The Biogeography Of Indigenous Rhizobia Associated With Faba Bean And Its Underlying Formation Mechanism

The nitrogen-fixing rhizobium-legume symbiosis is an important component of sustainable agriculture. The epidemic pattern of these facultative symbionts both within host tissues and in the surrounding environment is helpful for context-dependent application of rhizobial inocula.Rhizosphere soils and nodules of faba bean (Vicia faba L.) were collected from 44 sites in three ecoregions (northwest, east, and southwest) in China. The isolates purified from nodules have been identified as six species: Rhizobium anhuiense, Rhizobium laguerreae, Rhizobium leguminosarum,Rhizobium pisi/Rhizobium fabae, Rhizobium sp.l, and Rhizobium sp.2. Restriction Fragment Length Polymorphism (RFLP) analysis of nodD gene further uncovered five genotypes of nodD among these species. High-throughput sequencing analyses of 16S rRNA, rpoB, and nodD in rhizosphere samples were used to investigate the biogeography of bacterial community, rhizobial species and nodulation gene, respectively. Although total bacterial or rhizobial communities in rhizosphere were not significantly differentiated across ecoregions, biased geographical distribution of facultative microsymbionts of faba bean exists in rhizosphere and nodules. R. laguerreae and R. anhuiense were predominant in different samples with contrasting pH or salt content. The abundance of R. laguerreae correlated positively with pH and salt content in soils, while R. anhuiense abundance correlated negatively with these edaphic factors. Low pH and salt content were more preferred by R. anhuiense.However, R. anhuiense can outcompete R. laguerreae in certain sterilized soils where R. laguerreae originally dominated, and vice versa (soil pH 5.78～8.27; salt content: 0.54～1.47 g·kg-1). Therefore,it is possible that other undetermined abiotic and biotic factors might also affect the relative abundance of two Rhizobium species in soils. Contrasting bacterial taxa associated with either R. laguerreae or R.anhuiense in soils. The number of bacterial taxa positively related to R. anhuiense is more than that of R.laguerreae, and the alpha diversity of samples dominated by R. anhuiense is also significantly higher than that of R. laguerreae. The biogeographical pattern of nodD was clearer than that of rhizobial species in both rhizosphere soils and nodules. Competitive nodulation experiments demonstrated a hierarchical selection on nodD genotypes and their genomic backgrounds by faba bean cultivars. Taken together, abiotic and biotic factors in soils and the selection by legume hosts are either indirectly or directly involved in shaping rhizobial species-level taxonomic biogeography.This study, by using high-throughput sequencing method and high-resolution markers, has revealed the epidemicity of rhizobia in rhizosphere soils of faba bean in terms of both species and nodulation gene across three ecoregions in China. The role of edaphic factors and host plants in shaping the biogeographical patterns of faba bean rhizobia was also investigated. The findings obtained herein can provide guidance in our practice of selecting appropriate inoculant candidates, since rhizobial germplasms present in legume rhizosphere soils can be effectively uncovered using procedures described in this thesis.