The project proposes a comprehensive study of the evolution of male olfactory processing structures dedicated to sexual communication in honeybees of the genus Apis, at the species and at the subspecies level. To this goal, it will use a range of techniques including neuroanatomy, neurophysiology, molecular biology and behaviour. Honeybees constitute a prominent scientific model and a worldwide crucial agricultural agent for pollination and honey production. Although they have been intensively studied, central aspects of their mating behaviour and physiology are utterly unknown. Honeybees display striking mating behaviour, males (drones) gathering high in the air at discrete congregation areas, where they engage in scramble competition to mate with virgin queens, at which point they die. Attraction to the queen is mainly mediated by olfactory cues. Queens produce a wealth of pheromones, best studied for their role on worker physiology and behaviour, thereby maintaining the cohesion of the hive. Of these, one component, 9-ODA, was shown to attract drones. However, the existence of co-attractants is unknown. We know nevertheless that the brain of the drone harbours in its primary olfactory centre, the antennal lobe, four pheromone-processing units, the macroglomeruli (MG). Previous work carried out by the Project Coordinator using in vivo calcium imaging demonstrated that the largest MG actually processes 9-ODA information, but the role of the remaining units is unknown. Aim 1 of this project is to unravel the pheromonal communication system of the drone in the standard species, the western honeybee Apis mellifera. The genus Apis comprises 9 sympatric and allopatric species found mostly in south-east Asia. They can be classified in three main phylogenetic groups: the dwarf honeybees (2 species), the giant honeybees (2 species) and the cavity-nesting honeybees (5 species including Apis mellifera). Due to their more simple organization and cognitive abilities, the dwarf species are thought to have diverged first and the cavity-nesting bees would represent the more derived species in the bee phylogeny. Previous work has described colony organization, task allocation and foraging behaviour among others in these species. However, very little is known about the evolution of sexual communication in honeybee, although the main attractant (9-ODA) seems to be conserved in most – but not all species. Moreover, the existence of co-attractants is unknown. Cross-species comparisons of the pheromonal components suggest that the more derived cavity-nesting species present a higher number of mandibular queen pheromone components. This enrichment of the queen pheromone through evolution, could find its parallel in an apparent complexification of the pheromone-specific processing units in the drone antennal lobe. Aim 2 of this project asks how sexual pheromonal communication evolved in bees of the Apis genus. Was the observed improvement in social organization and cognitive abilities accompanied by a higher elaboration of sexual communication? Variations in sexual pheromonal communication may also take place between subspecies and play a role in ongoing speciation phenomena. The western honeybee A. mellifera is distributed on a wide geographical range, with 24 different subspecies living in very different climates from cold temperate to tropical, from permanent humidity to semi-desert. The subspecies show strong behavioural differences concerning colony defence, swarming, orientation, recruitment and foraging behaviour, especially flower constancy. By contrast, little is known about possible adaptations of their mating behaviour, and no study has studied the specifics of olfactory sexual communication and odour representation in different subspecies. Aim 3 of this project will attempt to identify race-specific modifications of the anatomy and function of the pheromone-specific units of the drone antennal lobe.