Legumes form a large group of plants that constitute the third largest family of angiosperms, including near 20,000 species and 750 genera. Most of them have the ability to establish symbioses with diazotrophic bacteria, collectively known as rhizobia, which induce root nodules where biological nitrogen fixation takes place, conferring legumes a relevant ecological advantage. This group of bacteria that for many years was thought to be formed by a scarce number of genera and species within alpha proteobacteria, shows nowadays an important genetic diversity including species phylogenetically divergent both in core and symbiotic genes sequences. Together with rhizobia, other endophytic bacteria are present in legume nodules coexisting with rhizobial strains and their ecological role remains unknown in most cases, but they likely have an effect in plant health, plant growth or even in the rhizobia-legume symbiosis. In this review we present an overview of the associations of bacteria with legumes, the current available knowledge on the phylogenetic diversity of both rhizobia and endophytic bacteria inhabiting root nodules, and the symbiotic features used to define symbiovars in rhizobia.

Rhizobia are the common bacterial symbionts that form nitrogen-fixing root nodules in legumes. However, recently other bacteria have been shown to nodulate and fix nitrogen symbiotically with these plants. Neptunia natans is an aquatic legume indigenous to tropical and subtropical regions and in African soils is nodulated by Allorhizobium undicola. This legume develops an unusual root-nodule symbiosis on floating stems in aquatic environments through a unique infection process. Here, we analyzed the low-molecular-weight RNA and 16S ribosomal DNA (rDNA) sequence of the same fast-growing isolates from India that were previously used to define the developmental morphology of the unique infection process in this symbiosis with N. natans and found that they are phylogenetically located in the genus Devosia , not Allorhizobium or Rhizobium . The 16S rDNA sequences of these two Neptunia -nodulating Devosia strains differ from the only species currently described in that genus, Devosia riboflavina. From the same isolated colonies, we also located their nodD and nifH genes involved in nodulation and nitrogen fixation on a plasmid of approximately 170 kb. Sequence analysis showed that their nodD and nifH genes are most closely related to nodD and nifH of Rhizobium tropici , suggesting that this newly described Neptunia -nodulating Devosia species may have acquired these symbiotic genes by horizontal transfer.

The threatened caesalpinioid legume Dimorphandra wilsonii, which is native to the Cerrado biome in Brazil, was examined for its nodulation and N₂-fixing ability, and was compared with another, less-threatened species, D. jorgei. Nodulation and potential N₂ fixation was shown on seedlings that had been inoculated singly with five bradyrhizobial isolates from mature D. wilsonii nodules. The infection of D. wilsonii by two of these strains (Dw10.1, Dw12.5) was followed in detail using light and transmission electron microscopy, and was compared with that of D. jorgei by Bradyrhizobium strain SEMIA6099. The roots of D. wilsonii were infected via small transient root hairs at 42 d after inoculation (dai), and nodules were sufficiently mature at 63 dai to express nitrogenase protein. Similar infection and nodule developmental processes were observed in D. jorgei. The bacteroids in mature Dimorphandra nodules were enclosed in plant cell wall material containing a homogalacturonan (pectic) epitope that was recognized by the monoclonal antibody JIM5. Analysis of sequences of their rrs (16S rRNA) genes and their ITS regions showed that the five D. wilsonii strains, although related to SEMIA6099, may constitute five undescribed species of genus Bradyrhizobium, whilst their nodD and nifH gene sequences showed that they formed clearly separated branches from other rhizobial strains. This is the first study to describe in full the N₂-fixing symbiotic interaction between defined rhizobial strains and legumes in the sub-family Caesalpinioideae. This information will hopefully assist in the conservation of the threatened species D. wilsonii.