Asian Honey Bees: Biology, Conservation, and Human Interactions 9780674041622

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 Asian Honey Bees

 ASIAN HONEY BEES Biology, Conservation, and Human Interactions

B EN J AMI N P. O LD ROY D S I R I WAT W O N G S I R I

HARVARD UNIVERS I T Y P RESS Cambridge, Massachusetts, and London, England

2006

Copyright © 2006 by the President and Fellows of Harvard College All rights reserved Printed in the United States of America Illustrations by Jenny H. Ekman Design by Gwen Nefsky Frankfeldt Library of Congress Cataloging-in-Publication Data Oldroyd, Benjamin P. Asian honey bees : biology, conservation, and human interactions / Benjamin P. Oldroyd, Siriwat Wongsiri. p. cm. Includes bibliographical references and index. ISBN 0-674-02194-0 (alk. paper) 1. Honeybee—Asia 2. Honeybee—Behavior—Asia. 3. Honeybee—Conservation—Asia. I. Wongsiri, Siriwat. II. Title. QL568.A6O53 2006 595.79⬘9—dc22 2005055007

To Her Majesty the Queen of Thailand for her efforts to help save the wild bees of Asia

 Contents

Foreword by Thomas D. Seeley ix Preface

xiii

1

To Be a Honey Bee

2

Introduction to the Species 13

3

Evolution

4

Speciation and Biogeography

5

Dance Communication and Foraging 65

6

Reproduction, Swarming, and Migration 90

7

Worker Sterility, Kin Selection, and Polyandry 118

8

Nesting Biology and Nest Defense

9

Parasites, Pathogens, Predators, and a Plant 180

1

36

10

Human Interactions

11

Conservation

12

Concluding Remarks

52

148

209

231 249

Appendix A. A Simple Key to the Workers of the Genus Apis 259 Appendix B. A Simple Key to the Parasitic Mesostigmatan Mites of Asian Honey Bees

260

Appendix C. The Names of the Honey Bee Species in Asian Languages Glossary

265

References Index

329

273

262

 Foreword by Thomas D. Seeley

One species of honey bee, Apis mellifera, has been the subject of scientific observations since antiquity. William Morton Wheeler, a world authority on ants, once explained the ancient fascination with the honey bee as follows: “Its sustained flight, its powerful sting, its intimacy with flowers and avoidance of all unwholesome things, the attachment of the workers to the queen—regarded throughout antiquity as a king—its singular swarming habits and its astonishing industry in collecting and storing honey and skill in making wax, two unique substances of great value to man, but of mysterious origin, made it a divine being, a prime favorite of the gods, that had somehow survived the golden age or had voluntarily escaped from the garden of Eden with poor fallen man for the purpose of sweetening his bitter lot.” Over the past 150 years, the age of rigorous biology and modern beekeeping, a wealth of books and tens of thousands of technical articles have been published on the biology of honey bees and the craft of beekeeping. Today, Apis mellifera is by far the most intensively studied of all the social insects, a distinction that applies with special strength to the subjects of sensory physiology, learning and memory, communication, caste determination, nest micrometeorology, circadian rhythms, and the organization of work. The honey bee’s lofty status within biology was recently catapulted still higher when it joined the select group of organisms whose entire genomes have been sequenced. This species of bee is now a “model system” for studying the influence of an animal’s genetic constitution on its social behavior. Apis mellifera is not, however, the only species of honey bee found on

x

Foreword

planet Earth. The authors of Asian Honey Bees explain that the genus Apis is native to the Old World, and that while A. mellifera is found in most of Europe, the Middle East, and Africa, if we swing our attention eastward to tropical Asia and eastern Asia, we will find at least eight more species of honey bee, each a distinctive product of evolution by natural selection. These include the world’s largest honey bee, Apis laboriosa, fearsome denizen of the Himalayan mountains, and the world’s smallest honey bee, Apis florea, gentle dweller of the lowlands across most of tropical Asia. Each species, like A. mellifera, ranks among the highest eusocial insects (the higher polistine and vespine wasps, most ants, the stingless bees, and the higher termites), for in each species we find thousands of workers per colony, strong queen-worker dimorphism, extensive altruistic behavior, complex communication systems, precise nest thermoregulation, and other traits indicative of high social complexity. Each species also shares with A. mellifera the striking derived characters that are unique in the whole order Hymenoptera and that define the genus Apis: vertical combs of hexagonal cells built of nearly pure beeswax, waggle dances in which a bee performs a ritualized reenactment of an outward flight to a food source to indicate its location, and strangely modified male genitalia. And yet each species differs conspicuously from A. mellifera in its morphology and behavior, the product of an adaptive radiation of the genus Apis that occurred within its 35-million-year evolutionary history. This combination of similarities and differences relative to Apis mellifera makes the study of the Asian honey bees important. We know that A. mellifera is ultimately tropical in origin and that it succeeded in spreading into the north temperate zone of the Old World by virtue of certain preadaptations for living in a seasonally cold climate. These include the ability to make honey, which gives a colony a food store safe from spoilage through dearth periods; the ability to form a thermoregulated cluster, which enables a colony to fight deadly cold; and the ability to swarm, which eliminates the need for a hibernating solitary phase in the colony cycle. To identify the adaptive origins of such major features of honey bee social life as honey making, thermoregulating, and swarming, we need to know how each one serves colonies living in the tropics. The honey bees of the Asian tropics offer the richest opportunities for solving these puzzles. The honey bees of Asia also offer opportunities for remarkable discoveries. A glance through the pages of Asian Honey Bees reveals that their seri-

Foreword

ous scientific study is still nascent, having gained real strength only in the last 15 years. To cite just a few examples of the fresh findings, there is the lethal balling and 45°C baking of invading hornets by Apis cerana, the reporting of lurking predators by homecoming foragers in Apis florea, the returning to a particular nest site after completing a more than 100-km migratory journey in Apis dorsata, and the hibernating of whole colonies as exposed, combless, chilled clusters in Apis laboriosa. The fact that we have just begun to study these lovely and mysterious creatures should serve as an attractive point on the scientific horizon, one that will draw explorers to the midst of this still unfamiliar terrain. Because the Asian honey bees have ecological importance as pollinators for countless species, economic importance as the makers of honey, and spiritual importance for Hinduism and other religions in Asia, they deserve our attention for reasons of sheer utility as well as scientific curiosity. Indeed, their survival probably depends on our gaining a deeper ecological knowledge of these magnificent social bees. There is little doubt that some of the Asian honey bee species are threatened by forest clearing and excessive honey harvesting. Further harm may arise from destruction of nesting sites and introduction of exotic diseases. We humans now recognize that our own species is exquisitely adapted to the razor-thin biosphere covering the planet; hence our own survival depends on understanding and protecting the rest of life. This book helps us understand the Asian honey bees, a part of the natural world that we must protect as a matter of practicality and pleasure.

xi

 Preface

Most Western people who read this book will be familiar with the Western honey bee, Apis mellifera. This species has occupied a central place in the agricultural and social systems of Europe (and in the countries that have descended from there in America and Australasia), and has regularly featured in the literature, art, and religious life of Europeans throughout history (Crane 1999). But many readers will be only vaguely aware that the Western honey bee represents just a fraction of the total diversity of honey bees. This diversity is centered in Asia, particularly southeast Asia and the Indian subcontinent, where at least eight other species of honey bee are extant. Both the husbandry and the scientific investigation of A. mellifera have led to the publication of hundreds, probably thousands, of books on its biology, and on how to keep these bees for profit or as a hobby. But books on the Asian species, at least in English, are limited to just three. Two of these have more on A. mellifera than on the other species combined, probably because, until recently, most Asian species had received so little scientific attention that a book about their biology would have been a slim and speculative volume indeed. But the 1990s changed that. Teams in China, Japan, Malaysia, and Thailand, in particular, combined with increasingly interested scientists from Western countries, have contributed to an explosion of research into Asian honey bees. From 1994 to 2004, nearly 250 scientific papers were published on Asian honey bees. At least six international conferences were organized to discuss them, and a new scientific society, the Asian Apicultural Association, was formed to foster their study. Many of the scientific reports about Asian honey bees are most prelimi-

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Preface

nary in nature, often the result of flying visits from Western scientists attending conferences in the region who happened to notice something they found interesting. Nonetheless, we are now seeing publication of the results of much more long-term studies involving sophisticated techniques. Pleasingly, a number of long-term collaborations (such as that between the authors of this book) between Asian and Western-based scientists have developed, and some of these have been both productive and fun. In this book we hope that we have synthesised most of what is scientifically known about Asian honey bees. We emphasize the scientific findings, but have tried to cover the cultural and economic values of these bees as well. Our aim is to provide a reference volume for researchers working on Asian honey bees, a general introduction to their biology for interested beekeepers, and an analysis of the significance of Asian bees both to human affairs and to the environment for anyone interested in them. Most of the actual writing of this book was done while Ben Oldroyd was on sabbatical leave at the Dyce Bee Laboratory at Cornell University. We thank Nick Calderone for his hospitality, and the use of his facilities, including the salubrious accommodations of the infamous Dyce Trailer. While Oldroyd was at Cornell, the writing of the book was facilitated by the amazing reprint collection accumulated and meticulously catalogued by the late Roger Morse at the Dyce Lab, and by the unbelievable riches of the beekeeping collection of the Mann Library. We also thank Inga Fine, the interlibrary loans librarian at the Badham Library of Sydney University, for her patience and efforts beyond the call of duty on our behalf. Our research into Asian honey bees has received generous support over many years from the Australian Research Council, the Thailand Research Fund, and the Royal Thai Golden Jubilee Project. All the chapters were read several times by Madeleine Beekman, and her comments, ideas, and encouragement are greatly appreciated. The book is much better for the efforts of the following people who provided us with comments on one or more chapters or illustrations or with other assistance: Denis Anderson, Chanpen Chanchao, Mathew Crowther, Jessada Denduangboripant, Sureerat Deowanish, Orawan Duangpakdee, Michael Engel, Bill Hughes, Gerald Kastberger, Gudrun Koeniger, Niko Koeniger, Chariya Lekprayoon, Stephen Martin, Mitsuo Matsuka, Marina Meixner, Jun Nakamura, Piyamas Nanork, Gard Otis, Mananya Phiancharoen,

Preface

Masami Sasaki, Tom Seeley, Moushumi Sen Sarma, Siriporn Sittipraneed, Ratna Thapa, Graham Thompson, Dhama Visuthakawee, and Tadahara Yoshida. Malcolm Ricketts prepared the photographic plates. Niko Koeniger and Chariya Lekprayoon provided extensive help in the production of the keys given in the appendixes. The index was prepared by Clodagh Jones. We warmly thank you all for your efforts and time.

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 Asian Honey Bees



1 To Be a Honey Bee

In this chapter we introduce the subjects of this book, the honey bees, genus Apis. We define their essential characteristics—their taxonomic position, their gross morphology, and the common aspects of their behavior. Honey bees are social creatures and in order to understand them we need to appreciate that they live not as individuals, but as part of a society from which they are inseparable. We describe the fundamental structures of their societies, at the core of which is the very strong reproductive dominance of the queens, an unusual method of sex determination, and extreme multiple mating by the queen. We describe how these factors affect the genetic architecture of colonies. And finally we show how work is allocated among the various members of honey bee societies. The honey bees, classified as the tribe Apini within the family Apidae, are part of the grand order of membranous-winged insects, the Hymenoptera, which encompasses the ants, bees, wasps, and sawflies. There is only one genus of honey bee, Apis, which comprises the eight Asian species that are the subject of this book, and the Western honey bee, A. mellifera. A. mellifera has been the topic of so many other books that we will refer to it only in passing, treating it as the vanilla honey bee, to whose biology the biology of the Asian species can be compared. What defines the genus Apis? Some of the most diagnostic criteria for workers are: the long, erect hairs that cover the compound eyes, the strongly convex scutellum, the pollen press on the hind leg, the jugal lobe in the hindwing, and the strong wing venation that is stretched distally in the forewing so that the marginal cell is greatly elongated (see Figure 1.1 and Engel 1999; Michener 2000; Ruttner 1988). More obvious than morphol-

2

To Be a Honey Bee

Hairy eyes Elongated marginal cell

Scutellum

Jugal lobe Pollen press

Figure 1.1 A generic worker honey bee, showing some of the key morphological characteristics of the genus Apis.

ogy however, is a suite of behaviors that sets honey bees apart from all other bees, and indeed, all other life forms (see Table 1.1). All the honey bees are highly social, and this is their defining behavioral characteristic. Without contact with its nestmates a honey bee can survive for a few days at most, even when food is plentiful and the temperature amenable. A solitary honey bee certainly cannot reproduce: every individual is locked into colonial life, utterly dependent on the resources and succor of its colony. There are no hermit honey bees. This total interdependence of individuals has led many authors to think of honey bee colonies as “superorganisms” (Moritz and Fuchs 1998; Moritz and Southwick 1992; Queller and Strassmann 2002; Seeley 1989; Wheeler 1928), and in many ways the analogy is apt. A metazoan organism has specialist tissues for different tasks. The mouth and gut gather and digest food, the liver and kidneys remove waste, white blood cells defend against disease, the epidermis provides a barrier against the external environment, and so on. Within a honey bee nest different groups of bees engage in many parallel functions: Foragers search the environment, bringing food to the nest, where it is pro-

To Be a Honey Bee Table 1.1

The behavior that separates honey bees from other social bees.

Dance language for communication of direction and distance Nest of vertical comb made from wax secreted by workers; no nest covering Recycling of comb cells used for brood rearing; food storage cells and brood cells similar, and in some species the same New colonies usually founded by an old queen accompanied by a large swarm of her workers Clustering of workers during swarming and for thermoregulation Progressive feeding of worker larvae so they are fed as much as they need and no more Extreme multiple mating Nest thermoregulation by fanning and water evaporation from water collected in the field

cessed and stored by other bees in a way that is loosely similar to digestion. Nurse bees clean the nest, removing debris and dead or diseased brood, performing a function analogous to that of phagocytes in removing dead cells. And the external curtain of bees on a cluster has some of the properties of skin. Highly interdependent sociality such as the form seen in honey bees is often called “eusociality.” Exactly what is meant by eusociality is difficult to pin down because the term has been used to describe cooperative breeding behavior across such a broad range of taxa (including a shrimp, some aphids and thrips, a beetle, all termites, social Hymenoptera, and two mole rats) that the definition needs to be a broad one in order to accommodate the huge diversity of systems (Costa and Fitzgerald 1996; Crespi and Yanega 1995), and so is perhaps not useful (Sherman et al. 1995). Nonetheless, no one would argue that honey bees are not eusocial, and they fit smoothly into the criteria for eusociality listed below, which are similar to the original definitions given by Wilson (1971, 1975). First, eusocial species cooperate in the rearing of the young. The brood is reared en masse, each individual being cared for by a multitude of workers, and with no one larva (except those destined to be queens) receiving any special attention. Second, eusocial societies have a pronounced reproductive division of labor in which one or a few individuals monopolize reproduction while other individuals are effectively sterile for most or all of the time. The more reproduction is skewed toward particular individuals the more eusocial we consider the society. Although workers of all honey

3

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To Be a Honey Bee

bee species are effectively sterile, they retain vestigial ovaries. These worker ovaries are functional and can be activated to produce small numbers of eggs that are sometimes reared to be fully functional males (see Chapter 7). This contrasts with some ant and stingless bee species, where workers cannot ever produce eggs because they lack ovaries altogether. Species that tend to the other end of the continuum of sociality include many paper wasps in which a number of mated workers share reproductive dominance (Gadagkar 2001), and some stingless bee species in which workers lay many of the male-producing eggs (Drumond, Oldroyd, and Osborne 2000). Because of the reproductive division of labor, eusocial societies have queens. Queens are behaviorally, physiologically, and morphologically specialized for egg production (Figure 1.2). The more divergent queens are from their workers, the more “highly” eusocial we consider the species. In honey bees, queens are profoundly different from workers. For example, whereas workers of A. mellifera have about 6 ovarioles per ovary, queens have 160–180 (Snodgrass 1956). Queens lack the pollen baskets found on the hind legs of workers, and have shorter tongues. This means that they

Figure 1.2 An Apis cerana queen surrounded by her “court” of workers. Photo by S. Wongsiri.

To Be a Honey Bee

cannot forage at flowers, and must be fed by workers in order to obtain their sustenance. Third, eusocial societies have overlapping generations. This means that workers are surrounded by their sisters and brothers, not their own offspring. This has important consequences, because it means that a particular worker is just as related to a sister queen or brother drone born 6 months hence as she is to a sister queen or brother drone born today (Queller 1989). Thus workers need not see their colony produce offspring queens and drones in their own lifetime in order to successfully transmit their genes to the next generation. In evolutionary terms, it works equally well for a worker if her colony’s reproduction occurs long after she is dead as it does if reproduction occurs while she is alive. Cooperative brood care and caste dimorphism between workers and queens, as seen in honey bees, results in the unavoidable interdependence of all individuals. Honey bees are phylogenetically constrained to be social: there is no escape from society because neither queen, drone, or worker can reproduce or even exist on its own (Cameron and Mardulyn 2001). Unlike bumble bee and most ant queens, honey bee queens cannot forage, and are therefore completely incapable of starting a nest on their own; the support of a swarm of workers is obligatory. Similarly, the workers need a queen to reproduce because their own ovaries are vestigial and they lack a sperm-storage organ. Thus a worker is trapped by her body. She can never drop out of society to start a new one on her own. An orphaned honey bee worker has but two choices: get adopted by another colony or perish. Drones are even more dependent. Although if they leave the nest they could potentially find a mate, they would need to do it quickly because their short tongues and stout bodies mean that they are incapable of foraging and will starve a few hours after being evicted from a nest. Thus for a honey bee to escape social life is almost as unlikely as a human finger dropping off and successfully making its own way in the world. It doesn’t happen.

Drones Drones are recognizable by their large eyes and short tongues. They arise parthenogenetically from unfertilized haploid eggs. This means that both

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To Be a Honey Bee

unmated workers and queens can produce male-producing eggs. Queens control the fertilization of their eggs by means of a valve that regulates the passing of sperm from the spermatheca, where sperm is stored, into the median oviduct, where fertilization takes place (Camargo and Mello 1970). If the valve is opened, the egg is coated with a minute volume of semen containing just a few spermatozoa as it passes down the oviduct (Harbo 1979). If the valve is closed, the passing egg does not come into contact with any sperm and will develop into a male. Queens seem to have complete control of this process, and one rarely if ever sees a male developing where it shouldn’t owing to an improperly laid egg (Ratnieks and Keller 1998). In all species except A. dorsata and A. laboriosa, males are reared in special cells on the periphery of the brood nest. These special drone cells are about a third larger than those in which workers are reared. The peripheral location of the drone cells means that males probably experience more variable conditions during brood development than most workers do. Workers are reared in smaller cells away from the edge of the comb where temperature and humidity are better regulated. Rearing temperature is critical for the correct development of the brain of workers, and those reared at even slightly suboptimal temperatures are unable to forage correctly and become lost (Tautz et al. 2003). Perhaps this means that the neural capacities of drones are less valued than those of workers. In those species where the drone cell is larger than the worker cell the adult drone is also larger than the adult worker. In A. dorsata and A. laboriosa, where the rearing cell is the same size, the adult worker and drone are pretty much the same size. The sole function of the males is to mate: they do no work around the colony. The morphology of the honey bee penis is unique to the genus (Ruttner 1988). It is a large, fragile, and membranous structure that is totally contained within the abdomen. Upon mating (or gentle squeezing between the fingers), the penis emerges with explosive force (and often an audible snap). The everted honey bee penis is an amazing organ that is fully a fourth the mass of the insect. In addition to the bulb, on the tip of which the pink spermatozoa float on a puddle of white mucus, there are two or more bizarre, filamentous protrusions that are species-specific, and presumably provide a significant or insurmountable hurdle to interspecific matings. The rest of the structure defies our verbal description, so please

To Be a Honey Bee

see Figure 6.7 on page 113 if you are preemptively curious about the diversity of bee penises. The eversion of the penis kills the drone, for his abdomen almost turns inside out.

Queens Social insects have different morphological forms called castes. In some ants and termites there are significant morphological differences among workers (for example, soldiers and minor workers), but honey bees have only two female castes, the queen and the workers (Figure 1.2). Queen honey bees are about the same size as the drones, but their abdomens are more tapered and wasp-like. We think of queen honey bees as more basal hymenopteran females than the workers, for they lack the derived modifications of workers: they have no corbiculae for gathering pollen, they lack the glands that produce wax or brood food, the crop for nectar, and the barbed sting of the worker. (The barbs of the worker’s sting help it work into the flesh of its victim and hold the venom sack there.) In contrast, the queen’s sting is a curved needle that can easily be withdrawn, making it perfect for stinging other queens. The primary adaptations of queens as egg-laying supremos relate to their massively expanded ovaries, their spermatheca, the sperm-storage organ, and their variety of glands used not for feeding brood but for signaling workers and drones in a variety of ways about their presence. Queens are reared in special cells. In all species they are in shape, size, and texture like peanut shells, and hang downward from the comb, usually from the outer margin. Thus developing queens, like developing drones, probably experience a more variable environment than most workers do. Like workers, queen honey bees begin life from a fertilized egg laid by the queen. The caste fate of an egg is determined entirely by its rearing conditions. If the queen lays the egg in a queen cell and the workers choose to rear it (rather than eat it), the resulting larva is so lavished with food that it literally floats in it for its entire larval life. The food is comprised entirely of secretions of the hypopharangeal glands of the workers, and this ad libitum feeding with the enriched food results in a developmental pathway that leads to a queen. If, on the other hand, the egg is laid in a worker cell, it is progressively provisioned with a more spartan diet that includes

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To Be a Honey Bee

regurgitated pollen and honey in addition to the glandular secretions. This feeding regimen dictates development of the egg as a worker. How the influence of diet and rearing cell so utterly changes the developmental pathway of female honey bees remains a profound mystery of developmental biology, though progress in understanding it is being made (Evans and Wheeler 1999).

Sex Determination and Kin Structure We mentioned above that drones arise from unfertilized eggs and females from fertilized ones. This means that (like all Hymenoptera) honey bees are haplodiploid: the females are diploid and the males haploid. In all species thus far examined—A. dorsata and A. florea (Fahrenhorst 1977), A. cerana (Hoshiba and Okada 1986), and A. mellifera (Hoshiba and Kusanagi 1978)—the basic number of chromosomes is 16, so males have 16 nonhomologous chromosomes and females 16 pairs—32 chromosomes. Haplodiploidy dictates an unusual genetics in honey bees. It creates asymmetries of relatedness among colony members, an unusual system of sex determination and a peculiar meiosis in males. In females, meiosis is normal: maternal and paternal chromosomes pair, exchange information via crossing over, and eventually divide into four daughter haploid cells. In males, on the other hand, the first meiotic division is bizarre. The 16 chromosomes of the haploid spermatocyte replicate as usual so that each chromosome is comprised of two chromatids. The cell divides, but all the chromosomes go to one daughter cell. The other cell forms a bud of cytoplasm that is lost. The second meiotic division is then normal. The cell divides, the two chromatids of each chromosome separating to opposite poles to produce two haploid spermatozoa, each with exactly the same genetic information as the father, and half that of his mother (White 1973). Drones, then, can be regarded as packets of cloned male gametes of a queen. Haplodiploidy requires an unusual sex-determination mechanism as well. In honey bees the Complementary Sex Determining (CSD) locus provides the primary signal of sex determination (Beye et al. 2003). Individuals that are heterozygous at the CSD locus are female. Individuals that are hemizygous (that is, haploid) are male (Cook 1993). Individuals that are

To Be a Honey Bee

Queen

Drone offspring

Fathering drone 1

Worker subfamily 1

Fathering drone 2

Worker subfamily 2

Figure 1.3 A honey bee colony pedigree. The queen (dark shading) is the mother of all colony members. She mates with many drones, each of which fathers a subfamily of workers (gray shading). Workers of the same subfamily are supersisters (relatedness = 0.75). Workers of different subfamilies are half sisters (relatedness = 0.25).

diploid but are homozygous at the CSD are also male, but these are eaten at the first larval instar so that adult diploid males have never been seen in nature (Woyke 1980b). Presumably diploid males, if they were ever naturally reared to maturity, would be sterile (as they are in the stingless bees) and do no work, so it is adaptive to remove them early. This is made possible by the fact that Apis larvae are progressively provisioned in open cells. Thus diploid male larvae can be removed before there has been much investment in them (Ratnieks 1990b). The sex-determining mechanism has other important effects on honey bee biology. Foremost, the consequences of inbreeding are particularly severe in honey bees, because a queen mating with a single male carrying the same sex allele as herself would suffer loss of 50 percent of her diploid larvae being useless, sterile, diploid males. Colonies headed by such queens are unable to grow to swarming strength and are therefore unable to reproduce (Tarpy and Page 2002; Woyke 1980a). Not surprisingly, honey bee mating biology is well adapted to minimize the possibility of inbreeding and reduce the frequency of diploid males to a low level: queens mate on the wing with a large (6–100) number of unrelated males (Baudry et al. 1998; Palmer and Oldroyd 2000; Wattanachaiyingcharoen et al. 2003), so outbreeding is usually assured. Haplodiploidy and multiple mating mean that the workers of a honey bee colony, far from being clones, are an eclectic genetic mix. Consider Figure 1.3, which shows the major relationships among nestmates. Queens are related to both their sons (drones) and daughters (workers and daughter queens) by one half. The worker population is made up of groups of half sisters, divided by their paternities. The daughters of each drone are re-

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To Be a Honey Bee

ferred to as a “subfamily” or a “patriline,” and these are related by three fourths, because the genetic information that comes from their father is identical but that from their mother has a 50–50 chance of being shared. Workers sired by different fathers share no genetic material via their fathers, and are related as half sibs by one fourth.

Division of Labor An efficient society is characterized by specialization: not all individuals perform all tasks at all times. In human societies specialization usually follows from a combination of opportunity, aptitude, and training that allows most individuals to gain some or a great deal of expertise in their chosen vocation and thus become adept at performing a particular class of work. In addition to their expertise, human specialists will often invest in the necessary tools of their specialty (for example a welding set or a violin), which most other members of the society would not possess, or know how to use. Exchange of goods and services among the different specialists of society increases the overall wealth, and allows nonspecialists access to goods and services that they would not otherwise have. A society of interacting specialists needs methods of task allocation so that all the necessary tasks of the society are performed, for the efficiencies that accrue from specialization will be greatly diminished if, for example, the number of aromatherapists or professional football players becomes too large relative to the number of plumbers. In human societies people are allocated to tasks in two broad ways. First, controlling humans pass laws and regulations that help to specify the number of other humans that can engage in particular tasks. For example, the medical and legal professions regulate their own numbers by controlling those that can enter their ranks, while governments may seek to alleviate labor shortages or surpluses by a targeted immigration program or by increasing or decreasing the number of training places available at colleges. Second, task allocation is largely self-organized. If there is a genuine need for a certain specialization, then salaries will tend to rise for those that can do it, attracting workers from tasks where there is an oversupply of labor. Honey bee societies are also characterized by specialization that increases the overall efficiency of the society. The most important driver of

To Be a Honey Bee

specialization is the age of the individual (Lindauer 1967). Young workers perform in-hive tasks like cleaning cells, feeding larvae, and building comb, whereas older workers are involved in foraging. The endocrine system of the individual changes over her lifetime so that she is equipped with the right toolkit to perform each task. Young workers have active hypopharangeal glands so that they can feed younger brood. As the individual ages, her wax and poison glands become active, allowing her to build comb and guard the nest. The final developmental step to forager sees loss of most exocrine gland function, and changes in brain chemistry that allows the individual to learn the scents and flavors of nectars and the location of both flowers and the nest, and to be motivated to leave the nest to forage (Elekonich et al. 2001; Fahrbach and Robinson 1996; Giray et al. 1999; Giray and Robinson 1996; Huang, Plettner, and Robinson 1998; Robinson 1992; Schulz and Robinson 1999). Within the broad framework of age-specifying task in honey bee colonies, allocation of work is self-organizing. A self-organized system is one that acquires order and structure “through interactions internal to the system without intervention by external directing influences” (Camazine et al. 2001, p. 7). The organization of work in a honey bee colony is therefore fundamentally different from the organization of a human factory, in which a great deal of task allocation is directed by sub-bosses who allocate tasks according to broad directions given by bigger bosses. Take, for example, the well-studied mechanisms by which colonies allocate the available foragers between alternative sources of food (Camazine and Sneyd 1991; Seeley 1995; Seeley, Camazine, and Sneyd 1991). Upon returning from a successful foraging trip a forager may perform a waggle dance that informs other foragers about the location of the food. In broad terms the probability that she will perform such a dance depends first on her independent assessment of the quality of the food source and second, on how quickly she was relieved of the rewards of her foraging trip by house bees. If many rich sources of food are known to the colony, the successful forager will experience a delay in being unloaded, and this reduces the probability that she will dance, even if she perceives her forage patch to be extremely profitable. If, on the other hand, the house bees are underemployed they crowd round the successful forager, rapidly unloading her— and this increases the probability that she will dance, even for what she perceives as being a quite modest forage patch. The outcome of the inter-

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To Be a Honey Bee

action of these two phenomena is that the foragers which have found the most profitable patches are the ones that will dance and recruit new foragers to their patches. Furthermore, a forager that fails to find food at a patch will stop foraging there and be available for recruitment. The emergent property of these processes is that the most profitable forage patches attract more recruits and more dancers, which builds the number of foragers there. Less profitable patches attract no recruits, and will lose foragers, which leads to a gradual diminution of foraging effort at less profitable patches.

Summary Honey bees are a very distinct eusocial genus of the Hymenoptera. They live in large societies that in some ways can be regarded as superorganisms—the society as a whole is subject to natural selection—and within the society, groups of specialist individuals perform functions that are loosely analogous to the organs of metazoan organisms. There are three kinds of adult honey bees: queens, workers, and drones. Queens and workers are known as castes. They arise from the same kind of eggs, and their different and irreversible developmental trajectories are determined solely by their feeding and other treatment as larvae. Queens are diploid and mate with many haploid males. This leads to asymmetrical levels of relatedness within colonies and has important consequences. The reproductive interests of all individuals are not identical because some of the colony’s offspring are more related to particular individuals than to others. Colonies allocate work among workers on the basis of the principles of self-organization. Workers respond to proximate cues around them, which determine whether or not they will engage in a task. An emergent property of the interactions of many workers is that the colony as a whole tends to allocate workers to tasks of greatest need in a way that maximizes the colony-level gains of each individual’s labor.



