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Division of labor represents a major advantage of social insect communities that accounts for their enormous ecological success. In colonies of the honeybee, Apis mellifera, division of labor comprises different tasks of fertile queens and drones (males) and, in general, sterile female workers. Division of labor also occurs among workers in form of an age-related polyethism. This helps them to deal with the great variety of tasks within the colony. After adult eclosion, workers spend around three weeks with various duties inside the hive such as tending the brood or cleaning and building cells. After this period workers switch to outdoor tasks and become foragers collecting nectar, pollen and water. With this behavioral transition, workers face tremendous changes in their sensory environment. In particular, visual sensory stimuli become important, but also the olfactory world changes. Foragers have to perform a completely new behavioral repertoire ranging from long distance navigation based on landmark orientation and polarized-skylight information to learning and memory tasks associated with finding profitable food sources. However, behavioral maturation is not a purely age-related internal program associated with a change, for example, in juvenile hormone titers. External factors such as primer pheromones like the brood pheromone or queen mandibular pheromone can modulate the timing of this transition. In this way colonies are able to flexibly adjust their work force distribution between indoor and outdoor tasks depending on the actual needs of the colony. Besides certain physiological changes, mainly affecting glandular tissue, the transition from indoor to outdoor tasks requires significant adaptations in sensory and higher-order integration centers of the brain.
The mushroom bodies integrate olfactory, visual, gustatory and mechanosensory information. Furthermore, they play important roles in learning and memory processes. It is therefore not surprising that the mushroom bodies, in particular their main input region, the calyx, undergo volumetric neuronal plasticity. Similar to behavioral maturation, plastic changes of the mushroom bodies are associated with age, but are also to be affected by modulating factors such as task and experience.
In my thesis, I analyzed in detail the neuronal processes underlying volumetric plasticity in the mushroom body. Immunohistochemical labeling of synaptic proteins combined with quantitative 3D confocal imaging revealed that the volume increase of the mushroom body calyx is largely caused by the growth of the Kenyon cell dendritic network. This outgrowth is accompanied by changes in the synaptic architecture of the mushroom body calyx, which is organized in a distinct pattern of synaptic complexes, so called microglomeruli. During the first week of natural adult maturation microglomeruli remain constant in total number. With subsequent behavioral transition from indoor duties to foraging, microglomeruli are pruned while the Kenyon cell dendritic network is still growing. As a result of these processes, the mushroom body calyx neuropil volume enlarges while the total number of microgloumeruli becomes reduced in foragers compared to indoor workers. In the visual subcompartments (calyx collar) this process is induced by visual sensory stimuli as the beginning of pruning correlates with the time window when workers start their first orientation flights. The high level of analysis of cellular and subcellular process underlying structural plasticity of the mushroom body calyx during natural maturation will serve as a framework for future investigations of behavioral plasticity in the honeybee.
The transition to foraging is not purely age-dependent, but gets modulated, for example, by the presence of foragers. Ethyl oleate, a primer pheromone that is present only in foragers, was shown to delay the onset of foraging in nurse bees. Using artificial application of additional ethyl oleate in triple cohort colonies, I tested whether it directly affects adult neuronal plasticity in the visual input region of the mushroom body calyx. As the pheromonal treatment failed to induce a clear behavioral phenotype (delayed onset of foraging) it was not possible to show a direct link between the exposure to additional ethyl oleate and neuronal plasticity in mushroom body calyx. However, the general results on synaptic maturation confirmed my data of natural maturation processes in the mushroom body calyx.
