Clostridium caldaquaticum sp. nov., a thermophilic bacterium isolated from a hot spring sediment.

A novel anaerobic bacterium, designated SYSU GA19001T, was isolated from a hot spring sediment sample. Phylogenetic analysis indicated that the isolate belongs to the genus Clostridium, and showed the highest sequence similarity to Clostridium swellfunianum CICC 10730T (96.63 %) and Clostridium prolinivorans PYR-10T (96.11 %). Cells of strain SYSU GA19001T were Gram-stain-positive, spore-forming, rod-shaped (0.6-0.8×2.6-4.0 µm) and motile. Growth was observed at pH 5.0-9.0 (optimum, pH 7.0), 37-55 °C (optimum, 45 °C) and in NaCl concentrations of 0-2.0 % (optimum, 0 %). The genomic DNA G+C content was 31.62 %. The major cellular fatty acids of strain SYSU GA19001T were C14 : 0, iso-C15 : 0, C16 : 0 and summed feature 8. The prominent polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol. Meso-diaminopimelic acid was the diamino acid in peptidoglycan. Based on the results of phylogenetic, chemotaxonomic and phenotypic analyses, strain SYSU GA19001T represents a novel species of the genus Clostridium, for which the name Clostridium caldaquaticum sp. nov. is proposed. The type strain of the proposed novel species is SYSU GA19001T (=NBRC 115040T= CGMCC 1.17864T).

Sandaracinobacteroides hominis gen. nov., sp. nov., isolated from human skin.

Strain SZY PN-1 T, representing a novel Gram-negative, aerobic, non-motile, rod-shaped and yellow-pigmented bacterium, was isolated from a skin sample of a healthy Chinese male. Growth occurred at pH 6.0-8.0 (optimum, pH 7.0) and 10-37 â„ƒ (optimum, 30 â„ƒ) with 0-1.0% (w/v) NaCl in R2A agar. Comparative analysis of the 16S rRNA gene sequences revealed that strain SZY PN-1 T shared high similarities with two invalid-published species, "Sandaracinobacter sibiricus" RB16-17 (97.1%) and "Sandaracinobacter neustonicus" JCM 30734 (96.6%), respectively. Phylogenetic analysis of 16S rRNA gene sequences together with protein-concatemer tree showed that SZY PN-1 T formed a separate branch within the family Sphingosinicellaceae. The DNA G + C content of the strain SZY PN-1 T was 65.0% (genome). The polar lipid profile included phosphatidylethanolamine, phosphatidylglycerol, two sphingoglycolipids, diphosphatidylglycerol, five unidentified glycolipids, and seven unidentified lipids. The predominant fatty acids (> 10.0%) were identified as C18:1 ω7c and/or C18:1 ω6c, C17:1 ω6c, C16:1 ω7c and/or C16:1 ω6c. The major respiratory quinone was Q-10. Based on the phenotypic and genotypic features, a novel genus and species, Sandaracinobacteroides hominis gen. nov., sp. nov. is proposed, with type strain SZY PN-1 T (= KCTC 82150 T = NBRC 114675 T).

Thermaurantiacus tibetensis gen. nov., sp. nov., a novel moderately thermophilic bacterium isolated from hot spring microbial mat in Tibet.

Two bacterial strains SYSU G02173T and SYSU G03142 were isolated from hot springs in Tibet, China. Based on the results of nearly full-length 16S rRNA gene sequences and phylogenetic analyses, strains SYSU G02173T and SYSU G03142 were assigned to the family Sphingosinicellaceae, and were closest to Sandaracinobacter sibiricus RB16-17 T (96.04% and 96.12% similarity, respectively). Cells of the both new strains were observed to be motile rod-shape, Gram-staining negative. Growth occurred at pH 6-8 (optimal: pH 7.0) and 37-55 °C (optimal: 45 °C) with 0-1.0% (w/v) NaCl in T4 broth. The cells were found to be positive for oxidase and catalase activities. The major respiratory ubiquinone was Q-8. The major fatty acids were identified as summed feature 8 (C18:1 ω7c and/or C18:1 ω6c), C16:0, C14:0 2-OH. The major polar lipids were found to consist of sphingoglycolipid, phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, one unidentified phospholipid, one unidentified glycolipid, three unidentified aminolipids and two unidentified polar lipids. The DNA G + C contents of strains SYSU G02173T and SYSU G03142 were 71.8%. The average nucleotide identity (ANI) value between strain SYSU G02173T and SYSU G03142 was 99.98%. The amino acid identity (AAI) values between them and their closely related species were below 66.14%. The isolates are characterized by aerobic growth, a yellow endocellular pigment and a higher optimum growth temperature. The results showed that strains SYSU G02173T and SYSU G03142 represent a novel species of a novel genus in the family Sphingomonadaceae, and thus the name Thermaurantiacus tibetensis (type strain SYSU G02173T = KCTC 72052 T = CGMCC 1.16680 T) is proposed.