2 Introduction to the Species

Our primary goals for this chapter are to provide you with sufficient information to make a quick species identification of any Asian honey bee, on the basis of comb, nest, or worker morphology, to provide a reference for each species’ distribution, and to give a comparative description of each species’ behavior. In defining a species, we will use the “biological” species concept (Mayr 1942). That is, if two animals can mate and produce fertile offspring, then we will regard them as being of the same species. If not, even if the barriers to mating are behavioral, such as a different time of mating, we will allow that they are of different species. We will not, however, regard mere geographical separation as a mating barrier. The species definition that we have chosen to use in this book is often regarded as dated (Noor 2002). In particular, the biological species concept is clearly inappropriate for asexual species and extinct species, where it has no meaning. For this reason, most taxonomists now prefer the “phylogenetic species concept,” in which a species is defined as a distinct lineage of organisms that differ from all others by one or more “taxonomically important” traits (Cracraft 1989). The problem with the phylogenetic species concept is that it requires an arbitrary definition of what is taxonomically important. Its potential danger is that it can lead to the recognition of a plethora of new species (Isaac, Mallet, and Mace 2004). We think that the biological species concept is the more useful definition for extant honey bees, because the species boundary is explicitly defined, at least conceptually. It is also the more widely used by honey bee specialists. But we emphasize that in a world of biological continuum, the definition of a species is necessarily arbitrary (Hey 2001; Oldroyd 1993). For example, two of the

14

Introduction to the Species

Table 2.1

The genus Apis Linnaeus.

Group

Subgenus

Species

Author

Common name

Dwarf honey bees

Micrapis

Apis florea Apis andreniformis

Fabricius (1787) Smith (1858)

Red dwarf honey bee Black dwarf honey bee

Giant honey bees

Megapis

Apis dorsata

Fabricius (1793)

Apis laboriosa

Smith (1871)

Common giant honey bee Giant mountain honey bee

Apis cerana Apis koschevnikovi Apis nuluensis

Fabricius (1793) Enderlein (1906) Tingek, Koeniger, and Koeniger (1996) Smith (1861) Linnaeus (1758)

Cavity-nesting honey bees

Apis

Apis nigrocincta Apis mellifera

Eastern hive bee Red honey bee Mountain honey bee

Sulawesian honey bee Western honey bee

species that we have recognized in this book would be lost had we adopted the phylogenetic rather than the biological species concept, and several as yet unnamed ones would probably have been erected. Nonetheless, it is important to remember that whatever species names or definitions we had chosen to use and classify honey bees, their biodiversity would have remained unchanged. On the basis of our adopted species definition we recognize nine species of honey bee, in a single genus, Apis. The number of honey bee species proposed has waxed and waned over the last 100 years, reaching a peak in Maa’s (1953) profligate recognition of 3 genera and 24 species, and a trough in Buttel-Reepen’s (1906) overly frugal assessment of a single genus and 3 species. Even as recently as 1988, Ruttner’s authoritative text on honey bee taxonomy recognized but 4 species in 1 genus. The species we recognize and their authorities are summarized in Table 2.1. A formal taxonomic key to the workers of the species is given in Appendix A.

Species Identification In most cases, species identification of a nest is reasonably straightforward, and can be done by referring to the pictures in Figure 2.1 and the distribu-

Introduction to the Species

A

10 cm

Exit hole

C

B

50 cm

10 cm

Figure 2.1 The nest architecture of the three groups of honey bees. A: Dwarf bees. The comb is built around a small branch. B: Giant bees. The comb is built beneath a branch or overhang. C: Cavity-nesting bees. Three or more parallel combs are built, usually in a cavity.

tion maps that appear later in this chapter. Identification of an isolated worker can be much more challenging. Closely related species have similar morphologies, and identification often rests on qualitative rather than quantitative differences. We provide tables of those features that are most useful for species identification, and a complete key in Appendix A. We have avoided detailed morphological descriptions or the use of multivariate statistics to define species, because these are of little practical use in the field. Features that are often used in adult honey bee taxonomy are the size of the wings, the lengths and the angles formed by the veins therein, the length of the tongue, the number of hooks (hamuli) that join the hindwing with the forewing, and the morphology of the male genitalia. Figure 2.2 shows the important features of the wings and tongues that are used in the species identifications described below. Figure 6.7 shows the male genitalia that are known, but we do not use male genitalia for our diagnoses because they are so rarely available and difficult to describe (Koeniger, Mardan, and Ruttner 1990).

Three Groups of Honey Bees There are three groups of honey bee species: the giant honey bees that build a large, single comb in the open, the dwarf honey bees that build a

15

Introduction to the Species

width

Forew ing

Forew ing

16

B

length

A

A:B = cubital index Hamuli Distal abscissa vein M

Proboscis length

Drone basitarsi

A. andreniformis

A. florea

Figure 2.2 Diagnostic features for species identification. Top: The forewing and hindwing showing the hamuli, the two veins (A and B), the ratio of which is the cubital index, the distal abscissa of vein M, and the length and breadth of the wings. Bottom left: The proboscis length. Bottom right: The contrasting lengths of the “thumbs” on the basitarsi of the males of A. andreniformis and A. florea.

Introduction to the Species Table 2.2

17

Quick identification of honey bee subgenera (all sizes approximate). Subgenus

Characteristic

Micrapis

Megapis

Apis

Worker body length (mm)

7–10

16.5–17.5

10–14

Worker forewing length (mm)

6–7

12–15

7–10

Comb (see Figure 2.1)

Single, less than 25 cm across, built around small branch, never closely aggregated

Single, more than 30 cm across, built under branch, rock face, or building, often aggregated

Multiple, almost always in a cavity, never closely aggregated

small single comb in the open, and multicombed cavity-nesting honey bees of intermediate size (Table 2.1, Figure 2.1). These three groups are currently recognized as three subgenera of the genus Apis (Engel 1999). The subgenera are Megapis (giant) and Micrapis (dwarf) and Apis (cavity-nesting). In the past the three subgenera have sometimes been regarded as genera (Ashmead 1904; Maa 1953; Skorikov 1929), but given the small number of honey bee species, these divisions are hardly necessary. When one is citing a species, the use of a subgenus name is optional, and usually unnecessary for honey bees. Examples of usage of complete names, giving both the genus and subgenus designations, are Apis (Micrapis) andreniformis for the black dwarf honey bee and Apis (Apis) cerana for the eastern hive bee. The first step to identification is to classify the bee to one of the three groups, and this can be easily achieved by reference to Figure 2.1 and Table 2.2. The descriptions below and reference to our tables or Appendix A should then allow identification of species.

Dwarf Honey Bees, Micrapis (Ashmead 1904) These small bees are by far the most common honey bees over most of tropical Asia. The top of the single comb is built around a small tree branch. Below the branch hangs a shroud of tiny bees, and foragers can be seen coming and going from the platform formed by the crown of the comb.

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Introduction to the Species

Figure 2.3 A relocated nest of Apis florea in Bangkok, Thailand. Photo by B. Oldroyd.

The two dwarf honey bee species A. florea and A. andreniformis (Figures 2.3 and 2.4) are superficially similar, and were not generally recognized as separate until Wu and Kuang (1987) reinstated the name A. (Micrapis) andreniformis (Smith 1858). Wongsiri et al. (1990) verified the validity of the species status of the two dwarf bees by showing separate mating times. The male genitalia of A. florea and A. andreniformis are different (see Chapter 6), and the workers differ strongly in their behavior. For example, a colony of A. andreniformis is defensive of its nest, whereas A. florea is not particularly so. They even fan their nests differently: fanning A. florea face with their heads up the nest whereas A. andreniformis face down (Thapa and Wongsiri 1994). In the field, A. florea workers are distinguished from A. andreniformis workers by A. florea’s generally reddish-brown (rufous) appearance, and the pale yellow to white hairs on the hind tibia and basitarsis, rather than the black hairs characteristic of A. andreniformis (Smith 1858; Wu and Kuang 1987). A note of caution however: although A. florea is generally rufous, and A. andreniformis generally black, the scutellum is an exception,

Introduction to the Species

Figure 2.4 A nest of Apis andreniformis near Chanthaburi, Thailand. Photo by B. Oldroyd.

and the colors are reversed. Moreover, some colonies of A. andreniformis contain a few rufous workers, and we suspect that there must be some kind of color polymorphism present in that species. But even in these rufous A. andreniformis the first abdominal segment is always black, whereas it is always rufous in A. florea. Notwithstanding the distinctive coloration and behavior of A. andreniformis and A. florea, most measurements of external morphology of the dwarf species overlap, so although species means are usually significantly different (Rinderer et al. 1995), measurement of a single character on a single individual is rarely diagnostic. The notable exception is the cubital index (Figure 2.2), which is much higher in A. andreniformis than in A. florea, and never overlaps (Table 2.3). Apis florea is distributed from the Middle East, east to peninsular Malaysia, whereas A. andreniformis is distributed from Palawan (Philippines) to China and Myanmar (Figure 2.5; Otis 1996). The two species overlap in Southeast Asia (Figure 2.5). Because of confusion of A. florea and A. andreniformis, studies prior to 1986 in the area of sympatry, i.e. Thailand,

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Introduction to the Species Table 2.3

Distinguishing characters of the dwarf honey bees. All entries are the range of population means of workers except where otherwise noted. Where data are available for only one population, the standard deviation is given.

Characteristic

A. florea

A. andreniformis

Workers Body length (mm) Overall coloration Proboscis length (mm) Forewing length (mm) Forewing width (mm) Cubital vein A (mm) Cubital vein B (mm) Cubital index Hind wing length (mm) Hind wing width (mm) Hamuli number

7–10 Red-brown 3.11–3.37a,b 6.17–6.74a,b,c 2.12–2.32a,b 0.49 ± 0.04b 0.18 ± 0.03b 2.86–3.50a,b,c 3.17–4.83 b,c 1.36 ± 0.04b 10.5–13.2a,b

8–9 Black 2.797–2.798f 6.43–6.49f 2.17–2.21f 0.51–0.52f 0.09–0.08f 6.28–6.38f 3.22–3.23f 1.25–1.28f 10.38–10.88f

Drones Thumb on hind basitarsis (see Figure 2.2)

> half the basitarsisd

< half the basitarsisd

12.0 ± 3.3 e

10.0 ± 3.3 e

16.9 ± 5.3 e 0.8 ± 0.07 e

12.2 ± 3.6 e 1.7 ± 1.7 e

Worker cell dimensionse Depth of cell (cm) Width of 10 cells (cm)

0.93 ± 0.07 2.98 ± 0.15

0.76 ± 0.02 2.78 ± 0.23

Drone cell dimensionse Depth of cell (cm) Width of 10 cells (cm)

1.33 ± 0.07 4.88 ± 0.21

1.45 ± 0.71 4.18 ± 0.24

Queen cell dimensionse Depth of cell (cm) Internal diameter of cell (cm)

1.41 ± 0.15 0.47 ± 0.09

1.24 ± 0.26 0.54 ± 0.08

Comb dimensions Vertical length (from support bottom) (cm) Horizontal width (cm) Branch diameter (cm)

a. Ruttner (1988). Numerous colonies from Iran, Oman, Sri Lanka, India. b. Rinderer et al. (1995). Thailand, n = 42 colonies. c. Ruttner et al. (1995); 3–6 colonies each from Iran, Sri Lanka, Oman. d. Wu and Kuang (1987). e. Rinderer, Wongsiri, et al. (1996), A. andreniformis, Thailand, n = 15–19 colonies except for drone cells, 4–6 colonies. A. florea, Thailand, n = 38–42 colonies except for queen and drone cells, 5–10 colonies. f. Rinderer et al. (1995), Philippines, n = 4 colonies; Thailand, n = 36 colonies.

Introduction to the Species

A. florea A. andreniformis

Figure 2.5 The distribution of the dwarf bees. A. florea is in the west, A. andreniformis in the east. The species are sympatric in southeast Asia.

Laos, Cambodia, Vietnam and southern China may have the incorrect species designations. Even in Malaysia where only A. andreniformis is found, it was usually referred to as A. florea prior to 1986. Dwarf honey bee queens and drones are much larger than the workers. Apis florea workers average 25.5 mg, whereas drones are about 80 mg (Koeniger and Koeniger 1993; Koeniger, Koeniger, and Wongsiri 1989), and queens are 86 mg (Koeniger and Koeniger 1993), which means that drones and queens are about three times the size of workers, and exceptionally for the genus, queens are slightly larger than drones (Deowanish et al. 2000). It is always a wonder to us that these large queens, A. cerana–like in size and appearance, can squeeze their abdomens inside the tiny worker cells in order to lay eggs. Males of both species bear a cleft on their hind basitarsus, which is speculated to be deployed to hold the hind legs of the queen while the nuptial pair is flying (Figure 2.2; Ruttner 1988). Nonetheless, since the “thumb”

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Introduction to the Species

formed by the cleft is not articulated and therefore cannot be moved, we regard this hypothesized functionality as speculative but reasonable. A solid reason for positing the thumb’s function as a grasping structure arises from the fact that the penis of dwarf bee drones lacks the large bulbus of all the cavity-nesting and giant species (Koeniger and Koeniger 1990; Koeniger and Koeniger 2000b; Koeniger, Koeniger, and Wongsiri 1989). In the cavity-nesting and giant species the bulbus serves to briefly lock the queen and drone together in copula, and we might imagine that without something to hang on with, in-flight sex is a tricky affair; hence the proposed need for the thumb. A. florea males can be distinguished from A. andreniformis males because the thumb is much shorter in A. andreniformis (Figure 2.2). The combs of A. andreniformis and A. florea are quite different (Table 2.3, Figure 2.6). The average dimensions of A. florea combs are 25% larger than those of typical colonies of A. andreniformis (Table 2.3; Rinderer and Wongsiri et al. 1996). Where they occur, racemes of coconut palm inflorescences are a particularly favored nest site for A. florea, but seem to be avoided by A. andreniformis. When A. florea combs are extended they may incorporate other secondary branches within the comb structure. This has not been reported for an A. andreniformis nest, and we have never seen it. In A. florea, the crown of the comb, where honey is stored, is a messy affair. As a consequence of the comb-building process, the crowns do not contain a comb midrib (the structure formed by the bases of the cells on opposing sides of the comb). The first cells to be constructed are joined directly to the supporting branch (Akratanakul 1977) and are lengthened as the comb grows. Since the cells retain the same diameter along their entire length (they are not fluted), a gap forms between the cells toward the outer margin. These gaps are filled with additional cells, leading to the characteristically irregular arrangement of the cells of the crown of the nest, greatly distorted from the usual regular hexagons that we normally associate with honey bee combs (Wongsiri et al. 1997). A. andreniformis, in contrast, builds a midrib up from the supporting branch, which permits the construction of regular cells (Figure 2.6). In both species, the brood comb that is suspended below the supporting branch contains a midrib, and the regular hexagonal arrangement of cells that we expect of honey bee combs. In the mature nest the outer perimeter of cells is enlarged to provide for the rearing of drones, and eventually

Introduction to the Species A

B

Figure 2.6 The contrasting construction of A. andreniformis and A. florea combs. A: In A. florea, cells are joined directly to the support, with additional cells constructed to fill the gaps on the outer margins. B: In A. andreniformis, a midrib is built up from the supporting branch. After Rinderer, Wongsiri, et al. (1996).

queen cells will be built before the colony swarms. It is not known if the presence of drone cells means that the nest has climaxed, becoming incapable of further expansion of worker cells, or if the drone cells can be torn down and recycled to provide for more worker cells.

Giant Honey Bees, Megapis (Ashmead 1904) A populous colony of giant honey bees is one of the most impressive sights of the insect world. Although many ant, termite, and even bee colonies have many more individual workers, most of them are hidden from view inside their nests, or dispersed on foraging trails. But the magnificent exposed nest of a giant honey bee is truly awe-inspiring because a large proportion of the individuals are visible at once. By honey bee standards, individual workers are huge (17 mm long). In large colonies the combs are massive (1.5 m × 1 m) and the number of individual workers can be over 50,000 (Morse and Laigo 1969). While even a single colony is a thing of

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Introduction to the Species

Figure 2.7 An aggregation of A. dorsata colonies on a tree in northern India. Photo by G. Kastberger.

wonder, giant honey bee colonies have a strong tendency to be highly aggregated, sometimes with 200 or more colonies crammed onto a single water tower, tree, or rock face. Apart from their great size, the giant bees are distinguished from all other honey bees by their wings, which are fuscous and quite hairy. There are two species: A. dorsata, the common giant honey bee, which is found throughout south and southeast Asia (Figure 2.7), and A. laboriosa, the giant mountain honey bee, which is found in mountainous regions, particularly the Himalayas, above 1500 m (Figure 2.8). Neither Engel (1999) nor Ruttner (1988) recognized A. laboriosa as a species distinct from A. dorsata, holding that most morphological characters blend “naturally into one another” (Engel 1999). Nevertheless, the case for species status of A. laboriosa is quite strong. First, there is evidence of mating barriers between the two. Underwood (1990b) observed Nepalese A. laboriosa males taking what were presumably mating flights between 12:30 and 14:30 (albeit on 2 days only). Although A. dorsata males

Introduction to the Species

Figure 2.8. Some A. laboriosa colonies on a rock face in Nepal. Photo by G. Kastberger.

also fly during the day to orient and defecate (Mardan and Kevan 1989), their mating flights invariably occur just after dusk (Koeniger and Wijayagunesekera 1976; Rinderer et al. 1993; Tan et al. 1999). Frustratingly, Underwood was unable to make observations on the behavior of A. laboriosa drones after dusk owing to poor weather, so there is a tiny cloud over his critical observation. An additional barrier to intraspecific matings may arise from the male genitalia. Although McEvoy and Underwood (1988) felt that the genitalia of A. dorsata and A. laboriosa are indistinguishable, Koeniger et al. (1990) questioned this claim, suggesting that the perceived lack of distinctness was due to incomplete eversion of the A. laboriosa endophalli studied by McEvoy and Underwood. But M. Engel (personal communication) confirms McEvoy and Underwood’s assertion that the endophalli are indistinguishable. Second, investigations of the cuticular and sting-shaft hydrocarbons re-

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Introduction to the Species

veal strikingly different profiles (Carlson, Roubik, and Milstrey 1991) and may suggest that the alarm pheromones of A. laboriosa differ from those of all other Apis (Blum et al. 2000). Third, the communication dance performed by A. dorsata is strikingly different from that of A. laboriosa (Chapter 5; Kirchner et al. 1996). Fourth, DNA sequence divergence between A. laboriosa and A. dorsata is 10.6–11.5 percent, which strongly supports species status (Arias and Sheppard 2005). Table 2.4 gives morphological measurements that are useful but not always diagnostic in distinguishing A. laboriosa and A. dorsata. A. laboriosa workers are about 10 percent larger than A. dorsata workers and the brood cells are 14 percent larger (Underwood 1986). The dorsal surface of the thorax is completely covered in tawny yellow hairs in A. laboriosa, whereas in most A. dorsata, most of the hairs, except on the margins, are dark (Maa 1953; Sakagami, Matsumura, and Ito 1980). The ocelli of A. dorsata males are raised, possibly as an adaptation to mating at dusk. In A. laboriosa they are flat (Sakagami, Matsumura, and Ito 1980), presumably because these bees do not mate at dusk. The hairs on the abdomen are uniformly dark in A. laboriosa but can be yellow in A. dorsata. But the giant bees of the Philippines are also dark, so color is not diagnostic. Sakagami (1980) and Roubik et al (1985) draw particular attention to the larger size of the spiracular plate of A. laboriosa (Table 2.4). In addition to A. laboriosa and A. dorsata, two other species of giant honey bees have been described A. breviligula and A. binghami. A. breviligula (Maa 1953) is found northwest of the Merrill line in Luzon in the Philippines (Figure 2.9). This bee is completely black with gray stripes (personal observations of Oldroyd). A. binghami (Cockerell 1906) is found east of the Wallace line in Sulawesi and Butang, Indonesia (Figure 2.9). Little is known of the biology of these island populations, but interestingly, unlike their continental brethren, they do not appear to form nest aggregations (G. Otis, personal commmunication; Morse and Laigo 1969). The species designations of A. breviligula and A. binghami are based on geographic isolation, color, and some minor morphological differences from A. dorsata. But recognition of A. binghami and A. breviligula as separate species has not been broadly supported because of the similarity of their morphology (Engel 1999; Morse and Laigo 1969; Ruttner 1988), lack of DNA sequence divergence in the ND2 gene of the mitrochondria (Arias

Introduction to the Species Table 2.4

Distinguishing characters of the giant honey bees. All entries are the range of population means of workers except where otherwise noted. Where only one population is available, the standard deviation is given.

Characteristic

A. dorsata

A. laboriosa

4.5–6.7a,b,c,d 9.75–13.2a,b,c,d,e 4.1–4.6d,e 1.14 ± 0.06a 1.80 ± 0.05a 5.28–8.38c,e 7.8–9.84a,d,e 2.49–2.77a,d,f 22.55–26.35a,d,e

6.1–7.1a,b,c 13.0–14.5a,b,c 4.2–4.4a,f 1.30 ± 0.06a,b 1.96 ± 0.07a 8.49–9.82b,c 9.20a 2.54a,f 22.61a

0.76 ± 0.05a 1.29 ± 0.06a 0.79 ± 0.04a

0.90 ± 0.03a 1.35 ± 0.06a 1.0 ± 0.04a

Comb dimensionsl Vertical length (cm) Horizontal width (cm)

51.2–88.0g,h,i,j 40.4–68.9g,h,i,j

25–110k 30–80k

Brood cell dimensions Depth of cell (cm) Width of 10 cells (cm)

1.6–1.9h,i, 5.3–5.9h,i

1.8–2.0m 6.0–6.3j

Morphological characters Proboscis length (mm) Forewing length (mm) Forewing width (mm) Cubital vein A (mm) Cubital vein B (mm) Cubital index Hindwing length (mm) Hindwing width (mm) Hamuli number Spiracular plate (metasomal tergum 7) (mm) Width Length Length of barbed portion of the sting (mm)

a. Sakagami (1980), various locations. b. Maa (1953), various locations. c. Trung et al. (1996), Vietnam. d. Sharma and Thakur (1999), India, several locations. e. Mujumdar and K. K. Kshirsagar (1986), India, many locations. f. Sakagami’s table actually lists A. dorsata as 2.55 and A. laboriosa as 2.61. This appears to be a misprint. g. Calculated from Morse and Laigo (1969), Philippines. h. Doedikar et al. (1977), India, means of several studies. i. Thakar and Tonapi (1961), India. j. Chinh et al. (1996), Vietnam. k. Underwood (1986), Nepal. l. Note that despite these figures mature colonies tend to have a greater horizontal width than they do vertical length. m. Calculated as one half of the comb thickness reported by Underwood (1986).

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Introduction to the Species

A. dorsata A. dorsata breviligula A. dorsata binghami A. laboriosa

Figure 2.9. Approximate distributions of A. dorsata and A. laboriosa. A. laboriosa is confined to mountainous regions. The A. dorsata population on Luzon in the Philippines can be regarded as a subspecies A. dorsata breviligula (Maa 1953). The population on Sulawesi and Butang is often referred to as A. dorsata binghami Cockerell (1906).

and Sheppard 2005), and a lack of solid information on mating behavior. There is, however, some evidence that these subspecies mate at dusk, as do A. dorsata. Males of both island populations have the raised ocelli of A. dorsata (Maa 1953), suggesting nocturnal mating. During Morse and Laigo’s extensive study (1969) of A. d. breviligula, they never saw drone flights. Since all their observations were in daylight hours, this is circumstantial evidence that A. d. breviligula mates at dusk, well after Morse and Laigo had repaired to their guest house for the evening. Otis et al. (2000) saw A. d. binghami males flying at dusk. Confirmation of the mating times of the giant bees of the Philippines and Sulawesi is needed. In the meantime, we recommend that these bees be regarded as the subspecies A. dorsata binghami and A. d. breviligula and not as distinct species. The size dimorphism between castes of the giant honey bees is much less pronounced than in other Apis. Queens (20 mm) are slightly longer

Introduction to the Species

than workers (17 mm); drones slightly shorter (16 mm) (Morse and Laigo 1969). In both A. dorsata and A. laboriosa the cells used to rear drones are identical in size to those used to rear workers (Qayyum and Nabi 1968; Underwood 1986). As with other species, queens tend to lay unfertilized, drone-destined eggs on the bottom margin of the comb, but they are often commingled with fertilized worker-destined eggs. If the comb is being rapidly extended, then arc-like bands of predominantly drone brood will be found between wider bands of worker brood (Chinh, Tan, and Thai 2004). In many cases, especially after brood cells have been reused, drone brood appear to be randomly distributed among the worker brood (Wattanachaiyingcharoen et al. 2001). Worker cell cappings are invariably flush with the top of the brood cell. It has been stated that the capping of the drone cells can be slightly raised relative to those used to rear workers (Qayyum and Nabi 1968; Thakar and Tonapi 1961; Underwood 1986). But in our experience there is no consistent difference in the cappings of cells containing drones and workers. In fact, a larger drone capping is not expected because drones are minutely shorter than workers. Unlike the combs of dwarf honey bees, in which the crown of the comb almost always encircles the support, the combs of A. dorsata are always built on the undersurface of a stout branch or an overhang of a rock face or building, and A. laboriosa combs only on rock faces. Where A. dorsata nests are found in trees, the diameter of the supporting branches varies from 12 to 30 cm (Morse and Laigo 1969) or much larger (personal observations). A slightly sloping branch is preferred (Tan et al. 1997). The largest A. dorsata nest found by Morse and Laigo (1967) in the Philippines was 1.67 m along its branch and 0.66 m in depth, while the average dimensions were 0.7 m along the branch and 0.51 m deep. Honey is stored in one corner of the comb nearest the uppermost section of comb in an area about 10 by 20 cm in a large nest. The honey-storage area is usually quite obvious because the cells are highly elongated: up to 15 cm long. The amount of honey stored is quite modest, up to about 10 kg, and often much less or none.

Cavity-nesting (Hive) Honey Bees, Apis sensu stricto (Linnaeus 1758) Cavity-nesting honey bees are medium-sized bees with forewing lengths of 7–10 mm (Table 2.5). They are usually, but not exclusively, found in tree hollows, and build three or more parallel combs affixed to the roof of the

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30 Table 2.5

Introduction to the Species Distinguishing characters of the cavity-nesting honey bees. All entries are the range of population means of workers except where otherwise noted. Where only one population is available, the standard deviation is given.

Characteristic

A. cerana

A. nigrocinctab

A. koschevnikovid

A. nuluensisi

Overall worker coloration Proboscis length (mm) Forewing length (mm) Forewing width (mm) Cubital vein A (mm) Cubital vein B (mm) Cubital index Hamuli number

Variable 4.06–6.25a,b,c,d 7.57–9.05a,b,c,d 2.66–3.13a,b,c,d 0.44–0.51b,d 0.12–0.15b,d 2.24–4.25a,b,d 16.7–19.6a,b,c,d

Yellow 4.98 ± 0.093 8.12 ± 0.17 2.73 ± 0.067 0.51 ± 0.016 0.14 ± 0.0064 3.84 ± 0.23 17.86 ± 0.68

Reddish to rufous 5.15–5.45 8.54–8.88 2.90–3.08 0.53–0.63 0.07–0.11 5.65–9.54 16–19

Black — 8.08 ± 0.09 2.78 ± 0.07 0.48j 0.12j 3.77 ± 0.12 18j

Worker cell dimensions Depth of cell (cm) Width of 10 cells (cm)

1.01e 4.20–4.87e,f

— 4.5h

1.02h 4.5i

— —

Drone cells Pore in capping

Present

Absent

Present

—

a. Kshirsagar and Ranade (1981), a review article citing studies of numerous colonies from numerous locations in India. The ranges given probably encompass the full variability across Asia. b. Hadisoesilo et al. (1995), Sulawesi. A. cerana, n = 3 colonies; A. nigrocincta, n = 10 colonies. c. Hepburn, Radloff, et al. (2001). 279 colonies from the southern Himalayas. d. Rinderer et al. (1989), Sabah. A. koschevnikovi, n = 9 colonies; A. cerana, n = 4 colonies. e. Inoue et al. (1990), Sumatra. f. Ruttner (1988). g. Unpublished measurement of Soesilawati Hadisoesilo, Sulawesi (n = 1 colony). h. Unpublished measurements of B. Oldroyd and S. Tingek, Sabah (n = 2 colonies). i. Tingek et al. (1996), Sabah. Numerous workers from flowers. Sample size unspecified. j. Single measurement from the holotype.

cavity (Figures 2.1 and 2.10). These bees can adapt to living in cavities in human structures and in purpose-made hives, and their nesting habit means that they can potentially colonize temperate or mountain areas with prolonged winters or cold temperatures. Apis mellifera tends to be slightly larger than the Asian species, and can be readily distinguished from A. cerana by the absence of the distal abscissa of vein M in the hindwing (Figure 2.2). The biological species status of A. mellifera and A. cerana was demonstrated by Ruttner and Maul (1983), who showed that although A. mellifera drones can naturally mate with A. cerana queens, A. mellifera–A. cerana hybrids created by artificial insemination are not viable.

Introduction to the Species

Figure 2.10 A domesticated colony of A. cerana in Thailand. The colonies of all the cavitynesting bees are similar. Photo by B. Oldroyd.

