Nitrogen-Fixing Rhizobial Strains Isolated from Common Bean Seeds: Phylogeny, Physiology, and Genome Analysis

Abstract

ABSTRACTRhizobial bacteria are commonly found in soil but also establish symbiotic relationships with legumes, inhabiting the root nodules, where they fix nitrogen. Endophytic rhizobia have also been reported in the roots and stems of legumes and other plants. We isolated several rhizobial strains from the nodules of noninoculated bean plants and looked for their provenance in the interiors of the seeds. Nine isolates were obtained, covering most known bean symbiont species, which belong to theRhizobiumandSinorhizobiumgroups. The strains showed several large plasmids, except for aSinorhizobiumamericanumisolate. Two strains, oneRhizobium phaseoliand oneS. americanumstrain, were thoroughly characterized. Optimal symbiotic performance was observed for both of these strains. TheS. americanumstrain showed biotin prototrophy when subcultured, as well as high pyruvate dehydrogenase (PDH) activity, both of which are key factors in maintaining optimal growth. TheR. phaseolistrain was a biotin auxotroph, did not grow when subcultured, accumulated a large amount of poly-β-hydroxybutyrate, and exhibited low PDH activity. The physiology and genomes of these strains showed features that may have resulted from their lifestyle inside the seeds: stress sensitivity, a ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO) complex, a homocitrate synthase (usually present only in free-living diazotrophs), a hydrogenase uptake cluster, and the presence of prophages. We propose that colonization by rhizobia and their presence inPhaseolusseeds may be part of a persistence mechanism that helps to retain and disperse rhizobial strains.