Given the result that dendritic plasticity is a major contributor to neuronal plasticity in the mushroom body calyx associated with division of labor, the question arose which proteins could be involved in mediating these effects. Calcium/calmodulin-dependent protein kinase II (CaMKII) especially in mammals, but also in insects (Drosophila, Cockroach), was shown to be involved in facilitating learning and memory processes like long-term synaptic potentiation. In addition to presynaptic effects, the protein was also revealed to directly interact with cytoskeleton elements in the postsynapse. It therefore is a likely candidate to mediate structural synaptic plasticity. As part of my thesis, the presence and distribution of CaMKII was analyzed, and the results showed that the protein is highly concentrated in a distinct subpopulation of the mushroom body intrinsic neurons, the noncompact Kenyon cells. The dendritic network of this population arborizes in two calyx subregions: one receiving mainly olfactory input – the lip – and the collar receiving visual input. This distribution pattern did not change with age or task. The high concentration of CaMKII in dendritic spines and its overlap with f-actin indicates that CaMKII could be a key player inducing structural neuronal plasticity associated with learning and memory formation and/or behavioral transitions related to division of labor. Interestingly CaMKII immunoreactivity was absent in the basal ring, another subregion of the mushroom body calyx formed almost exclusively by the inner compact Kenyon cells and known to receive combined visual and olfactory input. This indicates differences of this mushroom body subregion regarding the molecular mechanisms controlling plastic changes in corresponding Kenyon cells.
How is timing of behavioral and neuronal plasticity regulated? The primer pheromone ethyl oleate was found in high concentrations on foragers and was shown to influence behavioral maturation by delaying the onset of foraging when artificially applied in elevated concentrations. But how is ethyl oleate transferred and how does it shift the work force distribution between indoor and outdoor tasks? Previous work showed that ethyl oleate concentrations are highest in the honeycrop of foragers and suggested that it is transferred and communicated inside the colony via trophallaxis. The results of this thesis however clearly show, that ethyl oleate was not present inside the honey crop or the regurgitate, but rather in the surrounding tissue of the honey crop. As additionally the second highest concentration of ethyl oleate was measured on the surface of the cuticle of forgers, trophallaxis was ruled out as a mode of transmission. Neurophysiological measurements at the level of the antennae (electroantennogram recordings) and the first olfactory neuropil (calcium imaging of activity in the antennal lobe) revealed that the primer pheromone ethyl oleate is received and processed as an olfactory stimulus. Appetitive olfactory conditioning using the proboscis extension response as a behavioral paradigm showed that ethyl oleate can be associated with a sugar reward. This indicates that workers are able to perceive, learn and memorize the presence of this pheromone. As ethyl oleate had to be presented by a heated stimulation device at close range, it can be concluded that this primer pheromone acts via close range/contact chemoreception through the olfactory system. This is also supported by previous behavioral observations.
Taken together, the findings presented in this thesis revealed structural changes in the synaptic architecture of the mushroom body calyx associated with division of labor. For the primer pheromone ethyl oleate, which modulates the transition from nursing to foraging, the results clearly showed that it is received via the olfactory system and presumably acts via this pathway. However, manipulation experiments did not indicate a direct effect of ethyl oleate on synaptic plasticity. At the molecular level, CaMKII is a prime candidate to mediate structural synaptic plasticity in the mushroom body calyx. Future combined structural and functional experiments are needed to finally link the activity of primer pheromones like ethyl oleate to the molecular pathways mediating behavioral and synaptic plasticity associated with division of labor in Apis mellifera. The here identified underlying processes will serve as excellent models for a general understanding of fundamental mechanisms promoting behavioral plasticity.