Diversity and Distribution of Anaerobic Ammonium Oxidation Bacteria in Hot Springs of Conghua, China.

Anaerobic ammonium oxidation (anammox) is an important process of the nitrogen cycle, and the anammox bacteria have been studied in a wide variety of environments. However, the distribution, diversity, and abundance of anammox bacteria in hot springs remain enigmatic. In this study, the anammox process was firstly investigated in hot springs of Conghua, China. Anammox-like bacterial sequences that closely affiliated to "Candidatus Brocadia," "Candidatus Kuenenia," "Candidatus Scalindua," "Candidatus Anammoxoglobus," and "Candidatus Jettenia" were detected. Several operational taxonomic units (OTUs) from this study shared low sequence identities to the 16S rRNA gene of the known anammox bacteria, suggesting that they might be representing putative novel anammox bacteria. A quantitative PCR analysis of anammox-specific 16S rRNA gene confirmed that the abundance of anammox bacteria ranged from 1.60 × 104 to 1.20 × 107 copies L-1. Nitrate was a key environmental factor defining the geographical distribution of the anammox bacterial community in the hot spring ecosystem. Dissolved inorganic carbon had a significant influence on anammox bacterial biodiversity. Our findings for the first time revealed that the diverse anammox bacteria, including putative novel anammox bacterial candidates, were present in Conghua hot spring, which extended the existence of anammox bacteria to the hot springs in China and expands our knowledge of the biogeography of anammox bacteria. This work filled up the research lacuna of anammox bacteria in Chinese hot spring habitat and would guide for enrichment strategies of anammox bacteria of Conghua hot springs.

Seramator thermalis gen. nov., sp. nov., a novel cellulose- and xylan-degrading member of the family Dysgonamonadaceae isolated from a hot spring.

Two anaerobic bacteria, designated strains SYSU GA16112T and SYSU GA16107, were isolated from a hot spring in Tengchong County, Yunnan Province, south-west PR China. Phylogenetic analyses based on 16S rRNA gene sequences showed that strains SYSU GA16112T and SYSU GA16107 belong to the family Dysgonamonadaceae. Cells of strains SYSU GA16112T and SYSU GA16107 were Gram-stain-negative, rod-shaped and non-motile. The major fatty acids (>10 %) of strains SYSU GA16112T and SYSU GA16107 were identified as anteiso-C15 : 0 and anteiso-C17 : 0 3OH. The polar lipid profile of strain SYSU GA16112T was found to consist of phosphatidylethanolamine, two unidentified aminophospholipids, two unidentified phosphoglycolipids, two unidentified aminolipids and one unidentified polar lipid, while that of strain SYSU GA16107 consisted of phosphatidylethanolamine, two unidentified polar lipids, three unidentified aminophospholipids, two unidentified phosphoglycolipids and one unidentified aminolipid. The genomic DNA G+C contents of strains SYSU GA16112T and SYSU GA16107 were determined to be 41.90 and 41.89 %, respectively, and the average nucleotide identity value between them was 99.99 %. Based on their morphological and physiological properties, and results of phylogenetic analyses, strains SYSU GA16112T and SYSU GA16107 are considered to represent a novel species of a novel genus, for which the name Seramator thermalis gen. nov., sp. nov. (type strain SYSU GA16112T=CGMCC 1.5281T=KCTC 15753T) is proposed.

