Geographic Differentiation of the Thelephora ganbajun Mycosphere Across Multiple Sites and Its Associations with Fruiting-Body Metabolic Phenotypes

Thelephora ganbajun is a highly valued edible ectomycorrhizal fungus, but the relationships between its natural distribution and growth and soil environmental conditions remain poorly understood. In this study, mycosphere soils and adjacent control soils were collected from seven geographic sites in Yunnan Province, China. Soil physicochemical properties, enzyme activities, untargeted metabolomics, and bacterial and fungal amplicon sequencing were integrated to characterize geographic variation in the belowground environment and its associations with fruiting-body metabolic phenotypes. Nutrient levels and enzyme activities differed between mycosphere and control soils, and the magnitude of these differences varied among sites. A total of 5006 metabolites were annotated, including 77 differential metabolites shared across all seven sites, although their directions of change were strongly site-dependent. Bacterial and fungal communities in mycosphere soils exhibited significant geographic differentiation. The first two axes of distance-based redundancy analysis cumulatively explained 50.04% and 56.73% of the variation in bacterial and fungal communities, respectively. Soil pH, alkali-hydrolyzable nitrogen, and available phosphorus emerged as key environmental factors, with the latter two significantly associated with both bacterial and fungal community structure. Procrustes analysis further revealed concordant geographic differentiation of the two microbial communities. The volatile, non-volatile, and free amino acid metabolic phenotypes of fruiting bodies were associated with different components of the belowground environment. These findings reveal geographic variation in the belowground environment of T. ganbajun during a single fruiting season and its associations with fruiting-body metabolic phenotypes, providing candidate environmental indicators for future habitat assessment.