The bacterial diversity of the Tuber indicum×Pinus yunnanensis mycorrhizosphere was studied. The mycorrhizosphere in T. indicum×Pinus yunnanensis was collected in the field and the culturable bacteria were purified from the soil. The 16S rRNA genes of the purified bacteria were sequenced and a phylogenetic tree was constructed to analyze the population composition and structure of the culturable bacteria. Roche 454 high-throughput sequencing was used to determine the population composition and structure of all bacteria in the soil. (1) 793 culturable bacteria, representing 3 geni and 61 OTUs (Operational Taxonomic Units), were purified from the mycorrhizosphere. OTUs of the genus Pseudomonas accounted for 86% of the total OTUs, OTUs of the genus Acinetobacter accounted for 9.8% of the total OTUs, and OTUs of the genus Streptomyces accounted for 6.5% of the total OTUs. Pseudomonas was the predominant culturable bacteria in the mycorrhizosphere of T. indicum×Pinus yunnanensis. (2) 8937 sequences of the bacterial V1-V3 region, representing 2073 OTUs, were analyzed by using Roche 454 high-throughput sequencing. Proteobacteria, Actinobacteria, and Acidobacteria were the three dominant phylums, which accounted for 65.9% of the total OTUs. Flavobacterium, Rhizobium, and Pseudoxanthomona were the dominant geni, which accounted for 33% of the total OTUs. The population composition of culturable bacteria of T. indicum×P. yunnanensis mycorrhizosphere was simple; the dominant genus was Pseudomonas. The bacterial diversity of T. indicum×P. yunnanensis mycorrhizosphere was very high; the bacterial population was complex.