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Introduction to the Species

The four Asian cavity-nesting species (A. cerana, A. koschevnikovi, A. nuluensis, and A. nigrocincta) began their divergence from a cosmopolitan A. cerana type within the last 2 million years (see Chapter 4). The relatively recent origin of the Asian cavity-nesting species means that morphological and indeed genetic differences between them are subtle. On the island of Borneo there are three species of cavity-nesting bees: A. koschevnikovi and A. nuluensis in addition to the cosmopolitan A. cerana. A. koschevnikovi is also present in Java, Sumatra, peninsular Malaysia, and southern Thailand (Figure 2.11). But since it seems to require rainforest habitat, it is now rare outside of Borneo owing to deforestation (Otis 1996). Evidence for the species status of A. cerana, A. koschevnikovi, and A. nuluensis is fourfold. First, all species have strongly divergent mating times and physical separation of their mating places (Koeniger, Koeniger, and Tingek 1994a; Koeniger and Koeniger 2000b; Koeniger et al. 1996b; Koeniger et al. 1994; Koeniger et al. 1988; Mathew and Mathew 1990; Tingek, Koeniger and Koeniger 1996; see Chapters 4 and 6), even when the drones are reared in the nest of their conspecifics (Koeniger, Koeniger, and Tingek 1994). Second, although A. cerana can sometimes be induced to rear a replacement queen from larvae of A. koschevnikovi (Koeniger, Koeniger, Tingek, and Kelitu 1996), interspecific hybrids between A. koschevnikovi and A. cerana produced via artificial insemination have low fertility and hybrid colonies are probably nonviable (Koeniger, Koeniger, and Tingek 1996). Third, the male genitalia of A. koschevnikovi differ from those of A. cerana (see Chapter 6; Mathew and Mathew 1990). Fourth, both morphological traits (Fuchs, Koeniger, and Tingek 1996; Rinderer et al. 1989; Ruttner, Kauhausen, and Koeniger 1989; Tingek, Koeniger, and Koeniger 1996) and molecular analyses of mitochondrial and nuclear DNA (Arias et al. 1996; Arias and Sheppard 2005; Tanaka, Roubik, et al. 2001; Tanaka, Suka, et al. 2001) support recognition of A. koschevnikovi as a species distinct from all other cavity-nesting bees. However A. nuluensis, A. nigrocincta, and A. cerana are recently diverged, and their phylogenetic relationships are unresolved (Arias and Sheppard 2005). A. koschevnikovi (formerly A. vechti, Maa 1953) is reddish in overall appearance, with an amber gold labrum (Mathew and Mathew 1990). The body is 10–15 percent longer than that of sympatric A. cerana. The cubital index is large but highly variable (Tingek et al. 1988), and the extension of

Introduction to the Species

A. cerana A. koschevnikovi A. nigrocincta A. nuluensis

Figure 2.11 The distribution of the cavity-nesting bees A. cerana, A. koschevnikovi, A. nuluensis, and A. nigrocincta (Hepburn, Smith, et al. 2001; Otis 1996). A. cerana is a diverse species, and may have cryptic speciation (Hepburn, Smith, et al. 2001; see Chapter 3), especially in India. A. nuluensis is confined to the highlands of Borneo, and its existence is only known from the Crocker Range in Sabah. We have indicated a broader range based on elevation, but it is speculative beyond the Crocker Range.

the radial vein of the hindwing is also highly variable (0.08–0.30 mm). A. koschevnikovi males have a hairy fringe on the outer margin of the tibia and basitarsis of the hind leg (Mathew and Mathew 1990; Tingek et al. 1988). A. koschevnikovi workers are reluctant to ventilate their colony by fanning, but when they do so, they face outward from the colony. The species is prone to intraspecific robbing (Mathew and Mathew 1990). Apart from its mating time, almost nothing is known about the biology of A. nuluensis. The species is confined to mountainous regions above 1,500 m and has been reported only from the spectacular Mount Kinabalu, near Kota Kinabalu, which forms part of the Crocker Range in the Malaysian state of Sabah in Borneo. Whether it is present in all mountainous

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Introduction to the Species

Figure 2.12 Drone comb capping of A. nigrocincta (left) and A. cerana (right). Mature drone cells of A. cerana and A. koschevnikovi have a pore in their capping. Drone cells of A. nigrocincta do not (Hadisoesilo and Otis 1998). Note that recently capped drone cells do not contain the pore in any species.

areas of Borneo is not known. We have seen an A. nuluensis colony 2.3 km from the beginning of the summit trail of Mount Kinabalu National Park, and numerous foragers at the tree line at the Labon Rata mountain huts at 3,700 m, and at the Mesilau nature resort within the park. A third speciation within the cerana group appears to have occurred on the islands of Sulawesi and Sagihe (Damus and Otis 1997; Hadisoesilo and Otis 1996, 1998; Hadisoesilo, Otis, and Meixner 1995; Smith et al. 2003). There, in addition to isolated occurrences of A. cerana, a slightly larger, more yellow species is found, A. nigrocincta (Hadisoesilo, Otis, and Meixner 1995). The species status of A. nigrocincta is confirmed by mating flight times that diverge from those of A. cerana (Chapter 5; Hadisoesilo and Otis 1996), which suggests that A. cerana rarely mate with A. nigrocincta. The other species-specific trait is that A. nigrocincta lacks the pore in the capping of mature drone cells (Figure 2.12; Hadisoesilo and Otis 1998). In our experience this pore is present in all mature drone cells of A. cerana and A. koschevnikovi. To our knowledge, a brood comb of A. nuluensis is yet to be studied, so the presence of the pore in that species cannot be confirmed. Other evidence for the species status of A. nigrocincta comes from multivariate morphometrics (Damus and Otis 1997) and molecular analysis of a noncoding region of the mitochondria, which shows that A. nigrocincta has two mitochondrial haplotypes that have not yet been found in any other cavity-nesting species (Smith et al. 2003). The

Introduction to the Species

male genitalia, however, cannot be distinguished from those of A. cerana and A. nigrocincta (Hadisoesilo 1997), and the molecular phylogeny of Arias and Sheppard (2005) showed A. nigrocincta nested within A. cerana.

Summary There are three main groups (subgenera) of honey bees. The dwarf honey bees (Micrapis) build a single comb that surrounds a branch or twig. There are two species, the red dwarf honey bee (A. florea) and the black dwarf honey bee (A. andreniformis), both broadly distributed in tropical Asia. The giant honey bees (Megapis) build a single massive comb under a branch or cliff overhang. The slightly larger giant mountain honey bee, A. laboriosa, is confined to mountainous regions, whereas the closely related common giant honey bee, A. dorsata, is tropical. The cavity-nesting honey bees (Apis) have multiple comb nests, usually built in cavities. The main species are the western honey bee A. mellifera and the eastern honey bee A. cerana. The red honey bee, A. koschevnikovi, is distributed through rainforests of peninsular Malaysia and Borneo. The mountain honey bee, A. nuluensis, is reported only from the Crocker Range of Borneo. The Sulawesian honey bee, A. nigrocincta, is confined to the islands of Sulawesi, Sagihe, and Mindanao.

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3 Evolution

We begin this chapter with a discussion of the origins of bees in general. This starting point will demonstrate how ancient bees are and put the rather limited radiation of honey bees into phylogenetic perspective. We then discuss the much debated evolutionary history of the “corbiculate Apidae,” the four modern tribes of bees to which the honey bees belong. We end the chapter with a discussion of the radiation of the honey bees themselves, focusing in particular on the question of whether cavity nesting or open nesting is the ancestral condition of the honey bee tribe. This question is crucial to a proper understanding of the evolution of the dance language, a topic that we will address in Chapter 5.

The Beginning of Bees More than 200 million years ago, the present-day continents of Africa, Antarctica, Australia, South America, and the Indian subcontinent, were joined in a vast landmass known as Gondwana (Hallam 1994). About 130 million years ago, flowering plants, the angiosperms, first appeared in Gondwana (Crane, Friis, and Pederson 1995). Unlike the gymnosperms from which they diverged, the angiosperms did not rely solely on wind to move their male sex cells (pollen) from one flower to another. Rather, they exploited a more efficient vector: that of enticing animals, primarily insects, to visit their flowers, thus releasing plants from relying on the vagaries of the weather to transport their pollen. Presumably, exploiting di-

Evolution

rect transfer of pollen between flowers by insects allowed angiosperms to reduce the biological resources they devoted to pollen production, and helped these forerunners of modern plants to spread their pollen upwind as well as down. Among the insect visitors to these early angiosperm flowers were some short-tongued, hairy, spheciform (ground-burrowing) wasps (Engel 2001a), the adults of which had come to rely on eating pollen, and thus on moving from flower to flower to find it (Engel 2001a; Michener 2000). Many plants in their turn moved beyond offering a little extra pollen to feed their sex-cell vectors, by evolving nectar-producing organs that provided their pollinators with a source of carbohydrates. About 120 million years ago, this relationship became dependent (Engel 2001a), giving rise to the incipient bees that gave up all food sources other than those provided by flowering plants. The bees developed special morphologies, particularly increased hairiness, to more efficiently collect pollen (Michener 2000). So began the mutualistic relationship that now characterizes both bee and angiosperm life histories, in which bees provide pollination services and flowers provide enticements of food to attract them (Crane, Friis, and Pederson 1995; Grimaldi 1999). The bees, then, diverged as a monophyletic clade from the sphecid wasps in the early Cretaceous period 120–130 million years ago (Engel 2001a; Winston 1987). Some species have become specialists on plants of limited distribution or form, while others like the honey bees have become generalists, feeding from any plant species that has something to offer them. Today there are some 17,000 named species of bee, a little more than half of the estimated number of extant species (Michener 2000). The Gondwanan origin of bees ensured that bees, like most other insect families, enjoy a worldwide distribution through vicariance (Croizat 1982). Figure 3.1 shows the approximate phylogenetic relationships among the extant families of bees, and their estimated divergence times.

The Corbiculate Bees Around 90–100 million years ago, well before the break-up of Gondwana, a new way of packing and carrying pollen emerged among bees, and this

37

38

Evolution

ees of b n i g Ori

Cretaceous

Old

est f

o

ee ssil b

rn ode of m n i Orig y bees e hon

Tertiary

150 140 130 120 110 100 90 80 70 60 50 40 30 20 10

0

Colletidae Halictidae Andrenidae Melittidae Megachilidae

Apidae Corbiculate Apidae Figure 3.1 The phylogeny of the bee families and their approximate divergence times. The “corbiculate Apidae,” which include the honey bees, are part of the Apidae. After Engel 2001a; Grimaldi and Engel 2005.

led to the clade we now know as the corbiculate Apidae, which are part of the subfamily Apinae (Michener 2000). Corbiculate bees are long-tongued bees that have pollen baskets, or corbiculae. A corbicula is a patch of stout bristles surrounding a bare depressed area on an expanded hind tibia onto which the bee packs pollen, using a little nectar to make it stick. When full, the baskets take on a characteristic tear-drop shape, often brightly colored by the pollen. Today there are four tribes of corbiculate bees, the orchid bees (Euglossini),the stingless bees (Meliponini), the bumble bees (Bombini), and

Evolution

A

Bombini

Meliponini

B

Apini

Meliponini

Apini

Euglossini

Bombini

Euglossini

Figure 3.2 Two alternative phylogenies of the corbiculate bees. Phylogenies are based on DNA sequences (A) or morphology and behavior (B). The contrast between A and B is whether the bumble bees group with stingless bees (A) or with the orchid bees (B). The root of tree B is at the dot. The root of tree A is uncertain, but occurs at or to the right of the dot.

the honey bees (Apini) (Figure 3.2). In Table 3.1, and below, we present a comparative survey of the four tribes.

Orchid Bees Orchid bees (Euglossini) are large bees that build resinous nests, often underground. Their distribution is confined to South and Central America (Michener 2000). Though some species nest communally, they are generally solitary with individual females raising their own brood in separate nests (Dressler 1982; Michener 1974, 2000). Rarely, if ever, do daughters remain in the nest to assist the mother in rearing the next generation of brood. Intriguingly, male orchid bees forage on orchid flowers and

39

40 Table 3.1

Evolution Characteristics of the corbiculate bees. Present in all

Characteristic Vertical combs of hexagonal cells built of wax secreted by workers Multiple use and reuse of comb cells for both brood rearing and food storage Progressive provisioning of brood with food comprised predominantly of glandular secretions Extreme polyandry and worker policing Symbolic dance language for communicating location of food Nest defense via barbed sting Water collection Strong queen/worker caste dimorphism and reproductive division of labor Colony founding via reproductive swarming Food transfer via trophallaxis and food storage Collection of plant resin (propolis) Nest thermoregulation Pollen carried on corbiculae

Euglossini, Bombini, Meliponini, Apini

Bombini, Meliponini, Apini

Meliponini, Apini

Apini X X X

X X

X

X X X

X

X X X

X X

X X

X X X

X X X

some other sources of oily scents, diligently collecting various aromatic chemicals much like pollen-foraging females, and storing them in a storage chamber on the hind tibia (Robinson 1984). It is unclear what they do with the chemicals, but they may be used to attract mates or other males to leks (Dressler 1982).

Bumble Bees (Bombini) Bumble bees have a huge range throughout the Americas and through much of Asia (at high altitudes), Europe, and northernmost but not southern Africa (Goulson 2003). The absence of native bumble bees in Australia

Evolution

and sub-Saharan Africa may indicate that the Bombini arose after the break-up of Gondwana (Michener 2000). Bumble bees are highly social, but with significant differences from stingless bees and honey bees. Foremost in this regard is that almost all bumble bee species have annual rather than perennial colonies: nests are founded by a single mated female who rears her first generation of brood unaided by workers (Alford 1975). This means that bumble bee queens do not show the same degree of caste dimorphism as do queens of the stingless bees and honey bees, since queens must forage to provide for their first batch of brood, and therefore possess a long tongue and pollen baskets. The primary differences between queens and workers are the presence of a spermatheca in queens, their greater size, and their ability to enter diapause, for which they lay down large fat reserves. Although workers cannot mate, they can lay haploid male-producing eggs. Because of this potential, reproductive division of labor is not complete in all species. In Bombus hypnorum, workers produce over 20 percent of the males (Brown, Schmid-Hempel, and Schmid-Hempel 2003). In other species, many workers have activated ovaries, though their actual reproductive success can be low because worker oviposition is delayed until the end of the season, when brood rearing is disrupted, and because in temperate species, opportunities for mating by the late-season worker-laid males are limited (Cnaani, Schmid-Hempel, and Schmidt 2002). Bumble bees regulate the temperature of their nests to a considerable degree (Goulson 2003; O’Donnell and Foster 2001), and produce wax to build brood cells and nectar pots. In at least one species, B. terrestris, a successful forager can alert her nestmates to the existence of a profitable patch of flowers by excitedly running around the nest, but cannot provide directional or distance information about it (Dornhaus and Chittka 1999, 2001).

Stingless Bees Stingless bees (Meliponini) live in societies that are quite similar to those of honey bees. Their colonies comprise 500–100,000 individuals, usually domiciled in natural cavities. In most species queens are far larger than workers, and in all species queens lack corbiculae. Reproductive division

41

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Evolution

of labor is strict in most species (Green and Oldroyd 2002; Palmer et al. 2002; Tóth, Queller, Imperatriz-Fonseca, and Strassmann 2002; Tóth, Strassmann, Imperatriz-Fonseca, and Queller 2003), but workers produce male-destined eggs in others (Beig 1972; Drumond, Oldroyd, and Osborne 2000; Imperatriz-Fonseca and Kleinert 1998; Sommeijer, Chinh, and Meeuwsen 1999). One of the more fascinating behaviors of the stingless bees is the strange “rituals” that accompany oviposition. These have been interpreted as stylized aggressive interactions between the queen and her workers, in which the queen asserts her reproductive dominance over the workers (Drumond et al. 1999; Sakagami 1982; Tóth et al. 2004; Tóth, Strassmann, Nogueira-Neto, Imperatriz-Fonseca, and Queller 2002). The process of oviposition has three recognizable components, and the variants of these components have been analyzed phylogenetically (Drumond, Zucchi, and Oldroyd 2000; Zucchi et al. 1999). First, workers construct one or a small number of brood cells. Second, the cells are mass provisioned with brood food consisting of regurgitated crop contents. Third, the queen lays in the cell(s), whereupon the workers immediately seal them. The brood develops within the capped cell(s) with no further feeding involved. Stingless bees have some capacity to thermoregulate their nests. They secrete wax for building brood combs, honey pots, and the “batumen” layer, a membrane of wax that they use to insulate and protect their entire nest, and wall off unused areas of the cavity that they occupy. They have variable ability to communicate the location of food sources via scent marking, dancing, and acoustic signals (Nieh 1998, 2004) and to allocate foragers to the most profitable feeding sites (Biesmeijer and Ermers 1999; Nieh 2004). They establish new nests via fission in which a portion of the workers and a young queen move from the old nest to a new one over several weeks. Stingless bees have a generally tropical distribution in Africa, Asia, Australia, and South and Central America. One or two species extend into subtropical regions in Australia and Brazil.

Honey Bees Honey bees (Apini) have a natural distribution in Asia and Europe and one species, Apis mellifera, has radiated into Africa. The major behavioral differences between the honey bees and the other corbiculate bees are extreme multiple mating by queens (Palmer and Oldroyd 2000), the sym-

Evolution

bolic dance language for communication among workers about location of food (von Frisch 1967) and nest sites (Lindauer 1955), precise nest thermoregulation (Jones et al. 2004), reuse of brood cells for brood rearing or honey storage, and reproduction via swarms that make a clean break from the parental nest, rather than via slow fission as in the stingless bees. In queenright nests there is functional worker sterility (Barron, Oldroyd, and Ratnieks 2001). In the honey bees, larvae develop in open cells, and the nurse workers determine the amount of food they are fed. The workers can therefore dictate whether the developing female becomes a worker or a queen. In contrast, the mass provisioning of brood cells by stingless bees means that developing stingless bee females can have at least some power over whether they will develop as a small but hopeful queen or as a worker (Bourke and Ratnieks 1999; Ratnieks 2001; Wenseleers, Ratnieks, and Billen 2003). In one genus, the Melipona, workers and queens are of identical size and are reared in identical brood cells. Thus because the adult workers cannot interfere with the developing brood, a female larva has almost complete power over whether she will develop as a queen or a worker. This leads to a “tragedy of the commons,” in which queens are massively overproduced, which necessitates mass slaughter of excess young queens and a huge waste of colony resources (Wenseleers and Ratnieks 2003).

Relationships among the Corbiculate Bees Since there are four tribes of corbiculate bees, there are 15 possible rooted trees that can be drawn among them (Li and Graur 1991). At least 9 of these possible trees have been suggested as the correct interpretation of corbiculate bee evolution over the last 20 years (Ascher, Danforth, and Ji 2001; Thompson and Oldroyd 2004), which illustrates the extreme confusion currently surrounding corbiculate bee phylogeny. Despite this confusion, alternative phylogenetic hypotheses tend to fall into one of two groups; those that support a close relationship between honey bees and stingless bees and those that support a close relationship between bumble bees and stingless bees. The first (A in Figure 3.2) repeatedly emerges from molecular data (Cameron 1993; Cameron and Mardulyn 2001; Koulianos et al. 1999; Lockhart and Cameron 2001; Sheppard and McPheron 1991)

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Evolution

and at least one morphological analysis (Winston and Michener 1977). It groups the stingless bees with the bumble bees in one clade, but does not define the relationship between the honey bees and the orchid bees. The second (B in Figure 3.2) repeatedly emerges from analysis of morphological (Engel 2000a, 2001b; Michener 1974; Prentice 1991; Roig-Alsina and Michener 1993) and behavioral characters (Noll 2002), and some combined morphological and molecular data sets (Ascher, Danforth, and Ji 2001). It groups the eusocial honey bees and stingless bees together in one clade, and the bumble bees and presocial orchid bees in another. The topologies in Figure 3.2 cannot both be correct, but obtaining the correct phylogeny is essential to our understanding of the evolution of corbiculate bees in general and Asian honey bees in particular. In order to understand evolution within the Asian bees we need to know which is the most basal species and which is the most derived. And the only way to do this is to correctly determine the sister clade to Apis—the stingless bees or a bumble bee–orchid bee clade. So which phylogeny in Figure 3.2 is more likely to be correct?

Resolving the Phylogeny of Corbiculate Bees Construction of a phylogeny from a morphological or molecular data set requires different assumptions. We briefly review these assumptions in order to show that neither approach is inherently superior. Readers seeking more details of methods of phylogeny reconstruction are referred to Hillis et al. (1996), Hall (2004), or Felsenstein (2004). Molecular analyses have the virtue of being based on unambiguously homologous characters: a particular gene in one tribe is the same gene in another tribe. Although duplications and pseudogenes can sometimes lead to paralogous genes being analyzed in the different lineages (Moritz and Hillis 1996), this is unlikely with the well-characterized genes that have been used in the bee phylogenies. Generally speaking, individual character states are unambiguous and easy to define, as in “base position 239 is a T” (Li and Graur 1991; Swofford et al. 1996). But even here, there can be ambiguity, because molecules vary in length and must be aligned by inserting gaps in the sequences so that base position 239 really is homologous. Where genes are widely diverged the alignment can be difficult.

Evolution

Building molecular phylogenies requires assumptions about the way in which DNA molecules evolve, and the data itself can hold limited information, because in deeply diverged lineages like the tribes of corbiculate bees, repeated base substitutions at the same site can lead to loss of phylogenetic signal at that site (Li and Graur 1991). Thus differences or similarities between molecules at a particular site may have no biological meaning. The main way to avoid this problem when one is analyzing a deeply diverged phylogeny is to use a slowly evolving gene. To this end, Mardulyn and Cameron (1999) have used the eye pigment opsin, and Cameron and Mardulyn (2001) the large subunit of the ribosome. These are critical genes and might therefore be expected to evolve slowly. Second, DNA molecules may evolve at different rates and in different ways in different lineages, at different codon positions and even in different parts of genes (Griffiths 1997), which leads to erroneous conclusions (Jermiin et al. 1996; Nei 1996). But if the differences and biases among lineages are properly understood, then maximum-likelihood model-based approaches to tree building can largely compensate for them (Huelsenbeck and Crandell 1997). Even then, when trees are characterized by long branches among clades and short branches among taxa within clades, maximum likelihood procedures can return the wrong trees by a phenomenon called “long branch attraction,” an emergent property of the algorithms that are used to search for the best trees, in which taxa that are strongly diverged from the rest tend to get grouped together (Swofford et al. 1996). Morphological analyses require their own but different set of assumptions. Most important, the person undertaking the analysis must make judgments about the characters he or she will use for the analysis. First, characters that vary together because they are part of the same structure should be avoided. Second, characters must be measured on homologous structures across the taxa considered. This can sometimes be difficult because unrelated structures can appear similar in distant taxa owing to convergent evolution. Ambiguities about homology arising from convergent evolution are particularly important in resolving the corbiculate bee phylogeny because the stingless bees and the honey bees are highly eusocial whereas the orchid bees are solitary. Thus we would expect that honey bees and stingless bees might share characters related to social life via convergent evolution, even if they do not share evolutionary history.

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Evolution

As yet we have no satisfactory resolution of the two alternative phylogenies given in Figure 3.2. Both phylogenies are simultaneously incompatible, but consistent with their respective data sets.

Corroborating Evidence Additional support for phylogeny A comes from a reanalysis of all possible topologies using the four available molecular data sets (Thompson and Oldroyd 2004). This analysis showed that in fact only one data set (that of cytochrome b, cyt b) could reject any of the 15 possible trees that can be constructed to show relationships between the four tribes. Data from the other genes are useless for the phylogenetic analysis of the corbiculate bees. Nonetheless, cyt b statistically rejects all but 3 topologies and all of these support a close relationship between the tribes Bombini and Meliponini, that is, phylogeny A (Figure 3.2). Independent support for phylogeny B (Figure 3.2) comes from the fossil record. Engel (2001a, 2001b) examined 152 bee specimens from Baltic amber that are estimated to be about 45 million years old. He found a plethora of apparently advanced social bees (believed to be such because the reduced abdomen suggests a nonreproductive worker) encompassing eight extinct genera that morphologically fall between the honey bees and the stingless bees. Moreover, there are reports of compression fossils from the same amber-containing deposits that are clearly honey bees (Engel 1998; Nel et al. 1999) and some that are clearly stingless bees (Engel 2001a). Thus the fossil record suggests that the stingless bees are the sister group to the honey bees, with clear intermediate forms between the two. Phylogeny B is also more consistent with biogeography than phylogeny A. The orchid bees are confined to South and Central America (Michener 2000) and presumably evolved there, whereas the honey bees evolved in Eurasia (Ruttner 1988). Both the stingless bees and bumble bees are broadly distributed. Thus phylogeny B is consistent with bumble bees and orchid bees diverging from a common ancestor in the Americas and stingless bees and honey bees diverging in Eurasia, followed by range expansion by both bumble bees and stingless bees. In contrast, we find phylogeny A impossible to reconcile with biogeography unless we posit that the honey bees and orchid bees diverged before the break-up of Gondwana, and then

Evolution

went extinct in the regions where they are now absent. This seems an unlikely (though possible) scenario. To obtain a proper understanding of the evolution of the honey bees, the correct phylogeny of the corbiculate bees is required. In order for this to happen it will be necessary for the molecular, morphological, and paleontological-based hypotheses for the evolution of the corbiculate bees to be brought into accord. But in the absence of such agreement it is necessary to make an assessment of probabilities. Since independent data from biogeography and paleontology suggest that phylogeny B, based on morphology (Figure 3.2), is more likely than phylogeny A, based on similarities of cyt b DNA sequences, we tend to favor phylogeny B of Figure 3.2. Yet given the power of molecular phylogenetics, we consider it imprudent to dismiss phylogeny A out of hand.

Evolution of the Honey Bees Under hypothesis A the honey bees are distantly related to the stingless bees. For this to be true either two independent origins of eusociality are required or, alternatively, the common ancestor of the corbiculate bees was eusocial, and there has been a reversal to “primitive” eusociality in the bumble bees and solitary life history in the orchid bees. Eusociality has evolved independently in termites, wasps, bees, ants, beetles, thrips, aphids, spiders, scorpions, and naked mole rats (Crespi and Choe 1997), so we do not regard two origins of eusociality as being particularly unlikely in the corbiculate bees. But reversals from eusocial forms to solitary forms are extremely unlikely (Cameron and Mardulyn 2001; Thompson and Oldroyd 2004). We conclude therefore that under the phylogeny supported by cyt b, the honey bees arose from a solitary ancestor related to the orchid bees. The time and place of this divergence is unclear. Phylogeny B, supported by morphology, behavior, and biogeography, suggests that the two highly eusocial tribes, the stingless bees and the honey bees, are sister taxa. This suggests that the ancestor of the honey bees was eusocial, and shared many morphological and behavioral features with extant honey bees and stingless bees. The common ancestor of the bumble bees, stingless bees, and honey bees would have been “primi-

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tively” eusocial (Michener 1974; Wilson 1971), with the nest founder dominating reproduction, but lacking the strongly divergent queen and worker castes. The earliest known fossil bee is from Mesozoic amber sampled from New Jersey (Michener and Grimaldi 1988). Originally dated as 80 million years old (Michener and Grimaldi 1988), Engel (2000b) suggests that the fossil is slightly more recent, 65–70 million years old. This female bee had an extremely small abdomen, strongly suggesting that it could not lay eggs, and was therefore a worker of a eusocial species (Michener and Grimaldi 1988). It is similar morphologically to workers of the modern African stingless bee genus Dactylurina (Engel 2000b). If Engel’s inference is correct, eusocial behavior emerged early in the corbiculate bees. Because of the ambiguous evolutionary history of the stingless bees and the weak fossil record beyond 50 million years, the time when a creature that we would recognize as being a honey bee first appeared is uncertain. If the honey bees arose from a common ancestor of the orchid bees or from the orchid bees, we have no way of dating that divergence other than by DNA divergence. If the honey bees diverged from a common ancestor of the stingless bees, then honey bees must be relatively ancient, more than 70 million years old, because stingless bees apparently date to that time (Engel 2000b). Oligocene Europe was subtropical, and the biota was Indo-Malayan (Hallam 1994). Global cooling at the middle of the Miocene saw extinction of the ancient honey bee and stingless bee species in temperate regions; they persisted only in tropical Asia. The extinction of the ancient European honey bees indicates that they were open-nesting, single combed species, for if they had developed cavity nesting and multiple combs, they should have survived the Miocene cooling in Mediterranean refugia (Ruttner 1988). Engel (1999) suggests that the modern open-nesting species first appeared in southeast Asia 6–10 million years ago, presumably a remnant of a more broadly distributed Eurasian species, possibly A. armbrusteri (Armbruster 1938). Cavity nesting emerged slightly later, possibly in the Himalayan region, where there were a diversity of biotypes and climates, which may have facilitated evolutionary change (Ruttner 1988). Cavity nesting and multiple combs allow for precise thermoregulation with less energetic cost, so the new cavity-nesting species could colonize temperate

Evolution

as well as tropical regions. This ancestral cavity-nesting species spread throughout tropical and temperate areas of Asia. This cavity-nesting species eventually radiated into temperate regions of Europe and Asia. The European lineage became isolated from the Asian lineage by the expansion of desert areas of the Middle East. In isolation it evolved into modern A. mellifera, which is now distributed throughout most of Europe, Scandinavia, and Africa. The Asian lineage gave rise to A. cerana and its related species. The time of divergence of A. cerana and A. mellifera is unclear, but it may have been soon after the emergence of cavity nesting. The ND2 gene of the mitochondria shows 17–19 percent divergence between A. cerana and A. mellifera, which suggests (based on 2 percent divergence per million years) a divergence time of 3 million years ago (Arias and Sheppard 1996, 2005). We will return to a discussion of how the species of the Asian cavity-nesting bees evolved in Chapter 4.

Phylogeny of the Honey Bees As with the tribal-level phylogeny, the phylogeny of the honey bees has been controversial with considerable divergence of the topologies derived from molecular data sets (Willis, Winston, and Honda 1992) and those inferred from behavioral and morphological data (Alexander 1991a, 1991b; Koeniger 1976). Recently, however, trees derived from molecular, behavioral, and morphological data have been brought into agreement, and a phylogeny similar to that shown in Figure 3.3 is broadly accepted (Engel and Schultz 1997; Raffiudin and Crozier 2000, Arias and Sheppard 2005). This phylogeny clusters the closely related Asian cavity-nesting species (A. cerana, A. nigrocincta, and A. nuluensis) in a shrubby tip. Outside of this clade are A. koschevnikovi and the European A. mellifera. Outside of these are the two giant species A. dorsata and A. laboriosa. The most basal clade comprises the dwarf open-nesting species A. florea and A. andreniformis.

Cavity-Nesting Lost or Gained? Despite our strong inclination toward the phylogeny of Figure 3.3, we invite the reader to give due consideration to N. Koeniger’s alternative view

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A. florea A. andreniformis A. dorsata A. laboriosa A. mellifera A. koschevnikovi A. nuluensis A. nigrocincta A. cerana Figure 3.3 A phylogeny of the genus Apis. A complete phylogeny of all species of honey bee is not yet available. The figure shows the most likely topology in the authors’ opinion. The general topology is based on Arias and Sheppard (2005) and Engel and Schultz (1997). Although A. cerana, A. nigrocincta, and A. nuluensis are good species according to our criteria, their relative positions in the Apis phylogeny are not well resolved (Arias and Sheppard 2005; Smith et al. 2003; Tanaka, Roubik, et al. 2001).

(1976) of the evolution of Apis. He argues that the basal honey bee species are the cavity-nesting clade and that the open-nesting clades are derived, that the root of the Apis phylogeny should be reversed from that of Figure 3.3. In telling support of this argument he points to the fact that the stingless bees, orchid bees, and bumble bees—the three outgroups of Apis—are cavity-nesting, and that cavities are a limiting resource (Inoue, Adri, and Salmah 1990). Thus there is strong selective pressure for tropical bees to move out of cavities, but (Koeniger argues) little incentive for them to move into them. Given the strong support for the phylogeny of Figure 3.3 we cannot agree that the cavity-nesting clade is basal, for to do so would give too strong a weight to the character of cavity nesting in the construction of the Apis phylogeny: the weight of all other characters points to the phylogeny of Figure 3.3. Awkwardly, however, if the phylogeny of Figure 3.3 is correct, the basal honey bees all lived in the open whereas the three outgroups all

Evolution

live in cavities. This gives us one of two equally (un)likely evolutionary scenarios. Either the ancestor of honey bees nested in cavities and this character was lost twice in the lineages leading to the florea-andreniformis and the dorsata-laboriosa branches, or the ancestor lived in the open, which would require that cavity nesting to be regained in the lineage leading to the cerana-mellifera clade. Like most other authors (Alexander 1991b; Engel and Schultz 1997; Ruttner 1988), we support the latter hypothesis: that the cavity nesting is the derived condition for Apis and that the ancestral Apis was open-nesting. Tellingly, even temperate A. mellifera occasionally nests in the open or in highly exposed cavities like caves, which suggests that it can revert to its ancestral condition. The extinction of the earliest European honey bees in the Miocene cooling gives further support to the notion that the ancestral state of honey bees was open-nesting (Ruttner 1988). We also suggest, contrary to Koeniger, that there are considerable advantages to cavity nesting even for tropical bees, including improved nest defense and thermoregulation.

