution or defense. In that case, laboratories should select reasonable propositions to represent both viewpoints. To eliminate bias, analyst decisions on the use of conditioning profiles, the assumed number of contributors, or when to calculate LRs for multiple pairs of propositions must be guided by laboratory policies and documented in the case file. There needs to be a mechanism to reassess the data under additional propositions upon request by the defense, or if new information, e.g. a logical known contributor becomes available (Gill et al. 2018). Note that likelihood ratios for observing the evidence as explained above only pertain to the source of the DNA detected in a mixture and cannot be extended to detected body fluids or alleged activities, see Section 64.1.10 on reporting for a more detailed distinction.

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Several bodies have issued guidelines on how to validate probabilistic genotyping software (Coble, Buckleton et al. 2016; Haned et  al.  2016; SWGDAM  2016b). The software developer will be responsible for the accuracy of all statistical calculations and the appropriate choice of all modeling parameters, while the forensic laboratory must demonstrate that the software is performing as expected on a range of casework-­type samples through their internal validation, and both types of validations have been published (Coble and Bright 2019). Collaborative studies and LRAS comparisons have had encouraging results regarding reproducibility. Two studies using STRmix and EuroForMix for the same mixtures with variables like reference database and propositions kept constant generated LR values mostly in the same order of magnitude. Both studies also tested two semi-­continuous programs. Lacking peak height information these programs must consider all possible genotype combinations as equally likely and therefore calculated much lower LR values (Alladio et al. 2018; J.S. Buckleton et  al.  2018). A comparison on two open source LARs, LikeLTD and Euroformix, systematically compared modeling assumptions made in both programs and stated that despite different choices results were similar (You and Balding 2019). Courts are interested to know if a reported LR appropriately reflects the strength of the evidence in a specific case. Probabilistic genotyping can be evaluated by comparing LARS to previously accepted manual mixture interpretation results. One internal validation reports good concordance: for good quality mixtures where a person of interest had not been excluded, the software program returned LRs ranging from 8.7 × 108 to 1.8 × 1019. Values were lower, but still positive, for mixtures with peak heights below the manual allele call stochastic threshold. Of 53 manual exclusions, 51 were confirmed using probabilistic genotyping, one was inconclusive (LR = 1), and one four-­person mixture resulted in support for an inclusion (LR = 450). The latter was caused by a possible masked allele disregarded as stutter during manual interpretation (Moretti et al. 2017). Another useful test for the reliability of an LR result is the ability of the software to discriminate between true contributors and a large contingent of known non-­ contributors. Non-­contributors showing LRs supporting a positive association can be considered the false-­positive rate for a specific mixture or a set of mixtures (Gill et al. 2018). More complex mixtures have a higher incidence of fortuitous allele sharing and show more false positives, e.g. in one study on 2–5-­person mixtures only the 4–5-­person mixtures generated positive LRs for non-­contributors (0.05%), and only 0.004% of these had LRs greater than 1000 (Marsden et al. 2016). Another study deliberately selected non-­contributors with high levels of allele sharing but also found relatively few false positives, and the largest LR obtained was 120. This was different for first degree relatives, where the results would have been very misleading. For some of the four-­person mixtures simulated fathers and brothers yielded false positives with maximum LR values of 64 000 000 and 220 000 (Benschop et al. 2019). In a complex mixture or cases with low LRs, this type of non-­contributor assessment can weigh the probative value of a positive LR for a suspect comparison. Most probabilistic genotyping programs offer built-­in non-­contributor

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The term “Rapid DNA” refers to an integrated workflow starting with a biological sample being added to a cartridge/instrument combination that accommodates DNA lysis, PCR amplification, and capillary electrophoresis without user intervention in typically 90 minutes or less. Two of the currently available commercial instruments, the ANDE 6C Rapid DNA Analysis System from the ANDE Corporation and the RapidHIT ID from ThermoFisher Scientific, have a small footprint and all consumables in single use chips or cartridges that can be stored at room temperature (Salceda et al. 2017; Carney et al. 2019). This facilitates instrument placement in a crime scene van or a non-­ laboratory space like a police precinct. Data analysis is performed with expert system-­type software that produces various output files and typically flags samples as either passing, requiring review, or failing (Carney et al. 2019). The RapidLink system on the RapidHit instrument allows for remote data review (Salceda et  al.  2017). Collaborative studies and internal validations for buccal swabs have found acceptable size separation suitable for 1 bp differences and success rates of >90% for the database loci in STR multiplexes like the ANDE internal FlexPlex 27 and ThermoFisher Globalfiler Express (Ragazzo et al. 2020; Romsos et  al.  2020). Possible oral contaminants such as chewing gum, coffee, or tobacco did not inhibit the reaction, and it is possible to remove the sample from the cartridge to repeat testing in the laboratory (Wiley et al. 2017; Carney et al. 2019). The rapid DNA or “lab on a chip” approach requires more cellular material to produce results, and despite the clear advantages for time-­ sensitive investigations it is currently not recommended for low template crime scene samples and possible mixtures that could be consumed during testing (Thong et  al.  2015). It is unclear how to incorporate positive and negative controls and to make results court admissible, and the evidence may need to be retested in an accredited laboratory, which raises concerns about the exculpatory nature of result discrepancies (NDAA  2018). Nevertheless, manufacturers are already offering updated chips/ cartridges for disaster victim identification and evidence samples (Murakami et al. 2020; Turingan et al. 2020). A joint position statement by the European Network of Forensic Science Institutes and the FBI Scientific Working Group on DNA Analysis Methods outlines validation and quality assurance

Though more time-­consuming than the current PCR-­STR and especially the rapid DNA workflow, there is a range of potential improvements MPS can bring to forensic STR typing (Alonso et al. 2018; Bruijns et al. 2018; de Knijff 2019; Ballard et al. 2020). Human identification for crime scene evidence and human remains will benefit from the higher power of discrimination caused by increased allele variability and the improved performance for degraded DNA due to smaller amplicon sizes. Testing can be made more informative by simultaneous detection of other markers like lineage or identity SNPs, tissue-­specific RNA targets, and phenotype markers. On the other hand, processing references and evidence samples with MPS is slower and more expensive. Another factor holding up implementation for casework is the lack of uniform nomenclature rules and population data for new sequence variant alleles (Alonso et  al.  2017). The forensic genetics research community is working on these issues. The history of DNA sequencing and several prototypes for new sequencing chemistry and instruments are well described by these reviewers (Børsting and Morling 2015; Bruijns et al. 2018). While the terms next-­generation sequencing (NGS) and massive parallel sequencing (MPS) are often used synonymously, in reality the field has gone through multiple generations of new sequencing approaches and the term MPS is less ambiguous. The two MPS instruments currently being validated for forensic applications are the Illumina MiSeq FGx from Verogen and the Ion Torrent PGM and Ion S5 from ThermoFisher. Rather than using random primer and whole genome amplification, forensic MPS essays are based on targeted amplification with specific primers to generate libraries of tagged amplicons that become the template for clonal amplification and sequencing by synthesis (Bruijns et al. 2018). The multiplex kits being evaluated by forensic laboratories all include the database markers listed in Table  64.1, with additional autosomal STRs and other marker types added. The Verogen ForenSeq DNA Signature Prep kit has a primer set A with 27 autosomal STRs, 24 Y STRs and X STRs, and 94 identity-­informative SNPs. A performance test on reference samples found full concordance to capillary electrophoresis allele calls once stutter thresholds were adjusted (Moreno et  al.  2018). The ThermoFisher Precision ID GlobalFiler NGS STR, STR Panel v2 covers 31 autosomal STRs with one Y-­ Amelogenin and two more Y-­chromosome targets for sex determination. Again, there were some technical issues but a pilot study found 99.61% concordance with capillary electrophoresis results (Tao et  al.  2019). The PowerSeq  46  GY STR kit from Promega combines 23 autosomal STRs and 23 Y-­STRs. A study on reference samples also incorporating time-­saving automation steps found full concordance for all loci (Montano et al. 2018). The MPS process generates amplicons of various lengths and requires alignment of these sequence fragments to a known

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Massive parallel sequencing for STRs

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While STR testing via capillary electrophoresis is likely to remain the “workhorse” of human identity testing in forensic casework, it is useful to review two parallel technical developments: integrated “Rapid DNA” testing bringing human identification to police precincts and crime scenes, and massive parallel sequencing (MPS), also called next-­ generation sequencing (NGS), allowing for simultaneous analysis of STRs, single nucleotide polymorphisms (SNPs), and other DNA targets.

requirements and emphasizes the need for trained and qualified DNA analysts to be involved in data interpretation (Hares et al. 2020).

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­ opulations, e.g. Korea, Spain, US Americans, China, and many p other countries (Novroski et al. 2016; E.H. Kim et al. 2017; Barrio et  al.  2019; Devesse et  al.  2020; Z. Wang et  al.  2020). A Dutch court has accepted at least one forensic report with STR MPS data. The case was a sexual assault with the defendant’s alleles being part of a minor component partially masked through allele sharing and stutter. The MPS data added more information by resolving some of the shared alleles (Verogen 2020). Not being restrained by the number of fluorescent dyes and feasible amplicon size ranges, MPS primers can be designed to create more uniform, shorter amplicons, e.g. the PowerSeq Auto/Y panel amplifies the same loci with 129–303 bases as opposed to 72–486 bases for the same company’s PowerPlex kits (Montano et al. 2018). This results in higher STR success rates for degraded DNA (Sharma et al. 2020). Prior to a more widespread case application, assays need to be validated for sensitivity, inhibitor tolerance, error rates, and reproducibility. Sensitivity and minor component detection will depend on the agreed upon read coverage and depth interpretation thresholds, as well as the ability to identify allelic drop-­out, and distinguish true alleles from stutter and other artifacts. Current studies indicate a lower level of detection of approximately 62 pg (Alonso et  al.  2018). Characterizing MPS-­STR alleles through the longest uninterrupted stretch of the same repeat captures most of the isoallele variation and has been used as an intermediate solution to develop probabilistic genotyping algorithms for data analysis and

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r­ eference sequence. Technical terms important for MPS performance are read length, coverage, and depth. A sequence of bases is called a read with the actual number of sequences being the read length. The coverage is the number of short reads that overlap and represent a specific DNA region, while depth is the amount of reads obtained for the same sequence (Bruijns et  al.  2018). Read depth is correlated to amount and quality of tested DNA and can be used to determine the ratio between major and minor components in a mixture. Data analysis parameters must include minimum read depth requirements to distinguish true alleles from background noise and stutter (Bleka et al. 2020). Alleles with the same length but different sequences are called isoalleles. Figure  64.5 shows a three-­person mixture displaying four capillary electrophoresis peaks but six different sequences with the 19.3 and 20 peaks consisting of two isoalleles each. Each allele except for one of the 19.3 isoalleles also displays a -­4 bp stutter peak and, as expected, the longest uninterrupted stretch (LUS) is the section missing one repeat (Alonso et al. 2018). The detection of repeat-­type rearrangements distinguishes many isoalleles mostly for compound and complex STRs, e.g. D12S391 and SE33 both show more than twice the number of sequence variants than repeat number-­based alleles (Gettings et al. 2015). Including the flanking region with potential SNPs adds another level of allele discrimination for each STR allele. Multiple studies show an increase in heterozygosity for most loci across multiple

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Figure 64.5  An example for MPS results showing isoalleles in a DNA mixture. Results are for locus D12S391 after conventional STR electrophoresis with Globalfiler (section A) and MPS with Precision ID Globalfiler NGS STR panel (section B). Horizontal lines for stutter peak 19 and alleles 19.3 and 20 indicate the presence of two different sequences. Image first published by Alonso et al. (2018).

Single nucleotide and insertion/deletion polymorphisms

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Forensic geneticists have long had an interest in binary DNA markers such as single nucleotide (SNP) or insertion/deletion (INDEL) polymorphisms. The human genome contains approximately one SNP per kb of DNA sequence making SNPs an abundant marker type with alleles that due to their low mutation rate (~1  in 100 000 000) can be considered identical by descent (Butler  2012). Depending on their population distribution and genetic associations, SNPs can be informative for externally visible traits or biogeographical ancestry as described in Section 64.3. Most SNPs are biallelic with only two possible alleles and three possible genotypes, which limits the power of discrimination for human identification and means that large markers sets are required to achieve levels of statistical power similar to STRs (Butler 2012). Clinical geneticists and researchers routinely use high-­density hybridization arrays to analyze large panels of several 1000 SNPs, while forensic geneticists have preferred to explore a variety of lower throughput chemistries (Sobrino et  al.  2005). Several forensic SNP panels were developed for a single-­ base extension assay (SNapShot) that uses fluorescent detection via capillary electrophoresis and can easily be implemented in forensic laboratories (Mehta et  al.  2017). Systematic selection of autosomal identity-­ informative SNPs (iiSNPs) requires markers to be independent of each other and possess high heterozygosity and similar allele frequencies across multiple biogeographic populations (K.K. Kidd et  al.  2006; Pakstis et al. 2010). One example of a forensic iiSNP panel is the SNPforID 52plex with a mean random match probability of at least 5.0 × 10-­19 in three different populations (Sanchez et al. 2006). The low mutation rate makes identity-­informative SNPs a useful supplementary tool for resolving paternity cases with suspected STR

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mutations, while the small amplicon size for SNP alleles is an advantage for human identification involving degraded DNA. Both of these advantages could be demonstrated for the SNPforID 52plex using SNaPshot (Børsting et  al.  2008,  2013). With MPS technology, forensic iiSNP panels can now be expanded and combined with STRs for simultaneous detection (Mehta et al. 2017). The abundance of SNPs throughout the human genome allows for very specialized selections of iiSNPs, like a panel deliberately chosen for its chromosomal location in the nucleosome area, showing a characteristic pattern of intercalated histone molecules. These histone–DNA complexes are believed to be protected from enzymatic cleavage and a study on artificially degraded DNA was able to show increased typing success for the 18 nucleosome SNPs compared to other autosomal SNPs (Freire-­Aradas et al. 2012). Bi-­allelic SNPs with only three genotypes display a high degree of allele sharing which is problematic for mixture detection and interpretation. Specifically targeting triallelic and tetraallelic SNPs increases the number of genotypes and expected heterozygosity and can improve mixture recognition (Westen et  al.  2009; Phillips et  al.  2015,  2020). Another approach to increase heterozygosity is to select SNPs so close to each other that they are inherited together creating a multiallelic microhaplotype (K.K. Kidd, Pakstis et al. 2014). A microhaplotype of, for example, four binary SNPs has 16 possible alleles and 136 possible genotypes, which is still less polymorphic than STRs but there are other advantages. Like most SNPs, microhaplotypes have low mutation rates, uniform amplicon sizes, and no PCR artifacts like stutter (Oldoni et  al.  2019). Microhaplotype Sanger sequencing results for DNA mixtures were difficult to interpret, but MPS with clonal amplification can distinguish each parental chromosome with its microhaplotype or phased genotype. This makes microhaplotypes a possible tool for mixture deconvolution, where in the absence of PCR artifacts a minor component can be detected after amplifying higher amounts of DNA than is recommended for STR analysis. The presence of unique alleles can possibly help determine the number of contributors in a mixture (K.K. Kidd et al. 2017; Bennett et al. 2019). STR markers worth mentioning are Another type of non-­ identity-­informative INDELs that have characteristics similar to SNPs but can be typed in multiplexes via capillary electrophoresis because alleles differ by size. The first forensic panel of 38 binary INDELs contained insertion deletions ranging from 2–5 bp segments and showed high degrees of heterozygosity across several populations (R. Pereira et al. 2009). A commercial product, the Qiagen Investigator DIPlex, combines 30  identity-­informative INDELs with allele size differences between 4 and 15 bp. The loci could be shown to be in Hardy Weinberg equilibrium for the tested populations and have sufficient power of discrimination and detection sensitivity for forensic casework (LaRue et al. 2012). Another non-­STR marker type is based on Alu repeats, retrotransposable elements dispersed throughout the genome, and can be characterized as binary insertion/null allele polymorphisms (Brown et al. 2017). Alu repeats are abundant in the genome and provide a rich source of polymorphic sites for large panels. A commercial product, the InnoTyper 21  kit, was specifically

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mixture interpretation (Just and Irwin  2018; Bleka et  al.  2020). Regarding the final nomenclature for MPS-­STR, recommendations are insisting that data must be backward compatible to STR profiles stored in forensic databases and that analysis software must allow for the complete sequence with the maximum consensus information to be exported and saved (Parson et al. 2016). Several initiatives have made nomenclature suggestions, and collaborative efforts are underway (Ballard et al. 2020). It will be difficult to capture all of the available sequence information in a single allele name. MPS-­STR data reduced to repeat number allele calls are already accepted in some forensic databases with an annotation that additional sequence information is available (de Knijff 2019; Butler and Willis 2020). Many practical concerns regarding data compatibility, informatics workflows, data retention, as well as mixture interpretation and reporting of results still need to be resolved, but the advantages are clear and MPS for STRs will be a future forensic genetics tool (de Knijff 2019).

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pinpoints when during gastrulation the mutation must have occurred. The feasibility of this approach was confirmed for five male twin pairs and their offspring and at least one criminal case with semen evidence. For criminal casework, only the reference deep samples were analyzed using the whole genome ultra-­ sequencing approach. Once located, each SNP was confirmed through targeted Sanger sequencing for the probative sperm sample (Rolf and Krawczak 2021). Mitochondrial DNA has the potential to accumulate somatic mutations over time, which manifest as heteroplasmy and could theoretically be used to distinguish maternal relatives including siblings and identical twins, but this is complicated by the fact that mtDNA genomes are not identical within the same individual (Butler  2012). How much point heteroplasmy (PHP) is detected by whole mtDNA genome sequencing depends on sequence read depth and minimum read thresholds, and the data must also be carefully scrutinized for sequencing artifacts and heteroplasmy caused by co-­amplification of nuclear sequences homologous to mtDNA. A study on mother child pairs using a 2% calling threshold found “differentiating” PHPs in 17 (44%) and “random” PHP in 29 (52%) out of 39 pairs. A differentiating PHP had to be present in both tested tissue types (buccal and blood samples) in one family member and absent in the other, while a random PHP was called if present only in one tissue type (Holland et  al.  2018). Another study on identical twins found PHPs in blood samples for 80% of their 10 twin pairs applying a 5% threshold (Z. Wang et al. 2015). In a published case study, a combination of mtDNA genome sequencing, allele-­ specific amplification, and ultra-­deep amplicon sequencing found a single difference between the twins that was also present in the evidence and could be confirmed in multiple reference tissues (Yuan et  al.  2020). All authors agree that because the frequency of mtDNA heteroplasmy differs across tissue types, a new approach to which reference samples to test and how to interpret detected differences is needed. Epigenetic effects cause DNA methylation of specific DNA sites to change, either in correlation to age (see Section 64.3.3 for age-­related research and more background on DNA methylation) or based on health and life style habits such as exercising or smoking (Vidaki and Kayser 2018). The methylome is of clinical interest and detection tools include commercially available microarrays like the Illumina Infinitum Human Methylation27 BeadChip covering 27 578 CpG dinucleotides sites. This chip was used to screen blood from 22 pairs of monozygotic twins, and all could be distinguished based on differential methylation. The authors suggested a panel of 92 candidate CpG sites with different methylation levels for most of the 22 twin pairs for further research (C. Li et al. 2013). Another systematic search effort used an expanded version of a larger methylation BeadChip with 485 512 CpG target dinucleotides to test blood samples and buccal swabs. The authors found large sets of candidate twin differentially methylated CpG sites (tDMSs), but only a few of these were of value after subsequent validation with real-­time PCR and smaller amounts of blood or saliva mimicking forensic casework samples (Vidaki, Díez López et al. 2017; Vidaki et al. 2018). These results are ­promising but will

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designed for degraded DNA and utilizes fluorescently labeled primers to amplify short fragments of 60–125 bp. A comparative study on compromised mock casework samples and artificially degraded DNA demonstrated an average of 81% Innotyper 21 genotype recovery over only 4.2% of STR profiles (Brown et al. 2017). Large identity-­informative SNPs panels have a clear role in kinship and missing person investigations where relatedness for distant relatives needs to be resolved. Previously, the inability to reliably recognize and interpret mixtures had limited the use of stand-­alone binary allele markers in stain identification. With the advent of MPS and the ability to combine iiSNPs and STRs panels, casework samples can now be more easily subjected to SNP analysis and it remains to be seen how useful data will be for forensic evidence. Several different commercial MPS panels have been developed based on previously published iiSNPs and are being characterized by the forensic community (Bruijns et al. 2018). Selecting published SNPs is a valid strategy to ensure that each new iiSNP has allele frequencies for different populations, and the whole panel was evaluated for Hardy Weinberg equilibrium and linkage equilibrium so that random match probabilities can be calculated. Potential levels of discrimination far exceed current systems. Combing the 94 iiSNP and 28 autosomal STRs in the ForenSeq kit raised the mean RMP in four populations increased from 4.49 × 10-­38 to 1.03 × 10-­35 for the SNP panel to a range of 1.57 × 10-­74 to 8.92 × 10-­71 after including STR sequence variation (Churchill et al. 2017). Forensic validations need to address detection thresholds and reproducibility of SNP allele calling. Ballard et al. (2020) describe research on different commercial iiSNP primer panels on all three MPS platforms mentioned above. Results mostly showed good detection sensitivity and data concordance, but read coverage was often imbalanced for different markers and some SNPs consistently displayed allelic imbalance or drop-­ out (Ballard et al. 2020). More technical optimization, work on interpretation guidelines, and collaborative studies on reproducibility across laboratories and instruments are required and in progress. At the same time, another generation of MPS instruments with new challenges and/or advantages is being developed (Ballard et al. 2020).

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Molecular discrimination of identical twins Human identification with standard PCR-­STR or sequencing tools cannot distinguish between identical twins who inherit the same DNA from their parents. Differences can only arise through embryonic somatic mutations or epigenetic changes accumulating with age (Vidaki et  al.  2018). Ultra-­deep massive parallel sequencing can succeed in identifying somatic mutations, as demonstrated by a proof of principle study where five SNPs were only shared by the child and one of the identical male twins analyzed as the putative fathers. All five SNPs were present in the child’s blood and the true father’s sperm tissue. Some but not all SNPs were also detected in the father’s blood and buccal mucosa (Weber-­Lehmann et al. 2014). The presence in different tissues

require more follow-­up on how to reliably test trace amounts of DNA, account for tissue-­specific differences, and come to a statistically robust identification of twin individuals.

64.1.8  Haplotype markers and their application Genetic polymorphisms located on cellular structures other than autosomal chromosomes occupy a special place in forensic DNA testing. Y-­chromosome-­specific STR loci and SNPs are a useful tool in sexual assault cases, while sequence polymorphisms found on the mitochondrial genome are tested for samples with low nuclear DNA content such as hair shafts.

multiple genotypes (Gopinath et al. 2016). These multi peak loci and any duplications at other loci should not be confused with a chromosomal anomalies can cause loss of DNA mixture. Y-­ amplification signal through large-­scale deletions of the target areas, for example for loci like DYS458. This STR is located 1.13 Mb distant from the Amelogenin gene on the short arm of the Y-­ chromosome and has been shown to be absent for Amelogenin Y negative males (Jobling et al. 2006). Gender misidentification and loss of Y-­STR signal can complicate data analysis in cases involving male DNA mixtures. The Y-­chromosome is inherited in uniparental fashion without recombination. All Y-­STR loci are considered linked, and the product rule cannot be applied. In order to estimate the frequency of a particular Y-­STR haplotype in the human population, it is necessary to establish large databases of unrelated individuals for defined loci combinations and count how often each haplotype has been observed. The largest repository for Y-­STR haplotype frequency data is curated by the Institute of Legal Medicine and Forensic Science of the Charité in Berlin and can be accessed via YHRD (http://www.yhrd.org). The database represents 32 global metapopulations from all geographical areas. Table 64.4 lists how many haplotypes are available for different Y-­STR combinations. Each increase in the number of STRs produces higher powers of discrimination between different paternal lineages to a point where more than 90% of all haplotypes in the database occur only once (singleton) and database searches routinely cannot find an exact match (Butler 2012; Willuweit and Roewer 2015). A matching haplotype may be rare in a larger population, but in a specific case paternal inheritance combined with residency history could cause a haplotype to form a local cluster of not only close but also remote male relatives (Caliebe and Krawczak 2018). A promising approach to distinguish between male relatives and address this problem is to target rapidly mutating Y-­STR loci typically characterized by large repeat numbers (Ballantyne et al. 2010). A set of 13 rapidly mutating Y-­STRs could be shown to discriminate 27% of father–son pairs, 44% of brother and grandfather–grandson relationships, and 55% of cousins (Ballantyne et al. 2014; Adnan et  al.  2016). Some of the commercial kits listed in Table  64.4 include rapidly mutating loci. While these markers are extremely valuable for evidence associations, results must be interpreted with caution for paternity and kinship investigations (Kayser 2017). As already mentioned, Y-­STR exclusions are unambiguous; however, a match will also include all paternal relatives and an unknown number of other unrelated local male residents. For the few more common haplotypes, a population database can provide a good approximation of the paternal lineage frequency, but estimates are likely to be too conservative for singletons or haplotypes not represented in the database (M.M. Andersen et al. 2013). The YHRD database allows users to select between several different statistical calculations for haplotype frequency estimates. One is the counting method also in use for mtDNA frequency determinations (n/N) or an augmented version where the observed haplotype is added to both the observation and the database (n + 1/N + 1). In both cases, the results are corrected using a

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Y-­chromosome-­specific STR loci share their structure with autosomal STRs and were first described as being highly polymorphic in the early 1990s (Roewer et al. 1992). Early forensic applications emphasized how the absence of competing PCR product allows for successful detection of male DNA on semen positive vaginal swabs without interference of the female victim’s DNA. While for autosomal STR markers and capillary electrophoresis a minor component cannot be detected below ratios of 1:25 or 1:50, Y-­specific STRs will yield specific alleles for samples previously only showing the victim’s genotype and deliberate mixtures female DNA at a ratio of 1:2000 or higher (Roewer and Epplen 1992; Prinz et al. 1997). This has led to an application in sexual assault cases where differential lysis was either not possible or unsuccessful and autosomal results yielded none or only a few minor male DNA peaks. This approach can detect male epithelial cells in azoospermic semen, and after oral sexual assaults, and has enabled investigators to obtain probative results for semen evidence after extended postsexual assault intervals for more than 3 days (Soares-­Vieira et al. 2007; Mayntz-­Press et al. 2008). It is important to note that the Y-­chromosome with its specific STR alleles is inherited in uniparental fashion so that all male descendants from the same father are expected to share the same Y-­STR allele combination or haplotype. This means a Y-­STR result can reliably exclude a person of interest, but a positive association does not identify a single individual as the possible semen source but rather his paternal lineage (Roewer  2009). Several reviews and best practice recommendations provide valuable information on how Y-­chromosome markers support forensic applications (Gusmão et al. 2006; SWGDAM 2014; Kayser 2017; Roewer et al. 2020). As for autosomal STRs, Y-­STR testing has been optimized and made more informative by combining multiple loci in large multiplexes. Figure 64.6 displays a result for the AmpFlSTR Y Filer Plus kit. Note the presence of a single peak at most loci except for DYS385 and DYF387S1. DYS385 contains a duplicated region with two separate STR stretches that can either have the same length (resulting in one peak) or show different repeat numbers (resulting in two peaks). DYF387S1 is a multicopy marker with

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DYS3891

DYS635

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DYS627

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DYS437 150

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DYS533 330

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38 Figure 64.6  Genotyping results for a single male individual using the Y-­chromosome-­specific Yfiler Plus multiplex. All peaks are labeled with the allele corresponding to the repeat number. Red triangles indicate the allele range borders for each locus. Note the two peaks for DYS385 and DYF387S1.

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Table 64.4  Y-­STR haplotype population database size.

Minimal Haplotype (9 loci) (Kayser et al. 1997)

DYS19, DYS385a/b#, DYS389I, DYS398II, DYS390, DYS391, DYS392, DYS393

321 471

PowerPlex Y (12 loci) (Krenke et al. 2009)

All of the above and DYS437, DYS438, DYS439

280 843

AmpFlSTR Yfiler (17 loci) (Mulero et al. 2006)

All of the above and DYS448, DYS456, DYS458, DYS635, Y-­GATA-­H4

261 122

PowerPlex Y23 (23 loci) (Thompson et al. 2013)

All of the above and DYS481, DYS533, DYS549, DYS570a, DYS576a, DYS643

89 577

AmpFlSTR Yfiler Plus (27 loci) (Gopinath et al. 2016)

All of the above except DYS549 and DYS643, with added DYS449a, DYS460a, DYS518a, DYS627a, DYF387S1a/b#a

89 554

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c­onfidence interval calculation (Willuweit and Roewer  2015). Also offered are the kappa method using the number of singletons to adjust the search result (Brenner 2010) and the discrete Laplace method modeling the existence of shared ancestry clusters around central haplotypes, thus expanding the database beyond actually tested samples (M.M. Andersen et  al.  2015). Depending on regional residency history, certain Y-­haplotypes may be enriched in a region and the social circle of a matching person of interest. In these cases, an estimate of how often a paternal lineage was seen in a population database of unrelated individuals has limited value. Proposed solutions are independent of database counts and use simulations incorporating Y-­STR allele evolution and phylogenetic lineage growth models to predict the number of males sharing a haplotype (M.M. Andersen and Balding 2017; Caliebe and Krawczak  2018). These and other methods are under review. Regardless of how a frequency estimate was obtained, the preferred reporting of a positive association is the likelihood ratio method with an explicit disclaimer that paternal relatives are likely to share the same Y-­STR profile (Roewer et al. 2020). In some forensic reports, Y-­STR data may need to be combined with other genetic polymorphisms. Sexual assault cases, for example, could have partial autosomal STR results attributed to the same male individual matching the Y-­ STR profile. Both marker types are expected to be independent of each other and

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Mitochondrial DNA (mtDNA) resides outside of the cell’s nucleus, and a single cell may have hundreds or thousands of mitochondria with each carrying up to 10  mtDNA copies (Amorim et al. 2019). This increased copy number is the main reason mtDNA polymorphisms can still be detected in samples with low or degraded amounts of nuclear DNA (hair shafts or bone tissue) where autosomal testing failed. Mitochondrial DNA is inherited in uniparental fashion through the maternal line, limiting the power of discrimination to maternal lineages but making mtDNA testing a valuable tool for testing extended pedigrees and tracking human migration (Butler  2012). There are many fundamental differences between nuclear and mitochondrial DNA including the circular shape and increased mutation rate. For more details, how this affects forensic applications and a brief history of mtDNA typing, refer to Butler (2012). MtDNA polymorphisms are single nucleotide changes (SNPs) most commonly detected through DNA sequencing. The mtDNA control region, the largest non-­coding stretch of the molecule, contains three hypervariable regions that comprise 1121 bp of the complete 16 569 bp mitochondrial genome. Historically, two of these hypervariable regions, HVI (positions 16024–16365) and HVII (positions 73–340) were the main target of forensic assays, but sequencing additional areas like HVIII (340–576) was shown to increase power of discrimination and it has been recommended to target the whole control region (CR, positions 16024–576) for both casework and databasing (Lutz et  al.  2000; Andréasson et al. 2007; Parson et al. 2014). Instead of reporting all bases in a sequence, mtDNA typing results are expressed in comparison to a human reference sequence that was first established and then updated in Cambridge and is thus called the revised Cambridge Reference Sequence (rCRS) (Andrews et  al.  1999). Figure  64.7 displays the commonly used data format of only listing the differences to the rCRS. For many years, the most common test method for mtDNA SNPs was Sanger sequencing, often with various primer sets that

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# DYS385 and DYF387S1a/b each count as two loci. a  Markers with mutation rate >1% and characterized as rapidly mutating.

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can be multiplied with appropriate corrections for population substructure (J.S. Buckleton and Myers 2014). At least one scientific body requires laboratories to first establish independence in their relevant populations (SWGDAM 2014). More problematic is the interpretation of a male-­to-­male DNA mixture. Y-­STR testing can be useful and confirm the presence of multiple male individuals, but as with autosomal STRs allele sharing limits exact determinations of the number of contributors. Using peak height information some mixtures can be deconvoluted to formulate major and minor component haplotypes. Several software solutions have been suggested to establish possible haplotype contributions for non-­deducible mixtures (Wolf et  al.  2005; M.M. Andersen et al. 2015), but laboratories may deem such mixtures not suitable for comparison or limit reporting to exclusions (SWGDAM 2014). Probabilistic genotyping tools will be required to deal with complex Y-­STR mixtures and allelic drop-­out (Coble, Andersen et al. 2016).

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Number of haplotypes in YHRD database as of 18 November 2020

Multiplex system

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Figure 64.7  Mitochondrial DNA sequence alignment with the revised Cambridge reference sequence (rCRS). The highlighted position displays a base call difference between the test sample and the rCRS. This will be reported as 16193 T.

sequencing approach for forensic casework (Ballard et al. 2020). A commercially available MPS kit, PowerSeq CRM Nested System from Promega, is based on the Eichmann and Parson (2008) primer panel and was shown to be 20 times more sensitive than the FBI laboratory’s current Sanger sequencing assay (Brandhagen et  al.  2020). Other companies such as Verogen, ThermoFisher, and Qiagen also offer either control region or whole genome kits that pool hundreds of primers to generate amplicons mostly below 150 bp in size. All of the primer pairs display overlapping regions, and many primers have degenerate sequences covering known SNPs in the annealing site for a range of populations. These and other custom designed assays demonstrated good sequence concordance to Sanger sequencing data and high success rates for reference samples, forensic hair evidence, and ancient bone samples (e.g. Parson et al. 2015; Cuenca et al. 2020), for a comprehensive review see (Forsythe et al. 2020). For very old or extremely damaged samples, target enrichment through probe capture methods can improve results even more (Forsythe et al. 2020). The high level of sensitivity requires that bone and hair samples are thoroughly cleaned and decontaminated prior to analysis and that all testing is performed in a contamination-­free environment with appropriate controls. Low levels of background contamination can be considered when interpreting the data and do not necessarily invalidate the results (Carracedo et  al.  2000; Parson et  al.  2014). Data quality requirements emphasize the need to have redundant coverage of each area and sequences in both directions to recognize sequencing artifacts and not call phantom mutations The alignment of unknown samples to the

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decrease amplicons length and improve success rates for degraded samples (Forsythe et al. 2020). See Amorim et al. (2019) for case examples involving unidentified remains. Some alternative methods for HV1/HV2 and the coding regions targeted known polymorphisms through hybridization or primer extension assays (e.g. Gabriel et  al.  2003; Vallone et  al.  2004). Despite the lower density of SNPs outside the CR, analysis including the whole mtDNA genome allows for better discrimination of maternal lineages, which is one reason to start using massive parallel sequencing (MPS) technology more feasible for longer sequences for forensic mtDNA casework (Just et  al.  2015; Parsons and Coble  2001). Both with Sanger sequencing and MPS, mtDNA testing is most valuable for complicated kinship investigations and biological material with limited amounts of nuclear DNA such as bone matrix and hair shafts. The advent of DNA-­based hair comparisons provided a useful tool to evaluate microscopic hair examination results. In a study set of 97 hairs, all morphology-­ based exclusions were confirmed, but 9 of 80 positive associations were excluded by DNA thus firmly establishing mtDNA testing as the preferred method for hair analysis (Houck and Budowle 2002). Starting out with higher copy numbers makes mtDNA testing very sensitive, and for Sanger sequencing rates of obtaining full or useful partial profiles on head hair have been shown to be greater than 90%, with diminishing success for very small fragments (below 0.5 cm) or other types of body hairs (Melton et al. 2005). Adding additional primers to reduce amplicon size made full Sanger sequences feasible for even lower amounts and more degraded samples (Eichmann and Parson 2008). Massive parallel sequencing (MPS) is now slated to replace the traditional Sanger

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Due to its maternal inheritance, mtDNA is not subject to recombination, must be treated as a haplotype, and a large haplotype database is needed to generate population frequency estimates for matching mtDNA types. As for Y-­STR haplotypes, there are only a few common mtDNA haplotypes within populations of unrelated individuals with most of sequences being either rare or unique (singletons) in the database. With MPS poised to expand testing beyond the control region, it is important to also establish corresponding population databases (Irwin et al. 2011) while making sure data quality meets forensic reliability standards (Bandelt and Salas  2012).The largest publicly accessible database has been compiled by the European DNA Profiling Group and is hosted by the Institute of Legal Medicine in Innsbruck Austria. Release 13 of this EMPOP database (http:// www.empop.org) in November 2020 contains over 46 963 HVI/ HVII haplotypes, 38 361 control region, and 4289 entire mtDNA genomes from seven metapopulations (Parson and Dür  2007). EMPOP offers searches for results listing nucleotide strings and differences to rCRS, a haplogroup browser, and several quality control tools. For samples without matches in the database, the search also returns the number of nearest neighbors with up to two sequence differences. Frequency estimates are provided with augmented counting (n + 1/N + 1) or (n + 2/N + 2) with a confidence interval. More advanced statistical methods exploring the phylogenetic information for frequency estimates are in progress (Parson et al. 2014).

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rCRS must follow strict rules to accommodate insertion and deletions and generate unambiguous results. The relevant rules are incorporated in Sanger or MPS data analysis software packages, which need to be validated prior to forensic casework use. Incorrect alignment can cause SNP calling errors, but these can be detected by mapping the result to expected SNP positions for established haplotypes. This phylogenetic alignment is especially important for database samples but also recommended for casework results (Bandelt and Parson 2008; Parson et al. 2014). A phenomenon called sequence heteroplasmy challenges the interpretation of mtDNA data, especially mixture detection and associations between known and unknowns. MtDNA sequence heteroplasmy is defined as the presence of more than one mtDNA sequence type in an individual with the difference either being a single nucleotide (point heteroplasmy, PHP) or an insertion/deletion (length heteroplasmy, LHP). The occurrence of sequence heteroplasmy varies between tissue types, is based on a combination of inherited sequences and somatic mutations, and is higher for metabolically active tissues such as skeletal muscle or tissues with a developmental bottleneck like hair (Calloway et al. 2000). Single nucleotide heteroplasmy either manifests itself through a combination of two different base calls for the same position like in the famous case of the Russian Tsar or single sequence differences across multiple body tissue types or between individual hairs for the same individual (Butler 2012). Length heteroplasmy is characterized by various amounts of bases comprising a stretch of repeated bases, for example the HV1 region C-­stretch spanning nucleotide positions 16184–16193. For many individuals, a T base at position 16189 interrupts this C-­stretch. If this T is not present, polymerase replication slippage will result in loss or addition of additional cytosines and length heteroplasmy, forcing the DNA analyst to apply additional primer sets for sequence generation (Butler  2012). The same phenomena causes LHPs in HVII and HVIII (Parson et al. 2014). For Sanger sequences, individual peak heights do not reflect the ratio between a major or minor component for samples with heteroplasmy and DNA mixtures. This is different for MPS where read depth is correlated to starting DNA concentrations so that phasing and phylogenetic information can be used to deconvolute a mixture (Churchill et al. 2018). PHPs or minor base pair variants in mixtures could be called for either 1:10 or 1:100 mixtures depending on software settings and read depth required for the variant allele to be distinguished from noise (H. Kim et al. 2015; Cho et al. 2018; Churchill et al. 2018). A study on 512 individuals reported heteroplasmy rates in the control region of at least one PHP in 9.98% of individuals, which is higher than the 6% previously reported for Sanger-­type sequencing (Strobl et al. 2019). Enhanced heteroplasmy detection with deep coverage and a low variant calling threshold has the potential to even discriminate between maternal relatives (Holland et al. 2018). On the other hand, these interindividual sequence differences caused by PHPs can mislead forensic investigators, which why current intertype sequencing require more pretation guidelines for Sanger-­ than one sequence difference for an exclusion, a rule that needs to be revised for MPS data (Carracedo et al. 2000; Parson et al. 2014; SWGDAM 2019).

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64.1.9  DNA database searches Forensic DNA databases DNA typing is a powerful tool for direct comparisons between profiles obtained from evidence items and specific reference samples, but the main investigative value is based on large DNA databases containing DNA profiles from previously convicted offenders or other subjects of interest. According to a 2019 Interpol survey polling their 194  member countries, 89 reported using DNA profiling in police investigations and 70 countries have national DNA databases. These countries are storing over millions of DNA profiles, with the majority being from known individuals such as convicted offenders, arrestees, or other subjects, and a smaller percentage from crime scene evidence (Interpol 2019). Since the inception of the first national DNA databases – starting with the United Kingdom in 1995 – thousands of unknown evidence profiles have been matched to a known individual (Butler 2012). Focusing databases on persons with prior criminal convictions is justified through the documented high level of recidivism for certain crimes, and it has been reported that for high-­volume crime types in the US for every DNA conviction 7.4–7.8 additional crimes are prevented (Wickenheiser  2004; Siegel and Narveson 2009). A meta-­analysis by Struyf et al. (2019) summarizes more recent research on database effectiveness and confirms the importance of database searches for violent and

IDENTIFICATION

Explanation

Data compatibility

• DNA profiles must be based on loci shared among databases • Data must be stored in a sharable file format

Data quality

• DNA profiles must be generated in a laboratory with a quality system in place (proficiency testing, technical review, and contamination control) • All profiles must have results for a minimum number of loci • DNA mixtures are only allowed under certain criteria • Data entry or import is controlled for errors • Forensic evidence profiles must be relevant to the crime under investigation

Data protection

• Addition of DNA profiles from known individuals is governed by specific laws • Identifying information is stored separately from the genetic profiles • Genetic profiles are not based on coding regions • Penalties are attached to unauthorized name release or unauthorized additional testing • Data retention policies are in place • A legal framework for removing profiles exists • Computer networks are secure and access is limited

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­ atabases show the highest level of effectiveness in a close geod graphical area. Therefore, every state or country can maximize its database efficiency by ensuring collaboration and contributions at the local level (Butler 2012). For smaller countries and with increasingly open borders, international data exchange becomes important, e.g. in the Prüm treaty from 2005 several European countries have agreed to allow searches against each other’s databases. In addition, there is also the option to submit individual legal assistance requests to other jurisdiction through Interpol, who maintain a DNA-­database and DNA-­Gateway project for the transfer of DNA profiles between two or more countries (Butler 2012). Database searches grow increasingly complex with larger databases. Comparisons are not just a matter of finding identical profiles. Due to the presence of loci with unresolved mixtures, possible null alleles, or other factors, search stringencies must be modified to allow for potential associations of samples without 100% allele agreement. These lower search stringencies will also minimize false-­negative search results caused by errors in the DNA profiles. It is important for all database searches to not only rely on the electronic data but to also verify a database hit through a direct comparison with the original data, and if possible duplication of the reference sample. For lower stringency searches, a hit is only the first step in declaring a true match or positive association between the two samples. A review of the original crime scene DNA profile with the underlying allele peak height information may not confirm the match, and it is important to avoid premature notifications to investigative agencies or other interested parties (Butler 2012). The European Network of Forensic Science Institutes has issued a detailed review and accompanying recommendations on DNA databases (ENFSI DNA Working Group  2016). Recommendations include specific instructions on DNA-­typing-­ related issues such as entry of new alleles, the verification of hits to DNA mixtures discussed above, or ensuring quality through laboratory accreditation. The document also draws on previous statistical studies to assess the likelihood of finding fortuitous matches, which correlates to the size of the database and the expected population frequency of the specific genotype combination of a DNA profile. The number of STR loci that were successfully typed strongly affects the genotype frequency, and the agreed upon database core loci expansion addresses the need to prevent accidental matches in larger databases, especially if offender databases also contain samples from relatives (Tvedebrink et al. 2012; Hares 2015). Database profiles generated with older STR multiplexes are less discriminatory and may not be eligible for international comparisons anymore. Whenever possible, but definitely after a database match, profiles should be tested with the expanded marker set to decrease the possibility of an adventitious match and provide updated random match probabilities (ENFSI DNA Working Group 2016). It is important to note that the statistical weight of a DNA match is not weakened by the size of the database and the fact that the match was found through a database search (Balding 2005). It will be the role of the court to evaluate additional non-­ DNA factors for database matches (Gill et al. 2015).

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property crimes. The cited studies also supported the view that DNA samples from offenders should be collected as early as possible, e.g. for minor offenses and possibly immediately after arrest (Struyf et  al.  2019). Database design requires several key decisions addressing computer technology, resources, scientific approaches, and legal and ethical concerns. Table  64.5 outlines basic quality requirements for an effective but not overreaching DNA database. In general, databases contain several separate indices and searches can generate the following types of hits: “case to case,” where one evidence profile matches another which indicates that both crimes belong to a series, and “case to known,” where an evidence profile matches to a convicted offender or other subject, thus providing a name for further investigation. In addition to data pertaining to criminal investigations, many countries have databases for storing and searching DNA profiles from unidentified bodies against personal effects and family references from missing persons. National laws regulate sample eligibility and retention (Butler 2012). Published statistics for the US show that more than 85% of convicted offender hits obtained within the US CODIS (Combined DNA Index System) database occur at the state level. This trend was consistent from 2000 to 2010 and illustrates how DNA

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Table 64.5  Key quality factors for a DNA database.

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The term “partial match” refers to results of a low stringency search in a law enforcement database not showing an exact match, but a high degree of allele sharing indicating the evidentiary DNA may have been left by a relative of the convicted offender in the database. Partial matches are unintended, but the potential of identifying a perpetrator through a relative in the database has also led to the development of dedicated software for “familial searching,” a deliberate search for high levels of allele sharing (Wickenheiser 2019). For STR profiles, allele sharing cannot identify relatives beyond first or second degree relatives like parents, children, and siblings. Only SNPs with their low mutation rate can detect remote relationships and are the basis for “forensic genealogy” searches of SNP genotypes collected for private ancestry tests (Kennett 2019). Involving relatives in a criminal investigation, this way is controversial. Genetic privacy expectations must be balanced with public safety needs, and policies regulating database use have limited familial searches to unsolved serious crimes. A set of guidelines developed in California mandates additional Y-­STR typing to narrow down the pool of candidate families. Investigation procedures must ensure that no family member is stigmatized through false suspicions and the family’s reputation is not damaged (Myers et  al.  2011). Familial searching has been successfully used for years, for example identifying the “grim sleeper,” a serial killer who is believed to be responsible for 16 homicides in Los Angeles, CA, through his son who was a convicted offender (Butler 2012). A laboratory in the UK reports on 41 police investigations where familial searching led to the identification of a perpetrator or suspect. The same authors also provide information on ethics, governance, and status of familial searching in other countries (Maguire et  al.  2014). Many jurisdictions are concerned about function creep and database legitimacy and have banned familial searches. Interestingly enough, Kennett reports on a case in Florida ultimately solved with “forensic genealogy” that could have been solved earlier because the perpetrator’s brother was a convicted offender (Kennett 2019). Forensic genealogy targets direct to consumer (DTC) genetic databases originally created to enable hobby genealogists to research their genetic family background and locate possible relatives. Scientifically, testing is based on large SNP microarrays simultaneously detecting 400 000–900 000 SPNs and making inferences about relatedness by the amount of DNA sharing, specifically looking at short chromosomal stretches with linked SNP alleles identical by descent (IBD segments). This approach can potentially identify distant relatives from second to ninth cousins, but recombination will disrupt IBD segments and is random in nature which means that the amount of DNA shared varies even at the same level of relatedness. Up to 10% of third cousins and 50% of fourth cousins may not share any detectable IBD segments (Greytak et al. 2019). Forensic genealogy depends on individuals voluntarily publishing their DNA profile. Customers of companies, such as, e.g. 23andMe, AncestryDNA, MyHeritage, or

FamilyTreeDNA (FTDNA), are given access to each company’s pool of SNP genotypes to search for possible relatives and can make their own profile available to be searched by others. A separate website, GEDmatch, accepts personal data uploads from different DTC ancestry tests and offers different privacy options. Customers can decide to have their profile public, which means they not only see their own search results, namely which other customers share DNA with their own sample, but also vice versa (Kennett  2019). With new customers adding data every day, a contact request from a possible relative may come years later. For one of the first successful forensic genealogy cases, the Golden State Killer case, investigators posed as private customers and uploaded SNP microarray data for a sexual assault kit sample as if it was a family reference. After the publicity surrounding the arrest in this and other cases, both GEDmatch and FTDNA officially agreed to law enforcement use of their databases. Currently, several forensic providers offer SNP microarray testing for samples in missing person cases and violent crimes, and new microarray assays are being optimized for forensic evidence typing (Kennett 2019). A search result pointing toward a candidate family always needs to be investigated through public registration records to identify an individual of the right age that could have had access to the crime scene at the time. Finally, a standard STR test must confirm that the person of interest could have been the source of the evidence DNA. See Greytak et al. (2019) for a series of case examples. Genealogy as a hobby is most popular in the USA, Australia, and United Kingdom. A simulation study of the 1.28  million MyHeritage database revealed that for 60% of Americans of European descent, a database search would find a third cousin or closer relative (Erlich et al. 2018). This data collection affects everybody’s genetic privacy and raises a series of data security and ethical concerns. While FTDNA and GEDmatch now ask their customers for permission to share data with law enforcement, this decision does not affect only them but also their relatives. Working on genetic pedigrees always has the risk of uncovering misattributed paternities or undisclosed adoptions, potentially misleading to the investigator and being harmful to families (Kennett  2019). Like with familial searching, DNA data may incriminate innocent individuals, and whole families may become subject to intrusive police investigations. Nevertheless, public surveys have revealed a high level of public support (>70%) of allowing law enforcement to search DTC databases for violent crimes, missing person cases, and crimes against children. A framework for responsible use of this tool needs to be established and could for example require a court or commission to decide on the justification of a DTC database search (Guerrini et al. 2018).

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64.1.10  DNA result reporting and testimony Forensic DNA reports and testimony must follow the same high-­ quality standards guiding analytical testing. The main goal is to clearly communicate all results in an unbiased manner not

IDENTIFICATION

Table 64.6  Evidence hierarchy of propositions with DNA examples. Terminology

Examples

III

Offense

Mr. B. raped Ms. A. Mr. B. had consensual intercourse with Ms. A.

II

Activity

Mr. B. had sexual intercourse with Ms. A. Mr. B. had only social interactions with Ms. A.

I

Source

The semen came from Mr. B. The semen came from some other man

Sub-­source

The DNA came from Mr. B. The DNA came from some other man.

Sub-­sub-­source

The major component of the DNA mixture came from Mr. B The major component of the DNA mixture came from some other man

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­ verstating their significance. Before deciding on a report foro mat, it is useful to consider the different levels of concern in a forensic case and how DNA results can contribute. Table  64.6 shows a DNA version of the hierarchy of propositions that investigators and the court are interested in, modified from Cook et  al. (1998), Evett et  al. (2000), and Taylor et  al. (2014). The highest level “offense” pertains to guilt or innocence, and forensic geneticists typically do not contribute here, but other experts may be able to testify to injuries or physical damage observed at the crime scene. DNA reports traditionally address either the source of a body fluid (source level) or, if lacking a body fluid attribution, the source of the DNA (sub-­source or sub-­sub-­ source for mixtures). A source level conclusion needs to be considered with care and only applies if the evidence is a single source sample with a confirmatory body fluid identification (Gill et al. 2018). As soon as there is an indication of a mixture, source level reports can be very misleading. There is currently no mechanism to determine which individual contributed each body fluid, but emerging MPS mRNA sequencing may be able to provide this information in the future (Ingold et al. 2020). Sub-­ source reporting and sub-­sub-­source reporting are appropriate in the investigative phase and for cases processed for DNA database entry that do not have a suspect. After an arrest has been made, reporting only on the DNA result leaves many questions open and there is a risk that courts misinterpret the sub-­source level association and apply the statistical weight assigned to obtaining the DNA match to the alleged criminal activity (Biedermann et  al.  2016). Accordingly, there is an increasing demand for DNA reports to address the activity level of the hierarchy of propositions and provide an assessment of how the DNA could have been deposited on an item (Gill et al. 2018, 2020). The European Network of Forensic Science Institute has issued reporting guidelines asking for evaluative reports on likely activity scenarios whenever possible, with technical reports on the source or sub-­source levels to be limited to cases with not enough information for specified propositions (ENFSI 2015).

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The activity connected to the deposit of an evidentiary DNA sample determines the probative nature of a DNA result, and ­corresponding assumptions have always guided crime scene processing, sample triaging, database upload, and case prosecution. One example here is the expectation of a husband’s DNA to be present on various items in a shared house, meaning evidence collection will differ between a domestic violence or a home invasion crime scene. Aside from preexisting DNA on household items, weapons, tools, and clothing, another concern affecting probative value is indirect transfer. Especially for trace DNA not associated with a body fluid, there may be a plausible scenario how DNA could have been transferred to the evidence without the matching DNA source being at the crime scene. The need to distinguish between unrelated and probative DNA traces has triggered a large body of research on which type of DNA is deposited by touching, the prevalence and persistence of preexisting DNA, the effectiveness of different modes of DNA transfer, and the overall success in recovering trace DNA (Burrill et al. 2019; Gosch and Courts 2019; van Oorschot et al. 2019). All of these factors, termed TPPR for transfer, prevalence, persistence, and recovery, also apply to biological fluids and have an effect on where and how much DNA is found at a crime scene and can update competing propositions on the alleged activity. DNA left behind after handling an item derives from a combination of shed skin cells, sebaceous secretions or other body fluids picked up through face touching and cell-­free DNA present, e.g. in sweat. This biological material can be either from oneself or for unwashed hands, non-­self-­DNA transferred from other surfaces and persons (Szkuta et al. 2017; Burrill et al. 2019). How much DNA individuals leave behind, their “shedding propensity,” varies considerably with some very low and others very high, but mostly intermediate shedders (Daly et  al.  2012; Kanokwongnuwut et al. 2018). The probability of transferring DNA also depends on evidence surface characteristics and the type and duration of contact. Rough surfaces, more friction, and extended contact will generally result in higher DNA deposits. For indirect transfer, it is easier for wet or moist body fluids to transfer from one object to another. For trace DNA, a porous substrate-­like fabric will likely pick up DNA from a smooth surface, but not vice versa (van Oorschot et al. 2019). Prevalence and persistence have been studied for example by testing for foreign DNA under fingernails, sampling frequently touched common surfaces, or testing clothing and bedding after having been laundered. The finding that clothing in a washing machine can transfer spermatozoa and detectable amounts of DNA should be taken into account, e.g. when dealing with domestic child abuse cases (Butler and Willis  2020). The recovery aspect becomes important when transfer rates generated by another laboratory are being used to assess the probability of transfer in a specific case. How well DNA is recovered and detected depends on the different laboratory protocols. While this is a well-­documented concern, in a real application some uncertainty in the data is acceptable and may not have a major impact on the likelihood of one proposition over the other (Biedermann et al. 2016). One collaborative study on transfer, persistence, and recovery on touched cable ties allowed

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64.2  Paternity Testing Ruediger Lessig and Mechthild Prinz

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Paternity and kinship testing are based on fundamental laws of genetics and have a long history of research and casework. The basis for this type of testing was established when Landsteiner first described the ABO blood group polymorphisms in 1901. It took several years for serological marker results to be accepted as evidence in court, but subsequently polymorphic enzymes and other proteins present in blood became an important research topic and the standard technique for kinship analysis until the late 1990s (Patzelt 2004; Adams 2008). The evaluation of DNA polymorphisms for relationship testing started immediately after Alec Jeffreys first published his findings on human minisatellite polymorphisms in 1985 (Jeffreys, Wilson et  al. 1985). The first case where this new test was applied was an immigration case, where it had not been possible to determine if the tested woman was the mother or the aunt of the child in question. By testing the whole family including several assumed siblings for minisatellite restriction fragment length polymorphisms (RFLP), the woman could be shown to be the mother (Jeffreys, Brookfield et al. 1985). Allele designations and biostatistical treatment for multi-­locus and single locus RFLP DNA polymorphisms remained a challenge and, as for identity testing, single locus STR markers were shown to be more suitable for routine paternity testing (Henke et al. 1999). Compared to the previous serological markers, DNA polymorphisms are more abundant in the genome, as non-­coding regions generally more polymorphic, can be typed using a single electrophoretic system, and do not require a blood sample as the test material. The ability to test saliva and buccal swabs as the substrate allows for easier collection by non-­medical personnel and is suitable for newborn children, where the collection of an intravenous blood sample is not always possible. Unfortunately, this also allows for clandestine sample collection and secret paternity test submissions without the knowledge of all parties involved. Tests are also sensitive enough to detect circulating fetal DNA for non-­invasive prenatal paternity testing (Wagner et al. 2009). This raises many ethical issues that are discussed below. Overall, the question of disputed paternity has been an issue throughout human history. This is a multifaceted societal topic and genetic disease, and genealogy studies have shown a percentage of non-­ paternity of approximately 1–2% per generation (Larmuseau et  al.  2013). This uncertainty in biological family trees affects many areas like historical lineage studies, mass disaster victim identification, forensic genealogy, and family studies on genetic disease. All previously discussed types of DNA polymorphisms also find an application in relationship testing: • Autosomal polymorphic microsatellites, short tandem repeats (STR). • Autosomal diallelic single-­ nucleotide or insertion/deletion polymorphisms (SNP/INDEL). • Gonosomal polymorphic microsatellites, Y-­or X-­STRs.

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their participants to use their own protocols, and while they all had different DNA detection rates, the final LRs for the tested propositions were very similar. The effort to generate data to be used in activity level assessment includes simulated case studies re-­enacting scenarios proposed in real investigations and systematic exploration of individual factors like, e.g. sweating on transfer probabilities (Fonnelop et al. 2017; Samie et al. 2019). Gosch and Courts have created a database of relevant studies and with others call for harmonization and general availability of TPPR research (Gill et al. 2018; Gosch and Courts 2019; van Oorschot et al. 2019). The question of how the DNA got there, will be asked in court and a formalized approach of addressing this through mutually exclusive pairs of propositions devised in consultation with the prosecution and defense is superior to offering speculative explanations on the witness stand. Explaining the results after the fact can result in an open-­ended list of speculative opinions that are not limited by known circumstances and cannot be tested. Propositions on the other hand should be formulated independent of the result and must have a stated framework of circumstances and assumptions (Evett et al. 2000). Even if the defense is not sharing their version of the case, it should be possible to assess reasonable scenarios. Both ENFSI and the ISFG provide useful guidance on how to formulate propositions and incorporate experimental data in LR calculations, see also, e.g. Gittelson et  al. (ENFSI  2015; Gittelson et  al.  2016; Gill et  al.  2018). Bayesian Networks (BNs) are a useful framework for complex situations requiring the assessment of multiple variables, and several worked examples have been published (e.g. Biedermann and Taroni 2012; Gill et al. 2020). Again, empirical data from TPPR research are the basis for the alleged activity assessment, and the applicability of different studies from other laboratories has been questioned. The field is working on addressing these concerns, and it may be possible to reduce the complexity of the calculation through simulation studies that can identify the variables with the most impact (Samie et al. 2020). Jurors and judges will need guidance on how to understand the science behind DNA test results and the meaning of a likelihood ratio. DNA experts should avoid jargon but also not oversimplify their findings. It is tempting to use a verbal equivalent for likelihood ratio results, e.g. moderate, strong, very strong support for a proposition, and several verbal scales have been suggested. This can be misunderstood, especially for weak evidence, and it is recommended to always introduce the numeric expression at the same time (Gill et al. 2020). When asked how a result could have been affected by laboratory error or contamination, experts must make an effort of an honest assessment. This can be very case specific, clearly an isolated DNA result seen only once at the scene has a higher risk of having been introduced erroneously than a series of samples with the same profile obtained from multiple items. Bayesian networks as described above may be a useful tool here (Kloosterman et al. 2014). Overall, it is the responsibility of the expert to educate the court.

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It is important to only use polymorphisms with well-­characterized chromosomal locations and population-­specific databases that have been tested for Hardy Weinberg equilibrium (HWE) and linkage disequilibrium (P.M. Schneider  2007). Slight deviations from independence will occasionally be observed for individual loci, which may be caused by the test population being too small. If loci are located on the same chromosome and known to be linked, this has to be considered in the calculation. Laboratory setup and report writing for paternity laboratories should follow the recommendations of the International Society for Forensic Genetics (ISFG) (Morling et al. 2002) and relevant accreditation bodies, e.g. the American Association of Bloodbanks (AABB 2019). It is recommended to select markers with an average combined exclusion chance of more than 0.9999. The minimum number of loci requirement evolved from at least 12 to higher numbers now obtainable with the currently available large multiplex kits (see Table 64.1). For STR markers, three or more inconsistencies for loci on different chromosomes allow a statement of exclusion of the assumed lineage. In any situation with less than three exclusions, a mutation event has to be considered and additional polymorphisms such as more STRs, bi-­allelic markers, or if applicable, gonosomal systems must be tested to get a reliable result. The suspected mutation has to be incorporated in the biostatistical calculation (Brenner  2004; Gjertson et  al.  2007). Mutation rates for autosomal STR loci have been shown to be locus and allele specific and fall mostly in the 0.1– 0.4% range (Butler  2015). Mutations are caused by replication slippage and, much like stutter, are dependent on the longest uninterrupted stretch of same sequence repeat units. In a large study of over 45 000 paternity trios, 97% of the mutations identistep mutations, with one repeat difference fied were single-­ between the parental and the offspring allele (Sun et  al.  2014). Mutation rates for SNPs are low enough (approximately 1 in 100 000 000) that alleles are considered to be identical by descent (Butler 2012). Another seemingly genetic inconsistency that can affect both STRs and biallelic markers are null alleles caused by polymorphic single nucleotide positions in the primer annealing site (see factors influencing PCR results in Table  64.2). Null alleles, also called silent alleles, will result in a false homozygote genotype, are generally rare, and can be prevented by redundant primer sets in the test assay (Butler  2012). In a paternity scenario, a finding

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where the child and the alleged father are opposite homozygotes, the transmitted allele could be silent and not detected. The conflict can be resolved by retesting with different primers. If that is not possible, the assumed explanation for the inconsistent result must be included in the biostatistical evaluation (Gjertson et al. 2007). Markers with lower mutation rates like SNPs or INDELs have been shown to successfully resolve inconclusive paternity results caused by mutations (Pinto et al. 2013). A study on 50 paternity trios with a panel of 40 INDELs achieved a mean posterior probability of paternity of 99.994% and a power of exclusion similar to the 13 CODIS STRs (Pimenta and Pena 2010). It is important to note that despite the advantage of the lower mutation rate a smaller panel based solely on SNPs may not be sufficient for many more complicated kinship cases (Amorim and Pereira  2005). As described in the previous section, several well-­characterized large panels of identity-­informative SNPs or INDELs are now available for both electrophoresis and massive parallel sequencing instruments. The already mentioned SNPfor ID 52 plex was tested for paternity testing, and none of the 124 investigated trios showed a mutation (Børsting et al. 2008). Massive parallel sequencing offers the most information in a single test. A panel of 90 autosomal and 34 Y-­chromosome SNPs was successfully validated for relationship testing in an ISO-­accredited laboratory (Buchard et al. 2016). MPS also enables phased detection, meaning parental chromosomes can be distinguished from each other, of linked SNPs forming a microhaplotype. With a low mutation rate and much higher power of discrimination, this marker type is a useful tool for relationship testing (K.K. Kidd, Pakstis et al. 2014). Large SNP, INDEL, or microhaplotype panels are especially powerful in deficiency cases and can resolve extended pedigrees (Lareu et al. 2012).

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polymorphisms (SNP/INDEL). • Mitochondrial DNA sequence polymorphisms. It is important to note that no marker system should be used for kinship testing until a sufficient amount of data is available. One recommendation asks for data on a minimum of 500 meioses to establish relevant parameters such as Mendelian inheritance, mutation rates, occurrence of silent alleles, and allele or haplotype frequencies (P.M. Schneider 2007).

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64.2.2  Male and female lineage markers Gonosomal markers also offer advantages in deficiency cases where one or both parents are unavailable or unwilling to be tested. Second degree relatives such as grand parents or siblings may not show enough allele sharing for independently inherited autosomal STR or SNP alleles. The Y-­and X-­chromosomes both contain STR loci that could be shown to be as polymorphic as autosomal STRs. These gonosomal markers provide additional information about male or female relationships and have been successfully applied to routine kinship testing and historical investigations (Diegoli 2015; Kayser 2017; I. Gomes et al. 2020). determining region on the Y-­ chromosome (SRY) The sex-­ codes for the testis-­determining factor and is thus responsible for human maleness; this chromosome is uniparentally inherited only from father to son. In contrast to other chromosomes, the Y-­chromosome exists in a hemizygote state without an equivalent counterpart and contains only a small recombining region. All commonly typed Y-­STRs are located in the non-­recombining areas and are inherited as a haplotype. Disregarding mutations, it is expected that father and son carry the same allele at each Y-­STR location, and this haplotype can be traced for the paternal line

Amelogenin 22

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over multiple generations. Mutations for most Y-­STRs are similar to autosomal STRs and will cause mismatches when testing multiple loci and multiple generations (Decker et al. 2008; Goedbloed et  al.  2009). Note that Y-­chromosome haplotypes are generally not informative if other family members in the paternal lineage have to be considered as putative fathers, except where rapidly mutating Y-­STRs were able to differentiate between male relatives (Ballantyne et al. 2012). Y-­chromosomal DNA is of course also home to many SNPs locations that characterize lineage-­specific haplotypes and can be applied to historical investigations and modern kinship testing (Onofri et al. 2006; Kayser 2017). X-­chromosomal markers can be informative to follow the female or male line if female children are included in the investigation. This type of marker is not commonly used in forensic stain analysis and has not been described in this section above. Female individuals carry homologous X-­chromosomes, similar to an autosomal chromosome pair. Only one X-­chromosome is actively transcribed for gene expression and all additional copies are silenced which explains why monosomies, trisomies, and polysomies of the X-­chromosome are still compatible with life. Unexpected and undetected aberrant gonosomal karyotypes may occur in the offspring, thereby affecting the accuracy of kinship testing using these markers. Figure  64.8 shows a chromosomal map of several X-­STRs (ChrX-­STR.org 2020). With male individuals carrying only one copy, the X-­chromosome has reduced rates of recombination, and STRs in close proximity of each other must be considered linked (I. Gomes et  al.  2020). The X-­chromosome STRs shown in Figure 64.8 have been assigned to several linkage groups. Like autosomal STRs, individual X-­STRs show a range of mutation rates, mostly around 0.001%, with the majority being one-­step mutations. A recent study found one locus (DXS10135) with a higher mutation rate (0.01%), which disqualifies this locus for kinship investigations. The same authors also report that the majority of observed mutations (75%) was of paternal origin making the paternal mutation rate 5.2 higher than the overall (Pinto et al. 2020). Paternity cases involving the common trio constellation of mother, offspring, and alleged father can usually be solved with a set of autosomal STRs alone and do not require any additional or alternative markers. However, in paternity cases involving close blood relatives as alternative alleged fathers, the exclusion power of autosomal STRs is substantially decreased and X-­based STRs or INDELs may be superior (C. Gomes et al. 2012). This can help solve cases of attempted fraud, where for example the brother of the alleged father poses as his brother during sample collection. X-­chromosomal alleles are also not inherited from father to son and could resolve paternity cases where the two alleged fathers have this relationship. Brothers, in contrast, share a given maternal X-­allele with a probability of 0.5, which equals the probability that two alleles are shared identical by descent (IBD) for autosomal loci. For three unlinked loci, the chance to be identical by descent would be 0.5 × 3 = 0.125. However, when the markers are closely linked, they do not segregate independently and would instead represent a single haplotype that is again shared with a probability approaching 0.5 (Szibor et  al.  2003). For a detailed

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DXS7424 DXS101 DXS6797 DXS7133 GATA172D05 DXS9908 DXS7127 DXS8377 DXS10011

HPRTB DXS10101 DXS10103 DXS10146 DXS10134 DXS10147 DXS7423

Figure 64.8  X-­chromosome ideogram with X-­STR locations and linkage groups. For X-­chromosome STRs, genetic distance and recombination rates are too low to ensure independent inheritance. Source: Image courtesy of Jeanette Edelmann.

table of IBD allele sharing probabilities for different degrees of relationships, see Gomes et al. (2020). The major advantage of X-­markers arises in deficiency paternity cases, e.g. when a sample from a putative father is not available and DNA from paternal relatives has to be analyzed instead. When female individuals have the same father, they also share the same paternal X-­chromosome. An investigation of X-­markers of two sisters or half-­sisters can exclude paternity, namely through the presence of four different alleles or haplotypes, even when none of the parents is available for testing. Autosomal markers

Marker

Child 1

LG I

DXS6807 DXS10148 DXS10135a DXS8378

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14 25.1 26 12

11 25.1 19 10

14 25.1 24 11

11 24.1 23 11

14 25.1 26 12

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DXS7132 DXS10079 DXS10074

12 21 16

14 22 17

12 21 16

14 22 17

12 21 16

14 22 17

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DXS10159 DXS10161 DXS10162 DXS10163 DXS10165 DXS10164

25 25 17 B17 20 10

25 26 21 B17 20 12

25 25 17 B17 20 10

25 26 21 B17 20 12

25 25 17 B17 20 10

25 26 21 B17 20 12

Xq21

DXS6800 DXS6801 DXS6809 DXS6789

16 11 31 21

16 12 33 22

16 11 31 21

16 12 33 22

16 11 31 21

16 12 33 22

Xq22

DXS7424 DXS101 172D05

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16 25 11

13 18 8

16 25 11

13 18 8

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DXS10103 HPRTB DXS10101

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18 14 32

18 13 29.2

19 14 32

18 13 29.2

19 14 32

DXS10146 DXS10134 DXS10147 DXS7423

28 34 8 14

39.2 36 9 15

28 34 8 15

28 36 8 15

28 34 8 14

39.2 36 8 15

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64.2.3 Biostatistics The underlying principle of STR-­based paternity testing is the independent co-­dominant inheritance of the alleles. In a regular trio case, this means that each allele not inherited from the mother, the obligate paternal allele, must be present in the DNA

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Table 64.7  X-­chromosome typing results in sib ship case.

