Wen Feng Chen

Unique community structure and biogeography of soybean rhizobia in the saline-alkaline soils of Xinjiang, China

To investigate the community composition and biogeography of soybean rhizobia in Xinjiang, a total of 151 strains were investigated with RFLP and phylogenetic analyses of 16S rRNA gene, 16S–23S intergenic spacer, three housekeeping genes (atpD, glnII and recA), and two symbiotic genes (nifH and nodC), as well as cross-nodulation. Two rhizobial species, Bradyrhizobium liaoningense and Sinorhizobium fredii, were found as dominant groups in communities of soybean rhizobia in Xinjiang, whereas three Rhizobium genomic species, B. yuanmingense and B. japonicum, were minor groups. These genomic species showed clear correlations with eco-regions, and their symbiotic genes were identical or very similar to those of the reference strains for the corresponding species. Conclusively, the dominant soybean rhizobia S. fredii and B. liaoningense in Xinjiang might be introduced from other Chinese regions, but they have been selected as the rhizobia adapted to the saline-alkaline soils. The high pH, salinity, and phosphate concentration in soil might be the environmental factors determining the biogeography of these bacteria. It is worth mentioning that a novel Rhizobium species that may have acquired the symbiotic genes from a Bradyrhizobium lineage was identified.

Diversity and Biogeography of Rhizobia Isolated from Root Nodules of Glycine max Grown in Hebei Province, China

A total of 215 rhizobial strains were isolated and analyzed with 16S rRNA gene, 16S–23S intergenic spacer, housekeeping genes atpD, recA, and glnII, and symbiotic genes nifH and nodC to understand the genetic diversity of soybean rhizobia in Hebei province, China. All the strains except one were symbiotic bacteria classified into nine genospecies in the genera of Bradyrhizobium and Sinorhizobium. Surveys on the distribution of these rhizobia in different regions showed that Bradyrhizobium japonicum and Bradyrhizobium elkanii strains were found only in neutral to slightly alkaline soils whereas Bradyrhizobium yuanmingense, Bradyrhizobium liaoningense-related strains and strains of five Sinorhizobium genospecies were found in alkaline–saline soils. Correspondence and canonical correspondence analyses on the relationship of rhizobial distribution and their soil characteristics reveal that high soil pH, electrical conductivity, and potassium content favor distribution of the B. yuanmingense and the five Sinorhizobium species but inhibit B. japonicum and B. elkanii. High contents of available phosphorus and organic matters benefit Sinorhizobium fredii and B. liaoningense-related strains and inhibit the others groups mentioned above. The symbiotic gene (nifH and nodC) lineages among B. elkanii, B. japonicum, B. yuanmingense, and Sinorhizobium spp. were observed in the strains, signifying that vertical gene transfer was the main mechanism to maintain these genes in the soybean rhizobia. However, lateral transfer of symbiotic genes commonly in Sinorhizobium spp. and rarely in Bradyrhizobium spp. was also detected. These results showed the genetic diversity, the biogeography, and the soil determinant factors of soybean rhizobia in Hebei province of China.

Diverse rhizobia associated with soybean grown in the subtropical and tropical regions of China

Aiming at investigating the species composition and the association between ribosomal/housekeeping genes and symbiotic genes of rhizobia nodulating with soybean grown in the subtropical and tropic regions of China, a total of 252 rhizobial strains isolated from five eco-regions was characterized. Four genomic groups, Bradyrhizobium japonicum complex (including B. liaoningense, B. japonicum and a B. japonicum related genomic species) and B. elkanii as the major groups, B. yuanmingense and Sinorhizobium fredii as the minor groups, were identified by the ribosomal/housekeeping gene analyses. The symbiotic gene phylogenies were coherent with those of the housekeeping genes in these four genomic groups, indicating that the symbiotic genes were mainly maintained by vertical transfer in the soybean rhizobia. In correspondence analysis, the Bradyrhizobium species were not significantly related to the eco-regions, possibly due to the similar climate and soil conditions in these regions.

Diverse bacteria isolated from root nodules of wild Vicia species grown in temperate region of China

In the present study, a total of 154 bacterial strains isolated from nodules of eighteen Vicia species mainly grown in the temperate Chinese provinces were characterized by ARDRA, ITS PCR–RFLP, BOX-PCR, sequencing of 16S rDNA, nodC, nifH, atpD and glnII, and nodulation tests. The results demonstrated that most of the R. leguminosarum strains were effective microsymbionts of the wild Vicia species, while genomic species related to Rhizobium gallicum, Mesorhizobium huakuii, Ensifer meliloti and Bradyrhizobium spp. were symbiotic bacteria occasionally nodulating with Vicia species. In addition, fourteen strains related to Agrobacterium, Phyllobacterium, Ensifer, Shinella and R. tropici, as well as 22 strains of R. leguminosarum might be nodule endophytes without symbiotic genes. Diverse symbiotic gene lineages were found among the test strains and a strong association was found among the symbiotic gene types and genomic species, indicating the absence of lateral gene transfer. These results greatly enlarged the rhizobial spectrum of Vicia species.