Summary Bees evolved about 130 million years ago from spheciform wasps. The corbiculate bees emerged about 90–100 million years ago, and then diverged, leaving four extant tribes, the orchid bees, the bumble bees, the stingless bees, and the honey bees. The phylogeny of the corbiculate bees is controversial, with strongly different topologies emerging from molecular and morphological-behavioral data sets. The ancestor of the honey bees probably emerged about 35–40 million years ago in the Indo-European region. Oligocene-Miocene cooling caused honey bees to go extinct in Europe, but they persisted and speciated in tropical Asia. The ancestor of modern open-nesting Apis probably emerged about 10 million years ago. Slightly later, possibly 6 million years ago, an A. cerana–like bee that could efficiently thermoregulate owing to cavity nesting and multiple combs emerged, possibly in the Himalayan region. It spread east and north, becoming ubiquitous throughout Asia. About 2–3 million years ago, during a Pleistocene warming, this bee expanded west into Europe and thence into Africa to become A. mellifera.

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4 Speciation and Biogeography

In Chapter 3 we suggested that the common ancestor of the modern open-nesting honey bee species emerged about 10 million years ago, and the cavity-nesting species slightly more recently. In this chapter we consider the causes of more recent speciation events: the separation of the dwarf bees into A. andreniformis and A. florea, the giant bees into the lowland tropical form A. dorsata and the mountain-dwelling cool-climate species A. laboriosa, and the divergence of the Asian cavity-nesting bees into A. cerana, A. nigrocincta, A. nuluensis, and A. koschevnikovi. We also discuss some of the details of within-species biogeographic diversity.

The Causes of Speciation Events The means by which new species arise is one of the most discussed and modeled issues of evolutionary biology (see, e.g., Turelli, Barton, and Coyne 2001). Undoubtedly there are a variety of mechanisms that can lead to sufficient genetic divergence between two populations for biologists to regard the two populations as separate species. The simplest model of speciation, allopatric speciation, involves nothing more complicated than long-term separation of gene pools via physical partitioning of populations by a geographic barrier such as a mountain range, sea, or region of hostile habitat (Mayr 1942, 1963). With enough time, genetic drift and/or natural selection toward locally adapted types can cause the separated populations to become so diverged that hybrids between the two are nonviable, or the two incipient species simply cannot recognize each other as potential mating

Speciation and Biogeography

partners. The speed with which two disparate populations diverge into different species can be enhanced by a variety of mechanisms, some of which appear to have accelerated the recent radiations within the Asian honey bees. We will consider these accelerators of speciation below, but begin with a discussion of how changes in sea levels over the last 15,000 years led to the breaking up of a contiguous land mass into a series of islands, now known as the Malay Archipelago. This change in geography caused multiple instances of isolated honey bee populations, creating possibilities for speciation events. The creation of islands has also greatly complicated the current distribution of Asian honey bee species.

The Biogeography of Honey Bees in Asia The most recent period of glaciation began about 2 million years ago, and reached a peak 18,000 years ago, when much more of the world’s water was sequestered in polar ice and in glaciers than it is today (Heaney 1991). An important effect of this cold period was to reduce the world’s oceans by as much as 200 m below current levels, thus exposing land bridges between many of the current islands of the South China Sea (Figure 4.1). Notable features of the mid-Pleistocene map of Asia were the connections of the present-day islands of Japan, Taiwan, Borneo, Sumatra, Java, Timor, and Palawan to continental Asia. These bridges allowed honey bees of all three groups (dwarf, giant, and cavity-nesting) to colonize what are now islands. Even where land bridges were incomplete, such as between Borneo and Sulawesi, Luzon and Palawan, the Andaman Islands and Myanmar, and Mindanao and Borneo, distances between islands were much reduced relative to today’s distances, and this seems to have allowed honey bees to colonize some regions even where there was no direct land link. How they did this is unknown. The maximum distance an Apis swarm will travel over sea is unclear, but instances of migratory swarms crossing stretches of water exceeding 10 km have been reported (e.g., Hannabus 1939; Roubik 1989; Roubik 1990; Thornton and New 1988). This may actually be a low estimate, because theoretical calculations suggest that fully laden honey bees (those with a honey stomach full of food) can cover distances of about 100 km (Otis, Winston, and Taylor 1981; Seeley 1985). Furthermore, it is conceivable that a swarm or colony could colonize a new area by floating

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Speciation and Biogeography Merrill Line

Wallace Line

Sundaland

Figure 4.1 Approximate landforms of southeast Asia 18,000 years ago. The great land mass known as Sundaland encompassed the Malay Archipeligo, Borneo, and Palawan, and was joined to the Asian mainland. Sunderland was never joined to Sulawesi (Indonesia) or to Mindoro or Luzon (Philippines). These ancient straits led to considerable geographical separation of biota, now recognized as the Wallace and Merrill Lines, respectively.

across the sea on a log. There are frequent reports of A. cerana and A. mellifera swarms hitchhiking into Australian ports on a ship. In addition to natural colonization, migrations in the last 40,000 years may have been assisted by humans, and we suspect that the ubiquity of A. cerana across the South China Sea islands may owe much to human intervention. Around 10,000 years ago, the world was warming again, and the oceans began to refill with the water from the retreating icecaps, creating new islands and isolating the biota thereon (Heaney 1986, 1991). Where island populations were reduced to a small number of individuals, so that allelic diversity was heavily bottlenecked, or where the environment of the isolated group selected for a particular type, conditions that are thought to be conducive to the development of new species were present. On the other hand, small, isolated populations are vulnerable to extinction either by catastrophic annihilation via natural disaster, demographic failure (the inability to find mates), or the effects of inbreeding. Furthermore, isolated populations were probably reunited with the more cosmopolitan group, from which they had diverged from time to time, which would lead to extinction

Speciation and Biogeography

of incipient species via competition or hybridization. Thus many islands probably suffered species loss as well as having the potential for creating new ones. Given the multiple opportunities for allopatric honey bee speciation in the Malay Archipelago, it is perhaps a little surprising that there are not more honey bee species—and perhaps there are some that remain to be discovered. At present, however, there appears to be evidence for only a single allopatric speciation event caused by isolation on an island: the divergence of A. nigrocincta from an ancestral cerana type on Sulawesi and neighboring islands. All (or nearly all) other speciation events of the extant honey bees have apparently been independent of the late Pleistocene island formation and almost certainly predate it. Speciation of the dwarf bees seems to have predated island formation because both A. florea and A. andreniformis are present on continental Asia and on Java (Otis 1996, fig. 2.5). Current distributions may indicate that the precursor dwarf species had a continuous range from Sundaland to India. We speculate that at the height of early Pleistocene cooling, the population was divided by a region of inhospitable habitat in the area that is now Assam India and the northern part of Myanmar, and that this gave rise to A. florea in the west (presumably in India) and A. andreniformis in the east. Subsequently, with late Pleistocene warming, A. florea bees extended their range east into southeast Asia, colonizing Java and Sumatra before they were cut off by rising sea levels, and thus never making it to Borneo, the Philippines, or Sulawesi. Similarly A. andreniformis spread from Sundaland to colonize continental Asia as far west as Assam. Speciation of the giant bees also appears to be independent of island formation. Apis laboriosa is apparently the result of selection for a mountain ecotype that could have become separated from A. dorsata in some area of the Himalayas. It appears to have spread from its place of origin through the continuous mountain habitats of Asia as far east as northern Vietnam (Trung, Dung, and Ngan 1996). The events that led to speciation in the cavity-nesting bees are more complex and even more speculative. Because A. koschevnikovi is now present on Borneo, Java, Sumatra, mainland Malaysia, and southern Thailand (Figure 2.11), we must infer that this species diverged from the ancestral cavity-nesting species before island formation. The trigger of this diver-

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gence is difficult to explain. Potentially a temporary but prolonged isolation in a hot, wet, forested habitat led to speciation of A. koschevnikovi, which subsequently spread out to occupy what is now Borneo, Java, Sumatra, and the Malay peninsula. Owing to the great depth of the Cotabato-Sagihe trench, Sulawesi has never been joined either to Borneo to the west or Mindanao to the north (Hamilton 1979); this created the famous Wallace Line, which roughly divides the Eurasian biota to the west and the Australian (Gondwanan) biota to the east (George 1981). Apis nigrocincta probably evolved in isolation on the island following a rare colonization event by a tiny founder population of A. cerana. The modern A. cerana population on Sulawesi is probably derived from a relatively recent introduction, probably assisted by humans, from the islands to the north. The hypothesis that A. nigrocincta is indeed the result of a relatively recent allopatric speciation event (Arias and Sheppard 2005) could potentially be evaluated by tests for a genetic bottleneck based on microsatellite loci (Cornuet and Luikart 1996). A. nuluensis is confined to the mountains of Borneo (Fuchs, Koeniger, and Tingek 1996; Tingek, Koeniger, and Koeniger 1996). For most, if not all, of its range, the species lives in isolation from A. koschevnikovi and A. cerana, presumably because A. koschevnikovi and the ecotype of A. cerana present in Borneo is not competitive in the cooler climate of high altitudes. (A. cerana lives at similar altitudes in Nepal.) We can think of several alternative histories of the cavity-nesting bees on Borneo. First, A. nuluensis may have arisen from a temporarily isolated ancestral cerana population (Tanaka, Roubik, et al. 2001). Subsequently, A. cerana and A. koschevnikovi recolonized the lowland areas from other parts of Sundaland. Again, human assistance, possibly by Chinese traders, cannot be ruled out, but natural recolonizations are not unlikely. Second, A. nuluensis may have arisen de novo in the unique highland environment of the Crocker Range, from an A. cerana ancestor. This area, arising from fragments of Gondwanaland, has remnants of cool-climate Gondwanan vegetation that differ markedly from the surrounding tropical low-land forests (Audley-Charles 1987). Possibly, this unique area allowed the development of a new species via selection for a mountain ecotype. Finally, and least likely, A. nuluensis may be a remnant of a cooler period and potentially be found in other parts of Sundaland where the elevation is sufficient for its needs.

Speciation and Biogeography

How Honey Bee Mating Biology Reinforces Speciation We have seen how the changing geography of southeast Asia probably created multiple opportunities for the development of new honey bee species, but have also noted that most honey bee speciation events appear to have been independent of island formation. We will now consider mechanisms that have probably reinforced biological separation of honey bee species, wherever they have occurred, without the need for strong physical barriers to interbreeding. These mechanisms involve idiosyncrasies of the honey bee mating system, which appear to have constituted a potent force for honey bee speciation. Honey bees mate on the wing in well-defined locations known as drone congregation areas, or DCAs (Koeniger and Koeniger 2000b; Koeniger et al. 1994; Loper, Wolf, and Taylor 1987; Pechhacker 1994; Ruttner and Ruttner 1966). When a queen is 3–8 days old she may take an orientation flight to learn the location of her nest, or she may undertake the first of 1–5 mating flights in which she visits a nearby DCA and mates with several drones (Koeniger, Koeniger, and Tingek 1994a,b; Koeniger, Koeniger, and Wongsiri 1989; Ruttner, Woyke, and Koeniger 1972; Tan et al. 1999; Tarpy and Page 2000). Drones from many colonies join these aggregations (Baudry et al. 1998), which often comprise thousands of individuals (Loper, Wolf and Taylor 1987). DCAs are perennial and one at number 2 oval at Sydney University has persisted for at least 25 years. Of importance for our discussion of reinforcement of speciation, the location of a DCA is self-generating. Drones do not attract each other to the DCA via pheromones, and the presence of a queen is not required to form a DCA. DCAs are reformed after a seasonal absence of some months in the precise location of the previous season. Thus the location of a DCA is apparently dictated by certain physiographic characteristics of the landscape that attract the males (Loper, Wolf, and Taylor 1987, 1992; Pechhacker 1994). Males of each species are attracted by different cues, so the precise location of the DCA is a self-organized outcome of an interaction of genotype and environment (Koeniger and Koeniger 2000b). Selection for the preferred mating time and place can act as a powerful force for generating a mating barrier between two incipient species that

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find themselves in different environments. For example, the preferred mating place and time may be dictated by the need to avoid a frequent predator or simply by the need to utilize the optimal temperature and light environment for mating. Once selection has shifted a local population toward a particular time and place of mating, selection should then act to improve mating success within that environment (Boughman 2002). This could occur if one or both sexes became conspicuous within that environment (Endler 1987, 1992), so that they can easily attract mates, or the kinds of pheromones or calls made could be optimized for effective transmission in the preferred mating place (reviewed in Boughman 2002). Thus the A. dorsata’s habit of mating just after dusk may have been initially driven by predator avoidance. Given this, adaptations that assist males in first finding a DCA and then finding a queen would be strongly selected. Perhaps not surprisingly then, A. dorsata DCAs form under the canopy of tall trees that emerge significantly above the surrounding canopy of shorter trees (Koeniger et al. 1994). Presumably these are the easiest places to find after dark, and are safe from predators such as bats. In contrast, the closely related A. laboriosa probably mates in the midafternoon but at an unknown location (Chapter 2; Underwood 1990b). This makes sense if, in the absence of any predators, selection has shifted the mating time of A. laboriosa to the warmest part of the day—necessary in its mountain habitat. But in any areas of sympatry, these species would fail to mate, which would enforce reproductive isolation and facilitate further divergence. We conclude, then, that selection for the optimal time and place for mating probably assisted reproductive isolation of incipient honey bee species, keeping them separate when reunited with the species from which they had diverged. We will return to the evolution of mating biology in Chapter 6.

Intraspecific Biodiversity Although island biogeography appears to have played only a small role in the evolution of the honey bee species, it has been important for the generation of within-species diversity. In contrast to the Western honey bee A. mellifera, where hundreds of papers and at least two books (Hepburn and Radloff 1998; Ruttner 1988) have been written on A. mellifera bio-

Speciation and Biogeography

diversity and ways of measuring it, the study of within-species diversity of the Asian honey bees is in its infancy. Still, recent efforts, particularly by Deborah Smith and her colleagues utilizing phylogeographic approaches to the study of mitochondrial DNA sequences of Asian honey bees, are now providing important insights into how various ecotypes are currently distributed. The taxonomy of A. mellifera is mired by the recognition of a plethora of poorly defined subspecies. Fortunately, the Asian species have for the most part been spared from the confusing, and often dubious, nomenclatures that have blighted the taxonomy of A. mellifera—there are over 150 subspecies names (Engel 1999), with new ones still being added (e.g., Sheppard et al. 1997). Although it is undoubtedly true that cosmopolitan species such as some of the honey bees do evolve widely differing ecotypes, both in terms of behavior and appearance (Hepburn, Smith, et al. 2001), it is doubtful that naming them as subspecies is useful (Oldroyd 1993). In this book we shall refrain from contributing to the imposition of subspecies names on the Asian bees, but where appropriate we will note some of these names for completeness. Much of what we know about the within-species phylogeography of Asian honey bees stems from the recent analysis of mitochondrial genomes. Mitochondrial genomes are circular DNA molecules, which in honey bees are about 17,000 base pairs long (Crozier and Crozier 1993). There are multiple mitochondria in every cell, which makes mitochondrial DNA extraction and preservation less difficult than it is for nuclear DNA, of which there is only one copy in the nucleus. Mitochondrial genomes are particularly suited to the examination of recent evolutionary events because they evolve quickly, particularly so in their noncoding regions, which are not subject to purifying selection (Palumbi 1996). Within the mitochondrial genome of most A. mellifera exists a noncoding A+T rich region that lies between the cytochrome oxidase (COII) gene and the leucine t-RNA (Crozier and Crozier 1993). This region is extremely variable in A. mellifera, and provides a wealth of haplotype diversity that can be used to examine phylogenetic questions (Garnery et al. 1993). The region is less variable in Asian honey bees, but is still proving useful for within-species analyses (Cornuet and Garnery 1991; de la Rúa et al. 2000; Deowanish et al. 1996; Franck et al. 2000; Garnery, Cornuet, and Solingnac 1992; Garnery et al. 1993; Hepburn and Smith et al. 2001;

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Sihanuntavong, Sittipraneed, and Klinbunga 1999; Smith and Hagen 1996; Smith et al. 2000; Takahashi et al. 2002; Tanaka and Roubik et al. 2001).

Intraspecies Diversity of the Cavity-nesting Bees In the last chapter we referred to the Asian cavity-nesting species as a shrubby tip on the phylogenetic tree of the honey bees (Figure 3.3). We will now examine this tip in some detail at both the species and the withinspecies level. Based on the noncoding region, analysis of mitochondrial diversity in A. cerana and its sibling species A. nigrocincta has shown two major haplotype families. The first appears to be isolated to India and Sri Lanka and may be associated with the yellow “plain” bees recognized by Indian beekeepers (Smith and Hagen 1996). The number of bees sampled is not yet sufficient to determine if the “plain” bees are always associated with this haplotype, but if so, restricted gene flow between the “hill” and “plain” ecotypes suggests that they are cryptic species. The second major lineage includes all other A. cerana and A. nigrocincta (Smith et al. 2000). This second group is so divergent from the first that sequences from the two families cannot be confidently aligned. Within the second group of haplotypes there are at least five lineages, one of which is A. nigrocincta of Sulawesi. The distribution of these five lineages shows a strong biogeographical pattern (Figure 4.2; Smith and Hagen 1996; Smith et al. 2000) that is quite clearly correlated with Pleistocene geography (Figure 4.1). The “Mainland Asia” sublineage has a broad range, being found from India to Japan. In India, there are several variants of the haplotype, which seems to include the black “hill” bees, as opposed to the yellow “plain” bees discussed above. To the east, however, there is remarkably little variation within this sublineage. This suggests a recent selective sweep by bees carrying a single mitotype that spread east from India into southeast Asia and then northward to China, Korea, and the Primorsky district of Russia (Smith et al. 2000). Interestingly, this mitotype is also found in Japan (Kimura et al. 2000). This may suggest a Pleistocene colonization event, since the Japanese islands of Kyushu, Shikoku, and Honshu were joined to Korea at that time. On the other hand, the Pleistocene climate in Japan would have been extremely cold for honey bees, and it is perhaps more likely that the Japanese A. cerana population derives from a more recent

Speciation and Biogeography

DA MAINLAN

61

SIA

India Negros Palawan

Luzon Leyte Cebu Mindanao Sangihe

India Mainland Asia

Malaysia Sulawesi

Sundaland south of Kra ecotone Palawan, Mindanao, Cebu A. nigrocincta—Sulawesi, Sangihe Luzon, Mindanao, Leyte, and Negros

Figure 4.2 A phylogenetic tree showing the main mitochondrial haplotypes of A. cerana and A. nigrocincta and the estimated distribution of these haplotypes across Asia (Smith et al. 2000). The haplotypes of the cavitynesting bees on the Philippine islands of Mindoro and Panay have not yet been identified. The patterns and shadings in the phylogeny refer to the patterns and shadings in the map. Some regions have more than one haplotype lineage.

human-assisted migration from China or Korea after increasing temperatures severed Japan from the mainland. The distribution of the Mainland group abuts that of the “Sundaland” group in Thailand (Sihanuntavong, Sittipraneed, and Klinbunga 1999; Smith et al. 2000) and in Myanmar (Smith 2002), where both types occur. The Sundaland haplotype is additionally found on Ko Samui (Thai-

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land), in peninsular Malaysia, on Borneo, Java, Bali, and Lombok, and on the Philippine islands of Palawan, Mindanao, and Cebu, strongly echoing Pleistocene geography (Figure 4.2; de la Rúa et al. 2000; Smith et al. 2000). Also reflecting Pleistocene geography are the unique haplotypes found on Luzon and Negros (de la Rúa et al. 2000) and the A. nigrocincta haplotype on Sulawesi (Smith et al. 2000). These islands were never joined by land to Sundaland, and thus their bee populations have probably arisen via ancient, pre-Pleistocene, highly bottlenecked migrations over sea. Thailand, possibly specifically the area around the Isthmus of Kra (10oN on the Thai-Myanmar border), is the site of two independent species boundaries (the northern limit of A. koschevnikovi and the southern limit of A. florea), and an area of mixing of the Sundaland and Mainland mitotypes of A. cerana (see above; Sihanuntavong, Sittipraneed, and Klinbunga 1999; Smith et al. 2000). The hybrid zone between two A. cerana subpopulations is also reflected in the nuclear genome (Sittipraneed et al. 2001; Sittipraneed, Sihanuntavong, and Klinbunga 2001). The reason for this “abrupt” change is unclear, but may be associated with the change from dense tropical rainforest to more seasonal open forest (Smith et al. 2000). The Bilauktaung mountain range may also have provided an impediment to gene flow (Smith et al. 2000), directing migrations of bees carrying the Sundaland mitotypes to the west into Myanmar, away from Thailand.

Intraspecies Diversity of the Giant Bees Little is known of the intraspecific variation of A. dorsata and A. laboriosa. A. dorsata has a huge range (Figure 2.9) and not surprisingly shows considerable morphometric and other variation across that range (see, e.g., Bhandari 1983; Maa 1953; Mujumdar and Kshirsager 1986; Sakagami, Matsumura, and Ito 1980; Sharma 1983; Sharma and Thakur 1999; Singh 1981). To our knowledge there have been no comprehensive morphometric or mitochondrial DNA studies conducted on giant honey bees, and for this reason it is our preference to say very little here about intraspecific variation within A. dorsata. But we can suggest three good places to start investigations on the within-species diversity of A. dorsata. These are the populations of the Philippines, Sulawesi, and the Andaman Islands, which lie south of Myanmar in the Bay of Bengal. Maa (1953) suggested that there are two putative giant bee species in the

Speciation and Biogeography

far east of the honey bee’s distribution. On the basis of detailed measurements of pinned specimens, Maa called the giant bees of the Philippine islands east of the Merrill Line—which include the main islands of Luzon, Mindanao, and Negros but not Palawan—Apis (Megapis) breviligula. In contrast, Morse and Laigo (1969) thought that the giant bees of the Philippines are not sufficiently different from A. dorsata elsewhere to warrant species status. Nonetheless, our feeling is that the giant bees of the Philippines are quite different from their conspecifics in Borneo and further west. First, in contrast to the yellow abdomen with black stripes that is typical of the common giant honey bee, A. d. breviligula is a black bee with gray abdominal stripes. Second, Filipino giant bees never form nesting aggregations (Moltás-Colting and Cervancia 2004; Morse and Laigo 1969). Thus there remains the tantalizing possibility that Maa was right and “A. breviligula” should be regarded as a separate species from A. dorsata. As we noted above, Mindanao, Luzon, and Negros were never connected to Borneo, and the possibility of human-assisted migration seems remote with these large and ferocious colonies. Therefore it seems most likely that the Philippine giant bees arose from a small colonizing population that arrived by sea via natural means. This implies that the founder population would have undergone an extreme genetic bottleneck that may well have caused significant genetic drift from the populations farther west. The question of the species status of A. breviligula, at least according to our species definition (Chapter 2), can probably be resolved by an examination of the drone flight time, which is still unknown despite considerable effort (regrettably focused at the wrong time of day—before dusk) by Morse and Laigo to observe it (1969). Maa (1953) also recognized a separate species of giant honey bees on Sulawesi, east of the Wallace Line, Apis (Megapis) binghami. We have been unable to find any information about these bees. Arias and Sheppard (2005) found only minor differences between the genomes of A. dorsata and A. binghami, and did not support species status for A. binghami. The presence of A. dorsata on the Andaman and Nicobar Islands (Ahmed 1989; Ahmed and Abbas 1985) is quite mysterious since these islands are at least 500 km from the nearest continental land mass, Myanmar. Presumably this population also arose from another rare founder event during the Pleistocene. Little is known about this population or its possible distinctness from mainland populations. Ahmed and Abbas (1985) found that the island bees had a slightly smaller brood cell size than that in other

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areas. A molecular analysis to determine the likely origin of the Andaman population would probably reveal much about abilities of A. dorsata to migrate across the sea.

Intraspecies Diversity of the Dwarf Bees Several studies have shown minor morphometric variation among A. florea populations (Bhandari 1983; Chaiyawong 2001; Ruttner, Mossadegh, and Kauhausen-Keller 1995; Sharma 1983; Tahmasebi et al. 2002), but in general the species appears to be remarkably uniform across its range. For example, mainland Thailand, which shows a diversity of A. cerana haplotypes, has but a single A. florea haplotype (Nanork 2001; Nanork, Deowanish, and Wongsiri 2000). As far as we are aware, the only morphometric study of A. andreniformis showed minor differences between a population in eastern Thailand and in Palawan in the Philippines (Rinderer et al. 1995).

Summary Because honey bees are endemic to southeast Asia, a region replete with islands and changing shorelines, there have been many opportunities for speciation events and the creation of locally adapted forms. But the available evidence suggests that island formation has contributed little to speciation within the genus. On the other hand, an understanding of ancient landforms largely explains the present-day distributions of Asian honey bee species, and the genetic and morphological variation within them. The better-studied cavity-nesting bees show a great diversity of mitochondrial haplotypes and a huge range of morphological variants. The ubiquity of A. cerana across Asia, particularly in Japan and in islands east of the Wallace Line, may be in part or whole due to anthropogenic movements of semidomesticated colonies. The giant and dwarf bees are less well studied than the cavity-nesting species, and the structure of their biodiversity is not well understood. Some populations of A. dorsata are extremely isolated (as on the Andaman Islands). Since human-assisted spread of giant bees is unlikely, the origin of these populations is a mystery.



5 Dance Communication and Foraging

When a honey bee forager has found a highly profitable place to gather food she will often perform a communication dance on her return to the nest, which stimulates her nestmates to search for her find. Information about the distance and direction of the goal is encoded in the dance. Unemployed foragers can read this information and then use it to help them find the forage patch that has been located by their nestmate (von Frisch 1967). The rules and grammar of the honey bee’s dance language are now sufficiently well understood that human observers can accurately decipher it. The acquisition of the ability of the honey bee to recruit nestmates and direct them to a food source via the dance language was undoubtedly a decisive event in their evolution. The dance language is particularly important for the Asian species and their ancestors because floral resources are spread patchily in a tropical forest, and the best resources cannot be efficiently located via individual searches (Dornhaus and Chittka 1999, 2004b). The sharing of information about superior forage patches via the dance language gives the honey bees a significant competitive edge over stingless bees, which can recruit nestmates to food only via chemical trails, acoustical signals, and visual attraction (Nieh 2004), and bumble bees, which cannot recruit nestmates beyond indicating that food is available (Dornhaus and Chittka 2004a). Note that the fitness benefit of the dance language is not that dance followers find food sources more easily than individual scouts do. Indeed, dance followers are generally less efficient at finding food than are independent scouts (Seeley 1993). Rather, dancing

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allows the colony as a whole to track the most profitable food sources available. That an animal as simple as a honey bee can possess a symbolic language of sufficient complexity to be able to convey distance and direction information is remarkable, and for this reason the dance language has been intensively studied (see Dyer 2002, von Frisch 1967, and Seeley 1995 for superb reviews). But although much is known about the dance language of A. mellifera, studies of the dance language of the Asian bees are surprisingly few. Nonetheless, given the information that is available, it is apparent that the major features of the honey bee dance language are conserved across the genus. We begin with a description of these general features, followed by a discussion of the species-specific details of the individual languages. On the basis of these descriptions we attempt to reconstruct how the dance language evolved. Finally, we conclude the chapter by describing the special dances that are used for coordinating drone flights in A. andreniformis, reviewing some estimates of typical foraging ranges of Asian honey bees, and discussing the various ways in which the species partition the available forage. Honey bees also use dances to coordinate finding new nest sites and making migrations. These nonfood dances are discussed in the next chapter.

General Features of the Dance Language In all species, dancing for a food or water source is presaged by a successful forager returning home and running excitedly through the nest to a special area, the “dance floor,” where dances take place, nectar is unloaded, and unemployed foragers congregate seeking information about productive foraging sites from their successful nestmates. This area can be viewed as the coordination center for the colony’s foraging effort. If at the dance floor the forager has difficulty finding house bees to relieve her of her load, then this signals her that there is no shortage of successful foragers, and she will not dance (Anderson and Ratnieks 1999; Seeley 1992). But if eager house bees rapidly approach her in order to relieve her of her load, the forager is informed that there is a shortage of nectar in the nest, and she is stimulated to perform a dance. The location of the dance floor differs among the three groups of honey

Dance Communication and Foraging

bees. In the dwarf bees the dance is performed on the comb or backs of the workers sitting on the crown of the nest. In the giant bees, dances are performed on the curtain of bees that shroud the nest. (Foragers may also dance on the comb beneath the curtain, but this has never been reported.) In the cavity-nesting bees, foragers dance on the vertical comb surface just inside the nest entrance. The different locations of the dance floor of the three groups of species dictate important species-specific idiosyncrasies of the dance language, which we will discuss below. Up to five or six unemployed foragers face the dancer, antennate her, and try to follow her steps. In doing so, they gain the information coded in the dance (Judd 1995). The duration of the dance and its vigor probably provide information on the forager’s perception of the value of the food: for a given distance, more excited dances with more waggle runs are performed for rich sources of food; shorter, more subdued dances are performed for less profitable patches (Seeley, Mikheyev, and Pagano 2000). The perception of what is danceworthy is not a fixed measure, for the quality of food required to elicit a dance varies. When food is scarce, even poor-quality food sources will elicit a dance. When food is plentiful, and especially when foragers have difficulty getting unloaded by house bees, only first-quality food sources will elicit a dance (Seeley 1992; Seeley, Camazine, and Sneyd 1991). For most dances the dancer runs in the famous figure-of-eight pattern (Figure 5.1). The dancer begins the dance by striding forward for a distance approximately 1.5 times her own length while vigorously shaking her body from side to side. This phase of the dance is usually called the “waggle run” and the dance is called a “waggle dance.” At the end of the run, the dancer makes a sharp turn to the left or right and returns, without wagging, to the point where she commenced the waggle run. She then retraces the waggle run, making a sharp turn in the opposite direction at the end of the run to again return to the waggle axis. Hence, with every second execution of the dance, she has traced out an approximate figure of eight (Figure 5.1). A parsimonious interpretation of the dance of a successful honey bee forager is that it is a symbolic representation of her most recent foraging trip (von Frisch 1967; Wilson 1971). The waggle run is the most important phase of the dance for transferring information. In effect, the follower bees rehearse the flight they must take to get to the food source being ad-

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Increasing distance

Figure 5.1 Forms of the honey bee dance: from left to right, round, waggle, and transition. Adapted by permission of the publisher from The Dance Language and Orientation of the Bees by Karl von Frisch, trans. Leigh E. Chadwick (Cambridge: The Belknap Press of Harvard University Press, copyright © 1967, 1993 by the President and Fellows of Harvard College), p. 61.

vertised by the dancer by following her waggle runs. The followers may also emit brief, high-pitched sounds that call on the dancer to stop (Dreller and Kirchner 1994; Towne 1985a), though the function of this is unknown.