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cannot provide such information. A positive proof of paternity is also possible without parental genotype information but is generally less reliable because sisters usually inherit only partially matching haplotypes from their mother. As will be shown below, the co-­inheritance of two identical maternal X-­chromosomes without recombination is not impossible, but rare. For mother– daughter relationships testing, X-­STRs or SNPs are equivalent to autosomal markers and do not provide any specific advantage. Testing mother–son kinship, however, is more efficiently performed using X-­STRs. The exclusion chance in such cases is identical within father–daughter tests (Szibor et  al.  2003). X-­chromosome SNPs and INDELs are also organized in linkage groups and must be interpreted accordingly, but then can become a very useful test system for extended kinships and immigration cases (Tomas et al. 2010; I. Gomes et al. 2020). The following example illustrates an application of X-­chromosomal STR markers to a case with three putative sisters with no parents. Over the time span of 3 years, three female children were found in a baby hatch of a hospital. The youth welfare office in cooperation with the adoptive parents wanted to know if the three girls were full siblings, half siblings, or unrelated. Expecting that autosomal STR typing alone would not provide enough information, the children were also tested using the 12 X-­STR loci of the Investigator Argus X-­12  kit (Qiagen) and an additional set of 15 not commercially available X-­STRs. The results are presented in Table 64.7. Table 64.8 lists the results and the different hypotheses considered for the biostatistical calculation. In summary, according to all results  – autosomal STRs, X-­STRs, and mtDNA – it had to be concluded that the three girls are most likely full siblings (Immel et al. 2011). The first set of recommendations on X-­chromosome applications in forensic science was published in 2017 (Tillmar et al. 2017). Mitochondrial DNA polymorphisms provide a marker set for the female lineage of a family tree. The circular mitochondrial DNA (mtDNA) genome is not located in the cell nucleus, but in the mitochondria in the cell plasma. Sperm heads do not contain plasma and carry mitochondria so that mtDNA genomes are transmitted uniparental maternal fashion solely via oocytes (Butler 2012). Again, this inheritance mode through the female line allows for kinship determinations over multiple generations as demonstrated by the identification of the family of the Russian Tsar (Coble et  al.  2009). As a female lineage marker, mtDNA cannot be applied to questions of paternity but can helpful in complex kinship cases or serve as an additional technique in war and disaster victim identification efforts (Prinz et al. 2007).

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Table 64.8  Relevant hypotheses in sib ship case with results. Hypotheses

LR result

H1

All three children are siblings

92.6 billion

H2

Children 1 and 2 are siblings and child 3 is unrelated

249 million

H3

Children 2 and 3 are siblings and child 1 is unrelated

7.33 million

H4

1 and 3 are siblings and children 2 is unrelated

5.07

H5

All the children are unrelated

1

profile of the biological father. If one of these prerequisite alleles is missing in the tested putative father and a mutation event is ruled out, this man must be excluded (Butler 2015). In a duo case, where the mother is not available for testing, any information about maternal origin is missing and all of the child’s alleles must be considered as potential paternal alleles. For this scenario, it can be difficult to reach the desired statistical paternity probability threshold (for example 0.9999) for a determination of paternity, especially if the results involve potential mutations or many alleles with a high population frequency (Poetsch et  al.  2013).

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This can be compensated with an increase in the number of tested polymorphisms, where the additional loci should be selected based on their specific power of exclusion, stability, and possible mode of inheritance. Likelihood ratio principles are the recommended approach to the biostatistical assessment of a paternity inclusion and the evaluation starts with hypothesis building (Gjertson et al. 2007). For a standard trio case, the competing mutually, exclusive hypotheses are as follows: H1: The tested man is the father. H2: A random man unrelated to the tested man is the father.

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As described by Essen–Moeller in 1938, these two hypotheses are evaluated using the allele frequencies of all detected paternal alleles and the likelihood ratio is formulated as follows: LR = Y/X Essen–Moeller converted this as follows:

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Other scenarios and subsequent calculation could involve sibship or similar kinship questions. The DNA Commission of the International Society for Forensic Genetics (ISFG) describes how to treat observed mutations and silent alleles (Gjertson et  al.  2007). Routine calculations assume that putative fathers are not related to each other or the mother and formulas must be adjusted for incest cases or related fathers (e.g. Evett and Weir  1998). Multiple academic, freeware, or commercially available software programs have been developed to handle kinship calculations, for example DNA-­View, familias, or EasyPA and others (see Butler  2015, p.  362, for more details). In addition to these specialized programs, companies such as SoftGenetics, Qiagen, or Biotype have added paternity modules to their STR analysis software. From a quality assurance point of view, it is important to carefully evaluate and validate a program’s performance (Drábek  2009). Likelihood ratios obtained for gonosomal markers such a Y-­STR, Y-­SNPs and mtDNA, and X-­chromosome typing results can be combined with autosomal result but this requires special considerations of population substructure and possible linkage disequilibrium between the marker types (J.S. Buckleton and Myers 2014; Tillmar et al. 2017).

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based on the family tree. For example, the list could be the following: H1: The tested man is the biological father of the child. H2: The brother of the tested man is the biological father of the child. H3: A random man unrelated to the tested man is the biological father of the child.

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This is equivalent to the paternity index commonly used in English-­ speaking countries (Butler  2015): paternity index PI = X/Y. Both values have the same information content and under the assumption of a prior probability can be converted into the posterior probability or probability of paternity. The prior probability, or prior odds, relates to the probability of paternity based on non-­genetic evidence and is routinely set to 1:1 or 50% for mostly pragmatic reasons. The assumption is that the tested man has a 50% chance to be the true father, which is somewhat neutral but is not universally endorsed. It is important to note that these prior odds are not applicable in DVI situations and can lead to an erroneous view of the evidence (e.g. Brenner 2006). The prior probability can be changed if the alleged father is a relative of a man who could also be the true father (e.g. cousin, uncle, etc.). Using the Bayes theorem, the prior probability and the values for X and Y result in the posterior probability as follows: Probability of paternity W

X /X Y.

The exclusion probability is the reverse of the probability of paternity:

Exclusion probability A 1 W .

It is not necessary to restrict the number of well-­defined mutually exclusive hypotheses to two scenarios. In more complex cases, involving relatives or deficiency cases with untested family members, it is possible to develop multiple hypotheses

64.2.4  Quality standards and ethical considerations Paternity testing is highly regulated business and many countries have established mandatory guidelines and accreditation programs. For European member countries, genetic testing for the determination of parentage falls under a 2008 regulation about accreditation and surveillance relating to the marketing of products (European Union  2008). Other countries like the USA have established specific national standards for laboratories to follow (AABB  2019). The basis for reliable parentage testing is a good-­quality management system covering scientific staff qualifications, laboratory best practices, proficiency testing, documentation, and reporting as required by ISO standard 17025 (Morling et  al.  2002). One quality requirement, which has additional legal implications, is a procedure guaranteeing the identity of the tested subjects and the traceability of the collected samples. This is routinely achieved by controlled sample collection with designated personnel checking passports or identity cards, taking photos and fingerprints of all sample donors, and starting the chain of custody. As already mentioned, one of the advantages of DNA-­based paternity testing is the ability to use saliva on buccal swabs instead of venous blood

IDENTIFICATION

64.3  Forensic DNA Phenotyping and Biogeographical Ancestry Determination

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Connecting the genotype to a phenotype for DNA found at a crime scene is the goal of forensic DNA phenotyping (FDP). Each cell’s DNA contains information needed to determine the appearance and all of the functions of our body. Testing for DNA polymorphisms is a powerful tool for associating a person of interest to a case but can only lead to an unknown perpetrator, if this individual or possibly a family member is stored in a DNA database. The ability to test the same biological evidence to create a suspect’s physical description adds a new tool to the investigation. Several recent review articles illustrate how much progress has been made in this area (de Cerqueira et al. 2016; Kayser 2015; Seo et al. 2017; Quillen et al. 2019; P.M. Schneider et al. 2019). FDP is benefiting from large genome-­wide association studies, technical advances in high-­volume sequencing, and continuing software development, but exact mechanisms of how the known genetic factors contribute to human appearance are still unknown. Many externally visible characteristics (EVCs) are believed to be multigenic and multifactorial traits, meaning a single associated gene will only contribute a small amount to the observed variation. Epigenetic regulation, gene interactions, and external factors such as nutrition, health, and age also have an effect and modify correlations between genotypes and forensic phenotypes. Note that DNA-­based biogeographic ancestry (BGA) determination is sometimes considered to be predictive of appearance. This can be very misleading, especially for individuals with mixed ancestry, and BGA testing should therefore not be considered to be part of FDP (Kayser 2015; Phillips 2015). Ancestry information in itself can provide valuable investigative leads and is discussed below. Reliable DNA targets for FPD should be universally applicable and be chosen to be independent of the ancestral background of the test subject (Kayser 2015).

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samples, and buccal swabs have become the preferred sample type. It is recommended to always take more than one swab so that results can be confirmed using the other sample. The ease of collection has caused many paternity laboratories to offer home collection kits for their customers. This option may work for private investigations where all parties agree to the identity of the samples, but the findings will generally not be admissible in a court of law. Lack of legal status also applies to results of so-­called “secret” paternity tests performed without consent or knowledge of all affected parties. The most common scenario involves a suspicious father who secretly collects biological material such as a toothbrush or used cotton swab from a child and has it compared to his own DNA without knowledge of the mother. The German Federal Constitutional Court has deemed this type of testing a violation of the child’s right to informational self-­ determination and has upheld a decision banning the results from a civil court case (Bundesverfassungsricht  2007). Subsequently, a German federal law issued in 2010 and updated 2019 clearly states that paternity and kinship testing is only allowed with consent of all test subjects; in cases where test subjects are deceased individuals, a family court can give permission (Bundesamt fuer Justiz  2019). The British government issues guidance on human samples, and the Human Tissue Act from 2004 prohibits the covert collection of biological samples such as hair and fingernails for DNA testing, with the exception of medical or criminal investigations (https://www.hta.gov.uk/ standards-­ and-­ guidance). Despite the obvious societal risks, many countries do not have any legal restrictions in place and more discussion on the ethical and legal implications of secret DNA collection and testing should be encouraged (Joh  2011). natal paternity testing. Earlier Another difficult area is pre-­ methods, such as chorionic villi sampling, posed a risk for the unborn fetus, which is not the case for non-­invasive circulating fetal DNA testing (Wagner et al. 2009). The German gene diagnostics law restricts pre-­natal paternity testing to pregnancies after sexual assault, and other countries may have similar restrictions in place (Bundesamt fuer Justiz 2019). The matter of consent and right to informational self-­determination is a human rights concern. One area where ethical guidance is needed is the increasing use of kinship testing on refugees and migrants (Farahany et al. 2019). In summary, DNA testing allows for reliable paternity and kinship determination as long as a sufficient number of polymorphisms are tested, and data are interpreted correctly. Test reports must contain the biostatistical results in sufficient detail to allow for an independent evaluation and clearly state all assumptions (Gjertson et al. 2007). Specific cases may need to state multiple results to deal with different ethnic group assumptions or address relatedness issues. Ongoing research into software applications and new marker panels is likely to increase the ability to clarify complicated and even remote biological relationships (e.g. Lareu et al. 2012). Deep levels of sequence analysis will even be able to distinguish between putative fathers who are identical twins (Weber-­Lehmann et al. 2014).

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64.3.1 Pigmentation The most advanced forensic phenotyping application is the prediction of human pigmentation (eye, hair, and skin color). The Forensic Science Service in Great Britain developed the first forensic test for a pigmentation trait, namely an assay covering several single nucleotide polymorphisms (SNPs) in the melanocortin 1 receptor gene associated with red hair (Grimes et  al.  2001). Human eye, hair, and skin color traits are all connected to the melanin pathway within a specialized cell type (melanocytes) and genes regulating melanin transport to either keratinocytes in skin or cortical sheath cells in hair (Sturm and Duffy 2012). Some genetic polymorphisms in the pathway have an immediate causal effect, and researchers have found specific alleles with high impact on the resulting phenotype. Examples are the HERC2 SNP rs12913832, where the homozygote genotype is linked to either blue or brown eyes (Sturm et  al.  2008), or the

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Many candidate markers and often overlapping test panels have been described for eye, hair, and/or skin color predictions. Initial SNP panels were expanded, validated for forensic use, or integrated in commercially available massive parallel sequencing kits (Kayser 2015). The first fully validated pigmentation assay tested in a collaborative exercise was the IrisPlex system for eye color (S. Walsh et al. 2011; Chaitanya et al. 2014). Prediction accuracy for this 6-­SNP panel and its statistical model was high for blue and brown eyes but not for intermediate colors such as green or variations of brown and blue that could be described as light brown or gray. These colors may require additional markers and will also benefit from different categorization and more objective classification schemes. Quantitative, objective eye color measurements under investigation include a digital quantification of hue and saturation values from photos (Liu et al. 2010), measuring blue and brown pixel density (PIE-­score, Pośpiech et  al.  2016) and digital capture of eumelanin, pheomelanin colored, and non-­ pigmented areas of the iris only (Wollstein et  al.  2017). As expected for any genetic trait, eye color determination is affected by epistasis or gene-­to-­gene interactions that complicate additive SNP allele-­based predictions (Pośpiech et  al.  2014). Eye color variation is mostly limited to populations of European origin, and a collation of published data shows a North to South gradient regarding blue and brown eye color with France and Germany presenting the largest proportion of individuals in the intermediate category (Katsara and Nothnagel 2019). Evolution of human pigmentation is an interesting topic, and many genes have been shown to be under selective pressure. Migration to the Northern hemisphere with less UV exposure has been cited as the reason for loss of melanin and the selection of lighter colors. A possible evolutionary advantage for blue eyes could be this trait’s linkage to a gene protecting against seasonal affective disorder, a depressive illness caused by the lack of sunlight (Sturm and Duffy 2012). Overall eye color determination, at least for blue and brown eyes, is reproducible across marker panels and has reached a sufficient level of certainty for forensic applications, but intermediate eye color reporting still varies based on color category definitions and software algorithms (P.M. Schneider et al. 2019).

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a hair and eye color SNP panel named HIrisPlex based on the original IrisPlex that was expanded to include 24 hair color SNPs characterized by Branicki et al. (2011). The assay generates hair shade probabilities (dark or light) and results for color categories (red, blond, brown and black) that are then combined to hair color predictions such as “red” or “black” but also “light blond,” “dark blond/light brown,” “dark brown/black,” or other combinations. Overall prediction accuracy was 69.5% for blond, 78.5% for brown, 80% for red, and 87.5% for black hair (S. Walsh et al. 2013). For blond and brown hair, predictions are complicated by the fact that an individual’s phenotype may differ between childhood and adulthood. Out of 14  light brown to black haired individuals, where HIrisPlex had incorrectly predicted their hair color to be light blond, about half were found to have had lighter hair as a child (S. Walsh et  al.  2013). A systematic investigation on children confirmed that if a brown haired child (age 6–13) had been blond in early childhood, there was a significantly higher chance for an incorrect blond hair color prediction. For children with reported hair color darkening, only in 28% the HIrisPlex genotype matched the darker hair color, while the majority were predicted as blond reflecting their preadolescent blond phenotype but not their current appearance (Kukla-­Bartoszek et  al.  2018). The exact trigger is still unknown, but these age-­dependent hair color changes have been demonstrated not only for European but also African American populations (Commo et al. 2012). This is a limitation for blond/brown hair color prediction that forensic scientists need to clearly communicate to their stakeholders. The appearance of gray and white hair is another age-­dependent factor disrupting genetic phenotype predictions. This change is triggered by a loss of melanin production in hair root tissue, shows a lower prevalence in African and Asian subjects, and can affect individuals in different age groups (Panhard et  al.  2012; Jo et  al.  2018). A recent search for gray-­hair-­informative genetic markers through whole exome sequencing identified two novel SNPs associated with the trait. But the authors had to conclude that these two SNPs and a group of 11 previously characterized markers explained less than 10% of the hair graying variation, while age alone accounted for more than 45% (Pośpiech et al. 2020). As with eye color, hair color prediction could benefit from a more quantitative phenotype characterization. Ongoing research is working on quantitative classifications, epistatic effects, and the genetic basis behind age-­related hair darkening and graying (Kayser 2015).

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SLC24A5 *THR111 allele coding for lighter skin and fixed across European populations (Quillen and Shriver  2011). While pigmentation is still a continuous and somewhat complex phenotype, compared to other EVCs, e.g. height, fewer genes have been shown to have a large effect and a smaller number of markers can lead to high prediction accuracy (Kayser 2015).

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Hair color Hair color is also a continuous trait with much of the variation residing among populations of European origin. As with eye color, prediction accuracy is highest for phenotypes at the end of the color spectrum, in this case red and black hair (Branicki et al. 2011). Walsh et al. (2013) completed a forensic validation for

Skin color Skin pigmentation varies across the globe, and most of the geographical differences can be explained with selective pressure toward more UV protection and darker skin in equatorial latitudes as depicted in Figure 64.9 (Dimasi et al. 2011). Several of the eye and hair color SNPs associated with the melanin pathway are also informative for skin color. Table 64.9 shows all genes and their SPNs for the HIrisplex-­S panel, a forensic predictive tool for all three pigmentation traits. Of the six eye color-­informative SNPs, four also affect hair and skin color, and two are associated

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Figure 64.9  Global skin color distribution for native populations. Colors based on the Luschan color chart with original data compiled by Biasutti in 1941. The chart shows the concentration of UV-­protective darker skin colors in equatorial areas. Source: From Dimasi et al. (2011). Republished based on Creative Commons Attribution License.

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Table 64.9  HIrisPlex-­S SNP panel for human pigmentation prediction. Gene name and function

OCA2

Oculocutaneous Albinism II; melanocyte P protein production likely involved in melanosome pH regulation and/or cross membrane transport.

HERC2

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OCA2 OCA2 OCA2 OCA2

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Gene

HERC 2; nonpigmentation gene; role as melanocyte-­specific enhancer for the OCA2 gene.

HERC2 HERC2 HERC2 HERC2 MC1R

MC1R MC1R MC1R

Melanocortin 1 receptor; controls melanocyte production of either eumelanin (brown/black pigment) or pheomelanin (yellow/red pigment).

SNP

Trait

Chr.

Ref allele

Alt allele

rs1800407

Eye, hair, skin Skin Skin Skin Skin

15

C

T

15 15 15 15

T G A G

C T T A

15

A

G

rs2238289 rs6497292 rs1129038 rs1667394

Eye, hair, skin Skin Skin Skin Skin

15 15 15 15

A A G C

G G A T

rs796296176

Hair

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A insert

–

rs11547464 rs885479 rs1805007

Hair, skin Hair, skin Hair, skin

16 16 16

G G C

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rs1800414 rs1470608 rs1545397 rs12441727 rs12913832

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Table 64.9  (Continued) Gene

Gene name and function

Trait

Chr.

Ref allele

Alt allele

rs1805008 rs201326893 rs1110400 rs2228479 rs1805005 rs1805006 rs1805009 rs1126809 rs3212355

Hair, skin Hair Hair, skin Hair, skin Hair Hair, skin Hair Skin Skin

16 16 16 16 16 16 16 16 16

C C T G G C G G C

T A C A T A C A T

Solute Carrier family 24 member 4; potassium-­dependent sodium/ calcium exchanger.

rs2402130 rs12896399 rs17128291

Hair, skin Eye, skin Skin

14 14 14

G G A

A T G

SLC24A5

Solute Carrier family 24 member 5; encodes an intracellular membrane protein.

rs1426654

Skin

15

A

G

SLC45A2 SLC45A2

Solute Carrier family 24 member 2; or membrane associated transporter protein (MATP); encodes a transporter protein that mediates melanin synthesis.

rs28777 rs16891982

5 5

C C

A G

TYR

Tyrosinase; codes for Tyrosinase, enzyme responsible for the first step in melanin production.

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Hair, skin Eye, hair, skin

rs1042602

Hair, skin

11

C

A

rs1393350

Eye, skin

11

G

A

Hair, skin

9

C

A

Tyrosinase-­related protein 1; may stabilize tyrosinase and maintain melanosome shape.

rs683

IRF4

Interferon regulatory factor 4; SNP in melanocyte-­specific IRF4 enhancer affects Tyrosinase expression.

rs12203592

Eye, hair, skin

6

C

T

EXOC2

Exocyst complex component 2; involved in vesicle membrane interactions and membrane transport.

rs4959270

Hair

6

C

A

KITLG

KIT ligand; ligand for tyrosine kinase, regulates many cell functions including melanogenesis.

rs12821256

Hair, skin

12

T

C

PIGU

Phosphatidylinositol Glycan anchor biosynthesis class U; component of the GPI transamidase complex.

rs2378249

Hair, skin

20

G

A

RALY

RALY heterogeneous nuclear ribonucleoprotein; interacts with several proteins during mRNA metabolism

rs6059655

Skin

20

A

G

ASIP

Agouti signaling protein; ASP binding to MC1R downregulates eumelanin synthesis increasing pheomelanin.

rs6119471

Skin

20

C

G

ANKRD11

Ankyrin repeat domain 11; facilitating protein interaction including histone deacetylases controlling gene activity.

rs3114908

Skin

16

T

C

DEF8

Differentially Expressed in FDCP 8 homolog; associated with photoallergic dermatitis.

rs8051733

Skin

16

A

G

BNC2

Basonuclin 2; codes probable transcription factor specific for skin keratinocytes.

rs10756819

Skin

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SLC24A4 SLC24A4 SLC24A4

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MC1R MC1R MC1R MC1R MC1R MC1R MC1R MC1R MC1R

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Color-­coded SNPs are informative for more than one trait. Gene names and functions from NIH National Library of Medicine (https://ghr.nlm.nih.gov/gene) and NCBI Resources (https://www.ncbi.nlm.nih.gov/gene/).

with skin pigmentation. The remaining markers include 30 SNPs for skin color with some of them also predictive for hair color and part of the previous HIrisplex kit (Chaitanya et al. 2018). Research has discovered promising SNP markers for skin color prediction in different global populations (Kayser  2015). An observer’s perception of skin color is clearly subjective, and while it is prudent to use quantitative measurements for marker discovery and scientific testing, it is challenging to report the

resulting forensic prediction skin color in a meaningful manner. Published reporting categories range from “not white,” “not dark” (Spichenok et  al.  2011) to “white,” “intermediate,” “black” (Maroñas et  al.  2014) to a five-­category system based on the Fitzpatrick scale (Chaitanya et al. 2018). The Fitzpatrick scale (a skin phototype classification) augments each color category with sun reactivity: skin type 1 “pale white skin – always burns, never tans,” type 2 “white skin – usually burns, tans minimally,” type 3

Massive parallel sequencing for pigmentation panels

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Massive parallel sequencing (MPS) described in Section 64.1 can simultaneously test thousands of SNPs, which accommodates larger marker panels and could possibly improve prediction accuracies for intermediate colors. The European VISAGE consortium tested primer sets for pigmentation and ancestry informative SPNs on different MPS platforms (Breslin et  al.  2019; Palencia-­ Madrid et  al.  2020; Xavier, de la Puente, Mosquera-­ Miguel, et al. 2020). One commercially available forensic kit for the MiSeq FGx (ForenSeq DNA Signature Prep kit, Verogen) incorporates a primer set for the 22 HIrisplex eye-­and hair-­color-­informative SNPs, which means many casework laboratories already have the ability to test for eye and hair color and need guidance on data interpretation and reporting. MiSeq FGx results are analyzed with ForenSeq Universal Analysis Software (UAS), which uses the HIrisPlex multinomial logistic regression model for eye and hair color determination (England and Harbison 2020). But there are differences: the UAS algorithm has not been updated with additional training data samples like the HIrisplex webtool and does not return the additional light versus dark classification (England and Harbison  2020). A systematic comparison of the UAS with other eye and hair color prediction tools, specifically the Bayesian approach incorporated in the Snipper tool used by Maronas et al. (2014), the HIrisplex webtool (Walsh et al. 2017), and the categorical systems used by Spichenok (2011), revealed differences in prediction accuracy. Errors were typically caused by intermediate phenotypes or the known discrepancy between genotype and adult hair color (Sharma et al. 2019). A study on ancient and contemporary Native American samples revealed discrepancies between HIrisplex S and Snipper 23 prediction tools (Carratto et al. 2020). These findings emphasize the need for further software development and validation for pigmentation predictions. Overall, current prediction probabilities for different pigmentation panels vary from area under curve (AUC) values greater than 0.9 for blue or brown eyes, red hair and black or white skin, to values around 0.7 for brown hair and light to intermediate skin categories (P.M. Schneider et al. 2019). The AUC value depends on how well a binary classifier performs and is established by plotting the true positive prediction rate against the false-­positive prediction rate for each marker. A value of 0.5 represents a random prediction, while 1.0  would be a perfect prediction (Kayser  2015). Larger SNP arrays theoretically offer more nuanced EVC predictions. A private laboratory in the US currently offers a phenotyping and ancestry test that is marketed directly to law enforcement agencies. This assay is based on more than 800 000 SNPs, proprietary data sets, and machine-­learning algorithms, but the lack of a published forensic validation is a concern to the scientific community (P.M. Schneider et al. 2019; Arnold 2020). The threshold of communicating a result to a law enforcement agency is less a scientific but more of an operational decision about how much uncertainty can be tolerated (P.M. Schneider et al. 2019). Ethical and legal implications of reporting on eye, hair and skin color, and other EVCs are discussed below.

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“light brown skin – sometimes mild burn, tans uniformly,” type 4 “moderate brown skin – burns minimally, always tans well,” and skin type 5 “dark brown skin – very rarely burns, tans very easily” to skin type 6 “deeply pigmented dark brown to darkest brown skin – never burns” (Fitzpatrick 1988). Many authors work with continuous quantitative measurements captured with spectrophotometers, either brightness (e.g. Stokowski et  al.  2007; Valenzuela et al. 2010; Maroñas et al. 2014) or red reflectance corresponding to the melanin index (pigmentation protection factor) (Crawford et  al.  2017; A.R. Martin et  al.  2017; Andersen et  al.  2020). Working with South Asian samples from India, Pakistan, Bangladesh, and Sri Lanka, Stokowski et  al. (2007) eliminated all intermediate types and only used the highest and lowest reflectance values within this cohort for their association study. Maroñas et al. (2014) screened 59 candidate skin pigmentation SNPs in a global sample. Two of the most informative markers were rs16891982 in SLC45A2 (classifier for intermediate color) and rs1426654 in SLC24A5 (differentiates black or white vs. intermediate), but other previously published associations could not be confirmed. Consistent with strong recent selective pressure, allele frequencies of skin-­color-­informative SNPs differ widely across ethnic groups and studies within a population may not identify the most informative markers (Kayser  2015). Population-­specific information may still be useful. Crawford et al. (2017) and Martin et al. (2017) identified a high degree of skin color variation in African populations with the Khoi San from Southern Africa being the most lightly pigmented group, whereas populations closer to the equator like the East African Nilo-­ Saharan populations showing much darker skin colors. Both studies confirmed the prominent role of the SLC24A5 gene in human pigmentation, but only Crawford et al. (2017) showed an effect for OCA2/HERC2 alleles and identified two new genes strongly associated with African skin tone variation (Crawford et al. 2017). The skin markers included in the HIrisplex S system (Table 64.9) were selected to be informative across different continental populations and based on the evaluation of 77 candidate SNPs from 37 genetic loci and more than 2000 individuals from 31 populations. All test subjects were categorized by the same experienced dermatologists based on the Fitzpatrick scale either during sample collection or based on photos (S. Walsh et  al.  2017). Using a single evaluator accounted for the subjective nature of skin color perception. A possible forensic reporting strategy based on a modified Fitzpatrick scale (five categories with type III and IV combined) was presented as part of the HIrisPlex S forensic validation (Chaitanya et al. 2018). If a skin color category reaches a probability of 0.9 or higher in the online prediction tool, a prediction can be reported as being the most probable type. For all others, the second highest probability should be evaluated to formulate a more detailed description, e.g. a result with “pale” as the highest category could either be “pale” to “very pale” indicating a lack of tanning ability or “pale” to “intermediate” denoting a possibly darker type especially in the summer. Similarly, a result with the same highest probability for the “dark” category could be either “dark” to “intermediate” or “dark” to “black” (Chaitanya et al. 2018).

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Other hair-­related features such a male baldness, widow’s peak, unibrow, and eyebrow and beard thickness share many genes with scalp hair morphology (Adhikari et  al.  2016; Pickrell et al. 2016). Male baldness has been widely explored in clinical studies. Initially, genetic inheritance was studied in connection with early-­onset androgenetic alopecia (AGA) and one of the strongest associations found was to an androgen receptor and ectodysplasin A2 receptor (AR/EDA2R) haplotype on the X-­chromosome (Kayser  2015). A forensic study on European males from seven countries selected five SNPs, including AR, to be part of their prediction model. The authors used a seven-­ category scale ranging from no recession of the hairline, over several categories for increasing frontal hair loss, to all hair being lost in front and on the crown. The assay achieved a prediction accuracy expressed as AUC of 0.761, but only for males older than 50 (Marcińska et al. 2015). Another study by Liu et al. (2016) tested

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The genetic reconstruction of facial features is a complex task and clearly the holy grail of forensic DNA phenotyping. Facial features that have been targeted include eyebrows (Adhikari et al. 2016), but also eyelid shape, specifically the monolid feature in Asian populations (Q. Wang et al. 2017), ear lobe attachment (Adhikari et  al.  2015), chin dimples (Pickrell et  al.  2016), and freckles (e.g. Hernando et al. 2018; Kukla-­Bartoszek et al. 2019). Genes for the latter two traits overlap with skin pigmentation genes. The melanocortin-­1 receptor (MC1R) gene, also important for red hair and light skin phenotypes, is considered a major contributor to the formation of freckles in Europeans regardless of skin type and hair color. In Spanish individuals, this marker helped to correctly predict the presence or absence of freckles in 74.13% of test subjects (Hernando et al. 2018). Kukla-­Bartoszek et al. (2019) studied Polish individuals with 113 candidate DNA markers and introduced an additional “medium freckled” category. Like Hernando et al. (2018), they found a positive correlation of freckles to female gender. Including gender as one of the predictors, the prediction model reached accuracies of AUC = 0.814 for heavily freckled, AUC = 0.686 for medium freckled, and AUC=0.771 for non-­freckled individuals. As for other phenotypic traits, the appearance of facial freckles is continuous, difficult to categorize, and changes with age. This may limit its value in forensic descriptions for all but the more extreme variations. Based on their experience with identical twins, most people will agree that facial features must have a high degree of inheritability. A person’s face is a complex combination of highly variable components that make us unique and recognizable but are difficult to capture for scientific analysis. A face is not only influenced by genes, but also by sex, age, and body mass index which will need to be taken into account. Anthropologists, forensic artists, and software developers have been working on various aspects of forensic facial imaging applications, e.g. face reconstruction from skulls, creation of police sketches, age progression for images, or facial recognition software (Stephan et  al.  2019). Molecular photofitting or the prediction of a person’s face from their DNA is now joining the field of forensic facial imaging. Targeted or GWAS studies have tested different populations to

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Some of the other external traits under investigation are related to hair morphology predicting, e.g. hair thickness or differentiating between straight and curly hair (Kayser 2015). EVCs not related to pigmentation are especially of interest in countries with less diversity in hair colors like Asia (Seo et al. 2017). A genome-­wide association study (GWAS) on Indonesian and Thai individuals identified a receptor gene (EDAR) to be associated with thicker hair (Fujimoto et al. 2008). The EDAR association to hair shape was confirmed in a South American cohort (Brazil, Chile, Colombia, Mexico and Peru) in a study that also verified the role of the trichohyalin gene (TCHH) in coding for curly versus straight hair (Adhikari et  al.  2016). The same authors reported that trichohalyn is expressed in cornifying hair follicle keratinocytes, affects the cross-­linking of keratin filaments, and identified the first genetic variants to be associated with curliness in Europeans (Medland et  al.  2009). Exploring a forensic application, Pośpiech et al. (2015) categorized hair phenotypes as either “straight,” “wavy,” or “curly” and tested SNP candidates from the TCHH and other genes described by Medland et  al. (2009) in individuals from seven European countries. Three of the markers were found to be informative and together resulted in better than 80% predictability for straight hair, but also a high number of inconclusive results. An updated assay based on 32 SNPs typed on an MPS platform now achieves prediction accuracies for “straight” versus “not straight” of 0.664 (AUC) in Europeans and 0.789 (AUC) in non-­ Europeans (Liu et  al.  2018; Pośpiech et al. 2018). It remains to be seen if this capability will be useful in casework. Hair shape is continuous with many intermediate categories from straight, to wavy, to very tight curls that can be subjective, and reporting language for perceived hair shapes still needs to be developed.

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25 SNPs from 12  loci in middle-­aged and elderly German and Dutch males including AGA patients. Prediction accuracy for AGA patients reached an AUC of 0.74, but for the remaining cohort age alone was the main predictor for baldness (Liu et al. 2016). Male pattern baldness (MBP) shows a later onset and has different genetic associations in Asian males (W.S. Lee and Lee 2012). The trait also has a generally lower incidence rate in African populations (Hillmer et  al.  2009), indicating more research is needed for a globally applicable prediction panel. As stated by Marcińska et al. (2015), the lower predictability of the trait in younger test subjects is due to the fact that MPB is an age-­ progressive condition. To be useful in a forensic context, testing for baldness would need to be combined with chronological age determination on the evidentiary samples.

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Male baldness

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markers are only associated with a very particular trait, such as the nasion position or nose wing breadth. Some face informative SNPs are located in genes known to cause clinical face abnormalities, e.g. PAX3 involved in the Waardenburg syndrome, a genetic disease characterized by complex symptoms such as loss of pigmentation, hearing loss, but also a wider than normal distance between the eyes (Paternoster et  al.  2012). Another example is SOX9, where a specific mutation causes campomelic dysplasia (CMPD), a rare genetic disease causing abnormal facial features, including a flatter face and a depressed nasal bridge (Cha et al. 2018). Since the amount of facial feature variance explained by a single SNP in candidate genes is very low, it is expected that only very large marker panels will be useful (Kayser 2015). With research on facial features still ongoing, how to report findings to forensic stakeholders is an open issue. Claes et  al. (2014) were working with a “base” face matching a subject’s gender and biogeographical background and modified this face with detected genetic variations (Claes et al. 2014). A combination of a genetically predicted face and an artist’s sketch is what Parabon, a US company marketing a “DNA Phenotyping” service called Snapshot, provides to their customers. As mentioned under eye and hair color, this company tests questioned samples with a panel of over 800 000 SNPs and uses proprietary databases with phenotypes and genetic profiles and machine-­ learning algorithms to predict physical appearance and ancestry (Arnold 2020). The resulting “composite profile” includes eye, hair, and skin color reported in color charts and biogeographical ancestry together with a picture of the unknown DNA source’s face. Despite the speculative nature of the artist’s rendition, the company’s website displays examples with predicted faces being similar to the identified DNA source (Parabon  2020). The major factor here seems to be biogeographical ancestry having such strong effect on facial features (Sero et  al.  2019; Arnold  2020). The use of this technology and future facial prediction models needs to be carefully monitored to avoid exaggerated expectations and racial discrimination.

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identify candidate markers (Kayser 2015). As with other phenotypic traits, feature extraction and categorization varies between different research groups but mostly starts with high-­resolution 3 D images (e.g. Claes et al. 2018; Xiong et al. 2019). Figure 64.10 shows commonly used frontal face landmarks used to characterize eye and nose variation (M.K. Lee et  al.  2017). Additional measurements can be facial size, head circumference, and facial shape corrected by size (allometry), or upper eyelid features (Cole et al. 2016; Cha et al. 2018). Research has identified several SNPs associated with specific facial traits, but so far these markers only explain a small percent of the variability, e.g. the 31 associated SNPs identified in Xiong et al. (2019) together explain 4.62% of the variation. An estimated 25% of the adult face’s appearance is determined by biological sex and biogeographical origin, with an additional effect of age and body mass index (BMI) (Sero et  al.  2019). Many facial feature

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Figure 64.10  Standard facial landmarks used in genetic association studies. Landmarks are labeled as follows: n: nasion; prn: pronasale; sn: subnasale; ls: labiale superius; sto: stomion; li: labiale inferius; sl: sublabiale; gn: gnathion; en: endocanthion; ex: exocanthion; al: alare; ac: alar curvature point; sbal: subalare; cph: crista philtri; ch: chelion; and t: tragion. For bilateral landmarks, left and right indicated by _l and _r after the landmark abbreviation. Source: From Lee, M.K. et al. (2017) supplemental material. Republished based on Creative Commons Attribution License.

Stature information can be an important trait to add to the description of a person of interest or set of unidentified remains. Anthropological methods can provide this information, if long bones (preferably the tibia) are present but estimates have a standard error around 30  mm (Duyar and Pelin  2003). Adult height is an inherited polygenic trait, but average height has been increasing through modern times, a trend that has been attributed to changes in nutrition and hygiene affecting fetal development, childhood, and adolescence (Perkins et  al.  2016). Nevertheless, twin studies reveal a high degree of heritability ranging from 84% to 94% (Silventoinen et al. 2003). Height is an example of a complex trait where each functional gene only contributes a small amount of information. A sequence of association studies demonstrates how successive addition of relevant regions increases the amount of explained height v­ ariation from 27 regions and 3.7% (Gudbjartsson et al. 2008), 54 regions

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­ re-­programmed-­selective gene expression in different developp ment stages and tissues, and enables the body’s adaptation to environmental influences (Freire-­Aradas et al. 2017). These parallel functions cause different changes in the epigenetic pattern of DNA methylation that can be distinguished as either the “epigenetic clock” or “epigenetic drift.” The term epigenetic clock refers to pre-­programmed methylation changes in specific regions of the genome strongly associated with the age of an organism. In addition to these systematic changes occurring in a similar fashion across all individuals, methylation levels also change in reaction to lifestyle influences such as diet or alcohol intake, different diseases, and other environmental factors leading to epigenetic drift (Jones et  al.  2015). Figure  64.11 illustrates this directional and random aspect of methylation patterns changing with age

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and 4–6% (Aulchenko et al. 2009), 180 regions and 10% (Lango Allen et  al.  2010) to 423  loci with 697 SNPs and 16% (Wood et al. 2014). As expected, the associated regions were enriched for genes relevant for pathways and tissue types in human skeletal growth such as collagen, chondrocyte differentiation, insulin-­like growth factor 1, and fibroblast growth factors, and included genes previously described for abnormalities or syndromic short stature (Wood et  al.  2014). Aulchenko et  al. (2009) published the first height prediction model with the outcome being either “very tall” or not. Their panel of 54 SNPs reached an AUC of 0.65, only slightly above the AUC = 0.5 value equaling a random prediction (Kayser 2015). The 180 variants identified by Lango Allen et al. (2010) were tested on a large Dutch population sample and shown to be informative for height with an AUC of 0.75 for the binary “very tall” to not tall prediction (Liu et al. 2014). A follow-­up study on the same Dutch cohort added 697 markers discovered by Wood et al. (2014) and improved the prediction accuracy to AUC= 0.79 (Liu et al. 2019). The category “very tall” was defined as exceeding 1.88 times of the standard deviation for average height, meaning the absolute value varies between different populations. The described prediction model also adjusted the quantitative phenotype with an age and sex correction factor, something that then would first need to be established for unknown evidence. So far authors only found a weak correlation (R2 = 0.21) between predicted and observed adjusted height in a continuous quantitative height model (Liu et al. 2019). In conclusion, meaningful genetic height prediction will require much larger SNP panels and can only be achieved if other characteristics such as sex or age are established at the same time.

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Determining the chronological age of an unidentified living person or unidentified human remains in criminal and civilian investigations has long been a task for forensic anthropologists and/or medical examiners. Now molecular methods expand age determination from skeletal remains to biological evidence found at crime scenes making this application a part of forensic DNA phenotyping (Freire-­Aradas et al. 2017; Jung et al. 2017). Adding the age as a descriptor while searching for a person of interest can narrow down the pool of subjects under investigation. Age is also a required modifier for predicting progressive features such as baldness, hair color, height, and face phenotypes (Kayser 2015). The most advanced assay for chronological age is based on epigenetics and age-­associated DNA methylation patterns in different biological tissues (H.Y. Lee et al. 2016). In a systematic comparison to other molecular approaches, DNA methylation markers provided the most accurate age predictions and are now the main research focus (Zubakov et al. 2016). DNA methylation is one component of the cell’s gene regulation system ensuring dynamic and cell-­type specific gene expression during embryonic development, childhood, and adulthood. This epigenetic system also includes mechanisms such as chromatin remodeling and non-­ coding RNAs, allows for

Age

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MZ twins (purple and green) more discordant with age

MZ twins (purple and green) discordance similar over time

Figure 64.11  Schematic representation of epigenetic drift versus the epigenetic clock. Lifestyle affects methylation sites that may appear age related in a single individual but are not consistent as shown, e.g. in twin studies (genetic drift). Other sites’ methylation levels change with age across all individuals (epigenetic clock). Source: From Jones et al. (2015). Republished based on Creative Commons Attribution License.

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age. Another explanation may be that while AR-­CpGs are selected as part of the epigenetic clock, the same sites are also affected by epigenetic drift, meaning their methylation level would reflect an individual’s biological rather than the chronological age (see Figure  64.11). A study on clinical patients was able to demonstrate this effect for some of the CpG sites of interest. Two of the tested AR-­CpG markers (ELOVL2 and C1orf132) performed well in both, patients and healthy controls, while two other sites (TRIM59 and KLF14) overestimated the age in the Alzheimer group indicating accelerated aging of these patients (Spólnicka et al. 2018). Gender has not had an effect on age prediction (e.g. Zbieć-­Piekarska et  al.  2015; Hamano et  al.  2016). There is a genomic background effect and not all markers are equally informative across ethnic groups, for example the methylation to age correlation for the KLF14 gene was weaker in Korean subjects than for Polish Europeans (Cho et  al.  2017). Age predictions based on five markers including two CpGs at ELOVL2 showed similar levels of accuracy in Middle Eastern, West African, and Middle European cohorts (Fleckhaus et al. 2017). On the other hand, a larger set of novel AR-­CpG sites revealed consistency, but also site-­specific differences and lack of age associations between middle-­ aged white and African American cohorts (Tajuddin et al. 2019). Applicability across ethnic groups is thus a forensic marker selection criterium. Methylation status is a quantitative trait not easily replicated across different detection platforms, something that needs to be taken into account for forensic applications. It is currently recommended that each laboratory generates their own model training data on expected methylation levels as detected by their instrumentation before testing unknown crime scene samples (P.M. Schneider et  al.  2019). Methylation levels are measured as the ratio between unmethylated and methylated bases at each CpG position (methylation β-­value) ranging from 0 for completely unmethylated to 1 for full methylation of all target molecules. This value changes gradually in age-­associated markers with aging either causing an increase or decrease in the β-­value (Freire-­ Aradas et al. 2017). Selecting markers for maximum degrees of absolute methylation difference can increase accuracy (Bekaert et al. 2015). The most common DNA methylation detection technologies are all based on sequence detection after bisulfite conversion. This method exposes denatured single-­stranded DNA to sodium bisulfite causing unmethylated cytosines to become deaminated and convert to uracil. During PCR, protected methylated sites maintain their C=G base pairing, while deaminated sites replicate as T=A pairs. The sequence difference between cytosine for methylated sites and thymine for the unmethylated equivalent can then be analyzed with several methods showing different levels of simultaneous coverage. Forensic research teams have successfully used a range of methods, e.g. real-­time PCR and high-­resolution melt curves, fluorescent single base extension (SNaPshot), sequencing by synthesis (pyrosequencing), mass spectrometry (EpiTYPER) system, and massive parallel sequencing (MPS) (Freire-­Aradas et al. 2017). A study comparing four of these detection methods for the same samples and AR-­CpG panel in three different laboratories found generally good concordance

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across different individuals. Studies on monozygotic twins with identical genomes are the most convincing evidence for the heritability of methylation patterns and at the same time demonstrate the increasing divergence with age caused by external factors (Fraga et al. 2005). The latter is a possible approach to distinguish between identical twins as the source of DNA evidence (see Section 64.1). DNA methylation consists of the addition of a methyl residue to cytosine bases followed by a guanine, and a methylated site in a DNA sequence is annotated as CpG. The change is reversible and prevents gene transcription. Genes with an accumulation of CpG sites (hypermethylated) are considered silenced, while active genes are characterized by a lack of CpG sites in the promoter region (hypomethylation) (Schübeler 2015). The initial observation that an individual’s chronological age correlated to systematic methylation changes was made in aging and longevity studies based on thousands of methylated sites. After a comprehensive analysis of methylation profiles in blood, Weidner et  al. (2014) identified a panel of 102 age-­related CpG (AR-­CpG) sites in blood with the five most informative sites being located in the following genes: EDAR-­associated death domain (EDARADD), integrin alpha 2b (ITGA2B), a member of the RAS oncogene family (RAB36), phosphodiesterase 4C, cAMP-­specific (PDE4C), and aspartoacylase (ASPA). A predictive model based on three of these regions resulted in age predictions with a mean absolute deviation (MAD) from true chronological age of ±5.4 years (Weidner et  al.  2014). Other research also on blood confirmed most of these targets and discovered additional AR-­CpGs for a gradual improvement of prediction accuracy (Freire-­ Aradas et  al.  2017). One promising locus, the ELOVL2 (elongation of very long chain fatty acid protein 2) gene, contains several different AR-­CpGs, all tending to be hypermethylated with increasing age with a methylation level range from 7% in cord blood samples to 91% in 99-­year-­old subjects (Garagnani et  al.  2012). Marker evaluation and/or discovery for new promising candidates is ongoing (Vidaki, Ballard et al. 2017; Alsaleh and Haddrill 2019; Tajuddin et al. 2019). A forensic age prediction system must validate its marker panel in combination with an applicable statistical model. Most earlier predictions algorithms used multivariate linear regression models, but not all CpG sites show a linear correlation to age. One example is the ELOVL2 locus where a quadratic regression model improved predictions accuracy (Bekaert et  al.  2015). Applying either support vector regression or quantile regression models MAD values for age determination on blood could be lowered to 2.8 and 3.07 years, respectively (Xu et  al.  2015; Freire-­Aradas et al. 2016). It is important to note that these mean deviations are overall values, while the true error for age predictions varies for different age groups. Prediction ranges are narrower in young individuals and broaden for older age groups. Breaking down age predictions for different age categories demonstrates how an overall MAD of 3.75 years varies from 2.32 for 0–19 years, 2.61 for 20–39 years, 3.62 for 40–59 years to 5.28 for the 60–91 age group (Bekaert et  al.  2015). The higher error for the older age group may be caused by the fact that methylation levels change less with

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accelerated aging indicative of disease and higher mortality risk (Fransquet et  al.  2019). This raises genetic privacy concerns which will be discussed below and underlines the need to limit marker selection to CpG sites not affected by disease and lifestyle.

64.3.4  Biogeographical Ancestry

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Human diversity has developed during the time of early human migration when subpopulations were separated by distance and physical barriers. This does not mean humans are genetically distant from each other, and there is much more variation within a human population (93–95%) than between populations (3–5%) (Rosenberg et  al.  2002; Phillips  2015). Nevertheless, each individual carries the genetic legacy of their ancestors, and parts of this information can lead back to the ancestral biogeographical origin. Exploring this biogeographic ancestry (BGA) for biological samples such as crime scene stains, research specimen, or skeletal remains has a variety of applications (Philipps 2015). For research, a BGA test could improve the accuracy of self-­declared ancestry classifications for database and clinical study samples. Ancestry can help narrow down the pool of potential relatives in disaster victim identification and forensic genealogy applications. In cold case and current forensic investigations, knowing this information may help locate a perpetrator. There is a lot of uncertainty, and it is very important to manage an investigator’s expectations about the value of the ancestry prediction. For example, one has to realize that BGA estimates will only place an individual into a geographical area not necessarily corresponding to modern political states. BGA is also not the equivalent of race or belonging to a specific ethnic group, both of which have a cultural component and are defined by a combination of origin, language, and religion (P.M. Schneider et al. 2019). Uniparental markers like mitochondrial DNA and Y-­chromosome haplotypes indicate a single ancestor predating historical records and thus are only representative of an individual’s biogeographical ancestry, if there was no admixture in the past generations. Mitochondrial DNA (mtDNA) represents the maternal lineage, and mtDNA genomes have been used to establish the out of Africa theory and track migration paths for modern humans (Cann et  al.  1987). Y-­chromosomes are inherited from father to son, and testing for population-­specific haplotypes has complemented mtDNA data but also revealed gender differences in migration patterns. One example is the all-­male colonization after Spanish ships reached South America that to this day chromosomes in is reflected by a prevalence of European Y-­ admixed indigenous individuals (Vieira-­Machado et  al.  2016). Both of these marker types are of immense interest in anthropological research and historical investigations but can be misleading in forensic investigations more interested in recent origin of a person of interest. Autosomal DNA contains a small segment of all of our ancestors and can reveal the contribution from the same or different populations over many generations. Ancestry informative DNA

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between all of them except for SNaPshot, which is semiquantitative and showed higher predictive errors. It was possible to increase prediction accuracy across technical platforms by normalizing the data via z-­score transformation (Freire-­Aradas et al. 2020). Collaborative studies and systematic comparisons of different software transformations and prediction models are in progress (e.g. Aliferi et  al.  2018; Smeers et  al.  2018; Pfeifer et al. 2020). Another limitation for forensic applications is the large amount of DNA necessary for reliable results. Bisulfite conversion and subsequent purification are known to compromise forensic evidence through degradation, loss of material, and incomplete sequence conversion. It was estimated that 10 ng of DNA subjected to bisulfite conversion result in approximately 2 ng of DNA available for PCR and sequencing (Aliferi et al. 2018). Optimized forensic protocols have identified the most efficient kits, added internal conversion controls, and lowered the amount of required DNA input. A systematic study on different bisulfite conversion kits demonstrated robust replication of methylation levels down to 20 ng of genomic DNA, while 10 and 1 ng DNA amounts showed increased variation possibly due to stochastic effects (Heidegger et al. 2020). Two other studies reported accurate methylation quantification and age predictions on the Verogen MiSeqFx and for SNaPshot for 10 ng original DNA input (Hong et al. 2017; Aliferi et al. 2018). Experiments establishing the lower limit of detection are required components for assay validations and will have to be repeated by each laboratory planning to offer this test. Methylation patterns are body fluid/tissue specific (see also Section 64.1), which means AR-­CpG sites in blood are not equally informative in other sample types such as saliva, semen, or bones. Applying an age prediction panel optimized for blood had some success for saliva, albeit with a higher mean absolute error, but none of the same CpG sites were methylated in sperm cell DNA (Aliferi et al. 2018). Saliva contains up to 80% leukocytes, which explains why blood and saliva share age-­informative CpG sites. Adding methylated sites from different genes improved predictions accuracy for saliva to MAD 3.15 years (Hong et al. 2017). Semen consists of seminal fluid, which contains some diploid epithelial cells, and haploid sperm cells thus containing two cell types with very different biological functions. Sperm cells are gametes with very specific methylations patterns, which explains the lack of age association with previously described somatic CpG sites. Marker discovery for semen identified a panel of three AR-­CpGs with the highest level of association seen in a site in the promoter region of the TTC7B (Tetratricopetide Repeat Domain 7B) gene. The overall accuracy (MAD) over all three sites was 5.4 years (H.Y. Lee et al. 2015). A follow-­up study modified the statistical model to improve the accuracy for fresh semen to MAD of 4.8 years and successfully typed amounts as low as 5 ng of DNA and a range of mock casework samples, some of which had been stored for 17 years (J.W. Lee et al. 2018). Other tissue types with published age prediction assays include bone, muscle, and teeth (Freire-­Aradas et al. 2017). Ethical issues regarding chronological age prediction relate, for example to inadvertent detection of

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and global travel, all causing admixture between previously separated populations. This trend not only makes BGA determination more complex, but also disrupts the correlation between ancestry and appearance. As described above, admixture causes a variety of different phenotypes, and genes directly involved in coding for external traits have better predictive power for appearance. Research on genomes derived from the Human Genome Diversity Cell Line Panel (HDP-­CEPH) and collections of indigenous populations has shown that even without knowing which DNA locations are ancestry informative, individual genomes can be sorted into clusters based on their degree of genetic similarity and their corresponding common BGA (Rosenberg et al. 2002; J.Z. Li et al. 2008). One software program commonly used for this type of analysis is STRUCTURE, a model-­based algorithm that will create a predefined number of clusters (K) combining samples with distinctive allele frequencies. How well these clusters represent biogeographical distribution depends on the test population and the sensitivity of the included markers. Allowing for five clusters (K = 5), large continental groups (Africa, Eurasia, East Asia, America, and Oceania) are separated, at K = 7 Eurasia is divided in Europe, Middle East, and South Asia (Rosenberg et al. 2002; Phillips 2015). In order to apply this type of genetic similarity modeling for BGA prediction of unknown samples, it is essential to target informative markers and have the appropriate reference samples available (Kersbergen et  al.  2009;

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markers (AIMs) are characterized by population-­specific allele frequencies with either unequally distributed or fixed alleles. Single nucleotide (SNPs) or insertion deletion (INDEL) sequence changes identical by descent are the most common AIMS, but a sufficient number of STR alleles can also reveal population-­ specific frequencies (L. Pereira et al. 2011). Genes that are subject to positive selection such as the light skin gene SLC24A5  in Europeans or the malaria protective trait in the Duffy antigen receptor (DARC) in Africans are often population specific, and SNPs associated with these genes are among the most informative AIMs (Phillips 2015). The most extreme geographical separation between the five continents is reflected in the genetic distance between indigenous populations in Eurasia, sub-­Saharan Africa, East Asia, America, and Oceania. Areas where populations meet, the continental margins show gradients of AIM allele frequencies and are more difficult to distinguish via genetic typing. Nevertheless, with proper marker selection an area like Eurasia can be subdivided in the subgroups Europe, Middle East, and Central South Asia. Figure  64.12 illustrates how SNP data for continental populations such as Europeans, East Asians and Africans form distinct clusters while related populations such as East Asian and American, and European and South Asian show overlapping margins (Phillips 2015). More recent history shaped human populations patterns through the slave trade, colonization, escape from prosecution, and different waves of emigration

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Figure 64.12  PCA plots generated showing genetic distances between either three or six reference populations. The plots were generated with SNIPPER with either (a) 34-­plex genotypes from 1000 genomes or (b) global 128 AIM panel genotypes from 1000 genomes and HGDP-­ CEPH Native American/Oceanian samples. Borders are added to indicate marginal overlap. The analysis placed the unknown sample (cell line 9947A) in the European cluster. Gray dots indicate partial profiles. M is an artificially admixed sample of European and East Asian origin. Source: From Phillips (2015). Republished based on STM signatory agreement.

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reported as “European or Middle Eastern/North African” could have come from either Portugal, Italy, Romania, Russia/Poland (Ashkenazi Jews), or Jordan/Palestine. An investigator may also find it misleading that Philippine ancestry is broadly classified as East Asian. Again truly mixed individuals were the most difficult and had to be reported out as uninformative, if more than three root populations had a >20% contribution (Jin et  al.  2018). A Precision ID casework assessment in Australia found the same issue with a lack of differentiation between Southern European and Southwest Asian (called Middle Eastern/North African by Jin et  al.  2018). The authors also noted a lack of separation between East Asians and indigenous Americans, and the lack of Australian aboriginal references (Al-­Asfi et al. 2018). A weakness of the ThermoFisher population likelihood tool is the fact that in absence of the appropriate reference population, the highest likelihood ratio identified by the software points to the next closest population and can be misleading. This uncertainty can be assessed by a z-­score test evaluating if any of the available reference populations are sufficiently close to the true ancestry of the unknown. This test is part of the Genographer software package that was shown to improve ancestry prediction efficiency for the precision ID panel (Mogensen et al. 2020). Several regional AIM panels resolve the observed lack of predictive power for continental margins. The long history of human settlements around the Mediterranean and South Western Asia (encompassing the Middle East with countries like Lebanon and Syria, and adjacent areas such as Iran, Afghanistan and the countries around Kazakhstan) has led to different cultures, but neighboring countries are also prone to trade, population exchange, and geneflow. Two separate approaches to find AIMS for the region, an evaluation of previously published aiSNPs (Bulbul et al. 2018) and a de novo search in relevant population samples (V. Pereira et al. 2019), both again identified the close relationship between Middle Eastern and North African populations and Southern Europeans. One set of 111 new AIMs had discriminatory potential and was estimated to yield reliable results in 90–100% of individuals, but only if North Africa was combined with the Middle East (V. Pereira et al. 2019). A similar long history of population exchange has shaped the genetic makeup of South East Asia with the “mainland” countries such as Burma, Thailand, Laos, Cambodia, and Vietnam, and island states like The Philippines and Indonesia. A Korean research team identified 100 AIMs for East Asian and South East Asian populations and were able to differentiate between populations from Indonesia, Thailand, Japan, Korea, and China, in addition to separating these groups from African, American, European, and South Asian populations (J.Y. Jung et  al.  2019). The MAPLex panel for Asia-­Pacific populations was build based on a core set of globally informative AIMs (Phillips, Parson et al. 2014) and used the deliberate strategy of adding multiallelic aiSNPs and ancestry informative microhaplotypes combining several SNPs, thus increasing the expected level of polymorphism in a population. To ensure consistent multiplex performance on compromised skeletal remains and casework evidence, the size of the microhaplotype amplicons was reduced to match the SNP markers. The

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Cheung et al. 2018). The importance of having sufficient population representation in the reference sequences is illustrated by an early case application, namely the 2004 terrorist attack on a train station in Madrid, Spain. In order to distinguish between domestic and foreign terrorists, the forensic laboratory had to first build a more complete North African reference database (Phillips et al. 2009). Efficient AIM selection is well explained by Phillips (2015). In addition to selecting core markers based on the largest possible distance, this core set needs to be supplemented by other AIMs indicating more recent human divergence. Forensic BGA assays originally employed SNAPshot technology for either SNPs or INDELs combining between 14 and 55 AIMs (Phillips et  al.  2007; J.R. Kidd et  al.  2011; R. Pereira et al. 2012; Gettings et al. 2014; K.K. Kidd, Speed et al. 2014), but like for FPD testing future forensic applications are more feasible on massive parallel sequencing (MPS) platforms. Right now, two commercially available BGA assays can be analyzed on MPS instruments. The Verogen ForenSeq Signature Prep Kit B combines 22 EVC markers with 55 ancestry informative SNPs (aiSNPs) described by K.K. Kidd, Speed et  al. (2014). The panel allows for a broad categorization of African, European, East Asian, and Admixed American descent. Error rates were very low for self-­declared African, European, and East Asian individuals, but the company’s prediction algorithm missed most of the admixture and, due to a lack of reference data, failed to predict South Asian ancestry (Sharma et al. 2019). The lack of informativeness for South Asians was also noted in a study of the local population in Singapore (Ramani et al. 2017). A follow-­up study succeeded in improving ancestry inferences in Singapore for East Asian (Chinese and Vietnamese), South-­East Asian (Malay and Indonesian), and South Asian (Indian and Bangladeshi) individuals. This additional level of discrimination was obtained by adding Chinese, Malay, and Indian genotypes as references and using another software tool. Philippine and Burmese descent could not be resolved (Ramani et al. 2020). The ThermoFisher Precision ID ancestry panel combines two panels of 55 aiSNPs (K.K. Kidd, Speed et al. 2014) and 128 aiSNPs from Kosoy et al. (2009). Data evaluation is based on seven geographic root populations and offers the calculation of the likelihood of each genotype in each reference group (Population Likelihood) and an estimate of the ancestral composition for each genotype (Admixture Prediction). As described above, ancestry inference is limited by the availability of reference data. A Danish study was able to correctly predict the ancestry for their Danish and Somali samples, but not a set of Greenlanders (V. Pereira et  al.  2017). A casework validation by a laboratory in Toronto, Canada, rejected the Population Likelihood method as impractical for reporting purposes and developed interpretation and reporting guidelines based on Admixture Prediction (Jin et al. 2018). Only components >20% were considered for reporting. The authors’ results highlight the limitations of global ancestry inference. All Northern Europeans were predicted as European, but for Southern Europeans the most common result was a double inclusion inferring European or Middle Eastern/ North African descent. Accordingly, an unknown sample donor

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Ethical concerns about forensic DNA phenotyping center on genetic privacy. Traditional DNA identity testing has avoided possible conflicts by selecting polymorphisms not associated with diseases and functioning genes. Some countries enforced this restriction through legal provisions prohibiting any forensic use of coding genes, and only allowing non-­coding areas to be subject to analysis (MacLean  2014). These regulations are now under revision to allow for the implementation of forensic DNA phenotyping, a process already completed in some countries, e.g. The Netherlands and Germany, and in progress in several other regions (Koops and Schellekens  2008; Scudder et  al.  2018; Slabbert and Heathfield 2018; P.M. Schneider et al. 2019). Individual genetic privacy protection needs to be balanced against the public’s interest in efficient crime fighting and closure for victims, their families, and the families of missing persons. Forensic scientists have argued that testing for EVCs, such as hair color or height, should not be a genetic privacy concern since all phenotypes are openly visible, part of civil identification documents, and often already part of an investigation after eyewitness testimony. Under these circumstances, providing a genetic prediction inferred from evidence will either confirm or dispute eye witnesses with the advantage of a validated error assessment (Kayser 2015; de Cerqueira et al. 2016). In addition, genetic data from unknown crime scene samples are not considered protected personal information until after the source has been identified (Scudder et al. 2018). There are still concerns about inadvertent

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generation of internal disease-­related information that warrant a privacy impact assessment and legislative attention (Scudder et al. 2018). The forensic community is aware that moving from identity-­ informative DNA polymorphisms to new markers reflecting gene expression and methylation status constitutes a paradigm shift. Research revealed that the public and stakeholders are interested in using EVC data for investigative purposes but are also aware of the need for additional legal regulations and proportionality measures. In a Swiss survey including individuals working in criminal justice (forensic scientists, police investigators, and legal professionals) and other professions, over 80% in both groups were in favor of forensic phenotyping in casework. Of the participants supporting FDP, the majority (54.2%) preferred to limit the use to more serious crimes (Zieger and Utz 2015). The threshold for when to apply FDP depends on each individual’s role in the criminal justice system and/or the direction of their advocacy. In the Swiss study, more police officers than other professionals were willing to consider asking for FDP results in burglaries (Zieger and Utz  2015). Interviews with a variety of civil society stakeholders also revealed divergent views. For individuals affiliated with victims or families of missing persons, the value of finding a perpetrator through FDP outweighed the risk of a possible genetic privacy loss, while minority right advocates often argued that FDP may open the door to more intrusive testing and should either not be allowed at all or only under very specific circumstances (Samuel and Prainsack 2019a). Members of the European Visage consortium involved in either FDP research or regulation all agreed to limit predictive testing to truly externally visible traits and exclude internal characteristics like disease associations from forensic use. The same group, especially the forensic geneticists, emphasized the need for reliability and validity of any tests (Samuel and Prainsack 2019b). Another concern voiced both by forensic scientists and genetic privacy advocates that the probabilistic nature of phenotyping results could cause misunderstandings by law enforcement professionals, who may already have exaggerated expectations of what a laboratory report can provide. Specific training for investigators should include correct interpretation of reports and socially sensitive use of the information. Narrowing down a suspect pool to individuals with similar appearance can target an ethnic minority group at risk of racial profiling. The risk that this could lead to law enforcement stigmatization of these populations needs to be taken seriously and mitigated by proportionality of use, legal regulations, and law enforcement training (Branicki et al. 2014; Wienroth 2018; P.M. Schneider et al. 2019). Genetic privacy protection for data generated as part of FDP investigations becomes more challenging, if tested sites also provide health-­related predictions. While forensic assays are designed to avoid disease associations, this cannot always be prevented and disease relevant data may be generated inadvertently (Bradbury et al. 2019). This means it may become necessary to design the FDP data analysis laboratory workflow to either isolate or obfuscate unintended health associations. If a laboratory scientist interprets health data and becomes aware of a disease risk, there may be an obligation to notify the individuals/families affected by

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resulting panel included 108 biallelic, 34 triallelic, 2 tetraallelic SNPs, and 20 microhaplotypes (Phillips et al. 2019; Xavier, de la Puente, Phillips et al. 2020). The new markers were able to better differentiate between West Eurasian, South Asian, and East Asian populations, including a separation between Middle Eastern and European individuals (Phillips et al. 2019). MPS technology has been shown to be suitable for degraded DNA and sensitive enough for casework-­type samples, but partial genotypes can lead to wrong BGA predictions, and reliable performance for each SNP amplicon needs to be monitored carefully. Validation studies determined the lower limit of detection for complete genotypes to be between 0.063 and 0.125  ng of DNA (Al-­Asfi et  al.  2018; Xavier, de la Puente, Phillips et  al.  2020). Optimized panel design can replace primers and polymorphisms with systemic problems (Eduardoff et al. 2016). The research presented above has identified current capabilities and limitations of forensic BGA determination. This type of testing will be a valuable addition to the forensic toolbox, but implementation requires careful consideration of interpretation and reporting rules. Schneider et  al. (2019) propose that the report has to include which reference populations were part of the prediction algorithm. Investigators need to understand that results only reflect an individual’s historical biogeographical origin, which due to admixture and migration may or may not be of value in a forensic investigation.

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Table 64.10  Ethical and operational in forensic DNA phenotyping. Response and/or mitigation

Genetic privacy   Risk Sensitive information relating to disease states, disease susceptibility, or mortality (off target phenotypes) is revealed during testing.

• Physical appearance itself is public information and less sensitive to disclosure than other genetic traits. • Limit marker selection to loci not associated with internal traits, especially diseases. • If disease markers are inadvertently part of larger test panels, restrict interpretation and reporting to EVCs. • Do not share genetic data that could be mined for more information.

Racial discrimination   Risk Reporting on specific traits like skin color could result in stigmatization of ethnic minorities.

• FDP results do not necessarily incriminate; they may exclude members of vulnerable ethnic minorities. • Balance risk by restricting use of FDP to serious crimes and missing persons investigations. • Law enforcement training must include how to use information in a sensitive fashion without victimizing a subset of the population. • Phenotype information should be kept internal unless public participation is required.

Probabilistic nature of results   Risk Insufficient understanding of the level of uncertainty associated with FDP results can lead to exaggerated expectations and misdirected investigations.

• Effective communication of scientific results is an important part of any forensic application. • Reports should be designed to include a glossary on scientific terms and clearly state inherent limitations. • Criminal justice professionals must be trained on the value of each applicable test and the meaning of information obtained • Evidence submitted for testing must be from a single source and clearly linked to the crime.

Data storage   Risk FDP information could be stored in digital databases and be subject to unintended use or data loss.

• FDP data are only of temporary value during the investigation until a person of interest can be subjected to identity DNA typing. • Limit FDP to one-­off examination and testing without database entry. • Regulate access to and use of data within the laboratory and by forensic investigators. • Only share phenotype predictions rather than genetic data with stakeholders.

Unreliable tests   Risk Assays lack scientific validity and/or are not adequately tested for reliability prior to implementation

• • • •

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Mandate developmental and internal validations for all FDP test chemistry and software. Require transparency on protocols and published error rates for assays. Standardize result interpretation, reporting, and uncertainty estimators for different tests. Develop controls, standard, and proficiency tests for FDP assays.

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this result (Scudder et  al.  2018). Again, in light of the possible presence of disease information, genetic data for FDP investigations should not be stored and/or shared with other agencies instead distributed results should be limited to predicted phenotypes (Scudder et al. 2018). Table 64.10 summarizes the ethical and operational concerns and suggested solutions mentioned above. Careful implementation with appropriate legislation, training, and management of expectations is clearly needed for this new chapter in DNA typing. A concern for chronological age predictions is the unintended exposure of the biological age and the increased mortality risk associated with each 5-­year increase in methylation age (Fransquet et  al.  2019). Age determination for living individuals can become problematic, if insurance companies use methylation data to hold individuals accountable for their lifestyle or if the test is used in sensitive applications such as age estimation for refugees (Abbott  2018; Bell et  al.  2019). Operational concerns must also emphasize which sample types are suitable for FDP. The scientific requirement here is that the biological item needs to be from a single donor; from an

i­ nvestigative perspective, testing must be limited to evidence unequivocally linked to the crime. How widespread and useful FDP will be in routine casework remains to be seen. The least controversial application is clearly the generation of physical characteristics to supplement anthropological data for unidentified skeletal remains and missing person identification. The enhanced description and improved age determination will allow anthropologists and/or law enforcement agencies to better search missing person records and locate candidate families.

