title: Bradyrhizobium stylosanthis sp. nov., comprising nitrogen-fixing symbionts isolated from nodules of the tropical forage legume Stylosanthes spp.
abstract: The introduction of legumes and nitrogen-fixing bacteria in tropical areas under pasture is a key factor for improvement of soil fertility. However, there are still very few studies concerning the symbionts of tropical forage legumes. We performed a polyphasic study with three strains representing the genus Bradyrhizobium (BR 446T, BR 510 and BR 511) isolated from the tropical perennial forage legume of the genus Stylosanthes. On the basis of 16S rRNA gene sequences, the three strains showed highest similarity with B. huanghuaihaiense, and in the analysis of the intergenic transcribed spacer (ITS) they showed less than 93.4 % similarity to all described species of the genus Bradyrhizobium. Multilocus sequence analysis (MLSA) with three, four or five (dnaK, glnII, gyrB, recA and rpoB) housekeeping genes confirmed that the BR strains belong to a distinct clade, with <96.5 % nucleotide identity with other members of the genus Bradyrhizobium. Average nucleotide identity (ANI) of genome sequences between strain BR 446T and B.huanghuaihaiense was below the threshold for species circumscription (90.7 %). DNA-DNA hybridization resulted in ΔTm values over 6.7 °C with the most closely related species. Similarities among the BR strains and differences from other species were confirmed by rep-PCR analysis. Interestingly, the BR strains were grouped in the analysis of nifH and nodC genes, but showed higher similarity with B. iriomotense and B. manausense than with B.huanghuaihaiense, indicating a different evolutionary history for nitrogen-fixation genes. Morpho-physiological, genotypic and genomic data supported that these BR strains represent a novel species for which the name Bradyrhizobium stylosanthis sp. nov. is suggested. The type strain is BR 446T (=CNPSo 2823T=HAMBI 3668T=H-8T), isolated from Stylosanthes guianensis.
authors.affiliation: Embrapa Soja, C.P. 231, 86001-970 Londrina, Paraná, Brazil.|Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, SBN, Quadra 2, BlocoL, Lote 06, Edifício Capes, 70.040-020 Brasília, Distrito Federal, 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: Embrapa Agrobiologia, C.P. 74.505, 23891-000 Seropédica, Rio de Janeiro, Brazil.
authors.forename: Jean Luiz Simões
authors.initials: JLS
authors.lastname: Araújo
authors.affiliation: Embrapa Agrobiologia, C.P. 74.505, 23891-000 Seropédica, Rio de Janeiro, Brazil.
authors.forename: Luc Felicianus Marie
authors.initials: LFM
authors.lastname: Rouws
authors.affiliation: Embrapa Agrobiologia, C.P. 74.505, 23891-000 Seropédica, Rio de Janeiro, Brazil.
authors.forename: Jerri Édson
authors.initials: JÉ
authors.lastname: Zilli
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: 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; D004269:DNA, Bacterial / Q000235:genetics; D021903:DNA, Ribosomal Spacer / Q000235:genetics; D007887:Fabaceae / Q000382:microbiology*; D005227:Fatty Acids / Q000737:chemistry; D005798:Genes, Bacterial; D058885:Multilocus Sequence Typing; D009586:Nitrogen Fixation; D009693:Nucleic Acid Hybridization; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D053204:Root Nodules, Plant / Q000382:microbiology*; D017422:Sequence Analysis, DNA
chemical_list: D004269:DNA, Bacterial; D021903:DNA, Ribosomal Spacer; D005227:Fatty Acids; D012336:RNA, Ribosomal, 16S