Tricholoma matsutake is an ectomycorrhizal fungus that has obligate symbiotic relationships with Pinus densiflora. Its fruiting body has a distinctive flavor and is traded at a high price. Thus, it has been a significant source of income for rural communities in Korea. We hypothesized that biotic factors considerably influence the formation of the T. matsutake mushroom, and the soils producing T. matsutake share similar microbial characteristics. Therefore, the present study aimed to detect the specific fungal and bacterial groups in T. matsutake production soils (shiro+) and nonproduction soils (shiro−) of the Bonghwa and Yanyang regions via next-generation sequencing. In a total of 15 phyla, 36 classes, 234 genera of bacteria, six phyla, 29 classes, and 164 genera of fungi were detected from four samples at both sites. The species diversity of shiro+ soils was lower than the shiro− samples in both the fungal and bacterial groups. In addition, we did not find high similarities in the microbial communities between the shiro+ soils of the two regions. However, in the resulting differences between the fungal communities categorized by their trophic assembly, we found a distinguishable compositional pattern in the fungal communities from the shiro+ soils and the shiro− soils of the two sites. Thus, the similarity among the microbial communities in the forest soils may be due to the fact that the microbial communities in the T. matsutake dominant soils are closely associated with biotic factors and abiotic factors such as soil properties.