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Carney, C., Whitney, S., Vaidyanathan, J. et  al. (2019). Developmental validation of the ANDE rapid DNA system with FlexPlex assay for arrestee and reference buccal swab processing and database searching. Forensic Science International: Genetics 40: 120–130. https://doi. org/10.1016/j.fsigen.2019.02.016 Carracedo, A., Baer, W., Lincoln, P. et al. (2000). DNA commission of the International Society for Forensic Genetics: Revised and extended guidelines for mitochondrial DNA typing. Forensic Science International 110: 79–85. https://doi.org/10.1016/j.fsigen.2014.07.010 Carratto, T.M.T., Marcorin, L., Debortoli, G. et  al. (2020). Insights on hair, skin and eye color of ancient and contemporary Native Americans. Forensic Science International: Genetics 48: 102335. https:// doi.org/10.1016/j.fsigen.2020.102335 Cavanaugh, S.E. and Bathrick, A.S. (2018). Direct PCR amplification of forensic touch and other challenging DNA samples: A review. Forensic Science International: Genetics 32: 40–49. https://doi.org/10.1016/j. fsigen.2017.10.005 Cha, S., Lim, J.E., Park, A.Y. et  al. (2018). Identification of five novel genetic loci related to facial morphology by genome-­wide association studies. BMC Genomics 19: 481. https://doi.org/10.1186/ s12864-­018-­4865-­9 Chaitanya, L., Breslin, K., Zuñiga, S. et al. (2018). The HIrisPlex-­S system for eye, hair and skin colour prediction from DNA: Introduction and forensic developmental validation. Forensic Science International: Genetics 35: 123–135. https://doi.org/10.1016/j.fsigen.2018.04.004 Chaitanya, L., Walsh, S., Andersen, J.D. et  al. (2014). Collaborative EDNAP exercise on the IrisPlex system for DNA-­based prediction of human eye colour. Forensic Science International: Genetics 11: 241– 251. https://doi.org/10.1016/j.fsigen.2014.04.006 Chapman, B.R., Blackwell, S.J. and Müller, L.H. (2020). Forensic techniques for the isolation of spermatozoa from sexual assault samples – A review. Forensic Science Review 32 (2): 105–116. Cheung, E.Y.Y., Gahan, M.E. and McNevin, D. (2018). Predictive DNA analysis for biogeographical ancestry. Australian Journal of Forensic Sciences 50 (6): 651–658. https://doi.org/10.1080/00450618.2017.1422 021 Cho, S., Jung, S.E., Hong, S.R. et al. (2017). Independent validation of DNA-­based approaches for age prediction in blood. Forensic Science International: Genetics 29: 250–256. https://doi.org/10.1016/j. fsigen.2017.04.020 Cho, S., Kim, M.Y., Lee, J.H. & Lee, S.D. (2018). Assessment of mitochondrial DNA heteroplasmy detected on commercial panel using MPS system with artificial mixture samples. International Journal of Legal Medicine 132 (4): 1049–1056. https://doi.org/10.1007/ s00414-­017-­1755-­7 Churchill, J.D., Novroski, N.M.M., King, J.L. et  al. (2017). Population and performance analyses of four major populations with Illumina’s FGx Forensic Genomics System. Forensic Science International: Genetics 30: 81–92. https://doi.org/10.1016/j.fsigen.2017.06.004 Churchill, J.D., Stoljarova, M., King, J.L. and Budowle, B. (2018). Massively parallel sequencing-­enabled mixture analysis of mitochondrial DNA samples. International Journal of Legal Medicine 132 (5): 1263–1272. https://doi.org/10.1007/s00414-­018-­1799-­3 Claes, P., Hill, H. and Shriver, M.D. (2014). Toward DNA-­based facial composites: Preliminary results and validation. Forensic Science

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Thomasma, S.M. and Foran, D.R. (2013). The Influence of Swabbing Solutions on DNA Recovery from Touch Samples. Journal of Forensic Sciences 58: 465–469. https://doi.org/10.1111/1556-­4029.12036 Thompson, J.M., Ewing, M.M., Frank, W.E. et al. (2013). Developmental validation of the PowerPlex® Y23 System: A single multiplex Y-­STR analysis system for casework and database samples. Forensic Science International: Genetics 7: 240–250. https://doi.org/10.1016/j. fsigen.2012.10.013 Thong, Z., Phua, Y.H., Loo, E.S. et  al. (2015). Investigative leads from DNA: Casework experience from the IntegenX RapidHIT 200 System. Forensic Science International: Genetics Supplement Series 5: e69–e70. https://doi.org/10.1016/j.fsigss.2015.09.028 Tillmar, A.O., Kling, D., Butler, J.M. et al. (2017). DNA Commission of the International Society for Forensic Genetics (ISFG): Guidelines on the use of X-­STRs in kinship analysis. Forensic Science International: Genetics 29: 269–275. https://doi.org/10.1016/j.fsigen.2017.05.005 Tobe, S.S., Watson, N. and Daéid, N.N. (2007). Evaluation of six presumptive tests for blood, their specificity, sensitivity, and effect on high molecular-­weight DNA. Journal of Forensic Sciences, 52, 102–109. https://doi.org/10.1111/j.1556-­4029.2006.00324.x Tomas, C., Sanchez, J.J., Castro, J.A. et al. (2010). Forensic usefulness of a 25 X-­chromosome single-­nucleotide polymorphism marker set. Transfusion 50 (10): 2258–2265. https://doi.org/10.1111/j.1537-­2995.2010.02696.x Turingan, R.S., Tan, E., Jiang, H. et al. (2020). Developmental Validation of the ANDE 6C System for Rapid DNA Analysis of Forensic Casework and DVI Samples. Journal of Forensic Sciences 65: 1056–1071. https:// doi.org/10.1111/1556-­4029.14286 Tvedebrink, T., Eriksen, P.S., Curran, J.M. et  al. (2012). Analysis of matches and partial-­matches in a Danish STR data set. Forensic Science International: Genetics 6: 387–392. https://doi.org/10.1016/j. fsigen.2011.08.001 Valenzuela, R.K., Henderson, M.S., Walsh, M.H. et al. (2010). Predicting phenotype from genotype: Normal pigmentation. Journal of Forensic Sciences 55: 315–322. https://doi.org/10.1111/j.1556-­4029.2009.01317.x Vallone, P.M., Just, R.S., Coble, M.D. et  al. (2004). A multiplex allele-­ specific primer extension assay for forensically informative SNPs distributed throughout the mitochondrial genome. International Journal of Legal Medicine 118: 147–157. https://doi.org/10.1007/s00414-­004-­ 0428-­5 van Oorschot, R.A.H., Ballantyne, K.N. and Mitchell, R.J. (2010). Forensic trace DNA: A review. Investigative Genetics 1: 14. https://doi. org/10.1186/2041-­2223-­1-­14 van Oorschot, R.A.H., Goray, M., Szkuta, B. et al. (2019). DNA Transfer in Forensic Science: A Review 38: 140–166. https://doi.org/10.1016/j. fsigen.2018.10.014 van Oorschot, R.A.H. and Jones, M.K. (1997). DNA fingerprints from fingerprints. Nature 387: 766–767. van Oorschot, R.A.H., Treadwell, S., Beaurepaire, J. et al. (2005). Beware of the possibility of fingerprinting tehcniques transferring DNA. Journal of Forensic Sciences 50: 1417–1422. Vandenberg, N. and Van Oorschot, R.A.H. (2006). The use of Polilights?? in the detection of seminal fluid, saliva, and bloodstains and comparison with conventional chemical-­ based screening tests. Journal of Forensic Sciences 51: 361–370. https://doi.org/10.1111/j.1556-­ 4029. 2006.00065.x

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Walsh, S., Liu, F., Wollstein, A. et al. (2013). The HIrisPlex system for simultaneous prediction of hair and eye colour from DNA. Forensic Science International: Genetics 7: 98–115. https://doi.org/10.1016/j. fsigen.2012.07.005 Wang, D.Y., Gopinath, S., Lagacé, R.E. et al. (2015). Developmental validation of the GlobalFiler Express PCR Amplification Kit: A 6-­dye multiplex assay for the direct amplification of reference samples. Forensic Science International. Genetics 19: 148–155. https://doi. org/10.1016/j.fsigen.2015.07.013 Wang, Q., Jin, B., Luo, X. et al. (2017). Association between BMP4 gene polymorphisms and eyelid traits in Chinese Han population. Forensic Science International: Genetics Supplement Series 6: e355–e356. https:// doi.org/10.1016/j.fsigss.2017.09.112 Wang, Z., Wang, L., Liu, J. et al. (2020). Characterization of sequence variation at 30 autosomal STRs in Chinese Han and Tibetan populations. Electrophoresis 41: 194–201. https://doi.org/10.1002/elps.201900278 Wang, Z., Zhu, R., Zhang, S. et  al. (2015). Differentiating between monozygotic twins through next-­generation mitochondrial genome sequencing. Analytical Biochemistry 490: 1–6. https://doi. org/10.1016/j.ab.2015.08.024 Weber-­Lehmann, J., Schilling, E., Gradl, G. et al. (2014). Finding the needle in the haystack: Differentiating “identical” twins in paternity testing and forensics by ultra-­deep next generation sequencing. Forensic Science International: Genetics 9: 42–46. https://doi.org/10.1016/j. fsigen.2013.10.015 Weidner, C.I., Lin, Q., Koch, C.M. et al. (2014). Aging of blood can be tracked by DNA methylation changes at just three CpG sites. Genome Biology 15: R24. https://doi.org/10.1186/gb-­2014-­15-­2-­r24 Westen, A.A., Matai, A.S., Laros, J.F.J. et al. (2009). Tri-­allelic SNP markers enable analysis of mixed and degraded DNA samples. Forensic Science International: Genetics 3: 233–241. https://doi.org/10.1016/j. fsigen.2009.02.003 Wickenheiser, R.A. (2004). The business case for using forensic DNA technology to solve and prevent crime. Journal of Biolaw and Business 7: 34–50. Wickenheiser, R.A. (2019). Forensic genealogy, bioethics and the Golden State Killer case. Forensic Science International: Synergy 1: 114–125. https://doi.org/10.1016/j.fsisyn.2019.07.003 Wiegand, P. and Kleiber, M. (1997). DNA typing of epithelial cells after strangulation. International Journal of Legal Medicine 110: 181–183. https://doi.org/10.1007/s004140050063 Wienroth, M. (2018). Governing anticipatory technology practices. Forensic DNA phenotyping and the forensic genetics community in Europe. New Genetics and Society 37: 137–152. https://doi.org/10.108 0/14636778.2018.1469975 Wiley, R., Sage, K., LaRue, B. and Budowle, B. (2017). Internal validation of the RapidHIT® ID system. Forensic Science International: Genetics 31: 180–188. https://doi.org/10.1016/j.fsigen.2017.09.011 Willuweit, S. and Roewer, L. (2015). The new Y chromosome haplotype reference database. Forensic Science International: Genetics 15: 43–48. https://doi.org/10.1016/j.fsigen.2014.11.024 Wolf, A., Caliebe, A., Junge, O. and Krawczak, M. (2005). Forensic interpretation of Y-­ chromosomal DNA mixtures. Forensic Science International 152: 209–213. https://doi.org/10.1016/j.forsciint.2004. 07.021

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Verdon, T.J., Mitchell, R.J. and Van Oorschot, R.A.H.H. (2014). Evaluation of tapelifting as a collection method for touch DNA. Forensic Science International: Genetics 8: 179–186. https://doi. org/10.1016/j.fsigen.2013.09.005 Verheij, S., Harteveld, J. and Sijen, T. (2012). A protocol for direct and rapid multiplex PCR amplification on forensically relevant samples. Forensic Science International: Genetics 6: 167–175. https://doi. org/10.1016/j.fsigen.2011.03.014 Vernarecci, S., Ottaviani, E., Agostino, A. et al. (2015). Quantifiler® Trio Kit and forensic samples management: A matter of degradation. Forensic Science International: Genetics 16: 77–85. https://doi. org/10.1016/j.fsigen.2014.12.005 Verogen. (2020). How Next Generation Sequencing Resolved a Difficult Case, Leading to the First Criminal Conviction of Its Kind (Case Study). https://cdn2.hubspot.net/hubfs/6058606/Verogen-­First-­NGS-­­Court-­ Case-­Study_Final_VD2019024_8.5x11-­web.pdf?__hstc=238609695. bed74b81cf4041e42adad16833ab8584.1576870704888.1576870 704888.1576870704888.1&__hssc=238609695.1.1576870704888 (last accessed November 12 2020). Vidaki, A., Ballard, D., Aliferi, A. et al. (2017). DNA methylation-­based forensic age prediction using artificial neural networks and next generation sequencing. Forensic Science International: Genetics 28: 225– 236. https://doi.org/10.1016/j.fsigen.2017.02.009 Vidaki, A., Díez López, C., Carnero-­Montoro, E. et al. (2017). Epigenetic discrimination of identical twins from blood under the forensic scenario. Forensic Science International: Genetics 31: 67–80. https://doi. org/10.1016/j.fsigen.2017.07.014 Vidaki, A., Kalamara, V., Carnero-­Montoro, E. et al. (2018). Investigating the epigenetic discrimination of identical twins using buccal swabs, saliva, and cigarette butts in the forensic setting. Genes 9: 252. https:// doi.org/10.3390/genes9050252 Vidaki, A. and Kayser, M. (2018). Recent progress, methods and perspectives in forensic epigenetics. Forensic Science International: Genetics 37: 180–195. https://doi.org/10.1016/j.fsigen.2018.08.008 Vieira-­Machado, C.D., Tostes, M., Alves, G. et  al. (2016). Uniparental ancestry markers in Chilean populations. Genetics and Molecular 4685-­ Biology 39 (4): 573–579. https://doi.org/10.1590/1678-­ GMB-­2015-­0273 Virkler, K. and Lednev, I.K. (2009). Analysis of body fluids for forensic purposes : From laboratory testing to non-­destructive rapid confirmatory identification at a crime scene. Forensic Science International 188: 1–17. https://doi.org/10.1016/j.forsciint.2009.02.013 Wagner, J., Džijan, S., Marjanovié, D. and Lauc, G. (2009). Non-­invasive prenatal paternity testing from maternal blood. International Journal of Legal Medicine 123: 75–79. https://doi.org/10.1007/s00414-­008-­0292-­9 Walsh, P., Metzger, D. and Higuchi, R. (1991). Chelex 100 as a medium for simple extraction of DNA for PCR-­based typing from forensic material. BioTechniques 10: 506–513. Walsh, S., Chaitanya, L., Breslin, K. et al. (2017). Global skin colour prediction from DNA. Human Genetics 136: 847–863. https://doi. org/10.1007/s00439-­017-­1808-­5 Walsh, S., Liu, F., Ballantyne, K.N. et al. (2011). IrisPlex: A sensitive DNA tool for accurate prediction of blue and brown eye colour in the absence of ancestry information. Forensic Science International: Genetics 5: 170–180. https://doi.org/10.1016/j.fsigen.2010.02.004

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You, Y. and Balding, D. (2019). A comparison of software for the evaluation of complex DNA profiles. Forensic Science International: Genetics 40: 114–119. https://doi.org/10.1016/j.fsigen.2019.02.014 Yuan, L., Chen, X., Liu, Z. et al. (2020). Identification of the perpetrator among identical twins using next-­generation sequencing technology: A case report. Forensic Science International: Genetics 44: 102167. https://doi.org/10.1016/j.fsigen.2019.102167 Zapata, F., Fernández de la Ossa, M.Á. and García-­Ruiz, C. (2015). Emerging spectrometric techniques for the forensic analysis of body fluids. TrAC – Trends in Analytical Chemistry 64: 53–63. https://doi. org/10.1016/j.trac.2014.08.011 Zbieć-­Piekarska, R., Spólnicka, M., Kupiec, T. et al. (2015). Development of a forensically useful age prediction method based on DNA methylation analysis. Forensic Science International: Genetics 17: 173–179. https://doi.org/10.1016/j.fsigen.2015.05.001 Zieger, M. and Utz, S. (2015). About DNA databasing and investigative genetic analysis of externally visible characteristics: A public survey. Forensic Science International: Genetics 17: 163–172. https://doi. org/10.1016/j.fsigen.2015.05.010 Zubakov, D., Liu, F., Kokmeijer, I. et al. (2016). Human age estimation from blood using mRNA, DNA methylation, DNA rearrangement, and telomere length. Forensic Science International: Genetics 24: 33–43. https://doi.org/10.1016/j.fsigen.2016.05.014

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Wollstein, A., Walsh, S., Liu, F. et al. (2017). Novel quantitative pigmentation phenotyping enhances genetic association, epistasis, and prediction of human eye colour. Scientific Reports 7: 43359. https://doi. org/10.1038/srep43359 Wood, A.R., Esko, T., Yang, J. et al. (2014). Defining the role of common variation in the genomic and biological architecture of adult human height. Nature Genetics 46 (11): 1173–1186. https://doi.org/10.1038/ ng.3097 Miguel, A. et  al. (2020). Xavier, C., de la Puente, M., Mosquera-­ Development and validation of the VISAGE AmpliSeq basic tool to predict appearance and ancestry from DNA. Forensic Science International: Genetics 48: 102336. https://doi.org/10.1016/j. fsigen.2020.102336 Xavier, C., de la Puente, M., Phillips, C. et al. (2020). Forensic evaluation informative MAPlex assay. Forensic of the Asia Pacific ancestry-­ Science International: Genetics 48: 102344. https://doi.org/10.1016/j. fsigen.2020.102344 Xiong, Z., Dankova, G., Howe, L.J. and Lee, M.K. (2019). Novel genetic loci affecting facial shape variation in humans. ELife 8: e49898. Xu, C., Qu, H., Wang, G. et al. (2015). A novel strategy for forensic age prediction by DNA methylation and support vector regression model. Scientific Reports 5: 17788. https://doi.org/10.1038/ srep17788

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Some methods, such as DNA analysis and fingerprinting, have managed to quantify the degree of correspondence of two biological samples using a statistical analysis of the frequency of the discrete characters (e.g. alleles for DNA and minutiae for fingerprint analysis). This can lead the expert to be able to express the probability of these two samples belonging to the same person as a number. Methods that allow the operator to draw such indications are very popular, especially after the introduction of Daubert rulings in the judicial context. These rulings state that the content of testimony in trial must be testable and have been tested through a scientific method, have been subjected to peer review, have a known error rate and have been widely accepted by the scientific community. These indications result in methods being used that provide a quantitative indication concerning their error, independently from the type of rational procedure used to reach such a result. For example, in DNA analysis, available data are based on the epidemiological frequency of the chosen alleles, which gives a general probability of finding two individuals with the same genetic profile. This number only expresses a probability, and it does not state that the decedent is actually the identity suspect. Other methods, mainly based on the study of morphology (e.g. osteological methods), cannot provide a real quantification of the obtained results; for example, expressing the probability of finding two persons with the same scar or tattoo is very difficult and depends on several variables. However, scars and tattoos are still the most frequently used elements for identification, along with bone and dental prostheses, bone calluses and the bone profile itself when adequate ante-­mortem data are available. All these methods are usually considered reliable enough to reach personal

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Personal identification is a key issue in forensic anthropology and consists of the correct assignment of an identity to an unknown corpse. Although it may seem an easy and automatic procedure, the mechanism of identification encompasses several difficulties concerning the methods and, above all, the significance of the identification process in court. The process of identification strives towards two conclusions: exclusion and positive identification. However, between them there is a series of nuances that can be summed up as different degrees of compatibility, or better yet, ‘consistency’. There are cases where there are no elements for exclusion and the concordant characters are very general or with a low identification potential, and therefore there is not enough evidence to reach a positive identification. How much is sufficient to identify an individual is a central question, which unfortunately is without a real answer in the case of most morphological methods. Opinions differ on how to express these shades of grey: for example, different experts make statements of ‘identification-­ proven bordering on certainty’ or ‘identification extremely likely/ probable/not determinable’. Other authors from the Forensic Imagery Analysis Group (FIAG, born within BAHID, the British Association for Human Identification) have established a common procedure for describing the level of ascertainment of personal identification, ranging between ‘no support’ to ‘powerful support’. These are all, however, subjective judgments that lack the apparent strength of a quantitative method but which are still useful and valid.

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personal identification does not exist in morphological methods. The occurrence of discordant characters may settle the case by excluding the identity. Many concordant characters, particularly if fairly common within the population, can only give a judgment of compatibility or possibility. However, there are some characters, rare among the population, that will allow one to express a judgment of high probability or certainty. Prosthetic devices may be useful, particularly if they bear a serial number that could trace the device back to a hospital, a surgeon or a specific patient (Figure 65.1). Usually, identification based on bone morphology is carried out by comparing an ante-­mortem X-­ray of the missing person and a post-­mortem X-­ray of the same anatomical region. As far as the comparison of ante-­mortem and post-­mortem X-­rays is concerned, every part of the body may provide useful information for identification; in particular, the head offers useful features such as frontal sinuses and cranial suture patterns. The frontal sinuses are widely used and studied, and different authors have reported cases of positive identification using X-­rays of the head and comparing the skeletal features. For frontal sinuses, there are algorithms that allow one to compare shape, size, septae, etc. with a total score. The comparison of frontal sinuses is one of the most reliable methods of identification since they are also different in homozygous twins. Recent clinical approaches have seen the use of computer tomography (CT) scans and magnetic resonance imaging (MRI) instead of plain radiographs (Figure  65.2). From these imaging analyses, an important aid to identification may be provided by the extraction of a 3D model of frontal sinuses through a segmentation procedure, nowadays widely applied to different clinical and surgical fields of research (Yushkevich et  al.  2006; Figure  65.3). In this way, 3D models of frontal sinuses can be extracted and registered to assess possible differences in surface of air cavities. The difference between two 3D surfaces can then be quantified through the calculation of point-­ to-­ point mean distance (Figures  65.4 and  65.5). The three-­dimensional analysis may provide more individual information than the mere bi-­dimensional silhouette on X-­rays, improving the chances of identification. The first studies on this novel method of identification are promising for what concerns the distinction between matches and mismatches (Gibelli et  al.  2019); moreover, the progressive diffusion of CT scans in the next future will increase the availability of such analyses as ante-­mortem material. The same procedure has also been successfully applied to other anatomical structures, including the sphenoid sinuses (Cappella et al. 2019) and the lumbar vertebrae (Decker and Ford 2019). The thorax has also proven very useful in several forensic cases because of the remarkable number of features offered by this area and the notable availability of thoracic X-­rays in the population. The most useful features are the normal anatomical ones, such as vertebral, costal and clavicular morphology. However, the comparison of the shapes of bones can be carried out for any area of the body. Comparison of the trabecular design of cancellous bone has even been attempted, with unclear results. In all these procedures, it is of paramount importance that the comparison be

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identification, but they do not provide a quantitative indication concerning the probability of having reached a correct identification. From a general point of view, scars, protheses, signs of ante-­ mortem trauma and so on are the most useful since they are all morphological features which, in their way, are unique. Many individuals may have a bone callus on the right radius, but that specific callus with its morphology only belongs to a single individual. The same is only valid for signs of thoracotomy – many individuals have undergone this surgical procedure for a number of reasons, but the specific arrangement of metallic sutures and the bony reaction is unique to each single person. Therefore, these characteristics are the most suitable for a correct identification, even though they cannot be quantitatively expressed. For example, the information that an unknown decedent has a bone callus on the right radius is not enough to identify him or her since this characteristic may be shared by many individuals. However, if there is an ante-­mortem X-­ray that allows the morphological profile of the callus to be examined, this marker may be enough to reach a positive identification. The chances of another individual sharing the same morphology of a callus are not quantifiable, which puts osteological and odontological methods in a different category compared with DNA and fingerprints. In the USA, the recent Kumho rulings have softened the Daubert criteria for admission of anthropological testimony in trial, and in detail have made requisites more flexible, in order to meet the morphological nature of anthropological comparative methods. One of the general criteria states that expert witnesses can develop theories based on their observations and experiences and then apply theories to the case before the court. The importance of experience, therefore, enters the trial once again. FASE (the Forensic Anthropology Society of Europe) has recently made a statement on how reliable anthropology may be properly used for positive identification when all else fails (DeBoer H. et al. 2020), and facial traits as well as soft tissue markers are also regaining significance in more scientific manner (Caplova et al. 2018). Regardless of such philosophical, yet fundamental, matters, the morphology of bones may be fundamental for identification, and the use of osteology in the personal identification of both living and dead humans is discussed below.

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65.2  Identification of human remains Personal identification requires the analysis of specific and individualising characteristics observed in the unknown decedent and their comparison with similar data from the identity suspect. Identification may be reached by different methods. Morphological methods (forensic anthropology and odontology techniques), which compare the status and shape of teeth and bones, are advantageous because they are faster and less costly. However, as discussed, a unanimous and clear agreement on the quality and quantity of the characters necessary to achieve

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Figure 65.1  Femoral prothesis; in the red circle, the details of the series number are useful for identifying the subject.

Figure 65.2  Example of identification by comparison of the frontal sinuses: (a) post-­mortem X-­ray; (b) ante-­mortem X-­ray; (c) superimposition of the two profiles.

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Figure 65.3  Finding of metallic sutures on the sternal manubrium of skeletal remains provides signs of a surgical thoracotomy. The morphology of the steel sutures was useful in identifying the subject by comparison with an ante-­mortem chest X-­ray.

Figure 65.4  Example of the segmentation of frontal sinuses from a CT scan: in the lower left box, the 3D-­segmented model.

performed by someone who also has experienced in radiology since the issue of orientation and the correct interpretation of the various hues of grey in an X-­ray is fundamental. Signs of a thoracotomy are also useful and are beginning to be frequently found in the general population. The position and shape of the metallic

sutures may help in reaching a positive identification (Figure 65.6) since patients undergoing thoracotomy usually have several X-­ray scans that may be used as ante-­mortem material. A ‘last chance’ method, though not reliable on its own for identification, is craniofacial superimposition (Figure 65.7). This

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Figure 65.5  3D–3D surface superimposition of frontal sinuses extracted from CT scans belonging to the same individual (match): (a) extracted 3D models; (b) superimposed models; (c) results of point-­to-­point distance calculation between the two models: prevalence of green colour indicated correspondence between the two surfaces.

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Figure 65.6  3D–3D surface superimposition of frontal sinuses extracted from CT scans belonging to different individuals (mismatch): (a) extracted 3D models; (b) superimposed models; (c) results of point-­to-­point distance calculation between the two models: prevalence of yellow, red and blue colours indicated prevalence of discordant areas between the two surfaces.

consists of the superimposition of a photograph of the skull onto a similarly orientated photograph of the face of the living person. The correspondence between the two subjects is evaluated on the basis of matching several anatomical landmarks. After the selection of the images of the missing person, the best spatial orientation of the cranium should be achieved. Once the correct

orientation is gained, the superimposition is performed on both frontal and lateral views if possible. The criteria for the comparison in the frontal view are usually the following: 1.  The length of the cranium from bregma to menton must be included within the face (caution: the bregma may be covered by hair).

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bridge closely follow the contour of the thin layer of overlying skin. The nasal bones fall within the margins of the nose. 10.  The front nasal spine is situated posteriorly with respect to the base of the nose, close to the more posterior portion of the lateral septum cartilage. 11.  The prosthion should be posterior to the anterior margin of the superior lip. 12.  The pogonion is posterior to the indenting noticeable in the chin where the orbicularis oris crosses the chin muscle. Many other criteria for matching landmarks and general morphology exist although no common protocols have been agreed upon. Clearly, this superimposition has far less credibility than comparison methods based on dental or skeletal structures, which usually involve similar structures (in ante-­mortem and post-­mortem materials). In craniofacial superimposition, the soft tissues are compared with the skeleton, with consequent difficulties in localising the position of skeletal landmarks on the facial image (Figure 65.4). Supporters of this method report a reliability of 96% when both frontal and lateral superimposition is possible, but there is a large discrepancy in the degree of accuracy found in different investigations. This method should never be used on its own for identification, but it can be used for excluding an identity if gross incompatibilities are present. Craniofacial superimposition improved in recent years, along with other forensic fields of research, due to the introduction of the new technologies. In particular, there is now specific software that is able to match a photograph and a three-­dimensional model of the skull obtained by modern systems of three-­dimensional acquisition such as laser scanner and stereophotogrammetry. In conclusion, in order to identify a person, it is possible to compare the shape of bones, even if these methods have not yet been ‘standardised’ from a quantitative point of view. The comparison can be carried out between ante-­ mortem and post-­ mortem radiographs of different skeletal areas. If radiographs of the various bones are available, it is possible to compare the morphology of the skeletal elements visible in the radiograph with the

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temporal cranial line. 4.  The eyebrows generally follow the superior edge of the orbit to the medial and central third, and they continue superiorly to the lateral third, whereas the edge of the orbit deviates inferiorly. 5.  The orbits should contain the eyes entirely. 6.  The lachrymal groove, if distinguishable on the photo, should line up with the osseous groove. 7.  The width of the nasal bridge must correspond in the two images. 8.  The width of the nasal aperture should fall within the external margins of the nose. 9.  The nasal spine should be situated above the inferior edge of the medial nasal crus. 10.  The external auditory meatus should be medial to the tragus. 11.  The oblique line of the mandible, where visible, should correspond to the same line on the skull. 12.  The mandibular curve should be similar to the curve visible on the face. In the lateral view, the criteria are the following: 1.  The skullcap must correspond to the height of the head. 2.  Sometimes it is possible to observe the margin of the frontal process of the zygomatic bone that should match the one on the cranium. 3.  The margin of the zygomatic arch of the cranium is also superimposable to that of the face. 4.  The portion should be slightly behind the tragus and below the helix crus. 5.  The occipital curve should lie inside the margin of the nape. 6.  The anterior protuberance of the mandible is behind the chin: the chin shape should correspond to the mandible shape. 7.  The lateral margin of the eye is situated within the orbit. 8.  The profile of the glabella both on the cranium and the face must be similar.

9.  The prominence of the glabella and the depth of the nasal

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Figure 65.7  Example of craniofacial superimposition in the frontal and lateral views. The letters (frontal) and the numbers (lateral) indicate the landmark for which the correspondence between the face and skull is evaluated.

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consists of using a virtual camera with the same characteristics of the video surveillance system with which the images had been shot (De Angelis et al. 2007). The results demonstrate that height is a parameter that can be accurately estimated. The experimental data showed an average difference between the estimated and real height of about 10  mm together with a standard deviation of 10 mm. At the moment, height estimation is one of the most precise indications that may be drawn by two-­dimensional images if adequate corrections of optical distortions are set up. One limitation is that this procedure requires that the individual is represented in his or her entire figure from the feet to the head (Figure 65.8). Gait analysis also may provide some information as it is based on individual characteristics: however, prevalent literature agrees in recognising that gait analysis is still of limited scientific basis to be practically applied; main limits are the incomplete knowledge of intra-­and inter-­subject variability, limited databases of gait features, scarce data about dependency among different gait features, and the lack of standardisation in methods for assessing step movements (van Mastrigt et al. 2018). A more specific step towards personal identification is that of facial assessment. Classic literature refers to facial characters and indices (i.e. conventional anthropometry) in order to verify the resemblance between two individuals. Ear shape in particular has received some attention. It is, however, limited by the same pitfalls as for any character of the face in general. The danger in comparing faces on two-­dimensional images is only too well known: slight, imperceptible differences in orientation and facial expression, as well as inter-­observer differences in setting facial landmarks, may cause drastic errors in the study of indices or other parameters. Several morphological classifications and atlases of faces have been devised (Interpol, Vanezis, DMV) which are used to classify faces and thus verify the degree of resemblance of two faces. One of the most advanced morphological classifications was developed in the course of a project funded by the European Union, involving researchers from Germany, Italy and Lithuania, called DMV (from the universities of Düsseldorf, Milan and Vilnius, which participated in the project). Reference photographs of 900 males aged between 20 and 31 years in the three countries were acquired, and facial features were described and evaluated. These efforts led to the first edition of an anthropological atlas published in 2007, which includes detailed descriptions and drawings and photographic examples of 43  morphological features of male faces. In 2008, the atlas of male facial features was revisited, and population frequencies for the facial features were added (Ohlrogge et al. 2008). In 2009, an atlas of female facial features was developed (Ohlrogge et al. 2009). These two atlas editions include examples of 45 facial traits and allow a morphological assessment of both male and female faces (Figure 65.9). This model was tested in order to verify its reliability in the recognition of facial features. The intra-­observer mismatch percentages based on all features when using the DMV atlas ranged from 19% to 30%, whereas the inter-­observer error was approximately 39% (Ritz-­Timme et al. 2011). These results confirm that

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same osseous elements belonging to the cadaver or human remains. However, for specific exceptional morphological peculiarities such as osteophyte conformations, bone calluses, etc. the difficulty consists in determining which and how many of the morphological elements are sufficient for a definite identification in a normal, non-­pathological bone. The literature states that personal identification requires a meticulous comparison of details, but a minimum number of points does not exist. In some instances, one to four characteristics, without evident discrepancies, have been considered sufficient for identification. However, the peculiarity of the characteristic and the consistency between the ante-­mortem and post-­mortem shapes is often left to a subjective evaluation and to the experience of the operator. Prosthetic devices may be of relevant help in personal identification since most of them carry labels with numbers which can lead to a hospital or even to a single patient. However, laws concerning the registration of prosthetic devices vary among different countries and therefore this may not be as simple as it seems.

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Identification issues in forensic anthropology usually involve unknown decedents. However, age estimation in the living can be requested in cases of adoption, for ascertaining the status of imputability in sub-­adults, and retirement age in adults, and may be preliminary to a complete identification procedure. This issue has been discussed in later chapters of the book. In the last decade, however, anthropologists have been more and more involved in the identification of the living from images, for example from surveillance systems (Gibelli et al. 2016). Personal identification of the living from video surveillance systems, or from photographs or images in general, is in fact becoming more and more of an issue due to the number of terrorist attacks and crimes filmed on tape. Nowadays, judges often ask for ‘anthropometric’ experts in order to verify if a person seen on a specific video or photograph is the same person as the suspect. This topic deals with human diversity and strives to verify morphological and metric characteristics that make the physiognomy of one person distinct from that of another. This is an extremely difficult task for a series of reasons, which range from the frequent poverty of the images (little detail and poor resolution) to the lack of standardised protocols and studies in the field. Most experts agree that one of the preliminary procedures in cases of personal identification of the living consists of estimating the height of the individual, if relevant and possible. Height is a quantitative characteristic and, although it is not individualising, may provide useful information for comparison of the culprit and the suspect. The most recent studies concerning this topic highlighted that the reliability and potential of height estimation in personal identification depends on the rarity of the estimated perpetrator’s height and its closeness to the suspect’s height. In 2007, a new method for height estimation was published that

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65.3  Identification of the living

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Figure 65.8  Example of height estimation of a culprit from a video surveillance system: reconstruction of the three-­dimensional environment and estimation of the entire figure represented in the image.

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morphological classifications, although detailed and ameliorated with new facial features, are affected by an unavoidable subjectivity. The morphological approach therefore may provide a general and preliminary evaluation concerning the concordance (or discordance) between the culprit and the suspect, but it is not reliable enough to provide a definitive identification. The metric assessment of faces has also been employed through the decades using the measurement of facial dimensions and indices. However, this approach was severely criticised by a study that highlighted the unreliability of metrical measurements taken on two-­dimensional pictures for identification purposes (Kleinberg et al. 2007). More recently, another approach has found its place in the literature and practice, using a new method for personal identification from video surveillance systems. Superimposition can be performed with 2D images from the culprit and the suspect (2D–2D comparison) or between the 2D image of the culprit taken from the video surveillance system and a 3D virtual model of the suspect by a 3D optical digitiser (2D–3D comparison) (Figure 65.10). The 2D–2D comparison methods require that the images represent the suspect in the exact same position as the culprit in the video surveillance image – and this may be difficult to achieve. Moreover, the dangers of comparing faces on 2D images are well known: slight differences in orientation and facial expression as well as inter-­observer differences in set-

ting facial landmarks may cause drastic errors. The 2D–3D comparison method is therefore supposed to be more reliable for personal identification since it allows one to record 3D facial models and to create specific image database comparison procedures. Image acquisition is obtained by a laser scanner, with the reconstruction of a 3D model of the suspect; the comparison with the 2D image is performed by resizing, repositioning and reorientating single images with specific software until the best match with the culprit’s face is achieved. In this way, the differences in head orientation between the suspect and the culprit are considerably reduced. The 3D model of the suspect and its superimposition onto the 2D image of the culprit allows examiners to obtain an easy-­to-­read comparison and to assess the match as concern identification. The lack of a standardised method for assessing positive identification, however, still looms over every method. How much of a match is enough to identify? The quantitative assessment of a match between two profiles, for example, may be necessary to determine the probability rate of a correct identification. At the moment, there is no system detailing how similar two profiles should be in order to conclude that they belong to the same individual. A recent study aimed at devising a method of quantifying the match between 3D and 2D images based on the comparison of a simple projection of the face, the profile, derived from 2D

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Form of the chin contour in frontal view

FIG: 3.33.1.

FIG: 3.33.3.

2. Square

FIG: 3.33.4.

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Possible characteristics: 1. Round 2. Square 3. Pointed

Photographic example: 2. Square

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Figure 65.9  Example of morphological classification of facial features. Source: Reproduced with permission from Ohlrogge et al. (2009), © Verlag für Polizeiwissenschaft, Frankfurt, Germany.

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images and 3D models (Cattaneo et al. 2012). It was also trying to find the mean differences between two facial profiles in lateral view obtained from the same person and with the superimposition of different persons. The statistical analysis showed that the difference in areas seems to be a reliable method for evaluating identification correctly. In non-­correct matches, the difference in area was always higher than 157 mm2, whereas in correct matches the highest difference was between 43 and 133  mm2. A similar result was obtained by the overall difference of points between the 3D model and 2D image: in the case of non-­ matches, the difference was always higher than 1.96 mm, whereas in the correct matches this value was always lower than 1.9 mm. Both for areas and distances of all points, the incorrect and correct matches did not show any superimposition of values, and therefore seemed to provide a valid starting point for the diagnosis of identification. This experimental project showed that a clear method for identifying may exist, but requires more precise standardisation and further studies (Cattaneo et al. 2012).

65.4  The issue of quantification Anthropology naturally deals with identification as it ‘manages’ morphological and anatomical features, deriving both from genetics and environment, which represent a unique sample of markers useful to distinguish an individual from the other. Although anthropological methods can be considered as the most natural and immediate to provide identification, as mentioned, in the forensic practice, they suffer the limit of quantification as data concerning shape and position are difficult to describe through numbers, and therefore they usually cannot provide a percentage of correct (or incorrect) identification. This is the most relevant point of weakness of morphological methods in comparison with ‘quantitative’ methods such as DNA, which on the other hand can give a probability of positive identification, based on allelic frequencies. Quantification provides more readable results by judicial authorities and would allow skeletal methods of identification to gain popularity in comparison with genetic tests and other methods that provide a quantification of results.

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Figure 65.10  Example of a three-­dimensional model of the suspect by laser scanner superimposed on the face of the culprit from a video surveillance system image.

eral population. For example, signs of thoracotomy are enough to provide a conclusive identification (one has to think about the position of metallic sutures, or the type of bone remodelling, the shape of sternum, etc.) as are specific calluses and entheses on bones (Biehler-­Gomez and Cattaneo 2020). As previously mentioned, FASE’s recent statement has given clarity concerning anthropology as a tool for identification (DeBoer et al. 2020). Anthropologists find their main role today in forensics as co-­ workers on cases with pathologists or other experts for positive identification of human remains or of the living and as mentioned in Chapter 12 for scene of crime analysis, search and recovery of buried remains as well as building the biological profile (sex, age, stature, ancestry and pathology). A brief mention should be made in the role of osteology in aiding with trauma analysis, especially blunt, sharp force and gunshot wounds.

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Several attempts at giving a number to personal identification can be observed in literature: for example, some authors attempted at providing “rules of thumb” based on an arbitrary number of discrete traits needed to reach a conclusive diagnosis (Fuschman 1985). Another field of research concerns the validation studies, where the quantification is reached by the assessment of disagreement given by operators in assigning ante-­mortem and post-­ mortem materials (Rich et al. 2002; Koot et al. 2005; Mundorff et al. 2006; Stephan et al. 2011). The most problematic, and yet apparently natural approaches deal with the estimation of frequencies of specific skeletal traits, in order to replicate the same methodology by genetic tests, based on allelic frequencies. Yoshino et al. (1987) provided a classification of frontal sinuses into categories involving morphology and size and gave the frequencies of each type of frontal sinuses. However, the given percentages are far from confirming or even supporting a positive identification. Moreover, recent literature verified that morphological and metric characteristics of frontal sinuses are reciprocally correlated, thus increasing the frequency of specific profiles (Cameriere et al. 2020). Another contribution was given by Komar and Lathrop who provided the frequencies of different bone features (calluses, amputation, surgical devices, etc.) in different cemetery samples (Komar and Lathrop 2006): results found that the percentage of presentation of each feature is too high to provide by itself criteria for personal identification. These examples confirm that the uniqueness of skeletal features concern their specific shape and not their frequency in gen-

65.5 Not only identification: the analysis of trauma Identification is not the only task of forensic anthropologists, being the assessment of bone trauma one of the most difficult and tricky for the pathologist and at times the anthropologist’s expertise may be useful. In cases of traumatic signs, forensic anthropologists are usually requested to give information about different questions, from the age of production of lesions to the type of weapon used.

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tion of the fracture line with the osteon structure (Pechnikova et al. 2011) although their reliability is still to be verified. Once the fracture is assessed as peri-­mortem, the second step consists in verifying the number and sequence of lesions: in this case, Puppe’s rule which states that fracture lines from a blow are stopped by those produced by the pre-­existing ones (Puppe 1903) may help in determining which lesions were produced first. Finally, the anthropological analysis can focus on the specific weapon used for the production of lesions: the morphological characteristics may provide information concerning the type of lesion (blunt, sharp force trauma and gunshot wound) (Figures 65.11–65.13). Moreover, in cases of sharp force lesions, the macroscopic and microscopic information may help in

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One of the first issues which needs to be addressed concerns the classification of traumatic lesions within the categories of ante-­ mortem, peri-­mortem or post-­mortem. Post-­mortem fractures include lesions that have occurred on dry bone, characterised by low elasticity, with well-­defined fractures and lighter edges (in colour) than the remaining bone surface, and are relatively easy to recognise as well as ante-­mortem fractures, which show signs of remodelling (Berryman and Haun  1996). The main issue concerns peri-­mortem lesions: criteria used to make this diagnosis mainly deal with the morphological aspect of the fracture, usually showing ‘elastic’ breaks at its edges, with fragments attached one to the other (‘green bone’ aspect; Blau and Ubelaker 2009). However, the definition peri-­mortem covers a wide period, between the period immediately before and after death, as long as bone elasticity can be verified: several attempts have been performed to provide a more precise definition of timing of lesions, based on the search for haemorrhagic signs (Cattaneo et al. 2010) and interac-

Figure 65.12  Example of peri-­mortem sharp force trauma on scapular spina. Source: Modified from Cappella et al. (2014).

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Figure 65.11  Example of peri-­mortem blunt trauma on the distal end of the left tibia. Source: Modified from Cappella et al. (2014).

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Figure 65.13  Example of peri-­mortem gunshot force trauma on left temporal bone (a) and details of the lesions on the intra-­cranial surface (b). Source: Modified from Cappella et al. (2014).

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A pilot study. American Journal of Forensic Medicine Pathology 31 (1): 22–26. Cattaneo, C., Cantatore, A., Ciaffi, R. et al. (2012). Personal identification by the comparison of facial profiles: Testing the reliability of a high-­resolution 3D-­2D comparison model. Journal of Forensic Science 57 (1): 182–187. Cattaneo, C., Tidball Binz, M., Penados, L. et al. (2015). The forgotten tragedy of unidentified in the Mediterranean. Forensic Science International 250: e1–e2. Criminisi, A., Zissermann, A. and Van Gool, L. (1998). A new approach to obtain height measurement from video. In: Proceedings of the SPIE, Boston, Massachusetts, USA, Vol. 3576, 1–6 November. San Jose, CA. Daubert v. Merrell Dow Pharmaceuticals, Supreme Court of the United States, 113 S. Ct. 2786 (1993). De Angelis, D., Sala, R., Cantatore, A. et  al. (2009). A new computer-­ assisted technique to aid personal identification. International Journal of Legal Medicine 123: 351–356. De Angelis, D., Sala, R., Cantatore, A. et  al. (2007). New method for height estimation of subjects represented in photograms taken from video surveillance systems. International Journal of Legal Medicine 121: 489–492. De Boer, H.H., Obertová, Z., Cunha, E. et al. (2020) Strengthening the role of forensic anthropology in personal identification: Position statement by the Board of the Forensic Anthropology Society of Europe (FASE). Forensic Science International [Epub ahead of print]. Decker, S.J. and Ford, J.M. (2019). Forensic personal identification utilizing part-­ to-­ part comparison of CT-­ derived 3D lumbar models. Forensic Science International 294: 21–26. Edelman, G. and Alberink, I. (2009). Comparison of body height estimation using bipeds or cylinders. Forensic Science International 188 (1–3): 64–67. Fischman, S.L. (1985) The use of medical and dental radiographs in identification. International Dental Journal 35: 301–306. Gibelli, D., Cellina, M., Cappella, A. et al. (2019). An innovative 3D–3D superimposition for assessing anatomical uniqueness of frontal sinuses through segmentation on CT scans. International Journal of Legal Medicine 133 (4): 1159–1165. Gibelli, D., Mazzarelli, D., Porta, D. et al. (2012). Detection of metal residues on bone using SEM-­EDS – Part II: Sharp force injury. Forensic Science International 223: 91–96. Gibelli, D., Obertovà, Z., Ritz-­Timme, S. et al. (2016). The identification of living persons on images: A literature review. Legal Medicine 19: 52–60. Goos, M.I.M., Alberink, I.B. and Ruifrok, A.C.C. (2006). 2D/3D image (facial) comparison using camera matching. Forensic Science International 163: 10–17. Grivas, C.R. and Komar, D.A. (2008). Kumho, Daubert, and the nature of scientific inquiry: Implications for forensic anthropology. Journal of Forensic Sciences 53 (4): 771–776. Izumi, T. and Uerki, Y. (2004). Improvement of precision by introducing vitality degree to the height measurement system using stereo vision. In: Proceeding of the SICE Annual Conference, Sapporo, August 4–6, 2004. Kleinberg, K.F., Vanezis, P. and Burton, A.M. (2007). Failure of anthropometry as a facial identification technique using high-­quality photographs. Journal of Forensic Science 52 (4): 779–783.

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Al-­Amad, S., McCullough, M., Graham, J. et  al. (2006). Craniofacial identification by computer-­ mediated superimposition. Journal of Forensic Odontostomatology 24 (2): 47–52. Alberink, I. and Bolck, A. (2008). Obtaining confidence intervals and likelihood ratios for body height estimations in images. Forensic Science International 177 (2/3): 228–237. Alunni-­Perret, V., Muller-­Bolla, M., Laugier, J.P. et al. (2005). Scanning electron microscopic analysis of experimental bone hacking trauma. Journal of Forensic Science 50 (4): 796–801. Berryman, H. and Haun, S. (1996). Applying forensic technique to interpret cranial fracture patterns in an archaeological specimen. International Journal of Osteoarchaeology 6 (1): 2–9. Biehler-­Gomez, L. and Cattaneo, C. (2020). Interpreting Bone Lesions and Pathology for Forensic Practice. London, UK: Academic Press. Blau, S. and Ubelaker, D.H. (2016). Handbook of Forensic Anthropology and Archaeology. Walnut Creek, California: Routledge. Birngruber, C.G., Kreutz, K., Ramsthaler, F. et al. (2010). Superimposition technique for skull identification with Afloat® software. International Journal of Legal Medicine 124 (5): 471–475. Cameriere, R., Scendoni, R., Lin, Z. et  al. (2020). Analysis of frontal sinuses for personal identification in a Chinese sample using a new code number. Journal of Forensic Sciences 65 (1): 46–51 Caplova, Z., Obertova, Z., Gibelli, D. et al. (2018). Personal identification of deceased persons: An overview of the current methods based on physical appearance. Journal of Forensic Sciences 63 (3): 662–671. Cappella, A., Castoldi, E., Sforza, C. and Cattaneo, C. (2014). An osteological revisitation of autopsies: Comparing anthropological findings on exhumed skeletons to their respective autopsy reports in seven cases. Forensic Science International 244: 315.e1–e10. Cappella, A., Gibelli, D., Cellina, M. et  al. (2019). Three-­dimensional analysis of sphenoid sinus uniqueness for assessing personal identification: A novel method based on 3D–3D superimposition. International Journal of Legal Medicine 133 (6): 1895–1901. Cattaneo, C., Andreola, S., Marinelli, E. et  al. (2010) The detection of microscopic markers of hemorrhaging and wound age on dry bone:

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distinguishing between a saw or a knife (Reichs 1998; Symes 2005; Symes et al. 2010; Thompson and Inglis 2009). Furthermore, the reconstruction of the weapon involved may come from the use of scanning electron microscopy (SEM) that provides additional data about morphology and size of bone lesions (Alunni-­Perret et al. 2005) and the search for residues that may have been eventually left by the weapon through energy-­ dispersive X-­ray spectrometry (EDS) technology; however, for the latter point, caution must be taken concerning possible environmental contamination (Gibelli et  al.  2012; Pechnikova et al. 2012). Finally, from data collected on the type of lesion and the weapon used, hypotheses about the cause of death may be elaborated according to the position of the lesions, the reconstruction of organs eventually involved and the possible consequences.

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Symes, S.A. (2005). Morphology of saw marks in human bone: Introduction and examination of residual kerf contour. In: K.J. Reichs (ed.), Forensic Osteology, Advances in the Identification of human Remains. Springfield, USA: Charles C Thomas. Symes, S.A., Chapman, E.N., Rainwater, C.W. et al. (2010). Knife and saw toolmark analysis in bone: A manual designed for the examination of criminal mutilation and dismemberment. research report submitted to the U.S. Department of Justice. Stephan, C.N., Winburn, A.P., Christensen, A.F. and Tyrrell, A.J. (2011). Skeletal identification by radiographic comparison: Blind tests of a morphoscopic method using ante mortem chest radiographs. Journal of Forensic Sciences 56 (2): 320–332. Tanner, J.M., Whitehouse, R.H. and Cameron, N. (1983). Assessment of Skeletal Maturity and Prediction of Adult Height (TW2  Method). London: Academic Press. Thompson, T.J. and Inglis, J. (2009). Differentiation of serrated and non-­ serrated blades from stab marks in bone. International Journal of Legal Medicine 123 (2): 129–135. Van den Hout, A. and Alberink, I. (2010) A hierarchical model for body height estimation in images. Forensic Science International 197 (1–3): 48–53. Vanezis, P., Lu, D., Cockburn, J. et al. (1996). Morphological classification of facial features in adult Caucasian males based on an assessment of photographs of 50 subjects. Journal of Forensic Science 41 (5): 786–791. van Mastrigt, N.M., Celie, K., Mieremet, A.L. et al. (2018). Critical review of the use and scientific basis of forensic gait analysis. Forensic Sciences Research 3 (3): 183–193. Wang, L., Tan, T., Ning, H. and Hu, W. (2003). Silhouette analysis-­based gait recognition for human identification. IEEE Transactions on Pattern Analysis and Machine Intelligence 25 (12): 1505–1518. Yoshino, M., Imaizumi, K., Miyasaka, S. and Seta, S. (1995). Evaluation of anatomical consistency in craniofacial superimposition images. Forensic Science International 74 (1/2): 125–134. Yoshino, M., Miyasaka, S., Sato, H. and Seta, S. (1987). Classification system of frontal sinus patterns by radiography. Its application to identification of unknown skeletal remains. Forensic Science International 34 (4): 289–299. Yoshino, M., Taniguchi, M., Imaizumi, K. et al. (2005). A new retrieval system for a database of 3D facial images. Forensic Science International 148 (2/3): 113–120. Yushkevich, P.A., Piven, J., Hazlett, H.C. et al. (2006). User-­guided 3D active contour segmentation of anatomical structures: significantly improved efficiency and reliability. Neuroimage 31 (3): 1116–1128.

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Komar, D. (2003). Lessons from Srebrenica: The contributions and limitations of physical anthropology in identifying victims of war crimes. Journal of Forensic Sciences 48 (4): 713–716. Komar, D. and Lathrop, S. (2006). Frequencies of morphological characteristics in two contemporary forensic collections: Implications for identification. Journal of Forensic Sciences 51 (5): 974–978. Koot, M.G., Sauer, N.J. and Fenton, T.W. (2005). Radiographic human identification using bones of the hand: A validation study. Journal of Forensic Sciences 50 (2): 263–268. Lee, J., Lee, E.D., Tark, H.O. et al. (2008). Efficient height measurement method of surveillance camera image. Forensic Science International 177 (1): 17–23. Lynnerup, N. and Vedel, J. (2005). Person identification by gait analysis and photogrammetry. Journal of Forensic Science 50 (1): 112–118. Mundorff, A.Z., Vidoli, G. and Melinek, J. (2006). Anthropological and radiographic comparison of vertebrae for identification of decomposed human remains. Journal of Forensic Sciences 51: 1002–1004. Ohlrogge, S., Arent, T., Huckenbeck, W. et al. (2009). Anthropological Atlas of Female Facial Features. Frankfurt: Verlag für Polizeiwissenschaft. Ohlrogge, S., Nohrden, D., Schmitt, R. et al. (2008). Anthropological Atlas of Male Facial Features. Frankfurt: Verlag für Polizeiwissenschaft. Pechnikova, M., Porta, D. and Cattaneo, C. (2011). Distinguishing between perimortem and post-­mortem fractures: are osteons of any help. International Journal of Legal Medicine 125 (4): 591–595. Pechnikova, M., Porta, D., Mazzarelli, D. et al. (2012). Detection of metal residues on bone using SEM-­EDS. Part I: Blunt force injury. Forensic Science International 223 (1–3): 87–90. Puppe, G. (1903). Traumatische todesursachen. In Gerichtliche Medizin. Jena: Nabu Press. Rich, J., Tatarek, N.E., Powers, R.H. et al. (2002). Using pre-­and post-­ surgical foot and ankle radiographs for identification. Journal of Forensic Sciences 47: 1319–1322. Ritz-­Timme, S., Gabriel, P., Obertovà, Z. et al. (2011). A new atlas for the evaluation of facial features: Advantages, limits and applicability. International Journal of Legal Medicine 125 (2): 301–306. Rosenbloom, A.L. and Tanner, J.M. (1998). Misuse of Tanner puberty stages to estimate chronologic age. Pediatrics 102 (6): 1494. Schmeling, A., Olze, A., Reisinger, W. and Geserick, G. (2001). Age estimation of living people undergoing criminal proceedings. Lancet 358: 89. Schmeling, A., Reisinger, W., Geserick, G. and Olze, A. (2006). Age estimation of unaccompanied minors. Part I. General considerations. Forensic Science International 159 (Suppl): S61–64. Schmeling, A., Reisinger, W., Loreck, D. et al. (2000) Effects of ethnicity on skeletal maturation: Consequences for forensic age estimations. International Journal of Legal Medicine 113: 253.

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different materials, preparation methods, and endodontic, orthodontic and surgical treatments may have been used (Rötzscher and Reimann 1975). Depending on the quantity and quality of the findings, the degree of variation and the reliability of the comparison are increasing. Different calculation models show the possibility of more than 2.5 billion variations (Fiala 1968; Sopher  1986). As well as, the loss of teeth or dental treatments and other influences such as nutrition or trauma are able to cause abrasions and change the configuration of an individual’s teeth. This means that after several years the teeth of a human are highly individualised, comparable to a fingerprint, and a dental chart can be used for forensic purposes. Another important part of forensic odontology is the legal aspect involved in treating patients. Every dentist needs knowledge in this area and the relationship between dentist and patient includes ethical and deontological issues. There will be no further discussion of this here since laws and regulations are different in each country.

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Forensic odontology is an interdisciplinary field between forensic medicine and dentistry, with several forensic aspects playing a role in both fields. This establishes a close connection between forensic pathology and forensic odontology. Apart from forensic odontology, the international literature also uses different terms such as forensic dentistry or forensic odontostomatology. One of the most important applications of forensic odontology is the identification of unknown persons. Dental charts for adults are highly individualised, making the comparison of ante-­mortem and post-­mortem data one of the primary identification methods for either mass disaster victim identification or individual cases of unknown human remains. Another application of this variability is the comparison of bite marks left in violent crimes with specific suspects. The reliability of bite mark comparisons varies based on the evidence substrate and other factors, and cases of incorrect associations have been documented, meaning this application requires highly specialised expertise (Bowers  2006; Bowers and Pretty 2009). Wherever possible, DNA testing of the bite mark saliva has replaced this analysis. The high level of differentiation of a human dental chart is based on several aspects. Every adult normally has 32 teeth, every young child has 20 milk teeth, with or without anomalies, misalignments and other findings, and every tooth has five surfaces. From a mathematical point of view, this situation allows for 3.8 × 104 different restorations (Hausmann et  al.  1997). Any type of dental treatment multiplies the individualisation criteria since

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Handbook of Forensic Medicine, Volume 3, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

66.2  Dental charting After the odontological examination, the characteristics of an individual’s teeth, as well as specific features such as bridges or other dental work, are described in a dental chart or dental record. High-­ quality documentation is a prerequisite for ­reliable identification, and the more the findings are documented, the higher the achievable identification index. The

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structure and the individual treatment (in this case, root fillings) were sufficient for the identification (Figure 66.3). Ante-­mortem data collection is the other important part of the identification process. Depending on local legislation, the extent and retention of dental records will differ. If a patient was an emergency case and visited the dental practice only once, documentation may be limited. Also, in most cases, the dentist will document only missing or diseased teeth for a new patient and omit charting previous fillings and other treatments, making these data sets incomplete. This means a lot of information may be missed. A critical appreciation of the possible pitfalls of dental records is very important. It is well known that ante-­ mortem data sets from a dental practice may include errors, and a review of ante-­mortem data after the 2004 South Asian tsunami revealed that of 106  data sets, 51% had to be returned because they were either incomplete or of unacceptable standard (Kieser et  al.  2006). One of the most common mistakes is

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investigation of an unknown body’s teeth depends on the condition of the body, such as the amount of destruction or decomposition, and the findings are recorded in an electronic or paper-­based format. An additional tool for getting a full dental data set can be a radiological examination. Currently, the X-­ray technique is a standard method and gets reliable results. Computed tomography (CT) or magnetic resonance imaging (MRI) scans are also options, but the quality of the pictures still raises some issues as shown in Figures  66.1 and 66.2, although several techniques to get more and better information using CT have been reported (Thali et al. 2006). Modern CT or MRI examinations are more expensive and in order to compare data with digital X-­rays, specific software is required. Depending on the quantity of ante-­mortem available, a radiological investigation can be very helpful. In one example of an identification, X-­ray data from a single tooth led to a positive result. The highly individual anatomical

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Figure 66.1  Computer tomography scan of the upper and lower jaw. Source: Courtesy of Dr L. Oesterhelweg, Institute of Legal Medicine, Charite Berlin.

Figure 66.2  Orthopantomogram (X-­ray) of the upper and lower jaw. Source: Courtesy of Dr L. Oesterhelweg, Institute of Legal Medicine, Charite Berlin.

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until the 18th year of age; during puberty, there is an acceleration in dentition changes. The second period starts after the completion of dentition when degenerative changes start taking place. Within the dentition period, the following four stages have been defined: 1. Development of the crown. 2. Development of the root. 3. Development of the enamel. 4. Eruption. Dentition is completed after the occlusal surface of the tooth reaches the height of the neighbouring teeth and the growth of the root has ended. There are three methods for forensic age estimation – morphological, biochemical and radiological. Using these methods, it is important to know that the chronological age of a person can differ from the biological. One frequently used scheme was established by Schour and Massler (1958) with variations by Ferembach et  al. (1979) (Figure  66.4). Ciaparelli (1985) determined that the data are reliable for male children, but that there is a difference for females, who may be up to 6 months younger than the estimate. The older a child is, the less accurate the estimation is likely to be. One of the most important studies in the field of age estimation was the Gustafson method that used regressive changes such as secondary dentin formation, periodontal recession, attrition, apical translucency, cementum apposition and external root resorption  – all typical age-­ related phenomena. Compared with other studies that investigated only one age-­ related trait, the advantage of this method was that different changes of the tooth structure were combined. The Gustafson method was the first to use a statistical approach and required thin slices of the tooth to be available for microscopic investigation. Four stages of changes were established in this study by assigning a value between 0 and 3 for every phenomenon (Figure 66.5). The sum of all the values showed the correlation to the age to be r = 0.91 (with correction) (Figure 66.6) using the formula Age = 11.43 + 4.56x (where x is the sum of the points of the phenomena). The reliability of the estimation could be increased if more than one tooth was used. In such cases, the final result was established by using the mean value of the intermediate results (Gustafson 1947, 1950, 1955, 1966). Recently, research has focused on finding additional anatomical tooth structures that can be utilised for determining the age of a person. The pulp cavity, for example, is reduced as a result of secondary dentine deposits, and measurements of this reduction can be used as an indicator of age. Statistical analyses have shown a correlation between age and the different ratios of the pulp cavity (Kvaal et al. 1995; Paewinsky et al. 2005; Meinl et al. 2007). Other studies have shown the potential for age estimation using anatomical dental structures. Solheim (1988a) reported on the colour of teeth, which change with age and differ between males and females. Another aspect is the attrition of teeth; the premolars show the highest correlation with age with a coefficient of r = 0.68. In addition, attrition is much more

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Figure 66.3  (a) Ante-­mortem X-­ray of a missing person submitted from a dental practice showing fillings, root filling and apicoectomy. (b) Post-­mortem X-­ray taken from an unknown body showing the identical situation as shown in panel (a).

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attributing a description to the wrong side. This makes it very important to not only rely on the chart but also request the radiographs, casts and other documents from the dental practice. Ante-­mortem orthopantomograms may be of great value in these cases (Du Chesne et  al.  2000). Only by looking at the X-­rays or photographs, the forensic odontologist will be able to clarify apparent discrepancies and avoid false exclusions of a putative identification. Casework experience has shown that, in order to avoid errors, it is necessary to provide special training to the personal transferring the data from a patient record in a dental practice to an ante-­mortem form.

66.3  Age estimation One of the additional aspects of forensic dentistry is the use of a progressive dentition timeline to estimate the age of a person. Dentition is genetically determined and different age-­related periods have been established. The first period during childhood includes the dentition of the milk and permanent teeth

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Figure 66.4  (a, b) Development of the dentition over a person’s lifetime. Sources: Data from Schour and Massler (1958) and Ferembach et al. (1979), © American Dental Association, Chicago.

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methods are too complicated. The Gustafson method, in ­particular, requires a lengthy tooth preparation and microscopic examination. This can be applied to individual identification cases but is not feasible in instances with a high number of unknown bodies. In cases of mass disasters, these methods are too time-­consuming and normally not used. Another point of criticism is a strong subjective component relying on the experience of the investigator  – not only the accuracy of the measurements but also the skills of the investigator may have an influence on the results. An age estimation method using biochemical changes examines the degree of asparagine acid racemisation in dentin. Ogino et al. (1985) reported this method for the first time and obtained promising results, with a high correlation to the real age of up to r = 0.99. Ritz et  al. (1993) also investigated root dentin and showed a close relationship between the extent of aspartic acid racemisation and age. This is important in those cases where a large portion of the coronal dentin is absent, for instance following dental treatment. The method is only used in special cases since it is fairly expensive and requires special equipment.

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Figure 66.5  Classification of changes of the teeth following Gustafson (1947, 1950, 1955, 1966). A, attrition; C, apposition of cementum; P, changes of the periodontium; R, resorption; S, secondary dentine; T, transparency. Sources: Adapted from Gustafson (1947, 1950, 1955, 1966).

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prevalent in men (Solheim 1988b). Measurements of the non-­ uniformity of the root and the surface of the root show a strong correlation to the age: r = 0.44 and r = 0.68, respectively (Solheim and Kvaal 1993). Some of the reported methods have been criticised on several points. One of the most frequent criticisms is that the

The identification of an unknown person can be based on different primary and secondary methods. Being a primary method, odontological identification is one of the most important and reliable tools in the field. As described before, individual dental records form highly variable data sets that can be compared with each other. Some scenarios, for example existing teeth that are reported as missing in the ante-­mortem documentation, enable the exclusion of a possible identity. It is

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Figure 66.6  (a) The regression line for age estimation and (b) the relation between point values and the age of 41 individuals. Sources: Adapted from Gustafson (1947, 1950, 1955, 1966).

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recommended that a combination of several findings is used for a reliable exclusion comparable with other primary methods of identification. If the dental chart comparison indicates a positive association and thus a possible identity, the certainty depends on the overall complexity of the findings. Associating two charts with a treatment with combined dentures can give highly reliable ­information, but there is no source of statistical information about the distribution of different dental treatments in different populations and age groups. This means it is not possible to statistically evaluate the significance of a match as is done for cases identified using DNA-­based methods. Efforts in establishing databases capturing dental chart data and frequency of tooth characteristics have not yet been successful. Treatment choices and distributions change frequently, the extent of treatments is age-­dependent, the coding varies, and more complex modifications cannot be captured easily. The only epidemiological information available is the decayed missing filled teeth (DMFT) index established to track caries prevention (Korts et al. 1978), but this index is of limited value for forensic dentists. There are still no data on, for example, how frequent implants in the molar region of the right side of the upper jaw are in a population. Since there is no established statistical calculation to support the strength of a dental identification, the final conclusion may depend on the experience and subjective opinion of an investigator. This means training and extensive quality control, including a review of all findings, are necessary. The following case examples illustrate some problems encountered in fieldwork.

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Figure 66.7  Lower jaw with combination of treatments in an unidentified body.

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Case example I The first case represents a false exclusion. An unknown body was examined and the following dental data set was established: upper jaw toothless and full denture; lower jaw showing a combination of treatments with a block of crowns in the front and a partial denture coloured clamps (Figure  66.7). Fingerprints from the with gold-­ body had been compared with the fingerprints from the home of the possible victim and had led to a positive identification but a review of the dental records showed several exclusions. The ante-­ mortem record reported a single tooth to be present in the upper jaw and missing teeth in the front of the lower jaw. The original documentation of the dental practice clarified the discrepancy for the upper jaw. The transfer of the data had been incorrect: the dentist had documented the extraction of the tooth using only an abbreviation and thus the extracted tooth was not registered as missing. For the lower jaw, the medical record reported missing teeth in the front and a bridge. This dental treatment had not been performed by the dental practice providing the ante-­mortem data; therefore, their documentation of the lower jaw was incorrect. The block of crowns in the front had been misinterpreted as a bridge (Figure  66.8). The described discrepancies would normally have been exclusion criteria and would have prohibited an identification based on dental charts. This illustrates that reliable ante-­mortem data are as critical for the identification process as error-­free post-­ mortem data (Kieser et al. 2006). Source: Based on Kieser et al. (2006).

Figure 66.8  Cast from dental practice providing the ante-­mortem data for a missing person.

66.5  Bite marks The forensic odontostomatological investigation of bite marks is not common in forensic science casework. The state of the dentition, the degree of break down and/or repair of the teeth may create a bite mark with a high level of individuality. In some cases, this may allow identification of the biter (Ligthelm and van Niekerk 1994; Saglam Atsü et al. 1998; Lessig and Benthaus 2003; Lessig et al. 2006). Most bite marks are obtained from cases of sexual violence. Some may be defensive bites placed on the attacker by the victim, and, though rarely seen, some bite marks may be self-­inflicted.

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mented in the dental chart provided by the dental practice (Figure 66.9), the existing crowns and missing teeth gave sufficient information for a reliable identification.

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Figure 66.9  Ante-­mortem dental record.

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Case example II The second case example shows a positive identification with typical ante-­mortem information. Even though not all findings were docu-

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Generally, bite marks consist of superficial abrasions and/or subsurface haemorrhage, or bruising of the skin because of the bite (Endris 1979). Though the mechanism is not clearly understood, the pattern of the injury is affected by the force and length of time of the bite, in combination with other mechanical and physiological factors. Barbenel and Evans (1977) have discussed the influence of the skin lineages. Bite marks may be found in cases of sexual violence in typical areas of the human body – the genitals and breasts – but also in cases of child abuse. In such cases, the number of bites can be very high; Trube-­Becker (1973) reported a case with 17 bite marks. Occasionally, bite marks are obtained in various types of food such as chocolate, chewing gum, fruits, vegetables and similar (Endris  1979; Saglam Atsü et  al.  1998; McKenna et  al.  2000) although solid food is more likely. Aboshi et al. (1994) reported the identification of a suspect arsonist by means of bite marks in cakes that were found at the scene of the crime. A missing upper right central incisor was proved to be in the patterned injury. Bernitz et al. (2000) reported a case of murder where there was a bite mark in a piece of cheese. The pattern-­associated comparison between the impression and a study model of the suspect was able to identify the perpetrator. In this case, fingerprints and DNA evidence were not found at the crime scene, and the court was reluctant to accept the validity of the investigation method. The

forensic odontostomatologist involved in the court case proved the validity of the method by producing several bite marks in cheese, butter and cooked potato; pairwise comparisons were made by two odontologists. These examiners correctly identified all the true matches as well as selecting the dental models for which there were no corresponding impressions. The characteristics of human bites are superficial abrasions and/or subsurface haemorrhage looking like an arch. These are caused by the incisors, canine and premolars. The abrasions and/ or haemorrhage caused by the canines are in the shape of points. If the perpetrator has had major dental work, additional specific marks can be expected, which differ between bridges, crowns and dentures. Crowns and bridges have a ceramic delusion while dentures are characterised by braces. These peculiarities can be responsible for specific wounds and additional markers for identification. Depending on the part of the body and the constitution of the skin, the bite mark can be distorted. Frequently, this can be the reason for problems when analysing bite marks. To prevent mistakes in pattern-­associated comparison, it is recommended that bites should be simulated at similar body parts using the study casts of the suspect (Lessig 2001; Lessig and Benthaus 2003) or using digital techniques for a stepwise dynamic comparison (Sakoda et al. 2000; Thali et al. 2003). Sheasby and MacDonald

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late, and experts are usually not involved in recording the bite marks. Thus, the record of patterned injuries is based on recommendations for DNA analysis and may ignore guidelines for other types of investigations and the recommendations for photographic evidence. If DNA identification fails, forensic odontostomatological investigations should be considered in all cases of bite injuries. The presented cases suggest that almost all investigations can be carried out successfully. To standardise the analysis of bite marks, the American Board of Forensic Odontostomatology (ABFO) established guidelines in 1986. These guidelines are important for the investigation and should be carried out in all routine casework: 1. History: Obtain a thorough history of any dental treatment carried out after the suspected date of the bite mark. 2. Photography: Extra-­oral photographs should be taken, including full face and profile views; intra-­orals photographs should include frontal views, two lateral views and an occlusal view of each arch. Often it is useful to include a photograph of maximal mouth opening. If inanimate materials, such as foodstuffs, are used for test bites, the results should be preserved photographically. 3. Extra-­oral examination: Record and observe soft tissue and hard tissue factors that may influence biting dynamics. Measurements of maximal opening and any deviations on opening or closing should be made using a special scale rule (Figure 66.11). 4. Intra-­oral examination: Salivary swabs should be taken. The tongue should be examined to assess size and function. The periodontal status should be noted with particular reference to mobility. Prepare a dental chart if possible.

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(2001) recommended a classification to emphasise the need of a scientific approach for the interpretation of the types of distortion. They introduced the terms primary and secondary distortions. Primary distortion is defined by the dynamics of the bite. Secondary distortion can be divided into three categories: (i) time-­ related distortion when a bite changes with the time elapsed subsequent to the bite being made, (ii) posture distortion and (iii) photographic distortion. It is also important to differentiate human and animal bites as well as to identify the kind of human bite. Human bites can be classified into defence and suction types. To record a bite mark, an exact photographic documentation and one-­to-­one transfer to onto a transparent sheet are indispensable (Figure 66.10). Swabbing of the bite injury is important to recover trace evidence. Stains of saliva or human cells for a DNA analysis should be collected whenever possible (Wright and Dailey 2001; Lessig and Benthaus 2003; Lessig et al. 2006). Bite marks are often difficult to notice and may be overlooked. Sometimes it is problematic to detect and record these patterns. The uniqueness of the human dentition and analytical techniques usually allow an exact identification of the perpetrator (Pretty and Sweet  2001b), although the quality of the bite mark is an important factor. New approaches with digital techniques overlaying the bite mark and patterns of a suspect facilitate an exact investigation. Bite mark and study casts can be compared using three-­dimensional images (Thali et al. 2003; Blackwell et al. 2007). It is important to have a good-­quality offender trace recorded by specialists only (Pretty and Sweet 2001c). Unfortunately, due to the strategy of getting DNA evidence first, opportunities for forensic odontostomatological investigations are often used very

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Figure 66.10  Copy of bite mark from victim transferred to an acetate sheet.

Figure 66.11  Scale recommended by the American Board of Forensic Odontostomatology.

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s­ uccessfully matched to the case patterns. These cases show that it is worth trying bite mark analysis even in cases of bad documentation (Bowers 2006; Bowers and Pretty 2009).

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that meets the American Dental Association specifications. The occlusal relationship should be recorded. 6. Sample bites: Whenever possible, sample bites should be made into an appropriate material, simulating the type of bite under study (Figure 66.12). 7. Study casts: Casts should be prepared using type II stone. Additional casts should be made by duplicating the master casts. Pretty and Sweet (2001a) sent out a questionnaire to 69 American odontologists examining bite marks. Their purpose was to examine adherence to the guidelines. Twenty-­eight (41%) of the odontologists were members of the American Society of Forensic Odontology (ASFO). The study found that methods differed between examiners although, in general, the odontologists adhered to the guidelines. The materials employed by the odontologists were acceptable and defendable in court. Forensic dentists who neglected the guidelines could face harsh criticism when testifying in court (Pretty and Sweet  2001b). Pretty and Turnbull (2001) reported a case showing a lack of dental uniqueness between two suspects. The dental arrangement of the suspects was similar, and it was impossible to determine the biter positively. Thus, the central dogma that every bite mark is unique was disproved. In another study, the effectiveness of digital bite mark overlays was verified. Different examiners were asked to compare scanned photos of bite marks with impressions of suspects. They found that although the experience of the examiners had an influence on the results, the method overall had a high level of reliability (Pretty and Sweet 2001b). Using patterns of the study casts of a suspect and fixation of the bite registration was also found to be the basis for successful analysis (Endris 1979) (Figure 66.13). Impressions of the suspect made in material similar to that involved in the case were

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Figure 66.13  Bite mark with overlaid acetate sheet of the mark from the study casts of the putative perpetrator. The numbers represent the specific teeth that could be identified as second incisor of the right upper jaw (12), second incisor of the left upper jaw (22), second incisor (32) and eye tooth of the left lower jaw (33), and second incisor (42) and eye tooth of the right lower jaw (43).

