Steroidobacter gossypii sp. nov., isolated from cotton field soil.

A Gram-negative bacterium, designated S1-65T, was isolated from soil samples collected from a cotton field located in the Xinjiang region of PR China. Results of 16S rRNA gene sequence analysis revealed that strain S1-65T was affiliated to the genus Steroidobacter with its closest phylogenetic relatives being 'Steroidobacter cummioxidans' 35Y (98.4 %), 'Steroidobacter agaridevorans' SA29-B (98.3 %) and Steroidobacter agariperforans KA5-BT (98.3 %). 16S rRNA-directed phylogenetic analysis showed that strain S1-65T formed a unique phylogenetic subclade next to 'S. agaridevorans' SA29-B and S. agariperforans KA5-BT, suggesting that strain S1-65T should be identified as a member of the genus Steroidobacter. Further, substantial differences between the genotypic properties of strain S1-65T and the members of the genus Steroidobacter, including average nucleotide identity and digital DNA-DNA hybridization, resolved the taxonomic position of strain S1-65T and suggested its positioning as representing a novel species of the genus Steroidobacter. The DNA G+C content of strain S1-65T was 62.5 mol%, based on its draft genome sequence. The predominant respiratory quinone was ubiquinone-8. The main fatty acids were identified as summed feature 3 (C16:1ω6c/C16:1ω7c), C16 : 0 and iso-C15 : 0. In addition, its polar lipid profile was composed of aminophospholipid, diphosphatidylglycerol, phosphatidylethanolamine and phosphatidylglycerol. Here, we propose a novel species of the genus Steroidobacter: Steroidobacter gossypii sp. nov. with the type strain S1-65T (=JCM 34287T=CGMCC 1.18736T).

Characterization and Distribution of Agar-degrading Steroidobacter agaridevorans sp. nov., Isolated from Rhizosphere Soils.

The environment of plant rhizosphere soil differs from that of non-rhizosphere soil due to the secretion of mucilage polysaccharides from the roots. This environment is regarded as one of the preferential habitats for agar-degrading bacteria. In a previous study, agar-degrading Steroidobacter agariperforans KA5-BT was isolated from agar-enriched agricultural soil using diffusible metabolites from Rhizobiales bacteria. Based on the hypothesis that similar characteristic bacteria still exist in the rhizosphere, isolation was performed using rhizosphere soils. Agar-degrading SA29-BT and YU21-B were isolated from onion and soybean rhizosphere soils. The 16S rRNA genes of these strains showed ≥98.7% identities with the most closely related strain KA5-BT. However, differences were noted in polysaccharide utilization, and average nucleotide identities were <95-96% against strain KA5-BT, indicating that they are different species from S. agariperforans KA5-BT. To investigate the distribution of bacterial sequences affiliated with novel strains, a primer set was designed and a meta-analysis of the 16S rRNA gene was performed. Sequences were widely distributed in rhizospheres throughout Japan, but varied in plant- and region-dependent manners. Regarding phenotypic characterization, distinguishable features were observed in growth temperatures, pH, and dominant fatty acids. SA29-BT and YU21-B grew at 15-40°C and pH 6.0-12 and contained C16:0 as the dominant cell fatty acid, whereas KA5-BT showed no growth at 40°C and pH 12 and contained a moderate amount of C16:0. Based on these characteristics, SA29-BT (JCM 333368T=KCTC 72223T) and YU21-B (JCM 333367=KCTC 72222) represent novel species in the genus Steroidobacter, for which the name Steroidobacter agaridevorans sp. nov. is proposed.

Steroidobacter soli sp. nov., isolated from farmland soil.

A Gram-stain-negative bacterial strain, designated JW-3T, was isolated from a soil sample collected from farmland in Yantai, Shandong Province, PR China. Cells of strain JW-3T are motile rods and strictly aerobic, showing catalase- and oxidase-positive reactions. Strain JW-3T could grow at 16-37 °C (optimum, 30 °C), at pH 6.0-9.0 (pH 7.0) and in the presence of 0-1 % (w/v) NaCl (0.5 %, in Luria-Bertani broth). The major fatty acids were summed feature 3 (C16 : 1 ω7c and/or C16 : 1 ω6c; 35.5 %), iso-C16 : 0 (16.7 %) and C12 : 0 (10.8 %). The major respiratory quinone was ubiquinone-8 (Q8). The polar lipids of strain JW-3T consisted of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, four unidentified phospholipids, two unidentified lipids, two unidentified glycolipids and a partial unidentified aminophospholipid. Strain JW-3T was most closely related to Steroidobacter agariperforans KA5-BT with 97.67 % 16S rRNA gene sequence similarity. Results of phylogenetic analyses, based on 16S rRNA gene sequencing, showed that strain JW-3T forms a distinct phylogenic lineage within the genus Steroidobacter of the family Sinobacteraceae. The DNA G+C content of strain JW-3T was 62.57 mol%, based on its draft genome sequence. Average nucleotide identity values and digital DNA-DNA hybridization values for draft genomes, between strain JW-3T and strain KA5-BT, were 84.54 and 30.80 %, respectively. Based on its phenotypic, chemotaxonomic and molecular features, and DNA-DNA hybridization results, strain JW-3T represents a novel species of the genus Steroidobacter, for which the name Steroidobactersoli sp. nov. is proposed. The type strain is JW-3T (=CCTCC AB 2018184T=KCTC 62820T).

