In vitro host relationships of ectomycorrhizal Tricholoma kakishimeji and closely related species reflect their habitat characteristics.

Tricholoma kakishimeji, a poisonous fungus containing the toxic compound ustalic acid, has sometimes been misidentified as closely related species (T. stans, T. matsushimeji, T. kakishimejioides) under the name T. ustale in Japan until recently. Tricholoma ustale s. str. was not found in Japan according to a recent study, and it has been only recorded in Europe. Here, we report the first comprehensive morphological comparison of ectomycorrhizae among these four Tricholoma species. Several cultured strains of these species were inoculated onto Pinus densiflora in vitro. The resulting ectomycorrhizal pine seedlings were subsequently used as mother plants to establish an ectomycorrhizal system on Fagaceae plants. Although all tested fungal strains formed ectomycorrhizae on pine, mycorrhizal colonization by T. kakishimejioides was limited. On Quercus hosts, T. matsushimeji exhibited discontinuous Hartig net development, whereas T. kakishimeji and T. stans produced distinct Hartig nets. Additionally, ectomycorrhizal biomass development on oak hosts was limited in T. stans and T. matsushimeji. These findings correspond to the habitat characteristics of these fungal species. Ectomycorrhizae of these Tricholoma species sampled from natural forests showed morphological and anatomical characteristics similar to their in vitro ectomycorrhizae, including species-specific hyphal arrangements of the mantle and rhizomorphs. We propose that the ectomycorrhizal specificity of Tricholoma can be experimentally assessed in relation to their genetic background on pine and oak hosts, as well as the phyloecological characteristics of these fungal species.