The genus Bradyrhizobium is considered as the probable ancestor lineage of all rhizobia, broadly spread in a variety of ecosystems and with remarkable diversity. A polyphasic study was performed to characterize and clarify the taxonomic position of eight bradyrhizobial strains isolated from indigenous legumes to Western Australia. As expected for the genus, the 16S rRNA gene sequences were highly conserved, but the results of multilocus sequence analysis with four housekeeping genes (dnaK, glnII, gyrB and recA) confirmed three new distinct clades including the following strains: (1) WSM 1744T, WSM 1736 and WSM 1737; (2) WSM 1791T and WSM 1742; and (3) WSM 1741T, WSM 1735 and WSM 1790. The highest ANI values of the three groups in relation to the closest type strains were 92.4, 92.3 and 93.3 %, respectively, below the threshold of species circumscription. The digital DNA–DNA hybridization analysis also confirmed new species descriptions, with less than 52 % relatedness with the closest type strains. The phylogeny of the symbiotic gene nodC clustered the eight strains into the symbiovar retamae, together with seven Bradyrhizobium type strains, sharing from 94.2–98.1 % nucleotide identity (NI), and less than 88.7 % NI with other related strains and symbiovars. Morpho-physiological, phylogenetics, genomic and symbiotic traits were determined for the new groups and our data support the description of three new species, Bradyrhizobium archetypum sp. nov., Bradyrhizobium australiense sp. nov. and Bradyrhizobium murdochi sp. nov., with WSM 1744T (=CNPSo 4013T=LMG 31646T), WSM 1791T (=CNPSo 4014T=LMG 31647T) and WSM 1741T (=CNPSo 4020T=LMG 31651T) designated as type strains, respectively.

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Strains of the genus Bradyrhizobium associated with agronomically important crops such as soybean (Glycine max) are increasingly studied; however, information about symbionts of wild Glycine species is scarce. Australia is a genetic centre of wild Glycine species and we performed a polyphasic analysis of three Bradyrhizobium strains—CNPSo 4010T, CNPSo 4016T, and CNPSo 4019T—trapped from Western Australian soils with Glycine clandestina, Glycine tabacina and Glycine max, respectively. The phylogenetic tree of the 16S rRNA gene clustered all strains into the Bradyrhizobium japonicum superclade; strains CNPSo 4010T and CNPSo 4016T had Bradyrhizobium yuanmingense CCBAU 10071T as the closest species, whereas strain CNPSo 4019T was closer to Bradyrhizobium liaoningense LMG 18230T. The multilocus sequence analysis (MLSA) with five housekeeping genes—dnaK, glnII, gyrB, recA and rpoB—confirmed the same clusters as the 16S rRNA phylogeny, but indicated low similarity to described species, with nucleotide identities ranging from 93.6 to 97.6% of similarity. Considering the genomes of the three strains, the average nucleotide identity and digital DNA–DNA hybridization values were lower than 94.97 and 59.80 %, respectively, with the closest species. In the nodC phylogeny, strains CNPSo 4010T and CNPSo 4019T grouped with Bradyrhizobium zhanjiangense and Bradyrhizobium ganzhouense , respectively, while strain CNPSo 4016T was positioned separately from the all symbiotic Bradyrhizobium species. Other genomic (BOX-PCR), phenotypic and symbiotic properties were evaluated and corroborated with the description of three new lineages of Bradyrhizobium . We propose the names of Bradyrhizobium agreste sp. nov. for CNPSo 4010T (=WSM 4802T=LMG 31645T) isolated from Glycine clandestina, Bradyrhizobium glycinis sp. nov. for CNPSo 4016T (=WSM 4801T=LMG 31649T) isolated from Glycine tabacina and Bradyrhizobium diversitatis sp. nov. for CNPSo 4019T (=WSM 4799T=LMG 31650T) isolated from G. max.

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Since early studies, the history of prokaryotes taxonomy has dealt with many changes driven by the development of new and more robust technologies. As a result, the number of new taxa descriptions is exponentially increasing, while an increasing number of others has been subject of reclassification, demanding from the taxonomists more effort to maintain an organized hierarchical system. However, expectations are that the taxonomy of prokaryotes will acquire a more stable status with the genomic era. Other analyses may continue to be necessary to determine microbial features, but the use of genomic data might be sufficient to provide reliable taxa delineation, helping taxonomy to reach the goal of correct classification and identification. Here we describe the evolution of prokaryotes’ taxonomy until the genomic era, emphasizing bacteria and taking as an example the history of rhizobia taxonomy. This example was chosen because of the importance of the symbiotic nitrogen fixation of legumes with rhizobia to the nitrogen input to both natural ecosystems and agricultural crops. This case study reports the technological advances and the methodologies used to classify and identify bacterial species and indicates the actual rules required for an accurate description of new taxa.

Bradyrhizobium is a heterogeneous bacterial genus capable of establishing symbiotic associations with a broad range of legume hosts, including species of economic and environmental importance. This study was focused on the taxonomic and symbiovar definition of four strains – CNPSo 4026T, WSM 1704T, WSM 1738T and WSM 4400T – previously isolated from nodules of legumes in Western Australia and South Africa. The 16S rRNA gene phylogenetic tree allocated the strains to the Bradyrhizobium elkanii supergroup. The multilocus sequence analysis (MLSA) with partial sequences of six housekeeping genes – atpD, dnaK, glnII, gyrB, recA and rpoB – did not cluster the strains under study as conspecific to any described Bradyrhizobium species. Average nucleotide identity and digital DNA–DNA hybridization values were calculated for the four strains of this study and the closest species according to the MLSA phylogeny with the highest values being 95.46 and 62.20 %, respectively; therefore, both being lower than the species delineation cut-off values. The nodC and nifH phylogenies included strains WSM 1738T and WSM 4400T in the symbiovars retamae and vignae respectively, and also allowed the definition of three new symbiovars, sv. cenepequi, sv. glycinis, and sv. cajani. Analysis of morphophysiological characterization reinforced the identification of four novel proposed Bradyrhizobium species that are accordingly named as follows: Bradyrhizobium cenepequi sp. nov. (CNPSo 4026T=WSM 4798T=LMG 31653T), isolated from Vigna unguiculata; Bradyrhizobium semiaridum sp. nov. (WSM 1704T=CNPSo 4028T=LMG 31654T), isolated from Tephrosia gardneri; Bradyrhizobium hereditatis sp. nov. (WSM 1738T=CNPSo 4025T=LMG 31652T), isolated from Indigofera sp.; and Bradyrhizobium australafricanum sp. nov. (WSM 4400T=CNPSo 4015T=LMG 31648T) isolated from Glycine sp.

The genomes of five elite strains identified as growth promoters of lowland rice ( Oryza sativa L.) in Brazil were sequenced. They ranged in size from 3,695,387 bp to 5,682,101 bp, encompassing genes of saprophytic ability and stress tolerance. Genome taxonomy enabled their classification as Priestia megaterium , Bacillus altitudinis , and three putative new species of Pseudomonas , Lysinibacillus , and Agrobacterium .