Eurasian spruce bark beetle detects anti-attractant lanierone using a highly expressed specialist odorant receptor, present in several functional sensillum types

Background. Insects detect odours using odorant receptors (ORs) expressed in olfactory sensory neurons (OSNs) in the antennae. Odours important to fitness are believed to be detected by selective and abundant OSNs; hence, ORs with high antennal expression. However, little is known about the function of highly expressed ORs in beetles, since few ORs have been functionally characterized. Here, we aimed to functionally characterize the most highly expressed OR (ItypOR36) in the Eurasian spruce bark beetle Ips typographus L. (Coleoptera, Curculionidae, Scolytinae), a major pest of spruce trees. We hypothesized that this OR would detect a compound important to beetle fitness, such as a pheromone component. We next investigated the antennal distribution of this OR using single sensillum recordings (SSR) and in situ hybridization, followed by field- and laboratory experiments to evaluate the behavioural effect of the discovered ligand. Results. We expressed ItypOR36 in HEK293 cells and challenged it with a large panel of ecologically important odours. The OR responded exclusively to the monoterpene-derived ketone lanierone with high sensitivity. Lanierone is used in chemical communication in North American Ips species but had never been studied in relation to the ecology and sensory physiology of I. typographus. Single sensillum recordings revealed a novel and abundant lanierone-responsive OSN class with the same response profile as ItypOR36. These OSNs were co-localized in sensilla together with seven different previously described OSN classes, suggesting a promiscuous OSN co-localization principle in bark beetles. We further revealed lanierone to be a potent anti-attractant, strongly reducing attraction of I. typographus to its aggregation pheromone in the field at low release rates. Conclusions. Our study highlights the importance of the so-called ‘reverse chemical ecology’ approach to identify novel semiochemicals for ecologically important insect species. Our finding that OSN co-localization is not stereotypic as in Drosophila suggests fundamental organisational differences in the peripheral olfactory sense between insect orders. The lack of evidence that I. typographus produces lanierone and its anti-attractant effect imply that lanierone is used in interspecific pheromone inhibition and that the compound is a good candidate for incorporation in semiochemical-based protection of susceptible spruce stands.