Distance Information Over most distances, the duration of the waggle run is linearly correlated with distance, such that circuits are completed quickly for nearby sources, and slowly for more distant ones. This shows that each circuit of the dance represents the flight, rather than the whole dance. Thus a dance for a short distance can be protracted because it contains many circuits, whereas a dance indicating a great distance may be brief because it contains very few (albeit protracted) circuits. Occasional dances are extremely protracted, with many tens of circuits lasting several minutes in total. But most dances consist of just a few circuits. The waggle run becomes more and more hurried the closer the food goal. At some point, when the goal is close to the colony, the linearity of the dance curve is broken, and the dances become more hurried (Figure 5.2). This inflection point in the dance curve is known as “the break point” (Dyer 1987; Sen Sarma, Esch, and Tautz 2004). Eventually, as the goal gets even closer to the nest, the dancer is apparently unable to make the return runs quickly enough, and the figure-of-eight pattern becomes distorted

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Distance (m) Figure 5.2 The relationship between distance and dance tempo for A. florea, A. dorsata, and A. cerana. The dance circuit duration increases with distance for all species. Left: Lindauer (1956), working in Sri Lanka, found that the dance distance curves varied with the species. Illustration redrawn with the permission of the copyright owners from M. Lindauer, “Über die Verstänigung bie indischen Bienen,” Zietschrift für Vergleichende Physiologie 38 (1956): 521–527; copyright Springer journals, 1956. Right: Dyer and Seeley (1991a), working in northeast Thailand, found no differences in the dance distance curves for these same species. Redrawn with permission of the copyright owners from F. C. Dyer and T. D. Seeley, “Dance dialects and foraging range in three Asian honey bee species,” Behavioral Ecology and Sociobiology 28 (1991): 227–233; copyright Springer journals, 1991.

into what is known as a transition or sickle dance. At even closer distances, the dance form loses its figure-eight pattern altogether. This form of the dance is known as a round dance, wherein the bee runs quickly in excited circuits, sometimes wagging her abdomen, sometimes not (Figure 5.1; von Frisch 1967). Nonetheless, at least in A. mellifera, some distance and direction information is encoded even in these dances (Jensen, Michelsen, and Lindauer 1997; Kirchner, Lindauer, and Michelsen 1988), though whether recruits can use it is unclear. To determine the relationship between distance and the frequency of circuits, an experimenter normally trains a number of foragers to a dish containing concentrated (2 M) scented sugar syrup. After 10–20 bees are faithfully foraging at the dish, a meter or so from the colony, the bees are individually marked with paint dots or numbered tags. A second observer then records the number of dance circuits made by the marked bees as they advertise the rich source of food. The dish is then gradually moved out by 10-m intervals, and the dances of about 10 bees recorded at fre-

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quent intervals. Curves, similar to those of Figure 5.2, are obtained from these kinds of studies (Boch 1957; Dyer and Seeley 1991a; von Frisch 1967; Gould 1982; Lindauer 1956; Punchihewa et al. 1985; Sen Sarma, Esch, and Tautz 2004). The slope of the curve varies depending on the ecotype being studied and where the experiment is conducted. Much has been made of the differences in slope of these curves that relate distance to dance tempo, and the different slopes have been termed “dialects” of the honey bee dance language (Dyer and Seeley 1991a; von Frisch 1967; Gould 1982; Lindauer 1956; Punchihewa et al. 1985). Recall that a fast dance indicates a nearby food source whereas a ponderous one indicates a distant one (Figure 5.2). Clearly there are limits to the distances the dance language can indicate, imposed by the ability of dance participants to dance quickly or slowly enough. Thus an adaptively tuned dance curve would have the fastest possible dances for nearby sources, and the slowest feasible dances for the most distant sources that a bee of that ecotype is ever likely to visit. This idea led to the hypothesis that the variation between dance curves that have been observed between species of Apis (Lindauer 1956) and between the various subspecies of A. mellifera (von Frisch 1967) has been shaped by natural selection to reflect the typical foraging ranges of each ecotype. Thus in environments in which foragers are sometimes obliged to forage at great distances from their nests, natural selection will act to flatten the dance curve, so that dance circuits don’t become too lengthy and imprecise at the outer limit of the foraging range, and to reduce the time required for a dance. By contrast, in environments where flight ranges are short, a steeply rising dance curve provides more precise information about distances (Figure 5.3). A. florea are small bees and A. dorsata are large. This fact led Lindauer (1956) to the idea that A. florea would have a small foraging range, and that their dance calibration curve would accordingly rise steeply, providing accurate distance information over a relatively short range of foraging distances. Conversely, larger A. dorsata was predicted to have a large foraging range, and a commensurately slowly rising dance curve, leading to different “dance dialects” in these species. Data gleaned by Punchihewa et al. (1985) and Lindauer (1956) supported the adaptive dance dialect hypothesis. They showed that in Sri Lanka the dance curves of A. dorsata, A. florea, and A. cerana were significantly different, and varied as one

Dance Communication and Foraging

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Flight distance Figure 5.3 The tuning of dance dialects to maximize precision of distance information over typical foraging ranges. The optimal relationship—the one that will provide the most accurate information to recruits over typical foraging ranges—between flight distance and circuit duration should maximize the slope of the line subject to the constraint that the maximum circuit duration should not be too long (Gould 1982). The maximum circuit duration should correspond to the typical maximum flight distance (Dyer 2002; Dyer and Seeley 1991a; von Frisch 1967). Redrawn with permission from the Annual Review of Entomology, vol. 47, © 2002 by Annual Reviews, www.annualreviews.org.

might predict on the basis of the bees’ size and inferred foraging range. Dyer and Seeley (1991a), however, were unable to replicate these findings in Thailand, where they found that A. florea, A. dorsata, and A. cerana have dance circuit–distance curves that are not significantly different from one another (Figure 5.2). Another aspect of the dance dialects of different species is the transition points between round dances and waggle dances. In A. mellifera, the transition points between round dances and waggle dances are genetically influenced (Johnson et al. 2002; Rinderer and Beaman 1995) and highly variable. Boch (1957) reported that some ecotypes (e.g., Middle Eastern A. m. fasciata) show a transition from round dances to transition dances when the goal is about 3 m from their colony, and converge on true waggle dances when the goal is about 10 m away, whereas eastern European A. m. carnica perform round dances up to 20 m and do not converge on waggle

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dances until 85 m. The Asian species seem to converge on waggle dances within a few meters of their colony: 5 m for A. cerana, 2.5 m for A. dorsata, and less than 1 m for A. florea (Towne 1985b). This may indicate that the dance dialect of the Asian species (at least in south India, where they were studied by Towne) are tuned to provide accuracy over shorter distances than are those of A. m. carnica. But Sakagami (1960) reports a much longer transition point for Japanese A. cerana, and Sen Sarma et al. (2004) could find no consistent differences in the transition points of A. florea and A. m. carnica. These variable results suggest that the transition point is strongly influenced by both genetic and environmental factors. Therefore it is important that studies of dance dialects be conducted in the same environment. Interestingly, Esch et al. (2001) showed that distance perception by foragers is determined by the amount of optical information that passes the eye of the bee as she flies along. Thus a bee that flies over a complex landscape with great visual stimulation perceives that she is flying a long way, whereas a bee flying over a plain surface, such as a lake, perceives that she is barely moving at all. This means that the environment in which the dance dialect experiment is conducted must have considerable bearing on the shape of the curve, and possibly more so than the race or species of bee that is being studied (Esch et al. 2001). We should add here that until recently the “optic flow” hypothesis for the perception of distance by a flying bee competed with an “energy expenditure” hypothesis, in which it is presumed that a bee estimates distance by determining how much flying effort is required to reach the goal (Esch and Burns 1996; von Frisch 1967). When bees fly upwind or uphill, or are forced to walk for a bit or to carry small weights, they invariably indicate a longer distance in their dance than if they are unimpeded. The impediments, however, also cause the bee to fly closer to the ground, which increases the rate of optic flow past the eye. The issue has now been resolved by a series of ingenious experiments in which bees were trained to fly up tunnels of varying visual texture (Esch et al. 2001; Srinivasen et al. 2000). In a tunnel of great visual texture bees can be fooled into indicating a distance of hundreds of meters in their dances, when in fact they have traveled only 10 m. These experiments clearly show that optic flow is more important than energy expenditure in a forager’s perception of her foraging trip.

Dance Communication and Foraging

Information about Direction When we walk through an unfamiliar landscape we may use an arbitrary reference point on the horizon (called north) and a compass bearing to maintain a trajectory that is a precise angle from the arbitrary reference point. Furthermore, if we wish to tell other people how to get to a place that we know of, we can tell them to maintain the same bearing as we did, and how far to walk. With repeated trips to the same location we eventually become familiar with landmarks along the way, and can then maintain the correct bearing without constantly referring to our compass because we know where we are by reference to the now-familiar landmarks. The mechanism that foraging bees use to maintain their flight trajectory is remarkably similar to the one that we humans use. The fundamental difference between our way and the bees’ way of navigating is that their arbitrary reference point is not north, but that point on the horizon that is indicated by an imaginary line drawn down perpendicularly from the sun— the azimuth (Figure 5.4). As a bee flies toward her goal she keeps track of her travel direction by maintaining a precise angle from the sun’s azimuth as she flies along. Even when the sun isn’t visible because it is obscured by cloud or forest, a bee can still maintain her trajectory by referring to known landmarks and interpolating where the sun would be if she could see it. If there is any blue sky visible, then the bee can also estimate the sun’s location by using the pattern of the polarized light coming from that patch of sky (Dyer and Gould 1981). When reenacting her flight on the dance floor for the benefit of her nestmates, the successful forager does the same thing: during the waggle run she orients her body at the same relative angle from the current position of the sun’s azimuth, as she did during her recent flight. Follower bees learn the angle at which to fly away from the sun by assessing the angle at which the dancer waggles. In the dwarf bees where the dancer usually has a flattish surface to dance on, the dancer can maintain the correct angle by orienting her trajectory with the sun during flight, and maintaining an identical angle in during the waggle run. This means that she points at the food on her waggle run. But in the giant bees and the cavity-nesting bees, the dancer compensates for the fact that she usually dances on a vertical surface rather than a horizontal one—she can’t point directly. So it is at

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A

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C Figure 5.4 How direction information is conveyed by the waggle run in dwarf bees (A), giant bees (B), and cavity-nesting bees (C). In dwarf bees the forager dances on a horizontal plane and directly “points” at the goal. In giant bees and cavity-nesting bees the angle of the waggle run deviates from the vertical at the same amount as the sun’s current azimuth from the direction of the goal.

Dance Communication and Foraging

this point in our description of the dance language that we will need to discuss the dwarf, giant, and cavity-nesting species individually.

Dwarf Bees Here our description relates only to A. florea, for the dances of A. andreniformis have never been described. We assume, perhaps naively, that they are similar. Laden foragers can land on any part of the nest (Dyer 1987), but more often than not they land on the upper section of the curtain of bees that covers the vertical surfaces of the comb (Koeniger et al. 1982). Irrespective of where the food source is located, any waggle runs performed on the curtain of bees are always oriented straight upward toward the crown of the nest (Dyer 1987), where the dance floor proper is located. The purpose of this directionally meaningless phase of the dance may be to alert potential recruits that the forager is about to dance, and to collect an audience before the informative stage of the dance begins. Each of the vertical waggle runs on the curtain ends a little farther forward of where it started, so the dancer quickly arrives at the crown. Here the forager changes the direction of her waggle runs so that they point directly in the direction of the food source (Figure 5.4; Dyer 1987; Koeniger et al. 1982). The process of starting each waggle run a little forward of the previous one continues on the crown (Koeniger et al. 1982), so that the dancer quickly traverses it. On arriving at the opposite edge the worker may continue over the edge so that the dance ends with waggle runs oriented straight downward at the point where any recruits leaving the nest to fly for the food should fly from (Dyer 1987). Otherwise the dancer may pause at the edge of the crown and execute several waggles on the crown that point directly toward the food (Koeniger et al. 1982). The importance of the sun to the orientation of the A. florea dance can be demonstrated by placing the nest in a box or building so that the dancers cannot see it (Koeniger et al. 1982). This causes dances to become disoriented (Lindauer 1956). Dances can be reoriented by providing a surrogate sun with the aid of a mirror (Figure 5.5; Koeniger et al. 1982). Turning the mirror through α degrees changes the angle of the reflected sun perceived by a dancing bee by 2α (Koeniger et al. 1982). Sure enough, when

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Figure 5.5. A: An experiment demonstrating the influence of the position of the sun on the orientation of dances by A. florea (Koeniger et al. 1982). A colony was placed in a room with no view of the sky, which caused dances to become disoriented. Foragers could exit the room via an open window. An image of the sun provided by a mirror allowed foragers to orient their dances. Moving the mirror through 45o changed the perceived position of the sun by 90o. The orientation of waggle dances also changed by 90o, which shows that A. florea foragers use the current position of the sun to orient dances. B: An experiment showing that that A. florea can use a terrestrial substitute for the sun to orientate dances (Dyer 1985a). Here the bees that had no direct view of the sun used a conspicuous striped board to orient their dances. Changing the position of the board through 180o changed the orientation of dances by the same degree.

Dance Communication and Foraging

Koeniger et al. moved their mirror through 45°, many of the dancers promptly turned their dance angle through 90° (Figure 5.5). Any communication system that requires participants to know where the sun is positioned is vulnerable to cloudy days. Hence A. florea have a back-up if the sun is obscured by cloud. A. florea can interpolate the current position of the sun from the time of day and from landmarks whose location they have learned. Dyer (1985a) deployed a conspicuous striped board that was visible to dancers that had no direct view of the sky and so could not directly use the position of the sun to orient their dances. Moving the board changed the orientation of dances in the same way as moving the Koeniger’s mirror changed them. Dyer’s brilliant experiment shows an important aspect of how bees perceive the world. The direction of everything is perceived to be as a deflection from the calculated or perceived current azimuth of the sun. Can’t see the sun? Figure out where it should be from the time of day and the location of landmarks. Note that this system does not require all bees to use the same landmarks. There is only one gold-standard landmark (the current position of the sun). Individual bees are free to use whatever terrestrial cues they like to figure out the common celestial reference point. During the wagging run of the A. florea dance, the dancer holds her abdomen aloft and her wings are spread. Follower bees tend to stand back from the dancer more than in the cavity-nesting species, and this may be because they can see the vibrating abdomen of the dancer and gather information from it without being in direct physical contact with her. A. florea usually performs dances on a more or less horizontal surface. To compensate for the slight curvature of the crown of the nest the dancer adjusts the angle of her head so that she always perceives that she is dancing in the same horizontal plane (Dyer 1985a, 1987) (other species are not known to do this). But if a nest is rotated so that the crown is no longer horizontal, dancing is severely disrupted. If the nest is rotated through 180°, the dance floor is immediately moved to the pointed tip (previously the bottom of the nest), where foragers recommence dancing on the (highly restricted) horizontal plane. If the crown of the nest is covered by a notebook or something similar, so that dances cannot be performed in the horizontal plane, dances become completely disoriented, or the bees will start dancing on top of the notebook (Lindauer 1956). Nonetheless, A. florea

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Figure 5.6 Dances of A. florea on a vertical plane, such as occur on the surface of swarms. The waggle run points directly at the goal (indicated by the bull’s-eye) and is unaltered by the current position of the sun (Dyer 1991b).

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workers execute directionally meaningful dances on the vertical surface of combless swarms (Dyer 1985a; Nanork et al. 2005). When dancing on a vertical plane on the surface of a swarm, A. florea point at the goal directly on their waggle run (Dyer 1985a). Thus if the goal is directly ahead, the waggle run is oriented straight upward. If it is directly behind the dancer, the waggle run is oriented straight downward. This means that the angle of the waggle differs depending where on the swarm the dance occurs (Figure 5.6).

Giant Bees The giant bees build their nest beneath a supporting tree or cliff overhang, and so, for many nests, there is no horizontal surface upon which to create a dance floor. Instead, dances always occur in the vertical plain on the backs of curtain bees in a particular area known as the “mouth” (Morse and Laigo 1969). At the mouth, the curtain is much looser than it is elsewhere, so that arriving foragers can easily penetrate the curtain and find the comb beneath. In A. florea, the current position of the sun makes no difference to the angle of the waggle run. A. florea dancers always “point” at the goal on their waggle run, and only use the sun’s position as a guide for where to point. In contrast, dancing giant bees do not directly point at the direction

Dance Communication and Foraging

of the goal during the waggle run. Instead, in an important evolutionary step, dancers use a symbolic reference point from which dances are oriented. Giant bees all agree that a waggle run pointing straight up the comb points to a food source in the direction of the sun’s current azimuth. Dances that do not point straight up the comb tell of food sources that can be found by deviating from the sun’s azimuth to the same degree as the waggle run of the dance is deflected from straight up (Figure 5.4; Lindauer 1956). Hence giant bees have substituted negative geotaxis for positive phototaxis as their guide when dancing. This means that the angle of the waggle run of dances for a particular goal varies throughout the day to compensate for the sun’s movement, but at any moment in time all dances for a particular goal will have the same angle. Presumably the evolutionary shift from pointing directly to pointing in relation to a symbolic reference point occurred in response to normal forager dances occurring on a vertical plane. When the simpler A. florea form of the dance language is performed on a vertical surface, it requires precise adjustments of the dance angle depending where on the surface of the swarm the dance occurs (Figure 5.6). Thus both the dancer and the followers need to know where they are on the swarm in order for information to be successfully transferred. In contrast, the dance for a particular goal performed by A. dorsata or the cavity-nesting bees has the same angle from the vertical wherever it is performed in the nest. A second important adaptation in the dance of A. dorsata is a wing buzz emitted during the waggle-run phase of each dance circuit (Dreller and Kirchner 1994, 1995; Kirchner 1993; Kirchner and Dreller 1993). Although dancing A. dorsata hold their abdomens aloft and their wings spread during dancing as A. florea do, the species commonly forages and dances on moonlit nights (Figure 5.7; Dyer 1985b), when it is presumably difficult for followers to see the dancer clearly. Possibly, dancing may also occur under the curtain of bees, where it would be difficult for followers to see the dancer. Presumably to compensate for reduced visual cues, dancers generate 50 ms sound pulses of about 130 Hz by dorsoventral vibration of their wings during the waggle run (Kirchner and Dreller 1993). A. dorsata can detect sounds in this range (using the Johnson’s organ at the base of the antenna) and can learn to use sound of this frequency to locate a food reward (Dreller and Kirchner 1994). It is assumed that the sounds help bees attending the dance to follow the dancer and to thereby derive information from her. The duration of the buzz sound corresponds to the duration of

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Figure 5.7 A. dorsata workers foraging well after dark at 20:00 hrs on a kapok (Ceiba pentandra) inflorescence. Photo by S. Wongwirat.

the waggle run and thus to the distance to the goal (Dreller and Kirchner 1994). Interestingly, the mountain-dwelling A. laboriosa does not forage at night, presumably because it is rarely warm enough. This species apparently does not emit sounds when dancing (Kirchner et al. 1996). If this report is accurate, (it could be that the buzz is of such low frequency that it can’t be detected), it is quite good circumstantial evidence that the sound component of the dance language has developed in A. dorsata to facilitate information transfer in the dark. A. dorsata has a further adaptation to help it forage at night. The ability of an insect to see at night is limited by the number of photons that can be captured by a single ommatidium. But A. dorsata can resolve objects at light intensities below that which is theoretically possible given the diameter of their ommatidia (Warrant, Porombka, and Kirchner 1996). This implies that the A. dorsata brain has the ability to pool inputs from several ommatidia, and form a neural representation based on this collective information. Interestingly, even though A. dorsata will forage at night only when the moon is out in half phase or more, they do not use it as their celestial reference point for nocturnal foraging. Rather, they interpolate the sun’s

Dance Communication and Foraging

movement as a point of reference for dancing throughout the night (Dyer 1985b).

Cavity-nesting Bees The dances of cavity-nesting bees are performed in the dark, on a vertical comb surface of a multicomb nest. They therefore require the same adaptations as A. dorsata: the ability to convert the angle of the goal from the sun’s azimuth to the angle from the vertical, and the ability to convey their wagging motions in the dark via sounds. In addition, they delineate a particular area of the nest as the official dance floor, so that dancers and recruits can be efficiently brought together. This area is probably marked by pheromones so that dance participants can locate it (Tautz and Lindauer 1997). The 300-Hz wing vibrations during the waggle runs of A. cerana and A. mellifera are significantly more pronounced than those of A. dorsata (Dreller and Kirchner 1994; Kirchner and Dreller 1993), and it has been argued that this is because information transferred during the waggle run is received by follower bees via vibrations in the air. Evidence supporting this view includes reduced efficiency of dances from a mutant strain with short wings (Kirchner and Sommer 1992). In addition, dancing bees may use substrate vibrations to attract the attention of other bees in the dark. Evidence from high-speed video recordings of the waggle run of the dance shows that A. mellifera grasps the comb with all six legs and really gives it a shake at about 13 Hz (Tautz, Rohreseitz, and Sandeman 1996). The dance floor of A. mellifera tends to be on unsealed brood combs, which may resonate better than sealed brood combs (Sandeman, Tautz, and Lindauer 1996; Tautz 1996). It should be noted, however, that comb vibrations are not necessary for information transfer in A. mellifera. Dances that are performed on the surface of combless swarms are highly accurate (Weidenmuller and Seeley 1999) and are accurately perceived (Seeley 2003; Seeley and Buhrman 1999).

Evolution of the Dance Language One of the most important insights about the dances of cavity-nesting bees is that they have retained many of the characteristics of the A. florea

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dance form despite 6–10 million years of speciation. If an observation hive containing an A. mellifera colony is turned on its side so that the combs are horizontal, foragers are unable to perform oriented dances. If, however, the real sun or a point light source is provided, they can use that cue as a reference point from which to orient their dances and, like A. florea, align their waggle runs so that they point directly at the goal. In other words, if A. mellifera (and presumably the other cavity-nesting bees) are provided with a horizontal surface and a view of the sun, they can revert to the ancestral form of the dance language. On swarms, where all species are obliged to dance in a vertical plane in the open, dances are often oriented directly to the goal, and do not use gravity to orient the dance. Given this and the phylogeny of the genus presented in Chapter 3, we can reconstruct a scenario for the evolution of the genus (Figure 5.8; Dyer and Seeley 1989; von Frisch 1967). In the ancestral species of the genus a successful forager probably behaved similarly to a successful bumble bee or stingless bee forager, running excitedly through the nest. Before taking off it probably stood on the edge of the comb shaking its body while aligning itself with reference to the sun in the direction of the goal. It may have taken off and returned to the nest. Recruits probably followed it some or all of the way to the food source, as they do in some stingless bee species (Nieh 2004). The dance became more ritualized, and approached the form currently seen in the dwarf bees, with bees directly pointing at the goal during the waggle run and using celestial cues and landmarks for orientation. The species used time compensation to predict the movements of the sun when it was obscured. In the lineage leading to the giant bees and the cavitynesting bees, gravity was added as a reference to the sun’s current azimuth because no horizontal plane was available for dancing, and accuracy of dances can be improved by using a standard waggle run angle to indicate the sun’s azimuth. Sound, generated by wing vibrations, arose as an adaptation to dancing in darkness. Kirchner et al. (1996) suggested that sound generation evolved in the common ancestor of A. dorsata and the cavity-nesting bees. This would require loss of sound production in A. laboriosa. An equally parsimonious hypothesis would be that sound production has been independently acquired in the cavity-nesting bees and A. dorsata, or that A. laboriosa can use sound production when needed.

Dance Communication and Foraging Dwarf bees

Giant bees

Dances in total darkness

Slope compensation

Buzz running to indicate direction, celestial cues for dance orientation

Cavity-nesting bees

Dances in the vertical plane, sound production

Figure 5.8. A reconstruction of the evolution of the dance language based on the most likely phylogeny of the genus Apis.

The Curious Dances of A. andreniformis Drones The dwarf black bee mates at about midday. From about 11:00, drones start congregating on the crown of the nest in tight huddles of five to six males. (A. florea drones aggregate in the same way, but later in the day.) As soon as the sun has passed its highest point the males become highly excited. A few individuals will then perform a dance, the dancer being followed by three to four of his brothers in exactly the same way as workers follow a forager’s round dance (Rinderer et al. 1992). At the end of the dance the dancer takes off, followed by his dance partners. Such dances have never been reported in any other species. This extraordinary observation is further evidence that the original function of the dance was to stimulate nestmates to take off with the dancer. Why A. andreniformis males take off in small groups rather than singly as they do in other species we don’t know. Perhaps the brothers support each other’s mating attempts at the DCA, or the coordinated flight provides herd protection from predators such as weaver ants that often catch departing drones. The only drone dances that have been observed are identical in form to the round dances performed by workers (Rinderer et al. 1992). This may indicate that the DCA was close to the colony observed, and that waggle dances would be performed for a more distant DCA. Since the location of

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DCAs of A. andreniformis are unknown, it is possible that the drone dances could be helpful in locating a DCA.

Foraging Ranges When one is creating distance calibration curves by moving a feeding station farther and farther from the nest, there comes a point where foragers will abandon it. When bees are desperate for food, this abandonment distance is presumably near the end of the species-specific foraging range. Lindauer (1956) could train cerana to only about 350 m from their nest, but Darade et al. (1989) trained them 950 m and Dyer and Seeley (1991a) and Dhaliwal and Sharma (1974) trained them beyond 1,200 m. This suggests that the foraging range of A. cerana is at least 1 km. A second way to determine typical foraging ranges is to use the technique of beelining. Beelining involves placing a bait of comb containing scented sugar syrup in likely bee habitat and waiting for it to be located by wild bees. Such a bait is usually found by a forager, which then recruits her nestmates to the bait. The nest is located by gradually moving the bait in the direction of the flying bees returning to their nest. Using this technique, Seeley et al. (1982) found that most A. florea nests were within 200 m of the place where the first forager was attracted to the bait, and A. cerana within 1 km. A better sample of typical foraging ranges is obtained by using a distance calibration curve such as those in Figure 5.2, to estimate the distances being indicated by dances for natural forage sites (Sasaki, Takahashi, and Sato 1993; Visscher and Seeley 1982). Such estimates have been obtained for A. cerana, A. florea, and A. dorsata in Thailand (Dyer and Seeley 1991a) and Sri Lanka (Punchihewa et al. 1985). We have reproduced Dyer and Seeley’s results in Figure 5.9. These show that the typical foraging ranges are different. In A. cerana half of the dances indicate foraging sites less that 196 m from the colony, 269 m for A. florea, and 864 m for A. dorsata, about four times the typical foraging range for A. cerana. In A. cerana, 95 percent of dances indicate food less than 1 km from the colony, 1.5 km in A. florea, and 4 km in A. dorsata. Nonetheless, some rare dances suggested extremely distant foraging places; over 15 km for A. florea and 21 km for A. dorsata. In Japan, the average foraging range for A. cerana was estimated to be 2.1 km (Sasaki, Takahashi, and Sato 1993). This suggests

Proportion of dances (%)

Dance Communication and Foraging 40 30 A. cerana A. dorsata

20

A. florea

10 0 1

2

3

4

Distance (km) Figure 5.9. The distribution of distances indicated by dances for A. cerana, A. dorsata, and A. florea in northeast Thailand. Some dances for the most distant sites have not been included. Redrawn with the permission of the copyright owners from F. C. Dyer and T. D. Seeley, “Dance dialects and foraging range in three Asian honey bee species,” Behavioral Ecology and Sociobiology 24 (1991): 227–233; copyright Springer journals, 1991.

that all species are capable of great foraging ranges if required, but generally work within a few hundred meters of their nest. Dyer and Seeley’s results suggest that there is no correlation between body size and typical foraging range. Although the dances for the greatest distances were seen in the largest bee, A. dorsata, the next greatest distances were indicated by the smallest bee, A. florea. Experimental determination of a distance calibration curve also allows estimation of typical flight speeds of foragers. Towne (1985b), working in India, found these flight speeds for foragers working at an artificial feeder (mean ± s.d.): A. cerana, 7.9 ± 1.1 m/sec; A. dorsata, 7.7 ± 0.5 m/sec; A. florea, 6.1 m/sec. Dyer and Seeley (1987) found similar flight speeds for A. cerana and A. dorsata (c.a. 7.2 m/sec) but that A. florea was much slower: 4.8 m/sec. The speeds estimated for the Asian species are similar to those estimated for A. mellifera, 7.4. ± 0.6 m/sec (Wenner 1963).

Forage Partitioning In much of tropical Asia at least three honey bee species (and often several stingless bee species) are sympatric and apparently successful. Given the

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very high density of honey bee nests in some areas (Inoue, Adri, and Salmah 1990; Rinderer et al. 2002; Salmah, Inoue, and Sakagami 1990), how do all the species coexist without severe interspecific competition? Part of the answer seems to be that foragers of the different species specialize on particular floral resources. Worker size plays an important role in determining which floral resources are available to a species. Obviously flowers of highly specialized morphology may have nectaries that are inaccessible to some bee species: small bees may not be able to reach the nectaries of flowers with very long corolla tubes, and large bees may not be able to gain access to some very small flowers with deeply hidden nectaries. But most flowers have nectaries and stamens that are easily accessible, and resource partitioning arises from more subtle causes. Large bees such as A. dorsata, which have high energetic requirements while foraging, can profitably utilize only floral resources with high food value. Smaller bees with a lower energetic requirement when foraging can profitably exploit more dispersed resources that are not available to larger bees (Oldroyd, Rinderer, and Wongsiri 1992; Roubik 1989). Moreover, the quantity of food available may change over the course of a day, owing to exploitation or to changes in nectar production, and this means that the profitability of a plant species for different bee species also changes over the day (Hubbel and Johnson 1978; Real 1981; Schlising 1970). Oldroyd et al. (1992) studied the appearance of A. cerana, A. dorsata, A. florea, and A. andreniformis on several inflorescences of the king palm Archontophoenix alexandrea located near Chanthaburi, Thailand. This palm produces copious quantities of pollen overnight, and this resource is a useful prize for the first honey bees that can reach it in the morning. In our study, the first bees to arrive, just before dawn, were A. cerana, followed by A. dorsata shortly after dawn. Less than an hour after dawn virtually no A. cerana or A. dorsata remained, but were replaced by large numbers of A. florea, A. andreniformis, and often some stingless bees (Trigona spp.) as well. By midday, nearly all bees except the Trigona had abandoned our palm inflorescences. How can we explain this pattern? In order to fly a honey bee needs to achieve a thoracic temperature, TTh, of at least 27°C (Dyer and Seeley 1987; Heinrich 1979b). The ability of a forager to achieve this critical temperature in its flight muscles early in the morning depends on the ambient temperature, TA, the colony temperature, TC, and her ability to achieve a differential between TA and TTh via

Dance Communication and Foraging

production of metabolic heat. Once in flight, the forager will lose heat via convection, and this will be primarily determined by body mass and wind speed. Because their nests are multicombed and somewhat insulated, cavitynesting bees such as A. cerana can probably maintain a higher proportion of foragers with a TTh above 27°C than the open-nesting species. Presumably this is why A. cerana are the first bees to forage in the morning: being a cavity-nesting species they are able to achieve nest temperatures that allow significant numbers of foragers to fly early when ambient temperatures are relatively low. Furthermore, A. cerana can generate thoracic temperatures 14–21°C above TA, whereas A. dorsata can generate only 9–16° (Dyer and Seeley 1987). Thus A. cerana maintains the highest differential between TTh and TA, which must help them forage on cool mornings. A. dorsata arrive second, presumably because they are large bees that live in large colonies, and are therefore able to generate sufficient metabolic heat to be able to release foragers on cool mornings despite being opennesting. The dwarf bees cannot release foragers until later in the day when ambient temperatures exceed 20°C (Dyer and Seeley 1987), and often do not do so until temperatures are much higher. The larger bees rapidly deplete the resource, and because of their greater metabolic costs cannot forage profitably after this depletion. The dwarf bees, being smaller, can still forage profitably, despite the depleted resource (Oldroyd, Rinderer, and Wongsiri 1992). In addition to temporal partitioning of resources, foragers of different honey bee species seem to partition resources by being distributed heterogeneously in space. For example, in Tenom, Borneo, Rinderer, Marx, et al. (1996) found heterogeneous proportions of A. andreniformis, A. dorsata, A. cerana, and A. koschevnikovi at different heights in a single yellow flame tree, Peltophorum pterocarpum. Rinderer et al. proposed that bees of the different species have “stratum fidelity,” preferring to forage in different areas of the forest canopy. We suggest that heterogeneous availability of nectar is a more likely explanation. Bees of all species can exploit flowers of high nectar secretion, but the larger bees cannot exploit the less profitable flowers on the tree. Another possibility is that larger bees have greater difficulty in not overheating when foraging in direct sunlight, preferring to forage in shadier parts of the stratum. The lethal TTh for insects is about 46°C (Dyer and Seeley 1987). This means that foragers of these larger species need to stop foraging or start dumping heat by exposing nectar on the

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tongue as an evaporative cooler at ambient temperatures above 30°C (Dyer and Seeley 1987), as A. mellifera do (Heinrich 1979a). But since the smaller A. florea generate thoracic temperatures only 4–5°C above TA (Dyer and Seeley 1987), this means that they can forage on all but the hottest days without the need to dump heat, which perhaps allows them to forage on the tops of the canopy, where the larger bees are disadvantaged. It may also be that large bees avoid the tops of trees in response to predation (Roubik 1993). In at least two areas resources are partitioned between two similar species by their physical separation. In the Himalayan Range and its extensions into southeast Asia, A. dorsata exploits lowland forests whereas A. laboriosa nests in upland forests and mountain pastures from 2500 m to 3,200 m and forages up to 4,100 m (Roubik, Sakagami, and Kudo 1985). In the Crocker Range of Borneo, A. nuluensis has floral resources above 2,500 m to itself, whereas A. cerana and A. koschevnikovi compete with A. andreniformis and A. dorsata at lower elevations. In Sumatra, the opennesting species are not found above 1,000 m, whereas a cavity-nesting species, probably a mountain-adapted ecotype of A. cerana, is found above 2,000 m (Salmah, Inoue, and Sakagami 1990). Presumably these mountain species have special adaptations for cold that allow them to exploit resources free of competition from other honey bees. Despite some partitioning of floral resources based on species size and location, similarly sized A. florea and A. andreniformis compete directly throughout much of southeast Asia, and A. cerana and A. koschevnikovi compete in Borneo. The direct competition between the dwarf bees probably arises from their relatively recent range expansion such that their formerly allopatric ranges now overlap. The competitive interactions between A. koschevnikovi and A. cerana have not been studied, and perhaps there is more partitioning than first meets the eye.

