Metagenomic characterisation of fungal communities associated with Scots pine bark beetles: mechanisms of selective antagonism and monoterpene tolerance

Bark beetle-associated fungi contribute to beetle nutrition, detoxification, and interactions with conifer hosts; however, their composition and function across development and environments remain poorly understood. We characterised fungal communities of two pine-feeding Ips bark beetles, I. sexdentatus and I. acuminatus, in larval, pupal, and adult stages, and in wild versus laboratory populations, using high-throughput ITS2 amplicon sequencing combined with qPCR and functional assays. Both beetles harboured a stable core mycobiome dominated by Kuraishia, Ogataea, Ophiostoma, Graphilbum and Cyberlindnera, while adults showed species-specific differences and wild beetles, especially I. acuminatus, exhibited greater diversity than laboratory populations. Beetles shared more taxa with unfed control wood than with gallery wood, indicating acquisition during feeding and concurrent restructuring of the wood mycobiome. Monoterpene bioassays on beetle-associated yeasts revealed that mixtures of -pinene, 3-carene and terpinolene suppressed growth more strongly than single compounds, suggesting synergistic inhibition. Yeasts selectively antagonised entomopathogenic fungi and expressed complementary cell-wall-lytic and digestive activities, consistent with combined roles in pathogen suppression and plant-polymer deconstruction. Our results show that Ips mycobiomes are conserved yet dynamic across life stages and environments, and emphasise the importance of multi-terpene and interaction assays for understanding bark beetle-fungus-conifer interactions. ImportanceOur study reveals that pine-feeding bark beetles co-evolved in close association with a stable core mycobiota that supports nutrient acquisition, detoxification, and chemical signalling, while additional fungal partners shift in response to beetle development, environment, and host context. By demonstrating higher mycobiome diversity in beetle larvae and wild populations, substantial overlap between beetle- and wood-associated fungi, differential sensitivity to monoterpene blends and selective antagonism of yeasts towards entomopathogenic fungi, our study unravels key ecological and mechanistic principles shaping beetle-fungus assemblages under conifers, paving the way for more in-depth functional investigations.