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

Two strains (TRM95111 and TRM95001) 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 TRM95111 and strain TRM95001 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 JC85 and R. halotolerans AB21 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 TRM95111 and the closest strain R. subbaraonis JC85 was 21.8%, respectively. The dDDH value between strain TRM95001 and the closest strains R. tarimense PL-41 was 27.1%. Whole-genome average nucleotide identity (ANI) values of the strain TRM95111 were 75.6-79.3% and strain TRM95001 were 79.2-83.8%, compared to their closely related strains. The G + C content values of strain TRM95111 and TRM95001 were 65.1 and 60.7 mol%, respectively. Two isolates contained predominant quinone of Q-10 and the major fatty acids was Cω7c and they were sensitive to 1 μg of amikacin and kanamycin. The polar lipids of strain TRM95111 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 TRM95001 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 TRM95111 and strain TRM95001 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 TRM95111 (=CCTCC AB 2021116 =JCM34826) and TRM950011 (=CCTCC AB 2021095 =JCM34967), respectively.