Functional evolution of a bark beetle odorant receptor clade detecting monoterpenoids of different ecological origins

Abstract

Insects detect odors using an array of odorant receptors (ORs), which may expand through gene duplication. How specificities evolve and new functions arise in related ORs within a species remain poorly investigated. We addressed this question by functionally characterizing ORs from the Eurasian spruce bark beetle Ips typographus, in which antennal detection and behavioral responses to pheromones, volatiles from host and non-host trees, and fungal symbionts are well described. In contrast, knowledge of OR function is restricted to two receptors detecting the pheromone compounds (S)-(-)-ipsenol (ItypOR46) and (R)-(-)-ipsdienol (ItypOR49). These receptors belong to a species-specific OR-lineage comprising seven ItypORs. To gain insight into the functional evolution of related ORs, we characterized the five remaining ORs in this clade, using Xenopus oocytes. Two receptors responded primarily to the host tree monoterpenes (+)-3-carene (ItypOR25) and p-cymene (ItypOR27). Two receptors responded to oxygenated monoterpenoids produced in relatively large amounts by the beetle-associated fungi, with ItypOR23 specific for (+)-trans-(1R,4S)-4-thujanol, and ItypOR29 responding to (+)-isopinocamphone and similar ketones. ItypOR28 responded to the pheromone E-myrcenol from the competitor Ips duplicatus. Overall, the OR responses match well with those of previously characterized olfactory sensory neuron classes except that neurons detecting E-myrcenol have not been identified. The characterized ORs are under strong purifying selection and demonstrate a shared functional property in that they all primarily respond to monoterpenoids. The variation in functional groups among OR ligands and their diverse ecological origins suggest that neofunctionalization has occurred early in the evolution of this OR-lineage following gene duplication.