Root nodule symbiosis in Lotus japonicus drives the establishment of distinctive rhizosphere, root, and nodule bacterial communities

Significance

Legumes are known as pioneer plants colonizing marginal soils, and as enhancers of the nutritional status in cultivated soils. This beneficial activity has been explained by their capacity to engage in symbiotic relationship with nitrogen-fixing rhizobia. We performed a community profiling analysis of Lotus japonicus wild type and mutants to investigate the role of the nodulation pathway on the structure of the root-associated bacterial microbiota. We found that several bacterial orders were almost entirely depleted from the mutant roots, and that an intact symbiosis is needed for the establishment of taxonomically diverse and distinctive bacterial communities in the root and rhizosphere. Our findings imply that a symbiosis-linked bacterial community, rather than dinitrogen-fixing rhizobia alone, contributes to legume growth and ecological performance.

Abstract

Lotus japonicus has been used for decades as a model legume to study the establishment of binary symbiotic relationships with nitrogen-fixing rhizobia that trigger root nodule organogenesis for bacterial accommodation. Using community profiling of 16S rRNA gene amplicons, we reveal that in Lotus, distinctive nodule- and root-inhabiting communities are established by parallel, rather than consecutive, selection of bacteria from the rhizosphere and root compartments. Comparative analyses of wild-type (WT) and symbiotic mutants in Nod factor receptor5 ( nfr5), Nodule inception ( nin) and Lotus histidine kinase1 ( lhk1) genes identified a previously unsuspected role of the nodulation pathway in the establishment of different bacterial assemblages in the root and rhizosphere. We found that the loss of nitrogen-fixing symbiosis dramatically alters community structure in the latter two compartments, affecting at least 14 bacterial orders. The differential plant growth phenotypes seen between WT and the symbiotic mutants in nonsupplemented soil were retained under nitrogen-supplemented conditions that blocked the formation of functional nodules in WT, whereas the symbiosis-impaired mutants maintain an altered community structure in the nitrogen-supplemented soil. This finding provides strong evidence that the root-associated community shift in the symbiotic mutants is a direct consequence of the disabled symbiosis pathway rather than an indirect effect resulting from abolished symbiotic nitrogen fixation. Our findings imply a role of the legume host in selecting a broad taxonomic range of root-associated bacteria that, in addition to rhizobia, likely contribute to plant growth and ecological performance.