There is evidence that pheromones are communicative signals in animals. However, the existence and function of human pheromones are still under discussion. During the last years several substances have been labeled as putative human pheromones and especially 4,16–androstadien-3-one (androstadienone), found in male and female sweat, became subject of intense investigation. In contrast to common odors androstadienone presumably modulates human physiological and psychological reactions. Data suggest that androstadienone might influence the processing of visual cues, specifically faces or affective stimuli, via projections from the fusiform gyrus and the amygdala. Moreover, attentional processes may be modulated, which is supported by explicit and implicit behavioral data. This thesis includes three experimental studies examining effects of androstadienone exposure on behavioral and cortical reactions to visual and emotional stimuli. The main hypotheses were that androstadienone might influence human behavior to and perception of visual cues. The first study sought to clarify androstadienone effects on attention-related reactions as well as on behavioral tendencies. Motoric approach-avoidance reactions in response to happy and angry facial expressions were investigated in 30 women and 32 men. Participants either inhaled androstadienone or a control solution, without knowing the real content, while performing the following task: they had to push away or to pull towards them a joystick as fast as possible in reaction to either an angry or a happy cartoon face, which was presented on a computer screen. Results showed that androstadienone modulated the participant´s task performance by accelerating the reaction speed compared to the control compound. Faster reactions were observed particularly when reacting to angry faces but not when reacting to happy faces. This might be explained by the finding that human body odors, the source of androstadienone, were found to activate the human fear system, i.e. modulating fear-related attentional processes. Therefore, the quicker reaction towards angry faces with exposure to androstadienone could be due to an enhanced allocation of attentional resources towards fear-related cues like angry faces. Results also showed that androstadienone enhanced men´s approach tendency towards faces independent of emotional expressions. This observation might be explained by androstadienone´s former shown ability to improve attractiveness ratings of other persons. In this regard, the endogenous odor might enhance evaluations of faces in men and, thus, might improve their willingness to approach social stimuli. In contrast to men, women already showed in the control condition higher approach tendency towards faces. Therefore, androstadienone might rather maintain than enhance the approach score in women. In the second study event-related brain potentials (ERPs) triggered by social and non-social visual stimuli were investigated by means of electroencephalography. In a double-blind between-subjects design 51 women participated. Twenty-eight women inhaled androstadienone, whereas 23 women inhaled a control solution. Four different picture categories, i.e. real faces, pictures with couples, pictures with social and non-social scenes, each including three different valence categories, i.e. positive, negative and neutral, should clarify the stimulus type or context androstadienone is acting on. The androstadienone compared to the control odor did not influence brain responses significantly. Explorative analyses, however, suggested that androstadienone influences the processing of faces. While in the control group angry faces elicited larger P300 amplitudes than happy faces, the androstadienone group showed similar P300 amplitudes concerning all emotional expressions. This observation tentatively indicates that the endogenous odor might indeed affect the neuronal responses to emotional facial stimuli, especially late components reflecting evaluative processes. However, this observation has to be verified and further investigated, in particular whether androstadienone caused reduced responses to angry faces or enhanced responses to happy faces. The third study investigated androstadienone effects on face processing especially in men. ERPs elicited by happy, angry and neutral cartoon faces, which were presented on a computer screen, were measured while 16 men, not knowing the applicated odor, inhaled either androstadienone or a control solution. Exposure to androstadienone significantly increased later neuronal responses, the P300 amplitude. This belated component of the ERP reflects attention allocation and evaluative processes towards important stimuli. Therefore, androstadienone might facilitate central nervous face processing by enhancing attention towards these stimuli. In sum, the current results corroborate the notion of androstadienone as an active social chemosignal. In minute amounts and not detectable as an odor it influenced cortical and motoric reactions. Therefore, it might be concluded that androstadienone indeed affects cognitive functions like attentional processes and in turn affects our behavior. The current results further support the notion that androstadienone acts like a human modulator pheromone, namely modulating ongoing behavior or a psychological reaction to a particular context, changing stimulus sensitivity, salience and sensory-motor integration. However, these conclusions remain tentative until further replication takes place, best in ecologically valid environments. Furthermore, one has to keep in mind that the current studies could not replicate several previous findings and could not verify some hypotheses assuming communicative effects of androstadienone. Thus, the main assumption of this thesis that androstadienone is an active chemosignal is still challenged. Also, whether the term “pheromone” is indeed suitable to label androstadienone remains open.
