Ectomycorrhizal fungi (EcMF) form diverse communities and link different host plants into mycorrhizal networks, yet little is known about the magnitude of mycobiont diversity of a single tree individual. This study addresses species richness and spatial structure of EcMF in the root system of a single European aspen (Populus tremula) individual in an old-growth boreal mixed forest ecosystem in Estonia. Combining morphological and molecular identification methods for both plant and fungi, 122 species of EcMF were recovered from 103 root samples of the single tree. Richness estimators predicted the total EcMF richness to range from 182 to 207 species, reflecting the observation of 62.3% singletons and doubletons within the community. Fine-scale genetic diversity in Cenococcum geophilum indicates the presence of 23 internal transcribed spacer genotypes. EcMF community was significantly spatially autocorrelated only at the lineage level up to 3 m distance, but not at the species level. Proximity of other hosts had a significant effect on the spatial distribution of EcMF lineages. This study demonstrates that a single tree may host as many EcMF species and individuals as recovered on multiple hosts in diverse communities over larger areas.