Honeybees Learn Odour Mixtures via a Selection of Key Odorants

Abstract

BackgroundThe honeybee has to detect, process and learn numerous complex odours from her natural environment on a daily basis. Most of these odours are floral scents, which are mixtures of dozens of different odorants. To date, it is still unclear how the bee brain unravels the complex information contained in scent mixtures.Methodology/Principal FindingsThis study investigates learning of complex odour mixtures in honeybees using a simple olfactory conditioning procedure, the Proboscis-Extension-Reflex (PER) paradigm. Restrained honeybees were trained to three scent mixtures composed of 14 floral odorants each, and then tested with the individual odorants of each mixture. Bees did not respond to all odorants of a mixture equally: They responded well to a selection of key odorants, which were unique for each of the three scent mixtures. Bees showed less or very little response to the other odorants of the mixtures. The bees' response to mixtures composed of only the key odorants was as good as to the original mixtures of 14 odorants. A mixture composed of the other, non-key-odorants elicited a significantly lower response. Neither an odorant's volatility or molecular structure, nor learning efficiencies for individual odorants affected whether an odorant became a key odorant for a particular mixture. Odorant concentration had a positive effect, with odorants at high concentration likely to become key odorants.Conclusions/SignificanceOur study suggests that the brain processes complex scent mixtures by predominantly learning information from selected key odorants. Our observations on key odorant learning lend significant support to previous work on olfactory learning and mixture processing in honeybees.

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“…Three groups of 60 foragers each were conditioned using 12 trials over 2 days, with six trials per day. Group one was conditioned with a mixture of 14 common floral odorants (floral mix) 20 , group two with the floral odour linalool and group three was exposed to air (no odour learning), thereby serving as baseline control. This procedure provided a focused olfactory learning experience for the bees of the first two groups, and ensured formation of a robust LTM.…”

“…Nonetheless, when we compare bees trained to single odour (linalool) with bees trained to a floral mix (14 odours) using the PER assay, we see a similar qualitative pattern of molecular regulation compared with the common control irrespective of conditioning treatment (Supplementary Data 1). Linalool is part of the floral mix, and we know that honeybees learn complex scent mixtures based on a reduced number of key odorants 20 , with linalool possibly being one of the key odorants for the mix we used. Hence, floral mix and linalool treatments may indeed elicit a similar qualitative pattern of molecular response in the brain.…”

“…3a). This increase could be due to more odorants being learnt during conditioning to floral mix 20 compared with the single odour, and the mixture being perceived as a much greater or more concentrated stimulus than linalool alone.…”

“…Foraging honeybees (Apis mellifera) were captured at the entrance of an outdoor hive located on campus of the University of Queensland, Brisbane, Australia. Bees were cold-anaesthetized and harnessed in plastic tubes for the proboscis extension response (PER) assay as previously described 20,62 . Only bees that were active and extended their proboscis in response to sugar were used.…”

“…Odours were delivered using a custom-built olfactometer 20 . Odours were either a mixture of 14 common floral odorants diluted 1:10 in hexane (floral mix, composition and odorant origin as previously described 20 , or linalool diluted 1:10 in hexane (Sigma)). The specific conditioning procedure varied according to experiment, as described below.…”