BAKTERIELLE ENDOSYMBIONTEN DER BIENENWÖLFE Symbiontische Interaktionen zwischen verschiedenen Arten stellen allgegenwärtige und essentielle Bestandteile natürlicher Systeme dar und haben wahrscheinlich die Evolution jedes rezenten Lebewesens beeinflusst. Insekten als die diverseste Metazoen-Klasse der Erde profitieren von dem außerordentlichen metabolischen Potenzial vieler Mikroorganismen in einer großen Anzahl mutualistischer Assoziationen. Die große Mehrheit der bisher untersuchten Symbiosen zwischen Insekten und Mikroorganismen stellen Interaktionen dar, in denen die Wirte durch die Symbionten mit essentiellen Nährstoffen versorgt werden. Es sind jedoch auch einige Fälle bekannt, in denen symbiontische Bakterien eine wichtige Rolle für die intraspezifische olfaktorische Kommunikation spielen oder zur Verteidigung gegen Pathogene oder Parasitoide dienen. Die vorliegende Arbeit untersucht eine hoch spezialisierte Assoziation zwischen einer Grabwespen-Art, dem Europäischen Bienenwolf (Philanthus triangulum, Hymenoptera, Crabronidae), und Bakterien aus der Familie der Actinomyceten. Die bakteriellen Symbionten sind an einem einzigartigen Ort zu finden: Sie werden in den Reservoiren spezialisierter Antennendrüsen weiblicher Bienenwölfe kultiviert. Das Weibchen sezerniert vor der Eiablage große Mengen dieser Bakterien in die unterirdischen Brutkammern. Wenn die Bienewolf-Larve einige Tage später ihre Nahrungsaufnahme an den von der Mutter als Nahrungsvorrat bereitgestellten Honigbienen beendet hat, nimmt sie die Bakterien auf und spinnt sie in ihren Kokon mit ein. Dort erfüllen die Symbionten eine wichtige Funktion, indem sie den Schimmelbefall herabsetzen und dadurch die Überlebenschancen der Larve im Kokon während der langen und gefährlichen Winterruhe signifikant erhöhen. Experimente, in denen Bienenwolf-Weibchen ohne die Bakterien aufgezogen wurden, und Beobachtungen an Bienenwolf-Larven deuten darauf hin, dass die Symbionten vertikal von der Mutter an die Töchter weitergegeben werden. Vermutlich werden die Bakterien während des Schlupfes oder kurz davor vom Kokon in die Antennendrüsen-Reservoire aufgenommen. Phylogenetische Untersuchungen von Wirten und Symbionten sowie Transfer-Experimente mit den Bakterien wären notwendig, um herauszufinden, ob ein horizontaler Austausch der Symbionten zwischen verschiedenen Bienenwolf-Arten möglich ist. Genetische Analysen zeigen, dass die Symbionten einer unbeschriebenen Art der Gattung Streptomyces innerhalb der Actinomyceten angehören. 16s rDNA Primer und eine fluoreszenzmarkierte Oligonukleotid-Sonde wurden entwickelt, um die Bienenwolf-Symbionten mittels PCR und Fluoreszenz-in-situ-Hybridisierung (FISH) spezifisch nachweisen zu können. Mit Hilfe von PCR und Sequenzierungen der 16s rDNA konnten nah verwandte Endosymbionten in den Antennen von 28 Arten und Unterarten der Gattung Philanthus festgestellt werden, nicht aber in anderen Gattungen der Unterfamilie Philanthinae (Aphilanthops, Clypeadon, Cerceris), so dass die Symbiose auf die Gattung Philanthus beschränkt zu sein scheint. Phylogenetische Untersuchungen auf der Grundlage nahezu kompletter 16s rDNA-Sequenzen belegen, dass die Symbionten aller analysierten Bienenwolf- Arten eine monophyletische Gruppe innerhalb der Gattung Streptomyces bilden, was darauf hindeutet, dass die Symbiose hoch spezifisch ist und wahrscheinlich das Ergebnis einer langen Koevolution und Kospeziation darstellt. Anhand von Sequenzunterschieden zwischen den Symbionten lässt sich das Alter der Assoziation zwischen Philanthus und Streptomyces auf etwa 26-67 Millionen Jahre schätzen, was der Entstehung der Gattung Philanthus entsprechen könnte. Auf der Basis von 16s rDNA Sequenzen und Ultrastruktur-Daten wurden die Antennensymbionten