Entomologia Experimentalis et Applicata 83: 87–92, 1997. 87 c 1997 Kluwer Academic Publishers. Printed in Belgium. Discrimination of oilseed rape volatiles by the honeybee: combined chemical and biological approaches M. H. Pham-Del` egue 1 , M. M. Blight 2 , V. Kerguelen 1 , M. Le M´ etayer 1 , F. Marion-Poll 3 , J. C. Sandoz 1 & L. J. Wadhams 2 1 Laboratoire de Neurobiologie Compar´ ee des Invert´ ebr´ es, INRA, BP 23, 91440 Bures-sur-Yvette, France; 2 Department of Biological and Ecological Chemistry, IACR, Rothamsted, Harpenden, Hertfordshire, AL5 2JQ, England; 3 Station de Phytopharmacie, INRA, Route de St Cyr, 78026 Versailles cedex, France Accepted: December 12, 1996 Key words: honeybees, oilseed rape, volatiles, olfactory discrimination, learning Abstract Honeybees (Apis mellifera L.) were individually subjected to a classical conditioning procedure in order to obtain an olfactory conditioned proboscis extension response. To relate the behavioural response directly to antennal detection abilities, a technique was developped for coupling proboscis extension responses and electroantennogram recordings, with the stimulation being provided by the effluent of a gas chromatograph (GC). Bees were conditioned with a six-component mixture being part of oilseed rape (Brassica napus L.) floral volatiles, and tested with the indi- vidual components separated by GC. Responses of the conditioned bees were compared to those of unconditioned bees. No behavioural response was obtained in the control group, neither to the individual components nor to the mixture. Conditioning induced behavioural responses for three components, and an increase of electroantennogram responses for all components. A second experiment was conducted with an air entrainment extract of oilseed rape flower volatiles. Behavioural responses of conditioned and unconditioned bees were recorded. Responses obtained from conditioned bees tested with the air entrainment extract showed six groups of behaviourally active GC peaks. Unconditioned bees showed the same pattern of responses but at a lower level. The coupled technique described here appears to be a reliable tool for locating active components in a synthetic as well as in a natural mixture of floral volatiles. The effects of conditioning on odour discrimination and on its sensory correlates are discussed. Introduction Honeybee foragers visit plants to collect nectar and/or pollen for their own energetic needs and to provide the colony with food. Orientation to food sources is mainly mediated by floral volatiles, olfactory signals being particularly well learnt by honeybees compared to visual cues (Kriston, 1973; Menzel, 1984). Plant volatiles are complex chemical blends which may fluc- tuate according to species, varieties, or phenology (e.g. in sunflower: Eti´ evant, et al., 1984; Pham-Del` egue et al., 1989). Now honeybees are able to select appropri- ate floral sources by adapting their behaviour to chan- ging stimuli. Complex odour recognition might rely on the discrimination of a limited range of relevant com- ponents, as shown with free-flying bees visiting food sources scented with alfalfa volatiles (Waller et al., 1974) or sunflower volatile extracts (Pham-Del` egue et al., 1986). To contribute to the understanding of natur- al odour recognition in the honeybee, we investigated honeybee responses to oilseed rape flower volatiles. Oilseed rape was chosen as a model since it is a crop plant intensively visited by honeybees which are attrac- ted by its abundant production of nectar and pollen (Mesquida et al., 1988). Previous work on sunflower volatile blends recog- nition showed that behaviourally active components could differ from the range of components detected at the antennal level (Thi´ ery et al., 1990). Vareschi (1971), and more recently Akers & Getz (1992, 1993), have categorized individual components according to the selectivity of receptor neurons and behavioural