| Twenty-six strains of rhizobia were isolated and purified from the nodules of 2 cultivars of rice bean (Vigna umbellata L.) collected from the different soils located at various ecological sites in Hubei province, China. These strains and 8 reference strains of other genus and species of rhizobia were comparatively studied by determining their biological characteristics. In addition, the phylogeny of these strains also was tentatively studied.The generation time of all strains tested was between 2.70 and 11.45 hours in YMA liquid medium, which showed great diversity. According to the result of acid-alkali endurance, salt stress, resistance of intrinsic antibiotics, utilization of carbon and nitrogen sources, the biodiversity of rhizobia isolated from rice bean was shown with the dendrogram through cluster analysis. At the similarity of 77%, all strains could be divided into 3 groups. Group I consisted of slow-growing strains. Fast-growing strains fell into groups II and III. The groupsIII were further divided into different subgroups at the similarity of 80%. Similarities of some strains clustered into a subgroup were related with geographical sampling sites.Results of host-plant range test shown that some strains of rhizobia isolated from Vigna umbellata L.could form nodules on Vigna, Arachis, Glycine and Phaseolus. it can be concluded that the rhizobia isolated from Vigna umbellata L. is not an independent cross-inoculation group.Six types of plasmid profiles of fast-growing strains were elucidated with Echardt's method. The correlation of plasmid type and isolated place of strains were analysized.The genetic diversity and phylogeny of the isolated strains were further studied by using the methodlogy of 16S rDNA PCR-RFLP and 16S-23S rDNA IGS PCR-RFLP. The Result of 16S rDNA PCR-RFLP revealed that all strains could be divided into 6 groups at the similarity of 90%. 3 strains isolated from nodules of Vigna umbellata L var. glaber in Hanchuan fell into group V and were clustered with R.leguminosarum USDA2370 at the similarity of 100%. The similarities of all strains of group VI were up to 100% with Sinorhizobium fredii USDA205 and were in a same "species" level. Other strains were divided into I, II and IV group indicated different divergences with reference strains. It can be referred that these strains maybe respectively belong to other taxonomic positions in phylogenetic branch.All the strains tested were divided into three IGS types according to size of 16S-23S rDNA. At the similarity level of 52%, all the strains isolated from nodules of rice bean could be divided into two groups. Group I was composed of slow-growing strains and could be further divided into two branches: IA and IB, at the similarity of 79%. Group II was composed of fast-growing strains also could be further divided into 2 branches, IIA and IIB,at the similarity of 62%. The branch IIA consists IIA1, IIA2 and IIA3 clusters at the similarity of 72%. The strains of branch IIB were highly related with Sinorhizobium fredii USDA205.For some large groups, the results of 16S rDNA PCR-RFLP and 16S-23S rDNA IGS PCR-RFLP were in good agreement with that of numerical analysis of the phenotypic characteristics. From the above results, it can be concluded that the rhizobia strains isolated from the nodules of rice bean have genetic diversities. 5 strains from Hanchuan, including GAP2502, HCY5105, HCY5202, HCY5503 and HCY5609, had distinct difference with other strains. It indicated that these strains could represent a new phylogenetic branch of Rhizobium. the extensive analysis for the taxonomic position is under way for verification.The partial 16S rDNA sequences of representative strains CYR4243, CYY3302, HCY1101 and HCY5202 were analysized. The results showed that 4 strains respectively fell into different phylogenetic branches. CYR4243 shared 99.87% sequence homology with Sinorhizobium fredii type strain USDA205. HCY1101 was highly related with Rhizobium leguminosarum bv.trifolii and Rhizobium leguminosarum bv.viceae, HCY5202 s...
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