der Bienenwölfe als neues Taxon ‚Candidatus Streptomyces philanthi’ beschrieben, wobei die Symbionten verschiedener Wirtsarten als Ökotypen behandelt und nach der Wirtsart benannt wurden (z.B. ‚Candidatus Streptomyces philanthi triangulum’). Wie die Bakterien von der Assoziation mit Bienenwölfen profitieren, ist noch unklar. Auf jeden Fall wird ihnen vom Wirt eine unbesetzte und wahrscheinlich konkurrenzfreie ökologische Nische in den Antennen sowie eine zuverlässige Weitergabe an die nächste Generation garantiert. Außerdem sprechen einige Hinweise für eine Versorgung der Bakterien mit Nährstoffen durch den Bienenwolf: (1) Weibchen legen manchmal mehrere Brutkammern pro Tag an und sezernieren jedes Mal große Mengen an Bakterien; die Bakterien müssen sich also in den Drüsen-Reservoiren schnell vermehren, um den Vorrat an Symbionten wieder aufzufüllen. (2) Die Reservoire sind von Typ 3-Drüsenzellen umgeben, die die Bakterien mit Nährstoffen versorgen könnten. (3) Eine der Reservoir-Wände weist eine netzartige Struktur auf, die möglicherweise den Eintritt von Hämolymphe und damit von Nährstoffen in das Reservoir zulässt. Dies wird durch chemische Analysen der Kohlenwasserstoffe in der Hämolymphe und in dem Antennendrüsen-Sekret untermauert, die sehr ähnliche Zusammensetzungen aufweisen. Die Assoziation zwischen Bienenwölfen und Streptomyceten stellt den ersten bekannten Fall einer Symbiose dar, bei der Bakterien in den Antennen von Insekten kultiviert werden, und sie repräsentiert eines von wenigen Beispielen für Actinomyceten als Symbionten von Insekten. Weitere Untersuchungen evolutionärer und ökologischer Aspekte dieser Symbiose werden wertvolle Erkenntnisse über die Bedeutung von Actinomyceten für die Pathogen-Abwehr bei Insekten liefern und könnten sogar zur Entdeckung neuer Sekundärmetabolite mit antibiotischen Eigenschaften für die Verwendung in der Humanmedizin führen. CHEMISCHE KOMMUNIKATION UND PARTNERWAHL BEIM EUROPÄISCHEN BIENENWOLF Chemische Signale stellen sowohl die älteste als auch die am weitesten verbreitete Form von Kommunikation zwischen Organismen dar. Bei Insekten spielen Pheromone eine essentielle Rolle für die intraspezifische Kommunikation, und eine Vielzahl aktueller Untersuchungen belegt die Bedeutung olfaktorischer Signale für die Balz und Paarung. Die meisten dieser Studien konzentrieren sich jedoch auf Weibchen-Pheromone, während von Männchen produzierte Pheromone trotz ihrer ökologischen und evolutionären Bedeutung für die Partneranlockung und Partnerwahl bisher wenig Beachtung gefunden haben. Männchen des Europäischen Bienenwolfes etablieren und verteidigen Territorien, die sie mit einem Kopfdrüsen-Sekret markieren. Dieses Sekret wirkt höchstwahrscheinlich als ein Sex- Pheromon und lockt paarungsbereite Weibchen an. Da Männchen-Territorien meist aggregiert in der Nähe von Weibchennestern auftreten, haben die Weibchen die Möglichkeit, zwischen verschiedenen potenziellen Paarungspartnern zu wählen. Die chemischen Analysen der vorliegenden Arbeit zeigen, dass die Zusammensetzung und Menge des männlichen Markierpheromons vom Verwandtschaftsgrad, der Herkunft, dem Alter und der Größe der Männchen abhängen. Das Pheromon beinhaltet demnach Informationen über eine Vielzahl von Eigenschaften der Männchen, die für die Weibchenwahl von Bedeutung sein könnten. Sowohl die genetische Distanz („optimal outbreeding“) als auch die allgemeine genetische Qualität („good genes“) eines Männchens könnte die Partnerwahl der Bienenwolf-Weibchen beeinflussen. In dieser Arbeit für den Europäischen Bienenwolf entwickelte polymorphe Mikrosatelliten legen den Grundstein für Vaterschaftsanalysen und ermöglichen so die Durchführung und Auswertung von Experimenten zur Weibchenwahl bei dieser Art.