Foraging Pheromones? Foraging-age honey bees produce a variety of species-specific pheromones in their mandibular glands. One of these, 2-heptanone, appears to have a defensive function (Chapter 8; Morse et al. 1967; Suwannapong 2000), and is used to alert nestmates to the presence of an intruder and to help defending bees find an intruder that has been bitten by nestmates. In high

Dance Communication and Foraging

amounts it repels foraging A. cerana (Naik et al. 1997), but whether it is used in this way in nature is not known. The pheromone 1-eicsoanol is present in high amounts in the mandibular glands of A. andreniformis, A. florea, A. dorsata, and A. cerana and is attractive to foraging bees (Suwannapong 2000). Thus 1-eicsoanol and some of the other mandibular pheromones may help recruits locate profitable flowers.

Summary The dance language of the honey bee was an important evolutionary step, allowing tropical species to efficiently exploit patchy resources in a tropical forest. The language involves a stylized reenactment of a successful forager’s flight to the forage patch. During the dance the dancing bee strides forward vigorously shaking her body and, in A. dorsata and the cavitynesting bees only, vibrating her wings. The duration of this waggle run of the dance is correlated with the distance to the goal. This allows researches to estimate the typical foraging ranges of a species. Such studies show that for all species, most bees forage less than 300 m from the nest. Nonetheless, some dances indicate flights of distances exceeding 10 km. The direction of the food goal is encoded in the angle of the waggle run relative to the current position of the sun or to straight up, depending on whether bees dance on a horizontal or vertical surface, respectively. A. florea dances on the horizontal plane of the crown of the nest. Her run points directly at the food source and is aligned by using the sun’s azimuth as a navigation point. In the giant bees and the cavity-nesting bees the dance has been transformed to the vertical plane such that the angle of the waggle run from the perpendicular indicates the angle of the flight direction from the sun’s current azimuth. Dances of these species are accompanied by sounds (wing vibrations) that help bees that follow the dancer in the dark. Honey bees live sympatrically but their variation in size means that their preferred forage patches and places are slightly different. These differences may lead to reduced interspecific competition. Foraging-age honey bees produce a variety of pheromones in their mandibular glands that are attractive to other bees. They may use these pheromones to help recruit nestmates to profitable forage patches.

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6 Reproduction, Swarming, and Migration

In honey bees, reproductive swarming is the process by which colonies divide to make offspring colonies. During reproductive swarming the old queen leaves her colony, accompanied by about half of the workers. The old queen and her entourage of workers found a new, offspring colony, while the original nest is inherited by a daughter queen, which takes over as the natal colony’s egg layer. Before reproductive swarming, the colony must rear new individuals: queens, workers, and drones. We describe how new individuals are reared and their development. Queens must mate, and we describe the behavior of young queens before and after their mating flights, and speculate about the evolution of the mating system. In addition to reproductive swarming, Asian honey bees often undergo migratory swarming in which the colony abandons its nest and moves on in search of more profitable pastures. These movements can be triggered by predation or parasitism, by a dearth of food, by a lack of shade, or, in the case of A. laboriosa, by seasonal cold. We describe how absconding is coordinated.

Rearing New Individuals The production of a new worker, drone, or queen begins when an egg is laid in a cell of the brood comb (Figure 6.1). In all species, the egg matures for 3 days, during which time the embryo develops within it. On the third day, the chorion is cast off by undulations of the larva within the egg,

Reproduction, Swarming, and Migration Table 6.1

Development time (days) of the brood of some honey bee species. Larval development means the time from egg hatch to cell capping, pupal development the time from cell capping to adult emergence. Data are unavailable for the other species. Worker

Queen

Drone

Species

Egg

Larva

Pupa

Egg

Larva

Pupa

Egg

Larva

Pupa

A. floreaa A. ceranab A. dorsatac A. melliferad

3 3 2.9 3

6.3 5 4.6 6

11.2 11 10.9 12

3 3 2 3

6.8 4–5.5 4.5 5

7.7 6–7.5 7 5

3 3 2.9 3

6.7 6 4.6 7

12.8 14 14.3 14

a. Sandhu and Singh (1960). b. Rahman (1945), Lap and Chinh (1996), Rosenkranz and Engels (1994a), Punchihewa (1994), Dung et al. (1993). c. Qayyum and Nabi (1968). d. Moritz and Southwick (1992).

and the tiny c-shaped larva is revealed. The larva grows rapidly for 5–6 days, undergoing five molts (Table 6.1; Snodgrass 1956). The mature larva stretches out in its cell, spins a cocoon, and metamorphoses into an adult, a process involving such a profound rearrangement of the body plan that the mature bee bears no resemblance to the wormlike larva that gave rise to it. This pupal stage lasts 7–14 days, depending on the caste and species (Table 6.1). In the cavity-nesting bees the larva’s brood cell is capped by the workers with an air-porous capping of wax just before the last larval instar. In A. dorsata (and presumably A. laboriosa) the cell is capped during the last larval instar after the larva is stretched out in its cell (Qayyum and Nabi 1968). Although there is similarity of development times across the genus (for those species for which data are available), there is great diversity among the castes, with queens maturing 35 percent faster than drones. The decreased development time is entirely due to the more rapid development of queen pupae (Table 6.1), which is presumably mediated by the mass provisioning of queen larvae with higher-quality food than is fed to workers or drones. One of the first things a newly emerged queen does is search the brood combs for any of her sister queens that may be present. During this patrol, queens of A. florea (Wongsiri et al. 1997), A. andreniformis, A. cerana, and A. koschevnikovi (Otis, Patton, and Tingek 1995) emit bursts of sound last-

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Figure 6.1 An egg (left) and a larva (right) developing in a comb of an A. cerana nest. Photo by M. Sasaki.

ing 2–4 sec. In A. cerana and A. koschevnikovi, these “tooting” calls have a fundamental frequency of about 2,700 Hz and may end with the queen beating her abdomen on the comb (Otis, Patton, and Tingek 1995). Tooting causes workers in the colony to freeze (Winston 1987; Wongsiri et al. 1997), and presumably signals the presence of the tooter to both workers and other queens. Queens of A. koschevnikovi and A. cerana that have yet to emerge from their cells may “quack” (the sound is readily distinguished) in reply to the “toots” of the emerged queen at a frequency of 2,290 to 2,890 Hz (Otis, Patton, and Tingek 1995). Otis et al. suggest that these auditory exchanges between confined and emerged queens may encourage the emerged queen to leave with an afterswarm should there be sufficient workers available for a second colony division. In A. mellifera and A. cerana, patrolling virgins search for other queens, and use auditory signals to indicate their presence. If a virgin queen meets another queen a fight ensues, and only one of them will be the victor (Figure 6.2). If not prevented from doing so by the workers, a virgin will tear a hole in the side of an unemerged queen cell and sting the occupant

Reproduction, Swarming, and Migration

Figure 6.2 A fight between two A. cerana queens. Photo by M. Sasaki.

(Wongsiri, Lai, and Sylvester 1990). This has been taken as evidence that natural selection strongly favors the first queen to emerge and a rapid development time (deGrandi-Hoffman et al. 1998). But we have sometimes seen half a dozen emerged A. florea virgins in the same nest, and have rarely seen queen cells destroyed by virgin queens (Wongsiri et al. 1997). Nonetheless, queens of all species, including A. florea, have a short development time. Perhaps this suggests that the compelling reason for the short development time of queens is not the threat of being killed by a slightly older sister, but colony-level selection for rapid queen replacement, for a colony without an egg layer is severely disadvantaged by the cessation of brood rearing. Drones, by contrast, have a leisurely development, taking about 15 percent longer than workers (Table 6.1). Clearly this is not because of cooler conditions on the periphery of the brood nest, because fast-developing queens are also raised there, and because the development time of A. dorsata drones, reared in cells commingled with their sister workers, nevertheless have a slower development time than workers.

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Reproductive Swarming Tropical Asian honey bee colonies usually begin their growth trajectory from a cluster of broodless and combless adult bees that has arrived at a nesting site as a migratory or reproductive swarm. This differs from temperate A. mellifera and A. cerana, in which colonies are more sedentary and usually begin their growth from an overwintered cluster of adults that has considerable preexisting infrastructure built by their sisters in the previous season: there will be a set of brood and honey combs and probably the last of the food stores for the growing colony to utilize. A. dorsata colonies are always established from scratch, the arriving swarms ignoring the valuable wax of the past season’s colonies, even if the arriving swarm nests just centimeters away from an old comb (Woyke, Wilde, and Wilde 2000). A. florea also never directly reuse a comb, but may plunder wax from abandoned nests (Akratanakul 1977). Ignoring existing comb contrasts strongly with the behavior of A. mellifera swarms, which are positively attracted to cavities that contain abandoned combs, which they utilize willingly (Schmidt 2001). Presumably there is a tradeoff between the value of the wax and the risk of contracting diseases or parasites from it. Combs may be more valuable to cavity-nesting species, because there is much more comb per bee than in the open-nesting species. The behavior of reproductive swarms of the Asian honey bees after they have left the natal nest is not well studied, whereas we know quite a lot about the behaviour of swarms of A. mellifera (see Lindauer 1967, Seeley 1985, and Winston 1987 for reviews). Whether or not the behavior of reproductive swarms of all Asian species will turn out to be precisely the same as that of A. mellifera remains to be seen, but there are several clues suggesting that the process is quite similar. In A. mellifera a reproductive swarm clusters a few tens of meters from its natal colony. In less than an hour, scout bees begin searching for suitable cavities in which to construct the swarm’s new home (Seeley and Buhrman 1999). Successful scouts report the location of suitable nesting cavities to other bees by performing communication dances on the surface of the swarm cluster in much the same way as they would for food sources (Lindauer 1967). Each scout dances for a particular nest site a finite number of times, and can recruit other scouts to its discovery (Camazine et al.

Reproduction, Swarming, and Migration

1999; Seeley and Buhrman 1999). The duration of the dance is positively correlated with the scout’s perception of nest site quality (Seeley and Buhrman 1999). With each visit to the site, the number of circuits declines, but this decline is slower for good sites than for poor ones (Seeley and Buhrman 1999). Although nest scouts do not directly compare alternative sites to evaluate their respective qualities (Seeley 2003), an emergent property of scouts dancing more enthusiastically for quality sites than for poor ones is that the number of scouts visiting and dancing for the best nest site builds exponentially while the number visiting less suitable sites exponentially declines (Myerscough 2003; Seeley 2003). This process may take several days, but when consensus is reached among the scouts, the swarm promptly departs for the new home, led by the scouts. The scouts apparently guide the swarm by flying quickly through the slower-moving swarm, returning to the rear after flying through it (Lindauer 1955; Janson, Middendorf, and Beekman 2005). By visually following their neighbors and the scouts, the swarm moves toward the target. Avitabile et al. (1975) speculated that the scouts guide the swarm by pheromonal attraction, but by gluing shut the Nasanov gland of every bee in three swarms, Beekman, Fathke, and Seeley (2005) showed that visual, rather than pheromonal, attraction is more likely. It seems parsimonious that the process of nest site selection is similar for the Asian species. For example, we have seen A. cerana reproductive swarms settle 20–30 m away from the natal nest, stay for a few days, and then depart for a new nest site, just as A. mellifera would (Figure 6.3). We have also seen dances indicating a nest site in artificial swarms of A. florea. But as yet there is only circumstantial evidence that swarms of the opennesting species utilize an interim cluster and the competitive nest site selection process that characterizes house hunting in A. mellifera swarms (Seeley 2003; Seeley and Buhrman 1999). Akratanakul (1977) observed the departure of a reproductive swarm of A. florea, which settled about 20 m from its natal colony “in a shady tamarind tree.” No further observations were made on this swarm, so we do not know if this was an interim cluster (as Akratanakul assumed), or if the swarm stayed there permanently. Akratanakul also observed a number of combless clusters of A. florea that were probably absconding swarms. He observed communication dances on the surface of these swarms, and assumed that these were part of a nest site selection process. In 2004 we es-

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Figure 6.3 A swarm of A. cerana in Japan. Photo by M. Sasaki.

tablished three combless swarms of A. florea in Bangkok by shaking the workers from the comb near their queen, which we had tied, in a cage, on a tree. After 12 hours we released the queen and observed the departures of the swarms. In each case departure occurred at about 11:00 after directionally coordinated dances were observed. This may suggest that A. florea select a nest site from their temporary swarm in a way similar to that of A. mellifera. Presumably scouts mark a suitable branch with a pheromone and recruit other scouts to it. But the possibility remains open that A. florea reproductive swarms do not form an interim cluster, and move directly to their new nest site, which is selected via competitive dancing before swarm lift-off. They may only use a temporary cluster when the adults abscond from the nest after a disturbance. A. dorsata colonies that are ready to cast a reproductive swarm are characterized by a “cone” of young bees hanging down from the lower edge of the nest (Woyke, Wilde, and Wilde 2000). Colony reproduction appears to be of two kinds (Lindauer 1956). The first mechanism appears to be similar to that in A. mellifera and is described by Woyke et al. (2000). The cur-

Reproduction, Swarming, and Migration

tain becomes disordered and a large number of bees suddenly become airborne with a loud “wuuum.” These airborne bees may be aggressive. The airborne swarm is about 20 m in diameter. It is not known whether these swarms form a temporary combless cluster or whether they move directly to a new nest site. The distance that these reproductive (as opposed to migratory) swarms generally move is also unknown, but at least one such swarm settled more than 500 m from the natal nest. Lindauer (1956) and Woyke et al. (2000) suggest that the swarms fly “directly to a previously selected place.” We could find no reports of communication dances being performed on the surface of these reproductive swarms. In the second mechanism, known as “budding,” a group of workers gradually separate from the nest, and form a new colony about 1 m from the old (Lindauer 1956). Budding, if it is common, would tend to lead to the establishment of mother and daughter colonies along a branch. But genetic investigations (Oldroyd, Osborne, and Mardan 2000; Paar et al. 2004a) have shown that adjacent colonies are not related as mother and daughter, which suggests that the budding observed by Lindauer in India may be quite rare.

Migration and Absconding Not all swarming is related to reproduction. Asian honey bees do not usually store great amounts of honey. Their strategy is to put their biological surplus into reproductive swarms rather than storing honey that may be stolen at any time by their many predators. This means that with their small honey stores they are vulnerable to starvation if there is a prolonged shortage of nectar and pollen. For this reason, the response of most Asian bees to declining resources is to abscond from the existing nest and migrate to areas where forage is more abundant. Even the tropical ecotypes of cavity-nesting A. cerana are much more mobile than temperate A. mellifera. In tropical areas A. cerana readily absconds in the face of food shortage or disturbance (Punchihewa 1994; Sasaki 1990; Woyke 1976). At least in tropical areas, A. cerana can exist as mobile clusters of broodless and combless adults for several weeks at a time (personal observations). Absconding is presaged by cessation of pollen collection and brood rearing (Punchihewa 1994). After the last brood has emerged, the adult bees fill their honey stomachs, and swarm off to eventually establish a new

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nest at a new location, abandoning the comb and any remaining honey and pollen. In most cases, a colony of open-nesting bees will migrate at least twice a year. Cavity-nesting bees are also prone to absconding (Nakamura, Wongsiri, and Sasaki 1991; Punchihewa 1994), but perhaps not quite to the same degree.

The Remarkable Migrations of the Giant Bees The most distinctly migratory species are the giant bees. A common seasonal pattern of A. dorsata at an established communal nest site is the arrival of swarms at the end of the wet season, the building of combs, a period of rapid colony growth, and the production of reproductive swarms by the stronger colonies (Itioka et al. 2001; Kahona, Nakamura, and Amir 1999). In many mountainous areas the availability of pollen declines toward the end of the dry season (Aggarwal and Kapil 1989; Thapa et al. 2000; Wongsiri et al. 1996). In response to this, brood rearing is curtailed, and the adult bee populations begin to decline. Combs may be attacked by wax moths and other predators (see Chapter 9), and the populations of the parasitic mites Tropilaelaps clareae and T. koenigerum may be high. At that time most colonies will abscond from the site and begin a migration to a different locality, often to a plains area, where new nests are established for the wet season (Koeniger and Koeniger 1980). Often, though, a few colonies will remain at the dry-season site. The distance traveled by these migrating swarms is not known but can be inferred from the fact that certain areas are utilized at different times of the year (Ahmed 1989; Dyer and Seeley 1994; Itioka et al. 2001; Kahona, Nakamura, and Amir 1999; Koeniger and Koeniger 1980; Pandey 1974; Sattigi and Kulkarni 2001a, 2001b; Sheikh and Chetry 2000; Thapa et al. 2000; Underwood 1990a; Venkatesh and Reddy 1989), which suggests that colonies move between the different areas. Furthermore, populations of A. dorsata are genetically homogeneous over a broad range (Paar et al. 2004a), which indicates genetic mixing via migration. Koeniger and Koeniger (1980) postulated that migrating A. dorsata colonies in Sri Lanka may travel up to 100 km, and it is possible that migrations are much longer. In Nepal, A. laboriosa appears to divide its year between the warm zone from 1,200 m to 2,000 m in winter, where it often exists in combless

Reproduction, Swarming, and Migration

clusters, and the cold zone, above 2,000 m, in summer (Roubik, Sakagami, and Kudo 1985; Underwood 1990a; Woyke, Wilde, and Wilde 2001b). Migrations probably take several days or weeks. Along the route the swarm clusters in trees overnight. Workers forage for food from the bivouacked swarm (Koeniger and Koeniger 1980), and presumably the swarm can move only when it has sufficient energy reserves. Quiescent, broodless swarms are frugal in their energy needs, and can survive with minimal supplies for long periods, perhaps months (Underwood 1990a). Swarm movements are presaged by waggle dances by scout bees that indicate the direction of intended movement (Dyer and Seeley 1991a; 1994; Koeniger and Koeniger 1980). The waggle runs of these dances are extremely prolonged, which suggests that the distances indicated are very great. We do not, however, know whether the dances indicate the distance to be traveled for the entire migration, for just the day of travel, or for no specific distance. Nor is the typical distance traveled per day known. In one case a migrating swarm probably traveled 5 km during a 1-hour flight, but swarms can travel at about 20 km per hour (Koeniger and Koeniger 1980). Two independent studies in Borneo (Neumann et al. 2000) and Assam, India (Paar, Oldroyd, and Kastberger 2000), showed that giant honey bees return to their natal nest site (the same building or tree, but not the same comb) after seasonal absence, and in one case (Paar, Oldroyd, and Kastberger 2000) after two seasons of absence! These studies used microsatellite DNA fingerprinting to show that the queen of the returned colony was the same as the queen of the departed colony, and that the chance of a random queen having the same genetic match is impossibly low (much less than 1 in a million). This remarkable observation raises two fascinating questions: how do they do it and why do they do it? Consider the problem: A colony has been absent for 6 months (or 18 months), has built a comb at another site, probably many tens of kilometers away. It seems unlikely that workers live for more than 3 months (Chinh, Tan, and Thai 2004; Otis, Mardan, and McGee 1990), so the only individual that could have lived at or even visited the old nest site is the queen herself. This might suggest that the queen guides the swarm to her old nesting place, but this seems extremely unlikely. First, Woyke et al. (2000) have shown that a small number of workers visit the old nest site 1 or 2 days before the arrival of the main swarm and the queen. If it is the queen that guides the swarm, how do these workers find the correct site?

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Second, if it is indeed the queen that guides the swarm, what is the purpose of the “migratory dances” (Koeniger and Koeniger 1980) performed by workers on bivouacked swarms that indicate the direction of travel? We suspect that, rather than any individuals actually remembering the location of the old nest site and guiding their nestmates to it, traveling swarms follow a set of standard rules that often result in a migration trajectory that often ends up at the old nest site. Consider the simplest case of a migration up a mountain at the onset of the dry season. We speculate that colonies respond to increasing temperatures by moving to higher elevations or traveling in some environmentally triggered direction (Thapa et al. 2000). As with the typical flights of food foragers (Lindauer 1967) or drones seeking a drone congregation area (Loper, Wolf, and Taylor 1992), the path traveled by the swarm probably follows conspicuous features of the landscape such as a watercourse, road, or forest margin. Near the old nest site, the traveling swarm responds to the same signals as it did in the previous year—the air is cooler, the forage abundant— indicating a good area to settle and build a comb. The final nesting location may be beacon-like to the traveling swarm: a tall emergent tree, a cliff, a water tower, and the presence of other colonies may be attractive. A. dorsata seem to have preferences for the height (Reddy and Reddy 1989; Sattigi and Kulkarni 2001c; Starr, Schmidt, and Schmidt 1987) and aspect (Reddy and Reddy 1993) of the object upon which they build combs. Combined, these factors may all add up to the strong likelihood that a swarm will chose the same site as it did in the previous year—even though a forest or town viewed by human eyes is replete with seemingly suitable nesting places. The fidelity can be quite extraordinary. For example, a window eave we have periodically studied at the District Health Care Center in the city of Chiang Mai is occupied by an A. dorsata colony (the same one perhaps?) year after year, possibly for as many as 20 years. Furthermore, trees used for nesting and trees used for the establishment of DCAs have similar features: they emerge strikingly from the surrounding landscape. N. and G. Koeniger were actually able to predict sites of DCAs by searching for tall emergent trees from a high vantage point (N. Koeniger personal communication). Following the same rules as other traveling swarms may also lead to the strong tendency for A. dorsata colonies to be highly aggregated—indeed, many animal aggregations are thought to arise in this way (Parrish and

Reproduction, Swarming, and Migration

Edelstein-Keshet 1999). It may also mean that aggregations tend to move together as a group between two seasonally utilized nest sites. Paar et al. (2004a) showed that in the A. dorsata of Assam, India, although there is significant genetic differentiation among aggregation sites, there is still sufficient gene flow among aggregations to prevent inbreeding and significant genetic partitioning of the population. The existence of the communication dances and the arrival of bees at the final nesting place a day or so before the arrival of the main swarm indicates that scouts travel ahead of the main swarm, choosing the approximate location for the bivouac and the location of the final nest site, and guiding the swarm along the way. If so, this behavior would additionally increase the probability of swarms reusing their former nest sites. Scouts are attracted to Nasanov pheromones, secretions of the Nasanov gland, which lies between the last and second-to-last segment of honey bee workers (Snodgrass 1956). Workers use Nasanov pheromones to attract flying nestmates to join a cluster (Free 1987; Free et al. 1981). Artificial lures made of synthetic Nasanov secretions are highly attractive to scout bees, and if present, greatly increase the probability that a nest box will attract a swarm of A. mellifera (Schmidt, Slessor, and Winston 1993). Thus scout workers may be drawn toward existing colonies because of the presence of Nasanov pheromones (Oldroyd, Osborne, and Mardan 2000; Oldroyd, Smolenski, Lawler, et al. 1995). How else might A. dorsata swarms be attracted back to their old nest sites? One possibility is that they are attracted (at least over short distances) by the smell of the remains of their old comb (Paar, Oldroyd, and Kastberger 2000). Wax combs contain chemical signatures that honey bees can use as cues for nestmate recognition (Breed et al. 1995; Breed et al. 1998; Breed, Williams, and Fewell 1988), and presumably these same cues could be used by A. dorsata to recognize their old comb. But because A. dorsata do not reuse their old comb (Woyke, Wilde, and Wilde 2000), it seems unlikely that they use some chemical signature of their wax to relocate their nest sites. This hypothesis could be easily tested by manipulating the location of some abandoned combs to see if returning swarms are attracted to location or comb scent. A second possibility is that the assumption, based on Chinh et al. (2004) and Otis et al. (1990), that A. dorsata workers live for only 3 months is incorrect. Workers of the open-nesting species spend large amounts of time

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hanging quietly in the curtains of bees that protect their nest, and because of this a small proportion of workers may live far longer than 3 months. (One could even imagine a special “memory” guild of workers that do no work but retain knowledge of previous nest sites on behalf of the colony). Measuring the longevity of A. dorsata workers by releasing several thousand marked, day-old workers into a colony before a migration event would seem a useful way of testing the hypothesis that some workers live long enough that they could possibily guide their swarm back to their home of the previous season. A third, and probably fanciful, possibility is that queens perform a communication dance to direct scout workers to the former nest site. There does not appear to be any physical impediment to a queen dancing, and indeed A. andreniformis drones have been observed performing round dances (Rinderer et al. 1992), which shows that the behavior is not confined to workers alone. Honey bees have extraordinary capacities to remember geographical locations (Lindauer 1963), and so queen guidance is not inconceivable. But despite many days of observing migratory swarms of A. dorsata, Koeniger and Koeniger (1980) did not observe queens dancing. There remains a lot to understand about the migrations of A. dorsata. Abundant evidence suggests that colonies track the best available forage, with the number of colonies present rapidly increasing whenever there is abundant nectar (Itioka et al. 2001; Pandey 1974; Tan et al. 1997). How do colonies know that a distant area is productive? Is it that there are always swarms on the move, which settle only when forage conditions are favorable? Can swarms smell the perfume of flowers over great distances? Or do colonies maintain a group of very long distance (>20 km) scouts that really do have the ability to guide their swarm to a productive area? With the right questions, much could be learned through simple observations and some paint marks.

The Migrations of Dwarf and Cavity-nesting Bees Colonies of the dwarf bees are almost as peripatetic as those of the giant bees, with most nests undergoing at least one migration per year (Akra-

Reproduction, Swarming, and Migration

tanakul 1977; Nakamura, Wongsiri, and Sasaki 1991; Wongsiri et al. 1997). We do not yet know if A. florea colonies utilize the same nest site year after year as A. dorsata do, but we do know of some sites that are repeatedly used by A. florea (for example, a mango tree in Wongsiri’s garden in Bangkok has been seasonally occupied for over 10 years). A. florea migrations apparently track abundant forage as do those of A. dorsata. For example, Soman and Chawda (1996) report a migration of A. florea colonies into an area from which they had been absent for 10 years in order to utilize an unusual honey flow. In the Middle East, A. florea colonies migrate so that they have a warm aspect in winter and a shaded one in summer (Dutton and Simpson 1977). There is a definite tendency for dwarf bee colonies to be aggregated (Rinderer et al. 2002). A. florea and A. andreniformis are prone to absconding after disturbance (personal observations) or loss of shade (Seeley, Seeley, and Akratanakul 1982). Unless the comb is lost during an attack by a predator, absconding is an organized event over 10 days—long enough to let sealed brood emerge and to curtail the production of new larvae. In A. florea at least, the new nest site is chosen before the colony abandons the original nest, since scout bees dance for the new nest site and orchestrate the departure for it (Seeley, Seeley, and Akratanakul 1982). Combless A. cerana swarms are commonly found, and the species is prone to absconding, especially when disturbed by humans or ants, and during nectar dearths (Nakamura, Wongsiri, and Sasaki 1991; Punchihewa 1994). Absconding in A. cerana is presaged by the cessation of all activity. Some workers perform an “absconding dance,” a series of long (up to 80 sec) wag-tail runs with no return phase that appear to motivate the colony to abscond (Sasaki 1990). Waggle runs of this duration indicate great distances—far beyond the normal foraging range of A. cerana (Sasaki 1990). The angles of the waggle run are directional in that they are of a consistent sun-compensated angle for each dance by an individual bee. But angles are different between different bees, so the angles indicated are probably of no significance (Sasaki 1990). The predictable migrations of cavity-nesting bees like A. cerana and A. koschevnikovi are often exploited by local beekeepers, who deploy trap hives at the base of mountains at the end of the dry season. As migrating swarms return to the plains they often nest in the hives placed by bee-

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keepers. The beekeepers may then move the trapped swarms to their coconut plantations for the honey harvest (Nakamura, Wongsiri, and Sasaki 1991).