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References and further reading ABFO (American Board of Forensic Odontology) (1986). Inc: Guidelines for bite mark analysis. Journal of the American Dental Association 112: 383–386. Aboshi, H., Taylor, J.A., Takei, T. and Brown, K.A. (1994). Comparison of bitemarks in foodstuffs by computer imaging: a case report. Journal of Forensic Odontostomatology 12: 41–44. Barbenel, J.C. and Evans, J.H. (1977). Bite marks in skin – mechanical factors. International Journal of Forensic Dentistry 4: 6. Bernitz, H., Piper, S.E., Solheim, T. et al. (2000). Comparison of bitemarks left in foodstuffs with models of the suspects dentitions as a means of identifying a perpetrator. Journal of Forensic Odontostomatology 18: 27–31. Blackwell, S.A., Taylor, R.V., Gordon, I. et al. (2007). 3-­D imaging and quantitative comparison of human dentitions and simulated bite marks. International Journal of Legal Medicine 121: 9–17. Bowers, C.M. (2006). Problem based analysis of bitemark misidentifications: The role of DNA. Forensic Science International 195 (Suppl): S104–109. Bowers, C.M. and Pretty, I.A. (2009). Expert disagreement in bitemark casework. Journal of Forensic Science 54: 915–8. Ciaparelli, L. (1985). An Assessment of Dental Age in Essex Schoolchildren Using Panoral Radiographs with Forensic Applications. Dissertation Diploma in Forensic Odontology, London Hospital Medical College.

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c­ ontribution to forensic age estimation methods in adults. International Journal of Legal Medicine 119: 27–30. Pretty, I.A. and Sweet, D. (2001a). Adherence of forensic odontologists to the ABFO bite mark guidelines for suspect evidence collection. Journal of Forensic Science 46: 1152–1158. Pretty, I.A. and Sweet, D. (2001b). Digital bite mark overlays – an analysis of effectiveness. Journal of Forensic Science 46: 1385–1391. Pretty, I.A. and Sweet, D. (2001c). The scientific basis for human bite mark analyses – a critical review. Science and Justice 41: 85–92. Pretty, I.A. and Turnbull, M.D. (2001). Lack of dental uniqueness between two bite mark suspects. Journal of Forensic Science 46: 1487–1491. Ritz, S., Schütz, H.W. and Peper, C. (1993). Post mortem estimation of age at death based on aspartic acid racemization in dentin: its applicability for root dentin. International Journal of Legal Medicine 105:­ 289–293. Rötzscher, K. and Reimann, W. (1975). Die forensische Stomatologie. In: O. Prokop and W. Göhler (eds.), Forensische Medizin, 3rd edn, pp. 545–564. Berlin: Volk und Gesundheit. Saglam Atsü, S., Gökdemir, K., Kedici, P.S. and Ikyaz, Y.Y. (1998). Bitemarks in forensic odontology. Journal of Forensic Odontostomatology 16: 30–34. Sakoda, S., Fujita, M.W., Zhu, B.L. et al. (2000). Wounding dynamics in distorted bitemarks: two case reports. Journal of Forensic Odontostomatology 18: 46–51. Schour, I. and Massler, M. (1958). Chronology of the Development of the Dentitions, 2nd edn. Chicago: American Dental Association. Sheasby, D.R. and MacDonald, D.G. (2001). A forensic classification of distortion in human bite marks. Forensic Science International 122: 75–78. Solheim, T. (1988a). Dental color as an indicator of age. Gerodontontics 4: 114–118. Solheim, T. (1988b). Dental attrition as an indicator of age. Gerodontontics 4: 299–304. Solheim, T. and Kvaal, S. (1993). Dental root surface structure as an indicator of age. Journal of Forensic Odontostomatology 11: 9–21. Sopher, I.M. (1986). Grundsätzliche Begriffe der zahnärztlichen Identifikation: Der prämortale Befund. Die Individualität des Gebisses. In: I.M. Sopher (ed.), Forensische Zahnmedizin, pp. 65–66. Berlin: Quintessenz. Thali, M.J., Braun, M., Markwalder, T.H. et  al. (2003). Bite mark documentation and analysis: The forensic 3D/CAD supported photogrammetry approach. Forensic Science International 135: 115–121. Thali, M.J., Markwalder, T., Jackowski, C. et al. (2006). Dental CT imaging as a screening tool for dental profiling: Advantages and limitations. Journal of Forensic Sciences 51: 113–119. Trube-­Becker, E. (1973). Bißspuren bei Kindesmißhandlungen. Beitrage zur Gerichtlichen Medizin 31: 115–123. Wright, F.D. and Dailey, J.C. (2001). Human bite marks in forensic dentistry. Dental Clinics of North America 45: 365–397.

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Du Chesne, A., Benthaus, S., Teige, K. and Brinkmann, B. (2000). Post-­mortem orthopantomography – an aid in screening for identification purposes. International Journal of Legal Medicine 113: 63–69. Endris, R. (1979). Praktische Forensische Odonto-­ Stomatologie. Heidelberg: Kriminalistik Verlag. Ferembach, D., Schwidetzky, I. and Stloukal, M. (1979). Empfehlungen für die Alters-­und Geschlechtsdiagnose am Skelett. Homo 30: 1–32. Fiala, B. (1968). Identifikace osob podle chrupu – Forensni stomatologie. Prague: Státní Zdravotnické Nakladelství. Gustafson, G. (1947). Åldersbestämniningar pa tändar. Odontologisk Tidskrift 55: 556–568. Gustafson, G. (1950). Age determination on teeth. Journal of the American Dental Association 41: 45–54. Gustafson, G. (1955). Altersbestimmung an Zähnen. Deutsche Zahnärztliche Zeitschrift 10: 1763–1768. Gustafson, G. (1966). Forensic Odontology. London: Staples Press. Hausmann, R., Liebler, M. and Schellmann, B. (1997). Zur Personenidentifikation mittels Zahnstatus. Rechtsmedizin 7: 86–89. Kieser, J.A., Laing, W. and Herbison, P. (2006). Lessons learned from large scale comparative dental analysis following the South Asian tsunami of 2004. Journal of Forensic Science 51: 109–112. Korts, D.C., Poulsen, S. and Kingman, A. (1978). An evaluation of Knutson’s formula for estimating age-­specific DMF teeth. Community Dentistry and Oral Epidemiology 6 (4): 191–194. Kvaal, S.I., Kolltveit, K.M., Thomsen, I.O. and Solheim, T. (1995). Age estimation of adults from dental radiographs. Forensic Science International 74: 175–185. Lessig, R. (2001). Die forensische Stomatologie als Teil der rechtsmedizinischen Praxis. In: M. Oehmichen and G. Geserick (eds.), Osteologische Altersschätzung und Identifikation, Research in Legal Medicine, Vol. 26, pp. 67–74. Lübeck: Schmidt-­Römhild. Lessig, R. and Benthaus, S. (2003). Forensische Odonto-­Stomatologie. Rechtsmedizin 13: 161–168. Lessig, R., Weber, M. and Wenzel, V. (2006). Forensic odontological analysis of different types of bite marks. EXCLI Journal: Experimental and Clinical Studies (International Online Journal for Advances in Science) 5: 93–102. Ligthelm, A.J. and van Niekerk, P.J. (1994). Comparative review of bitemark cases from Pretoria, South Africa. Journal of Forensic Odontostomatology 12: 23–29. McKenna, C.J., Haron, M.I., Brown, K.A. and Jones, A.J. (2000). Bitemarks in chocolate: A case report. Journal of Forensic Odontostomatology 18: 10–14. Meinl, A., Tangl, S., Pernicka, E. et al. (2007). On the applicability of secondary dentin formation to radiological age estimation in young adults. Journal of Forensic Science 52: 438–441. Ogino, T., Ogino, H. and Nagy, B. (1985). Application of aspartic acid racemization to forensic odontology: post mortem designation of age of death. Forensic Science International 29: 259–267. Paewinsky, E., Pfeiffer, H. and Brinkmann, B. (2005). Quantification of secondary dentine formation from orthopantomograms  – a

IDENTIFICATION

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67 | The Doctor and the Law Handbook of Forensic Medicine, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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Brigitte Tag

Each nation has different values and perceptions about how humans should be both physically and mentally treated and what kind of human research, medical therapy and interventions are permitted. Where uncertainty exists with respect to moral and ethical concepts, it should not be the aim of legislation to proclaim any one of the valid positions to be generally binding and to exclude the others – the understanding of what legislation is, and should be, is subject to different insights that are continually changing. Moreover, only when the legislature provides a common purpose, when it is regarded by society as legally justifiable from an ethical point of view, there is a chance of it being accepted by the community.

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Knowledge of human diseases and adequate treatments, how the human body is structured and how it grows very quickly are discussed in this chapter. The hope is that all the information generated by medical research concerning the human body and health may give new insights which will be greater than the sum of each individual piece of knowledge. Against this background, the potential of medical treatment, handling of the corpse and human research must be evaluated from a legal point of view (Tag 2012a; Tag and Baur 2016; Deutsch and Spickhoff 2014; Lenk et al. 2014). Both information already gleaned, and new knowledge in the future will lead to new contexts and insights of the doctor–patient relationship, informed consent, decision-­making, protection of highly vulnerable persons and sensitive data. This progress is made more complex by the fact that findings from research concerning the human body and a broad range of diseases are made internationally and nationally, and need to be analysed, developed and integrated into an overall context. This leads to a confluence of various ethical and legal concepts and entails both challenges and burdens. Alongside the new insights and significant synergy effects of linking the generated data and databanks, there is a certain potential for conflict which must not be disregarded. The following is an attempt to highlight a selection of these topics. National and international law is not independent of the medical, technical, physical and sociological principles of medical treatment and its associated ethical issues.

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67.1  Legal relevance of medical treatment

Handbook of Forensic Medicine, Volume 3, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

67.1.1  Persons involved The number of people directly and indirectly affected by medical treatment and clinical research is considerable. Medical treatment, be it surgery, pharmaceutical intervention or prevention, e.g. in the case of vaccines, includes first those who use it to treat patients and/ or to perform medical research. Also, directly affected are the patients who are to be examined and treated, and indirectly not only their close family and social environs, but also broader circles. Those who bear the costs of this treatment (e.g. health insurance and research) and cooperating partners from industry and business who develop and distribute the relevant equipment and drugs are also involved. As illustrated by the COVID-­19 pandemic, the absence of examination and/or treatment may also result in large societal and economic costs being indirectly borne by the whole society.

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Box 67.1  Examples of liabilities involving medical activities (corresponding to the national regulations). • • • • • •

Civil liability Responsibility under criminal law Responsibility under labour law Responsibility under public law Responsibility under professional regulations Responsibility under disciplinary law, occupational ban

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For example, under Article 123, paragraph 1 of the Swiss Criminal Code (SCC) or sections 223 and 230 of the German Criminal Code (GCC), anyone who wilfully causes damage to the physical integrity or health of another shall, on a complaint, be liable to prosecution. Normally, there are also provisions that anyone who negligently causes damage to the physical integrity or health of another shall, on a complaint, be liable to imprisonment or a fine (e.g. see SCC, Article 125, paragraph 1; GCC, sections 229 and 230). In contrast to that which constitutes an offence of bodily harm in Germany, the protection provided for by Swiss law also includes mental health.1 In this, it is immaterial whether psychological impairment has a physical cause or whether it results in health impairments, provided that it reaches a certain materiality threshold. Normally, criminal codes protect healthy persons as well as those who are ill.2 Physical and mental integrity must be understood in a relative sense, especially as a person can have a physical or mental handicap from birth or it may be acquired during his or her lifetime. In Germany and Switzerland, medical malpractice claims are mostly settled with liability insurers (Tag 2012a; Teubner 2012)3. In many cases, there are mediation services and the social health insurers will have given expert opinions. In Switzerland, there is a huge difference if the malpractice took place in a public hospital or in a private hospital. In the first case, the Swiss cantons are liable for the malpractice, and in the second the Swiss Code of Obligations4 governs the malpractice and the doctors are liable for their acts and omissions (Aebi-­Müller et al. 2016). In some European countries, case law decides on medical diagnostic examination and medical treatment using the benchmarks that apply with respect to every other form of bodily harm or violence.5 The patient nevertheless has the right by reason of

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Distinctions may be made between several important issues related to medical treatment and clinical research. On the one hand, there are medical check-­ups, diagnoses, treatments and post-­treatments, and, on the other hand, there is human research. It is crucial to consider the use of the data thus generated, medical confidentiality, data protection and the right of the person concerned to know, as well as the right not to know. Furthermore, a distinction must be made between examinations and treatment of living persons and examinations of corpses. The right of the person involved to know or not to know requires a separate examination of the rights of access and information of third parties. Given this broad spectrum, it is hardly surprising that, as in the field of the doctor and the law, there is little international consensus on its contents and boundaries. The scope of permissible treatment, impermissible intervention and malpractice is correspondingly large. The complexity of the human body and causes of diseases and the current standard of lex artis make it more difficult. The judicature is drawn in by the undertow of these developments. This is shown for example in the field of criminal law: whether performing medical treatment constitutes an offence of bodily harm caused wilfully or negligently depends on the national legal preconditions and requires a solid knowledge of medical care and its significance. In the field of new medical treatments and medication as well as the increasing use of artificial intelligence in healthcare, it is not easy for the legislature to anticipate the degree of regulation needed and to provide the appropriate legislation. The net is thus widely cast. Nevertheless, the hugely different issues can be reduced to one common denominator. The following shows that it is not easy for traditional legal norms to do justice to modern developments, and for a variety of reasons they sometimes do not have the ability to meet the need for a rapid and simple means of adaption. It should be remembered that the provisions of the legislature and those governing medicine are of a fragmentary and subsidiary nature; this is also true concerning jurisdiction. Criminal law, in particular, covering so-­called ethical minimums and one of the most potent means of intervention for the state, should not become overwhelming when guiding the doctor– patient relationship. Otherwise, there is a danger that medical progress, and thus society’s legitimate interest in an effective and appropriate medical system, would be disproportionately stunted.

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67.1.2  Criminal relevance of medical diagnosis procedures and medical treatments as bodily harm or assault In many states, the criminal relevance of medical diagnosis procedures and medical treatments is determined according to provisions of criminal law. If there are no special provisions for malpractice in the case of violating the physical integrity of a patient, then offences against life and limb, in particular those of bodily harm and violence, constitute the focus of criminal liability (Box 67.1).

Swiss Federal Court, decision 20 April 1977, BGE (Published decision of the Swiss Federal Court) 103 IV 65, 70; decision 19 June 2008, BGE 134  IV  189, 192; decision 27 August 2008, 6B_517/2008, E.  3.1; Trechsel and Geth (2021); Roth and Berkemeier (2019). 2 Swiss Federal Court, decision 20 April 1977, BGE 103 IV 65, 70; Trechsel and Geth (2021). 3 See also: https://www.anwalt.de/rechtstipps/verdacht-­auf-­behandlungs­ fehler-­was-­tun_159790.html (last accessed 31 January 2022). 4 Swiss Code of Obligations (SCO), Articles 394 et seq. (provisions relating to the Agency Contract). 5 Swiss Federal Court, decision 14 September 1973, BGE 99 IV 208, 209 et seq.; decision 3 December 1998, BGE 124 IV 258, 260 et seq. 1

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Table 67.1  Some typical sources of problems concerning medical activity. Problem

Examples

Medical malpractice

Wrong anamnesis, examination, diagnosis Failed or wrong diagnostic assessment (lab, X-­ray, ultrasonic testing, CT, MRI, PET, etc.) Wrong method of treatment Risk taking without sufficient experience or specialised knowledge Danger due to fatigue Therapeutic consultation error, insufficient information Mistake in patient, in the operation field e.g. (arm, leg and kidney) and in the surgical procedure (e.g. abortion instead of prenatal diagnosis) Faulty medication, overdosage, mix of the remedy and lack of bedside monitoring Breach of hygiene and safety Complications overlooked Obsolete techniques for surgery, wrong bedding Wrong employment of equipment and omission of corrective maintenance Defective implantation of medical devices, organs and tissue Malpractice concerning injections, infusions and transfusions Failed or delayed hospital admission Failed or delayed consultation of experts Missing attention toward children Unnecessary surgery

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his or her dignity and autonomy in decision-­making to release by way of consent the doctor’s scope from the narrow confines placed on it by criminal law. That means as a rule that it is necessary for the patient to give his or her informed consent to the medical treatment. In cases where the actual informed consent of the patient cannot be secured (for example as a result of the physical or mental disposition of the patient or in cases of emergency where any delay is dangerous) or it cannot be obtained in time or at all, supposed consent  – or in exceptional cases the existence of a conflict of duties or necessity – can be invoked as justification. These legal principles are relevant to medical diagnosis and treatment where there is suspected bodily harm.

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In most countries, the main duties of the doctor  – to make an appropriate diagnosis, to treat well and to inform the patient correctly  – can be outlined more precisely by looking at the wide range of their responsibilities (Figure 67.1 and Table 67.1).

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Failed, wrong, delayed or missing information Missing or invalid consent

Errors in cooperation

Faulty organisation Errors in coordination Errors in communication Negligence concerning risk taking Delegation error: wrong delegation and wrong triage Monitoring error concerning the vertical division of work: failure to warn, wrong information, errors arising in the transmission of messages, error of observation/control, failed proofing and supervising documentation

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Figure 67.1  The well-­known duties of doctors. Some elements are considered to be core duties, for example history taking, accurate diagnosis, treatment, giving information and establishing informed consent, whilst other elements are regarded as ancillary to the core duties, for example error-­free medication, the operation and repair of medical equipment.

CT, computed tomography; MRI, magnetic resonance imaging; PET, positron emission tomography. Source: Tag (2012a).

general fundamental principles of law. For medical examination and treatment to be performed legally, lex artis as the standard of care and informed consent must be given.

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As the person to be examined faces the consequences of the possibly serious diagnosis and risks of a treatment and, as a rule, he or she is the only one who knows his or her personal priorities and values, the patient has the final say on whether an examination and medical treatment should be performed. The corresponding legal basis can be found in several bodies of rules and regulations, for example in Article 8 of the Convention for the Protection of Human Rights and Fundamental Freedoms (ECHR), Article 10 of the Convention on Human Rights and Biomedicine and Article 3 of the Charter of Fundamental Rights of the European Union (Box 67.2). In many European countries, the national constitution guarantees the power to decide what is done with one’s body, a right accorded every person by virtue of their existence and in the interests of a self-­determined life. Article 10, paragraph 2 of the Swiss Constitution states: ‘Every person has the right to personal liberty and in particular to physical and mental integrity and to freedom of movement’. Article 1, section 1 in conjunction with Article 2, section 2 of the Basic Law for the Federal Republic of Germany states that: ‘Human dignity shall be inviolable. To respect and protect it shall be the duty of all state authority [. . .] Every person shall have the right to life and physical integrity. Freedom of the person shall be inviolable. These rights may be interfered with only pursuant to a law’. Depending on the respective national legal situation, a large number of different legal provisions regulate informed consent in a specific and detailed manner.8 The right of self-­ determination provides the individual a set of values and the

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Medical indication covers the circumstances that, in addition to patient autonomy, form the basis for medical action. It regularly influences the decision for or against using certain diagnostic or therapeutic procedures. For a procedure to be indicated, it is not sufficient for a possible condition to be diagnosed. Rather, the patient’s personal data such as age, seriousness and duration of condition, mental and psychological state, personal resilience, acceptance of the condition and personal circumstances must also be taken into account. Linking medical diagnosis and therapy to an indication is part of the ethical–legal norms of the medical profession. In general, a method of diagnosis can be justified in medical terms if the diagnosis is followed by a specific preventative or therapeutic treatment. Whether the method is an appropriate means of recognising sicknesses or pathological deviations from the normal functions of the body must be assessed according to the experience and findings of the researchers and doctors working in the relevant field. Core knowledge that provides reliable information on the necessity of performing the particular method for diagnostic and therapeutic purposes will act concurrently as a guideline for legal evaluation. This principle is enshrined in Article 4 of the Convention on Human Rights and Biomedicine:6

tive requirements of lex artis.7 A diagnosis method or medical treatment is no longer capable of satisfying the standard where updated methods recognised by experts exist that entail less risk and/or place less of a burden on the person being examined or treated. In this case, the method previously used no longer meets the needs of appropriate treatment. Here, we can say yesterday’s standard constitutes today’s error. If the outdated diagnosis method or medical treatment leads to bodily harm, the objective elements of a crime of simple bodily harm or those of an offence of violence may be fulfilled, depending on the national legal requirements. For example, bodily harm through negligence (SCC, Article 125; GCC, section 229) may be fulfilled where the doctor as a result of insufficient diligence assumed that the specific methods were not harmful.

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The standards thus outlined, which allow and restrict medical diagnoses and treatments, are not set in stone. Because of progress the lex artis is subject to constant change. Furthermore, there is a wide range of examinations and treatments, from those providing maximum medical care to those providing so-­called basic care. Although the general tying of diagnosis and treatment to an indication is part of the ethical–legal norms of the medical profession, its scope of effect is ambivalent from a legal point of view. Besides, not everything that in medical terms or in keeping with the practice of a careful doctor is appropriate, expedient, measured or necessary, reflects the expectations and wishes of the patient. Thus, it can be that a patient effectively refuses an examination or a treatment although it is medically indicated. The doctor has an obligation to the person being examined or treated to use those methods that reflect the current state of the art. In determining diagnostic methods and treatments based on current knowledge, there is often a choice of several possible measures. However, the doctor breaches his or her duty when performing a diagnosis or selecting a therapy that no longer appears justifiable and which therefore does not satisfy the objec6

Chart of signatures and ratifications can be found at https://www. coe.int/en/web/conventions/full-­l ist/-­/ conventions/treaty/164/ signatures?p_auth=LZowqlUU (last accessed 31 January 2022).

For example, the Swiss Federal Court, decision 28 November 2003, BGE 130 IV 7, 12 (standard of care  – tentative diagnosis); decision 20 December 2005, BGE 132 III 359 (failed sterilisation); decision 24 April 2008, BGE 134 IV 175 E. 3.2; decision 23  March 2017, 6B_1031/2016 E. 6.4. 8 For example, in Switzerland each canton has its own rules. For example see Patients’ Act of the Canton of Zurich (813.13), Section 13 et seq. 7

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Article 3 of the Charter of Fundamental Rights of the European Union: right to the integrity of the person • Everyone has the right to respect for his or her physical and mental integrity. • In the fields of medicine and biology, the following must be respected in particular: ○○ the free and informed consent of the person concerned, according to the procedures laid down by law; ○○ the prohibition of eugenic practices, in particular those aiming at the selection of persons; ○○ the prohibition on making the human body and its parts as such a source of financial gain; ○○ the prohibition of the reproductive cloning of human beings.

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Article 10 of the Convention on Human Rights and Biomedicine: private life and right to information • Everyone has the right to respect for private life in relation to information about his or her health. • Everyone is entitled to know any information collected about his or her health. However, the wishes of individuals not to be so informed shall be observed. • In exceptional cases, restrictions may be placed by law on the exercise of the rights contained in paragraph 2 in the interests of the patient.

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Article 8 of the ECHR: right to respect for private and family life • Everyone has the right to respect for his private and family life, his home and his correspondence. • There shall be no interference by a public authority with the exercise of this right except such as is in accordance with the law and is necessary in a democratic society in the interests of national security, public safety or the economic well-­being of the country, for the prevention of disorder or crime, for the protection of health or morals or for the protection of the rights and freedoms of others.

that time the medical ­intervention into his or her body. The patient’s consent must be based on the awareness of the authorised person and has to be declared before the examination or treatment begins. In order to prove an act of true self-­determination, the person to be examined must be consciously aware, capable of understanding and prepared to bear the consequences of his or her decision. What is of interest here is the conflict between autonomy and care. Due to a lack of sufficient legal provisions concerning whether the patient is capable of such a judgment, the literature and the national case laws surrounding the subject have developed their own basic principles. Irrespective of legal capacity under civil law and responsibility under criminal law, the capacity to consent is determined subject to the existence of the actual capacity to understand and to judge the significance and consequences of the intervention and the permission to perform it. The facts of each individual case are decisive in evaluating the capacity to understand and to express a will (Aebi-­ Müller et al. 2016). In principle, persons of legal age have the capacity to consent insofar as no specific reasons exist to suspect the presence of psychological defects, consciousness deficiencies, etc. The rejection of medically indicated treatment alone does not constitute sufficient reason to conclude a lack of capacity to consent (Figure 67.2). Even decisions that contradict objectively reasonable considerations can be in the best self-­ determined self-­interest.

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Box 67.2  Legal rules on self-­determination.

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right to autonomy with regard to one’s own body and health. Examination and treatment is permitted following consent by the bearer of the legal interests. This consent represents whether, and to what degree, the person concerned permits at

Self-determination: information about medical care needed for patient selfdetermination

Deficiency of will Self-­determination of the person to be examined does not symbolise an absolute value, but rather moves on a scale between self-­determination and freedom, on the one hand, and dependence, on the other hand. A self-­determined decision in favour of the medical examination and treatment needs a decision that is made carefully, voluntarily and according to personal values. It should not run contra bonos mores; this term being taken from

Medical treatment (diagnosis, treatment, lege artis)

Range lex artis Therapeutic information concerning medical treatment

Figure 67.2  The three overlapping circles of duties concerning medical treatment, therapeutic information and information concerning self-­determination of the patient’s body and the necessary medical measures.

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‘Where, according to law, a minor does not have the capacity to consent to an intervention, the intervention may only be carried out with the authorisation of his or her representative or an authority or a person or body provided for by law. The opinion of the minor shall be taken into consideration as an increasingly determining factor in proportion to his or her age and degree of maturity.’

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The individual concerned shall as far as possible take part in the authorisation procedure.’

Due to their young age minors can be, but not necessarily, incapable of judgement. In fact, their capacity to understand and judge the significance and consequences of the medical examination and intervention depends in particular on the severity of the intervention and the patient’s intellectual and mental maturity. In Switzerland, the Swiss Federal Court rejects fixed age limits, with the consequence that age, type of medical intervention and its therapeutic necessity be taken into account in each individual case.10 Some legal scholars are of the opinion that children under the age of twelve years old should generally be considered as being unable to reason, whereas a mature twelve-­to fourteen-­year-­old boy or girl may give consent to a moderate, riskless intervention, if their capacity of judgement has been clarified individually. From the age of 16 years old on, the capacity of judgement may generally be presumed if there are no indications to the contrary. Notwithstanding the above, the capacity of judgement of such minors will still need to be assessed individually (Mirabaud et al. 2013; Büchler and Michel 2020). The Convention on Human Rights and Biomedicine (Article 6, paragraph 2: Protection of persons not able to consent) declares that:

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civil law.9 An act or omission that runs contrary to the mores of fair and just thinking people is deemed to be against good morals. As far as a medical therapy is concerned, the limits of good morals are reached only where the legal censure as an ethical minimum is derived clearly from the legal system. A meaningful, informative consultation and, thus, effective consent cannot occur if the patient  – whether temporarily or absolutely – is unable to understand and consider the main elements that are significant for a decision. Those who are psychologically ill and mentally disabled need special attention. Another question is whether the patient is capable of expressing his or her wishes and understanding. This question must generally be answered by the expert. Where such a deficit is present and where the patient cannot (any longer) manage his or her own affairs, the decision whether to perform a medical examination and treatment becomes a matter for the law or the appointment of an authority. Its consent cannot, however, be given freely, rather it must reflect the well-­being and, in principle, the wishes of the person in care. Regarding this question, Article 6, paragraph 3 of the Convention on Human Rights and Biomedicine (Protection of persons not able to consent) states that:

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In emergency situations, the appropriate consent often cannot be obtained. In this case, Article 8 of the Convention on Human Rights and Biomedicine (Emergency situation) states that:

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‘When because of an emergency situation the appropriate consent cannot be obtained, any medically necessary intervention may be carried out immediately for the benefit of the health of the individual concerned.’ However, advanced directives and their validity in their respective countries and other previously expressed wishes relating to a medical intervention by the patient also have to be taken into account (Article 9 of the Convention on Human Rights and Biomedicine).

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Article 41, paragraph 2 of the Swiss Code of Obligations states that: ‘A person who wilfully causes loss or damage to another in an immoral manner is likewise obliged to provide compensation’. Section 138, subsection (1) of the German Civil Code states: ‘A legal transaction which is contrary to public policy is void’.

The legal representative’s authorisation may be withdrawn at any time if his or her decision does not correspond to the best interest of a minor who is unable to reason.

Information Consent to a medical examination and treatment is an act of true self-­ determination only where it is combined with comprehensive information. The person charged with making the decision should be informed of all known significant factors, within the confines of reason. The decision to be taken is determined by a huge amount of factors with the result that the doctor’s sphere of responsibility – in relation to consultation – consists of correctly conveying and assessing the relevant factors, explaining appropriate opportunities offered by other diagnostic measures and providing expert support to the person seeking counsel in his or her decision for which he or she is individually responsible. The consultation must also cover the known successes and risks associated with the examination and treatment (Figure 67.3). Moreover, dynamic consent11 offers opportunities for Swiss Federal Court, 2 April 2008, decision BGE 134 II 235, 240. Budin-­Ljøsne et al. 2017 (human research), Mausbach 2020 (medical treatment) and Dankar et al. 2020 (genome sequencing).

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patient. In this case, his or her representatives have to be informed about all relevant issues.

I. In the therapeutic setting

The right not to know

ongoing communication to familiarise the patient step by step with the necessary treatment and to gradually incorporate new facts. Article 5 of the Convention on Human Rights and Biomedicine (General rule) states that:

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Professional secrecy: confidentiality Medical professional confidentiality is one of the oldest guarantees in the doctor–patient relationship and is an important part of medical professional ethics. Nowadays, the Hippocratic Oath has been updated by the Declaration of Geneva and the national ­professional rules of conduct concerning the duties of doctors and good medical practice (Box 67.4). All over Europe, there are different national and ethical regulations concerning medical secrecy. For example, in the UK, pro-

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‘An intervention in the health field may only be carried out after the person concerned has given free and informed consent to it.

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Figure 67.3  The two overlapping circles relate to medical treatment and human research. In the overlapping area, the medical duties as well as the duties concerning human research have to be considered. One example is drug testing during phase III of the drug development, when the drug is tested on a large number of patients but before the drug is licensed.

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II. In clinical research

The right to self-­determination does not automatically imply a corresponding obligation. The reverse of the right to information on the state of one’s own physical, mental and general health and body is the right to decide whether all findings should be provided. Each individual must be allowed to decide for him-­or herself the extent to which he or she wishes to be burdened by knowledge. If the patient chooses to refuse all information and instead places the fate of his or her health in the hands of the expert, this decision must be generally respected (Aebi-­Müller et al. 2016). However, the right not to know is not unconditional. Rejection of the right to know comes to a limit when a possible lack of information makes the patient vulnerable to third parties or when a lack of information may be dangerous for the patient. Although the requirements must be judged on an individual basis, a basic informative consultation with the person capable of understanding appears, in principle, to be requisite (Box 67.3).

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This person shall beforehand be given appropriate information as to the purpose and nature of the intervention as well as on its consequences and risks. The person concerned may freely withdraw consent at any time.’

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The patient or their representative must be given enough time before the intended examination and treatment in order to adequately consider the pros and cons before reaching a decision. As a general rule, the wording must be so chosen as to allow a medical layperson to understand the situation. Standard bulletins and datasheets containing information on how the examination is proceeded with and the associated risks are needed as a c­ omplement – but not as a surrogate – to the personal discussion. The existence and extent of a so-­called ‘therapeutic privilege’, enabling the doctor to restrict the information provided to his or her patient, seems to be doubtful in the light of the right to know and to decide for oneself whether to know the details (Christinat 2019). The acceptance and range of the ‘therapeutic privilege’ depend on national regulations.12 Nevertheless, the limits of such restrictions must be drawn very tightly and a majority of legal scholars are firmly opposed to such privilege (Hirsig-­ Vouilloz 2017). In practice, these restrictions come into consideration if a comprehensive informative consultation might lead to very serious adverse effects to the mental state or the health of the As regards Switzerland in particular, see Swiss Federal Court, 13  November 1980, decision BGE 105 II 284, E. 6, which first laid down the principle of the therapeutic privilege.

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Box 67.3  Legal rules on the right to know and not to know. Article 10 of the Convention on Human Rights and Biomedicine: private life and right to information • Everyone has the right to respect for private life in relation to information about his or her health. • Everyone is entitled to know any information collected about his or her health. However, the wishes of individuals not to be so informed shall be observed. • In exceptional cases, restrictions may be placed by law on the exercise of the rights contained in paragraph 2 in the interests of the patient. Article 6 of the Swiss Federal Act on Human Genetic Testinga: Right not to know • Every person has the right to refuse to receive information about his or her genetic status subject to Article 18 paragraph 2. Article 18 paragraph 2 of the Swiss Federal Act on Human Genetic Testinga: Right to self-­determination • The doctor must immediately inform the person concerned of the test result if there is an immediate physical danger to the person, to the embryo or to the fetus, which could be averted.  SR 810.12.

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Box 67.5  Section 203 of the German Criminal Code (GCC): Violation of private secrets.

Box 67.4  Medical ethics. Extract from the Hippocratic Oath (North 2009) All that may come to my knowledge in the exercise of my profession or in daily commerce with men, which ought not to be spread abroad, I will keep secret and will never reveal. Extract from the International Code of Medical Ethicsa A physician shall respect a patient’s right to confidentiality. It is ethical to disclose confidential information when the patient consents to it or when there is a real and imminent threat of harm to the patient or to others and this threat can be only removed by a breach of confidentiality.  https://www.wma.net/policies-post/wma-international-code-of-medicalethics/ (last accessed 31 January 2022). Sources: Adapted from North (2009); WMA International Code of Medical Ethics (2020).

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(1) Whoever unlawfully discloses another’s secret, in particular a secret relating to that person’s personal sphere or life or to a business or trade secret, which was revealed or otherwise made known to them in their capacity as 1. a physician, dentist, veterinarian, pharmacist or member of another healthcare profession which requires state-­regulated training to engage in the profession or to use the professional title; 2. a professional psychologist with a state-­recognised final scientific examination; [. . .] 7. a member of a private health, accident or life insurance company or a private medical [.  .  .] service incurs a penalty of imprisonment for a term not exceeding one year or a fine. (2) Whoever, without being authorised to do so, discloses another’s secret, in particular a secret relating to that person’s personal sphere of life or to a business or trade secret which was revealed or otherwise made known to them in their capacity as 1. a public official; [. . .] 6. a person who has been formally obliged by law to conscientiously meet his or her duty of confidentiality in the course of scientific research projects incurs the same penalty. [. . .] (4) Whoever, without being authorised to do so, reveals another’s secret which has become known to them in the exercise or on the occasion of their work as an involved person or in the performance of their duties as data protection officer for the persons referred to in subsections (1) or (2) incurs a penalty of imprisonment for a term not exceeding one year or a fine. [. . .] (5) Subsections (1)–(4) also apply if the offender, without being authorised to do so, discloses the another person’s secret following the person concerned’s death. [. . .]

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fessional confidentiality is described in Good Medical Practice (2013, last updated in 2019) Establish and Maintain Partnerships with Patients by the General Medical Council (GMC):13

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50. ‘You must treat information about patients as confidential. This includes after a patient has died.’

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The professional ethical obligations for nurses are set out in the Nursing and Midwifery Council (NMC) code of professional practice.14 Lord Goff 15 described the duty of confidence as follows:

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‘A duty of confidence arises when confidential information comes to the knowledge of a person in circumstances where he has notice, or is held to have agreed, that the information is confidential, with the effect that it would be just in all the circumstances that he should be precluded from disclosing the information to others.’

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In the UK, the right to confidentiality is protected by the Data Protection Act 2018. If medical secrecy is violated, an individual can sue through a civil court. That person can also complain to the Information Commissioner if there is a breach of the Data Protection Act 2018.16 In Germany, the medical secret is protected by the GCC (Box 67.5) as well as by the Data Protection Act, the German Civil Code and the Professional Code for Physicians in Germany of the http://www.gmc-­uk.org/guidance/good_medical_practice.asp (last accessed 31 January 2022). 14 The Code  – Professional standards of practice and behavior for nurses, midwifes and nursing associates: https://www.nmc.org.uk/ globalassets/sitedocuments/nmc-publications/nmc-code.pdf (last accessed 31 January 2022). 15 Spycatcher case (Attorney General v. Guardian Newspapers Ltd, No 2 [1990] 1 AC 109). 16 http://www.legislation.gov.uk/ukpga/2018/12/contents/enacted (last accessed 31 January 2022). 13

German Medical Association17 (section  9, Confidentiality) and the guidelines of the various State Chambers of Physicians. Section 203 of the GCC provides for punishment with imprisonment of no more than 1 year or a fine for every wilful18 breach of medical professional confidentiality, in principle particularly when the persons to whom the secret is disclosed are bound to secrecy on their part – for example as doctors or a body representing the medical profession (for more information see Tag 2022). Both the fact that a medical examination or treatment has been performed and that the information thus produced constitutes medical findings which, as a rule, are protected under section 203 of the GCC. A prerequisite is that the information is obtained during professional activities. If the doctor divulges the secret, he or she will not necessarily be punished. The code mentions the one limitation regarding professional secrecy: consent of the person concerned. A justification of the disclosure can also result from the general justification, for example necessity (section 34 of the GCC): https://www.bundesaerztekammer.de/fileadmin/user_upload/ downloads/pdf-Ordner/MBO/MBO-AE_EN_2018.pdf (last accessed 31 January 2022). 18 In this case negligence remains without criminal punishment, but can however be punished by disciplinary measures. 17

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disclosure can also result from the general justification motives (e.g. in the case of emergency). The SCC provides for medical professional confidentiality in Article 321 and research confidentiality in Article 321bis.19 The right of refusal of the medical profession to testify, contained in both the Swiss Civil Procedure Code and Criminal Procedure Code20, complements the protection provided by substantive criminal law. Article 321 of the SCC provides for punishment with imprisonment or a fine for every wilful21 breach of medical professional confidentiality, in principle particularly when the persons to whom the secret is disclosed on their part (for example as doctors or a body representing the medical profession) are bound to secrecy. However, not every instance of the doctor or assistant passing on this confidential information is punishable under criminal law. For example, it does not constitute a punishable act of disclosure when a doctor discusses the medical results with a patient and explains a possible course of treatment to him or her. Moreover, as mentioned before, the SCC recognises in particular four restrictions on professional confidentiality (for more information see Tag 2004; Mausbach 2010; Oberholzer 2019).

67.2 Legal aspects of personalised medicine

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Whoever, when faced with a present danger to life, limb, freedom, honour, property or another legal interest which cannot otherwise be averted, commits an act to avert the danger from themselves or another is not deemed to act unlawfully if, upon weighing the conflicting interests, in particular the affected legal interests and the degree of the danger facing them, the protected interest substantially outweighs the one interfered with. However, this only applies to the extent that the act committed is an adequate means to avert the danger. The right of refusal to give evidence of the medical profession, contained in the German Code of Criminal Procedure (GCCP), complements the protection provided by substantive criminal law (Box 67.6). Genetic data available to a limited number of people are also protected by the SCC from possible disclosure by medical practitioners. A prerequisite is that information is obtained as part of the medical practitioner’s professional activities. If the doctor divulges the secret, he or she will not necessarily be punished. The SCC mentions four limitations regarding professional secrecy: consent of the person concerned, consent of the supervisory authority, the right or obligation to provide information to authorities and duty to give evidence. A justification of the

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Box 67.6  German Code of Criminal Procedure.

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Section 53 of the GCCP: right to refuse testimony on professional grounds 1. The following persons may also refuse to testify: [. . .] 3. [.  .  .] doctors, dentists, psychological psychotherapists, psychotherapists specialising in the treatment of children and juveniles, pharmacists and midwives, concerning the information that was entrusted to them or became known to them in this capacity. [. . .] 3a. members or representatives of a recognised counselling agency pursuant to sections 3 and 8 of the Act on Pregnancies in Conflict Situations, concerning that information that was confided to them or which became known to them in this capacity; [. . .] 2. The persons designated in subsection (1), sentence 1 nos. 2 to 3b may not refuse to testify if they have been released from their obligation of secrecy. [. . .]

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Section 53a of the GCCP: Right of professional assistants to refuse testimony 1. Persons who, in the context of 1. a contractual relationship, 2. a measure preparatory to vocational training or 3. some other ancillary activity, are involved in the professional activity of persons who have the right to refuse testimony on professional grounds pursuant to section 53 (1) sentence 1 to 4, shall be equal to such persons. The decision as to whether or not such persons shall exercise their right to refuse to testify shall be taken by the persons with the right to refuse testimony on professional grounds, unless such a decision cannot be obtained within a foreseeable time. 2. Release from the obligation of secrecy (section 53 (2) sentence 1) shall apply equally to the persons involved referred to in subsection (1).

67.2.1  Definition and areas of application

Personalised medicine – which is also called precision medicine or individualised medicine – includes diagnostic, preventive and therapeutic measures that are optimally tailored to an individual person.22 Personalised medicine has the potential to have a lasting impact on national healthcare systems (e.g. Hüsing et  al.  2008; Mathur and Sutton  2017; Gavan et  al.  2018). Individual people and their personal healthcare needs are already the focal point of medicine. So, what makes personalised medicine so different? The answer to this question is relatively simple as personalised The Swiss Federal Act on Research Involving Humans and its implementing ordinances, which entered into force in January 2014, undertook significant amendments concerning the protection of research confidentiality. In particular, Art. 321bis SCC now refers to Art. 34 of the Swiss Federal Act on Research Involving Humans as regards the conditions for lifting secrecy in the context of research, absent a valid and informed consent. 20 See Article 163, paragraph 1 lit. b of the Swiss Civil Procedure Code and Article 171 of the Swiss Criminal Procedure Code. 21 Negligence remains without criminal punishment and can however be punished by disciplinary measures. 22 For more information, see Leopoldina, expert opinion ‘Individualized medicine’ 2018. https://www.leopoldina.org/uploads/tx_leopublication/ 2014_Stellungnahme_IndividualisierteMedizin.pdf; Swiss Federal Office of Public Health (FOPH), Fact Sheet Personalised Medicine dated April 2018. 19

THE DOCTOR AND THE LAW

identification of individual risks of incurring disease, differential interventions, and unique therapeutic measures’.

67.2.2 Challenges The challenges and potentials related to personalised medicine are tremendous. They have been extensively analysed in several studies, in personal expertise (Rippe et al. 2004; Hüsing et al. 2008; Maughan 2017) and in symposiums.24 The following focuses on some of the essential areas.

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Doctor–patient relationship

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First of all, the emergence of personalised medicine has given rise to various challenges in the doctor–patient relationship. Personalised medicine generates a considerable increase of knowledge and has resulted in fundamental changes in existing directives and standards being required. Furthermore, medical practitioners using personalised medicine have to be provided with clinical knowledge through expert information systems. Patients need to be competent to deal with personalised medicine because, if not, ill persons may regress to being ‘patronised patients’, becoming test subjects against their own will. There is a real risk that it might become obligatory to use these new possibilities in medicine and to pass results on to third parties – within healthcare, but also with regard to employers, health and life insurance companies and others.

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medicine mainly involves the exploration of molecular causes and interrelationships of diseases. It is widely known that the differential diagnosis is not always right and the chosen therapy might not be effective or is suboptimal. In the case of cancer treatments, systemic therapies might be successful in some patients – even leading to a complete remission – while in others chemotherapy cannot stop the lethal course of the disease and may not satisfactorily improve the patients’ quality of life. In some patients, rheumatoid arthritis can cause severe joint damage after a short amount of time, while in others this does not occur (Leipe et al. 2011; Kłak et al. 2016). The list of examples could be continued ad infinitum. They make clear that available diagnostic and therapy procedures may not always be suitable for individual patients. One reason for this might be that in evidence-­based medicine the effectiveness of therapies is evaluated by randomised, placebo-­ controlled studies. However, relatively large patient groups are necessary in order to make statistically significant statements. Statistical conclusions drawn on the meso level are then applied to individual patients on the micro level. This occurs, for example, in oncology when patients within and outside studies receive treatments according to fixed therapy protocols. Statistical results are not always true for individual cases and prescribed rigid therapies with standardised drug dosages and intervals might not always be the ideal treatment for individual patients. Although disease patterns might be similar, causes and effects on a molecular level and individual living standards can be completely different. In order to make better treatments possible, more biomarkers and drugs are increasingly used that have been developed with the help of genome research (Schmitz et al. 2008; Brittain et al. 2017). In this context, it is helpful to explain the meaning of ‘biomarkers’ (short for ‘biological markers’): a biomarker is a collective name for cells, genes, determined molecules, enzymes, proteins and hormones. According to a definition from the FDA-­NIH Biomarker Working Group’s glossary (2016),23 a biomarker is a ‘defined characteristic that is measured as an indicator of normal biological processes, pathogenic processes, or biological responses to an exposure or intervention, including therapeutic interventions. Molecular, histologic, radiographic, or physiologic characteristics are types of biomarkers’. A biomarker is not an assessment of how an individual feels, functions or survives. Well-­known examples of biomarkers are values that are collected during the analysis of a blood sample and can be indicators for the health situation of the patient, for example blood sugar or the number of thrombocytes or cancer cells (Riener  2011; Califf  2018). Thanks to such treatments, patients can be divided into ­subgroups leading to more precise diagnoses, therapies and follow-­up treatments. According to the German working paper Individualised Medicine and Health Care Systems (Hüsing et  al.  2008), five different concepts can be identified within individualised ­ ­medicine: ‘biomarker-­based stratification (group formation), genome-­based information about health-­related characteristics,

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Economic issues Closely related to this are the economic challenges of personalised medicine. In addition to socioeconomic conditions, the principle of cost efficiency in national healthcare systems creates challenges, constituting a principal barrier to the advancement of precision medicine. In practice, the economic case for precision medicine is a challenge to demonstrate (Gavan et al. 2018), which has a considerable impact on societal conditions of personalised medicine.

Research status It is a big challenge to explore and identify genes and molecular structures. Such developments make us hope and worry at the same time, mainly due to the following areas: the performance For example, the ICPerMed Conference ‘Personalised Medicine in Action’, 20-­21 November 2018, Berlin, Germany. See also https://www. icpermed.eu/en/ICPerMed-Conference-2018.html

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Medical treatment (diagnosis, treatment, lege artis)

Therapeutic information concerning medical treatment

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Figure 67.4  The three overlapping circles of duties concerning medical treatment, therapeutic information and information concerning self-­determination of the patient’s body and the necessary medical measures. However, if the patient does not want to know the whole truth concerning his or her illness or details of all the steps which are included in the medical treatment, he or she has the right not to know. This right is not limitless; as a minimum, the patient has to be informed about his or her state of health and the basic necessary steps of treatment. The so-­called therapeutic privilege – that is not fully informing the patient in order to avoid mental harm and to treat him or her without information and self-­determination – cannot be accepted in all situations.

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and validity of markers, the processing, storage and reliable ­handling of data, the readiness of research institutes to collaborate and a paradigm shift from financially attractive, all-­in-­one blockbuster medication and diagnosis procedures for large patient populations toward solutions for stratified groups.

67.2.3  Personalised medicine and the law

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Personalised medicine is also of legal concern. The recent years have seen many initiatives come to light, be it internationally – such as the International Consortium for Personalised Medicine (ICPerMed) bringing together over 30 European and international partners25 – or nationally – with the example of the Swiss Federal Office of Public Health (FOPH) which has launched a working group and issued a report in 2017 on developments in data-­driven medicine and the associated challenges.26 It is however beyond the scope of this chapter to discuss all the national and international legal implications and regulations. https://www.icpermed.eu/ (last accessed 31 January 2022). The FOPH’s report introduces the topic, highlights opportunities and risks, and presents an overview of current initiatives in the field of data-­driven medicine in Switzerland. It also points out the interfaces and interactions of the topic with the activities and tasks of the FOPH together with the associated challenges. The report is an ‘evolving working paper’ and is updated regularly by the working group.

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The following considerations are therefore limited to the most crucial points. Legal issues in personalised medicine mainly refer to two large thematic fields: the use of personalised medicine before and during a disease and the use within the context of medical research. Both fields may partially overlap. Medical intervention using personalised medicine has to be applied according to state-­ of-­ the-­ art science and lex artis (Figure 67.4), and general provisions of medical and healthcare law are applicable. However, an individual patient’s treatment can also be connected to a general research interest. This is, for example, the case when the safety and effectiveness of new drugs are evaluated in larger groups of patients in phase III randomised studies or checked during phase IV studies once the medication has been launched. In this case, not only general provisions of medical law apply, but also the legal and ethical provisions concerning medical products and medical devices. In contrast, clinical trials with active substances in phase I and phase II belong to research. According to their definition, phase I tests do not normally aim at direct proof of a therapeutic benefit, but it is the first test of an active substance on human beings. In phase II studies, active substances are used in patients who are affected by the target disease. Additionally, there are situations in medical routine where therapies are not part of the standard procedure and traditional methods fail. Therapies that are (still) not established might be applied outside clinical trials if the type and severity of the disease requires such action. Depending on national regulations, individual treatment methods are not always considered to be

THE DOCTOR AND THE LAW

Freedom of therapy

Genetic testing

In the case of personalised medicine based on molecular connections, it has to be highlighted that in some countries the consent to genetic testing is subject to special regulations.32 The Additional Protocol to the Convention on Human Rights and Biomedicine concerning Genetic Testing for Health Purposes33 provides for some rules on the topic as well (Box 67.7 Articles 1 and 2). Accordingly, human genetic testing may only be done if the person gives permission and non-­discrimination is ensured. Special emphasis is put on self-­determination, the right to know and not to know, non-­directive counselling for presymptomatic or prenatal genetic testing and tests for family planning purposes. The additional protocol provides that no one may be discriminated against on grounds of his or her genetic material. This guarantee is already enshrined under constitutional law for some nations. The additional protocol also explicitly regulates the right not to know. According to a provision of the protocol, every person who is capable of judgment has the right to refuse to receive information about his or her genetic status (Box 67.7 Article 16). However, the right to self-­determination does not imply the obligation to self-­determination. If the person concerned is incapable of judgment, his or her legal representative has to be informed and then has to decide while involving the patient to the highest possible extent. Appropriate prior information has to be provided to the person not able to consent in respect of whom the test is envisaged, to the extent of his or her capacity to

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In the context of treatment contracts, medical practitioners have, in general,27 the right to diagnose and treat according to the methods that they consider the most promising. They are free to apply methods of personalised medicine. However, they have to respect current state-­of-­the-­art science and recognised medical practices. They are obliged to inform the patient if there are several possible therapies with different impacts and chances. The mere fact that some of the relevant methods are not covered by basic insurance is certainly not a justification for not informing the patient about the alternatives. If there are several equally suitable therapies with different risks, the method that best guarantees improvement and has the least side effects should be chosen. This also applies to individual therapies within personalised medicine. It does not violate lex artis per se if a drug is administered under compassionate use or off-­label use.28 ‘Compassionate use’ stands for the use of a drug which might be effective, but has not been approved in patients in life-­threatening situations,29 and ‘off-­label use’ stands for drugs that are given to patients outside the approved indication.30

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In some countries, medical examination and treatment including body or health interventions for the patient is considered to be personal injury. In other countries special provisions exist. However, as a rule, medical intervention is justified by the explicit informed consent of the patient or his or her representative. This implies that medical treatment requires the prior informed consent of the patient as a rule. But there are some important exemptions: if the patient is not capable of judgment, in emergency cases, if the person had lost his or her capacity (e.g. as a consequence of dementia), or in the case of children and adolescents who are not yet capable of judgment. If so, the legal representatives have to be adequately informed about the disease pattern, therapy options and consequences.

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However, medical practitioners are more challenged when it comes to product surveillance or informing the patient.31

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research in the true sense and, subsequently, are not considered as clinical trials. In this case, individual nations have to point out the applicable provisions for these therapies, in order to be aware of existing legal grey areas or weak spots and perhaps determine the needs for action.

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As for cost reimbursement, special regulations may be applicable. See Swiss Federal Court, decision 14 September 2004, BGE 130 V 532, 542 et  seq.; decision 21 September 2005, BGE 131 V 349, 351; and decision 24 April 2008, BGE 134 IV 175 (off-­label use). 29 See Swiss Federal Court, decision 20  June 2008, BGer 6B_40/2008; decision 19 June 2013, BGE 139 V 375; and decision 9 May 2017, BGE 143 V 130. 30 See Swiss Federal Court, decision 14 September 2004, BGE 130 V 532, 542 et seq.; dosage decision 21 September 2005, BGE 131 V 349, 351; decision 23  November 2010, BGE 136 V 395 (off-­ label use and myozyme); decision 19 June 2013, BGE 139 V 375; and decision 7 June 2016, BGE 142 V 325. See also Articles 71a and 72a of the Swiss Federal Ordinance on Health Insurance (KVV) regarding the reimbursement of off-­label-­use drugs by insurers, which entered into force in 2011. 27

For more information about dealing with this issue in the German legislation, see the German Federal Court, decision 27 March 2007, VI ZR 55/05, Versicherungsrecht 2007, 1104, with further references. 32 For example, in Switzerland, the Swiss Federal Act on Human Genetic Testing (HGTA). 33 https://rm.coe.int/1680084824 (last accessed 31 January 2022). 31

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Genetic data available to a limited number of people are also protected by national regulations from possible disclosure by medical practitioners. The general provisions also apply to personalised medicine. In some national legal systems, the use of medical information for research purposes in the fields of medicine or healthcare is protected by criminal law, for example Article 321bis of the SCC, the application of which is limited to research into human diseases and concerning the structure and function of the human body. The anonymous use of data, on the basis of which no conclusions may be drawn as to the identity of the patient, is already deemed to not constitute an offence.34 By referring to Article 34 of the Swiss Federal Act on Research Involving Humans (Human Research Act), Article 321bis, paragraph 2 of the SCC provides for the conditions for the use and disclosure of the non-­anonymous use of such information – i.e. professional secret – obtained in the course of research activities. Such information may be disclosed if the requirements of the above-­mentioned legal provision are met and if an authorisation for disclosure has been obtained from the responsible ethics committee (Staempfli Haenni  2017; Oberholzer 2019). In order to conform to the personal rights guaranteed by the Swiss constitution and with the interests in authorising the use of such data, the Human Research Act, which entered into force in 2014, provides for a modified consent solution.35

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Article 13 – Tests on persons not able to consent Exceptionally, and by derogation from the provisions of Article 6, paragraph 1, of the Convention on Human Rights and Biomedicine and of Article 10 of this Protocol, the law may allow a genetic test to be carried out, for the benefit of family members, on a person who does not have the capacity to consent, if the following conditions are met: a. the purpose of the test is to allow the family member(s) concerned to obtain a preventive, diagnostic or therapeutic benefit that has been independently evaluated as important for their health, or to allow them to make an informed choice with respect to procreation; b. the benefit envisaged cannot be obtained without carrying out this test; c. the risk and burden of the intervention are minimal for the person who is undergoing the test; d. the expected benefit has been independently evaluated as substantially outweighing the risk for private life that may arise from the collection, processing or communication of the results of the test; e. the authorization of the representative of the person not able to consent, or an authority or a person or body provided for by law, has been given; f. the person not able to consent shall, in proportion to his or her capacity to understand and degree of maturity, take part in the authorization procedure. The test shall not be carried out if this person objects to it.

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Article 2 – Scope 1. This Protocol applies to tests, which are carried out for health purposes, involving analysis of biological samples of human origin and aiming specifically to identify the genetic characteristics of a person which are inherited or acquired during early prenatal development (hereinafter referred to as ‘genetic tests’). 2. This Protocol does not apply: a. to genetic tests carried out on the human embryo or fetus; b. to genetic tests carried out for research purposes. [. . .]

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Article 1 – Object and purpose Parties to this Protocol shall protect the dignity and identity of all human beings and guarantee everyone, without discrimination, respect for their integrity and other rights and fundamental freedoms with regard to the tests to which this Protocol applies in accordance with Article 2.

­ nderstand. Within narrow boundaries, the additional protocol u thus allows a genetic test to be carried out, for the benefit of family members, on a person who does not have the capacity to consent (Box 67.7, Article 13). Article 14 of the additional protocol states the requirements for the use of biological materials previously removed for another purpose when it is not possible, with reasonable efforts, to contact a person for a genetic test for the benefit of his or her family members. In this case, the national ‘law may allow the test to be carried out in accordance with the principle of proportionality, where the expected benefit cannot be otherwise obtained and where the test cannot be deferred’.

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Box 67.7  Additional Protocol to the Convention on Human Rights and Biomedicine, concerning Genetic Testing for Health Purposes

Article 16 – Respect for private life and right to information 1. Everyone has the right to respect for his or her private life, in particular to protection of his or her personal data derived from a genetic test. 2. Everyone undergoing a genetic test is entitled to know any information collected about his or her health derived from this test. The conclusions drawn from the test shall be accessible to the person concerned in a comprehensible form. 3. The wish of a person not to be informed shall be respected. 4. In exceptional cases, restrictions may be placed by law on the exercise of the rights contained in paragraphs 2 and 3 above in the interests of the person concerned.

Data protection Genetic data are also subject to European regulations (Box 67.8), in particular the General Data Protection Regulation (GDPR)36, See Article 2, paragraph 2 Human Research Act. https://www.bag.admin.ch/bag/en/home/medizin-und-forschung/ forschung-am-menschen.html (last accessed 31 January 2022). 36 Regulation (EU) 2016/679 of the European Parliament and of the Council of 27 April 2016 on the protection of natural persons with regard to the processing of personal data and on the free movement of such data, and repealing Directive 95/46/EC (General Data Protection Regulation): https://eur-­lex.europa.eu/legal-­content/EN/TXT/?qid=1 532348683434&uri=CELEX:02016R0679-­20160504 (last accessed 31 January 2022). 34

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Health insurance law

The doctor–patient relationship is still mainly regulated by the provisions of national health insurance laws. Some of the national acts on health insurance42 are based on the principle of compulsory membership and solidarity (Swiss Medical Board  2009; Gächter and Rütsche 2018). For example, all persons domiciled in Switzerland must be insured for sickness (Swiss Federal Act on Health Insurance (KVG), Article 3). These persons are free to choose any authorised health insurer (KVG, Article 4). The persons who do not comply with the obligation to be insured for sickness are being affiliated to an insurance company by the relevant Canton (KVG, Article 6). Insurance companies are obliged to accept contracts with every eligible person in their area of activity (Swiss Federal Act on the Supervision of Social Health Insurance, Article 5, lit. i). This means that eligible persons have to be offered a contract for mandatory Swiss health insurance, regardless of their personal health situation and disease risks. Insurance companies are not allowed to ask for disclosure of results from prior presymptomatic or prenatal genetic testing and tests for family planning purposes (Human Research Act, Article 27, paragraph 1). The companies determine the amount of insurance fees consistently and independently from individual disease risks (KVG, Article 61).43

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Article 6 of Regulation (EU) 2016/679 of the European Parliament and of the Council of 27 April 2016 on the protection of natural persons with regard to the processing of personal data and on the free movement of such data, and repealing Directive 95/46/EC (General Data Protection Regulation): Lawfulness of processing Processing [of personal data] shall be lawful only if and to the extent that at least one of the following applies: a. the data subject has given consent to the processing of his or her personal data for one or more specific purposes; b. processing is necessary for the performance of a contract to which the data subject is party or in order to take steps at the request of the data subject prior to entering into a contract; c. processing is necessary for compliance with a legal obligation to which the controller is subject; d. processing is necessary to protect the vital interests of the data subject or of another natural person; e. processing is necessary for the performance of a task carried out in the public interest or in the exercise of official authority vested in the controller; f. processing is necessary for the purposes of the legitimate interests pursued by the controller or by a third party, except where such interests are overriden by the interests or fundamental rights and freedoms of the data subject which require protection of personal data, in particular where the data subject is a child. Point (f) of the first subparagraph shall not apply to processing carried out by public authorities in the performance of their tasks. [. . .]

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Article 8 of the Charter of Fundamental Rights of the European Union: Protection of personal data 1. Everyone has the right to the protection of personal data concerning him or her. 2. Such data must be processed fairly for specified purposes and on the basis of the consent of the person concerned or some other legitimate basis laid down by law. Everyone has the right of access to data which has been collected concerning him or her, and the right to have it rectified. 3. Compliance with these rules shall be subject to control by an independent authority.

Data on health or intimate spheres are usually regarded as highly sensitive.39 For example, personal data may only be processed lawfully and the processing must, inter alia, be fair and transparent to the data subject (‘lawfulness, fairness and transparency’). Personal data must be collected for specified, explicit and legitimate purposes (‘purpose limitation’), must be adequate, relevant and limited to what is necessary in relation to the purposes for which they are processed (‘data minimisation’), must be accurate (‘accuracy’), kept in a form which permits identification of data subjects for no longer than necessary (‘storage limitation’) and processed in a manner that ensures appropriate security of personal data (‘integrity and confidentiality’).40 Additionally, consent must usually be given explicitly in the case of processing sensitive personal data or personality profiles.41

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Box 67.8  European regulations regarding data protection.

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which was adopted in 2016 and became enforceable in May 2018,37 as well as national acts of data protection.38 See also Directive (EU) 2016/680 of the European Parliament and of the Council of 27 April 2016 on the protection of natural persons with regard to the processing of personal data by competent authorities for the purposes of the prevention, investigation, detection or prosecution of criminal offences or the execution of criminal penalties, and on the free movement of such data, and repealing Council Framework Decision 2008/977/JHA: https://eur-lex.europa.eu/legal-content/EN/ TXT/PDF/?uri=CELEX:32016L0680&from=EN (last accessed 31 January 2022). 38 For more information see http://ec.europa.eu/justice/data-­protection/ law/index_en.htm (last accessed 31 January 2022). 37

See Article 9, paragraph 1 of the GDPR (Processing of special categories of personal data), and Article 3 point (a) of the Swiss Federal Act on Data Protection (Definitions). 40 See Article 5 of the GDPR (Principles relating to processing of personal data). 41 See Article 9, paragraph 2, particularly point (a) of the GDPR (Processing of special categories of personal data), and Article 4, paragraph 5 of the Swiss Federal Act on Data Protection (Principles). 42 In Switzerland notably the Swiss Federal Act on Health Insurance (KVG), SR 832.10; in Germany the Code of Social Law, Vol. V. 43 Supplementary insurance plans  – in addition to mandatory health, maternity, and accident insurance  – are subject to private law, even though they are carried out by insurance companies. In contrast to the mandatory health insurance, insurers are allowed to fix the insurance fees according to individual risks. 39

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treatment. Due to this, depending on the national regulations, insurers are obliged to carry out cost efficiency tests among medical practitioners and only pay for services in accordance with the law. Whether doctors fulfil the requirement of cost efficiency can be checked with statistical methods (Poledna 2009). This regulation not only ensures financing of the national health insurance, but also limits doctors in choosing therapies.50 Examples can be found in the guidance of the National Health Service (NHS) and the National Institute for Health and Care Excellence (NICE) in the UK;51 the recommendations of the German independent Institute for Quality and Efficiency in Health Care (IQWiG)52 and the recommendations of the Swiss Medical Board.53 The criteria of effectiveness, appropriateness and cost efficiency are of great relevance in the social insurance regulations of several countries. For this reason, in some countries medical boards and comparable service units have been established to protect healthcare consumers and the healthcare systems through evaluations of the effectiveness, appropriateness and cost efficiency of medical treatment. NICE, for example, is an independent organisation responsible for providing national guidance on promoting good health and preventing and treating ill health in the UK.54 In Germany, IQWiG is an independent scientific institute that investigates the benefits and harms of medical interventions for patients. The institute regularly provides information about the potential advantages and disadvantages of different diagnostic and therapeutic interventions.55 To evaluate the criteria of effectiveness, appropriateness and cost efficiency in medicine is a special challenge for personalised medicine. As services are adjusted to the findings in stratified patient groups or specific individual situations, traditional diagnosis and indication protocols might not always be meaningful enough. The way efficiency is evaluated will need to be adapted in the future. It might follow the model of how niche products such as orphan drugs are evaluated and dealt with.56 Whether personalised medical services are covered by additional insurance plans mainly depends on the risk evaluation when the contract is signed and the corresponding scope of services. People with higher genetic risks might be rejected. They will still have a basic health insurance; but the criteria of

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Mandatory Swiss health insurance pays the costs for diagnoses and treatment of diseases, including their consequences and those of preventative health check-­ups. The services are only provided if they are effective, appropriate and economical in the sense of KVG, Article 32. Comparable standards exist in some European countries although they differ in the details. In the opinion of the majority, a medical service is considered to be effective if it has – objectively speaking – the intended diagnostic, therapeutic and care benefits.44 In accordance with Article 32 paragraph 1  KVG, effectiveness has to be proved according to scientific methods. This is the case if a specific treatment ‘is considered to be appropriate by a broad group of researchers and practitioners of the medical science’.45 Usually, scientific (long-­ term) studies written according to international standards are the basis for evaluation. A retrospective judgment on one single case is not enough to assess effectiveness. The guidelines normally also apply for (niche) products. Although there are no long-­term studies required for them, they normally have to be largely considered to be appropriate in the medical scientific area and be effective in terms of national regulations on health insurance.46 Moreover, while the most reliable way of proving effectiveness remains clinical trials, other scientific methods – such as statistics – may also be used.47 In the field of complementary medicine, the examination of the effectiveness is carried out according to its own criteria (Swiss Federal Ordinance on Health Insurance, Article 35a). According to Article 118a of the Swiss constitution, complementary medicine must be taken into account, as a consequence of which a unilateral restriction to scientific and traditional medicinal methods is prohibited (Rütsche  2017; Oggier and Vokinger 2020). If there are several effective treatment possibilities, their appropriateness will be taken into account to decide on cost coverage (Eugster 2016). It is necessary to evaluate diagnostic and therapeutic benefits in each case and the risks related thereto.48 If the treatments are equivalent, the more cost-­efficient treatment should be chosen.49 The principle of cost efficiency requires service providers to reduce the services to an extent which is in the interest of the insurance policy holder and necessary for the purpose of the

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Effectiveness stands for the causal relationship between cause (medical measure) and effect (medical success) whereas a general suitability is enough to fulfill the objective (Swiss Federal Court, decision 11 December 2006, BGE 133 V 115, 117; and decision 28 February 2017, BGE 143 V 95, 98; Swiss Federal Office of Public Health (FOPH) Working Paper 2011; Gächter and Rütsche  2018; Oggier and Vokinger 2020). 45 Swiss Federal Court, decision 11 December 2006, BGE 133 V 115, 117. 46 Concerning the Swiss mandatory health insurance, see the Swiss Federal Court, decision 11 December 2006, BGE 133 V 115, 119. 47 Swiss Federal Court, decision 20 February 1997, BGE 123 V 53, 63. 48 Swiss Federal Court, decision 25 June 2001, BGE 127 V 138, 147, and decision 30 April 2004, BGE 130 V 299, 303. 49 Oggier and Vokinger 2020; Swiss Federal Court, decision 23 November 2010, BGE 136 V 395, 407. 44

Swiss Federal Court, decision 23  November 2010, BGE 136 V 395 (Myozyme); Rütsche 2017. 51 http://www.nice.org.uk/ (last accessed 31 January 2022). 52 https://www.iqwig.de/en/ (last accessed 31 January 2022). 53 https://w w w.swissmedicalb oard.ch/index.php?id=27&no_ cache=1&L=1 https://www.swissmedicalboard.ch/index.php?id=1 &L=1 (last accessed 31 January 2022). Concerning HTA, see Widrig and Tag 2014. 54 For further information see https://www.nice.org.uk/ (last accessed 31 January 2022). 55 https://www.iqwig.de/en/ (last accessed 31 January 2022). 56 Rarity of diseases and thus more complicated clinical trials are taken into account when evaluating the requirements for the scientific documentation in the approval process. 50

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Convention for the Protection of Human Rights and Fundamental Freedoms

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Since many areas of personalised medicine are being explored, existing provisions for research are applicable.57 In Europe, a huge range of provisions regulate human research. It is beyond the scope of this chapter to explain or describe all of them. The following is an attempt to highlight some important points. As the COVID-­19 pandemic well illustrates, medical research on human beings is essential for establishing the causes of diseases and developing new therapies. For example, new medications and treatment methods have to be tested for potency, tolerability and safety before they can be used in everyday medicine. The importance of a thorough process may also be illustrated by the public health crisis caused by the coronavirus, as for example it has been reported in 2020 that the American Food and Drug Administration (FDA) was facing political pressure to rush approvals for COVID-­19 treatments, posing significant risks to patients.58 There is no comprehensive and systematic regulation for medical research on human beings: the legal framework is fragmented, unstructured and sometimes contradictory. Legal policies regarding research on humans are often confusing. Existing national regulations are often scattered over a variety of laws. These circumstances are made more complex by the fact that findings from medical research on human beings are made both nationally and internationally, are analysed and  – sometimes in isolation, sometimes as bundled information – incorporated into in an overall context and evaluated. That this leads to a confluence of various moral concepts and ideals entails both opportunities and risks. In addition to new insights and significant synergy effects of linking data, a certain potential for conflict and the fear of known and unknown dangers must not be ignored. The requirements for a legal framework in the field of research on human beings have long been satisfied by so-­called alternative norms or soft law. Examples are the Nuremberg Code of 1947, the Helsinki Declaration of the World Medical Association of 1964 (including its amendments), the recommendations issued by the

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Council of Europe and the World Health Organization, the UNESCO Declarations on the Human Genome of 1997, on the Protection of Genetic Data of 2003 and on Bioethics and Human Rights of 2005, the ICH (International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use) guidelines on good clinical practice, the guidelines of the Organisation for European Economic Co-­operation and Development (OECD) on Human Biobanks and Genetic Research Databases and the Charter for Researchers issued by the European Commission. The Convention on Human Rights and Biomedicine and the Additional Protocol on Biomedical Research govern a lot of detailed requirements for human research. Both the convention and the protocol contain provisions regarding research on human beings. Some principles of the convention have been adopted into the national legal systems; however, some countries lag behind the provisions of international law. The need to improve national requirements arises primarily from constitutional law requirements on fundamental rights: the freedom of science, the freedom of expression, the fundamental rights of the test persons involved in research, the right to life, the prohibition of inhuman and degrading treatment, and the protection of private and family life.

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effectiveness, appropriateness and cost efficiency will be applied in order to protect the community  – based on the principle of mutual ­solidarity – from being overloaded. Due to this, personalised medicine might be able to provide therapies for determined diseases, but since these therapies cannot be covered by the insurance due to the high prices, patients might not receive the treatments. All the stakeholders will have to work together in order to find a solution to this challenge.

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An abridged version of the Convention for the Protection of Human Rights and Fundamental Freedoms is given in Box 67.9.

