title: Bradyrhizobium tropiciagri sp. nov. and Bradyrhizobium embrapense sp. nov., nitrogen-fixing symbionts of tropical forage legumes.
abstract: Biological nitrogen fixation is a key process for agricultural production and environmental sustainability, but there are comparatively few studies of symbionts of tropical pasture legumes, as well as few described species of the genus Bradyrhizobium, although it is the predominant rhizobial genus in the tropics. A detailed polyphasic study was conducted with two strains of the genus Bradyrhizobium used in commercial inoculants for tropical pastures in Brazil, CNPSo 1112T, isolated from perennial soybean (Neonotonia wightii), and CNPSo 2833T, from desmodium (Desmodium heterocarpon). Based on 16S-rRNA gene phylogeny, both strains were grouped in the Bradyrhizobium elkanii superclade, but were not clearly clustered with any known species. Multilocus sequence analysis of three (glnII, gyrB and recA) and five (plus atpD and dnaK) housekeeping genes confirmed that the strains are positioned in two distinct clades. Comparison with intergenic transcribed spacer sequences of type strains of described species of the genus Bradyrhizobium showed similarity lower than 93.1 %, and differences were confirmed by BOX-PCR analysis. Nucleotide identity of three housekeeping genes with type strains of described species ranged from 88.1 to 96.2 %. Average nucleotide identity of genome sequences showed values below the threshold for distinct species of the genus Bradyrhizobium ( < 90.6 %), and the value between the two strains was also below this threshold (91.2 %). Analysis of nifH and nodC gene sequences positioned the two strains in a clade distinct from other species of the genus Bradyrhizobium. Morphophysiological, genotypic and genomic data supported the description of two novel species in the genus Bradyrhizobium, Bradyrhizobium tropiciagri sp. nov. (type strain CNPSo 1112T = SMS 303T = BR 1009T = SEMIA 6148T = LMG 28867T) and Bradyrhizobium embrapense sp. nov. (type strain CNPSo 2833T = CIAT 2372T = BR 2212T = SEMIA 6208T = U674T = LMG 2987).
authors.affiliation: Embrapa Soja, C.P. 231, 86001-970 Londrina, Paraná, Brazil.|Universidade Estadual de Londrina, Department of Microbiology, C.P. 10.011, 86057-970 Londrina, Paraná, Brazil.
authors.forename: Jakeline Renata Marçon
authors.initials: JRM
authors.lastname: Delamuta
authors.affiliation: Conselho Nacional de Desenvolvimento Científico e Tecnológico, SHIS QI 1 Conjunto B, Blocos A, B, C e D, Lago Sul, 71605-001 Brasília, Distrito Federal, Brazil.
authors.forename: Renan Augusto
authors.initials: RA
authors.lastname: Ribeiro
authors.affiliation: Universidad Nacional Agraria La Molina, Av. La Molina s/n La Molina, Lima, Peru.
authors.forename: Ernesto
authors.initials: E
authors.lastname: Ormeño-Orrillo
authors.affiliation: Embrapa Meio Ambiente, C.P. 69, 13820-000 Jaguariúna, São Paulo, Brazil.
authors.forename: Marcia Maria
authors.initials: MM
authors.lastname: Parma
authors.affiliation: Embrapa Meio Ambiente, C.P. 69, 13820-000 Jaguariúna, São Paulo, Brazil.
authors.forename: Itamar Soares
authors.initials: IS
authors.lastname: Melo
authors.affiliation: Centro de Ciencias Genómicas, Universidad Nacional Autónoma de México, Cuernavaca, Morelos, Mexico.
authors.forename: Esperanza
authors.initials: E
authors.lastname: Martínez-Romero
authors.affiliation: Universidade Estadual de Londrina, Department of Microbiology, C.P. 10.011, 86057-970 Londrina, Paraná, Brazil.|Embrapa Soja, C.P. 231, 86001-970 Londrina, Paraná, Brazil.|Conselho Nacional de Desenvolvimento Científico e Tecnológico, SHIS QI 1 Conjunto B, Blocos A, B, C e D, Lago Sul, 71605-001 Brasília, Distrito Federal, Brazil.
authors.forename: Mariangela
authors.initials: M
authors.lastname: Hungria
mesh_terms: D015373:Bacterial Typing Techniques; D001482:Base Composition; D020369:Bradyrhizobium / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D001938:Brazil; D004269:DNA, Bacterial / Q000235:genetics; D007887:Fabaceae / Q000382:microbiology*; D005227:Fatty Acids / Q000737:chemistry; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D058885:Multilocus Sequence Typing; D009586:Nitrogen Fixation; D009693:Nucleic Acid Hybridization; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D017422:Sequence Analysis, DNA; D013025:Glycine max / Q000382:microbiology; D014329:Tropical Climate
chemical_list: D004269:DNA, Bacterial; D005227:Fatty Acids; D012336:RNA, Ribosomal, 16S