Rhizobium alarense sp. nov. and Rhizobium halophilum sp. nov. isolated from the nodule and rhizosphere of Lotus japonicus

Two strains (TRM95111T and TRM95001T) of Gram-stain-negative, aerobic, rod-shaped microbes were isolated from the nodule and rhizosphere of Lotus japonicus grown in the campus of Tarim University in Alar, Xinjiang, China. Strain TRM95111T and strain TRM95001T shared 93.1% 16S rRNA gene sequences similarity with each other and had 98.2 and 97.9% 16S rRNA gene sequence similarity to the closest species Rhizobium subbaraois JC85T and R. halotolerans AB21T by EzBioCloud blast, respectively. Phylogenetic analyses based on 16S rRNA gene sequences, housekeeping gene sequences and core-proteome average amino acid identity (cpAAI) showed that two strains belonged to the genus Rhizobium. The value of digital DNA–DNA hybridization (dDDH) between strain TRM95111T and the closest strain R. subbaraonis JC85T was 21.8%, respectively. The dDDH value between strain TRM95001T and the closest strains R. tarimense PL-41T was 27.1%. Whole-genome average nucleotide identity (ANI) values of the strain TRM95111T were 75.6–79.3% and strain TRM95001T were 79.2–83.8%, compared to their closely related strains. The G + C content values of strain TRM95111T and TRM95001T were 65.1 and 60.7 mol%, respectively. Two isolates contained predominant quinone of Q-10 and the major fatty acids was C18:1ω7c and they were sensitive to 1 μg of amikacin and kanamycin. The polar lipids of strain TRM95111T included unidentified aminophospho lipids (APL1-3), unidentified phospholipids (PL1-2), phosphatidylcholine (PC), unidentified lipids (L1-5) and phospholipids of unknown structure containing glucosamine (NPG), compared to the polar lipids of strain TRM95001T including unidentified aminophospho lipids (APL1-3), unidentified phospholipids (PL1-2), phosphatidylcholine (PC), unidentified lipids (L2-5), hydroxy phosphatidyl ethanolamine (OH-PE) and phospholipids of unknown structure containing glucosamine (NPG). Nodulation tests showed that two strains could induce nodules formation in L. japonicus. Based on the genomic, phenotypic and phylogenetic analyses, strain TRM95111T and strain TRM95001T are suggested to represent two new species of the genus Rhizobium, whose names are proposed as Rhizobium alarense sp. nov. and Rhizobium halophilum sp. nov. The type strains are TRM95111T (=CCTCC AB 2021116T =JCM34826T) and TRM950011T (=CCTCC AB 2021095T =JCM34967T), respectively.