The Production of Queens In all species, about 10 days before reproductive swarming, the workers construct 5–20 primordial queen cells (shallow waxen cups) on the margin of the brood nest. If conditions for swarming remain amenable—if there are abundant resources available—the queen will lay fertilized eggs, one in each cell cup (see, e.g., Qayyum and Nabi 1968), which the workers then rear to be queens by feeding the larvae a diet that is rich in quality and abundance (Beetsma 1979). A few days before the young queens are due to emerge from their cells, the old queen and about half the workers take to the air, and the swarm departs (Figure 6.4). Young queens may also be reared if the colony becomes queenless. Emergency queen cells (5–20) are constructed around young larvae, usually in the central area of the comb, in contrast to swarm cells, which are constructed around the periphery of the nest. Colony-level response to dequeening is extremely variable. In some A. florea colonies behavior changes very little, and the workers busy themselves rearing queens. Other colonies become highly agitated. Within a few hours of dequeening many workers leave the comb and cluster on adjacent branches, and do not produce queen cells but gradually disperse over a few days. This is almost always the response of A. andreniformis (personal observations; Koeniger et al. 2000). After emerging from their cells, virgin queens patrol the comb surface. When they are 3 days old the young queens become more adventurous. In A. florea colonies virgin queens will regularly be seen striding across the crown of the comb, often eliciting a strange dorsoventral shaking from the workers she comes in contact with (Koeniger, Koeniger, and Wongsiri 1989; Oldroyd et al. 1994b). Virgins 6 days old are often seen running across the crown, while pausing occasionally to clean their eyes and mouthparts. Brief (2 min) orientation and cleansing flights may occur in 6-dayold queens. Full mating flights of 15–30 min duration occur when queens

Reproduction, Swarming, and Migration

105

Figure 6.4 An A. florea nest that has recently swarmed. Mature queen cells are visible on the bottom right of the comb. Photo by B. Oldroyd.

are 6–8 days old (A. florea, Koeniger, Koeniger, and Wongsiri 1989; A. dorsata, Tan et al. 1999; A. cerana, Ruttner, Woyke, and Koeniger 1972, 1973; Woyke 1975).

Mating Mating occurs on the wing at drone congregation areas (DCAs) (Chapter 4). Drone congregations can sometimes be detected by listening for the buzz of the circling drones (provided one is not too distracted by mosquitoes) in likely places near tall emergent trees. The presence of the DCA is best confirmed by raising a drone trap into the area of the suspected DCA with the aid of a longish pole, helium balloon, or a length of fishing line looped over a tall tree. A drone trap consists of a lure baited with the primary constituent of A. mellifera queen pheromone, 9-keto-(E)2-decenoic acid (9-ODA) (Callow and Johnson 1960), or a virgin queen, and some device to capture drones such as flypaper (Fujiwara et al. 1994; Koeniger et al.

5.4 ± 0.5 0.5 ± 0.02 0.14 ± 0.01 n.d. 0.21 ± 0.03 0.16 ± 0.03 n.d.

10.3 ± 3.6 0.8 103 ± 41 28.8 ± 6.5 110 ± 4 116 ± 7 231 ± 17

0.3 ± 0.01 1.4 34.0 ± 0.0 18.1 ± 11.2 61.5 ± 3.1 106 ± 8 164 ±15

Queen

Worker 1.5 ± 0.3 2.9 ± 0.3 1.9 ± 0.03 0.9 ± 0.1 1.0 ± 0.2 0.75 ± 0.2 1.4 ± 0.2

9-HDA

0.2 ± 0.01 8.1 0.4 ± 0.2 1.8 ± 0.07 32.7 ± 3.0 23.1 ± 2.4 27.3 ± 2.4

Queen

3.2 ± 0.05 11.0 ± 1.0 56 ± 5 0.8 ± 0.2 3.5 ± 0.8 9.4 ± 1.2 95 ± 15

Worker

10-HDA

0.5 11.9 0.3 0.7 0.8 1.1 0.7

Queen

0.9 27.8 773.6 Very high 21.4 63.4 Very high

Worker

HDA/ODA ratio

Note: 9-ODA: 9-keto-(E)2-decenoic acid; 9-HDA: 9-hydroxy-(E)2-decenoic acid; 10-HDA: 10-hydroxy-(E)2-decenoic acid. In the cavity-nesting species and A. dorsata the ratio of 10-HDA to 9-ODA is very high in workers relative to the same ratio in queens, because 10-HDA is used in brood food as both a nutrient and a preservative. In contrast, the amount of 10-HDA present in the mandibular secretions of queens of the dwarf bees is high, particularly in A. florea, whereas the amount of 9-ODA is relatively low. This may suggest that 10-HDA is more important as a queen signal in A. florea than in the other species. Where no standard error is given, only one individual was analyzed.

Worker

Queen

9-ODA

Major components ( μg ± s.e.) of mandibular glands of mated queens and workers for some honey bee species (n.d. = not determined).

A. andreniformis (Plettner et al. 1997) A. florea (Plettner et al. 1997) A. dorsata (Plettner et al. 1997) A. cerana (Plettner et al. 1997) A. cerana (Keeling et al. 2001) A. nigrocincta (Keeling et al. 2001) A. mellifera (Plettner et al. 1997)

Species

Table 6.2

Reproduction, Swarming, and Migration

1994). (The drone traps suitable for A. mellifera [Taylor 1984; Williams 1987] are ineffective for catching drones of Asian species [Koeniger et al. 1994].) Drones of most (and probably all) species are attracted to 9-ODA. If they are present they are caught on the flypaper, and can then be examined to determine their species. Such traps are particularly useful for determining the precise location of each species’ DCA, since the number of drones caught increases the closer the trap is to the center of the DCA (Koeniger and Koeniger 2000b). We assume that on her mating flight the young queen flies to a drone congregation area. Drones are attracted to flying queens by a combination of sex pheromones that are secreted from the queen’s mandibular glands (Gary 1962) and the visual cue of a fast-moving, small, elongated dark object about 3 cm long (Strang 1970). The sex attractant, 9-ODA, is apparently conserved across the genus (Plettner et al. 1997; Shearer et al. 1976), which suggests that drones of all species are attracted to it (Butler, Calam, and Callow 1967). But the amount of 9-ODA found in the mandibular glands of queens of the dwarf species is quite low (Table 6.2; Plettner et al. 1997), and this may suggest that 9-ODA is not the primary sex attractant of A. andreniformis and A. florea. Further, males of the dwarf species have far fewer antennal receptors for 9-ODA than do the other species (Brockmann and Brückner 2001). Perhaps reduced attraction to 9-ODA is the reason that we do not yet know the location of the DCAs of the dwarf species: their males are not as strongly attracted to 9-ODA as are those of the other species. Surprisingly, mating seems to be a fairly orderly affair, with no obvious signs of contest among the males (Koeniger 1990). As the queen flies through the circling drones she is followed by a “comet” of about 20–100 interested males (Gries and Koeniger 1996; Koeniger 1990). A male flies up behind the queen, grasps her thorax with his front legs (causing her to loose height precipitously) and within seconds everts his penis into the queen’s sting chamber. This process instantly paralyses him, and he falls backward away from the queen. The next male then approaches and mates. The number of males that the queen mates with on one flight seems to be highly variable, but in A. dorsata it seems probable that she mates with at least 20 males on one flight (Wattanachaiyingcharoen et al. 2003), and may take 3–5 mating flights on successive evenings (Tan et al. 1999), which leads to extraordinarily high mating frequencies in some individuals (Wattanachaiyingcharoen et al. 2003).

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After her mating flight, the queen returns to her nest. Much of the sperm placed in her by the males is expelled during her return flight and during the night (Koeniger and Koeniger 1991), with the remainder being transferred to the spermatheca. That the queens mate with many more males than they apparently need is significant, because mating is associated with at least some risks (Moritz 1985; Palmer and Oldroyd 2000). We will discuss hypotheses for the evolution of multiple mating in detail in the next chapter.

Preventing Interspecific Hybrids Queens of all species are of similar size and shape, fly quickly, and secrete the same sex attractant, 9-ODA (though the amount is relatively low in the dwarf species). In many areas there are up to four sympatric species (Chapter 2), and drones of several species will attempt to copulate with the same queenlike lure (Koeniger and Koeniger 2000b). What mechanisms, then, act to prevent interspecific matings, for it seems likely that at least at a distance, a queen of any species would be attractive to a male of any other? Indeed, interspecific matings between A. cerana and A. mellifera have been recorded in Germany when A. cerana was imported there (Ruttner and Maul 1983). Koeniger and Koeniger (2000b) concluded that prezygotic barriers to interspecific hybridization are of three broad types: behavioral, copulatory, and fertilization. The behavioral barriers could include the time, place, and season of mating. Copulatory barriers could include variation in the cocktail of chemicals in the queen pheromone and the morphology of the genitalia. Fertilization barriers could include a failure of sperm transfer from the male to the spermatheca or death of the sperm within the spermatheca. We will consider each of these in turn.

Separation of Mating in Time and Space Honey bee swarms have limited food reserves and must have access to floral resources to survive. In order to build to swarming strength, a colony needs access to copious nectar and pollen resources over a protracted period. Because all colonies in a district experience the same environmental

Reproduction, Swarming, and Migration

40 A. andreniformis ?? A. dorsata

Height (m)

30

20 A. cerana 10

A. koschevnikovi

Figure 6.5 Drone congregation areas of three species in Tenom, Borneo. The location of the drone congregation areas of the dwarf bees is unknown. Redrawn from Koeniger and Koeniger (2000a).

conditions (including possible cues from day length, lunar month, or precipitation), there is usually a swarming season (Das and Rahman 2000; Lekprayoon and Wongsiri 1989; Lindauer 1956; Mahindre 2000; McLellan and Rowland 1986; Mossadegh 1990; Venkatesh and Reddy 1989; Woyke, Wilde, and Wilde 2000), in which most colonies of sufficient size will swarm, and drone production will peak, thus providing plenty of mates for the young queens. This means that the mating period is not well separated by season among the different species of Asian Apis. In Chapter 4 we described how, at any one location, the various species have specific times and places for mating. For example, at Tenom in Borneo (Figure 6.5), A. dorsata form their DCAs just after dusk under the spreading limbs of tall trees that emerge high above the main canopy of the forest (Koeniger, Koeniger, Keiltu, and Marden 1994). A. koschevnikovi males aggregate between 16:45 and 18:30 hrs (Koeniger, Koeniger, and Tingek 1994b)

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beneath the canopy of forest trees of average height (Koeniger, Tingek, et al. 1998), while A. cerana drones aggregate beside tree canopies (Koeniger and Koeniger 2000) between 14:00 and 16:15 hrs (Koeniger, Koeniger, and Tingek 1994a; Koeniger, Koeniger, et al. 1998). A. andreniformis drones aggregate in an unknown location just after noon (Koeniger et al. 2000) These differences in mating time and place must serve as an important barrier to interspecific matings (Chapter 4), and thus act as a mechanism to reinforce processes of speciation at Tenom. Although there is some overlap of the drone flight times, queen flights are more constrained (Koeniger, Koeniger, and Tingek 1994), so that it is extremely unlikely that a queen will meet a drone of another species during a mating flight. Interestingly, however, mating times and place are plastic, varying from locality to locality (Koeniger and Koeniger 2000a, 2000b; Otis et al. 2000). So, for example, in Sri Lanka A. cerana males aggregate beside a tree canopy (Punchihewa, Koeniger, and Koeniger 1990), whereas in Japan they congregate above a tree (Fujiwara et al. 1994; Yoshida 1994). Japanese A. cerana mate later in the afternoon than the A. cerana of Sulawesi, but earlier than those of Chanthaburi, Thailand, or Sri Lanka (Figure 6.6). In Germany, transplanted A. cerana males from Pakistan congregated in an open field, shunning the cover of the European trees (Ruttner and Ruttner 1966). Some of these differences are unsurprising, for the forests of Borneo and northern Japan are of a different nature, and drones must adapt to the local cues for DCA establishment as best they can. Nonetheless, the most significant factor that seems to affect the range of mating time is the presence of drones of other species (Figure 6.6). As the number of species present increases, the span of the mating time is decreased (Otis et al. 2000), which suggests that the presence of the drones of other species causes selection for premating isolation via temporal separation. On the other hand, in the absence of other species, the times of mating flights becomes broadened and tends to shift towards midafternoon (Figure 6.6). The most extreme cases would appear to be the shift of A. laboriosa from dusk mating to afternoon mating (Underwood 1990b), and A. nuluensis to the late morning (Koeniger et al. 1996b). Koeniger and Koeniger (2000b) point out that wherever honey bee species co-occur, the smaller species mate first (Figure 6.6). Koeniger and Koeniger have an intriguing hypothesis for this polarity, proposing that honey bee drones are cued to pursue mates that are larger than themselves, because this is the most efficient way for drones to distinguish queens from

Reproduction, Swarming, and Migration

1 Thailand 0 1

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aaaaaaaaaaaaaaaaaaaaaaa aaaaaaaaaaaaaaaaaaaaaaa aaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaa a a a a a a a a a a a a aaaaaaaaaaaaaaaaaaaaaaa aaaaaaaaaaaaaaaaaaaaaaa aaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaa a a a a a a a a a a a a

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Figure 6.6 The time of drone flight of Asian honey bees in various localities. Redrawn from Otis et al. (2000) from data in Hadisoesilo and Otis (1996), Koeniger et al. (1994, 1996b, 1988), Koeniger and Wijayagunasekera (1976), Rinderer et al. (1993) and Yoshida et al. (1994).

drones at the DCA. To avoid attempted matings with males of the larger species, there is strong selection for the smaller species to mate earlier than the larger species. (The reverse is not the case because the males of the smaller species are smaller than the males of the larger species.) On the basis of this hypothesis Koeniger and Koeniger (1990) predict that the dwarf A. florea, which has now colonized Sudan (Lord and Nagi 1987; Mogga

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and Ruttner 1988), will be selected to shift its mating time to before that of the indigenous A. mellifera—possibly before noon.

Barriers to Successful Copulation The genitalia of the male honey bee are extraordinary in their size, morphology, and great variety (Figure 6.7). The honey bee endophallus is very large and membranous, whereas the sclerotized portion is much reduced and simplified (Michener 2000; Ruttner 1988). Highly variable male genitalia across a small biological group such as honey bees is suggestive of sexual selection (Panhuis et al. 2001). Since variable male genitalia need to be accommodated by matching female genitalia in a “lock and key” mechanism, sexual selection on male genitalia can rapidly result in an insurmountable barrier to hybridization, thus playing an important role in speciation. Examination of Figure 6.7 suggests that matings between subgenera would be impossible, owing to the widely divergent penis morphology, but possible within subgenera. This may suggest that sexual selection has played a role in shaping the male genitalia of Apis, but the relatively conserved female sting chamber casts doubt on this view. The endophallus, as its name would suggest, is retained within the abdomen of the male, with no hint of its presence being shown externally. The three main identifiable regions are the vestibulum at the base, the bulbus, which is inserted into the queen, and the narrower cervix, which joins the two (Figure 6.7; Koeniger et al. 1991; Paliwal and Tembhare 2001; Simpson 1970; Snodgrass 1956). Upon mating, the various sections of the endophallus are successively (but very quickly) everted from the abdomen under pressure of hemolymph, mucus, and air forced into the organ by muscular contractions (Snodgrass 1956). In all species the process of copulation begins with the partial eversion of the vestibulum, which makes contact with the queen’s open sting chamber. The cornua then emerge, making contact with the queen and guiding the expanded bulbus into the queen’s sting chamber. The dorsal cornua enter the queen’s sting chamber and hold the queen’s sting as the paralyzed male falls backward away from her. Then the bulbus is extended, followed by ejaculation of the semen onto the tip of the bulbus deep within the queen’s genital tract, and deposit of the semen in the median and lateral oviducts (Koeniger 1984). The drone then falls away from the queen, and the cornua expand further (but

Reproduction, Swarming, and Migration

Dwarf bees

Cavity-nesting bees

A. florea

A. cerana A. nuluensis A. nigrocincta

A. andreniformis

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A. koschevnikovi bulbus notched lobe

cervix A. dorsata A. laboriosa

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Figure 6.7 The everted penises of the genus Apis. Redrawn from Koeniger et al. (1991).

not completely, as they do when the drone is squeezed between the fingers), pushing the male out of the queen. Each copulation takes but a few seconds (Koeniger 1990). In the cavity-nesting species the drone and the queen are held together in copula by the large bulbus (which tightly fills the queen’s sting chamber) and its protruding, notched, lobe (Figure 6.7), the dorsal cornua inside the queen. In the giant bees, the dorsal cornua have two lobes, a shorter one that enters the queen as in the cavity-nesting species, and a much longer one, absent in the cavity-nesting species, that may grasp her externally (Woyke 2000), or be used to push the drone out of the queen after eversion (G. Koeniger personal communication). The giant bees also have

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Figure 6.8 Woyke’s (2000) postulated position of the A. dorsata queen and drone during early (left) and late (right) copulation. Redrawn from J. Woyke, “Eversion of the drone endophallus and the probable mating process in Apis dorsata,” in Seventh International Conference on Tropical Bees: Management and Diversity, and Fifth Asian Apicultural Association Conference, Chiang Mai, Thaland, ed. S. Wongsiri, pp. 189–194; copyright 2000 International Bee Research Association, Cardiff; www.ibra.org.uk.

hugely enlarged ventral cornua that may serve to hold the queen, as shown in Figure 6.8. In the dwarf bees, the dorsal cornua and bulbus are quite small and pointed (Figure 6.7) and are presumably insufficient to securely hold the queen and her partner together. Instead, the males probably deploy the thumb of their hind legs (Figure 2.2) to grasp the hind legs of the queen (Koeniger and Koeniger 1991; Ruttner 1988). After mating, the cavity-nesting species leave a large plug of mucus and cornual secretions, and remains of the endophallus, in the queen’s sting chamber, and this is known to beekeepers as the “mating sign” (Koeniger 1986) because it can often be seen protruding from the sting chamber when a queen returns from a mating flight. The males of the dwarf bees do not have substantial mucus glands, and their mating sign consists only of a small plug of orange cornual secretions (Figure 6.9; Koeniger et al. 2000; Koeniger and Koeniger 2000b; Koeniger, Koeniger, and Wongsiri 1989). The mating sign of A. dorsata has not been observed (Koeniger and Koeniger 2000b; Tan et al. 1999), but since the males have large mucus glands, it seems likely to exist. In many insects mating plugs have evolved so that males can monopolize paternity—they prevent second fertilizations (Thornhill and Alcock 1983). But in honey bees this does not seem to be the case because, at least in the cavity-nesting species, the mating sign actually facilitates subsequent copulations by providing a visual cue that helps subsequent males

Reproduction, Swarming, and Migration

A

C

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Cornual secretion

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Figure 6.9 The mating signs of some members of the genus Apis. A: A. mellifera. B: A. cerana. C: A. koschevnikovi. D: A. andreniformis. Redrawn with permission from figure 5 of N. Koeniger and G. Koeniger, Apidologie 31 (2000): 313–339.

locate a queen (Koeniger 1990). The apparent ease with which males can remove the mating sign supports the view that the mating sign has not evolved to inhibit subsequent copulations. Rather, Koeniger and Koeniger (1991) argue that a male that successfully mates with a queen benefits if other males also mate with her, because queens that are multiply mated are more likely to produce a large colony capable of swarming (we discuss the hypothesized reasons for this in the next chapter). Such a system is potentially vulnerable to invasion by a selfish strategy in which males increase their individual reproductive success by not deploying the sign, and not encouraging further copulations, thus increasing their share of paternity of their partner’s offspring (Woyciechowski, Kabat, and Król 1994). We suggest that the mating sign probably functions as both a visual cue and, in all species except the dwarf bees, more important, as a barrier against backflow of semen. The dwarf bees inject semen directly into the median oviduct, and possibly the spermatheca itself (Koeniger and Koeniger 1990; Koeniger et al. 2000; Koeniger, Koeniger, and Wongsiri 1989), and this may be the reason why they do not require a plug of mucus to prevent backflow. Their mating sign of cornual secretions may help other males locate the queen with which they have just mated. But since it is small, it may not be visible and may serve no function beyond holding the pair together in copula.

A Lack of Barriers to Sperm Transfer During mating by cavity-nesting and giant honey bees, semen is deposited in the median and lateral oviducts of the queen (Koeniger 1984). Sperma-

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tozoa need to migrate from the oviducts into the spermatheca. Artificial insemination (Harbo 1986) mimics the mating process by allowing placement of semen collected from drones in the median oviduct. A. florea (Woyke 1993), A. cerana (Koeniger, Koeniger, and Tingek 1996a; Lap and Chinh 1996; Wongsiri, Lai, and Sylvester 1990; Woyke 1975) and A. koschevnikovi (Koeniger, Koeniger, and Tingek 1996a) queens have all been successfully artificially inseminated, in the sense that sperm injected into the queens successfully migrated to the spermatheca. Owing to the difficulty of obtaining virgin queens, artificial insemination has not been performed on a queen of a giant bee species to our knowledge. Artificial insemination allows experimenters to inseminate queens of one species with semen from another, and such experiments have often been performed (see, e.g., Koeniger, Koeniger, and Tingek 1996a; Phiancharoen et al. 2004; Ruttner and Maul 1983; Woyke 1993; Woyke, Wilde and Wilde 2001a). In all cases at least some spermatozoa migrate to the spermatheca, though this may not survive for more than a few weeks (Phiancharoen et al. 2004). Interspecific hybrids have extremely low viability. Either eggs are not fertilized and result in males, or if they are fertilized, they die during embryogenesis or tend to produce gynandromorph hybrid workers (Koeniger and Koeniger 2000b). These results clearly show that even if copulation occasionally occurs between honey bee species, there are strong barriers to fertilization that would functionally prevent interspecific hybridization.

Summary The development times of immature honey bees are highly conserved across species but different among castes, with queens developing much more rapidly than drones. Honey bees mate on the wing probably at speciesspecific locations (called drone congregation areas) and times. This separation provides a useful criterion for recognizing species, and helps to prevent interspecific matings. Mating times and places are, however, plastic across each species’ range. Reproductive swarming is not well studied in Asian honey bees, and it is not certain (though it is likely) that reproductive swarms of all species form temporary clusters while they search for the best available nest site.

Reproduction, Swarming, and Migration

Asian honey bees frequently abscond from their nests and undertake migrations that in A. dorsata may be as much as 100 km. The giant bees form huge aggregations of colonies and swarms return to use the same nest site season after season. The distribution of dwarf bee nests is also aggregated, but it is not known if colonies return to nest sites.

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7 Worker Sterility, Kin Selection, and Polyandry

Worker honey bees can lay unfertilized but perfectly viable eggs that, if properly reared, will develop into normal males fully capable of mating. Given that workers can lay eggs and that in queenless colonies they do lay eggs, many of which are reared, why don’t they lay eggs in colonies with a queen? In this chapter we will explore why there is conflict among workers, and between workers and the queen, over which individuals should lay the colony’s male-producing eggs, and how this conflict is generally resolved so that colonies function as cohesive units. These are general questions for the evolution of eusocial insects, but studies of Asian honey bees have provided important data and model systems for addressing these questions. The number of times the queen mates is a critical parameter for understanding intracolonial conflict. Thus the second major theme of this chapter is the evolution of multiple mating. Multiple mating is uncommon in the social Hymenoptera (Strassmann 2001), and single mating was almost certainly the ancestral condition for social insects. But extreme multiple mating (here defined as involving more than six mates) is universal in honey bees (Palmer and Oldroyd 2000). A. dorsata queens, for example, can mate over 100 times (Wattanachaiyingcharoen et al. 2003). What is the meaning of such seeming profligacy? Why do drones of some species produce more sperm than the queen can store, and why does the queen reject most of the sperm she receives during mating, retaining only a little from each drone?

Worker Sterility, Kin Selection, and Polyandry

The Evolution of Worker “Sterility” A defining characteristic of eusocial insects is worker sterility or subfertility, but the evolution of sterility is difficult to fathom. (Consider the relative fitness of an allele “for” sterility versus an allele that confers fecundity—other things being equal the fecundity allele should spread at the expense of a sterility allele). Hamilton (1964, 1972) pointed out that in haplodiploid insect societies, workers can actually gain more reproductive success (in terms of the number of copies of their genes they get into the next generation) by helping to rear their sisters rather than their daughters. Hamilton showed how the seemingly altruistic sterility of workers can actually be a major fitness benefit to them and therefore both logical and “selfish” in the evolutionary sense (Dawkins 1976). In the simplest case, where a queen is mated to a single haploid male, her female offspring are more related to each other (relatedness, r, = 0.75) than to their own daughters (r = 0.5). Here the benefits to sisters of refraining from personal reproduction of daughters and helping to rear the female brood of their mother are clear: their sisters are 0.75/0.5 = 3/2 times more valuable than their daughters. This is all very well for female offspring; what about male offspring? Here the fitness benefits to workers of sterility aren’t nearly so clear cut because workers are always more related to their own sons (r = 0.5) than to those of their mother (r = 0.25). But workers can still benefit from functional sterility if they can manipulate the sex ratio of their colony’s reproductives in favor of females in a ratio of three times the investment into females to each unit of investment to males, mirroring the relative kin value of sisters (0.75) to brothers (0.25) (Crozier and Pamilo 1996; Trivers and Hare 1976). Alternatively, workers can be functionally sterile with respect to female eggs, but contribute some or all of the male-producing eggs to the colony, perhaps removing haploid eggs laid by the queen. Although other nongenetic factors almost certainly played a role, the relatedness asymmetry between sisters and daughters in broods of monandrous haplodiploid females makes the development of eusocial life histories more likely to evolve in hymenopteran insects than in diploid species, and probably explains why eusociality is much more common in the

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Hymenoptera than in other animals (Crozier and Pamilo 1996). It is important, however, to note that monandry has been replaced by high levels of polyandry in queens of the honey bees, and several other “advanced” eusocial insects, including army ants (Kronauer et al. 2004), leafcutter ants (Villesen et al. 2002), harvester ants (Rheindt et al. 2004), and vespid wasps (Foster and Ratnieks 2001). An interesting exception is the stingless bees, which for the most part are monandrous (Palmer et al. 2002; Peters et al. 1999). In those species that are polyandrous, workers are obliged to rear their half sisters (r = 0.25) as well as their more valuable full sisters (r = 0.75). Despite this, functional worker sterility persists in these polyandrous species. Why? We think that the most important reason is that these species are irreversibly eusocial: queens cannot reproduce without workers and workers cannot reproduce without queens (see Chapter 1). It is difficult to see how an advanced eusocial species could be successfully invaded by mutations that resulted in solitary life histories (Thompson and Oldroyd 2004; Wilson 1971). (On the other hand, mutations favoring polyandry can easily invade populations of advanced eusocial insects [Ratnieks 1988; Ratnieks 1990b].) The unlikelihood of a reversal to a solitary life history is particularly high in species, such as stingless bees, honey bees and army ants, that reproduce by colony fission rather than by a solitary queen, as do most social wasps and ants and all bumble bees. In fission-reproducing species, individual queens are incapable of foraging, let alone providing for a brood; to reproduce, such queens must have an entourage of workers. Conversely, workers are trapped in their subfertile role because they have lost their ability to mate. This permanently excludes workers from producing fertilized eggs that could give rise to females. (In rare exceptions, honey bee workers can produce female offspring by thelytokous parthenogenesis [Onions 1912; Tucker 1958], rather than the normal arrhentokous parthenogenesis that gives rise to males.) An inability to produce female offspring (or male offspring in thelytokous individuals) means that workers are reliant on their queen to produce offspring of both sexes, and mutations promoting a solitary worker cannot invade these populations. For these reasons, it is not surprising that there are no examples of reversals from advanced eusociality to solitary life histories. (But reversals in “primitively” eusocial species, those without a divergent queen and worker caste, are quite common [Wcislo 1977]). Certainly there are instances of

Worker Sterility, Kin Selection, and Polyandry

parasitic worker reproduction in advanced eusocial species (here defined as those with a strongly and irreversibly divergent queen and worker caste), including examples from honey bees (Martin, Beckman, et al. 2002; Oldroyd, Smolenski, et al. 1994). In some ants, the queen caste has been lost, to be replaced by a reproductive “gamergate” worker (Monnin and Peeters 1997). But these examples do not constitute a reversal from eusociality to solitary life. Nonetheless, although we have no examples of reversals to solitary life history from highly eusocial life history, workers of most eusocial insect species have fully functional ovaries and lay many eggs when the colony is queenless (Michener 1974). In some monandrous stingless bee species, the majority of drones are sons of workers (Beig 1972; Drumond, Oldroyd, and Osborne 2000; Machado, Contel, and Kerr 1984). So why don’t honey bee workers and queens share the production of male eggs? An important part of the answer to this question came as recently as 1987, when Michael Woyciechowski (1987) and Francis Ratnieks (1988) independently pointed out that although direct production of sons is favored for each individual worker, workers are only distantly related to the sons of their half sisters and are therefore selected to minimize reproduction in fellow workers. To understand this theory, consider the focal worker, Alison, in Figure 7.1. Alison is always more related to her own offspring males (e.g., Andrew, r = 0.5) than to any other male in the colony, including sons of the queen (e.g., Quinton, r = 0.25) and sons of her full sister Anne (e.g., Anthony, r = 0.375). But she is particularly distantly related to sons of her half sister Betty (e.g., Ben, r = 0.125). Ratnieks (1988) pointed out that these asymmetries of relatedness, generated by multiple mating, mean that reproduction by any worker reduces the fitness of other workers, particularly that of half sisters. Reduced worker fitness has two origins. First, the relatedness of half sister’s sons is low relative to sons of the queen, meaning that workers that tolerate reproduction by their half sisters while they themselves remain sterile are severely disadvantaged. Second, reproductive workers probably contribute less work to the colony, reducing the colony’s output of males and swarms relative to colonies comprised solely of nonreproductive workers (Dampney, Barron, and Oldroyd 2004; Moritz and Hillesheim 1989; Ratnieks 1988). Given the costs of worker reproduction, Ratnieks (1988) predicted the existence of worker policing—any behavior in which workers act to sup-

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Quinton

Queen

Adrian

Bruce

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Figure 7.1 Relatedness of male offspring in a honey bee colony to the focal worker, Alison. Black shading = the queen; gray shading = workers.

press reproduction in other workers—in polyandrous species of haplodiploid social insects. Worker policing could take several forms, but aggression toward reproductively active workers or the selective removal of worker-laid eggs seem likely candidates. In the following year Ratnieks and Visscher (1989) showed that, precisely as Ratnieks (1988) had predicted, worker policing is an integral part of the life of honey bee colonies. They demonstrated worker policing by a simple experiment (Figure 7.2) in which they obtained worker-laid eggs from a queenless colony that contained large numbers of laying workers, and queen-laid eggs from a colony with a queen. They transferred these two kinds of eggs into a normal queenright discriminator colony. They then monitored the removal of the eggs over 24 hours. Eggs of laying workers were quickly removed, but many queen-laid eggs were retained. This shows that honey bee workers can discriminate between worker-laid eggs and queen-laid eggs and remove the former (Figure 7.2). Policing by aggression toward reproductively active workers probably exists in A. mellifera (Visscher and Dukas 1995), but is not very effective (Dampney, Barron, and Oldroyd 2002). We don’t know if aggression toward reproductive workers exists in any of the Asian species. The means by which honey bees distinguish queen-laid and worker laid eggs is not yet completely clear (Katzav-Gozansky, Soroker, and Hefetz 2002a; Katzav-Gozansky, Soroker, and Hefetz 2002b; Katzav-Gozansky et al. 2001; Katzav-Gozansky, Soroker, Francke, et al. 2003; Martin et al. 2004;

Worker Sterility, Kin Selection, and Polyandry

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Figure 7.2 Left: The method of a standard policing experiment. Worker-laid eggs and queen-laid eggs are transferred from their combs into a single drone comb from the tester colony, which is queenright. Rows containing eggs are identified by colored pins. The combs are examined every hour or so for the presence of eggs. Right: The results of the original worker policing experiment with A. mellifera performed by Ratnieks and Visscher (1989).