They can currently be found in the Law on Therapeutic Products, in the Stem Cell Research Act, the Human Research Act, the Federal Act on Human Genetic Testing, and the Law on Transplants in Germany and Switzerland. See also the recommendations on personalised medicine of the Swiss Academy of Medical Sciences (SAMW) dated 2019: https://www.samw.ch/de/Publikationen/Empfehlungen/Grundlagenpersonalisierte-medizin.html (last accessed 31 January 2022). 58 https://edition.cnn.com/2020/09/03/politics/white-house-fdacoronavirus-vaccine/index.html (last accessed 31 January 2022). 57

Convention on Human Rights and Biomedicine The Convention on Human Rights and Biomedicine regulates the basic principles for medical research on human beings. In order to protect humans participating in clinical trials for therapeutic products, and to ensure the quality of clinical trials, the Convention on Human Rights and Biomedicine contains special guidelines (Box 67.10). According to these guidelines, trial subjects have to be informed about the nature and purpose of the trial, all of the processes and investigations connected with the trial, the existence of other treatments, the anticipated risks, discomforts and benefits, their right to compensation in the case of harm attributable to the trial and their right to withdraw their consent at any time without impairment to their therapeutic care.59 Trial subjects must explicitly give their free consent in writing. Additionally, according to national requirements, ethics committees have to endorse the trial. Higher requirements are appliThe law does not make a distinction between healthy, volunteer test subjects and patients who might benefit due to participation because participating in a clinical trial is always linked to uncertainty (Dispatch Swiss Federal Act on Therapeutics Products, 1 March 1999, BB1 1999 3453, 3534 et seq.).

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Box 67.9  Convention for the Protection of Human Rights and Fundamental Freedoms.

Box 67.10  Convention on Human Rights and Biomedicine.

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Article 17: Protection of persons not able to consent to research 1. Research on a person without the capacity to consent as stipulated in Article 5 may be undertaken only if all the following conditions are met: i. the conditions laid down in Article 16, sub-­paragraphs i–iv, are fulfilled; ii. the results of the research have the potential to produce real and direct benefit to his or her health; iii. research of comparable effectiveness cannot be carried out on individuals capable of giving consent; iv. the necessary authorization provided for under Article 6 has been given specifically and in writing; and v. the person concerned does not object. 2. Exceptionally and under the protective conditions prescribed by law, where the research has not the potential to produce results of direct benefit to the health of the person concerned, such research may be authorized subject to the conditions laid down in paragraph 1, sub-­paragraphs i, iii, iv and v above, and to the following additional conditions: i. the research has the aim of contributing, through significant improvement in the scientific understanding of the individual’s condition, disease or disorder, to the ultimate attainment of results capable of conferring benefit to the person concerned or to other persons in the same age category or afflicted with the same disease or disorder or having the same condition; ii. the research entails only minimal risk and minimal burden for the individual concerned.

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Article 8: Right to respect for private and family life 1. Everyone has the right to respect for his private and family life, his home and his correspondence. 2. There shall be no interference by a public authority with the exercise of this right except such as is in accordance with the law and is necessary in a democratic society in the interests of national security, public safety or the economic wellbeing of the country, for the prevention of disorder or crime, for the protection of health or morals, or for the protection of the rights and freedoms of others.

Article 16: Protection of persons undergoing research Research on a person may only be undertaken if all the following conditions are met: i. there is no alternative of comparable effectiveness to research on humans; ii. the risks which may be incurred by that person are not disproportionate to the potential benefits of the research; iii. the research project has been approved by the competent body after independent examination of its scientific merit, including assessment of the importance of the aim of the research, and multidisciplinary review of its ethical acceptability; iv. the persons undergoing research have been informed of their rights and the safeguards prescribed by law for their protection; v. the necessary consent as provided for under Article 5 has been given expressly, specifically and is documented. Such consent may be freely withdrawn at any time.

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Article 3: Prohibition of torture No one shall be subjected to torture or to inhuman or degrading treatment or punishment.

Article 15: General rule Scientific research in the field of biology and medicine shall be carried out freely, subject to the provisions of this Convention and the other legal provisions ensuring the protection of the human being.

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Article 2: Right to life 1. Everyone’s right to life shall be protected by law. No one shall be deprived of his life intentionally save in the execution of a sentence of a court following his conviction of a crime for which this penalty is provided by law. 2. Deprivation of life shall not be regarded as inflicted in contravention of this article when it results from the use of force which is no more than absolutely necessary: a. in defence of any person from unlawful violence; b. in order to effect a lawful arrest or to prevent the escape of a person lawfully detained; c. in action lawfully taken for the purpose of quelling a riot or insurrection.

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Article 9: Freedom of thought, conscience and religion Everyone has the right to freedom of thought, conscience and religion; [. . .]

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Article 14: Prohibition of discrimination The enjoyment of the rights and freedoms set forth in this Convention shall be secured without discrimination on any ground such as sex, race, colour, language, religion, political or other opinion, national or social origin, association with a national minority, property, birth or other status.

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cable for people who need special protection, such as persons incapable of judgment or emergency patients.

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Human research in Switzerland Federal Act on Research Involving Humans Based on the amended article of the Swiss constitution, Article 118b, the Swiss Council of States adopted the Swiss Federal Act on Research Involving Humans on 30 September 2011 (Box 67.11).60 The latter entered into force on 1 January 2014. The Swiss Federal Act on Research Involving Humans is generally binding for health research on human beings. It establishes generally valid requirements for all research projects regarding human diseases and concerning the structure and function of the human body, which 60

https://www.bag.admin.ch/bag/en/home/medizin-und-forschung/ forschung-am-menschen/entstehung-humanforschungsgesetz.html (last accessed 31 January 2022).

Article 18: Research on embryos in vitro i. Where the law allows research on embryos in  vitro, it shall ensure adequate protection of the embryo. ii. The creation of human embryos for research purposes is prohibited.

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1. The Confederation shall legislate on research on human beings where this is required in order to protect their dignity and privacy. In doing so, it shall preserve the freedom to conduct research and shall take account of the importance of research to health and society. 2. The Confederation shall adhere to the following principles in relation to biological and medical research involving human beings: a. It is a requirement for any research project that the participants or their legal representatives have given their informed consent. The law may provide for exceptions. A refusal is binding in every case. b. The risks and stress for the participants must not be disproportionate to the benefits of the research project. c. A research project involving persons lacking the capacity to consent may be conducted only if findings of equal value cannot be obtained from research involving persons who have the capacity to consent. If the research project is not expected to bring any immediate benefit to the persons lacking the capacity to consent, the risks and stress must be minimal. d. An independent assessment of the research project must have determined that the safety of the participants is guaranteed.

permitted to make biological material and health-­ related data anonymous if they may result in information that might be of relevance for the participants regarding severe diseases. The Human Research Act explicitly regulates consent and information of the participants. Nowadays, the participant needs to have enough time before giving his or her consent. The affected persons have to give their consent before the extraction of biological material or the collection of health-­related personal data if it is intended for research purposes. The law also allows for the possibility of partial information. Participants shall only be informed partially, if – due to methodological reasons – it is not possible to completely inform the participant and if the research only bears minimal risks and impacts. Trial subjects have to be fully informed afterward. Highly vulnerable people The legislator developed a differentiated system regarding research on highly vulnerable people. The Human Research Act makes a distinction between children under the age of 14 years, teenagers between 14 and 18 years, persons who are incapable of judgment, pregnant women, emergency patients and prisoners. Additionally, a distinction is drawn between research with an expected direct use and research with benefits for outsiders. The legislator established several requirements for research – depending on the groups and the purpose. The corresponding parameters are informed consent of the affected person or his or her representative, right of veto of the person who is incapable of judgment, and the risks and impacts related to the research as well as the possible benefits for patients with the same disease. Details can be found in Article 21 et seq. of the Human Research Act. Handling data and biological material Handling data and samples gathered during research activities depends on whether it is genetic data or biological material. The affected person has to be informed and give his or her consent for open and encoded use. If it is used in an anonymous way, the person has to be informed previously and there must not be any objection. Missing consents regarding genetic and non-­genetic personal data and biological material can be replaced if it is not possible or is disproportionately difficult to comply with the normal collaboration requirements or if the affected persons cannot be exposed to them. There may not be any documented refusal and the interest of research must prevail over the interest of the affected person when exercising the right to self-­ determination. The decision about whether the requirements are fulfilled in a specific case is always up to the responsible ethics committee.62 Storing biological material and health-­related personal data for research purposes and the subsequent establishment of biobanks should be regulated in detail by a blanket law, pursuant to Article 43 of the Human Research Act.63 However, the Swiss Federal Council recently considered that there was no need for such regulation for the moment.

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Box 67.11  Article 118b of the Swiss Constitution: Research on human beings.

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involves persons, deceased persons, embryos and foetuses, biological material or health-­related personal data.61 Research in the context of personalised medicine is mainly based on the provisions provided by the Federal Act on Research Involving Humans. The following basic principles must be applied: preference of persons’ interests over interests of science and society, non-­discrimination, consent, information and right to refuse to receive information, prohibition of commercialisation of the body and its parts, and fulfilment of scientific requirements. In addition to the provisions regarding the protection of trial subjects, a research authorisation from the responsible ethics committee is necessary (Article 45 of the Human Research Act). General considerations The principle of subsidiarity of research on humans is applicable for research on people in general and on highly vulnerable persons. Participation in a research project with expected direct benefits has to be free of charge, while research which is of benefit to outsiders may be paid for adequately. It is

See Article 2, paragraph 1 of the Human Research Act. In contrast, the Swiss Federal Act on Therapeutic Products only establishes specific provisions for the therapeutic product sector (Swiss Federal Act on Therapeutic Products, Commentarial Report concerning the Prod-­ Draft to the Dispatch, February 2006, p.  126). See Article 53 of the Swiss Federal Act on Therapeutic Products, providing that: ‘For clinical trials of therapeutic products in humans, the Human Research Act of 30 September 2011 applies in addition to the provisions of this Act.’

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To conclude, general medical and public health law as well as the Swiss Federal Act on Research Involving Humans all cover personalised medicine with a partly solid, partly fragmentary legal basis. There are many obstacles to accessing patients or trial subjects and their health-­related and genetic data. Regulations aim to prevent such data from becoming mere commodities.

Jersey, Oregon, Vermont and Washington, as well as in the District of Columbia.67 However, the practice there is restricted to people with incurable diseases, requiring broad medical testing and consultation with physicians. It also requires that candidates be permanent residents of that particular state (or, as applicable, of the District of Columbia), be at least 18 years old, and be mentally competent. In Germany, the criminalisation of assisted suicide services was adopted in 2015. However, the ban was overturned by the Federal Constitutional Court in February 202068 because the prohibition of assisted suicide services violates the general right of personality (Article 2(1) in conjunction with Article 1(1) German constitution – or ‘Basic Law for the Federal Republic of Germany’) in its manifestation as a right to a self-­ determined death afforded to persons determined to end their own lives. While active euthanasia – or physician-­assisted death – is illegal in the United States of America, it is lawful in the Netherlands, Belgium and Luxembourg, as well as in Colombia and Canada.69 This entails strictly regulated procedures. In other European countries than those mentioned above, organisations offering medically assisted suicide are not yet widespread, which does not mean, however, that it is not possible. An increasing number of foreigners travel to Switzerland to make use of the assisted suicide service offered by these organisations, which could be prohibited in their own countries. This trend is called ‘suicide tourism’. The gradual establishment of organisations for medically assisted suicide did not arise ad hoc. Its roots date back to the middle of the 1980s when the German physician Julius Hackethal became involved with medically assisted suicide (Roxin  1984; Hirsch 1987) and campaigned for it to be allowed. On one occasion he placed potassium cyanide at the disposal of a woman who had fallen seriously ill with a facial cancer. She took it and died. Although a criminal investigation was initiated against Hackethal, he was not convicted. This was because the woman had taken the drug with her own hands in a self-­dependent way and therefore had control over the act of her suicide. In Switzerland, the associations for medically assisted suicide, Exit Deutsche Schweiz and Exit A.D.M.D. Suisse romande were founded in 1982, followed by Dignitas in 1998 and the German branch DIGNITAS-­

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Death is part of life and its biological limitations. With advancing mechanisation and medicalisation of dying, many people have become afraid of no longer being in control of the medical ­treatment at the end of their life. The choice of putting an end to one’s life would appear to be an alternative way out (Tag 2012b). In fact, the realisation of such a major decision often fails due to circumstances that require major effort as well as mental strength. Nowadays, however, in some countries of the world, it is possible to relatively easily obtain drugs with a lethal effect. In Switzerland, for instance, private organisations offer professionally assisted suicide, i.e. having physicians supplying lethal doses to terminally ill patients for self-­administration. Exit, the best known of these organisations for medically assisted suicide for official Swiss residents,65 accounts for approximately 130 000 members (according to Exit’s own statistics). Dignitas offers the opportunity of assisted suicide for foreigners in Switzerland.66 In order to better understand the end of life in Switzerland in all its dimensions, in 2011 the Swiss National Science Foundation launched a National Research Programme called ‘NRP  67, End of Life’. The results have been made available to decision-­makers in the healthcare system, as well as to politicians and professionals involved in the care of individuals at the end of life (Zimmermann et al. 2019). Assisted suicide is also legal in the American states of California, Colorado, Hawaii, Montana, Maine (since January 2020), New

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67.3  Assisted suicide, organised medically assisted suicide, and euthanasia

See Dispatch, Swiss Federal Act on Research Involving Humans, 21 October 2009, BBl 2009 8045, 8148. 65 http://www.exit.ch/ (last accessed 31 January 2022); members can only be Swiss or non-­Swiss with residence in Switzerland, cf. Article 3 of the statutes. The statutes are currently under consultation in order to be revised, so that  – inter alia  – only members with residence in Switzerland would be accepted. 66 http://www.dignitas.ch (last accessed 31 January 2022). 64

https://edition.cnn.com/2014/11/26/us/physician-assisted-suicidefast-facts/index.html (last accessed 31 January 2022). 68 ‘Criminalisation of assisted suicide services unconstitutional’ BVerfG decision dated 26 February 2020, 2 BvR 2347/15, 2 BvR 2527/16, 2 BvR 2354/16, 2 BvR 1593/16, 2 BvR 1261/16, 2 BvR 651/16. 69 Termination of Life on Request and Assisted Suicide (Review Procedures) Act,https://www.government.nl/topics/euthanasia/euthanasia-­assisted­suicide-­and-­non-­resuscitation-­on-­request (last accessed 31 January 2022); cf. Loi relative à l’euthanasie 28 May 2002; Loi relative aux soins palliatifs, 14 June 2002; Loi relative aux droits du patient, 22 August 2002, http://www.admd.be/information/legislation-­belge/ (last accessed 31 January 2022); Loi sur l’euthanasie et l’assistance au suicide, 16  March 2009, https://sante.public.lu/fr/droits/fin-vie/euthanasie/ index.html (last accessed 31 January 2022). 67

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other European countries, but not in the Low Countries, there is an explicit prohibition of active homicide, including direct active euthanasia – a deliberate killing in order to end the suffering of another person (Tag 2012a).71 This is the directed or at least knowing administration of a deadly drug or the application of another method leading to death, in the sense of a homicide. It can take place with or without the explicit request of the patient. In individual cases, direct active euthanasia is forbidden if no special regulations exist. In most countries, this form of euthanasia is punishable as murder, mercy killing on request or manslaughter (Art. 112, 113, 114 and Art. 115 SCC). If it is carried out with honourable motives (e.g. out of sympathy or based on a serious and insistent request of the future deceased), in several countries this offence carries a milder range of punishment than ‘normal’ intentional homicide. Criminal prohibition of homicide is applicable to everybody – to the treating physicians, to the nurses, to the caregivers in general, to the relatives and to the employees of an organisation for medically assisted suicide. However, as already mentioned, under special restrictions active medically assisted suicide is allowed in the Netherlands, Belgium and Luxembourg, as well as in Canada72 and Colombia73.

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Deutschland in 2005 (previously also Dignitate for a limited duration). In the year 1997, Exit International was founded in Australia (for more details see Venetz 2008). The activities of DIGNITAS Deutschland and other right-­to-­ die organisations gave rise to some harsh criticism in Germany. This led to the prohibition of assisted suicide services under German criminal law in 2015. After a long and controversial debate, this prohibition was overturned by the German Federal Constitution Court in February 2020. In Switzerland, the debate still persists as to whether organisations for medically assisted suicide should be state controlled. As far as organised suicide assistance is concerned, it should be said that it is more or less accepted within the limits of Swiss law. However, the limits of this permissibility are still being intensively debated in the course of a large number of questions (Zimmermann et al. 2019).

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This short introduction has shown that discussions about medically assisted suicide involve a lot of terms. For example, active medically assisted suicide, so-­called euthanasia, passive medically assisted suicide, indirect medically assisted suicide, direct medically assisted suicide, homicide at the request of the future deceased, aiding and abetting leading to suicide, organised medically assisted suicide, palliative medical treatment and care and so on. This diversity in terms and content has legal consequences because justice and injustice, criminal liability and impunity are often very close entwined. The average layperson can barely understand the juristic finesses.70 Therefore, the terms need to be explained briefly.

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Indirect active euthanasia describes the use of means to relieve suffering which may have side effects that may shorten life (e.g. when soothing drugs like morphine are being administered). The possibility that death might occur earlier than it would otherwise have done is taken into account. Whether it is possible in modern palliative medicine to dose so accurately that death is not hastened is under debate. Until now, the legal practice in many European countries assumed impunity concerning indirect active euthanasia, but this is now being actively discussed. In Switzerland, this type of euthanasia is usually viewed as permissible if some strict requirements are fulfilled (e.g. as reflected in the guidelines on the management of dying and death of 2018 of the Swiss Academy of Medical Sciences SAMS).74

Phase leading up to biological death

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In the medically assisted suicide, two periods of time are generally distinguished: the one leading to biological death and the one concerning the time before that. If death has not yet occurred but the course of the terminal disease has already passed the irreversible threshold of dying, the physician is neither entitled nor obligated to maintain the extinguishing life. His obligation to the patient consists of analgesia and facilitating a dignified death. Assistance in the dying phase is regarded as aiding the process of dying.

Previous phase

Active medically assisted suicide, known as  direct active euthanasia If the stage of inevitable death has not been reached, then there are many litigious issues. In Switzerland, Germany and many The German Ethics Council in 2006 proposed, therefore, to untangle the terms.

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In Switzerland, this form of euthanasia is punishable under Article 111 of the SCC (murder), Article 114 of the SCC (mercy killing on request) or Article 113 of the SCC (manslaughter). 72 https://laws-lois.justice.gc.ca/PDF/2016_3.pdf (last accessed 31 January 2022). 73 https://thebogotapost.com/step-­closer-­to-­legal-­euthanasia/5837/ (last accessed 31 January 2022). 74 See https://www.bj.admin.ch/bj/en/home/gesellschaft/gesetzgebung/ archiv/sterbehilfe/formen.html (last accessed 31 January 2022); The guidelines on the management of dying and death of the SAMS are available under  https://www.samw.ch/en/Publications/Medicalethical-Guidelines.html (last accessed 31 January 2022). 71

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Article 8: Right to respect for private and family life 1. Everyone has the right to respect for his private and family life, his home and his correspondence. 2. There shall be no interference by a public authority with the exercise of this right except such as is in accordance with the law and is necessary in a democratic society in the interests of national security, public safety or the economic wellbeing of the country, for the prevention of disorder or crime, for the protection of health or morals, or for the protection of the rights and freedoms of others.

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Article 9: Freedom of thought, conscience and religion 1. Everyone has the right to freedom of thought, conscience and religion; this right includes freedom to change his religion or belief and freedom, either alone or in community with others and in public or private, to manifest his religion or belief, in worship, teaching, practice and observance. 2. Freedom to manifest one’s religion or beliefs shall be subject only to such limitations as are prescribed by law and are necessary in a democratic society in the interests of public safety, for the protection of public order, health or morals, or for the protection of the rights and freedoms of others.

c­ oncerning the rights of the sick at the end of their life; termination of medical treatment at the request of a patient is then not subject to punishment when he or she is in an advanced or terminal phase of a serious and incurable disease – even if this termination accelerates death. If the patient no longer has a decision-­making ability and no advanced directive exists, then their legal representative – in most cases a next of kin or another person in a position of trust – has to be called in.

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Passive aid to the dying (e.g. through rejecting or discontinuing life-­ prolonging measures) is considered predominantly as an omission of medical aid which results in the occurrence of death. It is not governed by any specific statutory provisions and is not treated as a criminal offence provided certain conditions are fulfilled. The German Federal Court of Justice decided in 2010 that in the case of passive euthanasia it does not matter if an action or an omission is assumed.75 All actions that are related to a medical treatment should be summarised in normative/evaluative terms at the termination of treatment. This must embrace both the objective elements of action and the subjective goal setting of the doer to abort (an) already begun medical treatment measure(s) according to the will of the patient or to reduce its/their scope according to the will of the patient or his or her custodian. (In the latter case, an existent patient’s provision, the so-­called advanced directive, has to be considered.) Therewith, the distinction between an active and a passive action based on formal criteria has been mainly dismissed. As a result, new criteria have been established and the terms ‘medically assisted suicide’ and ‘termination of treatment’ are used. Medically assisted suicide through omission of treatment, limitation of treatment or termination of treatment assumes, according to the German Federal Court of Justice, that the affected person is life-­threateningly diseased and that the concerned measure is medically adequate for the preservation or prolongation of life. In this case, a justification of medically assisted suicide is possible through consent, implicit consent or an advanced directive. Intentional life-­determining actions that are taken beyond coherence with medical treatment of a disease cannot be justified through these requirements. This appraisal has to be endorsed as it lies within the judicious patient’s powers to determine which treatment he or she wants and which one they want to refuse. Even though his or her will is weighted differently in different countries, it is frequently agreed that the freedom to disease, the right to die in decent conditions and the right to one’s own death are protected by the rights to liberty and the respect for individual rights enshrined in the European Convention for the Protection of Human Rights and Fundamental Freedoms (Box 67.12). If national laws do not make the right to suicide a punishable offence, then the patient has the right to decide on the method and time of the termination of his or her own life. This applies to a person who is capable of exercising his or her will freely and to act accordingly. If he or she rejects a lengthening of life, then the medical obligation changes from the lengthening of life to end-­of-­life care. If this involves continuing life support measures against the patient’s will, then in many countries this arbitrary intervention is an assault, for example in Germany and Switzerland. In Austria, a criminal offence of arbitrary therapeutic treatment is possible. In France, there are regulations

Box 67.12  The European Convention for the Protection of Human Rights and Fundamental Freedoms.

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German Federal Court of Justice, decision 25 June 2010, 2 StR 454/09, Neue Juristische Wochenschrift (NJW) 2010, 2963 et seq.

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Participation in the suicide In some European countries, assisting suicide is a criminal offence. In other countries, such as France, while assisted suicide is not explicitly covered by criminal law provisions, existing laws are interpreted to the same effect. In some countries, the support of the self-­determined suicide of a person who is capable is not subject to litigation because being an accessory to the act is not an offence. Other conditions apply if a special offence exists for the collaboration and participation with a suicide. In Switzerland, Article 115 of the SCC states that the incitement and assistance to suicide are punishable offences when they are initiated by selfish motivated causes. Typically, these are egotistical reasons, for example when the offender targets a personal advantage. Organisations that offer assisted suicide within the framework of the Swiss legislation are not punishable provided they cannot be accused of having any self-­motivated reasons. It is still under scrutiny whether assisted suicide should or should not be part of a physician’s remit. If the aiding and abetting is done with impunity, then it does not matter if it is accomplished through an active action or an

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the prosecuting criminal authorities to effect binding directives defining in which cases the assistance should be subject to punishment and in which cases it is not. The new directive implied that relatives are not subject to prosecution when they act out of sympathy when the patient is terminally ill or has taken a decisive conscious decision to die and the aid has only been minimal. The legal debates continue because more severely ill persons have requested to commit assisted suicide.79

67.3.2  Some precedents

Piergiorgio Welby

Diane Pretty and Debbie Purdy

There are also situations when life-­sustaining apparatuses are switched off at the request of the person who wishes to die. Depending on the national legal situation and legal practice, the precedent is treated differently in different countries. Such a situation would traditionally count as passive euthanasia (see earlier). In most European countries, this is allowed when the person wishing to die demands it in a self-­determined way. However, the precedent of the Italian Piergiorgio Welby shows that this is not always easy. Piergiorgio Welby was taken terminally ill with myodystrophy, finally being totally immobilised and dependent on artificial respiration, but judicious. The requested termination of life-­sustaining measures was denied him by the Italian justice in spite of the fact that a passive medically assisted suicide had been demanded by him as a capable person’s wish to die. This self-­ determination is guaranteed by the Italian constitution in the case of medical treatments (Article 32, paragraph 2): ‘nobody may be forcefully submitted to treatment except as regulated by the law’. However, in another Italian case, a physician did comply with the request of a paralysed patient in December 2006 and turned off the respirator. A criminal procedure was initiated against him because of homicide at request; however, it ended with a verdict of not guilty (Becchi 2009). According to the general basic principles, a case of an allowed medically assisted suicide should have been accepted because the patient was life-­threateningly ill and the ventilation which was stopped was adequate for the preservation or prolongation of the life. As a consequence of this, the Roman Catholic Church refused a religious funeral.

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The precedent of the Briton Diane Pretty who was paralysed because of anterolateral sclerosis (ALS) demonstrates the problems and their development. In the year 2002, Diane Pretty brought a legal action to the European Court of Human Rights (ECHR). She wanted to make sure that her husband would be exempt from punishment if he helped her to kill herself. The reason was that Diane Pretty was still mentally capable but was no longer physically able to administer the deadly poison herself. If her husband had given her the deadly injection, then he would have been liable to prosecution according to the Suicide Act 1961 due to the assistance in the suicide. This would still have applied even where he submitted himself to the serious and explicit death wish of his wife and had ‘only’ been her ‘extended arm’. This is because incitement and assistance to suicide according to the Suicide Act 1961 were subject to punishment.76 The European Court of Human Rights rejected the application from Mrs Pretty because the UK Suicide Act 1961 was deemed not contrary to the sustaining articles of the ECHR.77 Diane Pretty died in a hospice a short time after that when under sedation which had been administered to her because of her heavy respiratory distress. The precedent of the Diane Pretty case has been of great concern to both the public and the politicians in the UK. It brought to light the situation where persons who are terminally ill, but are not able to commit suicide due to their disease, are strongly limited in their self-­determination at their life’s end. As a result, there was a controversial discussion about the Assisted Dying for the Terminally Ill Bill [HL] in 2006. This would have allowed physicians to aid medically assisted suicide at the request of the judicious patient. However, the development has not been completed because Debbie Purdy had brought another legal action to the House of Lords. She was suffering from multiple sclerosis and wanted to commit suicide with the aid of Dignitas, and with her husband accompanying her. However, in this case he would have been liable to prosecution. Debbie Purdy was entitled to make a legal claim.78 The highest English Board of Appeal then ordered

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omission. An indispensable condition for impunity is that the person who provides assistance should not take a dominant position with regard to suicide. Such impunity ends where the inducer or supporter is in control of the act. This might be the case if the inducer undertakes the deadly action by him-­or herself, but not if the victim attempts suicide as a result of misleading influences resulting in the loss of a balanced overview.

https://hudoc.echr.coe.int/eng# (last accessed 31 January 2022). ECHR, Fourth Section, case of Pretty v. the United Kingdom (application no. 2346/02), 29 July 2002. 78 http://news.bbc.co.uk/2/hi/uk_news/8265304.stm (last accessed 31 January 2022). 76 77

Eluana Englaro In the case where a person is no longer responsive (e.g. because the person is demented or in a coma), the question arises as to whether life-­sustaining measures can be stopped due to a patient’s previous provision or an implied consent of the ill patient. On this point, the legal situations of individual countries vary strongly. Partly this is because different laws exist, for example in Germany and in Austria, which regulate the requirements individually; elsewhere, however, equivalent regulations are found in the legislative procedures (e.g. in Switzerland) but interpretations vary. In many countries, patients’ provisions and implied consent for the termination of treatment fall under the general basic 79

For example, ‘Right-­to-­die man wins first step in legal battle’, BBC News, 27 January 2012.

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Although the role of the physician at the end of life primarily refers to the palliative support of the patient, there are different situations in which the patient wishes support from the physician, including assisted suicide. The relevant guidelines of the Swiss Academy of Arts and Sciences (SAMS) have been revised. The 2013 directive, ‘End of life care’, had the following requirements: the illness of the patient justifies the assumption that the end of life is near. Alternative possibilities of assistance were discussed and, if desired, used. The patient is able to judge, his wish is well-­balanced and without external pressure and is permanent. In addition, the examination must be carried out by an independent third party who does not necessarily have to be a doctor. The final action in the process leading to death must always be taken by the patient him- or herself. The new guidelines, ‘Management of dying and death’, which were adopted by the SAMS in June 2018, differ in particular from the requirement that the end of life is near. Instead, an ‘intolerable suffering’ shall apply.83 The extension in 2018 was based on the fact that ‘it is not the medically objectifiable condition that leads the patient to self-­determined suicide, but the subjectively experienced unbearable suffering’. This formulation is not without controversy. The FMH rejected the incorporation of the new guidelines into their professional medical law because there are no longer any objective, comparable and measurable criteria and euthanasia would be possible in the case of physical and psychological suffering that can be treated in principle. A physician would be authorised to provide suicide assistance to a patient who is only mildly ill with functional limitations or symptoms; provided the patient claims that he or she suffers unbearably from these.84 Further supervisory standards do not yet exist. So far, only Exit–Deutsche Schweiz has made an agreement with the senior public prosecution of the Canton of Zurich, which it did in the summer of 2009. These directives should prevent abusive practices, although the Swiss Federal Court has declared them to be unlawful.85 Nevertheless, these directives can be viewed as a reaction to the increasing pressure of society to either prohibit organisations for medically assisted suicide or to restrict their activities on the basis of predefined duties of care. Several nations, in particular Germany and Switzerland, are concerned about whether the activities of organisations for medically assisted suicide should be more regulated and, if so, in which legal area and to what extent. Such considerations cannot be limited to national regulations because of the free movement of persons wishing to die and their dying assistants. Such a discussion is answered very differently in the various countries. A broad range of arguments can be raised against

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The legal questions gain a new dimension if the aid is offered by professionals and when dying is not offered and accomplished in the individual case, be it by relatives, friends or a physician. If the aid is offered by a professional organisation for medically assisted suicide, then further problems arise. In Switzerland more and more people choose this way of dying. The following figures indicate that not only ethical but also financial questions are under consideration: Dignitas charges costs ranging from CHF 7500 to 10 500 per suicide, in addition to an admission fee and annual membership fee. Dignitas has accompanied more than 3027 persons to death since its foundation in 1998 so that it is not surprising that the society’s funds are growing.81 Exit also has considerable property assets, funded by membership fees, donations, capital gains from commercial papers and legacies. The expansion of organisations for medically assisted suicide in Switzerland, Germany and other countries is possible because the regulation of their activities is only rudimentary. Until now, only general regulations have been applicable to them, particularly the law of associations, the criminal law, the Medicinal Products Act and the Therapeutic Products Act or the Narcotics Act if SP is applied as the means for dying. The code of professional conduct of physicians is applicable if they provide their medical know-­how and facilities for the purposes of the organisation. In addition to federal and cantonal law, soft law, such as professional codes of conduct, is of central importance. The Code of Professional Conduct of the Swiss Medical Association FMH puts the decision on the assistance or its refusal concerning the patient’s end of life decisions under the physician’s conscience. They conclude that ‘assisted suicide is not a medical action to which patients could claim to be entitled; it is, however, a legally permissible activity’.82

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­ rinciples of law. This means that to some extent well-­meant p paternalistic considerations are used to decide about the continuation of a treatment. An example is the precedent of the Italian comatose patient Eluana Englaro. She had been in a coma for 16 years after a car accident which she had had as a 19-­year-­old, but she had given no advanced directive. There was a 9-­year struggle and a huge political discussion when her father requested the switching off of the life-­sustaining apparatuses. This was subsequently agreed to by the Italian Court of appeal in the year 2009,80 although it remained controversial. In this case, the court came to a decision due to the implied consent of the woman who had met an accident, although there were particular difficulties due to her longstanding comatose state.

http://www.economist.com/node/13110088 (last accessed 26 October 2020). http://www.dignitas.ch/images/stories/pdf/statistik-­ftb-­jahr-­wohnsitz-­ 1998-­2019.pdf (last accessed 31 January 2022). 82 Guidelines on the management of dying and death of the SAMS, 6.2.1: https://www.samw.ch/en/Publications/Medical-ethical-Guidelines. html (last accessed 31 January 2022). 80

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Act88 by providing a lethal dose of pentobarbital to an 86-­year old woman. The woman, healthy at the time, was determined to die at the same time as her gravely ill husband. The doctor was condemned to a suspended 120-­day fines sentence. The judgement was confirmed by the appellate court in April 2020, he appealed to the Federal Court. They decided that in the case of a “Bilanzsuizid” of a healthy person, there is no medical indication for the prescription of sodium pentobarbital, especially since the drug is not used “therapeutically” in the broadest sense. Thus, a conviction for dispensing sodium pentobarbital on the basis of the Therapeutic Products Act, TPA, was out of the question in the present case and proves to be contrary to federal law. Thus, the case was brought back before the lower court for a new judgement.89 Pursuant to both courts, the applicable legislation and rules on assisted suicide were not complied with, as the deceased woman did not suffer from any illness and was not at the end of her life. The judges indeed deemed that assisted suicide for ‘existential reasons’ is not permitted under Swiss law. While the sentence remains light, this conviction is a step in the right direction, confirming a certain framework for medically assisted suicide.

67.4  Research on corpses Whether research on corpses is permissible, for example neuroimaging, virtopsy, etc., is becoming ever more relevant and is seen as an urgent problem since many years (Tag 2006b). The exploitation of the dead body, the organs, cells and DNA represents a resource of increasing importance in generating new findings. Given the uncertain ethical and legal evaluation of the treatment of corpses, the following seeks to provide arguments in the interest of clarification. The intention is to find a valid balance between the legitimate concerns of the research community and the continuing personal rights of those whose corpses are to be examined and the rights of their next of kin. Whereas dying – as we understand it from a medical and technical point of view – describes an irreversible biological process from a certain point on, the speed of whose progression varies depending on the individual tissue type, death represents a more or less swiftly occurring event, depending on the circumstances of the specific case, within this entire process. The point at which a person can be considered to be still alive is a question which jurisprudence and philosophy have to clarify although the criteria for brain death have been defined in most national transplantation acts. Furthermore, the definition of the criteria of death involves not just biological and medical, but also legal and ethical features.

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­absolute prohibition.86 For example, self-­determination is essential because free capable people should be able to decide freely at the end of their lives. At the same time, apprehension exists that a prohibition would be unenforceable if the association has a high membership figure, as in Switzerland. An embedment in the criminal law could possibly lead to criminal prosecution being avoided. This has already been seen in former restrictive regulations for abortion which are not sustainable in the long run for states governed by the rule of law. Such a slippery slope and the possible misuse of the regulations should be taken seriously, although it does not lead to a moral capitulation of the state if no absolute prohibition is pronounced. It is clear, therefore, that for a country like Switzerland organisations for medically assisted suicide should not be forbidden. Instead, binding due diligence criteria should be defined that have to be adhered to by such organisations: for example, they must consider whether the person wishing to die has conceived and expressed his or her will freely, if it is well considered and if it remains in perpetuity. Assisted suicide has to be based on the decision of an autonomous subject. Therefore, if a request for assisted suicide is to be granted, mental capacity is of crucial importance. In addition to that, it has to be clarified how to deal with non-­judicious persons having or not having advanced directives. Another set of issues concerns the participation of physicians. This also raises the question whether the person has to be terminally ill, if severe chronic diseases are sufficient or if the suicide should be facilitated for healthy and depressive persons too. In addition, it has to be clarified how to deal with persons wishing to die. Is any assistance available other than suicide? Has this been discussed with him or her? Are palliative care hospices and medically assisted suicide mutually exclusive? Last but not least, it has to be resolved how one could counteract the commercialisation of suicide. In June 2011, the Swiss Federal Council decided to refrain from including specific regulations on organised assisted suicide in the criminal law. The Swiss Federal Council is of the opinion that any misuse can be dealt with under the current legal provisions: the current criminal law provisions, the Swiss Federal Therapeutic Products Act and the Swiss Federal Narcotics Act and conduct rules. The Swiss Federal Council continues to give priority to preventing suicide, to supporting palliative care, and the care and treatment of people with terminal, life-­threatening and chronic illnesses.87 Finally, the regional vice president of Exit in western Switzerland, Pierre Beck, was sentenced by a Geneva court in October 2019 for helping a healthy woman dying. More precisely, the doctor was found guilty of breaking the Therapeutic Products

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For Germany cf. particularly Federal Constitutional Court Judgment of 26 February 2020, 2 BvR 2347/15, 2 BvR 2527/16, 2 BvR 2354/16, 2 BvR 1593/16, 2 BvR 1261/16, 2 BvR 651/16; for Switzerland cf. the responses to the consultation concerning the amendment of Article 115 of the SCC:  https://www.bj.admin.ch/bj/de/home/gesellschaft/ gesetzgebung/archiv/sterbehilfe.html (last accessed 31 January 2022). 87 https://www.bj.admin.ch/bj/en/home/aktuell/news/2011/ref_2011-­ 06-­29.html (last accessed 31 January 2022). 86

Swiss Federal Act on Medicinal Products and Medical Devices of 15 December 2000 (SR 812.21). 89 Geneva Criminal Appeals and Revision Chamber, decision AARP/145/2020 dated 20 April 2020; see also: https://www.letemps.ch/ suisse/condamnation-vicepresident-dexit-suisse-romande-confirmeeappel and Decision Swiss Federal Court 09.12.2021, 6B_646/2020. (last accessed 31 January 2022). 88

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enjoy the same fundamental legal and ethical principles applied to other living persons.

67.4.1  Human research on corpses Current legal provisions

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Where total brain death has occurred, rules on the treatment of the corpse apply regarding the extent to which research may be performed on a deceased person being kept artificially ‘alive’. However, a search of national laws on the conditions or limits of performing research yields scant results.91 A browse through the European regulations produces similarly few answers. The conventions, additional protocols, directives and recommendations that address the topic of the corpse are fragmentary.92 The degree of legal uncertainty produced by such heterogeneous legislation is relatively high. Research helps to improve the definition of total brain death, the diagnoses and the therapies applied and promotes further development of medical knowledge, and thus helps other patients. Often the deceased patient offers his or her body for research purposes.

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Although the jurisprudence has confirmed the constitutionality of the concept of brain death, discussions on the key criteria of death have not yet been completely finished.90 Such discussions concern questions of whether the normative assessment of brain death is concordant with the actual death, which criteria and indications of death should be used to determine brain death, and whether provisions of national transplantation and human tissue acts can be transferred to other areas of the law. It became very clear during discussions about post-­mortal organ donation that a normative assessment was needed in determining complete brain death as the time of death. The fact that brain death occurring means that the persons biological death has occurred is normative. The signs of death play a central role. The clinical signs of death decide whether the removal of vital organs is permissible which causes the donor’s cardiovascular system to shut down. This means, once brain death has occurred, examinations that would lead to serious, fatal injuries in the living person can now be performed. Within this context, it must be noted that there is no international consensus on the criteria for determining death. The SAMS prescribes various observation periods in its guidelines on the determination of death with regard to organ transplantation and preparation of organ removals (SAMS 2017). The criteria to determine brain death may include, for example, proof of electrical inactivity of the brain (using electroencephalography), evoked potentials (i.e. stimulating the senses to evoke changes in the brain’s potential which can be read from the surface of the brain) and proof of cessation of intracerebral circulation (using Doppler or ultrasonic assessment, cerebral perfusion scintigraphy or the depiction of vessels (angiography)). Because of these varying criteria, it is possible that in some countries the time of death could be determined to have occurred at a point at which the person would have been regarded in another country as still alive. Another aspect must also be considered regarding non-­ heart-­beating donors (Grande and Pellegrini 2016) on one side, and the so-­called partial brain death or cortical death on the other side. Human research on dying patients and the deceased can be expected to produce new findings in this area. Those in an apallic state (i.e. patients who retain quantitative consciousness but whose qualitative consciousness, in contrast, cannot generally be recovered using current knowledge) or those who are anencephalous (i.e. a newborn whose head is marked by characteristic defects  – no cranium, cerebrum and meninges, possibly even no cerebellum, midbrain and diencephalons) are, as living persons, fully protected by the legal system. Despite the demand to examine these persons and thus to generate new findings, it must be taken into account at all times that they See the SAMS (Swiss Academy of Medical Science) (2017). Determination of Death with Regard to Organ Transplantation and Preparations for Organ Removal. Medical-­Ethical Guidelines, 16 May. Bern: SAMS: https://www.samw.ch/en/Publications/Medical-ethicalGuidelines.html (last accessed 31 January 2022).

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In Switzerland see the Swiss Federal Act on Research Involving Humans which entered into force on 1 January 2014. 92 The European Union Tissue and Cells Directives (EUTCD) are made up of three directives, the directive (2004/23/EC) that provides the framework legislation and two technical directives (2006/17/EC and 2006/86/ EC), which provide the detailed requirements of the EUTCD. Moreover, Directive 2015/565 regulates certain technical requirements for the coding of human tissues and cells, while Directive 2015/566 concerns the procedures for verifying the equivalent standards of quality and safety of imported tissues and cells. Finally, Directive 2010/45/EU of the European Parliament and of the Council of 7 July 2010 provides standards of quality and safety of human organs intended for transplantation. 91

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is primarily dependent upon the (assumed) will of the deceased and the attitudes of those authorised to care for the deceased. In general, the models for consideration are the solutions discussed during the preparation of the national transplantation acts: objection, declaration, information and models of narrow and extended consent. A closer look at the various models shows that they differ above all in the significance they accord to the self-­determination right of the person concerned and the subsidiary right to decide of those authorised to care for the corpse. The narrow or modified objection solutions, often favoured in medical practice as the right answer to a high demand for research on the corpse, is subject to constitutional concerns and needs to be considered in detail. If there is a lack of knowledge about this amongst the general public, it is likely that few will avail themselves of the right of objecting. In some senses, therefore, the deceased person is thus, while still alive, treated somewhat as an object of society or its interested branches. Furthermore, the objection solution encroaches upon the right not to have to deal with certain personal questions. The positive and negative freedoms of religion and philosophy are also of importance, and it must be remembered that not all religions are open to research on corpses (Box 67.13). The information and declaration solutions hardly differ in terms of breaching the post-­mortal self-­determination right and the rights of the next of kin to care for the deceased from the extended objection solution and are thus subject to criticism for the same reasons. Furthermore, there are some aspects of data protection that are in conflict with the declaration solution.

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‘erweiterte Zustimmungslösung’; in Germany its now called ‘Entscheidungslösung’)93 as a requirement for the removal of donated organs, tissues and cells. This position was in line with the consensus being sought at both a European and international stage of the Convention on Human Rights and Biomedicine, the corresponding Additional Protocol on Transplantation and Directive 2010/45/EU of the European Parliament and of the Council of 7  July 2010 on standards of quality and safety of human organs intended for transplantation. Nowadays, more and more countries are considering amending their laws on transplantation. To shorten the waiting period for an organ transplant, the Swiss Federal Council supports the principle of presumed consent in a broad sense which results in an opt-out solution. The Swiss Parliament decided in 2021 to introduce an opt-out solution for organ donation. In the absence of refusal, the removal of organs and tissues will be allowed. However, relatives will be consulted before any such removal if the will of the deceased is not clear. Moreover, they have the right to object to any removal. Furthermore, a new register will be created that will best keep track of persons’ refusals. However, a referendum against the new bill was successful. If there is not enough votes against the amendments, the amended law is expected to come into force in 2023 .94 In February 2020, the German Parliament rejected a Health Ministry proposal for introducing a system of presumed consent for organ donation95 and instead passed ‘The law to strengthen the willingness to make decisions on organ donation’, which will come into force on 1 March 2022.96 Some national criminal codes provide the corpse with protection from disruption of the right to peace as well as from malicious gossip and defamation. However, the usually fragmentary nature of both offences does not provide any real protection from unauthorised treatment of the corpse and consequently from impermissible research. The existing provisions do, however, mirror the increasing urgency for legislative action to provide more legal clarity and legal certainty. If no special national regulations exist, rules on the permissibility of performing research on the brain-­dead corpse must be deduced from general fundamental principles. In keeping with European provisions and the Convention on Human Rights and Biomedicine, it should be generally agreed that permission to perform brain diagnostics on a ‘fresh’ corpse

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German Transplantation Act, Sections 3 et seq.; Swiss Federal Transplan­ tation Act, Article 8. 94 https://www.bag.admin.ch/bag/de/home/medizin-und-forschung/ t r a n s p l a nt at i on s m e d i z i n / re c ht s e t z u n g s proj e kt e - i n - d e rtransplantationsmedizin/indirekter-gegenvorschlag-organspendeinitiative.html1 (last accessed 31 January 2022). 95 https://www.dw.com/en/german-­parliament-­explicit-­consent-­still-­ necessary-­from-­organ-­donors/a-­52022245; see also: http://www.drze. de/in-­focus/organ-­transplantation/legal-­aspects (last accessed 26 October 2020). 96 Federal Law Gazette Volume 2020 Part I No. 13, published in Bonn on 19 March 2020, p. 497 et seq. 93

Box 67.13  Charter of the Fundamental Rights of the European Union (2012/C 326/02). Article 1: Human dignity Human dignity is inviolable. It must be respected and protected. Article 3: Right to the integrity of the person 1. Everyone has the right to respect for his or her physical and mental integrity. 2. In the fields of medicine and biology, the following must be respected in particular: a. the free and informed consent of the person concerned, according to the procedures laid down by law; b. the prohibition of eugenic practices, in particular those aiming at the selection of persons; c. the prohibition on making the human body and its parts as such a source of financial gain; d. the prohibition of the reproductive cloning of human beings. Article 10: Freedom of thought, conscience and religion 1. Everyone has the right to freedom of thought, conscience and religion. This right includes freedom to change religion or belief and freedom, either alone or in community with others and in public or in private, to manifest religion or belief, in worship, teaching, practice and observance. ­ ccordance 2. The right to conscientious objection is recognised, in a with the national laws governing the exercise of this right.

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Box 67.14  Convention for the Protection of Human Rights and Fundamental Freedoms as amended by Protocols No. 11 and No. 14.

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Article 9: Freedom of thought, conscience and religion 1. Everyone has the right to freedom of thought, conscience and religion; this right includes freedom to change his religion or belief and freedom, either alone or in community with others and in public or private, to manifest his religion or belief, in worship, teaching, practice and observance. 2. Freedom to manifest one’s religion or beliefs shall be subject only to such limitations as are prescribed by law and are necessary in a democratic society in the interests of public safety, for the protection of public order, health or morals, or for the protection of the rights and freedoms of others.

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Article 14: Prohibition of discrimination The enjoyment of the rights and freedoms set forth in this Convention shall be secured without discrimination on any ground such as sex, race, colour, language, religion, political or other opinion, national or social origin, association with a national minority, property, birth or other status.

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Also being discussed at present are the permissibility and modalities of biobanks and databases. These are databases in which, among other things, the information and personal brain data produced by human research are stored. Such databases are an important resource in shedding light on the causes and mechanisms of different illnesses, especially those that are widespread among the general population. Data are sometimes collected specially for research purposes, sometimes they arise incidentally in medical or diagnostic connections. The information may be of value for biomedical research, in particular for epidemiology, and leads to progress in terms of the public good. Moreover, biobanks and databases are important sources of information for establishing the causes of diseases and could also represent a central support for monitoring and assuring quality in medicine. To this end, an examination may not be solely in the interests of the individual, but also in those of the larger collective. This poses questions of self-­ determination, solidarity, altruism and justice.97 In order to ensure that equal consideration is given to the interests of those involved, special rules should be drawn up on the preconditions and limits of recycling and collecting information gleaned from brain imaging. The central point of reference here is the right to self-­determination of the person from whom the information was generated. The effective consent of the person involved is needed not only to perform the research itself, but also to the related intention to save the information thus generated in order to make it later available for databank research

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Concerns remain that a declared objection will only be observed if the researcher is obliged to conduct further enquiry and that it will often prove impossible, or unreasonable given the amount of research it entails, to uncover the objection. This will lead to the danger of the (post-­mortal) interests of the person being ignored. If data generated by research are saved and transferred between databases and could lead to conclusions being drawn about surviving family members, there can be no reasonable doubt that the persons concerned should positively legitimise the procedure. Therefore Article 14 of European Recommendation CM/ Rec(2016)6 of the Committee of Ministers to member states on research on biological materials of human origin rightly declares that: 1. Biological materials should only be removed from the body of a deceased person for storage for future research with the consent or authorisation provided for by law. This consent or authorisation should have been preceded by appropriate information, including on the right to refuse. 2. Biological materials should not be removed for storage for future research activities if the deceased person is known to have objected to it. The model that takes these concerns best into account may be the consent solution, as self-­determination beyond death reinforces an individual’s set of values and thereby his or her interest in disposal over his or her own body. The fact that some national legislatures nevertheless decided in favour of the extended consent solution reflects a weighing of interests. Experience to date indicates that the narrow consent solution does not produce sufficient numbers of donors. This compromise, which seeks to balance the differing rights and interests, can be transferred with good reason to the field of research on corpses.

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Equality before the law is based on the concept of affording equal consideration to the subjective fundamental rights of all people – irrespective of sex, skin colour, culture, social origin, age, health and individual capabilities. Chapter III of the Charter of the Fundamental Rights of the European Union states some crucial principles concerning equality. Article 21 states: 1. Any discrimination based on any ground such as sex, race, colour, ethnic or social origin, genetic features, language, ­religion or belief, political or any other opinion, membership of a national minority, property, birth, disability, age or sexual orientation shall be prohibited. 2. Within the scope of application of the Treaties [establishing the European Community and on European Union] and without prejudice to any of their specific provisions, any discrimination on grounds of nationality shall be prohibited. The Convention for the Protection of Human Rights and Fundamental Freedoms also covers non-­discrimination in several articles (Box 67.14).

For more detail, see Directive 2004/23/EC of the European Parliament and of the Council of 31 March 2004 on setting standards of quality and safety for the donation, procurement, testing, processing, preservation, storage, and distribution of human tissues and cells.

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Article 7 – Confidentiality 1. Any information of a personal nature collected at the time of removal, storage or use of biological materials, or obtained through research, should be considered as confidential and treated according to the rules relating to the protection of private life. 2. Appropriate safeguards should be in place to ensure confidentiality at the time of removal, storage, use and, where appropriate, transfer of biological materials.

any, given by the person concerned, consent or authorisation to the proposed use should be sought and, to this end, reasonable efforts should be made to contact the person concerned. The wish of the person concerned not to be contacted should be observed. b. Where the attempt to contact the person concerned proves unsuccessful, these biological materials should only be used in the research project subject to an independent evaluation of the fulfilment of the following conditions: i. evidence is provided that reasonable efforts have been made to contact the person concerned; ii. the research addresses an important scientific interest and is in accordance with the principle of proportionality; iii. the aims of the research could not reasonably be achieved using biological materials for which consent or authorisation can be obtained; and iv. there is no evidence that the person concerned has expressly opposed such research use. 3. Any use of biological materials in an identifiable form should be justified in advance in the research protocol. 4. Non-­identifiable biological materials may be used in a research project provided that such use does not violate any restrictions defined by the person concerned before the materials have been rendered non-­identifiable and subject to authorisation provided for by law. 5. Biological materials from persons who, according to law, are not able to consent should only be used for research having the potential to produce, in the absence of direct benefit to the person concerned, benefit to other persons in the same age category or afflicted with the same disease or disorder or having the same condition, and if the aims of the research could not reasonably be achieved using biological materials from persons able to consent.

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Box 67.15  Chapter VI (use of biological materials in research projects) of European Recommendation CM/Rec(2016)6 of the Committee of Ministers to member states on research on biological materials of human origin. This Recommendation is a revision of Recommendation (2006)4 and takes into account new developments in the field of biobanking, such as the increasingly diverse origin of biological materials stored in collections, the difficulty to guarantee non-­identifiability of such samples, the increasing amount of research involving materials coming from different collections and the importance of research on biomaterials removed from persons not able to consent.98

experts and representatives of the patient or family members before the data is saved. Where there is no longer a viable connection between the data and the relevant persons, the protected interests of those persons are no longer affected. In order to protect the privacy of the deceased and his or her next of kin, the personal details must be as far as possible encrypted. The code itself and the encoded brain data have to be separately stored and administered. The bio-­and databanks must ensure that they comply with all the legal requirements of data protection. If an exception arises, it is necessary to consult the ethic commissions and abide by their decision.

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purposes (Box 67.15). This also applies to the ‘multiuse’ of data originally collected for medical purposes. There remains a grey area. In the past, research-­related use of personal data stored in bio-­and databanks was often performed without informed consent and no close relatives or persons of comparable status could be found in many cases. To ensure that these bio-­and databanks remain available, the process of obtaining informed consent must not be unnecessarily complicated. Situations similar to emergencies may arise, when attention must be paid to render the data, where possible, anonymous and to secure the consent of an independent commission comprising

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Article 13 – Right to withdraw consent or authorisation 1. When a person has provided consent to storage of identifiable biological materials for future research, the person should, without being subject to any form of discrimination, in particular regarding the right to medical care, retain the right to withdraw consent at any time, and, where possible, should also be able to alter the scope of that consent. When identifiable biological materials are stored for research purposes only, the person who has withdrawn consent should have the right to have, in the manner foreseen by law, the materials and associated data either destroyed or rendered non-­identifiable. The person who is considering withdrawing consent should be made aware of any limitations on withdrawal of his or her biological materials. [. . .] Article 21 – General rule 1. Biological materials should only be used in a research project if the latter is within the scope of the consentor authorisation given by the person concerned. 2. a. If the proposed use of identifiable biological materials in a research project is not within the scope of prior consent or authorisation, if For more information, see https://www.coe.int/en/web/bioethics/ biobanks/-/asset_publisher/lcb5Z6eMEwYb/content/research-onbiological-materials-of-human-origin-new-recommendations-tomember-stat-1?inheritRedirect=false (last accessed 31 January 2022).

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Article 22 – Independent review 1. Research should only be undertaken if the research project has been subject to an independent examination of its scientific merit, including assessment of the importance of the aim of the research, and verification of its ethical acceptability. The law may additionally require approval by a competent body. 2. Member states should apply the principles concerning ethics committees contained in Chapter III of the Additional Protocol concerning biomedical research (CETS No. 195) to the review of research within the scope of this recommendation. 3. Review procedures may be adapted to the nature of the research and the extent to which the persons concerned could be identified from their biological materials.

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In this context, the World Medical Association (WMA) adopted in 2016 the Declaration of Taipei99, which provides general definitions and ethical considerations regarding health databases and biobanks, as a complement to the Helsinki Declaration (Sprumont et al. 2016; Chassang and Rial-­Sebbag 2018). In particular, Article 24 of the Declaration of Taipei urges relevant authorities to formulate policies and laws that protect health data and biological material on the basis of the principles set forth in this document.

Article 21: Prohibition of financial gain The human body and its parts shall not, as such, give rise to financial gain. Article 22: Disposal of a removed part of the human body When in the course of an intervention any part of a human body is removed, it may be stored and used for a purpose other than that for which it was removed, only if this is done in conformity with appropriate information and consent procedures.

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67.5  Legal and ethical aspects concerning the handling of corpses and the display of human remains

Box 67.16  Chapter VII of the Convention on Human Rights and Biomedicine100: Prohibition of financial gain and disposal of a part of the human body.

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67.5.1  The dead body

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Death shows that there is a limit to everything concerning our life in this world. In the past, human remains were removed to the cemetery, and only a few corpses were used for research or some well-­preserved parts of the body – or in exceptional cases whole bodies  – were displayed in anatomical museums. The public has gradually found itself faced with issues of increasing interest regarding dead bodies and their anatomy, so it is hardly surprising to see a wide range of unresolved questions arising. The most important of these is whether and to what extent anatomical specimens of human origin should be accessible to public view, not only to doctors and medical students, but also to laypersons (Tag  2000b). The more detailed the bioethical, theological and social questions pertaining to the exhibition of human remains become, the clearer the need is to consider the legal ramifications. Nevertheless, it would be a mistake to believe that clear answers exist. The reasons for this are manifold. In the past, in particular, the body was primarily of interest to pathology, anatomy and, in certain ways, the fine arts. Outside this narrow range of interests, the treatment of bodies was focused on mourning and the precautions needed to protect the living from health risks. This changed when medicine and biochemistry discovered the corpse for their purposes. Up to recently, the discussion concerning the proper use of a corpse has focused primarily on organ and tissue transplants; the major issues involved here have been the debate on brain death and on finding solutions to problems of consent. Despite the Convention on Human Rights and Biomedicine (Box 67.16), this controversy has still not been fully resolved. To look to the law for clarity on this issue involves looking for guidelines in the drafting of laws (as well as the penalties to ensure their enforcement); it is evident that only a loose web of norms exists, which is by no means definite or definitive. Only a

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few anatomical acts exist, and they usually address very few questions regarding the adequate treatment of bodies. Rules about the treatment of bodies often have to be derived from fundamental general principles. However, this is not necessarily a disadvantage. In many areas of research and applied sciences, there are shifts in values and opinions, showing that our legal systems require a certain degree of flexibility to react to changes of this nature.

Legal considerations on this issue must begin by exploring the legal status of a corpse. Two fundamental questions must be examined: what obligations are there and to whom do they apply? What amount of legal protection is the body entitled to? When we know the answers to these, perhaps we will be able to find answers to other questions. Rights of disposition over a corpse presuppose a legal link to that corpse, which may seem simple at first glance. However, upon closer assessment it proves to be highly complex because written law has only recently begun to regulate the legal status of corpses. To understand the problems involved it is necessary to understand the legal link between a living person and his or her corpse. A living person is granted a unique legal protection on the basis of the unity of soul, spirit and body. This distinctive ­feature attributes the human being the right to dignity and to privacy. Death fractures the human life and throws the unambiguous nature of the former legal assessment into question. If a person’s life is ended, his or her remains are what firstly determine the nature of the legal object now under consideration. Usually, the lifeless body starts to decompose, which indicates that one’s m ­ ortal remains have come to be viewed as an object. This ­transformation can be seen in the area of post-­mortem organ and tissue donation and the broad range of uses for the Convention for the Protection of Human Rights and Dignity of the Human Being with regard to the Application of Biology and Medicine (Convention on Human Rights and Biomedicine).

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In principle, the above-­mentioned Additional Protocol, as well as many national regulations, accepts the appropriation and medical-­related handling of corpses or parts of them. This is reflected in the ever-­growing spectrum of the commercial and non-­commercial use of bodies and/or parts of them. All over the world, there are many anatomical museums and exhibitions – for example the so-­called touring ‘body-­world’ exhibition or the traditional anatomical museums (e.g. the Wellcome Museum of Anatomy and Pathology, London) and other displays of human remains (e.g. the skeleton of Jeremy Bentham, the famous philosopher and lawyer, which is preserved at University College, London). In fact, it can be said that the right to donate one’s body comes from the tradition of donating bodies to anatomical institutions. Such uses of dead bodies illustrate the fact that a corpse at some point becomes more of an object than a person. However, there is a huge discussion about the legal status of the corpse and the question whether human remains can be categorised as displaying certain characteristics of property or possession. ­ Although the deceased is no longer considered a person in his or her own right, some legal rights to which they were entitled during their lifetime remain for a defined period of time – depending on their circumstances of death (where, why, when, how, who, etc.). The continuity of human dignity beyond the grave means that, while corpses and human remains are objects, there are a lot of important constraints concerning an adequate handling of a human body. This provisional result seems to be consistent with the European Convention on Human Rights and Biomedicine and the Additional Protocol concerning Transplantation of Organs and Tissues of Human Origin (Box 67.18). Both these legal norms prohibit the commercial use of human remains, but they do not outlaw the right of ownership concerning the body. In addition to this, Directive 2004/23/EC of the European Parliament and of the Council of 31 March 2004 on setting standards of quality and safety for the donation, procurement, testing, processing preservation, storage and distribution of human tissues and cells is interesting (Box 67.19). These regulations provide that if the procurement and the processing of human tissues are regulated by law, then human matter can become third party property. If so, there is no reason why human remains should not be classified as a special form of property.

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corpse in human research. As the law of succession shows, in the case of death, all one’s belongings and possessions must pass into other hands. This makes sense when we recognise that the corpse has to be separate from its former state and become an object. Under common law, the starting principle of dead bodies is that ‘the body belongs to no one’ (corpus nullius in bonis), but recent developments show that we have to differentiate between status (whether an object or not) and property rights. From the moment death occurs, the body is ownerless. However, in Germany, Switzerland, Austria, the UK and several other European countries, the legal practice is of the opinion that every capable person is allowed to donate his or her future dead body to a department of anatomy or pathology and for organ, tissue and cell transplantation. These principles can be found in the Additional Protocol to the Convention on Human Rights and Biomedicine concerning Transplantation of Organs and Tissues of Human Origin of 2002 (Box 67.17).101 In Switzerland, Article 36 of the Human Research Act specifically provides that research may be carried out in deceased persons if, before their death, the persons concerned consented to the use of their body for research purposes. The applicable guidelines of the SAMS also provide for a clear ethical guidance regarding body donation.

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Box 67.17  Chapter IV of the Additional Protocol to the Convention on Human Rights and Biomedicine concerning Transplantation of Organs and Tissues of Human Origin: Organ and tissue removal from deceased persons.

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Article 16: Certification of death Organs or tissues shall not be removed from the body of a deceased person unless that person has been certified dead in accordance with the law. The doctors certifying the death of a person shall not be the same doctors who participate directly in removal of organs or tissues from the deceased person, or subsequent transplantation procedures, or having responsibilities for the care of potential organ or tissue recipients.

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Article 17: Consent and authorisation Organs or tissues shall not be removed from the body of a deceased person unless consent or authorisation required by law has been obtained. The removal shall not be carried out if the deceased person had objected to it. Article 18: Respect for the human body During removal the human body must be treated with respect and all reasonable measures shall be taken to restore the appearance of the corpse. Article 19: Promotion of donation Parties shall take all appropriate measures to promote the ­donation of organs and tissues.

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https://rm.coe.int/1680081562 (last accessed 31 January 2022).

67.5.2 Consent or authorisation required by law Defining the status of corpses purely in terms of property laws, however, is not convincing when addressing the issue of ownership of a human corpse. On the one hand, this would result in unlimited ownership and marketing rights over the corpse, which would not be acceptable when considering that the mortal remains were once a human being. On the other hand, property ownership carries with it the right of owners to do with their

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Article 13: Consent 1. The procurement of human tissues or cells shall be authorised only after all mandatory consent or authorisation requirements in force in the Member State concerned have been met. 2. Member States shall, in keeping with their national legislation, take all necessary measures to ensure that donors, their relatives or any persons granting authorisation on behalf of the donors are provided with all appropriate information as referred to in the Annex.

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Additional Protocol Chapter VI, Article 22 – Prohibition of organ and tissue trafficking Organ and tissue trafficking shall be prohibited.

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Additional Protocol Chapter VI, Article 21 – Prohibition of financial gain 1. The human body and its parts shall not, as such, give rise to financial gain or comparable advantage. The aforementioned provision shall not prevent payments which do not constitute a financial gain or a comparable advantage, in particular: • compensation of living donors for loss of earnings and any other justifiable expenses caused by the removal or by the related medical examinations; • payment of a justifiable fee for legitimate medical or related technical services rendered in connection with transplantation; • compensation in case of undue damage resulting from the removal of organs or tissues from living persons. 2. Advertising the need for, or availability of, organs or tissues, with a view to offering or seeking financial gain or comparable advantage, shall be prohibited.

Article 12: Principles governing tissue and cell donation 1. Member States shall endeavour to ensure voluntary and unpaid donations of tissues and cells. Donors may receive compensation, which is strictly limited to making good the expenses and inconveniences related to the donation. In that case, Member States define the conditions under which compensation may be granted. Member States shall report to the Commission on these measures before 7 April 2006 and thereafter every three years. On the basis of these reports the Commission shall inform the European Parliament and the Council of any necessary further measures it intends to take at Community level. 2. Member States shall take all necessary measures to ensure that any promotion and publicity activities in support of the donation of human tissues and cells comply with guidelines or legislative provisions laid down by the Member States. Such guidelines or legislative provisions shall include appropriate restrictions or prohibitions on advertising the need for, or availability of, human tissues and cells with a view to offering or seeking financial gain or comparable advantage. Member States shall endeavour to ensure that the procurement of tissues and cells as such is carried out on a non-­profit basis.

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Convention Chapter VII, Article 21 – Prohibition of financial gain The human body and its parts shall not, as such, give rise to financial gain.

Box 67.19  Directive 2004/23/EC of the European Parliament and of the Council of 31 March 2004 on setting standards of quality and safety for the donation, procurement, testing, processing, preservation, storage and distribution of human tissues and cells.

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Box 67.18  Chapter VII of the Convention on Human Rights and Biomedicine: Prohibition of financial gain and disposal of a part of the human body; and Chapter VI of the Additional Protocol: Prohibition of financial gain.

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property as they see fit: selling it, for instance, or excluding others from having any influence so long as this violates neither the law nor the rights of third parties. The law also allows for the purchase of movable goods in good faith provided they have been neither lost nor stolen. If all of these legal provisions were applied to human corpses, the consequences would be unacceptable. For this reason, some regulations argue in favour of extending the personal rights of privacy and against the unlimited application of property law to human remains.102 In light of this, decisions made by the deceased regarding the type and location of his or her funeral become important and the Additional Protocol to the Convention on Human Rights and Biomedicine concerning Transplantation of Organs and Tissues of Human Origin makes organ donation dependent primarily upon the consent of the deceased. If no consent – written or otherwise – has been given, the consent of the next of kin or that person whom the deceased has authorised is required. The presumed will of the deceased, however, must be taken into consideration – insofar as it is known. The basis for decisions such as these is implicated in For example, Swiss Federal Court BGer 1P.373/2002 /bie, decision 4 July 2003; German Federal Constitutional Court ‘Mephisto’, decision 24 February 1971, 1 BvR 435/68 – BVerfGE 30, 173; German Federal Court, decision 22 April 2005, 2 StR 310/04  – BGHSt 50, 80 et  seq. (Cannibal of Rothenburg).

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the Charter of Fundamental Rights of the European Union and takes the principles of the Convention on Human Rights and Biomedicine of the Council of Europe fully into account. Legal constraints regarding proper use of human remains are limited to a few selective statements. In some countries such as Germany and Switzerland, the authority to regulate dissection and body donation to anatomical institutions does not fall under federal jurisdiction. For this reason, the individual states not only have their own funeral laws, but also their own laws in governing dissection – or they may have no laws at all on this issue. In such a case, the Act on organ and tissue donation, removal and transplantation (e.g. German Transplant Act) is applicable, but does not necessarily cover all relevant aspects. Nevertheless, the main legal principle involved is clear: within the confines of current law every individual has the right to determine what will be done with his or her mortal remains. If the deceased made no use of that right while alive, then it is vested in the next of kin or, as the case may be, vested in the state (e.g. in the cases of so-­called ‘presumed consent solutions’). However, the next of kin are not free to decide as they like. The post-­mortal

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A well-­known sentence is often found inscribed above the portals of anatomical institutions: ‘Here (is the place where) death serves life’. This statement bears a new and important meaning if corpses are unquestioningly used for human research. In some older displays, in particular, there are specimens that were not voluntarily donated. What should we do with these human remains? Are we obliged to separate and to bury them as regulated by their native conventions and customs?103 Or is it permissible to continue to keep these specimens without restriction in the museum? Is it allowed to do human research with these specimens or to use them as a basis for medical products? (See Cheney  2007; Keller 2008; Madof 2010). If we cannot rule out the possibility that the human remains derive from individuals, who died a violent death under totalitarian regimes, then we have to search for adequate solutions. In most countries, there are no answers written by law, and those that do exist often only cover the ethical minimum. Ethics, as the general conviction of the right, the true and the socially acceptable, is more far-­reaching than written and unwritten rules. Law and ethics complement one other and need each other. Critics who denounce the handling of corpses in research and the exhibition of conserved bodies and skeletons as unacceptable, regardless of the decision of a body donor, generally back up their arguments by referring to the right to care for the corpse and to the feelings of reverence on the part of both survivors and the general public. The right to care for a corpse is a right reserved for the next of kin. Its purpose is to ensure that all laws applying to the care of corpses will be observed and to express the wishes of the dead person as expressed during their lifetime. In principle, survivors have to respect the decisions of the deceased and are not allowed to do whatever they see fit with the corpse. Nevertheless, a lot of different opinions exist on how to handle corpses and anatomical specimens. Human remains in anatomical museums that were provided without consent and donation, in most cases are relics of the past. Therefore, we have to solve primarily an ethical question: could and should we address yesterday’s injustices today? One solution would be the ‘fruit of the poisonous tree’ doctrine, where the fruits of the prohibited tree would not be harvested, that is the specimens should

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be removed as quickly as possible. However, this procedure is not enforceable by law so as there is no absolute protection of the corpse. To decide on an acceptable procedure means discussing several items as they are written in the 2003 Stuttgarter Recommendations (Arbeitskreis Menschliche Präparate in Sammlungen 2003). First, the fading memory of the deceased. There are differences if the human remains are part of an anonymous person who died a very long time ago104 or if they belong to a person who still has next of kin alive. The circumstances of the death could be another essential consideration. If a person lost his or her life solely because of his or her race, view of life or political reasons through violent measures, this injustice affects more than just the dignity of an individual.105 It is important for society that anatomical specimens deriving from such violent measures should not be used in human research or displayed in museums (Körtner 2001). The methods of display and preservation procedure are also essential to consider. In anatomy, the principle of anonymity must always be followed, with any descriptions focusing on anatomical information.

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personal rights dictate that they must take the wishes or the presumed wishes of the deceased into consideration. In conclusion, it is clear that the traditional use of a corpse has generally consisted of interring the dead, although our legal system does have general provisions for other uses of corpses.

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See, for example, the discussion that has taken place since years about the display of the skeleton of the ‘Irish giant’ in the Hunterian Museum in the Royal College of Surgeons, London (Doyal and Muinzer 2011; Nash 2018).