Steroidobacter flavus sp. nov., a microcystin-degrading Gammaproteobacterium isolated from soil.

A microcystin-degrading strain, designated CPCC 100154(T), was isolated from a forest soil sample collected from Hainan Island, South China. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain CPCC 100154(T) is affiliated to the family Sinobacteraceae in Gammaproteobacteria, with high 16S rRNA gene sequence similarities of 98.1, 96.6 and 96.6 % to Steroidobacter agariperforans JCM 18477(T), S. denitrificansis DSM 18526(T), and Povalibacter uvarum JCM 18749(T), respectively. In the phylogenetic tree based on 16S rRNA gene sequences, strain CPCC 100154(T) formed a stable phylogenetic subclade with S. agariperforans JCM 18477(T), which indicated that strain CPCC 100154(T) should be identified as a member of the genus Steroidobacter. The phylogenetic analysis based on partial gyrB gene sequences confirmed the affiliation of strain CPCC 100154(T) to the genus Steroidobacter. While the DNA-DNA hybridization value (47.0 ± 1.7 %) between the new isolate and its near phylogenetic neighbor S. agariperforans JCM 18477(T) was far below 70 %, which demonstrated that strain CPCC 100154(T) represents a different genomic species from S. agariperforans. The strain CPCC 100154(T) was found to be a Gram-stain negative, rod-shaped, motile with a polar flagellum, non-endospore-forming bacterium. Good growth was observed at 28-32 °C and pH 7.0-7.5. Polar lipids were identified to be diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, a unidentified phospholipid, an aminophospholipid and an unidentified aminolipid. The polyamine pattern was determined to contain spermidine as the predominant polyamine, moderate amounts of putrescine as well as traces of sym-homospermidine and spermine. The respiratory quinone was identified as ubiquinone-8. The major cellular fatty acids were found to include summed feature 3 (C16: 1 ω7C/C16: 1 ω6C) (46.1 %) and C16: 0 (29.6 %). The genomic DNA G+C content was determined to be 64.4 mol%. On the basis of the above polyphasic taxonomy evidence, a novel species, Steroidobacter flavus sp. nov. is proposed. The type strain is CPCC 100154(T) (=DSM 23339(T) =CGMCC 1.10759(T)).

The Growth Inhibition Effect on the Causative Bacteria of Bacterial Vaginosis by Bacterial Strains Isolated from the Vagina of a Healthy Woman

Two Gram-positive rod strains isolated from the healthy vagina of a woman were tested for the possibility as probiotics. One strain was identified as Steroidobacter denitrificans (YH1) and the other as Lactobacillus crispatus (YH2) by 16S rRNA partial sequencing. The Casman agar and Man-Rogosa-Sharpe (MRS) agar were mixed in same quantity, supplemented with 5% human rbc lysate (CMB agar). The Wilkins-Chalgren agar and MRS agar were mixed in same quantity (WCM agar). Gardnerella vaginalis was cultured in Casman broth, supplemented with 5% human rbc lysate and 1,000 x-diluted with normal saline. Bacteroides fragilis, Mobiluncus mulieris and Peptostreptococcus asaccharolyticus were cultured in Wilkins-Chalgren anaerobe broth and 2,000x-diluted. S. denitrificans YH1 and L. crispatus YH2 were cultured in MRS broth anaerobically and 100x-diluted. The diluted suspensions of B. fragilis, M. mulieris and P. asaccharolyticus were inoculated on WCM agar and G. vaginalis on CMB agar by cotton swabs. Ten microl aliquots of YH1 and YH2 were inoculated on the center of WCM agar and CMB agar. The growth inhibition zone diameters of B. fragilis, G. vaginalis, M. mulieris and P. asaccharolyticus by YH1 were 35 mm, 35 mm, 25 mm and 60 mm. The inhibition diameters by YH2 were 25 mm, 30 mm, 20 mm and 40 mm, respectively. These results implicate that S. denitrificans YH1 can be the stronger probiotics for the treatment of bacterial vaginosis than L. crispatus, compared inhibition zone diameters by YH1 and YH2.