Isolation of Clostridium from Yunnan-Tibet hot springs and description of Clostridium thermarum sp. nov. with lignocellulosic ethanol production.

Lignocellulose is considered a major source of renewable energy that serve as an alternative to the fossil fuels. Members of the genus Clostridium are some of the many microorganisms that have the ability to degrade lignocellulose efficiently to sugar, which can be further converted to biofuel. In this study, we isolated twelve Clostridium strains from hot spring samples of Yunnan and Tibet, of which isolates SYSU GA15002T and SYSU GA17076 showed low 16S rRNA gene sequence identity profiles to any of the validly named Clostridium strains (<94.0%). Studies using a polyphasic taxonomy approach concluded that the two isolates represent one novel species of the genus Clostridium, for which we propose the name Clostridium thermarum sp. nov., with SYSU GA15002T as the type strain of the species. Isolate SYSU GA15002T has an optimum growth temperature at 45°C. Fermentation of the substrates cellobiose, cellulose, xylan and untreated straw powder by this strain results in the production of ethanol, along with acetate and formate. The complete pathways for the conversion of cellulose and xylan to ethanol is also predicted from the genome of isolate SYSU GA15002T, which revealed a single step conversion of lignocellulosic biomass through consolidated bioprocessing. This paper is a comprehensive study encompassing isolation, polyphasic taxonomy, lignocellulose biodegradation and the genomic information of Clostridium in Yunnan-Tibet hot springs.

Network-directed efficient isolation of previously uncultivated Chloroflexi and related bacteria in hot spring microbial mats.

The perplexity of the complex multispecies community interactions is one of the many reasons why majority of the microorganisms are still uncultivated. We analyzed the entire co-occurrence networks between the OTUs of Tibet and Yunnan hot spring samples, and found that less abundant OTUs such as genus Tepidimonas (relative abundant <1%) had high-degree centricity (key nodes), while dominant OTUs particularly genus Chloroflexus (relative abundant, 13.9%) formed the peripheral vertexes. A preliminary growth-promotion assay determined that Tepidimonas sp. strain SYSU G00190W enhanced the growth of Chloroflexus sp. SYSU G00190R. Exploiting this result, an ameliorated isolation medium containing 10% spent-culture supernatant of Tepidimonas sp. strain SYSU G00190W was prepared for targeted isolation of Chloroflexi in the Tibet and Yunnan hot spring samples. 16S rRNA gene fingerprinting characterized majority of the colonies isolated from these media as previously uncultivated Chloroflexi, of which 36 are potential novel species (16S rRNA sequence identity <98.5%). Metabolomes studies indicated that the spent-culture supernatant comprises several low-molecular-weight organic substrates that can be utilized as potential nutrients for the growth of these bacteria. These findings suggested that limited knowledge on the interaction of microbes provide threshold to traditional isolation method.

Francisella salimarina sp. nov., isolated from coastal seawater.

Four strains (SYSU SYW-1T, SYW-2, SYW-3 and XLW-1) were isolated from seawater near the shore in Guangdong Province, China. Cells were Gram-stain-negative, aerobic, non-motile and non-spore-forming. Growth was observed at a temperature range of 16-40 °C (optimum, 32 °C), a pH range of 4-8 (optimum, pH 7) and in the presence of up to 10 % (w/v) NaCl. The major polar lipids were diphosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine and an unidentified phospholipid. The respiratory quinone was ubiquinone 8 (UQ-8), and the predominant fatty acids were C18 : 0 3-OH, C10 : 0, C14 : 0 and C18 : 1ω9c. Comparison of 16S rRNA gene and genome sequences confirmed that these strains represented a novel member of the genus Francisella, with less than 98.8 % 16S rRNA gene sequence similarity and less than 95 % genomic average nucleotide identity to recognized Francisella species. The phylogenetic tree based on 16S rRNA gene sequences and the protein-concatamer tree based on a concatenation of 28 protein marker sequences both indicated that the strains clustered with 'Francisella salina' TX07-7308 and 'Francisella marina' E95-16, but formed a distinct lineage group among the other members of the genus Francisella. The DNA G+C contents of the four strains were determined to be 32.9, 32.7, 32.9 and 32.9 %, respectively (genome). On the basis of phenotypic and genotypic features, the strains are considered to represent a novel species of the genus Francisella, for which the name Francisella salimarina sp. nov. is proposed. The type strain is SYSU SYW-1T (=CGMCC 1.17031T=NBRC 113781T).