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Aebi-­Müller, R., Fellmann, W., Gächter, T. et al. (2016). Arztrecht. Bern: Stämpfli. Arbeitskreis Menschliche Präparate in Sammlungen (Working Group on Human Remains in Collections) (2003). Stuttgarter Recommendations on the Treatment of Human Remains in Collections, Museums and Public Places, pp. 378–383. Deutsches Ärzteblatt, August. Heft Nr. 8 (https:// wissenschaftliche-sammlungen.de/de/service-material/materialien/ stuttgarter-empfehlungen-zum-umgang-mit-praeparaten-aus-menschlichem-gewebe-sammlungen-museen-und-oeffentlichen-raeumen2003)

See, for example, the Swiss Federal Act on Research Involving Humans, which states that human research with specimens that are older than 70 are allowed without consent. However, if the next of kin do not allow this research it must not be made (Article 36, paragraph 4). Human Tissue Act 2004, UK, Section 1 states: ‘Authorisation of activities for scheduled purposes (1) The following activities shall be lawful if done with appropriate consent . . .’ This does not apply to a body if – (a) it has been imported, or (b) it is the body of a person who died before the day on which this section comes into force and at least one hundred years have elapsed since the date of the person’s death (see subsection (5)). 105 German Federal Court, 15 March 1994, 1 StR 179/93, Neue Juristische Wochenschrift 1994, 1421 et seq. 104

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Fellmann, W. (2007). Arzt und das Rechtsverhältnis zum Patienten. In: M. Kuhn and T. Poledna (eds.), Arztrecht in der Praxis, 2nd edn, pp. 103–231. Zurich: Schulthess. Gächter, T. and Rütsche, B. (2018). Gesundheitsrecht – Ein Grundriss für Studium und Praxis, 4th edn. Basel: Helbing Lichtenhahn Verlag. Gardiner, D. (2016). How the UK overcame the ethical, legal and professional challenges in donation after circulatory death. QUT Law Review 16 (1): 125–134. Gardiner, D., and Sparrow, R. (2010). Not dead yet: Controlled non-­ heart-­beating organ donation, consent, and the dead donor rule. Cambridge Quarterly of Healthcare Ethics 19 (1): 17–26. Gavan, S., Thompson, A. and Payne, K. (2018). The economic case for precision medicine. Expert Review of Precision Medicine and Drug Development 3 (1): 1–9. German Ethics Council (2006). Selbstbestimmung und Fürsorge am Lebensende. https://www.ethikrat.org/fileadmin/Publikationen/ Stellungnahmen/Archiv/Stellungnahme_Selbstbestimmung_und_ Fuersorge_am_Lebensende.pdf (last accessed 31 January 2022). Grande, A. and Aseni, P. (2016). Management of hemodynamic and metabolic impairments in heart-­ beating donors. In: P. Aseni, A. Grande and L. De Carlis (eds.), Multiorgan Procurement for Transplantation  – A Guide to Surgical Technique and Management, pp. 53–67. Cham: Springer International Publishing AG. Grande, A.M. and Pellegrini, C. (2016). Non-­Heart-­Beating Donors. In: Aseni, P., Grande, A., and De Carlis, L. (eds.), Multiorgan Procurement for Transplantation. Cham: Springer, pp. 69–77 doi: 10.14503/ THIJ-16-6152. Gross, D., Kaiser, S. and Tag, B. (eds.) (2016). Leben jenseits des Todes? – Transmortalität unter besonderer Berücksichtigung der Organspende. Frankfurt/New York: Campus Verlag. Gross, D., Tag, B. and Schweikardt, C. (eds.) (2011). Who Wants to Live Forever?  – Postmoderne Formen des Weiterwirkens nach dem Tod. Frankfurt/New York: Campus Verlag. Hirsch, H.J. (1987). Behandlungsabbruch und Sterbehilfe. In: Festschrift für Karl Lackner, pp. 597–620. Berlin: Walter de Gruyter. Hirsig-­Vouilloz, M. (2017). La responsabilité du médecin, Aspects de droit civil, pénal et administratif. Bern: Staempfli Verlag AG. Hüsing, B., Hartig, J., Bührlen, B. et al. (2008). Individualised Medicine and Health Care System. TAB Report No. 126. Berlin: Office of Technology Assessment at the German Bundestag. Hüsing, B., Jähnke, L. and Tag, B. (2006). Impact Assessment of Neuroimaging: final report. Zurich: Vdf Hochschulverlag. Keller, M. (2008). Ausgeschlachtet, Die menschliche Leiche als Rohstoff. Berlin: Ullstein Buchverlage. Kłak, A., Paradowska-­Gorycka, A, Kwiatkowska, B. and Raciborsk, F. (2016). Personalized medicine in rheumatology. Reumatologia 54 (4): 177–186. Körtner, U.H.J. (2001). Der unbezähmbare Tod. In: Wetz, F.J. and Tag, B. (eds.), Schöne neue Körperwelten. Der Streit um die Ausstellung, pp. 241–265. Stuttgart: Klett-­Cotta. Leipe, J., Schramm, M.A., Grunke, M. et al. (2011). Interleukin 22 serum levels are associated with radiographic progression in rheumatoid arthritis. Annals of the Rheumatic Diseases 70 (8): 1453–1457. Lenk, C., Duttge, G. and Fangerau, H. (eds.) (2014). Handbuch Ethik und Recht der Forschung am Menschen. Göttingen/Ulm: Springer.

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Aseni, P., Grande, A. and De Carlis, L. (eds.) (2016). Multiorgan Procurement for Transplantation – A Guide to Surgical Technique and Management. Cham: Springer International Publishing AG. Barnikol, M. (2018). Die Regelung der Suizidbeihilfe in den neuen SAMW-­Richtlinien. Schweizerische Ärztezeitung 99 (41): 1392–1396. Becchi, P. (2009). Piergiorgio Welby–Therapieverweigerung oder Euthanasie? Bioethica Forum 2 (1): 43–45. Blechta, G., Colatrella, P., Rüedi, H. and Staffelbach, D. (eds.) (2020). Basel Commentary on the Swiss Federal Act on Health Insurance and the Swiss Federal Act on the Surveillance of Health Insurance. Basel: Helbing and Lichtenhahn. Brittain, H., Scott, R. and Thomas, E. (2017). The rise of the genome and personalised medicine. Clinical Medicine 17 (6): 545–551. Büchler, A. and Michel, M. (2020). Medizin  – Mensch  – Recht, Eine Einführung in das Medizinrecht der Schweiz, 2nd edn. Zurich: Schulthess. Budin-­Ljøsne, I., Teare, H.J.A., Kaye, J. et al. (2017). Dynamic consent: A potential solution to some of the challenges of modern biomedical research. BMC Medical Ethics 18: 4. Califf, R.M. (2018). Biomarker definitions and their applications. Experimental Biology and Medicine 243 (3): 213–221. Chassang, G. and Rial-­Sebbag, E. (2018). Research Biobanks and Health Databases: The WMA Declaration of Taipei, Added Value to European Legislation (Soft and Hard Law). European Journal of Health Law 15 November, 25 (5): 501–516. Cheney, A. (2007). Bodybrokers, Inside America’s Underground Trade in Human Remains. New York: Broadway Books. Christinat, R. (2019). Le procès en responsabilité civile médicale, Mise en œuvre en procédures civile et administrative. Basel/Neuchâtel: Helbing Lichtenhahn – Faculté de droit de l’Université de Neuchâtel. Deutsch, E. and Spickhoff, A. (2014). Medizinrecht. Arztrecht, Arzneimittelrecht, Medizinproduktrecht und Transfusionsrecht, 7th edn. Berlin: Springer. Doyal, L. and Muinzer, T. (2011). Should the skeleton of ‘the Irish giant’ be buried at sea? British Medical Journal 343 (7597). Dankar, F., Gergely, M., Malin, B. et al. (2020). Dynamic-­informed consent: A potential solution for ethical dilemmas in population sequencing initiatives. Computational and Structural Biotechnology Journal 18: 913–921. Eugster, G. (2016). Krankenversicherung. In: U. Meyer (ed.), Schweizerisches Bundesverwaltungsrecht (SBVR), Soziale Sicherheit, 3rd edn, pp. 399–823. Basel: Helbing and Lichtenhahn. Eugster, G. (2009). Überarztung aus juristischer Sicht. In: T. Gächter and M. Schwendener (eds.), Rechtsfragen zum Krankheitsbegriff, pp. 97–145. Zurich: Schulthess. Federal Department of Justice and Police (2006) Euthanasia and Palliative Care – Does the Confederation need to act?, https://www.bj.admin.ch/ ejpd/en/home/latest-news/news/2006/2006-05-311.html (last accessed 31 January 2022). Federal Department of Justice and Police (2007). Supplementary Report on the Report: Euthanasia and Palliative Care – Does the Confederation Need to Act?, Ergänzungsbericht zum Bericht “Sterbehilfe und Palliativmediz” in https://www.bj.admin.ch/bj/de/home/gesellschaft/ gesetzgebung/archiv/sterbehilfe.html (last accessed on 31 January 2022).

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Schmitz, G., Endres, S. and Götte, D. (eds.) (2008). Biomarker–Bedeutung für den medizinischen Fortschritt und Nutzenbewertung. Stuttgart: Schattauer Verlag. Schwarzenegger, C. (2018) Verfassungsrechtliche Aspekte der Sterbehilfe und die Konsequenzen für das Strafrecht. In: Schwarzenegger, C. and Ida, M. (eds.), Autonomie am Lebensende  – Kultur und Recht, Die aktuelle Diskussion in der Schweiz und in Japan, pp. 9–23. Zurich/St. Gallen: Dike. Schwarzenegger, C. and Ida, M. (eds.) (2018). Autonomie am Lebensende – Kultur und Recht, Die aktuelle Diskussion in der Schweiz und in Japan. Zurich/St. Gallen: Dike. Spickhoff, A. (2018). Medizinrecht, Kommentar, 3rd edn. Munich: C.H. Beck. Sprumont, D., Joye, C. and Pilottin, A. (2016). Biobanques: il est urgent d’investir dans une loi fédérale. Bulletin des Médecins suisses 97 (48):1693–1695. Staempfli Haenni, A. (2017). Annotations Article 321bis of the Swiss Criminal Code. In: A. Macaluso and L. Moreillon (eds.), ‘Romand’ Commentary on the Swiss Criminal Code, vol. II, 1st edn. Basel: Helbing Lichtenhahn. Swiss Federal Council (2009). Dispatch Swiss Federal Act on Research Involving Humans, 21 October 2009, BBl 2009 8045. Swiss Federal Council (2002). Dispatch Swiss Federal Act on Human Genetic Testing (HGTA), 11 September 2002, BBl 2002 7361. Swiss Federal Office of Public Health (FOPH) (2011). Operationalisierung der Begriffe Wirksamkeit, Zweckmässigkeit und Wirtschaftlichkeit nicht gefunden. FOPH Working Paper, 21 July 2011. Swiss Medical Board (2009). Beurteilung medizinischer Verfahren, Methodischer Ansatz. Zurich: Swiss Medical Board. Swiss National Advisory Commission on Biomedical Ethics (NEKCNE) (2005). Assisted Suicide. Report No. 9/2005. Zurich: Swiss Medical Board nicht gefunden. Tag, B. (2000a). Der Körperverletzungstatbestand im Spannungsfeld zwis­ chen Patientenautonomie und lex artis. Berlin: Springer. Tag, B. (2000b). Legal considerations regarding body donations, plastination and human dignity. In: G. von Hagens (ed.), Anatomy Art, Fascination Beneath the Surface, Catalogue. Heidelberg: Körperwelten. Tag, B. (2002). Ausstellungen zur Anatomie: ‘Hier dient der Tod dem Leben’. Deutsches Ärzteblatt 99 (15): A-­1001. Tag, B. (2004). Die Verschwiegenheit des Arztes im Spiegel des Strafgesetzbuches und der Strafprozessordnung des Kantons Zürich. Schweizerische Zeitschrift für Strafrecht 122 (1): 1–20. Tag, B. (2006a). Legal relevance of neuroimaging in brain research. In: B. Hüsing, L. Jäncke and B. Tag (eds.), Impact Assessment of Neuro­ imaging, Final Report, pp. 156–173. Zurich: Vdf Hochschulverlag. Tag, B. (2006b). Neuroimaging on brain dead persons. In: B. Hüsing, L. Jäncke, and B. Tag (eds.), Impact Assessment of Neuroimaging, Final Report, pp. 174–190. Zurich: Vdf Hochschulverlag. Tag, B. (2010). Sterbehilfe–betrachtet im Lichte des Strafrechts, Vom Recht auf einen menschenwürdigen Tod oder: darf ich sterben, wann ich will? In: T. Fuchs, A. Kruseand G. Schwarzkopf(eds.), Menschenbild und Menschenwürde am Ende des Lebens, pp. 153–179. Heidelberg: Universitätsverlag WINTER. Tag, B. (2012a). Medizin und Recht. In: B. Madea, F. Mußhoff and B. Tag (eds.), Kurzlehrbuch Rechtsmedizin, pp. 15–79. Bern: Hogrefe.

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68 | Personal Injury Assessment Handbook of Forensic Medicine, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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Personal Injury Assessment

in terms of the social and forensic importance attributed to it, and today it is a specific area of legal medical and forensic expertise, often included in clinical forensic medicine, a branch of forensic medicine that is growing in importance. Forensic medical experts are increasingly called upon to examine and pronounce upon a range of situations, sometimes very complex, related to questions of personal injury in the area of criminal, civil, labour and administrative law, amongst others. This type of expert activity has particularities related to the legal system of the nation in question, and methods and procedures may vary from country to country as a result of legal specificities and different forensic systems. However, despite these specificities, the expert evaluation remains a medical act, which means that the methodology used needs to be technical and scientifically grounded, otherwise it runs the risk of becoming abstract and fictitious, which does not serve the interests of science and justice. What is at stake is nothing less than the need to grant space to doctors so that they can define what may or may not be assessed, and select the necessary tools, without having to use instruments that are scientifically inappropriate (Magalhães and Pinto da Costa 2007). In fact, clinical forensic medicine is currently facing ever more complex demands as a means of providing scientific evidence and is a rapidly expanding area. This means that its materials and methods are constantly having to adapt not only to new technologies and scientific discoveries but also to changes taking place in the sociocultural and legal spheres. The methods used in expert personal injury evaluation thus need to be defined by a body of medical experts specialised in this area.

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The very first kind of legal medicine practised in Mesopotamia over 4000 years ago involved the assessment and reparation of personal injury caused by third parties (Margeat  1988). The Nippur Tablets (2050 BC), Hammurabi Code (1750 BC), Talion Law, and Mishnah and Lex Aquilia are just some of the historical examples of legislation that contains references to methods of assessing and repairing personal injury (Geerts  1962). In fact, damage to bodily integrity has always been considered particularly important, requiring a form of punishment that corresponds to the social and individual suffering incurred. Over time, this punitive reaction was joined by another concern, namely to repair victims for the injury suffered; indeed, these two forms of reaction (punishment and reparation) only became completely autonomous with the appearance of the modern nation state. Questions such as whether or not the personal injury caused by third parties should be punished and/or repaired, how and with what means it should be done and who is responsible for implementing it, and for assessing and evaluating the consequences, have solicited very different responses throughout history in accordance with the type of society involved and the sociocultural values and principles underpinning its respective legal system (Magalhães  1998). The development of the welfare state also opened the way to other means of assessing personal injury, particularly when non-­traumatic in origin, in order to grant particular benefits and advantages to handicapped citizens. This whole matter has evolved, not only scientifically and technically, but also

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The general aim of personal injury assessment is to define – in technical and scientific terms, and from a medical–biological perspective and within a particular legal framework  – any temporary or permanent injury or damage liable to prosecution under criminal law and/or reparation, or which could entail certain benefits (such as tax or social benefits) (Magalhães et al. 2010c). As regards injury caused by third parties, the objective of reparation has evolved over time, moving away from mere retribution for the damage inflicted towards a greater focus on the rehabilitation and reintegration of the victim, whatever the type or aetiology of the event. This evolution has occurred naturally, not only as a result of policies aiming at the protection and promotion of human rights but also for economic reasons, given the socioeconomic costs of permanent consequences of trauma and diseases (i.e. by focusing on the victim’s rehabilitation and reintegration, it aims to promote a faster return to active life and, where possible, productive activity). This principle of reparation applies to all kinds of traumas and pathologies; hence, it makes no sense, at least from the scientific point of view, for there to be different forms of assessment and reparation in accordance with the facts that produced the injury (Hamonet and Magalhães 2001). However, since the early 20th century, almost all countries have proclaimed their agreement with universal human rights, recognising citizens’ rights to health, safety and integration (non-­exclusion), based on the fundamental right of equal treatment for all. Hence, the legal systems of these countries usually provide means of reparation, with a view to the social and professional reinsertion of their citizens. In more severe cases (where there are serious disturbances to the person’s functioning, activities and participation), the main aim of the assessment and reparation process should be to help the victim to return to the kind of life he or she had before the event, approached in a way that is not standardised but rather adapted to the particularities of their status and situation. Thus, the recoverable losses should not only be limited to anatomical–physiological sequelae, but should also include the multiple consequences of the event upon the victim’s daily activities, affective and family life, and professional situation. The essential acts of life are not limited to satisfying physiological needs; people have other aspirations, such as the need to communicate, acquire knowledge and engage in pastimes and leisure activities, all of which should be taken into account. The objective of all personal injury assessment and reparation procedures should be to try to reduce the damage incurred with a view to improving the victim’s physical, mental and social well-­being, reintegration into family, social and professional life, and general quality of life (Hamonet and Magalhães  2001). In other words, injury assessment in such cases should proceed in a way that is fair and appropriate to the victim’s real needs, taking into account the two fundamental principles enshrined in most legal systems of the world: that everyone fully enjoys the same rights, and in the case of personal injury, the aim should be to return the victim to the situation they would have enjoyed had the injury not occurred, as far as that is possible.

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Consequently, and given the complexity of current damage claims, this kind of intervention, once the exclusive responsibility of the medical expert (frequently the forensic physician), is becoming increasingly more inter-­and transdisciplinary, involving different medical specialities, the social services and psychology (as the object of evaluation is the person). The medical expert often has to request complementary opinions and reports from specialists in other areas, while nevertheless remaining responsible for the final assessment as a whole. Personal injury assessment is a medical act and is therefore subject to the rules and norms of the art of medicine. This means that the expert must display respect for the dignity of the injured person and an attitude of constant understanding and sympathy towards that person and his or her family, as well as availability in terms of time – something that is often essential for the proper diagnosis of the case. The expert also has to remain detached and impartial at all times, aiming to establish the truth of the facts under investigation so that justice may be properly done. As has already been mentioned, the legal principles regulating the assessment and reparation of personal injury vary from country to country, even in areas that are historically and culturally close. The European Union is paradigmatic in this respect, as its 27 member states offer a mosaic of different procedures and models concerning the assessment and reparation of personal injury and the parameters of damage to be evaluated. Thus, it is important to reflect on the present status of questions relating to personal injury assessment, the concepts inherent to it and the methodologies most commonly used, taking account of the specific requirements of different areas of law.

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68.2  Aim of personal injury assessment and reparation

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This matter traditionally made use of concepts such as ‘bodily harm’, ‘bodily damage’ or ‘harm to physical integrity’ expressions that suggest an exclusive relation with injury on the organic level. The concept of injury usually involves damage that is primarily biological (to the body), but which can also be manifested as disturbances to the victim’s capacities/functions, life situations/activities and participation. Thus, what is really at stake is the broader concept of personal injury, which requires a concrete, global and personalised approach to assessment and reparation in order to assist the injured person’s rehabilitation and reintegration into family, social and professional life (Hamonet and Magalhaes 2001; Magalhães and Pinto da Costa 2007). Indeed, this concept overlaps with the notion currently proposed by the World Health Organization (WHO) in its International Classification of Functioning, Disability and Health (2004).

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• Its type. • The victim’s experience of it (in the peri-­and post-­traumatic

68.3  General aspects for forensic medical assessment

periods).

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The medical expert’s function is to fulfil the aims of the expert report as impartially and objectively as possible, presenting complex notions in terms that magistrates, lawyers, insurance company professionals as well as the victims may understand, so that decisions concerning the application of sanctions or the granting of reparation and/or state benefits are based on properly grounded and comprehensible expert information (Anciaux and Attanian 1993; Rogier 1993). In the exercise of their functions, the medical expert has full autonomy over his or her scientific appreciation and is responsible for drawing up the expert report (though this may include the opinions of other specialists). This autonomy is fundamental to ensure that the expert does not feel constrained in formulating the conclusions considered most appropriate to each case. Indeed, the person performing the evaluation is in the best position to accurately assess the damages incurred as he or she was the one who most closely observes the victim and analyses the case. This circumstance naturally brings great responsibility, although it is common, and indeed desirable, for the opinions of other specialists to be heard where possible, and for the most complex cases to be discussed in departmental meetings prior to submission of the report (Magalhães et al. 2008). However, the expert should respect the norms, models and methods legally in force in the country or state in which he or she is practising as this is the only way to achieve any kind of harmonisation of expert services and ensure equitable treatment in similar situations. This harmonisation is fundamental in order to warrant good administration of justice and does not in any way curtail the technical or scientific freedom of these experts nor prevent them from giving full rein to their professional capacities (Magalhães et al. 2008).

direct correlation between their severity and the seriousness of the consequences of the trauma). • The perception that the person has of the event and its consequences. This latter perception is related not only to the resulting personal injury, but also to the justice system and to personal factors, such as previous experience of other traumatic situations, former health status, consumption habits, cultural and religious characteristics, the victim’s responsibilities, and his or her resilience and motivation for rehabilitation/reinsertion (Magalhães 1998; WHO 2004). Finally, the consequences of trauma also depend on environmental factors, which constitute physical, economic, family, social and even cultural contexts that may influence (positively or negatively) both the consequences of the trauma and the handicaps resulting from it (Magalhães 1998; WHO 2004). In order to assess the damage in the most global and personalised manner and promote concrete and integral reparation in a way that is as closely aligned with the victim’s worldview as possible, the expert should consider the person as a whole, rather than as the sum of his or her body parts, including capacities/functions and life situations/participation and activities. These three levels may be defined as follows (Hamonet and Magalhães  2000; Magalhães and Hamonet 2001): 1. Body: Biological aspects with their morphological, anatomical, histological, physiological and genetic particularities. 2. Capacities or functions: Physical and mental capacities (actual or potential), taking into account the person’s age and gender, irrespective of the environment where they live. These arise in the wake of the sequelae on the level of the body and are influenced, positively or negatively, by personal factors (e.g. age, previous physical and psychic state, motivation and personal efforts at adaptation) and environmental factors (e.g. architectural barriers and the need for technical or human assistance). 3. Life situations or participation and activities: The confrontation (concrete or otherwise) between the person and the reality of their physical, social and cultural environment. These situations may relate to the activities involved in the victim’s daily, family or social life, leisure, education, work or others, within a framework of social participation. They arise as a consequence of the sequelae on the level of the body and its functions and because of personal and environmental factors. This three-­dimensional approach, which analyses not only bodily injury, but also the concrete functions and situations that have been affected by the traumatic event or pathology, is fundamental to the expert evaluation, enabling a better perspective of the losses but, above all, the needs of the person in the light of what remains of his or her potential. In cases where the person’s functioning is seriously impaired, the following notions are essential (Didier et al. 1988):

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68.3.1  Role of the forensic medical expert

• The injuries resulting from it (even if there is not necessarily a

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68.3.2 General methodology for personal injury assessment

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The same forensic medical examination methodology should be used in all kinds of personal injury assessment situations, irrespective of the branch of law under which it is taking place, although there may be differences in the discussion and conclusion sections of the final report in accordance with the scope of law in which the expertise is performed (e.g. criminal, civil or labour law). For this reason, it is essential to know the rules and objectives of the law that governs each type of expert activity and to be aware of the implications that the expert report may have on the judicial decision. Particularly in post-­traumatic cases, the expert must be able to understand the full impact of the trauma upon the injured party, considering:

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qualifications and functions.

3. Type of interventions (at home or outside). 4. Type of activities envisaged (e.g. monitoring of vital functions, administration of therapy, hygiene, dressing and feeding).

5. Site of interventions (e.g. home or adapted establishment). 6. Degree of assistance required (e.g. monitoring, encouragement, complementary care or total replacement).

7. Duration and frequency/timetable of interventions (i.e. number of hours per day).

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68.3.3 Forensic medical report: general aspects

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The expert evidence is presented in the form of a report that describes the findings of the examination performed and interprets those findings, drawing properly supported conclusions. The report should comply with specific norms (Boróbia 2006) to fulfil its purpose completely, which will vary in accordance with the branch of law in which it is taking place. The report contains two fundamental parts: 1. The identification and description of the damages incurred (temporary and permanent), based on an interview, documental evidence, physical examination and ancillary exams performed. 2. The interpretation and valuation of the findings and conclusions drawn (fully substantiated). The following should be borne in mind when preparing an expert report of this type (Magalhães et al. 2008): 1. The damages should be described in a way that is clear, rigorous, objective, detailed, systematic and comprehensible to non-­medical practitioners (though medical terminology should nevertheless be respected, particularly as regards the established anatomical and traumatological nomenclature). 2. Sources of information should always be given. 3. The concepts used should always be defined, as there is a lack of consensus regarding many of them (particularly some of the parameters of damage), even within the same professional area. 4. Methods and instruments used for the assessment (e.g. scales and tables) should be duly identified. 5. The method used for the identification and description of damage should be generally the same, irrespective of the legal framework involved. 6. The specific aspects of the evaluation relative to each branch of law are only considered in the discussion and conclusion sections of the report. In most legal contexts, the report carries great weight as evidence, conferring responsibility upon the medical expert. Hence, the expert must be very demanding with regard to the content and quality of the message transmitted in the report to the other professionals that will go on with the case. This message should enable the unequivocal reconstitution of all that was observed by the expert in order to enable the case to be discussed

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depending not only on personal but also on environmental factors, which means that it may be possible to reduce them by altering the environment. 3. Quality of life depends on taking advantage of the remaining possibilities. Hence, it is essential in personal injury assessment constantly to refer to the person’s environment and life context (Magalhães and Hamonet 2000). In the light of this, it should be possible to indicate interventions required in relation to the person (e.g. treatment and rehabilitation) and to the individual’s environment (e.g. loved ones, housing, place of study, work or recreation, transport or access to specially adapted services) in order to reduce their hardships and limitations (Hamonet and Magalhães  2001). This should help to promote the person’s reintegration and autonomy, even in the most serious cases, and where possible, stimulate a faster return to active life. When there are important and serious handicaps (irrespective of the branch of law under which the investigation is taking place), a descriptive multidisciplinary approach should be taken, ideally performed in the victim’s own environment (home and/or workplace), and should include a forensic medical assessment as well as an evaluation of the victim’s degree of dependence and need for specialised helpers, always bearing in mind his or her remaining capacities and autonomy with and without technical assistance. Dependence may be relative to different kinds of needs (Magalhães and Vieira 2008): 1. Medicinal aid: This corresponds to the constant need for regular medication (e.g. painkillers, anticonvulsants or antiepileptic drugs) without which the victim would not be able to overcome his or her difficulties in terms of functions and daily life situations. 2. Regular medical treatment: This corresponds to the regular use of medical treatments to prevent the recurrence or aggravation of sequelae (e.g. physiotherapy). 3. Technical aids: These refer to the constant need for technology in order to prevent, attenuate, neutralise or compensate for personal injury (from the anatomical, functional and situational perspectives), with a view to obtaining autonomy and independence in daily life activities; this may involve technical aids for physical damages (impairments), or for functional or situational damages (disabilities). 4. Adaptation of the home, workplace or vehicle: This corresponds to the need for technology on the level of architecture, furnishings or equipment so as to permit the victim to perform daily activities that they would otherwise be unable to do, thereby losing autonomy and independence. 5. Third-­party assistance: This corresponds to human assistance for the victim that has become dependent, to complement or substitute a particular function or daily life situation. Third-­party dependence should be characterised relative to the following (Magalhães and Vieira 2008): 1. Type of assistance required in accordance with the necessary social–professional category (professional or otherwise).

2. Number and type of third parties required, and their

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1. The most important thing is not what is lost but what remains. 2. The real-­ life consequences (situational level) are relative,

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include family pathological antecedents, if relevant. This aspect is essential to determine the causal nexus between the trauma and the damage as it involves an analysis of the person’s state prior to the trauma. The chapter ‘Present state’ should include ‘Complaints’, ‘Physical examination’ and ‘Ancillary examinations’: 1. Complaints (Magalhães and Hamonet 2001): If the examination is based on the three-­dimensional method of damage description, the complaints may be listed in terms of the effects of the trauma upon the person’s functions and life situations. There are multiple human functions, but the most common and relevant include: a. carriage, displacement and transfer; b. manipulation and grip; c. communication; d. cognition and affectivity; e. sphincter control; f. sexuality and procreation. Life situations are unlimited, but the most significant damage should be described in accordance with the following aspects: a. acts of daily living; b. affective and social life, and leisure activities; c. professional life or training. As in most cases these aspects are described by the victim, it is important that the interview be guided and systematic, with frequent use of open questions that do not condition responses. In the case of people with severe handicaps, a detailed description of these complaints is essential, focusing on (for each level relevant to the case) the degree of difficulty observed in performance of a particular function or situation. 2. Physical examination: The damage itself is described in the ‘Body’ level. The medical examination should begin with the person’s general state, followed by a rigorous guided description, indicating all the characteristics of the injuries or sequelae (Magalhães et al. 2010c): a. type, for example, ecchymosis, excoriation or lacerations (in the case of acute injuries); dysmorphia, dysmetria, muscular atrophy, limitation of joint amplitude or stability, limb axis deviation or rotation, loss of segment or organ, disturbances of strength, sensitivity and equilibrium (in the case of sequelae); b. precise location; c. colour (if relevant); d. exact size (e.g. measuring dimensions or degrees and comparing with the contralateral side, in the case of orthopaedic or neurological permanent consequences). Reference should also be made to any alterations arising from a previous state. The following body regions should be considered in the examination (Magalhães and Hamonet 2001): a. skull; b. face; c. neck; d. spine and spinal cord; e. thorax;

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in full detail, particularly when there are contradictions or other circumstances (Magalhães et al. 2008). A model of the personal injury assessment report may include the following chapters corresponding to the different phases of the procedure: ‘Introduction’, ‘Information’, ‘Present state’, ‘Discussion’ and ‘Conclusion’ (Magalhães et al. 2010c). The ‘Introduction’ identifies the case in question and the victim, and should also include the type and date of the forensic medical examination. The chapter ‘Information’ may include a ‘History of the event’, ‘Documental data’ and ‘Past medical history’: 1. History of the event: This describes the trauma and its consequences based on information provided by the victim or whoever is representing them. This part is essential to understand the experience of the trauma and to support the damages to be attributed, particularly the most subjective. It should include: a. date, place, mechanism, type and circumstances of the traumatic event; b. resulting injuries (general description, in accordance with the information provided by the person); c. medical establishments used, complications that arose and treatment provided; d. date of hospital discharge and of outpatient consultations; e. date of return to work, or other circumstances, such as change in professional activity or retirement. 2. Documental data: This describes the information given in the clinical records, fundamental for a precise understanding of the injuries suffered and treatment provided, particularly when there has been some time lapse since the event and when the expert did not observe the initial injuries. A timeline should be provided showing the relevant data, avoiding transcriptions but always indicating the source of the information. It is important to include: a. the date of the traumatic event; b. resulting injuries (described in an ordered fashion, going from top to bottom, left to right and outside in, making use of all clinical records and mentioning any discrepancy that might exist between them); c. the medical establishments where assistance was provided, complications that arose and treatments given; d. date(s) of hospital discharge; e. outpatient consultations (e.g. specialties, institutions, treatments, ancillary examinations performed and period of consultations with reference to discharge dates); f. damage parameters valued by other experts, where relevant. It is important to correctly transcribe dates and to make reference to any aspects that offer a perspective on how the clinical condition has evolved and its repercussions upon the activities of the victim. 3. Past medical history: This describes any pathological and/or traumatic personal antecedents that could be relevant or influence the final outcome of the permanent consequences relative to the case under analysis (based on information provided by the victim and/or in the clinical files); it may

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68.4 Specifics of forensic medical reports for personal injury assessment The medical expert should study, describe, interpret and value the specific damages covered by the branch of law and the legal system in which the evaluation is taking place. These may generally be described in the way laid out below (adapted to suit the particular case).

68.4.1  Forensic medical report under criminal law

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It may be necessary to photograph the injuries to complement these observations. The photographs that accompany the report should be mentioned in the text and contain scales that enable the size of the injuries/sequelae to be estimated. They should also give an idea of how the injuries/sequelae look in the context of the whole body (e.g. a series of photographs from the general to the particular may be required). These photographs should only be taken with the victim’s formal permission (Magalhães et al. 2010c). 3. Ancillary examinations: This section describes the conclusions of examinations requested by the expert, including the date and place where they were performed (e.g. orthopaedic or ophthalmic examination, report of imaging or electrophysiological tests). The chapter ‘Discussion’ should focus on the causal nexus between the trauma and the injury, and if such a nexus is admitted, define the date upon which the injury was deemed as having cured or consolidated from the forensic medical point of view. In this chapter, the various injuries should also be discussed in the light of the law underpinning the specific assessment. The ‘causal nexus’ corresponds to the relation of imputability (i.e. total or partial, direct or indirect) between a trauma and an injury. It is important to bear in mind the various assumptions involved in its valuation: a. consistency between the type of injury or sequelae and a concrete event/aetiology; b. consistency between the type of aetiology and type of injury or sequelae incurred; c. consistency between the site of the trauma and the site of the injury or sequelae; d. anatomical–clinical consistency between the trauma and injury or sequelae; e. temporal consistency between the event, injury and sequelae; f. exclusion of the possibility that the injury or sequelae may have pre-­existed; g. exclusion of the possibility that the injury or sequelae may have been caused by a mechanism other than the event. The causal nexus is considered partial or indirect when there is another associated cause, usually relating to a previous state. In these cases, the trauma due to the event in question may trigger a pre-­existing clinical condition or accelerate or aggravate its evolution (Vieira and Corte-­Real 2008). The ‘cure’ corresponds to the moment when the injury is deemed healed without there having resulted in any sequelae (restitutium ad integrum), while ‘consolidation’ is the moment after which no further significant clinical development may be expected in terms of sequelae, corresponding to the end of the period of temporary damages (Magalhães et al. 2010c). The valuation of different aspects of the damage or injury may be divided into two moments: period of temporary damage and period of permanent damage (Magalhães et  al.  2010c). The parameters of damage to be described in this chapter may vary according to different branches of law, different legal systems or different legal medicine organisations.

The chapter ‘Conclusions’, when definitive, should not be long. It usually consists only of the reference to the existence or inexistence of a causal nexus, the date of cure or consolidation, and the various parameters of the damage mentioned and substantiated in the ‘Discussion’ section. Unfortunately, it is not uncommon for the body requesting the report to read only the conclusion, with prejudice to a correct interpretation of the whole. Thus, to avoid major problems, the medical expert should take care that the conclusions thoroughly describe the injuries incurred by the person as a consequence of the traumatic event (Magalhães et al. 2008).

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f. abdomen (including pelvic contents and perineum); g. superior limbs; h. inferior limbs (including pelvic bone).

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Examinations performed in the sphere of criminal law mostly involve offences to physical integrity (e.g. assaults or road accidents) and physical abuse (e.g. domestic violence). The main objective of the examination is to help the public prosecutor or judge affirm or deny the existence of personal injury corresponding to a specific legal type of crime, by means of a technical and scientific analysis (Magalhães et al. 2010b). Thus, it should involve a detailed description of the injuries and their consequences for the victim (e.g. their nature and extent, the effects on the victim’s body and health or any danger to life that might exist) and provide information about the influence of the agent’s conduct in producing the result, the number of days required for cure or consolidation, the nature of the instrument(s) used to produce such injuries, and the correspondence between the alleged criminal practice and the result (Magalhães et al. 2010b; Oliveira et al. 2010). In this case, the injuries may be described and valued taking account of the terms of the respective criminal code in the country where the event took place, with a full justification given in the ‘Discussion’ chapter. However, victims may also be entitled to reparation for injuries incurred in events liable to criminal prosecution. In such situations, the aim of the assessment may be not to assist the decision regarding the type of crime, but rather to describe the respective damage (temporary and permanent) liable to reparation. From this perspective, the valuation of damage in the ‘Discussion’ chapter and the reference to them in the ‘Conclusion’ may follow the methodology usually used in civil law.

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Temporary total professional repercussion

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This is an economic injury corresponding to the period in which the victim is entirely unable to carry out his or her usual professional activity. It is also assessed in days, based on the clinical records (e.g. health services and insurance company), concrete clinical condition, information provided by the victim or his or her representative, his or her usual profession and the expected period of recovery. If the clinical records do not provide enough information to estimate these periods, the expert may make use of tables, such as those by Ramírez (1996) or Pérez (2006). In the case of students, the report should mention the period in which their education is interrupted. With regards to people with subsistence activities, the report should mention the period in which those activities were affected. There may be various periods of total deficit alternating with periods of partial deficit (e.g. intermittent hospitalisations).

Temporary partial professional repercussion This is an economic injury corresponding to the period in which the victim has begun to resume his or her usual professional activity though with limitations. It is assessed using the same criteria as before without reference to rates or points. It should be noted that periods of functional deficit do not always coincide with periods of professional repercussion.

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Examinations performed in the ambit of civil law involve mostly the assessment of damages resulting from road accidents, and, less frequently, from assault, amongst others. The aim is to help define the reparation to be awarded in each case, taking account of the various kinds of damage liable to reparation. Thus, for the effects of assessment and reparation, two types of damage are usually considered: economic and non-­economic. Economic damage corresponds to the real consequences of the injury upon the victim’s financial situation. It refers to those damages that are liable to objective assessment and pecuniary reparation, such as expenses and loss of earnings (temporary or permanent) directly due to the event, and may be classified as quantifiable (temporary damage) or equitable (permanent damage). It covers losses that may be repaired, if not directly (through the natural restoration or reconstitution of the situation existing prior to the injury), then at least indirectly (by means of the equivalent or pecuniary reparation). The economic consequences of the injury should be repaired as completely (Boróbia 2006) and objectively as possible by means of an exhaustive calculation of the real temporary and permanent losses (Lambert-­ Faivre  2004). In some countries, permanent impairment tables are used, although in several others this is not considered advisable as they are considered as arbitrary and fictitious, failing to reflect the victim’s real situation. Non-­economic damages are those that cannot be assessed in pecuniary terms because they affect intangible assets that do not form part of the victim’s estate. Hence, they can only be compensated by means of a pecuniary obligation imposed on the person responsible for the harmful event, constituting a form of compensation rather than indemnity. These involve matters relating to the person’s health, well-­being, freedom, beauty, physical perfection, honour and good name; thus, they cover consequences that are physical (e.g. certain kinds of pain and the reduction of functional potential), psychic and aesthetic, analysed generally and independently of economic losses. The non-­economic consequences of personal damage are described (as far as possible) in the forensic medical report, and depending on their severity (established by the physician), they may give rise to compensation. Some of the economic damages are quantified based on permanent non-­ impairment tables (e.g. American Medical Association Table (Rondinelli  2007) or Guide Barème Européen d Évaluation Médicale des Atteints à l’Intégrité Physique et Psychique (Pierre et al. 2010)). The injury or damage may be described and quantified in the report as temporary and permanent, bearing in mind that this may vary in accordance with the legal system and forensic organisation of the country in question. The damages listed below are those stipulated by the European Confederation of Experts in Bodily Damage Assessment (CEREDOC), which are more or less consensual in most countries of Europe and South America (though with some variations that should always be taken into account) (Vieira 2003). The terminology used in different countries to refer to various kinds of damages may also vary considerably as efforts have not yet been made to standardise them.

The period of temporary damages includes total temporary professional repercussion, partial temporary professional repercussion, total temporary functional deficit, partial temporary functional deficit and quantum doloris (Magalhães and Vieira 2010).

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Total temporary functional deficit This is a non-­economic injury and corresponds to the period in which the victim is prevented from autonomously performing acts of daily, family and social life, without any reference to his or her professional activity. It is quantified in days and coincides with periods of hospitalisation and/or of total rest. It is determined on the basis of the clinical records (which indicate the periods of hospitalisation and bedrest at home, with dependence upon third parties for daily life activities), information provided by the victim or his or her representative and the assessment of the concrete clinical condition (which should enable the expected recovery period to be defined). If the clinical records are insufficiently informative, the tables mentioned above may be used. There may be various periods of total deficit alternating with periods of partial deficit.

Partial temporary functional deficit This is also a non-­economic injury and corresponds to the period in which the victim may resume activities of daily, family and social life with some degree of autonomy though still with limitations. It is assessed using the same criteria, without reference to rates or points.

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Permanent professional repercussion

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This is a non-­economic damage that corresponds to the physical and psychic suffering experienced by the victim during the period of temporary damages. This is valued by means of medical interview designed to gauge the victim’s subjective experience of the trauma (peri-­and post-­ traumatic circumstances). The following valuation criteria should be taken into account: • Type of trauma and its circumstances. • Type and nature of injuries incurred and treatment given. • Length of hospitalisation. • Medical and surgical complications. • Duration and complexity of functional rehabilitation period. The victim’s feelings during this period should also be considered (such as anguish, anxiety, fear, awareness of risk to life, and suffering through being away from family and professional activities). It should be valued using a seven-­point scale of increasing severity (1/7–7/7). Tables may also be used indicating the amount of pain corresponding to each damage situation, such as the Thierry and Nicourt table. The value attributed should always be well described and properly justified. With regard to permanent damages, these include permanent professional repercussion, permanent functional deficit, permanent aesthetic damage, permanent repercussion on sexual activity, permanent repercussion on sporting and leisure activities, and dependences.

sporting activity, though it is independent of professional activities. It is assessed with relation to the individual’s integral capacity (100 points or 100%) and may in some cases involve total compromise of this capacity (e.g. vegetative state). In valuing this damage, consideration should be given to cases that may have resulted from a previous state. It cannot easily be objectified as an economic injury hence it should be valued in the same way for everyone, irrespective of their professional or occupational activity, using medical permanent impairment tables, while the compensation should be calculated on the basis of standardised systems (e.g. reparation tables or the value of the point). The permanent impairment tables are merely indicative in character, and therefore the expert still needs to properly justify the assessment made, particularly when this differs considerably from the values stipulated. It must be thoroughly substantiated through an accurate detailed description of the complaints and sequelae, and a description of the implications of it for the person’s autonomy and independence. In cases where foreseeable damage is not yet observable, a ‘future damage’ may be anticipated (involving exclusively the aggravation of sequelae as part of the inexorable and logical development of the clinical condition); the expert should justify this circumstance in the ‘Discussion’ part of the forensic report and mention it again in the ‘Conclusions’. However, as it is not always possible to foresee the evolution of certain sequelae objectively and rigorously in terms of ‘future damage’, given the multiple variables to be taken into account pertaining to the victim (e.g. age, pathological states and types of activities), injuries (type, severity and evolution) and treatment provided, in such situations it is better to suggest a subsequent evaluation.

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This corresponds to the effect of the sequelae upon the victim’s ability to exercise his or her usual professional activity (considering the concrete activity on the date of the event). The following situations may be found with regard to the sequelae: • Compatibility with the victim’s professional activity. • Compatibility with the professional activity though requiring additional effort. • Incompatibility with the victim’s professional activity though compatible with other professions in his or her area of training. • Incompatibility with the victim’s professional activity or with any other activity in his or her area of training. It therefore constitutes an economic damage to be repaired objectively; hence, it must be substantiated by means of an accurate detailed description of the related work issues. In this case, recourse to a permanent impairment table is not advisable because as an economic damage its concrete reparation is possible and, therefore, is not necessary to transform the medical evaluation into an abstract or fictional estimate of the damage, using rates or points.

Permanent functional deficit This is a non-­economic injury, often called ‘permanent impairment’, which corresponds to the definitive effects upon the person’s physical and/or psychic integrity, with repercussions on their daily life activities, including family and social life, leisure and

Permanent aesthetic damage This is a non-­economic damage and corresponds to the repercussions of the sequelae upon the victim’s self-­image and image before others. The damage may be static (e.g. a scar) or dynamic (e.g. lameness when walking), and account should be taken of its prominence, the suffering caused to the victim (considering age, gender, marital status and social–professional status) and the possibility of recovery through surgery. The difficulties resulting from the double subjectivity in this assessment (on the part of the expert and the victim) may be overcome through a detailed description of the sequelae with regard to their location, shape, size, prominence, texture, colour and number, and by documenting them photographically (with the victim’s prior permission) to enable them to be appreciated by the decisionmaker. Nevertheless, the assessment will have to be personalised as similar damages may have different repercussions in different cases. This damage may be valued using a seven-­point scale of increasing severity (1/7–7/7). The value attributed should always be well described and justified.

Permanent repercussion on sexual activity This is a non-­economic damage and corresponds to total or partial limitation on the level of sexual performance/gratification arising from the physical and/or psychic sequelae. It does not

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Many legal systems only really consider, in theoretical terms, the economic damage related to the loss of working capacity, although, in reality, this damage may involve non-­ economic aspects that are ultimately repaired as if they were economic. The report should describe and categorise these damages as temporary or permanent, giving particular attention to their repercussions upon the victim’s work and earning capacities. The damages should be described, and in certain cases quantified, in accordance with the legal system and forensic organisation of the country. For example, the following damages are the most consensual in Europe and South America though with substantial variations that should always be taken into account by the respective experts. Regarding temporary injury, we can consider total professional temporary impairment, partial temporary professional impairment, permanent impairment and dependences (Magalhães et al. 2010a).

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This corresponds to the period in which the victim is entirely unable to carry out his or her usual professional activity. It is assessed in days based on the clinical records (e.g. health services and insurance company), concrete clinical condition, information provided by the victim or his or her representative and upon the demands of the profession. If the clinical records do not provide enough information on this matter, this damage may be assessed based on the average period of impairment expected, taking account of the clinical condition, its normal evolution to cure or consolidation, the type of profession in question and tables providing estimates of these periods, such as the Ramírez (1996) or Perez (2006). With some clinical conditions, there may be various periods of total temporary professional impairment, resulting from complications, relapse, recurrence or surgery (such as for the extraction of osteosynthesis material).

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include aspects related to procreation. This damage is frequently undervalued or not valued at all, partly because of the overriding significance of other serious sequelae, but also due to the preconceptions and reservations that still surround the matter on the part of experts and victims. The expert should give special attention to the conditions of the interview, content of the account (e.g. type of trauma) and affective resonance, victim’s age and prior state (e.g. diabetes, kidney or liver insufficiency, vasculopathies), and physical or psychological damage incurred (trauma epiphenomena). This damage may be manifested through disturbances to the libido, discomfort, erectile dysfunction or problems with ejaculation or orgasm. It is advisable for such complaints to be supported by ancillary examinations where possible. The assessment should take account of the initial injuries, resulting complications and the results of the ancillary examinations performed. If it is not possible to identify any organic cause of damage, the expert should pronounce upon the plausibility of the complaints based on the information already given and the experience of the trauma. It should be described in the forensic medical report only with the victim’s prior permission. This damage is different from damage to reproductive capacity, which should be valued in terms of permanent effects on physical–psychic integrity. It may be valued using a seven-­point scale of increasing severity (1/7–7/7), and must always be well described and justified. In some cases, the damage resulting for the victim’s sexual partner may also be considered.

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Permanent repercussions on sporting and leisure activities

Dependences

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This is a non-­economic damage and corresponds strictly and specifically to the impossibility of the victim engaging in certain leisure or social activities that he or she does regularly and which represents a clear source of personal fulfilment and gratification. This may be valued on a seven-­point scale of increasing severity (1/7–7/7), and should always be well described and justified.

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Dependences are related to the victim’s needs, with repercussion on his or her independence and autonomy. They should be assessed along the lines mentioned above for people with severe handicaps, taking into account his or her best chances of rehabilitation and reintegration.

68.4.3  Forensic medical report under labour law The examinations performed under labour law are almost exclusively related to work accidents so it is important to take account of the concepts of the ‘work accident’, ‘workplace’ and ‘work time’, which may vary in accordance with each legal system. The aim is to help define the reparation to be awarded in each case, considering the various kinds of damages liable to indemnity.

Temporary partial professional impairment This corresponds to the period in which the victim has begun to resume his or her usual professional activity though with limitations. It is also assessed in days based on the clinical records, concrete clinical condition, information provided by the victim or his or her representative and upon the demands of the profession. If the clinical records do not provide enough information on this matter, the expert should proceed in the manner described above. However, the periods and rates of impairment attributed by the physician who has accompanied the victim are usually accepted as it is difficult for the expert retrospectively to assess these accurately and rigorously.

Permanent impairment Theoretically, this corresponds to the loss of work capacity as a result of one or more dysfunctions, as final sequelae of the initial injuries, with total dysfunction understood as the total permanent

INSURANCE MEDICINE

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68.5 Conclusions

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expert assessments in the field of administrative law, given the various supports and benefits granted by the state. These assessments occur in the context of social insurance, fitness for duty, disability evaluations, amongst others, and vary greatly from country to country in their principles and objectives. In most cases, they are outside the domain of forensic medicine despite the fact that they clearly constitute legal medical acts. Without prejudice to the methodology described above for a complete assessment, the expert should always bear in mind the specific aims of the situation in hand and the methodological rules in force in the country where this is taking place.

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All the expert activity developed in the sphere of forensic clinical medicine is encapsulated in the expert’s report that, though not absolutely binding for the decision-­maker, nevertheless carries considerable weight in the decision-­making process. Thus, the medical expert has a great deal of responsibility, and an inaccurate comment or reference may have significant repercussions upon the final decision. Indeed, the consequences of a criminal and/or reparation process may profoundly affect the equilibrium of those that see it as the last chance to obtain justice that may otherwise be denied them (Magalhães et al. 2008). Thus, it is important for the expert to listen attentively and try to understand the victim’s experience of the post-­traumatic period and his or her complaints, persistently seeking out information regarding the case, observing the clinical records in detail and with caution, performing thorough and careful examinations, accurately describing all the information obtained and observations made, and grounding final conclusions with objectivity and scientific rigour. It is also important to request and accept any pertinent advice and suggestions from experts with different opinions, and to know how to use the time that the process requires without undue haste, though without delaying too long in sending the report  – for while the process remains unsettled, the victim may suffer deprivations that could be reduced by the timely application of the justice he or she deserves (Magalhães et al. 2008).

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Dependences

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impairment for any kind of work. There may be different levels of permanent impairment: partial permanent impairment, total permanent impairment for one’s usual work and total permanent impairment for any kind of work. Permanent impairment is determined by taking account of the global sequelae of the particular case (e.g. body, functions and life situations, with particular emphasis on professional activity). In some countries, these sequelae are quantified using the published permanent impairment tables in force and may be mandatory in some legal systems. This option is not consensual because, as mentioned above, the professional repercussion aspect may be assessed and repaired in a concrete fashion, so the application of tables becomes a mere abstraction that often makes the assessment and reparation unjust. In fact, the application of tables is only justifiable in the case of non-­economic damages. In order to ensure maximum rigour in assessing the damages resulting from work accidents, guaranteeing the victim’s rights and ensuring jurisdictional appreciation, the process set-­up for this effect should contain the following components: • Professional inquiry, particularly into the individual’s work history. • Analysis of the job, with characterisation and quantification of the professional risks involved, whenever technically possible (this is to establish and quantify the causal agent in the work accident). • Clinical history, with mandatory reference to relevant medical and surgical antecedents. • Appropriate ancillary examinations. In cases where total permanent impairment for usual work is attributed, account should be taken of the injured party’s residual functional capacity for another profession compatible with that incapacity, in the light of his or her age, academic and professional qualifications, and the possibility of professional integration. Permanent impairment for another profession compatible with that incapacity should also be assessed. In the valuation of permanent impairments, it is important to bear in mind questions relating to the injured party’s prior state and its possible aggravation or repercussion on the injury under appreciation. This may be considered in another way by the labour legislation in different countries.

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Dependences are related to the victim’s needs, with repercussion in his or her independence and autonomy. They should be assessed along the lines mentioned above for people with severe handicaps, taking into account his or her best chances of rehabilitation and reintegration.

68.4.4 Forensic medical report under administrative law As well as the three different areas of personal injury assessment described above (clearly integrated in many countries within legal medical expert activity), there is increasing demand for

References and further reading Anciaux, P. and Attanian, E. (1993). L’ expert et le préjudice non économique. Revue Française du Dommage Corporel 19 (4): 345–361. Boróbia, C. (ed.) (2006). Informe pericial en la valoración del daño. In: Valoración del Daño Corporal. Legislación, Metodología y Prueba Pericial Médica, pp. 389–430. Barcelona: Masson. Didier, J.-­ P., Casillas, J.-­ M., Moigne, C. and Dentan, S (1988). La Classification Internationale des Handicaps: sa place dans l’ enseignement médical souhaits et réalités. In: J. Sanchez (ed.), Actes du Colloque CTNERHI ‘Classification Internationale des Handicaps: Du Conceptà l’ Application’, pp. 131–141. Paris: Diffusion PUF.

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Magalhães, T. and Vieira, D.N. (2008). A avaliação dos grandes traumatizados e a atribuição de terceira pessoa In: D.N. Vieira and J.A. Quintero (eds.), Aspectos Práticos da Avaliação do Dano Corporal em Direito Civil, pp. 131–147. Coimbra: Caixa Seguros e Imprensa da Universidade de Coimbra. Magalhães, T. and Vieira, D.N. (2010). [General recommendations to perform clinical forensic medicine reports related with bodily harm assessment in civil law]. Revista Portuguesa do Dano Corporal 20: 79–90. Margeat, H. (1988). La réparation du dommage corporel à l’horizon 1992. Revue Française du Dommage Corporel 14 (3): 403–408. Oliveira, C., Santos, B., Jardim, P. et al. (2010). [Danger to life’ forensic assessment]. Revista Portuguesa do Dano Corporal 21: 37–48. Papart, T. and Groutel, H. (2010). Guide Barème Européen d’ Evaluation Médicale des Atteints à l’ Intégrité Physique et Psychique, 2nd edn. Louvain-­la-­Neuve: Anthemis. Pérez, M.G.-­B. (2006). Nuevo Manual de Valoración y Baremación del Daño Corporal, 14th edn. Granada: Editorial Comares. Ramírez, L.B. (1996). Tiempos de Curación en Traumatologia. Madrid: Praxis 2000. Rogier, A. (1993). L’ avis médical et l’ expertise. Revue Française du Dommage Corporel 19 (2): 123–128. Rondinelli, R. (2007). Guides to the Evaluation of Permanent Impairment, 6th edn. Chicago: American Medical Association Press. Vieira, D.N. (2003). Assessment of bodily damage in civil law: harmonization of an expert protocol. Revista Portuguesa do Dano Corporal 12 (13): 29–36. Vieira, D.N. and Corte-­Real, F. (2008). Nexo de causalidade em avaliação do dano corporal. In: D.N. Vieira and J.A. Quintero (eds.), Aspectos Práticos da Avaliação do Dano Corporal em Direito Civil, pp. 61–83. Coimbra: Caixa Seguros e Imprensa da Universidade de Coimbra. WHO (World Health Organization) (2004). International Classification of Functioning, Disability and Health. Geneva: WHO.

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Geerts, A. (1962). L’ indemnisation des Lésions Corporelles à Travers les Siècles. Paris: Librairies Techniques. Hamonet, C. and Magalhães, T. (2000). Système d’ Identification et Mésure des Handicaps. Paris: ESKA. Hamonet, C. and Magalhães, T. (2001). La notion de santé. La Presse Médicale 30: 587–590. Lambert-­Faivre, Y. (ed.) (2004). L’ indemnisation des préjudices de la victime directe. In: Droit du Dommage Corporel. Systèmes d’ indemnisation, 5th edn, pp. 159–282. Paris: Dalloz. Magalhães, T. (1998). Estudo Tridimensional do Dano Corporal: Lesão, Função e Situação. Sua aplicação médicolegal. Coimbra: Almedina. Magalhães, T., Antunes, I. and Vieira, D.N. (2010a). [Recommendations to perform clinical forensic medicine reports related with bodily harm assessment in labor law]. Revista Portuguesa do Dano Corporal 20: 69–78. Magalhães, T., Corte Real, F., Costa Santos, J. and Vieira, D.N. (2010b). [General recommendations to perform clinical forensic medicine reports related with bodily harm assessment in penal law]. Revista Portuguesa do Dano Corporal 20: 63–68. Magalhães, T., Corte Real, F., Costa Santos, J. and Vieira, D.N. (2010c). [General recommendations to perform clinical forensic medicine reports related with bodily harm assessment]. Revista Portuguesa do Dano Corporal 20: 53–61. Magalhães, T., Corte-­Real, F. and Vieira, D.N. (2008). O relatório pericial de Avaliação do dano corporal em direito civil. In: D.N. Vieira and J.A. Quintero (eds.), Aspectos Práticos da Avaliação do Dano Corporal em Direito Civil, pp. 159–171. Coimbra: Caixa Seguros e Imprensa da Universidade de Coimbra. Magalhães, T. and Hamonet, C. (2000). [The personal damage]. Revista Portuguesa de Avaliação do Dano Corporal 10: 46–69. Magalhães, T. and Hamonet, C. (2001). Handicap assessment: Setting the grounds for an effective intervention in the community. Medicine & Law 20: 153–166. Magalhães, T. and Pinto da Costa, D. (2007). Avaliação do dano na pessoa em sede de Direito Civil. Perspectivas actuais. Revista da Faculdade de Direito da Universidade do Porto 4: 417–452.

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Burkhard Madea

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Madea, B. (2020). Asphyxiation, Suffocation and Neck Pressure Deaths. Boca Raton, FL: CRC Press. Taylor & Francis Group. Madea, B. and Brinkmann, B. (2003). Handbuch Gerichtliche Medizin, Bd. II.Berlin, Heidelberg, New York, Tokyo: Springer. Madea, B. and Luhmer, A. (2017). Schnittstelle Rechtsmedizin – Polizei – Rettungsdienst. Bonn: LUHRI-­Verlagsgesellschaft bR. Madea, B., Musshoff, F. and Berghaus, G. (2012). Verkehrsmedizin. Fahreignung, Fahrsicherheit, Unfallrekonstruktion, 2nd edn. Köln: Deutscher Ärzte-­Verlag. Madea, B., Musshoff, F. and Tag, B. (2011). Kurzlehrbuch Rechtsmedizin. Bern: Huber Verlag. Madea, B. and Saukko, P. (2008). Forensic Medicine in Europe. Lübeck: Schmidt-­Römhild. Madea, B. and Weckbecker, K. (2020). Todesfeststellung und Leichenschau für Hausärzte. Berlin-­Heidelberg: Springer Verlag. Madea, B., Prangenberg, J., Ulbricht, J. and Doberentz, E. (2022). Feststellung der Todesursache, Berlin: Lehmanns Media GmbH. Mant, A.K. (1984). Taylor’s Principles and Practice of Medical Jurisprudence, 13th edn. Edinburgh, London, Melbourne, New York: Churchill Livingstone. Maresch, W. (1988). Atlas der Gerichtsmedizin. Stuttgart, New  York: Thieme-­Verlag. Maresch, W. and Spann, W. (1987). Angewandte Gerichtsmedizin, 2nd edn. Wien, München, Baltimore: Urban & Schwarzenberg. Mason, J.K. (1978). The Pathology of Violent Injury. London: Edward Arnold Ltd. Mason, J.K. (1989). Paediatric Forensic Medicine and Pathology. London: Chapman and Hall Medical. Mason, J.K. (1993). Forensic Medicine: An Illustrated Reference. London, Glasgow, New York: Chapman and Hall Medical. Mason, J.K. (1993). The Pathology of Trauma, 3rd edn. London, Boston, Melbourne, Auckland: Edward Arnold. Mason, J.K. (1995). Forensic Medicine for Lawyers, 3rd edn. London: Butterworths. Mc Lay, W.D.S. (1996). Clinical Forensic Medicine. London: Greenwich Medical Media. Mc Lay, W.D.S. (2009). Clinical Forensic Medicine, 3rd edn. New York: Cambridge University Press. Messina, R. (2010). Autopsi medico-­legali. Technica e diagnostica. Editioni Minerva Medica, Torino. Mueller, B. (1975). Gerichtliche Medzin, 2nd edn. Berlin, Heidelberg, New York: Springer-­Verlag. Naeve, W. (1978). Gerichtliche Medizin für Polizeibeamte. Heidelberg: Kriminalistik Verlag. Neureiter, F.v., Pietrusky, F. and Schütt, E. (1940). Handwörterbuch der gerichtlichen Medizin und naturwissenschaftlichen Kriminalistik. Berlin: Springer-­Verlag. Norelli, G.A., Buccelli, C. and Fineschi, V. (2009). Medicina legale e delle assicurazioni. Padova: Piccin. Oehmichen, M., Auer, R.N. and König, H.G. (2006). Forensic Neuropathology and Associated Neurology. Berlin, Heidelberg, New York: Springer. Oehmichen, M. and Saternus, K.S. (eds). Research in Legal Medicine – Rechtsmedizinische Forschungsergebnisse. Several volumes between 1992 and 2006. Lübeck: Schmidt-­Römhild.

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Stark, M. (.2011). Clinical Forensic Medicine. A Physician’s Guide, 3rd edn. Totowa: Humana Press. Stark, M. (2020). Clinical Forensic Medicine: A Physician´s Guide, 4th edn. Springer: International Publishing. Stark, M.M. (2002). A Physician’s Guide to Clinical Forensic Medicine. Totowa, New Jersey: Humana Press. Strassmann, F. (1895). Lehrbuch der gerichtlichen Medizin. Stuttgart: Enke. Strassmann, F. (1911). Medizin und Strafrecht. Berlin: Langenscheidt. Suvarna, S.K. (2016). Atlas of Adult Autopsy. A Guide to Modern Practice. Switzerland: Springer International Publishing. Tedeschi, C., Eckert. W.G. and Tedeschi, L.G. (1977). Forensic Medicine, 3 Vol., Philadelphia, London, Toronto: WB Saunders Company. Teke, A. (2001). Medicina Legal, Segunda Edición. Santiago, Chile: Publicationes Técnicas Mediterraneo Ltda. Thomsen, J.L. (2004). Retsmedicin  – Nordisk Laerebog. Copenhagen: Fadl’s Forlag. Tsokos, M. (2004–2006). Forensic Pathology Reviews, Vol. 1–5. Totowa/ New Jersey: Humana Press. Van de Voorde, W., Decorte, R. and Cuypers, T. (2010). Handboek forensische Geneeskunde. Brugge: Die Keure. Villadiego, M.S. and Pampin, J.B. (2015). Histopatologia Forense. Madrid: Ministerio de Justicia. Wagner, S.A. (2017). Colour Atlas of the Autopsy, 2nd edn. Boca Raton: CRC-­Press, Taylor & Francis. Wayne, J.M., Schandl, C.A. and Presnell, S.E. (2018). Forensic Pathology Review Questions and Answers. Boca Raton: CRC Press, Taylor and Francis Group. Whitwell, H., Thorne, K., Kolar, A. and Harvey, P. (2015). Mason´s Forensic Medicine for Lawyers, 6th edn. Sussex: Bloomsberry Professional. Wirth, J. and Schmeling, A. (2012). Rechtsmedizin – Grundwissen für die Ermittlungspraxis, 3. Auflage. Heidelberg: Kriminalistik Verlag. Wirth, J. and Schmeling, A. (2020). Rechtsmedizin – Grundwissen für die Ermittlungspraxis, 4th edn. Heidelberg: Kriminalistik Verlag. Wirth, J. And Strauch, H. (2000). Rechtsmedizin – Grundwissen für die Ermittlungspraxis. Heidelberg: Kriminalistik Verlag. Wyatt, J., Squires, T., Norfolk, G. and Payne-­James, J (2011). Oxford Handbook of Forensic Medicine. Oxford: Oxford University Press. Zagra, M., Argo, A., Madea, B. and Procaccianti, P. (2011). Medicina legale orientatata per problemi. Milano: Elsevier.

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Rognum, T.O. (ed.) (2010). Laerbok i Retsmedisin, 2nd edn. Oslo: Gyldendal. Rosendahl, W. and Madea, B. (2017). Tatorte der Vergangenheit. Archäologie und Forensik. Darmstadt: Konrad-­Theiss-­Verlag, Wissenschaftliche Buchgesellschaft. Rutty, G. N. (2004-­2006). Essentials and Autopsy Practice, Vol. 1–3. London: Springer. Rutty, G.N. (2006). Essentials of Autopsy Practice. London: Springer. Rutty, G.N. (2008). Essentials of Autopsy Practice. London: Springer. Rutty, G.N. (2017). Essentials of Autopsy Practice Reviews, Updates and Advances. Cham: Springer International Publishing. Rutty, G.N. (2019). Essentials of Autopsy Practice: Reviews, Updates and Advances. London: Springer. Saternus, K.S. and Madea, B. (2007). Forensic Autopsy. Research in Legal Medicine, Vol. 36. Lübeck: Schmidt-­Römhild. Sathyavagiswaran, L. and Rogers, C.B. (2018). Multidisciplinary Medico-­ Legal Death Investigation: Role of Consultants. Elsevier Academic Press. Saukko, P. and Knight, B. (2004). Knight’s Forensic Pathology, 3rd edn. London: Edward Arnold. Sauvageau, A. and Geberth, V.J. (2013). Autoerotic Deaths. Boca Raton: CRC Press, Taylor & Francis Group. Schmidtmann, A. (1905). Handbuch Gerichtliche Medizin, 9. Auflage des Casper-­Liman`schen Handbuchs, 3. Bände August Hirschwald Berlin. Schneider, V. (1991). Farbatlas der Rechtsmedizin. Stuttgart New  York: Gustav Fischer Verlag. Schwerd, W. (1992). Rechtsmedizin, 5th edn. Köln-­Lövenich: Deutscher Ärzte-­Verlag. Sheaff, T.M. and Hopster, D.J. (2005). Post Mortem Technique Handbook, 2nd edn. London: Springer. Siegel, J.A. and Saukko, P.K. (2013). Encyclopedia of Forensic Sciences, 2n edn. Amsterdam: Elsevier. Siegel, J.A. and Saukko, P.K. (eds.) (2013). Encylopedia of Forensic Sciences. Waltham: Academic Press. Siegel, J.A., Saukko, P.K. and Knupfer, G.C. (2000). Encyclopedia of Forensic Sciences, 3 Vol, San Diego, San Francisco, New York, Boston, London, Sydney, Tokyo: Academic Press. Simpson, K. (1979). Forensic Medicine, 8th edn. London: E Arnold Ltd. Spitz, W.U. (1993). Spitz and Fisher’s Medicolegal Investigation of Death: Guideline for the Application of Pathology to Crime Investigation, 3rd edn. Springfield: Charles C Thomas Publ.

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Handbook of Forensic Medicine, Second Edition. Edited by Burkhard Madea. © 2022 John Wiley & Sons Ltd. Published 2022 by John Wiley & Sons Ltd.

