Bradyrhizobium tunisiense sp. nov., a novel rhizobial species isolated from Acacia saligna nodules.

Three slow-growing rhizobial strains, designated as 1AS2L^T, 1AS20L and 1AS5L, were isolated from nodules of Acacia saligna in Borj Cedria, northern Tunisia. These strains, which belong to the symbiovar cyanophyllae, were characterized using a polyphasic approach. Phylogenetic analysis of the 16S rRNA (rrs) gene placed these strains within the genus Bradyrhizobium, specifically in the superclade associated with Bradyrhizobium japonicum. Further phylogenetic analysis using concatenated sequences of the recA, atpD, glnII and gyrB genes (totalling 1,734 bp) positioned the strains in a distinct lineage, with Bradyrhizobium shewense identified as their closest related species, sharing a sequence identity of 95.2%. The type strain, 1AS2L^T, exhibited average nucleotide identity values of 89.10%, 89.08% and 89.00% with the type strains of the closest valid species: Bradyrhizobium frederickii, B. shewense and Bradyrhizobium ottawaense, respectively. Additionally, digital DNA-DNA hybridization values confirmed the novelty of strain 1AS2L^T, showing low similarity (38.0%-38.3%) with the type strains of the closest known species. Phylogenomic analyses based on up-to-date bacterial core genes, Type (Strain) Genome Server and the Genome Taxonomy Database (GTDB) pipelines further supported the uniqueness of the 1AS2L^T, 1AS20L and 1AS5L strains. The GTDB analysis also robustly clustered two strains (SZCCT0449 and NSD-1) with our strains, suggesting putative members of the proposed novel species. The differentiation of these novel strains from their closest phylogenetic neighbours was also corroborated by phenotypic, physiological and fatty acid content analyses. Based on genomic, phenotypic and biochemical data, we propose the establishment of a novel species, Bradyrhizobium tunisiense sp. nov., with strain 1AS2L^T (=LMG 33170^T=DSM 114401^T) as the type strain.