A hitherto unresolved problem is how workers are prevented from reproducing in large insect societies. The queen informs about her fertility and health which ensures sufficient indirect fitness benefits for workers. In the ant Camponotus floridanus, I found such a signal located on eggs of highly fertile queens. Groups of workers were regularly provided with different sets of brood. Only in groups with queen eggs workers refrain from reproducing. Thus, the eggs seem to inform the workers about queen presence. The signal on queen eggs is presumably the same that enables workers to distinguish between queen and worker-laid eggs, latter are destroyed by workers. Queen and worker-laid eggs differ in their surface hydrocarbons in a similar way as fertile queens differ from workers in the composition of their cuticular hydrocarbons. When I transferred hydrocarbons from the queen cuticle to worker eggs the eggs were no longer destroyed, indicating that they now carry the signal. These hydrocarbons thus represent a queen signal that regulates worker reproduction in this species. But the signal is not present in all fertile queens. Founding queens with low egg-laying rates differ in the composition of cuticular hydrocarbons from queens with high productivity. Similar differences in the composition of surface hydrocarbons were present on their eggs. The queen signal develops along with an increasing fertility and age of the queen, and this is perceived by the workers. Eggs from founding queens were destroyed like worker eggs. This result shows that founding queens lack the appropriate signal. In these little colony foundations chemical communication of queen status may not be necessary to prevent workers from reproducing, since workers may benefit more from investing in colony growth and increased productivity of large colonies rather than from producing male eggs in incipient colonies. If the queen is missing or the productivity of the queen decreases, workers start laying eggs. There is some evidence from correlative studies that, under queenless conditions, worker police each other because of differences in individual odors as a sign of social status. It can be expressed as either aggressive inhibition of ovarian activity, workers with developed ovaries are attacked by nest-mates, or destruction by worker-laid eggs. I found that in C. floridanus workers, in contrast to known studies, police only by egg eating since they are able to discriminate queen- and worker-laid eggs. Workers with developed ovaries will never attacked by nest-mates. This is further supported by qualitative and quantitative differences in the cuticular hydrocarbon profile of queens and workers, whereas profiles of workers with and without developed ovaries show a high similarity. I conclude that workers discriminate worker eggs on the basis of their hydrocarbon profile, but they are not able to recognize egg-laying nest-mates. Improving our knowledge of the proximate mechanisms of the reproductive division of labor in evolutionary derived species like C. floridanus will help to understand the evolution of extreme reproductive altruism involving sterility as a characteristic feature of advanced eusocial systems.