Microvirga arsenatis sp. nov., an arsenate reduction bacterium isolated from Tibet hot spring sediments.

Two novel Gram-stain negative, moderately thermophilic, aerobic, rod-shaped strains, designated 3D203T and 3D207, were isolated from hot spring sediment samples collected from Tibet, western China. Phylogenetic analyses based on 16S rRNA gene sequence similarities showed that two isolates belonged to the genus Microvirga and were most closely related to Microvirga makkahensis SV1470T (98.5% and 98.4%, respectively) and two strains had 99.8% similarity to each other. The average nucleotide identity (ANI) based on whole genome sequences of two strains and M. makkahensis SV1470T was 80.8% and 80.78%, respectively. Optimum growth was observed at 45 °C, pH 7.0 and 0.5% NaCl. They both could tolerate to high concentration arsenic. Ubiquinone 10 (Q10) was their predominant quinone. The differences of strains 3D203T and 3D207 were phosphatidyl dimethyl ethanolamine, phosphatidyl-N-methylethanolamine, phosphatidylglycerol, unidentified glycolipids and unidentified lipids. The major fatty acids (> 5%) were identified C18:1ω7c and/or C18:1ω6c, C18:0 and C16:0. The genomic DNA G + C contents of strain 3D203T and 3D207 based on whole genome sequences were 64.8% and 64.7%, respectively. Phenotypic, chemotaxonomic, phylogenetic and genomic analyses suggested that two strains represent a novel species of the genus Microvirga, for which the name Microvirga arsenatis sp. nov. is proposed. The type strain is 3D203T (= CGMCC 1.17691T = KCTC 72653T).

Rhabdaerophilum calidifontis gen. nov., sp. nov., a novel bacterium isolated from a hot spring, and proposal of Rhabdaerophilaceae fam. nov.

A novel thermotolerant bacterial strain was isolated from a hot spring in a Tibetan geothermal field. Phylogenetic analysis of the 16S rRNA gene sequence of the novel strain showed that it belongs to a distinct lineage far from any known taxa. The new isolate shared the highest pairwise sequence identity with Phreatobacter cathodiphilus S-12T (92.8 % similarity) according to the 16S rRNA gene sequences. Cells were Gram-stain-negative, aerobic, rod-shaped and formed white round colonies. The strain grew at the ranges of 28-45 °C (optimum, 37 °C), pH 5.0-7.0 (optimum, pH 6.0) and 0-2 % NaCl. The strain was positive for catalase and oxidase. The major respiratory quinone was ubiquinone Q-10. Polar lipids consisted of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol and phosphatidylcholine. The major fatty acids were summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c). The DNA G+C content was 68.3 mol%. Based on these distinguishing properties, this strain is proposed to represent a new species of a new genus Rhabdaerophilum calidifontis gen. nov., sp. nov., within a new family Rhabdaerophilaceae fam. nov. The type strain of the type species of Rhabdaerophilum calidifontis is SYSU G02060T (=KCTC 72351T=CGMCC 1.17070T).

Rhodobacter flagellatus sp. nov., a thermophilic bacterium isolated from a hot spring.

A thermophilic bacterium, designated SYSU G03088T, was isolated from Moincer hot spring, Tibet, PR China. Polyphasic taxonomic analyses and whole-genome sequencing were used to determine the taxonomic position and genomic profiles of the strain. Phylogenetic analysis using 16S rRNA gene sequence indicated that SYSU G03088T showed highest sequence similarity to Rhodobacter blasticus CGMCC 1.3365T (96.0 % sequence identity). The strain could be differentiated from most recognized species of the genus Rhodobacter by its slightly purple colony colour, distinct phenotypic characters and low ANI values. Cells were Gram-staining negative, and oval-to-rod shaped. Poly-ß-hydroxybutyrate and vesicular intracytoplasmic membrane structures were formed inside cells. Growth occurred optimally at 45 °C and pH 7.0. Ubiquinone 10 was the only respiratory quinone. The major fatty acids (>10 %) were C18 : 0, C18 : 1ω7c 11-methyl and summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c). The detected polar lipids of SYSU G03088T included diphosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol and phosphatidylmethylethanolamine. The DNA G+C content of SYSU G03088T was 67.7 % (genome). On the basis of the differences in the phenotypic characteristics and phylogenetic analyses, SYSU G03088T is considered to represent a novel species of the genus Rhodobacter, for which the name Rhodobacter flagellatus sp. nov. is proposed. The type strain is SYSU G03088T (=CGMCC 1.16876T=KCTC 72354T).

