Bark beetles locate fungal symbionts by detecting volatile fungal metabolites of host tree resin monoterpenes

Outbreaks of the Eurasian spruce bark beetle (Ips typographus) have decimated millions of hectares of conifer forests in Europe in recent years. The ability of these 3-6 mm long insects to kill mature trees over a short period has been ascribed to two main factors: (1) mass attacks on the host tree to overcome tree defenses and (2) the presence of fungal symbionts that support successful beetle development in the tree. While the role of pheromones in coordinating mass attacks has been well studied, the role of chemical communication in maintaining the fungal symbiosis is poorly understood. We previously demonstrated that I. typographus can recognize beneficial fungal symbionts of the genera Grosmannia, Endoconidiophora and Ophiostoma by their de novo synthesized volatile compounds. Here, we hypothesize that the fungal symbionts of the bark beetles might metabolize spruce resin monoterpenes of the beetles host tree, Norway spruce (Picea abies), and that the volatile products could be used as cues by beetles for locating breeding sites with beneficial symbionts. We show that Grosmannia penicillata and other fungal symbionts altered the profile of spruce bark volatiles by converting the major monoterpenes into an attractive blend of oxygenated derivatives. Bornyl acetate was metabolized to camphor, and - and {beta}-pinene to trans-4-thujanol and other oxygenated products. Other co-occurring non-beneficial fungi (Trichoderma sp.) also produce oxygenated monoterpenes, but in non-attractive ratios. Extensive electrophysiological measurements showed that the bark beetle possesses dedicated olfactory sensory neurons for oxygenated metabolites. Compounds such as camphor and trans-4-thujanol attracted beetles at specific doses in olfactory experiments, and the presence of symbiotic fungi enhanced attraction of females to pheromones. Finally, we show that colonization of fungal symbionts on spruce bark diet stimulated beetles to make tunnels into the diet. Collectively, our study suggests that the blends of oxygenated metabolites of conifer monoterpenes produced by fungal symbionts are used by bark beetles as attractive cues to find breeding or feeding sites containing their essential microbial symbionts. The oxygenated metabolites may aid beetles in assessing the presence of the fungus, the defense status of the host tree and the density of conspecifics at potential feeding and breeding sites.