ABSTRACT
Members of the Leguminosae form the largest plant family on Earth, with around 18,000 species. The success of legumes can largely be attributed to their ability to form a nitrogen-fixing symbiosis with specific bacteria known as rhizobia, manifested by the development of nodules on the plant roots in which the bacteria fix atmospheric nitrogen, a major contributor to the global nitrogen cycle. Rhizobia described so far belong exclusively to the alpha-subclass of Proteobacteria, where they are distributed in four distinct phylogenetic branches. Although nitrogen-fixing bacteria exist in other proteobacterial subclasses, for example Herbaspirillum and Azoarcus from the phylogenetically distant beta-subclass, none has been found to harbour the nod genes essential for establishing rhizobial symbiosis. Here we report the identification of proteobacteria from the beta-subclass that nodulate legumes. This finding shows that the ability to establish a symbiosis with legumes is more widespread in bacteria than anticipated to date.
Subject(s)
Betaproteobacteria/physiology , Fabaceae/microbiology , Nitrogen Fixation , Plants, Medicinal , Betaproteobacteria/classification , Betaproteobacteria/isolation & purification , Burkholderia/classification , Burkholderia/genetics , DNA, Bacterial , DNA, Ribosomal , Genes, Bacterial , Molecular Sequence Data , Mutagenesis , Phylogeny , Plant Roots/microbiology , RNA, Bacterial/genetics , RNA, Ribosomal, 16S/genetics , SymbiosisABSTRACT
Rhizobia described so far belong to three distinct phylogenetic branches within the alpha-2 subclass of Proteobacteria. Here we report the discovery of a fourth rhizobial branch involving bacteria of the Methylobacterium genus. Rhizobia isolated from Crotalaria legumes were assigned to a new species, "Methylobacterium nodulans," within the Methylobacterium genus on the basis of 16S ribosomal DNA analyses. We demonstrated that these rhizobia facultatively grow on methanol, which is a characteristic of Methylobacterium spp. but a unique feature among rhizobia. Genes encoding two key enzymes of methylotrophy and nodulation, the mxaF gene, encoding the alpha subunit of the methanol dehydrogenase, and the nodA gene, encoding an acyltransferase involved in Nod factor biosynthesis, were sequenced for the type strain, ORS2060. Plant tests and nodA amplification assays showed that "M. nodulans" is the only nodulating Methylobacterium sp. identified so far. Phylogenetic sequence analysis showed that "M. nodulans" NodA is closely related to Bradyrhizobium NodA, suggesting that this gene was acquired by horizontal gene transfer.