A xylan-degrading thermophilic and obligate anaerobe Xylanivirga thermophila gen. nov., sp. nov., isolated from an anammox dominant wastewater treatment plant, and proposal of Xylanivirgaceae fam. nov.

In our search for novel anaerobes with potential carbohydrate polymers degrading activity, we have isolated a xylan-degrading bacterial strain SYSU GA17129T from an anammox bacteria dominant wastewater treatment plant. Phylogenetic analysis of the 16S rRNA gene sequence indicated the strain SYSU GA17129T belong to the order Clostridiales and shared highest sequence identity with Caldicoprobacter faecalis DSM 20678T (89.9%). The strain was thermophilic, obligately anaerobic, non-motile and rod shaped. Optimum growth of the strain was observed at 45 °C, pH 8.0 and in the presence of 0.5% NaCl (w/v). The chemotaxonomic features of the strain SYSU GA17129T comprised of C14:0 FAME, iso-C15:0 FAME and C16:0 FAME as the major fatty acids (>10%), diphosphatidylglycerol, phosphatidylinositol mannoside, an unidentified phospholipid, three unidentified polar lipids and two unidentified glycolipids as its polar lipids, and meso-diaminopimelic acid (meso-DAP) as the diamino acid in peptidoglycan. The G + C content of the genomic DNA was 35.9%. The strain could be distinguished from other defined families within the order Clostridiales by the differences in phenotypic and physiological characteristics, distinct phylogenetic lineage in 16S rRNA gene- and genome-based phylogenies and low genomic relatedness index. Based on these distinguishing properties, strain SYSU GA17129T is proposed to represent a new species of a new genus Xylanivirga thermophila gen. nov., sp. nov., within a new family Xylanivirgaceae fam. nov. The type species of the new taxon is SYSU GA17129T (=KCTC 15754T = CGMCC 1.5282T). This strain is characterized within the order Clostridiales, class Clostridia of the phylum Firmicutes.

Rhabdothermincola sediminis gen. nov., sp. nov., a new actinobacterium isolated from hot spring sediment, and emended description of the family Iamiaceae.

One thermophilic bacterium, designated strain SYSU G02662T, was isolated from hot spring sediment sampled in Tibet, PR China. Polyphasic taxonomic analyses and whole-genome sequencing were used to determine the taxonomy position of the strain. Phylogenetic analysis using 16S rRNA gene sequences indicated that strain SYSU G02662T showed the highest sequence similarity to Actinomarinicola tropica SCSIO 58843T (95.1 %). The strain could be differentiated from other species of the family Iamiaceae by its distinct phenotypic and genotypic characteristics. Cells of strain SYSU G02662T were aerobic, Gram-staining-positive and short rodshaped. Growth occurred optimally at 45 °C and pH 7.0. In addition, meso-diaminopimelic acid was the diagnostic diamino acid in the cell-wall peptidoglycan. The respiratory quinone was MK-9 (H8), while the major fatty acids (>10 %) were C16 : 0, C17 : 0, C18 : 0 and iso-C16 : 0. The detected polar lipids included diphosphatidylglycerol, phosphatidylinositol mannoside and phosphatidylinositol. The G+C content of the genomic DNA was 70.5 % based on the draft genomic sequence. On the basis of phenotypic, genotypic and phylogenetic data, strain SYSU G02662T represents a novel species of a novel genus in the family Iamiaceae, for which the name Rhabdothermincola sediminis gen. nov., sp. nov. is proposed. The type strain of the proposed novel species is SYSU G02662T (=CGMCC 4.7688T=KCTC 49500T).