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ossification of clavicles  1032, 1033, 1034, 1035 physical examination  1029–38 radiation exposure in X‐ray examinations  1038–9 X‐ray examination  1038–9, 1039 sudden unexpected death  191 Adonis vernalis (pheasant’s eye)  1277 adrenal apoplexy  344 adrenal gland sepsis 343–4, 344 weight loss in starvation  674 adrenaline, in asphyxia  514–5 adrenergic syndrome  1125 adult respiratory distress syndrome  336, 407 adversarial legal systems  1058–9 adverse drug reactions  748, 748, 1015, 1099, 1244 adverse events in hospitalised patients  735, 736 AEIOU rule  61 African Court of Justice and Human Rights 28 age and driving  1384–5 older drivers  1384, 1387 young drivers  1385, 1429 age estimation  275–82, 275, 276 adolescents and young adults  1028–41 age marker concept  1032 dental examination  1030–2, 1031–2 ethnicity, influence of  1039–41 non‐ionizing imaging procedures  1035, 1036 ossification of clavicles  1032, 1033, 1034, 1035 physical examination  1029–38 radiation exposure in X‐ray examinations  1038–9 X‐ray examination  1038–9, 1039

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acetone 1271 analysis of  1171 blood levels  1185 elimination half‐life  1183, 1183–4 as marker of alcohol abuse  1444 respiratory toxicity and exposure threshold  1272 acetylcysteine 1111, 1117 6‐acetylmorphine chemical structure  1254 stability  1139, 1140 acetylsalicylic acid  789, 792 acid deaths  797 Aconitum napellus (monkshood)  1275, 1277 acrivastine, and driving fitness  1428, 1432 acrolein 616 actus rea (act/action)  314 acute coronary syndromes  1332, 1332 acute death (minutes to hours)  334–5 acute haemolytic transfusion reaction  745, 746 acute interstitial pneumonitis (AIP)  832 acute respiratory distress syndrome (ARDS)  339, 832 ad hoc tribunals  28 adhesion molecules  368–9, 369 adipocere 109 bodies recovered from water  138, 561 see also saponification administrative law, personal injury assessment  1634 adolescents age assessment  1028–41 age marker concept  1032 dental examination  1030–2, 1031–2 ethnicity, influence of  1039–41 non‐ionizing imaging procedures  1035, 1036

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abbreviated injury scale (AIS)  1455, 1456 ABC toxins  61, 62, 85 abdomen diving accidents  586 external examination  188 internal examination  192 abdominal injuries blunt force  403–406, 404–406 physical activity after  765–6 sharp force  456–7, 456, 457 abortion 699–700 illegal (criminal)  700–701, 700 causes of death  701–2 internally active agents  701 soapy abortion  701 legal 699 abortus imminens  699 abrasions  376, 380–2, 380, 382 elder abuse  1009 fingernail scratches  381, 382 genitalia 925 healing 1008 pressure/crushing 380–1 road traffic accidents  1460 tangential/brushing 380, 381 absorption  1155, 1158–60 factors affecting  1159–60 P‐glycoprotein transporter  1159–60 intestinal first pass metabolism  1159 pH value  1159 abused substances see illegal drugs; toxicology accidental death  78, 320 see also traffic accidents accommodation forensic DNA laboratories  48 forensic toxicological laboratories  49 accreditation 43–54 accumulated degree‐days  129, 271

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Page numbers in italics denote figures, those in bold denote tables

1642

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algor mortis  99–103 cooling curve  100, 100 correction for body weight  104, 105 temperature–time of death nomogram  99–101, 100, 103 see also body temperature aliphatic alcohols  1167–9, 1168 aliphatic diols  1171–3, 1187 aliphatic hydrocarbons  1269 alkalinisation 1112 allergic reactions anaphylactic shock  881 to blood components  746 alopecia, traumatic  1009 alprazolam chemical structure  1242 and driving fitness  1429, 1429 metabolism 1244 pharmacokinetics 1237 toxicological analysis  1417, 1419 alternating current  542, 640 alternobaric hypoxia  512 aluminium, sample requirements  1115 alveolar macrophages, drowning victims  566 Alysia manducator  356, 356 Amanita muscaria  85 Amanita phalloides (death cap mushroom)  85, 1275, 1277 American Academy of Psychiatry and the Law (AAPL)  1057, 1063 7‐aminoclonazepam, toxicological analysis  1417 7‐aminoflunitrazepam, toxicological analysis 1419 amitriptyline 789 and driving fitness  1429, 1429 toxicological analysis  1417 ammonia  616, 1267 respiratory toxicity and exposure threshold 1272 Amnesty International  27, 37 amniotic fluid embolism  351, 353 amperes 38, 640, 643 amphetamines  790, 1206–8 and driving fitness  1381, 1382, 1414 metabolism 1164 plasma concentrations  1208 stability  1140, 1141 structures 1207 toxicological analysis  1417, 1419 amputation fingers 377 self‐inflicted 1023 amyl acetate  1271

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as poison antidote  1176 poisoning 1176, 1185 properties 1389–90, 1390 tolerance 1401–2, 1401, 1402 toxicity 1176 alcohol abuse  772 appearance and social life of alcoholics  774 control of abstinence  1447–9 detection of  1447 and domestic violence  991–2 postmortem biochemistry  881 toxicological markers  1439–49, 1441 carbohydrate‐deficient transferrin  1442–3 chemical structure  1148 clinical laboratory parameters  1442 ethyl glucuronide  1441, 1441, 1444–5, 1448 ethyl sulphate  1441, 1441, 1444–5 fatty acid ethyl esters  1446, 1448 5‐hydroxyindolylacetic acid  1441, 1441, 1444 5‐hydroxytryptophol 1441, 1441, 1444 methanol/isopropanol/acetone 1444 phosphatidylethanol 1441, 1441, 1443 alcohol dehydrogenase  1163, 1168, 1187 alcohol‐related deaths  772–83, 1188–90, 1188 acute alcohol intoxication  772–3, 773 manner of  782–3 mixed alcohol‐drug intoxication  773–4 natural causes  774–8 non‐natural causes  778–7 bolus 781, 781 exposure 780 falls 780–1, 780 fires/smoking in bed  779–80, 779 traffic accidents  781–2 scene examination  779 alcoholic beverages, congener alcohols in 1181 alcoholic hepatitis  777 alcoholic ketoacidosis  775–6, 775, 1189 acetone/isopropanol blood levels  1185 alcohols (general)  1167–9 in alcoholic beverages  1181 aliphatic 1184, analysis of  1169, 1171 chemistry 1168–9, disposition and fate  1171–2 in doping  1286 metabolism 1172–4, 1174 see also specific alcohols aldehyde dehydrogenase  1172–5, 1172

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age estimation (continued) auricular surface (Lovejoy’s method)  280– 2, 280–2 child pornographic images  1028, 1041 and ethnicity  1038–41 eruption/mineralisation of third molars 1041 sexual maturation  1042 skeletal maturation  1040–1 expert reports  1041 forensic odontology  1577–9, 1578, 1579 adolescents  1031, 1042 and ethnicity  1041 older people  1042 fourth rib (Iscan’s method)  278–280, 279 historical aspects  1027 living individuals  1027–46 older people  1042 pubic symphysis (Suchey–Brooks method)  276, 276 quality assurance  1043 age‐related morbidity/mortality  1010 ageing 1005 agonal period  59–60, 60 AICAR  1286, 1289 air embolism  192–3, 200, 351–2, 352, 361, 585–7, 701, 746, 759 cardiac 331 transfusion‐associated 746 air weapons  461–2 aircraft accidents  769 airways alcohol effects  777 drowning victims  562 trauma 334–5, 335 AK‐47 guns  795–6, 796 alcohol  78, 1167–90 in alcoholic beverages  1181 amount in body  1179 analysis of  1169, 1171 blood alcohol concentration  1178, 1188–90, 1390, 1396–7 and driving fitness see driving under influence of alcohol effects 1401 immediate 772 long‐term 774 metabolic 774–5, 774 elimination rate  1178 intoxication  1390, 1396 metabolism 1174, 1175, 1176, 1177 pharmacodynamics 1236 pharmacokinetics 1176–8, 1176, 1178 physicochemical properties  1169, 1170

1643

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mouth and nose  543–4, 544 trachea and bronchi  545–6, 546 obstruction of respiratory tract  535–43 burking 537 head‐down position  539, 539 orthograde suspension  540 pneumothorax 536–7, 539, 539–41 positional asphyxia syndrome  538 respiratory paralysis  335, 642 thoracic compression  535–6, 535, 536 unstable thorax  541 oxygen deficiency  547–9 hypobaric  512, 549 normobaric/hyperbaric 547–9, 548 pathomorphology 519–22 evidential value  522 external 519, 520 histological 521–2 internal 521 radiological 522 pathophysiology 511–15 abnormal body positions  515 adrenaline effects  514–15 alternobaric hypoxia  512 arterial compression  513–14 brain glucose requirement  511 cerebral ischaemia  513–14 hypobaric hypoxia  512 hypoventilation 511–12 hypoxic brain death  511, 513 hypoxic sensitivity  511 intrathoracic pressure increase  514 jugular vein compression  514 normobaric/hyperbaric hypoxia  512 positional/crush 335, 336, 721 restraint  717, 721–2, 721 sexual abuse  929–30 stages of  515–16, 516 strangulation see strangulation subjective perceptions  516–17 agonal state  517 increased sexual response  517 pain and paraesthesia  517 visual impairment  517 aspiration 353–4 blood 354, 354, 355, 355 fluids 355 assessment of laboratory samples  46 assisted suicide  1605–10 asthma 781 astrocytes 412 staining methods  418 atherosclerosis  806, 826 and traffic accidents  1464–5

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apoptosis 415–16 apparent death  61–2 AEIOU rule  61 aquaporins  370, 569 Arbeitsgemeinschaft für Forensische Altersdiagnostik (AGFAD)  1027–8 argon ion laser  1507 Arias Stella phenomenon  698–700, 700 arms, dissection of  519 aromatic hydrocarbons  1269 arrhythmias 815–16, 816, 1333–8 bradycardia 1333–4 implantable cardioverter defibrillators  1334–8 supraventricular tachycardia  1334 ventricular tachycardia  903, 1334 see also specific types arrhythmogenic right ventricular dysplasia  75, 226 arrow wounds  491–2, 492 arsenic 1263–5 exposure 1263 poisoning 85, 111, 1117 reference concentrations  1264–5 sample requirements  1115 toxicity 1264 toxicokinetics 1264 arterial gas embolism  578, 582 arteritis 901 Asherman chest seal  336, 336 asphyxiation 508–49 auto‐erotic 710–11 plastic bag  547–8, 548 thoracic compression  535, 536 autopsy 517–19 anterior vertebral muscles  518–19 arms 519 artificial bloodless field  517 back and buttocks  519 hyoid bone, larynx and upper trachea  518, 518 neck 517–18, 518 nuchal muscles  519 oral cavity and nasopharynx  519, 519 children 411, 442, 951 classification 508, 509 definition 508, 509 external 508–9 internal 508–9 mechanical vs. non‐mechanical  509–11 natural vs. non‐natural  511 non‐fatal 516 obstruction of respiratory orifices  543–5 larynx 544–5, 546

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respiratory toxicity and exposure threshold 1273 anabolic steroids  1043, 1285 anaemia in children  944 anaesthetics, dissociative  789, 790 anamnesis  88, 1029 anaphylactic shock  881 ancestry informative SNPs  1541 aniline 1271–2 respiratory toxicity and exposure threshold 1273 animal predation  105, 105, 108–9, 109 anogenital lesions  921–30, 922–9, 925–6 anthropology/osteology  267–87, 1561–72 biological profile  273–86 age estimation  275–82, 275, 276 diagnosis of ancestry  282–3, 283 height estimation  283, 284 pathological markers  283, 284 sex determination  273–4, 274–5 stature estimation  283, 284 diagnosis of species  272–3, 273 disaster victims  294 facial reconstruction  286–7, 286, 286 geographical origin  287 identification of human remains  1562–7, 1563–6 identification of living  1567–9, 1568–9 PMI see postmortem interval recovery of remains  268–71, 269, 270 anticholinergic syndrome  1125, 1126 anticholinesterases 1280 anticoagulant rodenticides  1283–4 anticontamination precautions  47 antidepressants  789, 791 and driving fitness  1430–1, 1426 antidigoxin Fab antibody fragments  1110 antihistamines and fitness to drive  1427, 1431–3 H1 1246–7 antimony, sample requirements  1115 anus bruising 925 erythema 925 fissures, lacerations, scars and tags  927, 929 reflex anal dilatation  927 anxiolytics, and driving fitness  1426, 1429–30 aortic aneurysm  333 and traffic accidents  1464–5 aortic dissection 210, 827–8 aortic rupture  756, 828 aortic stenosis  203, 225 apnoea in children  943

1644

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metabolism 1244 pharmacokinetics 1237 stability 1140–1 toxicological analysis  1417 see also specific drugs benzoylecgonine and driving fitness  1381, 1382 plasma concentrations  1206 stability 1139 toxicological analysis  1418, 1419 berry aneurysm  397 and traffic accidents  1464–5 beta‐blockers 1286, 1287 beta2‐agonists 1285 Bichat’s fat pad, loss of  656 bilastine, and driving fitness  1428, 1432 bile specimen collection  1120 billiard ball effect  486, 486 Billroth, Theodor  15, 16 bioavailability 1158 biobanks 1613–15 biochemistry, postmortem  871–83 alcoholism 881 anaphylactic shock  881 diabetic coma  882 diagnostic value  873 genetic alterations  882 high excitation  881 hypoglycaemia  874, 877 hypothermia 881 liver function tests  877–8 renal failure  876–8, 879 water and electrolyte imbalances  880–1 biological fluids  872 cerebrospinal fluid  872, 873 pericardial fluid  872, 881 vitreous humour see vitreous humour biological profile  273–86 age estimation  275–82, 275, 276 auricular surface (Lovejoy’s method)  280–2, 280–2 fourth rib (Iscan’s method)  278–80, 279 pubic symphysis (Suchey‐Brooks method) 276–8, 278 diagnosis of ancestry  282–3, 283 height estimation  283, 284 pathological markers  283, 284 sex determination  273–4, 275 Biologischer Arbeitsstoff Referenzwert (BAR) value 1258 Biologischer Leitwert (BLW) values  1258 biomechanics 321–6 shear stress  326 tensile stress  321–6, 322–6 bipyridyl derivatives  1282 birth compression injury  958

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back, examination of  188 see also spinal cord; spinal injuries; spine bacterial myocarditis  814 ballistics 467–72 arrow wounds  491–2, 492 explosive injuries  492 autopsy 495 injury types  493–4, 494 mechanisms of injury  492 physics of explosion  492 reconstruction 495 exterior  466, 485 firearms 459–5, 460 billiard ball effect  486, 486 bone contact  470–1 deformation and fragmentation  470, 470 deformed lead pellets  486–7 energy deposit and transfer  471 head injuries  471–2, 472 high velocity missiles  471, 472 mechanisms of injury  468–9, 468 missile–tissue interaction  469–70, 470 sawn‐off shotguns  487 shock wave  469 yawing  467, 469–470, 470 interior 466 see also shooting incidents Bamberg Criminal Code  3, 6 banned substances  1284 bar hold  720, 720 barbiturates  788, 789, 1236, 1239–40 chemical structure  1240 pharmacokinetics 1237 toxicological analysis  1417 see also specific drugs barotrauma  376, 547 lightning explosive  657, 658 pulmonary  478, 581 see also diving accidents bathroom, deaths in  84 bathtub electrocution  652–3 battered child syndrome  970 Battle’s sign  944 beard growth, apparent  104 beatings 37 child abuse  972 Belgium, statutory limits for alcohol levels 1394 bending fractures  323, 324 benzene 1273 benzodiazepine‐related drugs  1236, 1244, 1247 benzodiazepines 788, 788, 1241 chemical structure  1242 and driving fitness  1381, 1382, 1429–30

N

atlanto‐occipital dislocation  400 atomic absorption spectrometry  1132 Atropa belladonna (deadly nightshade)  1275–6, 1277 atropine 1108 auricular surface method of age estimation  280–2, 280–2 Australia fitness to plead  1062 statutory limits for alcohol levels  1395 Austria 12 insanity standards  1064 statutory limits for alcohol levels  1394 auto‐erotic deaths  710–11 asphyxiation plastic bag  547–8, 548 thoracic compression  535, 536 categorisation 710 injuries to male genitalia  712–13 mechanisms of  710–11, 710 autolysis 105 autonomic nervous system in diving response 719 autopsy 28–30, 28–30, 517–19 benefits of  183 cause of death  70–6 clinical  184, 193–7 cardiac conductive system  195 children 195 fetal autopsy  195 legal background  194 microbiological 196 neuropathological 196, 197 occupational lung disease  196 performance of  194–5 special procedures  195–6 forensic see medicolegal autopsy forensic imaging  207–15 manner of death  81–3, 82 medical malpractice  751 molecular 564 postmortem changes see postmortem changes protocols 31–2 religious objections to  296 standards 43–5 terminology 313 see also specific modes of death autopsy report  197–8 autopsy room  184 autosomal markers  1156–8 AVID (Antiarrhythmics Versus Implantable Defibrillator) study  1336 axons injury 416 staining methods  418

1645

Index

N

O

C

R

FO

O

N

LY

confirmatory tests  64–7, 262 definition 62 diagnosis 62–4, 64 UK criteria  262–3 drowning 560 hypoxic  508, 511 neuropathological findings  444 brain injuries  390–400, 391–400 boxing‐related 430 children  433–5, 944, 945 pathophysiology 949–51, 950 closed 427–32, 428, 429 compression  950, 951 contrecoup lesions  397, 398, 418, 429 contusions 397, 398, 399, 429–30, 430 children 434 microscopic findings  429 survival time  430 cortical haemorrhage  398–9, 398 diffuse axonal injury  399, 416 impact (acceleration/deceleration) injury 949 intracerebral haemorrhage  430, 430 intracranial haemorrhage see intracranial haemorrhage; and specific types intracranial vessels  425–6 lacerations 419 children 434 open 426–7 and oxygen deprivation  444 penetrating 951 rotational impact deceleration  951 rotational (inertial)  950, 950 shearing/rotational 950 traumatic mechanisms  950 whiplash  433, 951 see also head injuries brain swelling  394, 396, 399, 416, 419, 943 breath alcohol analysis  1389, 1392–4, 1392, 1394 Brittain, R.P. 3 bromazepam, toxicological analysis  1419 bromide salts  1247 bromides 1247 sample requirements  1108 bromisoval 1247 chemical structure  1246 pharmacokinetics 1237 bromofluorocarbons 616 bronchial obstruction  545–7, 546 bronchiolitis 900 bronchopneumonia alcohol‐associated 780 prurulent 406 Brouardel, P.C.H. 11 Brugada syndrome see long QT syndrome

U

TO

R

U

SE

pregnant women  704 sternum 332 see also specific injuries blunt force torture  37, 37, 38 sequelae 39–40 body armour  491 body fluid identification  1499–502 body mass correction factor  115 estimation of  114 temperature correction  104, 105 body mass index (BMI)  679, 1536 body modification  303, 1021 body packers  216, 784–5 body temperature drowning victims  559 estimation of, rectal  113, 114 regulation 627, 628 see also algor mortis bog bodies  109, 111 Bohn, Johannes  9 bolus death  544–5, 545 alcohol‐related 780, 780 bomb curve  227 bombs, self‐made  493 Bonaventura‐Orfila, Mathieu‐Joseph  11, 11 bone diagnosis of species  272–3, 273 diaphyseal length and age  276 dispersal of  269 postmortem interval  272 specimen collection  1123 see also anthropology/osteology bone‐by‐bone radiographic methods  1030 bone marrow embolism 353 specimen collection  1123 bone marrow derived cells  370 borderline personality disorder  1021, 1022 bows 491 boxing injuries  430 Boyle’s law  578, 579 bradycardia 1333–4 brain autopsy 196 examination 416–18, 417 exenteration of  472 glucose requirement  511 imaging in non‐accidental head injury  961, 963 sepsis 343–4, 344 specimen collection  1121 weight loss in starvation  674 brain death  62–7 checklist 65–6 children 443–4, 443

TR IB

bismuth poisoning 85 sampling requirements  1115 bite marks  1580–3, 1581, 1582 animal predation  109, 109, 138, 138 elder abuse  1009 human 385, 972 sexual abuse  929 bitemporal hemianopsia  1351 black powder firearms  461 blank cartridges  465–6 blisters electrical 379 putrefactive 134, 135 bloating 106, 106, 107, 134 blood aspiration 354, 355, 355, 450, 545, 546 dried blood spot sampling technique  1443 drug stability in  1139–40 specimen collection  1119, 1143 blood alcohol analysis  876, 1390–2, 1390, 1391 blood alcohol concentration (BAC)  51, 405, 425, 570, 638, 764. 1091, 1101, 1396–7, 1398 and crash risk  1399 units 1391 blood components allergic reactions  746 bacterial contamination  746 blood doping  1286 blood stains  620, 1500 blood transfusion errors  745–7, 745, 746 blood vessels, alcohol effects  779 blow fly (Lucilia sericata)  127–8, 127 Bluestar 1498 blunt force injuries  37, 329, 376, 376, 380–400 abdomen and pelvis  403–6, 404–6 chest 400–3 child abuse  971–4, 972–3, 972, 974 elder abuse  1007–10, 1008, 1009 extremities 406–7 head 390–400 brain/cerebral injuries  397–400, 398–400 facial trauma  393–4, 293, 294 intracranial haemorrhage  394–7, 395–7 scalp 420–1, 421 skull fractures  190, 390–3, 390–3, 421–2, 421, 422 integument 380–400 abrasions  376, 380–2, 381, 382 contusions  376, 382–8, 383–8 lacerations  376, 388–90, 389 neck  192, 400

1646

Index

TR IB

U

TO

R

U

O

C

R

FO

LY

N

SE

cachexia 669, 670 cadaveric poisons  106 cadmium 1260–1 exposure 1260 poisoning 1117 reference concentrations  1261 sample requirements  1115 toxicity 1260 toxicokinetics 1260 ‘café coronary’ 781, 781, 1014 Caffey, John  939 Caisson’s disease  376 calcium 881 calcium disodium EDTA  1117 calibre of firearms  464, 464, 465 Calliphora vicina  131 camazepam 1237 Canada competence to stand trial  1062 insanity standards  1064 statutory limits for alcohol levels  1394 cancer 77 and alcohol consumption  778 cannabinoids 1286 cannabis  788, 790, 1197–200 and driving fitness  1381, 1411–13 toxicological analysis  1417 capacity see mental capacity caprettamento 796–7 captive bolt devices  462 car bombs  793, 798 car to car collisions  1460–4, 1462 frequency 1462 front‐end collisions  1461–3, 1462 rear‐end collisions  1463–4 seat belt injuries  384, 1461–2, 1463

side collisions  1463 car to pedestrian collisions  1455–60, 1456–60 impact 1455, 1457 loading 1455, 1459, 1464, 1464 lower leg injuries  1456, 1458, 1477 overrunning 1460, 1462 shoe heel abrasions  1458 shuffling off  1455, 1457, 1458–9, 1458 trace patterns on car  1457 tyre tread marks  383, 1461 carbamate insecticides  1282 carbamazepine and driving fitness  1433 poisoning 1111 toxicological analysis  1417 carbon dioxide  1266 effects of  616 respiratory toxicity and exposure threshold  1272 toxicity 581 carbon monoxide  1266 poisoning 85, 616, 780 divers 581 lividity 95, 95 smoke inhalation  614 respiratory toxicity and exposure threshold 1272 carboxyhaemoglobin  1110, 1487–8 carbromal chemical structure  1246 pharmacokinetics 1237 cardiac causes of sudden death see sudden cardiac death cardiac syncope  1339 cardioinhibitory reflex cardiac arrest (CiRCA)  329 cardiomyopathy 809–15, 814 dilated  813, 810 hypertrophic  813, 809–10 restrictive 810–11 stress 811–12, 612 cardiovascular disease, driving fitness  1329–41 arrhythmias 815–16, 816, 1333–8 coronary heart disease  1332–3 heart failure  1340–1 risk of harm formula  1331–2 syncope 1338–9 carisoprodol, toxicological analysis  1417 carotid artery compression  442–3 carotid sinus reflex  514 carotid sinus syncope  1338–9 carotid sleeper hold  533, 720

O

buspirone, and driving fitness  1426, 1430 butalbital, toxicological analysis  1418 butane 1208 respiratory toxicity and exposure threshold 1272 1,4‐butanediol  1168, 1187–8, 1187 metabolism 1176, 1187 toxicity  1176, 1236 2‐butanol, in alcoholic beverages  1181 n‐butanol, in alcoholic beverages  1181 t‐butanol 1168 buttocks, dissection of  519 butyl acetate  1271 respiratory toxicity and exposure threshold 1272 butylene glycol  1271 butyro‐1,4‐lactone 1209

N

bruises see contusions bucket handle fractures  948 bullet embolism  353, 490–1 bullets 464, 464 deformation 469, 469 determination of  484–5 deviation 467 fragmentation 467, 469, 470 infectivity 490 lead poisoning  491 mass and muzzle velocity  464, 464, 465 penetration 467 reflection 467 ricochet 467 yawing  467, 469, 470 buprenorphine  1147, 1203 chemical structure  1254 metabolism 1164 pharmacokinetics 1255 buried bodies  270 burking 537 Burma 32, 33, 783 burning torture  37 sequelae 39–40, 40 burns  379, 607 body checklist  619–20, 619 additional examination  620 adhesions of fire scene on body  619 autopsy 620 clothing 619 external examination  619–20 child abuse  979, 980 classification 609–11, 610, 610, 611 first degree  610, 610, 611 second degree  610, 610, 611 third degree  610–5, 610, 611 fourth degree  610, 611, 611 criminological aspects  620–1 elder abuse  1009 electrical  379, 641–43, 641–43 current marks (Joule burns)  641–2, 644 exposure time  607, 609 heat damage  607, 608 postmortem 136, 138, 611, 614, 612–13, 612, 616 microwave 643–4 mortality probability  612 self‐inflicted 1023 sources of  608 surface area  611, 612, 612 temperature  607, 609, 609 see also fire injuries burst fractures  324, 1460

1647

Index

N

O

C

R

FO

O

N

LY

skull fractures  434, 948, 948, 958 smothering 443 strangulation 441, 442 sudden unexpected death  191, 438 suffocation 442, 442 China diminished responsibility  1067 forensic medicine  5 positional torture  39 Chlamydia trachomatis  931, 935 chloral hydrate  1240 metabolism 1240 pharmacokinetics 1237 chlorazepate 1242 chlordiazepoxide 1146 metabolism 1242 pharmacokinetics 1237 chloride 880–1 chlorinated hydrocarbons  1269–70 chlorine 1268 respiratory toxicity and exposure threshold 1272 chlorophenoxy compounds  1281–2 choke holds  533 choking 544, 545 cholinergic syndrome  1124–5 cholinesterase poisoning  1111 chromatic skin discoloration  561 chromatography 1131, 1132 drug stability in  1147 see also specific types chromium, sample requirements  1115 chyme aspiration  546–7, 546 cigarette burns  40 in children  948 circulation, vital reactions  350–2 embolism see embolism haemorrhage see haemorrhage cirrhosis 777 citalopram 789 civil forensic psychiatry  1057 civil law competencies 1060–1 consent to treatment  1061 legal competency  1060 retrospective civil assessments  1060–1 testamentary capacity  1060–1 personal injury assessment  1630–4 dependences 1633 partial temporary functional deficit  1631 permanent aesthetic damage  1632 permanent functional deficit  1632 permanent professional repercussion  1632

U

TO

R

U

SE

chest injuries  335–6, 336, 400–3, 401–3 open wounds  335–6, 336 physical activity after  765, 766 pneumothorax and gastrothorax  336–7, 336 rib fractures  336, 366, 400, 401 sharp force  456, 456, 457, 766, 766 child abuse  939–63, 948 blunt force  971–4, 972, 972, 973, 974 burns 975, 976–8, 978 case history  984 contusions 945, 945 differential diagnosis  980–1 haematological disorders mimicking  981 criminology 971 definitions 970–1 differential diagnosis  979–80, 980–1, 982 documentation 984 epidemiology 970–1 eye injuries  978 fractures 974–5, 974, 977 historical aspects  939–40 interpretation of injuries  974, 974 lethal 982–3 Münchausen syndrome by proxy  980–2 non‐accidental head injury  939–84 physical examination  971 physical neglect  983 poisoning 983, 983 sexual see sexual abuse shaken baby syndrome see shaken baby syndrome child pornographic images, age estimation  1028 child protection  961–2, 962 child protection case conference (CPCC)  961 child welfare  1094 children abuse see child abuse apnoea 943 asphyxiation 441, 442, 951 brain death  443, 443 brain injuries  433–4, 944–6, 945, 950, 950 causes of death  888 clinical autopsy  194 head injuries accidental  958–9 non‐accidental 939–3 hypoxia 441, 442 ischaemia 439–40, 440, 442 permanent global  443, 443 juvenile pornography  933 sexually transmitted infections  930–2

TR IB

cartilage, specimen collection  1122 cartridges 463, 463 blank 465–6 tear gas  465–6 Casper, Johann Ludwig  3, 9–12, 12, 107, 108 cataracts 1346–7 assessment 1351 classification 1346 clinical symptoms  1346 development of  1346 epidemiology 1346 mesopic vision  1347 relevance for road users  1347 treatment 1346–7 catecholamine release  881 Catha edulis (khat/quat)  1276, 1277 cause of death  70–80, 71, 77, 314, 327–46, 328 autopsy findings  80–1 errors in diagnosis  83 infants and children  888 liability and time  314 postmortem biochemistry  871–83 sharp force injuries  448–57, 449 statistics 80, 77, 78 see also specific causes CBRNE weapons  296 cell embolism  351 central nervous system  412 depressants 788 examination 416–18 SIDS 895 stimulants 787, 788, 790 cerebral arterial gas embolism (CAGE)  576, 581, 582 cerebral hypoxia  515, 707 cerebral ischaemia  513–14 cerebral oedema  202, 399, 702, 944, 949 cerebral perfusion pressure  443, 944 cerebrospinal fluid  872, 873 specimen collection  1122 cervical spinal injuries  417, 432 cetirizine, and driving fitness  1428, 1095 charring 270 and fire duration  618–19, 619 chemical biological radioactive nuclear explosive weapons see CBRNE weapons chemical warfare  1095–6 chemokines 369, 369 chest external examination  584–5 internal examination  585 chest fluid specimen collection  1122

1648

Index

U

R

TO

U

TR IB

O

C

R

FO

O

N

LY

with cortical haemorrhage  428–30 shooting incidents  474 conductance 640 confidentiality 1593–5 personalised medicine  1595 congenital heart disease  74, 809 sudden death  819 Conium maculatum (hemlock)  1276, 1277 consent to treatment  1061 emergency situations  1592 information 1592–3 minors 1592 personalised medicine  1598 Constitutio Criminalis Carolina  3, 5 contra bonos mores  1591 contraction band necrosis  344, 344, 807 contrecoup lesions  397–9, 397, 398 controlled substances see illegal drugs contusions  376, 382–8, 383–8 ageing of  974, 974 anal 927 brain 397, 397, 398, 429–30 children  434, 950 microscopic findings  431 survival time  430 child abuse  944, 973 differential diagnosis  980–1 haematological disorders mimicking  980–1 colour changes  386–7, 387 elder abuse  1007 fingertip bruises  384–5, 384, 973 genitalia 622–624, 925 Convallaria majalis (lily of the valley)  1277 Convention Against Torture  35, 36 Convention on Human Rights and Biomedicine  259, 1590, 1592 copper, sample requirements  1115 cor pulmonale  72, 601 coronary heart disease  1332–3 hospital mortality and prognosis  1332–3 percutaneous coronary interventions  1333 stable 1333 and traffic accidents  1464–5 coronary stents  806 corpses buoyancy of  561, 562 condition of  191 definition of  69 disposal in water  472 examination see external examination handling of  1615–18 identification see victim identification position of  87 research on  1610–15

SE

and fitness to drive  1414 metabolism  1163, 1205–6, 1205–6 physiological effects  1204 plasma concentrations  1206 stability  1139, 1140–1 toxicological analysis  1417, 1419 cockroaches, skin lesions caused by  140 codeine 1202–3 chemical structure  1254 metabolism 1163, 1202 pharmacokinetics 1255 plasma concentrations  1205 toxicological analysis  1418 coffin birth  106 coin‐rubbing 382 colchicine  1165, 1272 Colchicum autumnale (autumn crocus)  964, 1277 cold injuries see hypothermia cold shock  558 cold stiffening  60, 98 collagens  365, 370, 370 Colombia 31 colour vision disorders  1348, 1350–2 colposcopy 921 coma 874 coma blisters  1240 Committee Against Torture  35 commotio cerebri  429 compensatory anti‐inflammatory response syndrome 339–40 competency civil law  1060–1 consent to treatment  1061 legal competency  1060 retrospective civil assessments  1060–1 testamentary capacity  1060–1 criminal law  1061–3 competence to stand trial  1061–2 determination and consequence  1063 duration of hospitalisation  1063 fitness to plead  1062–3 indications for hospitalisation  1063 no fitness to plead/competence to stand trial 1062 treatment refusal  1063 compound method for time of death  115–18, 116–18 compression fractures  326 computed tomography angiography  211–12 computed tomography (CT)  211–12, 457 sharp force injuries  457 concussion injuries with contusions  429–30, 429

N

civil law (continued) permanent repercussion on sexual activity 1632–3 permanent repercussions on sporting and leisure activities  1633 quantum doloris  1632 temporary partial professional repercussion 1631 temporary total professional repercussion 1631 total temporary functional deficit  1631 clavicles, ossification  1032, 1033, 1034 clearance 1156–7 clemastine, and driving fitness  1432, 1428 clinical autopsy  184, 193–7 cardiac conductive system  195 children 195 fetal autopsy  195 legal background  194 microbiological 196 neuropathological 196, 197 occupational lung disease  196 performance of  194–5 special procedures  195–6 clinical forensic medicine examinations 32 ISO/IEC  17025 53 clinical forensic toxicology  1091–3 child welfare  1094 driving under the influence  1091–3 drug‐facilitated sexual assault  1094 workplace drug testing  1093 clobazam chemical structure  1242 pharmacokinetics 1237 clomethiazole 1246 chemical structure  1246 pharmacokinetics 1237 clonazepam, toxicological analysis  1417 clorazepate 1237 and driving fitness  1426, 1426 clothing and textiles asphyxiation by  543, 544 burned bodies  619 examination of  185–6 shooting incidents entrance wounds  475–7, 476–7 fibres in wound  481 position of victim  483 cocaethylene, toxicological analysis  1417 cocaine  788, 790, 1204–6 absorption 1205 distribution 1205 and driving fitness  1381, 1381 elimination 1205–6, 1205–6

1649

Index

N

O

C

R

FO

cyclohexane 1273 cyclo‐oxygenase 369–70 cyclopyrrolones 788, 788 cytokines 369, 369

O

N

LY

dactyloscopy  291, 295 Dalton’s law  512, 548, 578 data handling  1604 data protection  1599 databases 1613–15 date rape  932, 1094 Datura stramonium (thornapple/Jimson weed/angel’s trumpet)  1276–7, 1277 De relationibus medicorum  8 De renunciatione vulmerum  9 death and dying  59–60 agonal period  59–60, 60 brain death  62–4 determination of death  60–7 manner of death  81–3, 82, 1090 time of death see time of death uncertain signs of death  61, 62 see also death certificate death certificate  61, 70, 78, 184, 185 cause of death  70–80, 71, 77, 314, 327–46, 328 autopsy findings  80–1 statistics 80, 77, 78 completion of  99 errors in  77, 78, 83 in‐custody deaths  725–6 manner of death  81–3, 82 terminology 70 death on operating table (exitus in tabula)  747–8 complications 748 occupational groups  747 type of intervention  748 decapitation 377 Declaration of Helsinki  259 Declaration of Human Rights  27 decollement  387, 1456 decomposition 107, 108 CT imaging  214 in grave (saponification)  110–11, 111 see also putrefaction decompression illness  580 decompression sickness  578–81, 603 decubitus ulcers in elder abuse  1013–14 differential diagnosis  1013 grading 1014 defence injuries  385, 386, 548 forearms/hands 455, 455–6 shooting incidents  489

U

TO

R

U

SE

cribra orbitalia  286 crime scenes  161 aircraft accidents  769 alcohol‐related deaths  779 DNA evidence  1497–9 drug addiction  785 initial examination  161–2 integrating chart for casework  117–19 potential health hazards  161 securing stains and evidence  163 sexual homicide  730 sharp force injuries  454–7, 465–7 tasks 163–4 time of death see time of death criminal forensic psychiatry  1057 criminal law competencies 1061–3 competence to stand trial  1061–2 determination and consequence  1063 duration of hospitalisation  1063 fitness to plead  1062–3 indications for hospitalisation  1063 no fitness to plead/competence to stand trial 1062 treatment refusal  1063 personal injury assessment  1625 criminal offences, faking of  1021 criminal responsibility  1063–70 determination and consequence  1069 diminished responsibility and overwhelming emotion 1067–9 insanity standards  1064–7 no insanity defence  1067 preventive detention  1069 rehabilitation and security  1069 criminality 317–20 homicide 317–19, 318, 320 non‐homicidal events  319 suicide  319, 320 torture see torture crossbows 491–2 crush asphyxiation  335, 336 crush‐tear injuries  326 CT see computed tomography cultural issues religious objections to autopsy  207 cupping 383 cyanides 1268–9 poisoning 85, 616, 618, 1266 lividity 96 respiratory toxicity and exposure threshold 1273 stability 1145 cyanoacrylate fuming  1498 cyber mobbing  997

TR IB

status of  1611–13 temperature of  99, 100–3, 104, 105 cortical haemorrhage  427–8, 429 Cosa Nostra see Mafia‐related deaths Council of Europe  35 Convention on Human Rights and Biomedicine 259 counterpressure injuries  519 COVID‐19 associated deaths  76 causes of death  76–80, 79 clinical courses  833 defence mechanisms  251 diagnostic methods infectious particles (virus culture)  834–5 viral proteins (electron microscopy/ immunohistochemistry) 834 viral RNA (RT‐qPCR/in situ hybridization) 834 epidemiology 832–3 handling of corpses  835 legal aspects of corpse examination  835 general aspects  832 medical and personal history  582 multimodal approach conventional autopsies  836–7 external examinations  836 minimal invasive autopsy  836 post‐mortem computed tomography  836 post‐mortem findings central nervous system  837–8 heart 838–9 kidney 839–40 liver 840–1, 841 post‐mortem computed tomography  837 respiratory system  838 post‐mortem stability  835 risk factors  833 SARS‐CoV‐2 positive deaths  836 virus, characteristics of  833 COX‐2 369 cranial sutures  281 craniofacial superimposition  1564–6, 1566 cranium fracture see skull fractures racial groups  283 sex determination  275 creatinine 878 cremation 151 examination of body in  153, 154 history 151 modern day  151 process of  152–3, 152 remnants of  153 cresyl violet  418

1650

Index

U

R

TO

U

TR IB

O

C

R

FO

O

N

LY

pharmacokinetics 1237 toxicological analysis  1428 diplopia  1347, 1349 diquat 1282 direct current  640 disaster victim identification (DVI)  291– 7 CBRNE weapons  296 dactyloscopy 291 DNA typing  292 documentation 293, 294, 295–6 forensic molecular biology standards  295 forensic odontology standards  295 international cooperation  292–4, 292 quality management  297 radiology standards  296 standardisation 294–5 team structure  294, 294 dismemberment 767, 768 dissecting aneurysm  426 distribution  1155, 1160–1 drug reservoirs  1160–1 protein binding  1160 dithiocarbamates 1283 diuretics 1286 Divergy, Alphonse  11 diving accidents  576–8 diving types  577–8, 577 breath‐hold diving  577 diving with breathing apparatus  577–8, 577 saturation diving  578 investigation of  582–4, 583 dive equipment  584 dive profile and history  583 environmental conditions  583 medical and personal history  582 main causes of death  581–2, 582 medicolegal autopsy  584–8 diatom test  586 external examination  584–5 forensic imaging  587, 588 gas detection  584–5 histology 585 internal examination  586 molecular autopsy  586 toxicology 586 physiopathology 578–81 decompression illness  580 decompression sickness  579–80 diving gas problems  580 pulmonary barotrauma and arterial gas embolism 578–9 diving response  558 DMPS 1111, 1117

SE

clinical symptoms  1361–2 drivers’ licences  1367 driving fitness  1361–4 driving performance  1366–70 patient education  1362 treatment 1364–6 insulin 1365–6 new agents  1366 oral agents  1364–5 diabetic coma  882, 874 glucose  874, 875 haemoglobin HbA1c 875–6 ketone bodies  776, 876 lactic acid  875, 876 urine examination  876 diabetic foot  1364 diabetic ketoacidosis, acetone/isopropanol blood levels  1185 diabetic nephropathy  1363 diabetic neuropathy  1363 diabetic retinopathy  1347–8, 1363 diacetylmorphine see heroin Diagnostic and Statistical Manual of Mental Disorders (DSM)  660, 707, 1059 diapedetic haemorrhage  350 diatom test for diagnosis of drowning  566–9, 567, 569–70 diving accidents  586 false positives  568 sample preparation  568 diazepam chemical structure  1242 and driving fitness  1426, 1426 metabolism 1244 pharmacokinetics 1237 toxicological analysis  1418, 1419 dichloromethane 1273 dicobalt edetate  1111 diethyl ether  1208, 1271 respiratory toxicity and exposure threshold 1273 diethylene glycol  1186–7, 1271 metabolism 1176 toxicity 1176 diffuse axonal injury  399, 416, 951 Digitalis purpurea (foxglove)  1277 Dignitas 1605 Dignitate 1606 digoxin poisoning  1110 dilated cardiomyopathy  810, 813 diminished responsibility  315–16, 1067–9 diphenhydramine chemical structure  1246 and driving fitness  1428, 1431

N

defibrillation marks  140 deficiency of will  1591–2 dehydration  376, 676–7 elder abuse  1015 vitreous humour pattern  677, 677 delirium, excited  723–5 delta sign  209 delta‐9‐tetrahydrocannabinol (THC)  788, 760, 1197 absorption 1198 distribution 1198–9 and driving fitness  1379, 1381 elimination 1199–200 metabolism  1197, 1199 plasma concentrations  1198, 1199 stability 1140 toxicological analysis  1417, 1419 dementia 77 Demirjian’s stages of third molar mineralisation 1030–1, 1031 dendrites, staining methods  418 Denmark no insanity defence  1067 presentation of psychiatric evidence  1062 statutory limits for alcohol levels  1394 dental age estimation adolescents 1030, 1030, 1033 Demirjian’s stages of third molar ­mineralisation  1030–1, 1031 dental charts  1575, 1576, 1577 dentistry see forensic odontology desferrioxamine 1111, 1117 desloratadine, and driving fitness  1428, 1433 determination of death  60–7 apparent death  61–2 dexchlorpheniramine, and driving fitness  1432, 1428 dexmedetomide 1247 pharmacokinetics 1237 diabetes acute complications  1361–3 hyperglycaemia 1362 hypoglycaemia 1362 chronic complications  1364 diabetic foot  1364 hypertension 1363 macroangiopathy 1363 microangiopathy 1363 neuropathy 1363 vascular disease  1363 classification 1359–61 gestational 1361 type  1 1360–1 type  2 1361

1651

Index

N

O

C

R

FO

O

N

LY

level of evidence  1330 medical aspects  1330 vision/visual fields  1345–52 effects of  1353 older motorists  1351–2 post‐eye surgery  1351 vision problems  1351–2 visual field disorders  1350–1 driving skills  1316 driving under influence of alcohol (DUIA)  1091–3, 1188–9, 1188, 1381 alcohol consumption and crash risk  1397–8 driver fatalities  1400–1, 1141 laboratory studies  1399–400, 1399, 1400 roadside surveys  1398–9 amounts of alcohol consumed  1397–8 characteristics of offenders  1395 age and gender  1395–6, 1364, 1395 blood alcohol concentration  405, 409–10, 1101, 1168, 1379, 1396–7 recidivism 1397, 1397 trends over time  1397, 1397 drug combinations  1100–1, 1111 medicolegal determination  1389–95 blood alcohol analysis  1390–2, 1390, 1391, 1394 breath alcohol analysis  1392, 1392, 1394 statutory alcohol limits  1394–5 total amount consumed  1396 driving under influence of illegal drugs  1081, 1196–209, 1200, 1381, 1409–19 amphetamines  1381, 1414 cannabis  1381, 1411–13 cocaine 1414 epidemiological data  1410–1, 1411 forensic toxicological analyses  1416–18, 1417–19 hallucinogens 1415 heroin and opioids  1413–14 mixing drugs  1416 roadside drug testing  1411 driving under influence of prescription medicines  1322, 1382, 1423–34 antidepressants 1430, 1426 antihistamines 1413, 1428 anxiolytics 1423, 1426 hypnotics 1424, 1425 drowning 355, 355, 556–73 children 904 cold shock  588 contributing causes  470–1 disposal of corpse in water  572 diving response  588

U

TO

R

U

SE

doping  1095, 1134, 1284 anabolic steroids  1285, 1286 available matrices  1284, 1286 banned substances and methods  1284 black‐market/non‐approved products  1290, 1291 health issues and fatalities  1290 peptide hormone analysis  1290 sampling procedures  1284–5 dothiepin, and driving fitness  1427 doxepine, and driving fitness  1427 doxylamine chemical structure  1246 pharmacokinetics 1237 dried blood spot sampling technique  1143, 1443 driving aptitude  1409 see also driving fitness driving behaviour  1336, 1413 Keskinen’s model  1318, 1321 Michon’s model  1317, 1319 driving capability  1315–16 drug testing  1419 driving fitness  1315–24, 1409 assessment 1322–4 detection of unfit drivers  1322 preconditions for driving licence  1329 cardiovascular disease  1329–41 arrhythmias 815–16, 816, 1333–8 coronary heart disease  1332–3 heart failure  1340–1 risk of harm formula  1331–2 syncope 1338–9 deaths associated with driving  1089 definition 1330 diabetes 1359–60 epidemiology 1375–80 analytical 1377–8 causal factors  1376, 1376 descriptive 1375–7 drugs 1380–1, 1381 quality assurance  1378 studies in  1378–9, 1379 epilepsy 1355–7 factors affecting age 1384–5 alcohol see driving under influence of alcohol character deficiencies  1385 diseases and indispositions  1383–4 illegal drugs see driving under influence of illegal drugs prescription medicines see driving under influence of prescription medicines

TR IB

DMSA 1111, 1117 DNA analysis  46, 1497–543 body fluid identification  1499–500 contamination 1497 disaster victims  304 DNA typing  1502–9 history 1502 kits 1504 mixture interpretation  1508–9, 1509 PCR amplification  1503–5, 1503, 1505 STR polymorphisms  1502–3, 1502, 1505–6, 1506–7 evidence collection  1499 kits 1499 extraction and quantitation  1500 forensic DNA databases  1521–2, 1522 haplotypical markers  1517–21, 1519, 1520 mitochondrial DNA  1519–20, 1520 Y chromosome‐specific STR loci  1517–19, 1518, 1519 ISO/IEC  17025 47–9 mixture interpretation  1508, 1509 paternity testing  1525–30 autosomal markers  1526 biostatistics 1528–9 male and female lineage markers  1526–8, 1526, 1528 quality assurance and ethical considerations 1529–30 population genetics  1510–12, 1510 SNPs 1513, 1528 stain analysis  1497–43 DNA profiling  1521 DNA transfer in shooting incidents  1524 doctor–patient relationship 1584, 1589 documentation autopsy report  193–7 child abuse  984 crime scenes  161 death certificate see death certificate disaster victim identification  293, 294, 295–6 errors in  1589 human rights violation  27 laboratory samples  46 domestic violence  318 aetiology 982 cycle of  993 diagnostic procedures  994 effects and consequences  991–3 gay and lesbian relationships  1001–3 and HIV  992–3 see also child abuse; elder abuse

1652

Index

U

R

TO

U

ear bleeding from  392, 655 tympanic membrane rupture  655 ear pulling  972 earthing/grounding injuries  642, 642 ebastine, and driving fitness  1428, 1433 ecstasy see amphetamines Ecuador 34 Egypt 4, 5, 34 elder abuse  1005–15 abandonment 1010 clinical evaluation  1010–12 legal capacity  1011 physical examination  1011–12 psychological, neurological and cognitive 1011 definition and scope  1005–7, 1006 financial exploitation  1010 forms of maltreatment  1007 global prevalence  1005 medicolegal investigations  1012–15 decubitus ulcers  1013–15, 1013, 1014 forensic autopsy  1012 forensic neuropathology  1015 starvation, malnutrition and dehydration  1015 violent homicide  1015 mimics of  1010 accidental trauma  1010

TR IB

O

C

R

FO

O

N

LY

chronic disease  1010 self‐neglect 1010 neglect 1009–1010 non‐physical 1010 physical 1007–9 blunt force injuries  1007–1010, 1008, 1009 thermal burns  1009 screening questions  1011 sexual 1009 electric current  640 electric eels  651 electric fences  645 electrical excitability of muscle  92–4, 93, 94, 94 electrical injuries  376, 379, 640 auto‐erotic 505 basic concepts  640, 640 electrocution 639–61 lightning  379, 652–61 electrical torture  38 sequelae 39 electrocution 639–61 atypical 643–9 earthing/grounding injuries  643, 643 electric fences  645 high voltage injuries  643, 644, 645 let‐go phenomenon  640 microwave burns  643 skin burns  641–3, 641–3 standard operating procedures  649–61 case history  649 postmortem examination  650 scene investigation  649–50 special procedures  650–1 startle reactions and reflex movements  641 stun guns and electro‐shock stun belts  645 Tasers 645–6 typical 639–3, 641–3 in water  651 bath 652–3 electric eels  651 swimming pools  651 underwater welding  651–2 electroencephalography 961 electrolyte imbalances  851 drowning victims  472 electron microscopy in drowning victims  566 electro‐shock stun belts  645 elimination rate constant  1157 ELISA  273, 365, 1133 embalming postmortem injuries  141 and specimen collection  1146 embolism  337–8, 351–52, 352 air see air embolism amniotic fluid  339, 351

SE

drug–drug combinations  1101–2, 1102 drug‐facilitated sexual assault  1094 drug screening  1129–3 analytical methods  1133–4 confirmation tests  1130 instability of drugs during  1146–8, 1147 instrumentation 1131–2, 1132, 1132 isolation step 1130–1 method performance  1134–5, 1135 target substances  1130 drug stability in samples  1138–50 blood from living individuals  1139–40 definitions 1138–9, 1138, 1139 evaluation of  1148–50 postmortem specimens  1141–3, 1142 processing and analysis  1146–8, 1147 sampling artefacts  1143–5, 1145 storage 1145–6, 1146 urine from living individuals  1140–1 drugs, driving under influence of  1089, 1200–4 dry drowning  559, 562, 565 Dunant, Henri  35 dural haematoma  442–2 Dusky test of competence  1062 duties of forensic medicine  19–22, 20 dysphagia, bolus death  545, 454

N

drowning (continued) dry  588–9, 564 future studies  572–3 homicide 572 incidence 559 laboratory diagnosis  566–9 aquaporins 569 artificial tracers  566 controversies 568 diatom method  566–9, 567, 569–70 electrolytes 569 exogenous substances  566 false positives  568 microbiological tests  569 molecular autopsies  569 sample preparation  568 natural deaths in water  571 organ effects  557 pathophysiology 577–9 postmortem changes advanced 561 early 560, 560 postmortem diagnosis  562–70 analytical morphometry  566 electron microscopy  566 forensic radiology  569–70 macroscopic changes  562–4, 563, 563 microscopic changes  565–6, 565 see also laboratory diagnosis, above postmortem submersion time  560, 563 pulmonary emphysema  345 sequence of events  529 site of death  561–17 cadaver buoyancy  561, 562 cadaver drifting  561–62 suicide 571–2, 572 victim identification  559 drug addiction  77, 783–92, 1196–209 autopsy findings  786–7, 786, 787 body packers  784–5 cause of death  787 chyme aspiration  546–7, 546 classic 783, 783 intravenous drug users  784, 784 legislation 783 manner of death  787 mechanisms of death  788–2, 789 prevalence 792 puncture wounds  787 scene examination  785 Western countries  784 withdrawal 784 see also illegal drugs; and specific drugs drug‐associated deaths  748–50 classification 749 types of error  749

1653

Index

N

O

C

R

FO

O

N

LY

exitus in tabula (death on operating table)  747–8 complications 748 occupational groups  747 type of intervention  748 expert evidence  763–770 alcohol‐related deaths  772–83 drug addiction  783–92 Mafia‐related deaths  792–8 physical activity after injuries  765–7 postmortem mutilation  767–70, 768, 769 priority and order of injuries  763, 764 survival time  765–7 expert witnesses  34; see also expert evidence explosive injuries  492 autopsy 495 car bombs  798 injury types  493–4, 494 mechanisms of injury  492 physics of explosion  492 reconstruction 495 exposure deaths in alcoholics  780 external examination  69–89, 70 cause of death  70–80, 71, 77, 314, 327–46, 328 autopsy findings  80–1 statistics 80, 77, 78 checklist 86 conditions/time of death  87 forensic autopsy  185 identification of corpse  87 postmortem changes  87 special circumstances deaths behind steering wheel  85 deaths by poisoning  85–8, 85, 86 deaths in bathroom  84 deaths in police custody  84 deaths in prison  84 deaths in psychiatric hospitals  84–5 multiple corpses  86 unexpected deaths in hospital  83 systematic 87–8 external genitalia  188, 193 external haemorrhage  350, 351 external lesions  186, 517 extrapyramidal syndrome  1126 extra‐uterine pregnancy  698–9, 699 extremities blunt force injuries  404–6 examination of  188 fractures in children  948, 948 physical activity after injury  769 sharp force injuries  455, 455–6 see also specific areas eye drying of  134, 134 injuries in child abuse  978

U

TO

R

U

SE

ethanol see alcohol ethchlorvynol 1247 chemical structure  1246 pharmacokinetics 1237 ethnicity and age estimation  1038–41 eruption/mineralisation of third molars 1041 sexual maturation  1042 skeletal maturation  1040–1 identification of  283–4, 283 ethyl acetate  1271 respiratory toxicity and exposure threshold 1272 ethyl glucuronide  1141, 1441, 1441 in hair  1445–7, 1446 as marker of alcohol abuse  1441, 1448, 1444–5 toxicological analysis  1419 in urine  1444–5 ethyl sulphate, as marker of alcohol abuse  1441, 1441, 1444–5 ethylene glycol  1168, 1184–6, 1270, 1270 analysis of  1171 metabolism 1176, 1185 poisoning 1111, 1185 toxicity 1176, 1185 European Charter of Fundamental Rights  1613 European Committee for the Prevention of Torture (CPT)  36 European Convention for the Prevention of Torture and Inhuman or Degrading Treatment or Punishment see European Committee for the Prevention of Torture European Convention for the Protection of Human Rights and Fundamental Freedoms  1590, 1602, 1603, 1607, 1613, 1615 European Convention on Human Rights  646, 1069, 1093, 1616 European Court of Human Rights  28, 1069, 1095, 1608 European Union Directive  2004/23/EG  258 European universities, foundation of  7 euthanasia active 1606 indirect 1606 passive 1606 evisceration 189–90, 189 exchange transfusion  1112 excited delirium  723–4 excretion  1155, 1164 exenteration cranii  472 exhumations 28–30, 28–30

TR IB

bone marrow  353 bullet  353, 490 cell/tissue 351 cerebral arterial gas embolism  578, 581, 582 fat  193, 338, 338, 351, 353 thromboembolism 338, 353 emedastine, and driving fitness  1428, 1432 encephalopathy in children  942, 742, 948 endocardial fibroelastosis  810, 899, 902 endocarditis, intravenous drug users  784, 784 endothelial cells, staining methods  418 endotoxic shock, post‐abortion  699 England and Wales diminished responsibility  1067 fitness to plead  1062 see also United Kingdom Englaro, Eluana  1608, 1609 enterohepatic circulation  1159, 1160 entomology see forensic entomology environmental conditions forensic DNA laboratories  48 forensic toxicological laboratories  49 environmental temperature  113 eosinophilic myocarditis  814, 815 epidemiology 1100 analytical 1377–8 cataracts 1346–7 child abuse  725–6 descriptive 1375–7 driving fitness  1375–80 heart failure  1340–1 hypothermia  888, 630 medical errors  736–8 postmortem mutilation  768 self‐harm 1021 SIDS 894–5, 894, 895 stillbirth 684 epidermal growth factor  368 epidural haematoma see haematoma, extradural epidural haemorrhage  333, 394, 395, 396, 423, 423 children 434 shaken baby syndrome  438, 904 epiglottitis 900 epilepsy 943 and driving fitness  1355 and general road safety  1357 and traffic accidents  1464–5 epiphyses, fusion of  275, 276 equipment, laboratory  46 Eschweiler–Clarke reaction  1146, 1146 esmirtazapine, and driving fitness  1425 eszopiclone 1244, 1244 pharmacokinetics 1237

1654

Index

FO

R

U

R

TO

U

TR IB N

C

O

N

LY

black powder firearms  461 captive bolt devices  462 non‐lethal projectiles  462–3 pen guns and signal guns  462 stud/nail guns  462 zip guns  462 firing marks  484 first‐order elimination  1157–8 first pass metabolism  1159 fitness to drive see driving fitness fitness to plead  1062 five‐point tie  39 flashfire deaths  618 floating injuries  137, 618 flotation test of lungs  354, 689 fluid imbalance  881 flumazenil 1111 chemical structure  1242 pharmacokinetics 1237 flunitrazepam (Rohypnol)  932 chemical structure  1242 and driving fitness  1424, 1425 pharmacokinetics 1237 toxicological analysis  1419 fluoride, sample requirements  1115 fluorine 1268 respiratory toxicity and exposure threshold 1272 fluoroacetate poisoning  96 fluoxetine 789 and driving fitness  1427, 1431 flurazepam, and driving fitness  1424, 1425 Fodere, François Emanuele  10 fomepizole  1111, 1174 forearm fractures  977 foreign bodies aspiration of  545, 546, 904 vaginal 926 forensic anthropology see anthropology/ osteology forensic autopsy see medicolegal autopsy forensic ballistics see ballistics forensic DNA databases  1521–2, 1522 forensic DNA laboratories, ISO/IEC  17025 47–9 forensic entomology  123–32 blow fly (Lucilia sericata)  127–8, 127 case reports  130–2, 130, 131 collection of insects  128 estimation of post‐mortem interval  128–30 identification of insects  128 storage of insects  128 succession 125–7, 126 see also specific species

O

facial reconstruction  286–7, 286, 287 facial trauma  393–4, 293, 294 road traffic accidents  1461 factitious disorders  877 factor IX deficiency  980–1 falanga  37, 40, 320 sequelae 39 falls 191 alcohol‐related 780, 780 toddlers 958–9 Fannia manicata  126, 126 Fannia scalaris  127 fat embolism  193, 338, 338, 351, 353, grading 353 fatal toxicity index  1089, 1090 fatty acid ethyl esters  1441, 1441 as marker of alcohol abuse  1445, 1448 febrile non‐haemolytic transfusion reaction 746 females see women femoral blood  871 femoral fractures  72, 330 femoral prosthesis  284 fentanyl 1204 pharmacokinetics 1255 fetishism  707, 713 Fettwachs see saponification fetus/fetal autopsy 194 maceration 688, 688 stillbirth 683–4 fexofenadine, and driving fitness  1433 fibroblast growth factor  368 fibronectin  365, 370, 370 Fidelis, Fortunatus  6, 8 fingernail scratches  381, 382 fingerprints 29 disaster victims  293, 294 fingers amputation 377 torture 38 fingertip bruises  384–5, 384, 973 Finland, statutory limits for alcohol levels 1394 fire gases  616 fire injuries  376, 376 burns  379, 607 charring  270, 618–19, 619

checklist 619–20 postmortem 136, 138, 611, 614, 612–13, 612, 616 smoking in bed  779–80, 779 see also specific injuries firearms 459–5, 460 acoustics 490 ammunition 463–6 blank and tear gas cartridges  465–6 bullets 464, 464 calibre 464, 464, 465 cartridges 463, 463 primer and propellant  463–4 shotgun 465, 465 special 465 barrel 459 breech and lock  459–60 bullets 464, 464 deformation 469, 469 determination of  484–5 deviation 467 fragmentation 467, 469, 470 infectivity 490 lead poisoning  491 mass and muzzle velocity  464, 464, 465 penetration 467 reflection 467 ricochet 467 yawing  467, 469, 470 classification 460–1, 460, 461 pistols  460, 461 revolvers 460–1, 460 rifles 461, 461 shotguns 461, 461 determination of  484 firing marks  484 muzzle imprint  484 exterior ballistics  466 angle of incidence  466–7 environmental conditions  466 maximum range  466 trajectory 466 incidents involving see shooting incidents interior ballistics  466 muzzle phenomena  466 recoil 466 intermediate targets  467–7 limiting angle  467 ricochet and yawing  467 wound ballistic consequences  467 muzzle modifications  490 sights 460 silencers 490 special types  461–2 air weapons  461–2

SE

eye (continued) petechiae 187 tache noir  104, 105, 134 see also visual field defects; visual problems eye movement disorders  1348, 1352 25 F  9 365

1655

Index

N

O

C

R

FO

LY

Frank, Johann Peter  12 freedom of therapy  1598 freon 1208 front‐end collisions  1461–3, 1462 frontal sinuses, in victim identification  1563 frost erythema  378, 631, 633 fungal growth  108 fungal myocarditis  814 fungicides 1283 dithiocarbamates 283 hexachlorobenzene 283 pentachlorophenol 283 phthalimides 283

O

N

G 16/1 365 GABA analogues  789, 788 gagging 543 galactosaemia 961 gamma‐butyrolactone 1209 gamma‐hydroxybutyrate  932, 1208–9 toxicological analysis  1418 Gardner–Diamond syndrome  980, 982 garrotting 529 see also ligature strangulation gas and dust explosions  494 gas chromatography–mass spectrometry  1132, 1147 gas embolism arterial 578, 582 post‐abortion 701 venous 579 gas–liquid chromatography  1169, 1390 gases in body cavities diving accidents  586–8 subcutaneous expansion in gunshot wounds 478, 479 fire 616 inhalation 300, 355, 556 occupational exposure  1272, 1272–3 partial pressure  579 respiratory toxicity  1273 toxicology 1266 see also specific gases gastric contents specimen collection  1122 and time of death  120–2, 121 gastric emptying  121 gastric juice, regurgitation of  140, 141 gastroenteritis, and sudden infant death  901 gastrointestinal tract absorption from  1159–60 drowning victims  565 multiple organ failure  345–6, 345 vital signs  355 gastrothorax 336–7

U

TO

R

U

SE

disaster victims  293 ISO/IEC  17025 53 forensic physicians  646 forensic psychiatry  1057–72 civil law competencies  1060–1 consent to treatment  1061 legal competency  1060 retrospective civil assessments  1060–1 testamentary capacity  1060–1 criminal law competencies  1061–3 competence to stand trial  1061–2 determination and consequence  1063 duration of hospitalisation  1063 fitness to plead  1062–3 indications for hospitalisation  1063 no fitness to plead/competence to stand trial 1062 treatment refusal  1063 criminal responsibility  1063–70 determination and consequence  1069 diminished responsibility and overwhelming emotion  1067–9 insanity standards  1064–7 no insanity defence  1067 preventive detention  1069 rehabilitation and security  1069 definitions 1057–8 examinations 1059–60 formal education in  1071 inquisitorial vs. adversarial legal systems 1058–9 landmark court cases  1072 legislation 1072 forensic reconstruction explosive injuries  494 shooting incidents  474, 477 forensic security hospitals  474–87 forensic toxicological laboratory standards see laboratory standards fourth rib method of age estimation  278–80, 279 fractures bending 324 burst 1462 child abuse  974–9, 974, 978 compression 326 healing 1009 mandible 394 Messerer’s 1456, 1458 pelvis 403–6 ribs 336, 336, 400, 401 shell 325 skull see skull fractures vertebrae 403, 403 France 10–11 statutory limits for alcohol levels  1384

TR IB

forensic genetics  30 forensic imaging  207–15 age estimation in adolescents  1028–41 CT 208–11, 209–11 CT angiography  211–2, 212 dental charts  1575, 1576, 1577 disaster victims  296 diving accidents  586, 587 drowning victims  569, 570 MRI 213–14, 213 non‐accidental head injury  961, 963 non‐ionizing procedures  1039 SIDS and SUDI  905 standards 296 surface scanning  216–18, 217 forensic medicine autopsy standards  43–5 as book science  7–10, 9 current problems  15–16 definition 3–4 development of  4 as experimental science  10–15 forensic neuropathology  412–44 brain 416–17 cell and tissue reactions  413–15 central nervous system  412 elder abuse  1015 head and spine  418–33 inflammation 416 paediatric 433–9 regenerative capacity  416 spinal cord and cervical spine  417–18 staining methods  418 tissue decay and reactivity  415 forensic observers/inspectors  30–1 forensic odontology  1575–83 age estimation  1577–9, 1578, 1579 adolescents  1031, 1042 and ethnicity  1041 older people  1042 bite marks  1580–3, 1581, 1582 dental charts  1575, 1576, 1577 dental investigation  192 dental schemes  292 dentition development  1578 disaster victims  293, 294 guidelines 295 identification of unknown persons  1579–80, 1579, 1580 decayed missing filled teeth (DMFT) index 1580, 1580 pink teeth in water‐related deaths  560 root transparency  282 third molar eruption/ mineralisation  1041 forensic pathology

1656

Index

U

TO

R

U

Haberda, Albin  12, 506 haematological disorders  980–1 haematoma chest 376 dural 442–2 epidural 423, 423 extradural  333, 394, 395, 396 heat‐induced 614, 614 periorbital  390, 392 scalp 366 streak‐like 384–5 subdural 333, 391, 395, 396, 396, 397, 423, 423 temporal muscle  367 tram‐like 384, 384 haemodialysis 1112 haemofiltration 1112 haemoglobin HbA1c 875–6 haemoperfusion 1112 haemopericardium 191, 331, 332 haemoperitoneum  72, 332, 405 haemophilia 980–1 haemorrhage  186, 350–1, 351, 352 catastrophic 300–4 diapedetic 350 external 350, 351 internal 332–3, 332, 333 intracerebral 430, 430, 472 intracranial 394–7, 395–7, 423–4, 423, 425 petechial see petechial haemorrhage postmortem blood loss  333–4 retinal  436–7, 939

TR IB

O

C

R

FO

O

N

LY

scalp 190 temporal bone  564 thorax 191 ‘walk and die’ 334 haemorrhagic–dysoric syndrome  522 haemothorax 332, 336, 492 hair 187 drug stability in  1144, 1145 ethyl glucuronide in  1444–5, 1448 specimen collection  861, 1145 hair pulling  972 hallucinogens  789, 790–1 and driving fitness  1415 hand grenades  494, 494 hanging  187–8, 522 accidental 528, 535 alcohol‐related 779 children 442 complete 523–4, 524, 525 definition 522 extravasation 525 frequency/occurrence 522–3 homicide 527–8 incomplete 523, 523, 524, 529 ligature marks  525, 525 Mafia‐related deaths  769–70 observations and time relationships  516, 516 pathomorphology 524–6, 524, 352, 529 petechial haemorrhage  352 physical activity after  765 strangulation devices  523 suicide  523, 528 haplotypical markers  1517–21, 1519, 1520 mitochondrial DNA  1519–20, 1520 Y chromosome‐specific STR loci  1517–19, 1518, 1519 hashish see delta‐9‐tetrahydrocannabinol head diving accidents  587 external examination  584–5 forensic neuropathology  418–33 internal examination  190 head‐down asphyxiation  539, 539 head injuries  390, 390–400 children accidental 958–9 non‐accidental 939–51 facial trauma  393–4, 293, 294 forensic aspects  419–20 intracranial haemorrhage  394–7, 395–7, 425–6, 426, 425 mechanical 418–33 physical activity after  765 physical trauma  418

SE

ethylene glycol see ethylene glycol propylene glycol  1271 glycoprotein, P‐, transporter  1159–60 glyphosate 1282 golden hour  334 gonorrhoea 931 granulation tissue  368 granulomatous myocarditis  814, 815 grazes see abrasions Great Britain see United Kingdom Greece, statutory limits for alcohol levels  1394 Greulich–Pyle radiographic atlas  1030, 1040 grief 249 gripping, forceful  972 ground fault interrupters  652 growth factors  369, 369 growth hormones  789, 792 gun cleaning accidents  490 gunshot wounds see firearms; shooting incidents Gustafson–Johanson procedure  282

N

gene doping  1286 genetic markers  1502, 1531 genetic testing  1598–9 genitalia abrasions 925 anatomy 922 anogenital lesions  921–30, 922–9, 925–6 bruising 922–923, 925, erythema 925 external 188 female dissection of  769, 770 hymen 923, 924, 926, 928 labial fusion  926 lacerations 925, 928 male, injuries to  712–13, 927 oedema 925 self‐injuries  454, 1021 Tanner stages of development  1041 geographical origin  287 Germany competence to stand trial  1062 consent to treatment  1061 diminished responsibility  1067 forensic society training  1071 insanity standards  1064–5 legal competency  1060 rehabilitation and security  1069 retrospective civil assessments  1060–1 statutory limits for alcohol levels  1394 testamentary capacity  1060–1 Transplantation Act (1997) 257–9 transplantation procedures  257–9 gestational diabetes  1361 ghost neurons  440 giant cell myocarditis  814, 815 glare sensitivity  1347, 1352 Glasgow Coma Scale  327 for children  943 glass injuries  389 glaucoma 1348 glial fibres, staining methods  418 gliding contusion injury  420, 430 glucocorticosteroids 1286 glucose blood levels  875 metabolism 873–4, 874 glufosinate 1282 glutethimide 1243, 1243 pharmacokinetics 1237 glycerol 1168 glycols 1270–71, 1270 butylene glycol  1271 diethylene glycol  1176, 1186, 1270, 1271

1657

Index

N

O

C

R

FO

O

N

LY

hormones, in doping  1285, 1290 hospitalisation on psychiatric grounds  1063 duration of  1063 hospitals forensic security  474–87 unexpected deaths in  83 hot‐air balloon accidents  549 HSPG 365, 370 human biomonitoring values  1258 human papillomavirus  931 human remains display of  1615–18 PMI see postmortem interval recovery of  268–71, 269, 270 use without explicit consent  1618 see also corpses human rights  27–42 ad hoc and permanent tribunals  28 autopsy protocols  31–2 clinical forensic examinations  32 courts 28 exhumations and autopsies  28–30 expert witnesses  34 forensic documentation  32–4 forensic observers/inspectors  30–1 international expert network  35 legislation 27 torture see torture training and research  34–5 humerus fractures  977 hunting accidents  489 hydrocarbons 1269–70 aliphatic 1269 aromatic 1269 chlorinated 1269–70 hydrocephalus 415 children 435 hydrochloric acid  616 hydrocodone 1254 toxicological analysis  1418 hydrocyanic acid  85, 616 hydrogen cyanide see cyanides hydrogen sulphide  1269 poisoning 85 lividity 96 respiratory toxicity and exposure threshold 1272 toxicity 1269 hydromorphone 1255 toxicological analysis  1418 hydroxizine, and driving fitness  1432, 1428 hydroxocobalamin 1111 5‐hydroxyindolylacetic acid  1441, 1441, 1444 5‐hydroxytryptophol 1441, 1441, 1444

U

TO

R

U

SE

predisposing factors  624 types of  624 heat wave  622 height estimation  283, 284 helicopter accidents  1483 helium, effects of  580–1 Henry’s law  578, 579 hepatitis 77 alcoholic 777 intravenous drug users  787 herbicides 1281–2 bipyridyl derivatives  1282 chlorophenoxy compounds  1281–2 organophosphates 1282 heroin 1201–3 chemical structure  1254 and driving fitness  1413–14 metabolism 1163, 1202 pharmacokinetics 1255 plasma concentrations  1203 stability  1139, 1140 hexachlorobenzene 283 hexane 1273 hide and die syndrome  630 high excitation  881 high resolution mass spectrometry  1132 high velocity missiles  471 high voltage injuries  643, 644, 645 hinge fracture of skull  389 Hippocratic Oath  1593, 1594 histology, diving accidents  587 history cremation 151 forensic medicine Italian town charters  7 Justinian enactments  6–7 Near East and China  4–6 timetable of  5–6 HIV/AIDS 930–3 and domestic violence  984 intravenous drug users  784 hoarding behaviour  1010 Hofmann, Eduard von  12, 13, 15, 506 hogtie prone restraint  721 Holzer blisters  85 homicide 317–19, 318, 320 homonymous hemianopsia  1351 homosexuals, violence against  1001–3 characteristics 1001–2 statistical problems  1002 ‘honour killings’ 319 HOPE (Heart Outcomes Protection Evaluation trial) study  1333 hormone antagonists  1285

TR IB

railway accidents  1477 sharp force  454–7 shooting incidents  329, 460, 460, 473–4 incapacitation after  473 skull fractures  190, 390–3, 390–3, 421–2, 421, 422 see also brain injuries health hazards, crime scenes  161 health insurance law  1600–2 heart alcohol effects  777 blunt force trauma  401, 402 cardiac conductive system  195 cardiac syncope  1338–9 congenital heart disease  74, 809 COVID‐19 clinical aspects and macromorphology  838–9 micromorphology:839 dissection of  190 intraseptal haemorrhage  656 malformations 902 multiple organ failure  345–6, 345 rupture 805, 818 sepsis 342, 342 sharp force injuries  766, 766 SIDS 899–90 specimen collection  1122 weight loss in starvation  674 see also sudden cardiac death heart failure  1340–1 causes and co‐morbidity  1340 classification 1340 driving restrictions  1341 epidemiology 1340 mortality and prognosis  1340–1 New York Heart Association Classification  1340, 1341 types of  1350 heat collapse  622, 623 heat cramps  622, 623 heat exhaustion  623 heat haematoma  613, 615 heat injuries  607–22 burns 607–17 causes of death  617–22 causes and symptoms  622 diagnosis 623–4 fire duration and charring  618–19, 619 hyperthermia see hyperthermia scalding  379, 621, 621, 621 heat rigor  618 heat shock  618 heat stroke  623, 622

1658

Index

TR IB

U

TO

R

U

O

C

R

FO

LY

N

SE

iatrogenic maternal death  703–4 ibuprofen  789, 792 ICAM‐1 365, 369 identification of corpse see victim identification identifying features  186 identity SNPs  1513 idiomuscular pad  87, 91, 92, 351 idiopathic thrombocytopenic purpura  980–1 illegal drugs  1081, 1196–209 and driving fitness see driving under influence of illegal drugs screening for see drug screening see also drug addiction; opioids; toxicology; and specific drugs imaging see forensic imaging; and specific techniques imipramine, and driving fitness  1425, 1427 immediate death (within minutes)  330–4 immersion time  123, 589–60 minimal, estimation of  560, 592 validity and reliability  592 water temperature and putrefaction  591–2, 591 immunisation, and SIDS  959 immunoassay  1132, 1133 immunohistochemical markers of wound age 365 immunohistology, wound age estimation  855 impact (acceleration/deceleration) brain injury 949 impedance 640 implantable cardioverter defibrillators  1334–8 risk of arrhythmia occurrence  1337–8 risk of arrhythmia recurrence  1335 risk of harm to patients/ bystanders  1336–7 risk of syncope  1335 in‐custody deaths  84, 717–19, 718 alcohol‐related 779 death certificate  725 investigation of  724 inborn errors of metabolism, screening for 961 incapacitation after shooting incidents  453–4

incidence 1375 inductively coupled plasma mass spectrometry 1132 infanticide  315, 683–93 autopsy 687 sample taking  692–3, 693 birth‐related changes  688 causes of death  690 diagnosis 683, 683 duration of life  687–8 history 683 incidence 684 investigation of mothers and neonates  686–3 proof of live birth  688–90, 689 viability and maturity  688, 688, 689 infants causes of death  888 mortality 888 sudden unexpected death (SUDI)  893 see also children infarction  77 infection  78, 339 infection screen  960 inflammation 416 brain 416 influenza, pathological findings  814 informed consent see consent to treatment inhalation 353 gases 330, 355 soot 355, 355, 355 inhalation trauma  356, 616 injuries 314 air embolism see air embolism blunt force  329, 332, 376, 376, 380–400 chest 336–7, 336 definition 313 fat embolism  193 head 335 neck 192 postmortem see postmortem injuries priority and order  763, 764 semi‐sharp force  377 sequelae 327–40 sexual 193 sharp force  377, 448 see also specific injuries inquisitorial legal systems  1058–9 insanity standards  1063 insecticides 1276–81 anticholinesterases 1280 carbamates 1280–1 organochlorines 1279 organophosphorus compounds  1280–1 pyrethroids 1281