Martin, Jones, et al. 2002), but it seems likely that queens mark their eggs with a chemical signal that cannot be produced by workers (Oldroyd, Ratnieks, and Wossler 2002). Nurse workers frequently inspect brood cells containing eggs, and remove those eggs that are not marked with the queen’s pheromonal mark (Halling and Oldroyd 2003). That the mark is a chemical and not a physical signal is known because if worker-laid eggs are briefly touched to the surface of queen-laid eggs the treated workerlaid eggs are more likely to survive in a worker policing assay than control worker-laid eggs (Ratnieks 1995) and because no physical difference between worker-laid and queen-laid eggs has been found (Katzav-Gozansky, Soroker, Kamer, et al. 2003; Martin, Jones, et al. 2002). An important observation is that workers do not selectively remove worker-laid eggs because they are less viable than queen-laid eggs. First, the viability of queen-laid eggs and worker-laid eggs produced in strong colonies is similar (Ratnieks and Visscher 1989). Second, if a colony is offered queen-laid eggs, half of which have been killed by exposure to carbon dioxide, there is no difference in the rates at which the two kinds of eggs are removed (Beekman and Oldroyd 2005).

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Figure 7.3 Dwarf bees and cavity-nesting bees both police worker-laid eggs. Right: A worker policing experiment on A. florea. The test drone comb containing grafted worker-laid and queen-laid eggs is suspended beneath the discriminator colony. Left: The results of worker policing experiments in A. cerana (top) and A. florea (bottom). The circles are the queenlaid eggs and the squares the worker-laid eggs. ND = next day (about 22 hrs). The data are from Halling et al. (2001) and Oldroyd et al. (2001). Photograph by P. Nanork.

Across the genus, successful worker reproduction is extremely low in colonies with a queen. In A. mellifera, sons of workers account for fewer than 1 percent of the drones produced by a colony (Visscher 1996). In the Asian species A. cerana, A. dorsata, and A. florea, no worker-laid male has ever been detected despite extensive sampling (Halling et al. 2001; Oldroyd et al. 2001; Wattanachaiyingcharoen et al. 2001). Worker policing via differential egg eating has been demonstrated in two Asian species, A. cerana (Oldroyd et al. 2001) and A. florea (Halling et al. 2001). The experimental design is essentially the same as in Figure 7.2, and the results are similar to those found in A. mellifera: worker-laid eggs are quickly removed, whereas many queen-laid eggs are retained (Figure

Worker Sterility, Kin Selection, and Polyandry

7.3). Evidence of effective worker policing in these phylogenetically diverse species, coupled with genetic evidence for the absence of any significant worker reproduction in any honey bee species (Halling et al. 2001; Oldroyd et al. 2001; Visscher 1989; Wattanachaiyingcharoen et al. 2001), strongly suggests that worker policing is universal in the genus, and that it was necessary for worker policing to evolve alongside polyandry to stabilize the inherent reproductive conflicts that arise from it (Oldroyd et al. 2001).

Queen Signals Given that worker policing is so effective, the possibility of successful worker reproduction is extremely limited. This leads to the prediction that workers should refrain from activating their ovaries whenever the queen is present, because doing so has a physiological cost and is pointless owing to policing (Wenseleers, Hart, and Ratnieks 2004). Queen honey bees inform workers of their presence via pheromones that they secrete from their mandibular and other glands (Free 1987; Plettner et al. 1993; Plettner et al. 1997; Winston and Slessor 1998; Wossler and Crewe 1999). These signals are acquired from the queen by the workers in her immediate vicinity, and then spread to other workers in the colony mainly via bodily contacts (Naumann et al. 1991; Seeley 1979; Watmough 1997). In the presence of these signals, the vast majority of workers refrain from activating their ovaries, which persist as vestigial threads. These signals of queen presence have been interpreted in three main ways. The first model regards the queen’s pheromones as a cocktail of chemical suppressors of reproduction in workers (see, e.g., Michener 1974; Zahavi and Zahavi 1997). Support for the chemical suppression model comes from the great chemical complexity of the pheromones themselves, and the ability of workers, especially reproductively active ones, to chemically mimic some of them (Katzav-Gozansky, Soroker, and Hefetz 2000; Simon, Moritz, and Crewe 2001). This complexity may be evidence of an evolutionary arms race between workers and queens (Foster, Ratnieks, and Raybould 2000; Katzav-Gozansky, Soroker, Francke, et al. 2003). The argument goes something like this: A mutation in workers allows them to escape chemical suppression and become reproductively active, parasitizing the colony with their eggs. As this mutation becomes more common,

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queen fitness is reduced, which creates strong selection pressure for queens to produce another chemical to counter the effects of the new mutation. The net effect is the diversity of chemical constituents found in the queen’s mandibular pheromones. But since other pheromones of the honey bee, notably the alarm pheromones, also show great chemical complexity (Free 1987), we feel that complexity in itself is weak evidence for either an arms race or a chemical suppression model for the control of worker fertility. The chemical suppression model is unlikely to apply to honey bees and other species where the queen is vastly outnumbered by the workers (Keller and Nonacs 1993; Seeley 1985). A queen would need to devote huge metabolic resources to the production of the suppressor chemicals, and protect herself from their effects. If it really comes down to biological warfare between the castes, it seems that such a battle is unlikely to be won by a single queen against 50,000 workers. This leads us to the second model for the role of queen pheromones. Under this model queen pheromones act not as a suppressor but as an honest signal of the queen’s presence and fertility. Workers respond to queen signals by not activating their ovaries, not because they are chemically coerced but because it is in their best interests not to do so (Keller and Nonacs 1993; Seeley 1985): in the presence of a highly fecund queen (and assiduously policing fellow workers) (Wenseleers, Hart, and Ratnieks 2004), a worker’s most profitable reproductive strategy is to contribute to the production of the queen’s offspring. The third model for the role of queen pheromones is Zahavi’s (1997, 2003) Handicap Principle, which is a kind of hybrid of the first two. Zahavi argues that in order for the queen’s signal to be respected by the workers as an honest indication of her fecundity, it must be metabolically expensive for the queen to produce it, for if it were not, any worker could mimic it. That is, the currency of the queen’s signal cannot be merely a cheap token, but must be backed by something more solid: a metabolic inefficiency that only queens, which are fed by workers, can afford to produce. Evidence from honey bees does not support the need for a handicap for workers to recognize and respond to the queen’s signal by not activating their ovaries. In the cavity-nesting species and A. dorsata, the main component of the worker’s mandibular gland secretions (which are used to feed young larvae) is either 9-hydroxy-(E)2-decenoic acid (9-HDA) or 10hydroxy-(E)2-decenoic acid (10-HDA), whereas the main component of

Worker Sterility, Kin Selection, and Polyandry

the queen’s mandibular pheromone is 9-keto-(E)2-decenoic acid (9-ODA) (see Table 6.2). There is no evidence that 9-ODA is biosynthetically more expensive to produce than HDA (Plettner et al. 1995, 1996). But 9-ODA is not a good food source for larvae (Gadagkar 1997), whereas it is good at attracting males (Chapter 5) and workers (Keeling et al. 2003; Pankiw, Winston, and Slessor 1994). This would argue that honey bees of these species do recognize the token 9-ODA as an honest queen signal and that it is not a handicap for queens to produce it. The dwarf bees, on the other hand, may use another mandibular gland component as the primary signal of the presence of royalty. With their smaller nests, and the high ratio of brood to adult workers, the queen’s presence may be more obvious to workers than it is in other species where a large proportion of the workers have reduced direct contact with the brood or the queen. It may even be possible for queens of the dwarf species to directly police their offspring. A. florea queens frequently patrol their nests and elicit a strange dorsoventral shaking (almost a bow) in workers as they pass (Oldroyd, Sylvester, et al. 1994). Perhaps the queen’s signal is transferred during these exchanges. Before we leave queen signals completely, we would like to make some final remarks. First, 9-ODA is not the only queen signal. Although 9-ODA is important at signaling the queen’s presence, it is not involved in the most important reproductive signal, the queen’s egg-marking signal. This as yet unidentified signal is the basis for worker policing, and ultimately the stability of colonial life in a polyandrous species (Barron, Oldroyd, and Ratnieks 2001; Ratnieks 1988). We predict that there should be no biologically possible mechanism by which reproductive workers can fake the queen’s egg-marking signal; that is, the queen’s egg-marking signal should be produced by some structure that is present in queens but absent in workers, or, when present in workers causes them to be attacked because it conflicts with some other worker signal. Obvious structures that are present in queens but not workers are the spermatheca, its associated glands, and the spermathecal duct. But surgical removal of the spermathecal gland seems to make no difference to egg survival (Koeniger 1970), which suggests that it is not the source of the unique queen egg-marking signal. Recent work by Pflugfelder et al. (2004) has demonstrated a cuticular hydrocarbon that is present on the dorsal abdominal surface of queens but not workers. This relatively simple hydrocarbon elicits queen fighting in all species of honey bee and is the primary signal by which queens recognize

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other queens (Pflugfelder, Koeniger, and Svatos 2004). Since queen cuticular hydrocarbons are used for distinguishing queen-laid and worker-laid eggs in the ant Campanotus floridanus (Endler et al. 2004), it could be that these compounds are also the primary queen egg-marking signal in Apis.

Conflicts in Queenless Colonies The queen lays male-producing eggs that are equally related to all workers. Her eggs offer a compromise position for workers, for while the queen remains the sole reproductive, no nonreproductive worker has higher reproductive success than any other (at least via male production). When honey bee colonies become queenless, the egalitarian queen-laid eggs are no longer being laid. Let us suppose that the colony is unable to rear a new queen, or that the replacement virgin queen is lost on her mating flight. Now the only possibility for the orphaned colony to reproduce is via the production of worker-laid males, which have a small but nonzero possibility of mating with queens from other colonies (Berg et al. 1997). Under these circumstances it is in the best interests of individual workers and of the queenless colony as a whole for some workers to activate their ovaries and lay eggs. The colony should then raise as many drones as possible before it finally perishes owing to a lack of replacement workers. We would predict, therefore, that queenless colonies would be more tolerant of worker reproduction than colonies with a queen. Nonetheless, each worker is still more related to her own eggs than she is to those of full sisters or half sisters (Figure 7.1). This leads to potential conflict among workers over which individuals will lay and which will work, and to potential conflicts among larvae about which of them should be reared to maturity. In A. mellifera and A. florea there is good evidence that the potential conflict among subfamilies predicted from kin selection theory is indeed played out in queenless colonies. Subfamilies (Figure 1.2) vary considerably in their reproductive success, with some subfamilies being more successful in producing eggs and having them reared by the colony than others (Martin, Oldroyd, and Beekman 2004; Nanork, Oldroyd, and Wongsiri 2004; Page and Robinson 1994). Martin et al. (2004) showed that workers in competitive subfamilies activate their ovaries early, and that their eggs have high survival rates. Workers in less competitive subfamilies have de-

Worker Sterility, Kin Selection, and Polyandry

layed ovary activation and/or reduced egg survival. But queenless workers eventually become more permissive of each other’s eggs, an indication of colony-level selection for reproduction of at least some drones (Miller and Ratnieks 2001). In A. cerana, workers rapidly change their reproductive status after dequeening. In most colonies, 50 percent of workers have eggs in their ovarioles 5 days after dequeening, and worker-laid eggs appear just 3 days after dequeening (Oldroyd et al. 2001). In contrast, successful worker oviposition does not occur in A. mellifera for 20–40 days after dequeening (Miller and Ratnieks 2001; Page and Erickson 1988; Ruttner and Hesse 1979). In A. florea, 1–2 percent of workers activate their ovaries within 4 days but most workers do not have active ovaries until 2 weeks after dequeening (Nanork et al. 2005).

The Behavior of Queenless Dwarf Honey Bees Colonies of dequeened dwarf bees rapidly stop foraging. The cluster breaks up and the bees spread along the supporting branch in loose festoons as if seeking to regain their lost queen (Figure 7.4). More than half of the A. florea and A. andreniformis colonies that we have dequeened have absconded within a few days, abandoning the brood that they might have used to rear a replacement queen. Still more colonies abscond as soon as the first virgin emerges. The reason that queenless dwarf bee colonies are so reluctant to stay around may be that they are vulnerable to reproductive parasitism by workers from other nests (Nanork, Paar, et al. 2005). In order to rear its own drones, a queenless colony must cease policing behavior in order to allow drones to be reared. This window of opportunity is apparently exploited by workers from other nests because genetic analysis shows that about 5 percent of the workers in a queenless nest are from other colonies. The foreigners have twice the rate of ovary activation as natal workers, and account for about 30 percent of a queenless colony’s drone offspring (Nanork, Paar, et al. 2005). It is not known where the foreign workers come from. Are they themselves from queenless nests, or are they true parasites from queenright ones? We suspect that they are from queenright colonies, because we have

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Figure 7.4 The disordered curtain of a queenless A. andreniformis colony. Photo by B. Oldroyd.

occasionally found queenless nests consisting solely of drone comb (Figure 7.5). Thus at least some queenless workers try and reproduce in their own nests rather than exploiting others. The cells of queenless dwarf bee nests are often packed with 10 or more eggs, and it would be very interesting to determine the maternity of eggs within cells. Are they the eggs of a single worker (parasitic or otherwise) that guards her own eggs, or are they from different workers? How is it determined which eggs are reared?

The Behavior of Queenless A. cerana As with A. mellifera (Ratnieks 1993), worker activation is extremely rare in queenright colonies of A. dorsata (Velthuis et al. 1971; Wattanachaiyingcharoen et al. 2001) and A. florea (Halling et al. 2001), but in A. cerana an exceptionally high proportion (about 5 percent) of workers have activated ovaries and probably lay eggs (Bai and Reddy 1975; Blanford 1923;

Worker Sterility, Kin Selection, and Polyandry

131

Figure 7.5 A queenless colony of A. florea. The comb consists solely of drone comb, and drones are being successfully reared. Presumably this colony arose from a queenless absconding swarm. In dequeened A. florea colonies, there is often massive worker egg laying, with many eggs per cell, and many eggs laid by parasitic workers from other colonies. Photo by B. Oldroyd.

Oldroyd et al. 2001). The 5 percent estimate is based on samples of random-aged bees, which suggests that a much higher proportion of the workers aged around 10 days (when ovary activation peaks) would have activated ovaries, perhaps as many as 50 percent. This proportion is similar to that of the anarchistic strain of A. mellifera maintained at Sydney University, which has been specifically bred for high rates of worker reproduction, and in which the pheromonal cues that normally regulate worker reproduction have been seriously disrupted (Barron and Oldroyd 2001; Barron, Oldroyd, and Ratnieks 2001; Oldroyd, Wossler, and Ratnieks 2001). We do not know if rates of worker ovary activation found in A. cerana are also found in the other Asian cavity-nesting species, or if A. cerana is unique. A. cerana does, however, provide a fascinating exception to A. dorsata, A. florea, and A. mellifera. Oldroyd et al. (2001) suggested that

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high rates of ovary activation, but limited reproductive success, of A. cerana workers may be an adaptation to frequent events of queenlessness in this species. There is not, however, any compelling reason to expect that A. cerana is any more prone to becoming queenless than other Apis species. Perhaps the high rates of worker oviposition in A. cerana are just an example of the interplay of selfish and altruistic reproduction: no one strategy is superior, and A. cerana has come down more on the side of individualism than the more altruistic A. mellifera, A. florea, and A. dorsata.

Determining Colony Kin Structure Multilocus genotyping of a large sample of workers at several microsatellite loci allows the kin relationships of a colony to be determined. In Figure 7.6, we illustrate how the genotypes of workers can be used to infer the genotype of the queen heading the colony and the genotype of all the males that mated with her. Following such a paternity analysis it is usual to estimate three parameters that describe the kin structure of the colony (Boomsma and Ratnieks 1996; Tarpy and Nielsen 2002). The first parameter is the mating frequency, k, which is simply the number of different fathering males detected in the worker sample. This is an estimate of the number of copulations by the queen, but is probably an underestimate because: (1) Not all males may be represented in the sample of workers (possibly because the sample size is too small or because the queen uses her sperm supply nonrandomly. (2) Some males may produce little or no viable sperm. And (3) the method may fail to detect some males because two or more have identical genotypes, or share an allele with the queen at each locus. The probability of not sampling a patriline, nd, can be estimated from nd = (1 − p)n, where n is the number of workers sampled and p is the proportion of the patriline in the worker sample (Foster et al. 1999). The parameter k can also be overestimated if there are “phantom” subfamilies arising from scoring errors or microsatellite mutations (Tarpy and Nielsen 2002). The second useful parameter is m, the Effective Paternity Frequency (Boomsma and Ratnieks 1996), also known as the Effective Promiscuity (Starr 1979). If all males contribute equally to the paternity of brood, then m = k. But this is never the case, with some males fathering more workers

Queen’s genotype

Locus 1

Locus 2

Locus 3

104/104

233/235

105/109

233/235

109/109

Drone’s genotype

105,233/235,109

104/105 105/110 105,235,110

235/235 109/110 104/100

233/233

105/107

100,233,107

109/107

103,240,107

109/109a 104/103

233/240

105/109b 109/115

103,240,109

103,240,115

Figure 7.6. How to determine mating frequency from multilocus genotyping of a worker sample. In this simple example, 8 workers were genotyped at three loci. The genotypes of the workers can be read from the figure by following the arrows. From the top, the first worker had the genotype 104/105 at the first locus, 233/235 at the second, and 109/109 at the third. The first task is to place the genotypes of each worker into a spreadsheet and then to sort them by genotype so that the spreadsheet looks roughly like the figure. The genotype of the queen is then determined for each locus from the worker sample. (Usually this requires at least 12 workers for a reliable determination.) For Locus 1, we infer that the queen is homozygous 104/104 because all workers carry at least one 104 allele. For Locus 2 the queen must be heterozygous 233/235 because some workers are homozygous 233/233 and some are homozygous 235/235. For Locus 3, the queen is probably heterozygous 105/109 because all workers carry one of these two alleles. After the genotype of the queen is determined, the genotype of the fathering male is determined for each worker by subtraction. These genotypes are shown in bold in the figure. Where a worker has the same genotype as the queen, the paternal allele cannot be determined, so both alleles are in bold. The final task is to determine the minimum number of male genotypes that could conceivably have fathered the array of worker genotypes. This is always a puzzle, but drawing a bifurcating tree similar to the figure is helpful. When the paternal allele is ambiguous, as in workers with genotype 105/109 at Locus 3 (markedb), it is usual to pool genotypes to produce the minimum number of fathering males. So in this case we infer that there is definitely a father of genotype 103,240,109 (marked a)and that there may well be a second father of genotype 103,240,105 (marked b). But the workers of genotype 105/ 109 at Locus 3 could also be fathered by a male of genotype 102,240,109. Thus, in the absence of other information, we declare a single father rather than two. There are at least two computer programs available to help with social insect pedigree reconstructions, including Easymate available at http:// www.bio.usyd.edu.au/Social_InsectsLab/Social_InsectsLab.htm.

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than others, and thus raising the average relatedness of workers. The parameter m is estimated as 1 ∑ p i2 where pi is the proportion of workers sired by the ith male. A correction for small sample size is often applied so that mc =

n −1 n

n ∑ p i2 −1 i =1

where n is the number of workers sampled (Pamilo 1993). Tarpy and Nielsen (2002) developed an improved estimate that is applicable to single colonies: mc =

(n −1) 2 k

∑ p i2 (n +1)(n −2) +3−n

i =1

which has an approximate variance Var(me) =

2k 2 (k −1)(n −2) 2 (n −1)n (n +1) 2 . (2k +(n −2)(n −1)) 4

The third parameter is r, the average relatedness of workers. Average relatedness varies between 0.75, the relatedness of haplodiploid female full sibs under monandry and approaches 0.25, the relatedness of half sibs, under high levels of polyandry according to the function r = 0.25 +

0.5 me

(Figure 7.7; Crozier 1970). With the exception of A. nuluensis, we have estimates of the kin structure of all honey bee species (Table 7.1) though undoubtedly these could be profitably refined. These figures show that mating frequency and effective mating frequency are high across the genus. Why?

Worker Sterility, Kin Selection, and Polyandry

Proportion of viable offspring

Relatedness

0.75

Figure 7.7 The relationship between effective mating frequency, relatedness, and brood viability. The top panel shows that worker relatedness decreases as a function of me, the effective mating frequency, according to the function r = 1/4 ⫹ 1/2me. The function asymptotes to 0.25. Over 90 percent of the change in worker relatedness occurs in the first six matings. The bottom panel shows the relationship between me and brood viability. The proportion of viable offspring (diploids heterozygous at the sex locus) varies as a function of me and the number of sex alleles, s. The expected proportion of viable offspring remains constant at 1 ⫺ 1/s, but the variance, (1/2me) (1/s) (1 ⫺ 2/s), (Adams et al. 1977; Page and Marks 1982), declines as 1/2me. Again, over 90 percent of the reduction in variance accrues in the first six matings. Redrawn with permission from K. Palmer and B. Oldroyd, Apidologie 31 (2000): 235–248.

0.5 me = 6

0.25

0

Mean + variance

1−1/s Mean − variance

5

10

15

135

20

Effective number of matings, me

The Evolution of Polyandry in Apis Clearly the switch from monandry to polyandry was a profound evolutionary transition, and so it is important that we get a good understanding of how it occurred. This probably explains why the causes of the evolution of polyandry is one of the most vociferously debated issues by social insect researchers (e.g., Kraus and Page 1998; Sherman, Seeley, and Reeve 1998), and it is with some trepidation that we weigh into the discussion, knowing that we may infuriate a few of our readers and irritate many more. As we discussed above, it seems likely that the ancestral condition of haplodiploid social insects was monandry. Furthermore, we would (somewhat controversially) argue that the risks of mating are nonzero to queens. For instance, we have seen bats feeding on A. dorsata drones returning from their mating flights in northeast Thailand, and the wasp Vespa affinis specifically hunts in A. cerana drone congregation areas (Koeniger, Koeniger, and Mardan 1994). It seems that predation like this must pose

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Worker Sterility, Kin Selection, and Polyandry Table 7.1

Numbers of detected fathers, k, effective paternity frequency, me, and average relatedness of workers, r, in honey bees. All entries are means of several colonies across all available studies ± s.d. Paternity data are from Tarpy et al. (2004) except for A. laboriosa, which is from Paar et al. (2004b). At the time of publication, data for A. nuluensis were not available. References to the primary literature are given in the table notes.

Species A. andreniformisa A. floreab A. dorsatac A. laboriosad A. ceranae A. koschevnikovif A. nigrocinctag A. melliferah

Paternity frequency (k)

Effective paternity frequency (me)

Coefficient of relatedness (r)

13.5 ± 4.5 11.6 ± 5.0 54.9 ± 31.5 34.4 ± 10.3 18.8 ± 5.6 16.2 ± 10.5 54.0 ± 11.5 12.0 ± 6.3

10.5 ± 1.9 7.9 ± 3.3 44.2 ± 27.1 19.9 ± 5.5 14.1 ± 3.9 13.3 ± 10.4 40.3 ± 23.4 11.6 ± 7.9

0.30 0.31 0.26 0.28 0.29 0.29 0.26 0.29

a. Oldroyd and Clifton et al. (1997). b. Oldroyd and Smolenski et al. (1995), Palmer et al. (2001). c. Oldroyd et al. (1996), Moritz et al. (1995), Wattanachaiyingcharoen et al. (2003). d. Paar et al. (2004b); see also Takahashi and Nakamura (2003). e. Oldroyd et al. (1998). f. Rinderer et al. (1998). g. Palmer et al. (2001). h. Mean of many studies calculated by Tarpy et al. (2004).

considerable mating risk to queens in all species. Why, then, the shift from monandry to polyandry? We argue that there must be considerable benefits associated with multiple mating; otherwise it would not have evolved given the costs involved.

The Initial Change from Monandry to Oligandry A shift from one mating to more than one effective mating is a profound evolutionary step for a eusocial insect, because the kin structure of the colony is so radically altered by it. The primary driver of the switch from monandry to polyandry almost certainly arose from the genetic load imposed by the method of sex determination (Page 1980; Palmer and Oldroyd 2000; Shaskolsky 1976). In honey bees and many other haplodiploids, sex is determined at a single sex-determining locus that is hemizygous in haploid males and heterozygous in diploid females (Cook and Crozier 1995). Diploid individuals that are homozygous at the sex locus are male, but in

Worker Sterility, Kin Selection, and Polyandry

honey bees these individuals are removed by nurse workers at the first larval instar (Woyke 1963). Thus a queen that mates with a single male carrying the same sex allele as she herself carries will suffer a 50 percent loss of her larvae. By mating with more than one male she will increase the probability of sharing a sex allele with at least one partner, but, crucially, will simultaneously decrease the overall proportion of her brood that are diploid males, because not all of her partners are likely to share a sex allele. To understand why, consider a population where there are s sex alleles at equal frequency and where queens mate only once. A queen is heterozygous at the sex locus so that there is a 2/s probability that one of her sex alleles will match that of the drone with which she mated. Thus the probability that a queen will mate with a male carrying a different sex allele and therefore having fully viable eggs is 1 − (2/s). This viability, however, is unevenly spread. Most colonies will have 100 percent viability, but in 2/s colonies, the queen will have mated with a male carrying the same sex allele as herself and suffer a 50 percent loss in brood viability (Page 1980). These unfortunate colonies, suffering low brood viability, are unlikely to ever generate a sufficiently large population to produce a swarm, and are therefore evolutionarily doomed. Thus low brood viability creates strong selection pressure to reduce that risk (Page 1980). A strategy to avoid this possibility is to mate with several males. As the number of males with which a queen mates increases, her expected brood viability approaches the population average (Figure 7.7; Page 1980; Palmer and Oldroyd 2000). Across a population the expected brood viability, V, is 1 − 1/s, and depends solely on the number of sex alleles (Page and Marks 1982). If there are 20 alleles in the population (which is realistic: see Adams et al. 1977; Ekbohm and Ebbersten 1979), the average brood viability is 95 percent, which is sufficient to allow a colony to develop to swarming strength. Thus by mating with many males, queens can eliminate the risk of being the mother of one of the 1 in 10 nonviable colonies that is expected to occur in a monandrous population with the same number of sex alleles (Page 1980). Polyandry can therefore be seen as a classic bet-hedging strategy against low brood viability.

The Shift to Extreme Polyandry Page and his colleagues (Kraus and Page 1998; Tarpy and Page 2000; Tarpy and Page 2002) have argued that the genetic load imposed by the sex lo-

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Worker Sterility, Kin Selection, and Polyandry Table 7.2

Summary of the five most plausible hypotheses for the evolution of extreme polyandry in honey bees.

1. Multiple mating favors sperm competition and thereby bet hedges against poorquality sperm (Madsen et al. 1992). 2. Queens can’t count, and often overshoot the optimal number of matings because extra matings entail little cost (Tarpy and Page 2000). 3. Multiple mating is favored because it increases the genetic variation within broods. This is beneficial because a mixture of genotypes promotes task specialization and an ability to buffer environmental stress (Oldroyd et al. 1992). 4. Multiple mating by queens is favored because it enables queens to store sufficient sperm to build up large and long-lived colonies (Cole 1983). 5. Multiple mating is selected because it mitigates against the effects of parasitism and disease (Sherman, Seeley, and Reeve 1988).

cus is all that is needed to explain polyandry in honey bees. But the benefits of multiple mating in generating sex-allele heterozygosity (Page 1980) quickly asymptote so that beyond six matings there is little additional benefit for each additional mating (Figure 7.7; Palmer and Oldroyd 2000). And yet all species mate many more than six times (Table 7.1). Why? In Table 7.2 we summarize what we believe are the only plausible hypotheses for the evolution of extreme polyandry in honey bees: the selective pressures that could have plausibly shifted mating frequencies well beyond 6 to 20 and sometimes many more. We suggest that all these selective pressures have probably contributed to the evolution of extreme polyandry in Apis, and that their combined effects provide very strong selection toward extreme polyandry. We discuss these hypotheses below. queen worker conflict over sex ratios Relatedness asymmetries within monandrous social insect colonies mean that the queen and the workers are in evolutionary conflict over the investment ratio in the colony’s reproductives: other things being equal, queens prefer an equal investment ratio (because they are equally related to sons and daughters), whereas workers prefer a female-biased one in the ratio of 3 female units to 1 male unit, mirroring the workers’ relatedness to their sisters (r = 0.75) and brothers (r = 0.25) (Trivers and Hare 1976). Under polyandry, the preferred investment ratio of workers becomes aligned with that of the queen because workers have approximately equal relatedness to their brothers (r = 0.25) and sisters (r = 0.25 + 0.5/me) (Moritz 1985;

Worker Sterility, Kin Selection, and Polyandry

Starr 1984). Therefore, queens can potentially manipulate their workers into raising the queen’s preferred investment ratio by mating with several males, a process over which they have power in that they have a “choice” to be monandrous or polyandrous (Moritz 1985; Queller 1993). In swarm-founding species like honey bees, the significance of the “conflict over sex ratios” hypothesis to the evolution of polyandry gets complicated. The honey bee population sex ratio is hugely (there are hundreds of males for every queen) male biased, which seemingly contradicts the prediction of female bias. Yet sex ratio investment should be measured not by merely counting the numbers of queens and drones but by assessing biological investment in the two sexes. Thus the apparent biological investment in a few grams of queens should be corrected by including an allowance for the mass of workers, food, and brood that must be left with an offspring queen when the colony swarms. But this argument is weak because if such a correction is made, the investment in queens hugely exceeds that for males for the mass of a swarm (a kilogram or so) is much greater than the mass of a few hundred drones in a colony. Thus the hypothesis of equalizing the preferred sex ratio investment between workers and queens is probably not relevant to honey bees. sperm selection Honey bee ejaculates vary greatly in the quality and quantity of spermatozoa that they carry, mature drones having many more viable sperm than younger ones (Harbo 1986). In most species, drones deposit sperm in or near the median oviduct (Koeniger et al. 1991). From there, sperm must migrate into the spermatheca. This seems like an ideal situation for queens to bias their sperm use toward more vigorous sperm that are presumably more likely to survive in the spermatheca for protracted periods (Woyciechowski and Krol 1996). Note that this is not sperm competition in the strict sense (Parker 1984), because there is little or no evidence that male honey bees compete for paternity via mechanisms that reduce the viability of the ejaculates of other males (Franck et al. 1999; Franck et al. 2002). Nor is it an example of post-copulatory female mate choice (Eberhard 1996) (also known as cryptic female choice), because honey bee paternities are extremely mixed, which suggests that spermatozoa of most copulations are used at least to some extent (Franck et al. 1999; Haberl and Tautz 1998; Sasaki, Satoh, and Obara 1995). Rather, the hypothesis is that

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Worker Sterility, Kin Selection, and Polyandry

honey bee queens mate with many males and thereby set up a competitive device for ensuring that only the most viable sperm of each male enter their spermathecae. We think that this hypothesis provides a plausible benefit for polyandry at low (50 ≈18 10–23 6 8.1 4.8 400

36,600 0.7 n.d. 60–90 15–35 n.d.