Updates on fungal communities associated with Tomicus destruens: Insights into potential contributors to pine decline in Mediterranean forests

Among the bark beetles threatening Mediterranean pine ecosystems, Tomicus destruens (Wollaston) (Coleoptera: Curculionidae: Scolytinae) stands out as one of the major pests. Its life cycle involves an often lethal reproductive phase under the bark of stressed pines and a maturation feeding phase in the shoots of both healthy and stressed individuals; the latter phase being a potential precursor to the former. The ecological impact of T. destruens is further heightened by its association with several fungi, which colonize plant tissues and contribute to the decline of pine stands. This study is an update on the fungal communities associated with T. destruens, assessing the potential role of insect-fungi associations in pine decline processes. To assess the diversity of taxa, fungal communities were isolated from 50 T. destruens adults, collected during the maturation phase from the shoots of 50 stone pines located in the Maremma Regional Park (Tuscany, Italy). The fungal taxa were isolated in pure culture and molecular characterization was performed following a multilocus DNA barcoding approach. End-point PCR was performed on extracted DNA, targeting the internal transcribed spacers (ITS) of ribosomal DNA. For finer taxonomic resolution, additional regions such as the elongation factor gene (TEF1) and fragments of the beta-tubulin gene (TUB2) were amplified and sequenced. The obtained sequences were compared with those deposited in international reference databases (GenBank, UNITE) for species assignments. This study contributes to the ongoing characterization of fungal communities associated with T. destruens, aiming to better define the dynamics between these fungi and their host trees. Preliminary results indicate a diverse array of fungal taxa, providing a basis for further taxonomic and functional studies. These findings supplement our current knowledge of the ecological significance of such associations and highlight areas for future investigation into Mediterranean forest health.