Darwin’s theory of sexual selection explains the evolution of flamboyant male traits through female choice. It does not, however, address the question why males typically court and females choose. This asymmetry is now thought to be the result of the dichotomy in reproductive expenditures: Females invest primarily in parental care and males invest predominantly in mate attraction or competition. Based on this view, several hypotheses for the origin and maintenance of female preferences have been proposed. They include the classical sexual selection models, i.e. female choice for direct and indirect benefits as well as the more recent concepts of female choice for genetic compatibility and receiver bias models. The complementary choice scenario assumes that females choose mates with regard to genetic compatibility. The receiver bias concept views male traits and female preferences within the framework of communication theory and encompasses various more or less distinct models, two of which are sensory exploitation and sensory trap. Both models postulate that male signals evolved in response to pre-existing perceptual biases of females. The sensory trap hypothesis additionally emphasizes that pre-existing female preferences for certain cues evolved in non-sexual contexts, like e.g. foraging. Males that mimic these cues and elicit a favourable out-of-context response by females may increase their reproductive success. This thesis examines the evolution of the pheromone communication in the European Beewolf Philanthus triangulum. Beewolf females are specialized hunters of honeybees and provision their progeny with paralyzed prey. Male beewolves establish and scent mark territories with a pheromone from a head gland to court females. The concordant occurrence of the otherwise rare alcohol (Z)-11-eicosen-1-ol in the male pheromone and in the alarm pheromone of honeybees, the exclusive prey of the females, suggests a sensory trap process as an explanation for the evolution of the male pheromone in P. triangulum. According to this hypothesis, we tested three predictions: First, foraging honeybees should emit eicosenol. Via chemical analysis we could show that honeybee workers in fact smell of eicosenol during foraging. The occurrence of eicosenol on the cuticle and in the headspace of honeybees is a new finding. Second, beewolf females should use eicosenol as a cue for prey detection or identification. Using behavioural assays, we demonstrated that prey recognition in beewolf females is accomplished by olfactory cues and that eicosenol is an essential cue in this process. The sensory sensitivity of beewolf females to eicosenol must be extremely high, since they perceive the trace amounts present in the head space of honeybees. This sensitivity may be due to specialized olfactory receptors on the antennae of beewolf females. An inventory of the flagellar sensilla of both sexes showed that females carry one type of sensillum that is missing in males, the large sensillum basiconicum. This chemo-sensitive sensillum most likely plays a role in prey recognition. The third prediction is that beewolf males incorporate bee-like substances, including eicosenol, into their pheromone, and possibly catch females in a sensory trap. A reanalysis of the male pheromone revealed, among others, eicosenol and several alkanes and alkenes as pheromonal compounds. Our own analyses of the chemical profiles of honeybee workers and beewolf pheromone disclosed a surprisingly strong resemblance between the two. Eight of the eleven substances of the male pheromone are also present on the cuticle and in the headspace of honeybees. Notwithstanding this similarity, the male pheromone does not function as a sensory trap for females. Nevertheless, the extensive congruence between the odour bouquets of the females’ prey and the male pheromone strongly suggests that the male signal evolved to exploit a pre-existing female sensory bias towards bee odour, and, thus represents a case of sensory exploitation. In addition to the above described scenario concerning mostly the ‘design’ of the male pheromone, we addressed possible indirect benefits female beewolves may gain by basing their mating decisions on signal ‘content’. We show that the pheromone of male beewolves varies between families and may, thus, contain information about the degree of relatedness between the female and a potential mate. Females could use this information to choose genetically complementary males to avoid inbreeding and the production of infertile diploid sons. Collectively, our results provide strong evidence for a receiver bias process in the evolution of the male pheromone of P. triangulum. They further indicate that the pheromone composition may subsequently have been influenced by other natural or sexual selection pressures, like e.g. complementary female choice.