Rhizobial Resource Associated with Epidemic Legumes in Tibet

A total of 128 bacterial test strains originated from Astragalus, Caragana, Gueldenstaedtia, Medicago, Melilotus, Oxytropis, Trifolium, and Vicia grown in Tibet were characterized phenotypically and genomically. Based upon the consensus of grouping results, they were identified as 16 putative species. Twenty-five test strains belonging to seven putative species of Agrobacterium, Bradyrhizobium, and Rhizobium might be nonsymbiotic bacteria and the remaining 103 test strains were symbiotic bacteria belonging to Mesorhizobium, Rhizobium, and Sinorhizobium meliloti. Although no novel taxon was detected in the symbiotic bacteria, several characters including the alkaliphilic psychrotolerance revealed that the Tibetan rhizobia could be ecotypes adapted to the local conditions. The results also demonstrated that frequent lateral transfer of symbiotic genes might have happened in the Tibetan rhizobia since nodC genes similar to that of S. meliloti were found in several Rhizobium test strains and all the Mesorhizobium species had very similar nodC genes despite their genomic background. All of these findings demonstrated that the Tibetan rhizobia were an important resource for further studies on rhizobial ecology and application.

Diversity and geographical distribution of rhizobia associated with Lespedeza spp. in temperate and subtropical regions of China

Eighty-eight root-nodule isolates from Lespedeza spp. grown in temperate and subtropical regions of China were characterized by a polyphasic approach. Nine clusters were defined in numerical taxonomy and SDS-PAGE analysis of whole cell proteins. Based upon further characterizations of amplified 16S rDNA restriction analysis (ARDRA), PCR-based restriction fragment length polymorphism of ribosomal IGS, 16S rDNA sequence analysis and DNA-DNA hybridization, these isolates were identified as Bradyrhizobium japonicum, B. elkanii, B. yuanmingense, Mesorhizobium amorphae, M. huakuii, Sinorhizobium meliloti and three genomic species related to B. yuanmingense, Rhizobium gallicum and R. tropici. The Bradyrhizobium species and R. tropici-related rhizobia were mainly isolated from the subtropical region and the species of Mesorhizobium, S. meliloti and R. gallicum-related species were all isolated from the temperate region. Phylogenetic analyses of nifH and nodC indicated that the symbiotic genes of distinct rhizobial species associated with Lespedeza spp. might have different origins and there was no evidence for lateral gene transfer of symbiotic genes. The results obtained in the present study and in a previous report demonstrated that Lespedeza spp. are nodulated by rhizobia with diverse genomic backgrounds and these Lespedeza-nodulating rhizobia were not specific to the host species, but specific to their geographic origins.

Effects of rhizobial inoculation, cropping systems and growth stages on endophytic bacterial community of soybean roots

The effects of rhizobial inoculation, soybean-maize intercropping and plant growth stages on the diversity of soybean root endophytic bacteria were evaluated. PCR-based terminal restriction fragment length polymorphism (T-RFLP) of 16S rRNA genes and analyses of cloned partial 16S rRNA gene libraries were used to study the endophytic bacterial communities. The root endophytic bacterial populations during the whole growing season was significantly affected by the three factors analyzed, with plant growth and rhizobial inoculation showing the highest and lowest impact. The alphaproteobacteria and betaproteobacteria were the most predominant endophytes of soybean roots. Some undefined bacterial lineages were also found, indicating that the root endophytic populations might constitute an important source for the isolation of novel bacteria. Bradyrhizobium liaoningense (used as inoculant) occupied most of the soybean nodules formed and Sinorhizobium americanum-related bacteria were detected as the main root endophytes. Results also suggest that bacteria might be transferred from nodules to roots (as endophytic bacteria) when the seeds become mature, which might be a possible process in the life cycle of rhizobia. Rhizobial inoculation and intercropping practices could increase the diversity of the soybean root endophytes, which might have a positive influence on the growth of soybean plants.

Bradyrhizobium spp. and Sinorhizobium fredii are predominant in root nodules of Vigna angularis, a native legume crop in the subtropical region of China.

Adzuki bean (Vigna angularis) is an important legume crop native to China, but its rhizobia have not been well characterized. In the present study, a total of 60 rhizobial strains isolated from eight provinces of China were analyzed with amplified 16S rRNA gene RFLP, IGS-RFLP, and sequencing analyses of 16S rRNA, atpD, recA, and nodC genes. These strains were identified as genomic species within Rhizobium, Sinorhizobium, Mesorhizobium, Bradyrhizobium, and Ochrobactrum. The most abundant groups were Bradyrhizobium species and Sinorhizobium fredii. Diverse nodC genes were found in these strains, which were mainly co-evolved with the housekeeping genes, but a possible lateral transfer of nodC from Sinorhizobium to Rhizobium was found. Analyses of the genomic and symbiotic gene backgrounds showed that adzuki bean shared the same rhizobial gene pool with soybean (legume native to China) and the exotic Vigna species. All of these data demonstrated that nodule formation is the interaction of rhizobia, host plants, and environment characters.