Two bacterial strains isolated from root nodules of soybean were characterized phylogenetically as members of a distinct group in the genus Ensifer based on 16S rRNA gene comparisons. They were also verified as a separated group by the concatenated sequence analyses of recA, atpD and glnII (with similarities <= 93.9% to the type strains for defined species), and by the average nucleotide identities (ANI) between the whole genome sequence of the representative strain CCBAU 251167(T) and those of the closely related strains in Ensifer glycinis and Ensifer fredii (90.5% and 90.3%, respectively). Phylogeny of symbiotic genes (nodC and nifH) grouped these two strains together with some soybean-nodulating strains of E. fredii, E. glycinis and Ensifer sojae. Nodulation tests indicated that the representative strain CCBAU 251167(T) could form root nodules with capability of nitrogen fixing on its host plant and Glycine soja, Cajanus cajan, Vigna unguiculata, Phaseolus vulgaris and Astragalus membranaceus, and it formed ineffective nodules on Leucaena leucocephala. Strain CCBAU 251167(T) contained fatty acids 18:1 omega 9c, 18:0 iso and 20:0, differing from other related strains. Utilization of L-threonine and D-serine as carbon source, growth at pH 6.0 and intolerance of 1% (w/v) NaCI distinguished strain CCBAU 251167(T) from other type strains of the related species. The genome size of CCBAU 251167(T) was 6.2 Mbp, comprising 7,581 predicted genes with DNA G+C content of 59.9 mol% and 970 unique genes. Therefore, a novel species, Ensifer shofinae sp. nov., is proposed, with CCBAU 251167(T) (=ACCC 19939(T) = LMG 29645(T)) as type strain. (C) 2017 Elsevier GmbH. All rights reserved.