Soil samples were collected at an altitude of 500, 1,060, 1,500, 1,950, 2,400 and 3,100m, respectively, from Shennongjia, a forest reserve in Hubei province (central China). Their corresponding pHs were 5.50, 4.91, 5.64, 5.28, 5.49 and 4.60. By using a plant trap method, a total of 25 soybean rhizobia were isolated from the soil above an altitude of 1,500 m and all identified to be Sinorhizobium fredii. Their genetic biodiversity was characterized by 16S-23S rDNA internally transcribed spacer (ITS) region polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and random amplification DNA (RAPD) analysis. All the tested strains produced a 2.1 kb 16S-23S rDNA ITS fragment. After digestion with three restriction endonucleases (HaeⅢ, MspI and CfoI), respectively, great variations in 16S-23S rDNA ITS PCR-RFLP patterns were observed. The tested strains could be differentiated into 11 ITS genotypes. The genotypes of rhizobia were not related to geographical location. Twelve primers were applied to RAPD analysis and a dendrogram was obtained, showing that all the strains (including reference strain S. fredii USDA205) were divided into two diverging groups. Moreover, each group could be further divided into two subgroups. Both RAPD and 16S-23S rDNA ITS PCR-RFLP analysis indicated that a high degree of genetic diversity existed among S. fredii strains isolated from Shennongjia virgin soils. Since Shennongjia is an unexploited forest region in central China and the gene centre of soybean is located in China, the symbiotic genes harboured by these strains may be of great importance and the rich diversity of these strains might contribute to the adaptation of soybean to an alpine environment.