Phenotypic Diversity And Analysis Of 16S RDNA RFLP On Rhizobia Isolated From Leguminous Plants In Some Mining Areas Of Northwest China

An aim of this experiment is to study the phylogeny of rhizobial strains, phenotypic diversity and the relationship between their resistance to heavy metals and phylogeny diversity. Thirty-eight rhizobial strains were isolated from the leguminous plants which grew in some mining areas of Northwest China. The genetic diversity of these strains was analyzed by using 16S rDNA PCR-RFLP and the complete 16S rDNA sequences of the representative strains to establish the phylogenetic tree. Their 119 physiological and biochemical characteristics were tested, including salt-tolerance, growth of the initial pH, growth temperature range, abilities of using single carbon and nitrogen source and resistance to antibiotic and chemical dyes.And response to increasing levels of Zn, Hg, Cu, Cd, Pb in media by these strains was observed. The result of the genetic diversity analysis that these strains belonged to Sinorhizobium, Rhizobium and Agrobacterium based on 16S rDNA phylogenetic tree. Nine strains were selected as representative in the phylogenetic tree. The majority of Soybean rhizobia and Vicia rhizobia belonged to S. fredii and R. leguminosarum, and Medicago lupulina rhizobia belonged to S. meliloti and S. xinjiangense. The similarity of CCNWSX1294 and R. sullae was 98.83%, having 13 base differences. CCNWSX1294 may be a potential new species. It is shown that 57.9% of all these strains could resist 5% NaCl. All strains are able to grow on the YMA medium with pH 9¹2. They had strong salinity and alkali tolerance capacity. 26.3% of strains could tolerate low temperature, and all strains had the characteristic of tolerating instantaneous high temperature. These strains could make extensive use of nitrogen and carbon sources. Different strains had different tolerance to varieties of heavy-metals, antibiotic and chemical dyes.The two strains CCNWSX1277 and CCNWSX1294 survived at Zn²⁺ (2.0 mmol·L^-1). Only one of the nine strains, CCNWSX1277 survived at Hg²⁺ (0.25 mmol·L^-1). Most of the nine strains survived at Cu²⁺ (1.6 mmol·L^-1). Only three representative strains could survive any exposure to Cd²⁺ of which CCNWSX1277 could survive at highest level (1.4 mmol·L^-1 Cd²⁺). All nine of these strains survived at 2.5 mmol·L^-1 Pb²⁺. Two representative strains, both belonging to Agrobacterium had strong and stable tolerance to all five kinds of heavy metals. But four representative strains belonging to Rhizobium and three representative strains belonging to Sinorhizobium had different tolerance to all five kinds of heavy metals. Overall, the genus order of the tolerance to all five heavy metals was Agrobacterium >Rhizobium>Sinorhizobium.