In the various groups of social bees, different systems of communication about food sources occur. These communication systems are different solutions to a common problem of social insects: efficiently allocating the necessary number of workers first to the task of foraging and second to the most profitable food sources. The solution chosen by each species depends on the particular ecological circumstances as well as the evolutionary history of that species. For example, the outstanding difference between the bumble bee and the honey bee system is that honey bees can communicate the location of profitable food sources to nestmates, which bumble bees cannot. To identify possible selection pressures that could explain this difference, I have quantified the benefits of communicating location in honey bees. I show that these strongly depend on the habitat, and that communicating location might not benefit bees in temperate habitats. This could be due to the differing spatial distributions of resources in different habitats, in particular between temperate and tropical regions. These distributions may be the reason why the mostly temperate-living bumble bees have never evolved a communication system that allows them to transfer information on location of food sources, whereas most tropical social bees (all honey bees and many stingless bees) are able to recruit nestmates to specific points in their foraging range. Nevertheless, I show that in bumble bees the allocation of workers to foraging is also regulated by communication. Successful foragers distribute in the nest a pheromone which alerts other bees to the presence of food. This pheromone stems from a tergite gland, the function of which had not been identified previously. Usage of a pheromone in the nest to alert other individuals to forage has not been described in other social insects, and might constitute a new mode of communicating about food sources. The signal might be modulated depending on the quality of the food source. Bees in the nest sample the nectar that has been brought into the nest. Their decision whether to go out and forage depends not only on the pheromone signal, but also on the quality of the nectar they have sampled. In this way, foraging activity of a bumble bee colony is adjusted to foraging conditions, which means most bees are allocated to foraging only if high-quality food sources are available. In addition, foraging activity is adjusted to the amount of food already stored. In a colony with full honeypots, no new bees are allocated to foraging. These results help us understand how the allocation of workers to the task of food collection is regulated according to external and internal nest conditions in bumble bees.
Division of reproductive labour in societies represents a topic of interest in evolutionary biology at least since Darwin. The puzzle of how helpers can be selected for, in spite of their reduced fertility has found an explanation in the kin selection theory: workers can overcome the cost of helping and of forgiving direct reproduction by rearing sufficiently related individuals. However, in the Hymenoptera, little is known on the proximate mechanisms that regulate the division of labour in colonies. Our knowledge is based on several "primitive" ants from the subfamily Ponerinae and two highly eusocial Hymenoptera species. In the former, the dominance hierarchies allowing for the establishment of individuals as reproductives are well understood. In contrast, the pheromonal mechanisms that help maintain their reproductive status are not understood. Similarly in "higher" ants, pheromonal regulation mechanisms of worker reproduction by queens remain largely unknown. The aim of this study is to determine the modalities of production, distribution and action, as well as the identity of the queen pheromones affecting worker reproduction in the ant Myrmecia gulosa. This species belongs to the poorly studied subfamily Myrmeciinae, which is endemic to the Australian region. The subfamily represents, together with the Ponerinae, the most "primitive" ants: their morphology is close to that of the hypothetical ancestor of ants, and the specialisation of queens is weaker than that of "higher" ants. Simple regulation mechanisms were therefore expected to facilitate the investigation. The first step in this study was to characterise the morphological specialisation of queens and workers, and to determine the differences in reproductive potential associated with this specialisation. This study contributes to our understanding of the link between regulation of division of reproductive labour and social complexity. Furthermore, it will help shed light on the reproductive biology in the poorly known subfamily Myrmeciinae. Queens were recognised by workers on the basis of cuticular as well as gland extracts or products. What is the exact function of the multiple pheromones identified and how they interact remains to be determined. This could help understand why queen "signal" in a "primitive" ant with weakly specialised queens such as M. gulosa appears to be as complex as in highly eusocial species. Primer pheromones act on workers? physiology and have long-term effect. Whether workers of M. gulosa reproduce or not is determined by the detection of a queen pheromone of this type. Direct physical contact with the queen is necessary for workers to detect this pheromone. Thus, the colony size of M. gulosa is compatible with a simple system of pheromone perception by workers based on direct physical contact with the queen. When prevented from establishing physical contact with their queen, some workers start to reproduce and are policed by nestmates. The low volatility of the cuticular hydrocarbons (CHCs), their repartition over the entire cuticle and the existence of queen and worker specific CHC profiles suggest that these chemicals constitute a queen pheromone. Importance of HC versus non-HC compounds was confirmed by bioassaying purified fraction of both classes of chemicals. This study demonstrates for the first time that purified HCs indeed are at the basis of the recognition of reproductive status. This supports the idea that they are also at the basis of the recognition of queens by their workers. As CHCs profiles of workers and queens become similar with acquisition of reproductive status, they represent honest fertility markers. These markers could be used as signals of the presence of reproductives in the colonies, and represent the basis of the regulation of division of reproductive labour.