O

divers 580 hypobaric  512, 547 normobaric/hyperbaric  512, 547–9, 548 hypoxic brain death  511, 513 see also asphyxiation hypoxic ischaemia, and subdural haemorrhage  959 hypoxic sensitivity  511 hypoxyphilia 517

N

hymen 923, 924 abnormalities 926, 928 hyoid bone, dissection of  518, 518, 520 hyperacute cervicomedullary syndrome  949 hyperglycaemia 1362 hypertension, in diabetes  1363 hypertensive heart disease, and traffic accidents 1464–5 hyperthermia 376 definition 622 and SIDS  892 hypertrophic cardiomyopathy  226, 813 and sudden infant death  903 hypnotics see sedatives and hypnotics hypobaric hypoxia  512, 598 hypoglycaemia  874, 877 in diabetes  1362 and driving  1367 factitious 877 and traffic accidents  1464–5 hyposphagma 520 hypostasis 94, 95, 95 drowning victims  560 hypothermia  60, 337, 376 clinical phases  629, 629, 630 cold water immersion  558 epidemiology  888, 630 frost erythema  378, 631, 633 integrating chart for casework  119 localised 630, 630 morphological/biochemical changes  630–1, 631 blood colour  631 core muscle haemorrhages  632 gastric mucosal haemorrhages  631–2, 633, 632 lipid accumulation  632, 634 lividity  96, 632 pancreas changes  632, 634 pathophysiology 627–9 duration of cold exposure  627–8 final cause of death  628 temperature regulation  627, 628 peripheral cold injuries  631 postmortem biochemistry  881 signs and symptoms  629 undressing/hide and die phenomena  630 hypoventilation 511–12 in children  943 hypovolaemic shock in children  743 hypoxia 334 alternobaric 512 of ascent  580 children 441, 442

1659

Index

N

O

C

R

FO

insanity standards  1065 rehabilitation and security  1069–70 statutory limits for alcohol levels  1394 town charters  7

LY

Japan competence to stand trial  1062 diminished responsibility  1068 insanity standards  1065 Joule burns  641–2, 641 jugular vein compression  514 Justinian enactments  6–7 juvenile pornography  933

O

N

Kastle–Meyer test  1499 Kawasaki syndrome  904 Kempe, Henry  970 keratin 5 365 Kerley–Ubelaker method  282 ketamine  789, 790, 932 ketazolam 1237 ketoacidosis, alcoholic  775–6, 775, 1189 ketone bodies  776, 876, 1183, 1184 ketotic coma  874 Khmer Rouge Tribunal  28 kicks 972 kidney blunt force trauma  407 COVID‐19 clinical aspects and macromorphology 839–40 micromorphology 840 multiple organ failure  345–6, 345 specimen collection  1121 weight loss in starvation  674 Kirchhofrosen 94, 95 Kosovo 29, 29, 30, 292 Krebs’ cycle  775, 776

U

TO

R

U

SE

iron deficiency 286 poisoning 1110, 1117 sample requirements  1115 Iscan method of age estimation  278–9, 276 ischaemia, in children  440, 442 permanent global  443, 443 ischaemic heart disease  388, 804–9, 805–806 coagulative myocytolysis  808 contraction band necrosis  344, 807 evolution of infarct necrosis  808 polymorphonuclear leukocytes  807, 807 and traffic accidents  1464–5 wavy fibres  808 ischaemic ulcers  1031 ischiopubic ramus  274 ISO/IEC  17025 47–53 clinical forensic medicine  53 forensic DNA laboratories  47–9 accommodation and environmental conditions 48 analytical testing, calibration and validation 48 documentation 48 measurement traceability  48 personnel qualifications  47–8 quality assurance  44 reporting results  47, 49 samples 48 forensic pathology  53 forensic toxicological laboratories  49–53 accommodation and environmental conditions 49 analytical testing, calibration and validation 49 documentation 52 personnel qualifications  49, 50–1 quality assurance  52 reporting results  52 samples 51, 52 isobutanol, in alcoholic beverages  1181 isocyanates 616 isomorphen/isomegalen diagram  129 isopropanol 1182–4, 1184 analysis of  1171 blood levels  1185 concentration‐time profile  1183 as marker of alcohol abuse  1443–5 metabolism 1176 physicochemical properties  1170 poisoning 1185 toxicity 1176 Italy diminished responsibility  1068

TR IB

insects see forensic entomology; and specific species instantaneous death  329–30, 329, 330 insulin in doping  1290 in treatment of diabetes  1365–6 insurance fraud  1023 intent 313–14 Inter‐American Court of Human Rights  28, 34 interleukin‐1α 369 interleukin‐1β 365 interleukin‐6 365 interleukin‐8 369 internal examination in forensic autopsy 188–2 internal haemorrhage  332–3, 332, 333 International Commission on Missing Persons (ICMP)  30 international courts  27 expert witnesses  34 International Crime Court  27 International Crime Tribunal of former Yugoslavia (ICTY)  27, 28, 34 International Criminal Tribunal for Rwanda 28 international guidelines  36, 43 autopsy standards  43–5 laboratory standards  46–7 Interpol Disaster Victim Identification Forms  186, 559 interstitial pneumonia  821, 900 intervertebral disc haemorrhage  349 intimate partner violence see domestic violence intracerebral haemorrhage  430, 430, 472 shooting incidents  474 and traffic accidents  1464–5 intracranial haemorrhage  394–7, 395–7, 425–6, 426, 425 contusion index  429 see also specific types intracranial pressure, raised, in children  943–4 intracranial vessels injuries to  425–6mechanically induced alterations 429 intracranial volume  413–14 intramyocardial haemorrhage  661 intrathoracic pressure, raised  514 intravenous drug users  784, 784 ion trapping  1122 Ireland, statutory limits for alcohol levels 1394 iris, pharmacological excitability  94, 94

labelling of laboratory samples  46 labial fusion  926 laboratory standards  46–7 anticontamination precautions  47 assessment  46, 51 equipment and measurement  49, 48, 51 evaluation and interpretation  47, 49 ISO/IEC  17025 47–53 labelling and documentation  46 proficiency testing  47 samples  46, 48, 51 validation  46, 49–51 labour law, personal injury assessment 1633–4 dependences 1633

1660

Index

FO

R

O

N

LY

circumstances 529 definition 528 frequency/occurrence 528–9 homicide 528, 530 ligature marks  358 observations and time relationships  516, 516 pathomorphology 529–30, 531 strangulation devices  529 suicide 531 lightning explosive barotrauma  657, 658 lightning injuries  379, 652–60 autopsy 656, 656 mechanism of injury  657, 658, 659 physics 654 scene investigation  654–6, 655 special procedures  656–7, 656 standard operating procedures  644, 649, 659 limbs see extremities limit of detection (LOD)  51, 1134 lipid accumulation in hypothermia  632, 634 liquid chromatography–mass spectrometry  1132, 1147 liquid ecstasy see gamma‐hydroxybutyrate lithium  789, 792 poisoning 1110 sample requirements  1115 liver alcohol effects  776 cirrhosis 777 fatty degeneration  776, 776 blunt force injuries  403, 404 COVID‐19 clinical aspects and macromorphology  840 micromorphology 840–1 multiple organ failure  345–6, 345 sepsis 342–3, 342 specimen collection  1120–1 weight loss in starvation  674 liver function tests  877–8 lividity 94–6, 95, 96 discoloration 96 hypothermia 96, 632 shift with movement  96 stages of  97 time course  97 water‐line 652 livor mortis  62, 87 poisoning 85 SIDS 897 lobar pneumonia  777, 781 long QT syndrome and SIDS  888–9

TR IB

U

TO

R

U

SE

clinical autopsy  193–7 competency see competency definitions 313 diminished responsibility  315 display of human remains  1615–18 drug addiction  783–92 expert evidence  763–70 forensic psychiatry  813 handling of corpses  1615–18 human rights  27 infanticide 315 injuries and causation  314 liability 313 medical errors  745 medical treatment  1587–95 as bodily harm/assault  1588–9 confidentiality 1593–5 medical indication and standards  1589, 1589, 1589 persons involved  1587–8 self‐determination 1590–1 mens rea (intent)  313–14 mental capacity  315 murder and manslaughter  315 personalised medicine  1595–602, 1597 areas of application  1595–6 challenges 1596 confidentiality 1600 data protection  1600 definition 1595–6 freedom of therapy  1598 genetic testing  1598–9 health insurance law  1600–2 informed consent  1598 research 1602 punishment for crime  316 research 21 research on corpses  1610–15 sexual abuse  932–3 thin‐skull rule  316 toxicology 1079–6 Lehrbuch der Gerichtlichen Medizin  12, 13 let‐go phenomenon  640 leucocyte sticking  342, 342 leukaemia 78 levocetirizine, and driving fitness  1428, 1433 lex artis  1588, 1589, 1590 liability  313, 314, 315 apportioning 316 Lichtenberg figures 659, 660 ligature marks hanging 525, 525 ligature strangulation  528 ligature strangulation  187, 380, 528–9, 529 accidental 531

N

C

O

labour law, personal injury (continued) permanent impairment  1633 temporary partial professional impairment 1633 temporary total professional impairment 1633 Lacassagne, Alexandre  11, 767 lacerations  376, 388–90, 389 anal 927 brain 419 children 434 elder abuse  1009 genitalia 622–624, 925 scalp 389 lactic acid  875, 876 Traub’s calculation  875 LADME principle  1155, 1156 laminin 365, 370 lanugo hair  351, 688, 688 laryngeal fracture  335 larynx dissection of  518, 518 obstruction of  544–5, 546 soot inhalation  355 late death (days to weeks/months)  337–40 Le Fort fractures  394 lead 1259–60 exposure 1259 chronic lead intoxications  1259 poisoning 85, 1111, 1117 from retained bullets  491 reference concentrations  1259 sampling requirements  1115 toxicity 1259 toxicokinetics 1259 lead pellets, deformed  486–7 Lebanese Criminal Code  1065 Lebanon insanity standards  1065 leg injuries railway accidents  1477 road traffic accidents  1456, 1456 legal capacity, older people  1011 legal competency  1060 legal highs  1085 legal issues  313–16, 1587–95 actus rea (act/action)  314 apportioning liability  316 assisted suicide and medically assisted suicide 1605–10 organised suicide  1609–10 precedents 1608–9 terminology 1606–8 causation, liability and time  314 child abuse  961

1661

Index

N

O

C

R

FO

maceration 134 drowning victims  560, 560 of fetus  688, 688 McNaughton’s rule see M’Naghten’s rule macrophages, immunohistochemical markers 365 macular degeneration  1349 MADIT (Multicenter Automatic Defibrillatory Implantation Trial) II trial  1337 Mafia‐related deaths  792–8 AK‐47s 795–6, 796 ambushes and massacres  797–8, 798 caprettamento 796–7 hangings 796–7

O

N

LY

epidemiology 736–8 data from arbitration committees  738–40, 729–41, 741 data from Institutes of Forensic Medicine 740–4, 740, 741 exitus in tabula (death on operating table) 747–8, 749, 748 incidence 736, 737 investigation 750 mechanical device‐related  751 medical disciplines involved  740, 741 medicolegal aspects  745 most frequent mistakes  740, 745 occupational group  742 outcome 745, 745 patient complaints  470 medical ethics  1594 medical malpractice  1589 medical treatment  1587–95 as bodily harm/assault  1588–9 confidentiality 1593–5 medical indication and standards  1589, 1589, 1589 persons involved  1587–8 self‐determination 1590–1 medically assisted suicide  1605–10 medicolegal autopsy  184, 185–92 diving accidents  586 diatom test  586 external examination  584–5 forensic imaging  587, 588 gas detection  584–5 histology 585 internal examination  588 molecular autopsy  586 toxicology 586 elder abuse  1012, 1013 external examination  185–8 clothing 185–6 conduct of  186–7 state of corpse  186 internal examination  188–92 evisceration 189–90, 189 mass disasters  291 special procedures  650–1 newborn children  193 trauma 192–3 see also specific body parts medicolegal research  21 Mees’ lines  85 mefloquine, and driving fitness  1433 Megaselia scalaris (scuttle fly)  126, 130 melatonin 1237 chemical structure  1245 melatonin receptor antagonists  1245

U

TO

R

U

SE

historical aspects  792–4 ritual homicides  796–7 role of women in Mafia system  794–5 sawn‐off shotguns  795, 795 strong acids  797 symbolism in  794 maggots, collection of  1123 magnetic resonance imaging (MRI)  213–14, 213 males see men malformations, and sudden infant death  902–3 malnutrition 670 elder abuse  1015 protein‐calorie 675 protein‐energy 675, 677 mandibular fractures  394 manganese, sample requirements  1115 manner of death  81–3, 82 manslaughter 315 marijuana see delta‐9‐tetrahydrocannabinol mass disasters classification 292–3, 292 closed 292 open 291–3 transport‐related 291–7 mass graves  28–30, 29–30 mass spectrometry  1132 maternal death in gestation  697 iatrogenic 703–4 mortality rate  697 pregnancy‐related death  698–702 pregnancy‐related risks  697–8, 698 MCP‐1 369 MDMA see amphetamines mechanical excitability of muscle  91–6, 95 mechanical trauma  376 Meckel’s diverticulum  189, 195 meconium 688 medazepam 1237 medical diagnosis, criminal relevance  1589 Medical Error Reporting System (MERS)  738, 740 medical errors  735–52 autopsy 751 blood transfusion errors  745–7, , 746 bodily damage caused  741 cause of accusation  741 causes of preliminary proceedings  742 classification 742 Clinical Liaison Service  752 definitions 735–6, 736 diagnosis of disease  740 drug‐associated deaths  748–50, 749, 749 duration and site of treatment  742

TR IB

and sudden infant death  888 and traffic accidents  1464–5 loprazolam, and driving fitness  1424, 1425 loratadine, and driving fitness  1428 lorazepam and driving fitness  1430, 1426 toxicological analysis  1417, 1419 lormetazepam, and driving fitness  1424, 1425 love bites  383, 385 Lovejoy’s method of age estimation  280–2, 180–2 lower limit of quantification (LLOQ)  51 lowest observed adverse effect level (LOAEL)  1258 Lubanga, Thomas  28 Lucilia caesar  131 Lucilia sericata (blow fly)  127–8, 127 Ludes, Bertrand  10 luminol test  272 lungs adult respiratory distress syndrome  336, 407 drowning victims  562, 563, 564, 564, 904 emphysema 604 floating test  352, 689 multiple organ failure  345–6, 345 neurogenic pulmonary oedema  339 pneumonia  77, 339 respiratory lung  339 sepsis 341, 342 specimen collection  1121 trauma 339 blunt force  401 penetrating injury  191 volume 578, 579 weight 564 see also pulmonary disease; respiratory tract lymphocytic myocarditis  813, 813, 814 lysergic acid diethylamide (LSD)  780, 789, 1126, 1133 stability 1141

1662

Index

FO

R

O

N

LY

motor vehicle accidents see traffic accidents mouth 187 dissection of  519, 519 obstruction of  543–5, 544 MRI see magnetic resonance imaging mucosa, drying of  103–5, 105, 134 multiorgan dysfunction  340 multiple corpses  86–7 multiple organ failure  345–6, 345, 346 mummification 109–10, 109, 110 Münchausen syndrome  1112 by proxy  970, 976–7 murder 315 muscle drowning victims  565 excitability electrical 92–4, 93, 94, 94 mechanical 91–6, 95 haemorrhages in hypothermia  632 specimen collection  1122 musculoskeletal system in diving accidents  585 mutilation mass disasters  291 postmortem 767–70, 768, 769 see also specific injuries muzzle imprint  484 muzzle modifications  490 Mycoplasma hominis  931 mydriasis 85 myelin, staining methods  418 myocardial infarction see ischaemic heart disease myocarditis  809, 812–15 classification 813 Dallas criteria  814 diagnosis 813, 814 eosinophilic 815 giant cell  815 granulomatous 815 infectious causes  812, 814 lymphocytic 813, 813, 814 polymorphonuclear 815 and sudden infant death  902 myocytolysis, coagulative  808 myofibroblasts, immunohistochemical markers 365 myopia  1345–7, 1345

TR IB

U

TO

R

U

SE

methohexital 1181, 1240 methotrexate poisoning  1111 methylecgonine, stability  1139 methylenedioxyamphetamines, metabolism 1164 methylphenidate (Ritalin), and driving fitness 1433 methyprylon 1243, 1243 pharmacokinetics 1237 Metoprolol CR/XL Randomised Intervention Trial in Congestive Heart Failure (MERIT‐HF) 1341 mianserine, and driving fitness  1092, 1427 microbiological autopsy  196 drowning victims  569 microglia 413 staining methods  418 microwave burns  643 midazolam chemical structure  1242 metabolism 1244 pharmacokinetics 1237 midbrain shearing  951 military service, faked disease/injuries  40, 733, 942 mines 494 minimal invasive autopsy (MIA)  836 Minnesota Protocol  29, 31, 646 minors, medical treatment of  1592 miosis 85 MIP‐1α 369 mirtazapine, and driving fitness  1425, 1427 mitochondrial DNA  1520 mizolastine, and driving fitness  1428, 1432 MK‐2866 1290 M’Naghten rule  1064, 1065 MNOP toxins  85 moclobemide, and driving fitness  1425, 1431 molecular autopsy  569 diving accidents  587 molecular biology disaster victims  293, 294 standards 294–5 wound age estimation  368–71 molecular genetics  48, 183, 1506 morphine 1200 absorption 1200 chemical structure  1254 distribution 1201 excretion 1201 metabolism 1201 pharmacokinetics 1255 plasma concentrations  1201 toxicological analysis  1418, 1419

N

C

O

men age estimation  278–9, 276 sex determination  273–4, 274–5 meningitis, and sudden infant death  901 mens rea (intent)  313–14 mental capacity  315 lack of  1591 mental competence  1059 consent to treatment  1061 meprobamate 1247 chemical structure  1246 pharmacokinetics 123 toxicological analysis  1417 mequitazine, and driving fitness  1428 mercury 1261–2 exposure 1261 poisoning 85, 1111, 1117 reference concentrations  1261 sample requirements  1115 toxicity 957 toxicokinetics 1261 mescaline  789, 790–1 mesopic vision  1347, 1352 Messerer’s fracture  1456, 1458 metabolic disorders screening for  961 and sudden infant death  902, 902 metabolism  1155, 1161–4 first pass  1177 genetic variation  1162 polymorphism 1162 methadone 1203–4 chemical structure  1254 metabolism 1162 pharmacokinetics 1255 toxicological analysis  1418, 1419 methaemoglobin 1110 methaemoglobinaemia 96, 792, 1267, methamphetamine see amphetamines methanol  1168, 1180–1 in alcoholic beverages  1181 analysis of  1171 concentration–time profile  1177 as marker of alcohol abuse  1442 metabolism 1176 physicochemical properties  1170 poisoning 1181 toxicity 1176 methanol poisoning  85, 1111 methaqualone 1245 chemical structure  1246 pharmacokinetics 1237 methionine 1111 Methodus testificandi  6, 8

nail marks  187 nails poisoning 85 specimen collection  1123 naphthol‐AS‐D‐chloracetate esterase stain 418

1663

Index

N

O

C

R

FO

O

N

LY

differential diagnosis  957–59, 957 evidence of malicious injury  948, 948 historical aspects  939–40 history at presentation  940 investigations 759–61 medical legalities  962 missed diagnosis  959 neuropathology 951–2 outcome 963, 963 parental and child risk factors  940–2, 941, 942 rebleeding 958 socioeconomic risk factors  940 non‐biological exhibits  1123 non‐discrimination 1613 non‐homicidal events  319 non‐lethal projectiles  462–3 non‐renal clearance  1157 non‐ST‐elevation MI (NSTEMI)  1331 nordiazepam 1238 toxicological analysis  1418, 1419 norfluoxetine 1147 normobaric/hyperbaric hypoxia  547–9, 548 Norway, statutory limits for alcohol levels  1394 nose 187 obstruction of  543–5, 544 novus actus intervenens  314 Nowak, Manfred  32 NSAIDs  789, 792 nuchal muscles, dissection of  519

U

TO

R

U

SE

neonaticide  683, 685–6, 686 incidence 684 see also infanticide neovascularisation 368 Nerium oleander  1276, 1277 Netherlands, statutory limits for alcohol levels 1394 neurogenic pulmonary oedema  339 neuroleptic malignant syndrome  791 neurological disease  816–28 intracerebral haemorrhage  430, 430, 472, 818–19, 818, 819 SUDEP 816–19 neurons 412 staining methods  418 neuropathic ulcers  1013 neuropathological autopsy  196, 197 neutrophilic myocarditis  815 New Zealand competence to stand trial  1062 insanity standards  1065 statutory limits for alcohol levels  1394 newborn children see neonates niaprazine 1246, 1246 nickel, sample requirements  1115 Nicrodes littoralis  126, 126 Nigeria insanity standards  1066 Nissl stain  418 nitrazepam 1238 and driving fitness  1425 nitric oxide  1267 respiratory toxicity and exposure threshold 1272 nitrobenzene 1272 respiratory toxicity and exposure threshold 1272 nitrogen dioxide  1267 respiratory toxicity and exposure threshold 1272 nitrogen narcosis  580 nitrogen phosphorous detector  1132 nitrous gases  616 nitrous oxide  1179, 1267 respiratory toxicity and exposure threshold 1272 no observed adverse effect level (NOAEL)  1258 non‐accidental head injury (NAHI)  939–63 biomechanics 952, 952–5 clinical diagnosis  952, 955–9 clinical examination and investigations 942–3, 942, 943 diagnosis 940–6

TR IB

narcotic (opioid) syndrome  1125 narcotic (sedative/hypnotic) syndrome  1124 narcotics see opioids Nasonia vitripennis  125, 125 nasopharynx, dissection of  519, 519 Natatolana woodjonesi (sea lice)  136, 137 National Aging Resource Center on Elder Abuse (NARCEA)  1007 National Association of Medical Examiners (NAME)  45–6, 724 natural causes of sudden death  774–8, 813 cardiac 803–16 non‐cardiac 616–28 see also specific causes natural poisons  1274–8, 1277 analytical methods  1277 see also poisoning; and specific poisons near‐drowning 62 torture 37–8 Near East  4–6 neck asphyxiation, dissection in  517–19, 518 external examination  187–8 internal examination  190–1 neck holds  533 accidental 535 carotid sleeper holds  566 choke holds  566 circumstances 534, 534 classification 534 definition 533 frequency/occurrence 534 pathomorphology 533–5 physical restraint  719–20, 720 neck injuries  192, 400 physical activity after  765 sharp force  454, 454 Necrobia rufipes  126, 126 Necrophorus vespilloides  126, 126 necrosis 415 necrotising fasciitis  340 needle biopsy  212 nefazodone, and driving fitness  1427, 1431 neglect  677–8, 983 children 678 older people  1009–10 self‐neglect 1010 see also starvation Neisseria gonorrhoeae  931 neonates forensic autopsy  193 normal genital variants  933 SIDS see sudden infant death syndrome see also children; infanticide; infants

occupational exposure  1272, 1272–3 occupational lung disease  196 octane 1273 oedema cerebral  202, 399, 702, 944, 949 genitalia 925 pulmonary 340, 897 oesophageal varices  547, 773, 781 ohms 640 Ohm’s Law  640 older people age estimation  1042 driving fitness  1385 vision problems  1351–2 violence against see elder abuse women sexually transmitted infections  1009 oligodendrocytes 413 staining methods  418 ophthalmology 961 opioids  77, 788, 788, 1200–4, 1253–4 chemical structures  1254

1664

Index

TR IB

U

TO

R

U

O

C

R

FO

LY

N

SE

P‐glycoprotein transporter  1159–60 paclitaxel 1277 paediatric neuropathology  433–9 ischaemia, hypoxia and asphyxiation  439–40, 440, 442 postnatal mechanical brain injury  433–4 painkillers  789, 791–2, 792 pale nerve cell injury  440, 443 ‘Palestinian hanging’ torture  38, 39 pallor in children  943 Paltauf bleeding  904 pancreas hypothermic changes  632, 632 weight loss in starvation  674 pancreatitis, alcohol‐induced  777 paracetamol  789, 791–2, 792 poisoning 1110, 1117 paraldehyde  1238, 1241 paraquat 1276 poisoning 1110, 1111 Paré, Ambroise  6, 7–8, 155 paroxetine, and driving fitness  1427, 1431 parrot perch torture  39 paternity testing  1525–30 autosomal markers  1526 biostatistics 1528–9 male and female lineage markers  1526– 8, 1526, 1528 quality assurance and ethical considerations  1529–30 pathological markers  283, 284 patient controlled analgesia (PCA)  750 PCR amplification see polymerase chain reaction amplification PEACE (Prevention of Events with Angiotensin Converting Enzyme Inhibition) trial  1333 pelvis fractures 404 sex determination  273–4, 277–8 pen guns  446 pentachlorophenol 283 pentobarbital 1238, 1240 percutaneous coronary interventions  1333 perianal bruising  927 perianal erythema  926, 923

pericardial fluid  872, 873 specimen collection  1122 periorbital haematoma  390, 392 peritoneal dialysis  1112 permafrost bodies  109, 111 personal injury assessment  1625–34 administrative law  1634 aim of  1626 civil law  1630–4 dependences 1633 partial temporary functional deficit  1631 permanent aesthetic damage  1632 permanent functional deficit  1632 permanent professional repercussion 1632 permanent repercussion on sexual activity 1632–3 permanent repercussions on sporting and leisure activities  1633 quantum doloris  1632 temporary partial professional repercussion 1631 temporary total professional repercussion 1631 total temporary functional deficit  1631 criminal law  1630 forensic medical reports  1630 general methods  1628–30 historical aspects  1625 labour law  1633–4 dependences 1633 permanent impairment  1633 temporary partial professional impairment 1633 temporary total professional impairment 1633 reparation 1630 role of forensic medicine expert  1630 personalised medicine, legal issues  1595–602, 1597 areas of application  1595–6 challenges 1596 confidentiality 1600 data protection  1600 definition 1595–6 freedom of therapy  1598 genetic testing  1598–9 health insurance law  1600–2 informed consent  1598 research 1602 personnel, qualifications forensic DNA laboratories  47–8 forensic toxicological laboratories  49, 50–51 Peru 34 pesticides 1278–4

O

oxygen deficiency  547–9, 618 and brain injury  444 hypobaric  512, 549 normobaric/hyperbaric 547–9, 548 oxygen toxicity  580, 581 ozone 1267 respiratory toxicity and exposure threshold 1272

N

opioids (continued) in doping  1286 and driving fitness  1381, 1413–14 narcotic (opioid) syndrome  1125 narcotic (sedative/hypnotic) syndrome  1124 pharmacokinetics 1255–6, 1255 pharmacology 1253 postmortem toxicology  1256 sources of  1253 tolerance and dependence  1254–5 toxicity 1256 use of  1253 see also specific drugs Optional Protocol under the Convention Against Torture (OPCAT)  36 oral cavity see mouth oral fluid  1144 organ donation  263 organic nitrites/nitrates (poppers)  789, 792 organised suicide  1609–10 Organization for the Prohibition of Chemical Weapons (OPWC)  1096 Organization for Security and Co‐operation in Europe (OSCE)  27 Organization of American States (OAS)  27 organochlorines 1279 organophosphates herbicides 1281–2 insecticides 1282 ORP‐150 369 orthograde suspension  540 orthopantomogram 1576 orthostatic hypotension  1339 osteology see anthropology/osteology osteons  273, 282 osteophytosis 285 overkill 453, 731, 732 overrunning railway accidents  1477 road traffic accidents  1459, 1460 overwhelming emotion  1067, 1069 oxazepam chemical structure  1242 and driving fitness  1426 fatal toxicity index  1090 metabolism 1242 pharmacokinetics 1237 toxicological analysis  1418, 1419 oxycodone chemical structure  1254 metabolism 1162 pharmacokinetics 1255 toxicological analysis  1418 oxydemetonmethyl poisoning  85

1665

Index

U

R

TO

U

TR IB

O

C

R

FO

O

N

LY

positional restraint see physical restraint postgraduate professional education  50–51 postmortem blood loss  333–4 postmortem changes  87, 91–133 algor mortis  99–103, 100–3, 104, 105 animal predation  109, 109 autolysis 105 bog and permafrost bodies  109, 111 decomposition 108, 108–9 drowning advanced 561 early 560, 560 drying of skin and mucosa  103–5, 105, 134 early 91 forensic entomology  123–32 injuries see postmortem injuries late (decomposition)  105 lividity 94–99, 95, 95, 96 stages of  91, 97 mummification 109–10, 109, 110 preservation processes  109 putrefaction see putrefaction rigor mortis  97–9, 97, 98, 98 saponification 110–11, 111 skeletonisation 108–9, 109 supravitality 91–6 time of death  111–25 post‐mortem computed tomography (PMCT) 836 postmortem injuries  133–42 animal predation  105, 105, 108–9, 109 bodies recovered from water  134, 135, 136, 137 drying of mucosal surfaces and eye  103–5, 105, 134, 135 embalming 141 fire victims  136–7, 138 handling, transportation and storage of body 128, 140 heat damage  136, 138, 611, 614, 612–13, 612, 616 incised wounds  377 maceration 134 pseudoligature marks  138–40, 140 putrefaction mimicking  134, 135 regurgitation of gastric juice  140, 141 resuscitation procedures  140–5, 141–2 urine leakage  141 vibices 141, 140 postmortem interval (PMI)  128–30 accumulated degree‐days/degree‐hours method  129, 271 determination of  271–2, 272 larval age  129–30 modelling of insect development  130

SE

piercings  187, 426 pinching 972 pink teeth  560 pistols 460, 461 pituitary gland, weight loss in starvation  674 placenta previa  684, 702 planimetric radiographic methods  1030 plaque jaune  398 plasma, specimen collection  1119 plasma elimination half‐life  1156, 1157, 1202 plasmapheresis 1112 plastic bag asphyxiation  547–8, 548 pleural exudate drowning victims  563 specimen collection  1122 PMI see postmortem interval pneumohaematothorax 1460 pneumonia  77, 820–3, 821 interstitial 821 lobar 777, 777 and sudden infant death  900 pneumothorax 536–7, 539, 539–41 poisoning 85, 85, 86, 376, 1107–26 child abuse  982, 983 clinical features  1108 clinical and laboratory liaison  1110–1112 diagnosis 1107–8 infants 906 interpretation of results  1115–18, 1117, 1117 investigations 1109, 1110 natural poisons  1274–8 samples and sampling  1112–15 general requirements  1112–3, 1113 postmortem collection  1114 trace elements  1114, 1115 treatment 1110, 1111, 1112 see also drug addiction; toxicology; and specific poisons Poland, statutory limits for alcohol levels  1394 police custody, deaths in see in‐custody deaths polymerase chain reaction (PCR) amplification  1503–5, 1505 polymorphisms in drug metabolism  1161 STR 1502–3, 1502 polymorphonuclear leukocytes ischaemic heart disease  807, 807 staining methods  418 polymorphonuclear myocarditis  814, 615 pontine myelinosis  775 population genetics  1510–12, 1510 pornography, juvenile  933 Portugal, statutory limits for alcohol levels 1394 positional asphyxia syndrome  538, 721

N

insecticides 1276–81 rodenticides 1283–4 petechial haemorrhage  187, 351, 383 asphyxiation 520–, 520 hanging  352, 524–6 ligature strangulation  528–9, 529 manual strangulation  532 SIDS 697, 895 see also contusions pH, and absorption  1159 phase I metabolic enzymes  1161 phase II metabolic enzymes  1162 phencyclidine  789, 790 toxicological analysis  1417 phenobarbital 1238, 1240 toxicological analysis  788 phenotype 1531 phenytoin poisoning 1111 toxicological analysis  1103 phosgene 1267 respiratory toxicity and exposure threshold 1272 phosphatidylethanol  1441, 1441, 1443 phosphine 1267 respiratory toxicity and exposure threshold 1272 photodiode array detector  1132 photogrammetry 378 photography 297 non‐accidental head injury  963 phthalimides 283 physical activity after injuries  765–7 abdominal injuries  766 chest injuries  765, 766 definition 765 head injuries  765 injuries to extremities  766 neck injuries  765 physical neglect see neglect physical restraint  717–26 auto‐erotic 710–11 elder abuse  1009 excited delirium  723–4 extrinsic factors  719–23 in‐custody deaths  717–19, 718 death certificate  725–6 investigation of  724 intrinsic factors  722–3 neck holds  719–20, 720 options for  719 restraint asphyxiation  721–2, 721 torture 38–9 physical violence  987 Physicians for Human Rights  27, 31, 32

1666

Index

U

R

TO

U

TR IB

O

C

R

FO

O

N

LY

railway accidents  1093, 1477 ramelteon  1238, 1245 and driving fitness  1425 rear‐end collisions  1463–4 rebreathing accidents  508, 548 recoil 466 recovery of remains  268–71, 269, 270 rectal temperature  113, 115death time interval 115 environmental temperature  113 measurement 113 re‐epithelialisation 368 Reference Dose (RfD)  1258 reflex anal dilatation  627 reflex syncope  1338–9 refractive errors and driving fitness  1345 rehabilitation of criminals  1069–70 religious objections to autopsy  296 renal clearance  1157 renal failure  77, 876–8, 879 research 1602 Convention on Human Rights and Biomedicine 1603 Convention for the Protection of Human Rights and Fundamental Freedoms  1603 on corpses  1610 data handling  1604 highly vulnerable people  1604 human rights  34–5 medicolegal 21 sample handling  1604 resistance 640, 640 respiration, vital reactions  352–5, 353, 354, 355 respiratory lung  339 respiratory orifices, obstruction of  543–5 larynx 544–5, 546 mouth and nose  543–4, 544 trachea and bronchi  545–6, 546 respiratory paralysis  335, 642 respiratory tract obstruction of  534–41 burking 537 head‐down position  539, 539 orthograde suspension  540 pneumothorax 536–7, 539, 539–41 positional asphyxia syndrome  538 respiratory paralysis  335, 642 thoracic compression  535–6, 535, 536 SIDS 899 thermal injuries  616 toxicity of gases and solvents  1272 see also specific parts

SE

pseudoligature marks  138–40, 140 psilocin/psilocybin  789, 490 psychiatric hospitals, deaths in  84–5 psychological torture  41 psychological/emotional abuse  970, 987, 989 psychosis 446, ptomaines  106, 769 pubic symphysis, in age estimation  276, 278 pugilistic position of burned bodies  614, 614 pulmonary barotrauma  578 pulmonary disease  818–24 asthma 781 pneumonia see pneumonia pulmonary embolism  78, 353, 406, 818, 821 and traffic accidents  1464–5 pulmonary emphysema  345 pulmonary oedema neurogenic 340 SIDS 897 pulseless electrical activity  376, 379 punch drunk syndrome  430 puncture marks  134, 135, 186 punishment for crime  316 pupil width  61, 88 poisoning 85 Puppe’s rule  324, 390, 422, 422, 484 Purdy, Debbie  1608 putrefaction  62, 87, 105, 106, 107, 109 blisters , 134, 135 chemistry of  107 infanticide 684 mimicking postmortem injuries  134, 135 morphological changes  97, 107, 107 progression of  107 and specimen collection  1123 and time of death  122, 123 transudates  106, 134, 134 and water temperature  591–2, 591 pyrethroid insecticides  1280, 1281 pyridine 1272 respiratory toxicity and exposure threshold  1273 pyrithyldion 1243, 1243

N

postmortem mutilation  767–70, 768, 769 classification 767 defensive 767, 769 definition 767 epidemiology 768 findings 769–70, 770 offensive 770 post‐traumatic stress disorder (PTSD)  40–1, 251–2, 997, 1068 potassium 880 in vitreous humour  118–20, 120 Praktisches Handbuch der Gerichtlichen Medizin 14 pregnancy Arias Stella phenomenon  698–700, 700 extra‐uterine 698–9, 699 gestational diabetes  1361 maternal death during  697 and partner abuse  1001 physiological risks  697–8, 698, 699 uterine fundus level  698 pregnancy‐related death  698–702 early pregnancy  698–702 abortion 699–700 extra‐uterine pregnancy  698–9, 699 preservation processes  109 pressure sores, drug addicts  789 pressure stasis  514 pressure ulcers  1031 Pretty, Diane  1583 prevalence 1375 preventive detention  1069 prison deaths in  84 self‐harm in  1023 prodrugs 1138, 1139 professional standards  1590 proficiency testing  47 prognathism 163, propane 1272 2‐propanol 1168 n‐propanol 1168 in alcoholic beverages  1181 physicochemical properties  1170 propellor injuries  136, 137, 377 propylene glycol  1271 prostheses 284 in victim identification  1563 protein binding  1160 protein‐calorie malnutrition  675 protein‐energy malnutrition  675 proteomics 882 protozoal myocarditis  814 Prussia 11–12, 12 Prussian blue  1117

Quaestiones medico‐legales  6, 9, 9 quality management disaster victim identification  297 forensic DNA laboratories  48 forensic toxicological laboratories  46 quantum doloris  1632 race see ethnicity radiation 376 exposure in X‐ray examinations  1038–9 radiology see forensic imaging

1667

Index

U

TO

R

U

N

O

C

R

FO

LY

N

SE

sabotage 1095–6 sacroiliac anchylosis  285 salicylate poisoning  1110 samples see specimen collection saponification 110–11, 111 Sarcophaga spp. 131 Savigny, Friedrich Karl von  62 sawn‐off shotguns  487 Mafia‐related deaths  795, 795 scalds  379, 621, 621, 621 child abuse  975, 976–9 differential diagnosis  980 time–surface temperature thresholds  609 scalp blunt force injuries  420–1, 420 haematoma 366 lacerations 389 scarification 783, 786 SCD‐HeFT (Sudden Cardiac Death in Heart Failure Trial) study  1337 schizophrenia  1021, 1066 school violence  995–7 motives and reasons  997 prevention and intervention  997 statistics 996 sciatic notch  274 scleral desiccation  134, 134 Scotland  21, 242, 260 Scottish Index of Multiple Deprivation (SIMD) 940, 941 scuttle fly (Megaselia scalaris)  126, 130 sea lice (Natatolana woodjonesi)  135, 136 seat belt injuries  384, 1461–2, 1463 sedatives and hypnotics  1236–48 aldehydes and derivatives  1240–1 barbiturates  788, 789, 1236, 1239–40 benzodiazepine‐related drugs  1241–3, 1242 benzodiazepines 773, 788, 1140, 1142, 1243–4

clinical and forensic aspects  1239 and driving fitness  1424, 1425 melatonin receptor antagonists  1245 narcotic (sedative/hypnotic) syndrome  1124 pharmacodynamics 1236 pharmacokinetics 933, 1237 piperidinedione derivatives  1243, 143 poisoning 933 tolerance, dependence and withdrawal 1239 see also specific drugs selectin, E‐ 365, 369 selectin, L‐ 365 selectin, P‐ 369 selenium, sample requirements  1115 self‐amputations 1023 self‐determination 1590–1 deficiency of will  1591–2 emergency situations  1592 information 1592–3 legal rules  1590 minors 1592 right not to know  1593 significance 1590–1, 1591 self‐harm 454, 455–6, 1021–5, 1022 body modification  303, 1021 classification 1021–3 definitions, phenomenology and causes  1021 diagnostic procedures  1024–5, 1025 epidemiology 1021 factitious disorders  877 faking of criminal offences  1021 genital self‐injuries  454, 1021 insurance fraud  1026 psychiatric diseases  1021 scars, piercings and cultural/religious self‐injuries  454, 1021 self‐burning 1023 suicidal acts  1021, 1021 see also suicide self‐made bombs  493 self‐neglect 1010 semi‐sharp force injuries  377 see also specific injuries sepsis 339–40 adrenal gland  344, 344 brain 343–4, 343 heart 343, 343 liver 342–3, 343 lung 341–2, 342 pathology 341 post‐abortion 699 severe 340

O

Rokitansky, Carl von  10, 12 Romano–Ward syndrome  903 rotational impact deceleration brain injury 951 rotational (inertial) brain injury  950, 950 midbrain shearing  951 vascular shearing  950 white matter shearing  951 rubbing alcohol see isopropanol rupatadine, and driving fitness  1428, 1432 Russian Federation diminished responsibility  1068 insanity standards  1066 Russian Penal Code  1066

TR IB

restraint see physical restraint restraint asphyxiation  719–20, 720 restrictive cardiomyopathy  810 resuscitation injuries  761 reticulin fibres, staining methods  418 retinal haemorrhage birth‐related 958 in children  946–9, 947 shaken baby syndrome  887, 904 retinitis pigmentosa  1349 reverse transcriptase polymerase chain reaction (RT‐PCR)  227, 230–2, 365 revolvers 460–1, 460 rib fractures  336, 366, 400, 401 child abuse  948, 948 rickets 285 ricochet 467 rifles 460, 461 right not to know  1593 right ventricular blood  872 rigor mortis  97–9, 97, 98, 98 causes 98 re‐establishment of  93 time course  98 vs. cold stiffening  98 RITA  3 (Randomised Intervention Trial of unstable Angina  3) study  1332, 1341 ritanserin, and driving fitness  1426, 1426 ritual homicides  796–7 RM 3/1 365 road traffic accidents  1455–60 abbreviated injury scale  1456 car to car collisions  1460–4, 1462 frequency 1462 front‐end collisions  1461–3, 1462 rear‐end collisions  1463–4 seat belt injuries  384, 1461–2, 1463 side collisions  1463 car to pedestrian collisions  1455–60, 1456–60 impact 1455, 1457 loading 1455, 1459, 1464, 1464 lower leg injuries  1456, 1458, 1477 overrunning 1460, 1462 shoe heel abrasions  1458 shuffling off  1455, 1457, 1458–9, 1458 trace patterns on car  1457 tyre tread marks  383, 1461 two‐wheeler vehicle to car collisions  1464–5, 1464 Roadside Testing Assessment (ROSITA) project 1410 rodenticides 1283–4 anticoagulants 1283–4 strychnine 1284 Rohypnol see flunitrazepam

1668

Index

U

R

TO

U

TR IB

O

C

R

FO

O

N

LY

fibres in wound  481 position of victim  483 CT imaging  214, 329 defence injuries  489 direction of gunshot  480–1 backspatter of skin  481 bone contact  481, 481 suicide vs. homicide  488 textile fibres  481 see also entrance wounds, below; exit wounds, below dynamic vs. static situations  484 entrance wounds  475–7, 476–7 atypical 480 morphology and range of fire  486, 486 prior exposure  489 sites of  487–8 epidemiology 490 exit wounds  480, 481 atypical 480 expert evidence  763–70 forensic reconstruction  474–7 methods 474–5 textiles and skin  475–7, 476 geometry of  488 head  329, 427 incapacitation 473 concussion and cerebral pressure  474 definition 473 delayed 474 head injuries  473–4 immediate 473 mechanisms 473 psychological factors  474 rapid 474 number of gunshots  487 propellant burns  378 range of fire  486, 486 contact wounds  378, 478, 478, 479 distant wounds  479 gunshot residues  477–8, 477–8 intermediate‐range wounds  348–9 near‐contact wounds  478 shot pattern  485 suicide vs. homicide  527–8 sequence of gunshots  484 shooter 482–3, 483 backspatter 481–3, 483 injuries and DNA transfer  483 position of  483–5 shooting hand  482, 483 steadying hand  482, 483 sooting  378, 479, 479 spine 432 subcutaneous gas expansion  478, 640

SE

subdural haemorrhage  438–9 shaken impact syndrome  949 shallow water blackout in divers  580, 581 shark bites  139 sharp force injuries  376, 377, 448–57 abdomen 456, 455–6 brain 951 cause of death  70–80, 71, 77, 314, 327–46, 328 chest 321, 455–6, 765, 766 crime scene  449–51, 450, 451 extremities 455–6, 455–6 head 454 homicidal 453 medicolegal findings  451–7 external investigation  451 individual stab wounds  451–2, 451, 452 number of lesions and patterns  453–4, 454 wound track  452–3 neck 454, 455 occurrence 448 prevention 457 self‐inflicted 454, 455–6 suicidal 453 weapon types  449, 449 see also specific injuries shear stress  325, 326 shearing/rotational brain injury  951 shell fractures  325 shock kidney  345, 353 shooting incidents  186, 192, 376, 378 accidental 489 discharge by mistake  489–90 gun cleaning accidents  490 hunting accidents  489 angles of gunshot  481–2, 482 backspatter shooter 482–3, 483 victim  482, 484 ballistics 459–95, 460 billiard ball effect  486, 486 bone contact  470–1 deformation and fragmentation  470, 470 deformed lead pellets  486–7 energy deposit and transfer  471 head injuries  471–2, 472 high velocity missiles  471, 472 mechanisms of injury  468–9, 468 missile–tissue interaction  469–70, 470 sawn‐off shotguns  487 shock wave  469 yawing  467, 469–470, 470 clothing and textiles entrance wounds  475–7, 476–7

N

sepsis (continued) SIRS 340–1, 341 spleen 343 septic shock  340, 341 septicaemia, and sudden infant death  190, 340, 959 septopyaemic abscess brain 343 heart 342 spleen 343 serial killers  731 serotonin syndrome  791–3, 1125–6 serum, specimen collection  1119–10 sex determination  273–4, 274–5 sexual abuse  919–36 checklist 935–6 conditions mistaken for  934 examination of victim  920–30 anogenital lesions  921–30, 922–9, 925–6 extragenital lesions  930 general procedures  920–1 findings caused by other conditions  935 incidence 939 interpretation of medical findings  933–3, 934 juvenile pornography  933 legal outcome  932–3 older people  1009 toxicological analysis  932 sexual homicide  729–44 crime assessment  541 crime scene evidence  730–1 definition 729–30 offender profiling  730–1 disorganised offender  730 organised offender  730 serial killers  731 sexual maturation  1042 sexual trauma  188, 193 sexual violence  987, 991 date rape  1094 drug‐facilitated 1094 evidence collection kit  1497 threats of  994 sexually transmitted infections  930–2 adults 932 children 931 older women  1009 shackling  39, 39 shaken baby syndrome  438, 904, 887, 949 cause of death  438 diagnosis 949 neurological classification of injuries  949 pathologies not causing  970 retinal bleeding  887, 904

1669

Index

N

O

C

R

FO

O

N

LY

spine examination  192, 417–18 forensic neuropathology  418–39 spleen blunt force injuries  404, 405 drowning victims  564 sepsis 343 specimen collection  1122 weight loss in starvation  674 ST‐elevation acute coronary syndrome (STE‐ACS) 1332 ST‐elevation MI (STEMI)  1332 stab wounds see sharp force injuries stain analysis  1497–43 body fluid identification  1499–502 DNA extraction and quantitation  1500 DNA typing  1502–9 evidence collection  1497–8 forensic DNA databases  1521–2, 1522 haplotypical markers  1517–21, 1519, 1520 novel techniques  1513–15 population genetics  1510–12, 1510 SNPs 1513, 1528 staining methods  418 standards autopsies 43–5 laboratory 46–7 standing trial, competence to  1061–3 starfish‐induced injuries  136 starvation  376, 669–79 body weight loss  691, 674 cachexia 669, 670 classification 674–6, 675 clinical and autopsy findings  623 death from  679–7 immediate cause of death  676–7, 677 physical condition prior to death  676, 676, 677 duration of  676 elder abuse  1015 energy requirements  670 malnutrition 670, 675 organ weight loss  670 status thymicolymphaticus  889 stereo vision  1350 sternum, blunt trauma  332 steroid abuse  789, 792 stillbirth 683–4 epidemiology 684 incidence 684 pathology 684–5 risk factors  685 stimulants  788, 790 in doping  1286

U

TO

R

U

SE

Smith, Sydney  3–4, 6 smoke inhalation  614 smoking in bed  779–80, 779 smooth muscle cell actin  365 smothering 508 children 443 see also asphyxiation; sudden infant death syndrome SNPs 1513, 1528 ancestry informative  1541 characteristics and current status  1528 identity 1513 lineage 1513 phenotype predictions  1513 soapy abortion  699–701 sodium 880–1 sodium chloride/nitrite/nitrate poisoning  96 sodium fluoride  1143 soft coverings, asphyxiation by  542, 544 solitary confinement  41 solvents 1269–2 occupational exposure  1272, 1272–3 respiratory toxicity  1273 see also specific solvents soot inhalation  355, 355, 355, 614, 615 sooting of gunshot wounds  378, 479 South Africa competence to stand trial  1062 diminished responsibility  1068 insanity standards  1063, 1066 Spain, statutory limits for alcohol levels  1394 specialists in forensic medicine  10 species, diagnosis of  272–3, 273 specimen collection  46 chain of custody  1124 DNA analysis  37, 1497–543 forensic toxicological laboratories  49–1, 51, 1118–24 handling  48, 51–2 labelling  46, 1124 poisoning cases  1110–12 postmortem 1141–3, 1142 storage  862, 1145–6, 1146 see also specific types spectrophotometry 961, 1132 spider’s web skull fracture  387, 422 spinal cord autopsy 197, 197 examination 417 spinal injuries  400 cervical spine  417, 432–3 closed 432 mechanical 432–3, 432 open 432–3 vertebral fractures/dislocations  402–3, 402

TR IB

suicide or homicide  164, 489, 1257 victim gun inside hand  489 position of  483–4 weapon see firearms wound characteristics  485–7 wound morphology  479, 482, 481 see also firearms short tandem repeats see STRs shotguns 461, 461 ammunition 465–6, 465 shoulder pin restraint  720 side collisions  1463 side effects see adverse drug reactions SIDS see sudden infant death syndrome signal guns  446 silencers 490 silver, sample requirements  1115 silver stains  418 Simon’s haemorrhage  506 single nucleotide polymorphisms see SNPs sinuses, drowning victims  564 SIRS see systemic inflammatory response syndrome sixpenny bruises  384–5, 384 skeletal maturation  1040–1 skeletal muscle see muscle skeletal survey in children  960 skeleton, examination of  192 skeletonisation 108–9, 108–9, 269 dispersal of bone segments  269 skin bleeding 85 blunt force injuries  380–400 abrasions  376, 380–2, 381, 382 contusions  376, 382–8, 383–7 lacerations  376, 388–90, 389 chromatic discoloration  560 drying 103–5, 105, 134 maceration see maceration multiple organ failure 345–6, 345 postmortem shrinking  379 putrefactive blisters  106, 134, 135 splitting in burned bodies  136, 137, 615 washer‐woman’s  15, 136–8, 137 wound healing  368, 1008 skull, fire effects  618 skull fractures  190, 390–3, 390–3, 421–2, 421, 422 children  433–4, 948, 948, 958 forensic neuropathology  421–2, 421, 422 Puppe’s rule  324, 390, 422, 422, 484 spider’s web pattern  387, 422 sleep disorders, and driving fitness  1383–4 small intestine, floating test  352

1670

Index

U

R

TO

U

TR IB

O

C

R

FO

O

N

LY

natural causes  803–28, 813 sudden unexpected death in infants (SUDI)  900–3 accidental and non‐accidental causes  904, 904 infection 901 malformations 902–3 metabolic disorders  901 practical investigation  905–8 bacteriology and virology  905–6 histology 905 metabolic and genetic tests  905 radiology 906 toxicology 906 sudden cardiac death  903–4 see also sudden infant death syndrome SUDEP see sudden unexpected death in epilepsy SUDI see sudden unexpected death in infants suffocation  376, 508, 509 children 441, 442 infants 895 see also asphyxiation suicide  319, 320 attempted  1022, 1023 drowning 570–1, 572 hanging  525, 528 incised wounds  377 ligature strangulation  531 maternal 703 medically assisted  1605–10 organised 1609–10 rebreathing 508, 548 shooting incidents  487 thoracic compression  535–6 see also self‐harm sulphaemoglobin 106 sulphur dioxide  616, 1267 respiratory toxicity and exposure threshold  1272 sunstroke  622, 623 supraventricular tachycardia  1334 supravitality  60, 91–9 duration of  92 iris, pharmacological excitability  94, 94 muscle electrical excitability  92–4, 93, 94, 94 mechanical excitability  91–6, 95 postmortem lividity  94–5, 94, 95, 96 stages of  97, 97 surface scanning  216–18, 217 surfactant, drowning victims  565 surgery, complications of  78 suriclone, and driving fitness  1430, 1426

SE

shaken baby syndrome  887, 904 subdural hygroma  424, 434–5 subendocardial haemorrhage  702 submarino torture  37 subpubic concavity  274 Suchey–Brooks method of age estimation  276–8, 278 sudden cardiac death  803–16 arrhythmias 815–16, 816, 1333–8 arrhythmogenic right ventricular ­dysplasia  75, 226 dilated cardiomyopathy  810 hypertrophic cardiomyopathy  226 in infants  903–4 ischaemic heart disease  73, 388 myocarditis 812–13 restrictive cardiomyopathy  810 stress cardiomyopathy  816 sudden death cardiac causes see sudden cardiac death neurological disease  816–29 pulmonary disease  819–20 sudden in‐custody restraint deaths (SIRD)  646, 717, 718 sudden infant death syndrome (SIDS)  225, 232, 241, 572, 887–99 autopsy findings  905, 895, 897 case definition  892–4, 893 circumstances of death  895–6, 894 clinical history  887, 888 epidemiology 894–5, 894, 895 co‐sleeping 895 sleeping position  892–3, 895 incidence 888 pathophysiology and aetiology  890–3, 891 possible causes  888–90 practical investigation  905–6 bacteriology 905 histology 905 metabolic and genetic tests  905 radiology 906 toxicology 906 risk factors  894 immunisation 959 sudden natural death while driving  1465–7 cause of death  1466–7 circumstances 1466 demographics 1466 epilepsy  1466, 1467 sleep‐related road traffic collisions  1467 sudden unexpected death adolescents 192 children  193, 260, 438–9 infants see sudden unexpected death in infants

N

storage of bodies  141, 140 of insects  128 of specimens  1124, 1145–6, 1146 strabismus 1349 strangulation 522–6 auto‐erotic 710–11 children 441–2, 442 hanging  187–8, 522 children 442 observations and time relationships  516, 516 petechial haemorrhage  352 infants 904 ligature 525, 525 observations and time relationships  516, 516 manual  380, 531–2 circumstances 529 definition 531 frequency/occurrence 531 pathomorphology 532–3, 532 marks 380 neck holds  533 see also asphyxiation streak‐like haematoma  384–5 stress cardiomyopathy  811–12, 816 stress proteins  369, 369 stretch marks  326 strontium, sample requirements  1115 STRs 1505 data analysis  1505–8, 1506–7 new technologies for testing  1513–15 polymorphisms 1502–3, 1502 typing kits  1504 strychnine 1284 poisoning 85 stud/nail guns  462 stun guns  645 subarachnoid haemorrhage  333, 396 children 434 Subcommittee on Prevention of Torture (SPT) 36 subdural haemorrhage  333, 391, 395, 396, 396, 397, 423, 423 ageing of  945–6 alcohol‐related 779 biomechanics 424 children  435, 944, 945 diagnosis 424 and hypoxic ischaemia  959 neonatal 958 pathologies not causing  970 post‐traumatic survival time  424

1671

Index

FO

R

C

O

N

O

N

LY

time of death  111–25, 314, 329 acute (minutes to hours)  334–7 compound method  115–18, 116, 114–18 determination of  112, 114 gastric contents  120–2, 121 immediate (minutes)  330–4 instantaneous 329–30, 329, 330 late (days to weeks/months)  337–40 objectives of estimation  113 potassium in vitreous humour  118–20, 120 putrefaction 122, 123 rectal temperature  113, 115 time of flight  1132 tissue embolism  351, 353 tissue remodelling  368, 1008 tissue retrieval  263 TNF‐α 365 Togo 32, 33 Tokyo Declaration  36 Tokyo War Crimes Tribunal  28 toluene 1208 respiratory toxicity and exposure threshold 1273 toluidine blue  921 tongue 187 haemorrhages 526, 527 torture  32, 35–42, 319–20 blunt force injuries  37–8, 37, 38 burning 38 definition 35 electrical 37 falanga  37, 40, 320 fingers 38 near‐drowning 37–8 positional 39 psychological 41 sequelae 39–40, 40 suspension 38 ‘telephono’ 38 victims examination 41 treatment 41–2 total clearance  1157 toxic elements  1257–9, 1258 human biomonitoring values  1258 see also specific elements toxicity 1100 evidence base for  1100–1102 toxicokinetics/toxicogenetics 1155–65 absorption  1155, 1158–9 bioavailability 1158 clearance 1156–7 distribution  1156, 1160–1 elimination rate constant  1157–8

SE

U

R

TO

U

tache noir  104, 105, 134 Tanner stages  933 Tardieu, Ambroise Auguste  11, 506, 939 Tardieu’s spots  521 Tasers 645–6 tattoos  163, 386, 1021 Taxus baccata (yew)  1277, 1277 Taylor, Alfred Swaine  3 teaching  15, 21 tear gas cartridges  462, 465, 466 teeth see forensic odontology ‘telephono’ torture  38 temazepam and driving fitness  1425 fatal toxicity index  1090 metabolism 1242 pharmacokinetics 1237 toxicological analysis  1418 temperature regulation  627, 628 temperature–time of death nomogram  99–101, 100, 103 temporal bone, haemorrhage  564 temporal muscle, haematoma  367 tenascin 365

tensile stress  321–6, 322–6 terfenadine, and driving fitness  1428, 1433 terrorism 1095–6 test results quality assurance  44, 52 reporting  47, 49 testamentary capacity  1060–1 tetrachloromethane 1273 textiles see clothing and textiles TGF‐α 365, 368 TGF‐β1 365 Thailand 32, 33 thallium 1263 exposure 1263 poisoning 85, 1117 reference concentrations  1263 sample requirements  1115 toxicity 1263 toxicokinetics 1263 thanatochemistry see biochemistry, postmortem thebaine 1254 theophylline poisoning  1110 therapeutic privilege  1593, 1597 thermal injuries burns see burns child abuse  975, 976–9, 977 respiratory tract  616 scalds see scalds Thiemann–Nitz radiographic atlas  1030 thin layer chromatography  1132 thin‐skull rule  316 thiopental 1238, 1240 thoracic compression  535–6 accidental 537 circumstances 536 definition 535 frequency/occurrence 535–6, 536 pathomorphology 536–7 suicide 537 thorax diving accidents  586 road traffic accidents  1459 unstable 541 in victim identification  1562–7, 1563 see also chest thromboembolism 338–9, 352, 353 post‐abortion 702 pulmonary 78, 354, 407, 819, 820, 821, 822 thrombosis 78 post‐abortion 703 thymus, petechial haemorrhage  897 thyroxine overdose  1111 tibial fractures  1458

TR IB

survival time  765–7 cerebral contusions  429–30, 430 see also time of death suspension torture  38–9 swallowing 356 Sweden no insanity defence  1067 presentation of psychiatric evidence  1062 rehabilitation and security  1070 statutory limits for alcohol levels  1394 swimming pools, electrocution in  651 Switzerland human research  1595–6 insanity standards  1066–7 syllogomania 1010 symbolism, Mafia‐related deaths  794 synapses, staining methods  418 syncope 1338–9 cardiac 1339 driving and  1339 implantable cardioverter defibrillators  1334–8 orthostatic hypotension  1339 prognosis 1339 reflex 1338–9 syphilis 931 and stillbirth  685 systemic inflammatory response syndrome (SIRS) 339–1, 341

1672

Index

U

R

TO

U

TR IB

O

C

R

FO

O

N

LY

car to pedestrian collisions  1455–60, 1456–60 two‐wheeler vehicle to car collisions  1464–5, 1464 sudden natural death while driving  1465–7 cause of death  1466–7 circumstances 1466 demographics 1466 epilepsy  1466, 1467 sleep‐related road traffic collisions  1467 traffic medicine definitions 1315 in forensic medicine  1315 operationalisation 1317 relevance of  1315 see also driving fitness training forensic psychiatry  1072 human rights  34–5 toxicology 1080 trajectory 466 tram‐like haematoma  384, 384 tramadol 1204 metabolism 1162–3 toxicological analysis  1418 transferrin, carbohydrate‐deficient  1442–3 transfusion‐associated circulatory overload (TACO) 746 transfusion‐related acute lung injury (TRALI)  746 transplantation 257–62 diagnosis of brain death see brain death in Germany  257–9 in Japan  261–2 organ donation and tissue retrieval  263 regulations and procedures, in Germany  257–9 in Switzerland  259–60 in UK  260–1 trauma see injuries traumatic asphyxia  725, 951 traumatology and criminality  317–20 definition 313 legal issues  313–16 trazodone 1090 treatment refusal  1063 Treponema pallidum  931 triazolam chemical structure  1242 metabolism 1242 pharmacokinetics 1237 trichloroethylene 1273 Trichomonas vaginalis  931

SE

non‐biological exhibits  1123 skeletal muscle  1121 special considerations  1123 urine 1120 vitreous humour  1120 stability and artefacts  1138–9 blood from living individuals  1139–40 definitions and causes  1138–9, 1138, 1139 evaluation of  1148–50 postmortem specimens  1141–3, 1142 processing and analysis  1146–8, 1147 sampling artefacts and drug stability  1143–5, 1144 storage 1145–2, 1146 urine from living individuals  1140–1 toxic elements  1257–9 see also specific substances toxidromes 1124–6, 1126 adrenergic syndrome  1125 anticholinergic syndrome  1125, 1126 cholinergic syndrome  1125 extrapyramidal syndrome  1126 narcotic (opioid) syndrome  1125 narcotic (sedative/hypnotic) syndrome  1125 serotonin syndrome  1125–6 Toxoplasma gondii  687, 1469 trace elements, sampling for  1114, 1115 trace evidence at crime scenes  337 trachea dissection of  518 obstruction of  545–9, 546 soot inhalation  615 tracheo‐oesophageal fistula  195 traffic accidents  191, 389, 1455–39 aircraft accidents  769 alcohol‐related  772–83, 1188–90, 1188, 1381 car to pedestrian collisions  1455–60, 1456–60 contributing factors  1388 expert evidence  763–70 mass disasters  291 cause and time of death  412 forensic medical examination  1627 pregnant women  704 railway accidents  375, 767 cause of death  1476, 1477, 1478 circumstances 1475–7 demographics 1474 train speed and injury pattern  1478 road traffic accidents  1455–60 abbreviated injury scale  1456 car to car collisions  1460–4, 1462

N

toxicokinetics/toxicogenetics (continued) excretion  1155, 1164 first‐order elimination  1157–8 LADME principle  1155, 1156 metabolism  1155, 1159, 1161 plasma elimination half‐life  1157 release of active compound  1155, 1158 volume of distribution  1156 toxicology 1167 adverse drug reactions  748–50, 749, 749, 1099–100 aircraft accidents  769 alcohols 1167–9 associations and organisations  1081 clinical forensic  1091–3 child welfare  1094 driving under the influence  1091–3 drug‐facilitated sexual assault  1094 workplace drug testing  1093 diving accidents  587 doping  1095, 1134, 1284 drug screening  1129–3 analytical methods  1133–4 confirmation tests  1130 instability of drugs during  1146–8, 1147 instrumentation 1131–2, 1132, 1132 isolation step 1130–1 method performance  1134–5, 1135 target substances  1130 education and research  1080 epidemiology 1100 fatal toxicity index  1089, 1090 illegal drugs  1081, 1196–209 intoxication see toxidromes laboratory standards  49–53, 1081 legal issues  1597 narcotics 1252–7 natural poisons  1274–8 pesticides and insecticides  1278–4 poisoning see poisoning postmortem 1071 sabotage, terrorism and chemical warfare  1094 sedatives and hypnotics  1236–48 sexual abuse  932 SIDS and SUDI  887 solvents and gaseous poisons  1266–9 specimen collection  1118–24 bile 1120 blood, serum and plasma  1119 brain 1121 kidney 1121 liver 1120 lungs 1121

1673

Index

FO

R

C

U

R

TO

U

TR IB

O

N

vaginal discharge in prepubertal girls  926 vaginal foreign bodies  926 VALIANT (Valsartan in Acute Myocardial Infarction Trial) study  1333 validation

N

LY

older motorists  1351 position and motility of eyes  1349–50 post‐eye surgery  1351 reduced visual acuity  1345–6, 1352 retinitis pigmentosa  1349 stereo vision  1350, 1352 vita minima  59, 61 vita reducta  61 vital processes  349, 350 vital reactions  349–62 circulation 350–2 embolism see embolism haemorrhage see haemorrhage digestion 356 respiration 352–5, 353, 354, 355 time course  349 vital signs  349, 350 vitamin deficiency, alcohol‐induced  774, 775 vitreous humour  872, 872 dehydration pattern  677, 677 glucose  874, 875 lactic acid  875, 876 potassium in  118–20, 120 specimen collection  1120 volatile solvents  1111 voltage 640, 640 volume of distribution  1156 vomiting chyme aspiration  546–7, 546 in SIDS  895 von Willebrand’s disease  980–1

O

ubiquitin 369 ultrasound, cranial  960 ultraviolet detector  1132 umbilical cord demarcation of  688 irregular edge  686 underfunding 21 underwater welding  651–2 United Kingdom  14–15 brain death diagnosis  263 forensic society training  1071 Human Tissue Act (2004) 260 retrospective civil assessments  1060–1 statutory limits for alcohol levels  1394 testamentary capacity  1060–1 transplantation procedures  260–1 United Nations Committee Against Torture  35 Declaration of Human Rights  27, 35–6 model autopsy protocol  44 United States competence to stand trial  1060 consent to treatment  1061 diminished responsibility  1068–9 forensic society training  1071 insanity standards  1067 legal competency  1060 rehabilitation and security  1070 retrospective civil assessments  1060–1 statutory limits for alcohol levels  1394 testamentary capacity  1060–1 urea 879 Ureaplasma urealyticum  931 urinary tract infection  77 urine diabetic coma  876 leakage postmortem  141 specimen collection  1120, 1144

forensic DNA laboratories  48 forensic toxicological laboratories  49 Valsalva manoeuvre  514, 920 vasovagal inhibition  330 vasovagal syncope  1338–9 VCAM‐1 365, 369 VEGF 368–70 venlafaxine 789 and driving fitness  1430–1, 1426 venous gas embolism  579 venous marbling  106, 107, 134, 135 venous ulcers  1031 ventilation 335 ventral arc  274, 274 ventricular tachycardia  1334 catecholaminergic polymorphic  903 idiopathic right ventricular outflow tract 904 vernix caseosa  193, 688, 702 vertebral disc haemorrhages  352, 527 vertebral fractures/dislocations  400–3, 403 vibices 141, 140 victim identification  30, 86–7, 186, 167–8 drowning victims  588–9 mass disasters  291–7 see also anthropology/osteology video surveillance systems, identification from 1567–8, 1568 Vieira, D.N. 19 violence against elderly  1005–12 against homosexuals  1001–3 domestic  318, 991–2 school 995–7 violent death classification of  375–80, 376–80, 376 see also specific injuries expert evidence  763–70 legal issues  313–16 older people  1015 viral myocarditis  814, 816, Virchow, Rudolph  44, 194 visual field defects  1350–1, 1031 assessment 1351 bitemporal hemianopsia  1351 classification 1350 examination method  1351 homonymous hemianopsia  1351 relevance for road users  1350 visual problems  1345 cataracts 1346–7 colour vision disorders  1348, 1350–2 diabetic retinopathy  1347–8, 1363 glaucoma 1348 macular degeneration  1349

SE

triclofos 1247 chemical structure  1246 pharmacokinetics 1273 triprolidine, and driving fitness  1432 tubal pregnancy  698–9, 699, 704 tuberculosis, alcohol‐associated  778 two‐wheeler vehicle to car collisions  1464–5, 1464 tympanic membrane rupture  647, 655 tyre tread marks  383, 1461

‘walk and die’ 334 Wallerian degeneration  416 warfarin 749 washer‐woman’s skin  15, 136–8, 137 water‐line lividity  652 water‐related deaths diving accidents  576–8 drowning see drowning electrocution 651–2 bath 652–3 electric eels  651 swimming pools  651 underwater welding  651–2 immersion time  123, 589–60, 560 postmortem injuries  136–40, 137, 140 waterboarding 38 Waterhouse–Friedrichsen syndrome  344, 701, 901 Welby, Piergiorgio  1608 Wernicke–Korsakoff syndrome (WFS)  775 whiplash injury  433, 939, 951 whipping 37, 38 sequelae 39–40

1674

Index

N

LY

Zacchia, Paolus  6, 9–10, 9 zaleplon 1238 and driving fitness  1381, 1425 zinc, sample requirements  1115 zip guns  462 zolpidem 1244, 1244 and driving fitness  1381, 1425, 1425 pharmacokinetics 1237 toxicological analysis  1417 zopiclone 1244, 1244 and driving fitness  1381, 1425 fatal toxicity index  1090 pharmacokinetics 1237 stability 1140 Zsako’s muscle phenomenon  91

FO

R

C

O

N

TR IB

U

TO

R

U

SE

X‐chromosomal markers  1527 X chromosome typing  1528 X‐rays age estimation in adolescents  1029–30, 1029 atlas methods  1030 bone‐by‐bone methods  1030 planimetric methods  1030 radiation exposure  1038–9 xenotransplants 259 xylene 1208 respiratory toxicity and exposure threshold 1273

Y chromosome‐specific STR loci  1517–19, 1519 young drivers  1384–5

O

skin wound healing  368 results and interpretation  365, 365–6, 366 wound ballistics  467–72 wound healing  368, 1008 wound tracks  452–3 Wydler’s sign  356

white matter shearing  951 whole bowel irrigation  1112 Widmark equation for alcohol consumed  1367–8 Wischnewsky’s spots  631, 632, 636 Wolff–Parkinson–White syndrome  1334 women age estimation  278–9, 276 genital mutilation  320 role in Mafia system  794–5 sex determination  273–4, 274–5 workplace drug testing  1093 World Anti‐Doping Agency (WADA)  1082, 1134, 1284 World Medical Association  36, 259, 1602, 1615 wound age estimation  364–7 markers 368–71, 369, 370 methods 364–5, 365 molecular biology  368–71