Diversity And Biogeography Of Endophytic Bacteria Isolated From Root Nodules Of Ammopiptanthus Grown In Northwest China

The northwestern regions of China are the unique areas for the grown of Ammopiptanthus, of which the vegetation are discretely and dominantly distributed. Both considering the distribution characters of this plant and the coverage of sampling locations, nines sampling sites, with different ecological environments, where are the areas of Ammopiptanthus being dominantly distributed, were covered by our study and the endophytic bacteria isolated from the sampled nodules were further explored to their diversity and biogeography. Finally, a total of 372 isolates were divided into 33 16 S r RNA species based upon the 16 S r RNA RFLP and 16 S r RNA Bayesian phylogenetic analysis. Of those isolates, the bacteria defined under the genera Rhizobium and Sinorhizobium, including Rhizobium galegae(82 isolates), Sinorhizobium meliloti(51), Rhizobium radiobacter(28), Rhizobium gallicum(21), Rhizobium sp.(15), Sinorhizobium fredii(13), Sinorhizobium arboris(2), Rhizobium herbae(3) and Rhizobium leguminosarum(4), are N2-fixing microsymbionts harbored by Ammopiptanthus, with R. galegae and S. meliloti as the dominant groups, implying the microsymbionts preferences of this plant, and S. arboris, R. herbae and R. leguminosarum as the minor groups, indicating their weakness in competing to nodulate with Ammopiptanthus. In contrast, the remaining 151 isolates were the other endophytic bacteria, which were classified into twenty-five species included in three phyla, with the phylum Proteobacteria as the predominant endophytes(147/151) associated with this plant. There were four genera including eighteen species found in Proteobacteria where Phyllobacterium brassicacearum, and Xanthomonas translucens(18) were in dominance. The other two phyla, corresponding to Firmicutes and Actinobacteria, which included seven species, were only defined among thirty-four isolates, with singletons in most of the species.Diversity and biogeography of Ammopiptanthus nodule endophytes were further explored based on the defined 34 species as the operational taxonomy units(OTUs). As a results of this, we found that alpha diversity were varied among sampling sites, with the influences of different edaphic and climatic factors. Moreover, GLM and GLDM models described that how the variation in alpha diversities among sites was affected by distinct edaphic or climatic factor. On the other side, the beta diversity analysis resulted in the correlations between the species composition or phylogenetic distance among sites and geographic distance or environmental heterogeneities, which suggested that both distance-decay effect and environmental filtering determined the compositional and phylogenetic diversity of the endophytic community composition, implying that the genes flow among endophytic communities were limited both at population and community level. As the results, the biogeographic distributions of endophytes were found in the three ecoregions where the edaphic, climatic factors and geographic distance had a significant difference.Furthermore, the definition of OTUs was also based on the individual level. All of the 372 isolates were genotyped by RAPD fingerprint analyses, which produced a unique genotype for each single isolate. The genotypic data was used to perform multivariate analysis and the results demonstrate the existence of the three-ecoregion biogeographic pattern in the endophytes at strain level, indicating that the evolutionary forces drove the biogeography of the endophytes, as revealed by the divergence and purify selection of some genomic fragment resulted from the influence of environmental variables.To investigate the relationship between environmental variables and the evolutionary distribution, 155 isolates belonging to thirteen dominant species were used to sequencing their rec A and atp D genes, and the evolutionary distance calculated from the Bayesian tree of the concatenation of this two genes was ordinated by a CCA model, with different environmental variables constrained in. The resulting biplot of CCA revealed that(i) those of the 155 isolates were evolutionary assembled in the three ecoregions, which was in agreement with the results of RAPD and taxon based analysis;(ii) the soil components of p H and available P as the dominant factors to determine the endophytes evolutionary assembled in the three ecoregions;(iii) according to the correlation between the certain environmental variables and endophytic species, the endophytes mainly found in ecoregion I were affected by natural environmental selection, while in contrast to this the endophytes tended to be distributed in ecoregion II were due to the anthropogenetic impacts, and ecoregion III possessed the endophytes having tightly biotic interaction with the host plant.