Thermospira aquatica gen. nov., sp. nov., a novel thermophilic spirochete isolated from a Tunisian hot spring, and description of the novel family Thermospiraceae.

A novel thermophilic, anaerobic bacterium, strain F1F22T, was isolated from hot spring water collected in northern Tunisia. The cells were non-motile, Gram-negative and helical with hooked ends, 0.5×10-32 µm in size. Growth of the strain was observed at 45-70 °C (optimum, 55 °C), in 0.0-1.0 % (w/v) NaCl (optimum without NaCl) and at pH 6.5-8.5 (optimum, pH 7.5). Yeast extract was required for growth, and the strain grew on glucose, sucrose and maltose. The major fatty acids were C16:0 (40.2 %), iso-C16: 0 (30.2 %) and C16 :0 DMA (14.5 %). The genome consisted of a circular chromosome (2.5 Mb) containing 2672 predicted protein-encoding genes with a G+C content of 43.15 mol %. Based on a comparative 16S rRNA gene sequence analysis, strain F1F22T formed a deeply branching lineage within the phylum Spirochaetota, class Spirochaetia, order Brevinematales, and had only low sequence similarity to other species of the phylum (lower than 83 %). Genome-based analysis of average nucleotide identity and digital DNA-DNA hybridization of strain F1F22T with Treponema caldarium DSM 7334T, Brevinema andersonii ATCC 43811T and Spirochaeta thermophila DSM 6578T showed values between 63.26 and 63.52 %, and between 20 and 25 %. Hence, we propose strain F1F22T as a representative of a novel family (Thermospiraceae fam. nov.), genus and species of Brevinematales: Thermospira aquatica gen. nov., sp. nov. (type strain F1F22T=JCM 31314T=DSM 101182T).

Bacterial diversity of an acid mine drainage beside the Xichú River (Mexico) accessed by culture-dependent and culture-independent approaches.

Xichú River is a Mexican river located in an environmental preservation area called Sierra Gorda Biosphere Reserve. Around it, there are tons of abandoned mine residues that represent a serious environmental issue. Sediment samples of Xichú River, visibly contaminated by flows of an acid mine drainage, were collected to study their prokaryotic diversity. The study was based on both cultural and non-cultural approaches. The analysis of total 16S rRNA gene by MiSEQ sequencing allowed to identify 182 Operational Taxonomic Units. The community was dominated by Pseudomonadota, Bacteroidota, "Desulfobacterota" and Acidobacteriota (27, 21, 19 and 16%, respectively). Different culture conditions were used focusing on the isolation of anaerobic bacteria, including sulfate-reducing bacteria (SRB) and arsenate-reducing bacteria (ARB). Finally, 16 strains were isolated. Among them, 12 were phylogenetically identified, with two strains being SRB, belonging to the genus Solidesulfovibrio ("Desulfobacterota"), while ten are ARB belonging to the genera Azospira (Pseudomonadota), Peribacillus (Bacillota), Raineyella and Propionicimonas (Actinomycetota). The isolate representative of Raineyella genus probably corresponds to a new species, which, besides arsenate, also reduces nitrate, nitrite, and fumarate.

Alkaliphilus serpentinus sp. nov. and Alkaliphilus pronyensis sp. nov., two novel anaerobic alkaliphilic species isolated from the serpentinite-hosted Prony Bay Hydrothermal Field (New Caledonia).

Two novel anaerobic alkaliphilic strains, designated as LacTT and LacVT, were isolated from the Prony Bay Hydrothermal Field (PBHF, New Caledonia). Cells were motile, Gram-positive, terminal endospore-forming rods, displaying a straight to curved morphology during the exponential phase. Strains LacTT and LacVT were mesophilic (optimum 30°C), moderately alkaliphilic (optimum pH 8.2 and 8.7, respectively) and halotolerant (optimum 2% and 2.5% NaCl, respectively). Both strains were able to ferment yeast extract, peptone and casamino acids, but only strain LacTT could use sugars (glucose, maltose and sucrose). Both strains disproportionated crotonate into acetate and butyrate. Phylogenetic analysis revealed that strains LacTT and LacVT shared 96.4% 16S rRNA gene sequence identity and were most closely related to A. peptidifermentans Z-7036, A. namsaraevii X-07-2 and A. hydrothermalis FatMR1 (95.7%-96.3%). Their genome size was of 3.29Mb for strain LacTT and 3.06Mb for strain LacVT with a G+C content of 36.0 and 33.9mol%, respectively. The ANI value between both strains was 73.2 %. Finally, strains LacTT (=DSM 100337=JCM 30643) and LacVT (=DSM 100017=JCM 30644) are proposed as two novel species of the genus Alkaliphilus, order Clostridiales, phylum Firmicutes, Alkaliphilus serpentinus sp. nov. and Alkaliphilus pronyensis sp. nov., respectively. The genomes of the three Alkaliphilus species isolated from PBHF were consistently detected in the PBHF chimney metagenomes, although at very low abundance, but not significantly in the metagenomes of other serpentinizing systems (marine or terrestrial) worldwide, suggesting they represent indigenous members of the PBHF microbial ecosystem.

Towards a congruent reclassification and nomenclature of the thermophilic species of the genus Pseudothermotoga within the order Thermotogales.

The phylum Thermotogae gathers thermophilic, hyperthermophic, mesophilic, and thermo-acidophilic anaerobic bacteria that are mostly originated from geothermally heated environments. The metabolic and phenotypic properties harbored by the Thermotogae species questions the evolutionary events driving the emergence of this early branch of the universal tree of life. Recent reshaping of the Thermotogae taxonomy has led to the description of a new genus, Pseudothermotoga, a sister group of the genus Thermotoga within the order Thermotogales. Comparative genomics of both Pseudothermotoga and Thermotoga spp., including 16S-rRNA-based phylogenetic, pan-genomic analysis as well as signature indel conservation, provided evidence that Thermotoga caldifontis and Thermotoga profunda species should be reclassified within the genus Pseudothermotoga and renamed as Pseudothermotoga caldifontis comb. nov. (type strain=AZM44c09T) and Pseudothermotoga profunda comb. nov. (type strain=AZM34c06T), respectively. In addition, based upon whole-genome relatedness indices and DNA-DNA Hybridization results, the reclassification of Pseudothermotoga lettingae and Pseudothermotoga subterranea as latter heterotypic synonyms of Pseudothermotoga elfii is proposed. Finally, potential genetic elements resulting from the distinct evolutionary story of the Thermotoga and Pseudothermotoga clades are discussed.

Characterization of Desulfovibrio salinus sp. nov., a slightly halophilic sulfate-reducing bacterium isolated from a saline lake in Tunisia.

A novel slightly halophilic sulfate-reducing bacterium, designated strain P1BSRT, was isolated from water of a saline lake in Tunisia. Strain P1BSRT had motile (single polar flagellum), Gram-negative, rod-shaped, non-spore-forming cells, occurring singly or in pairs. Strain P1BSRT grew at temperatures between 15 and 45 °C (optimum 40 °C), and in a pH range between 6 and 8.5 (optimum pH 6.7). The strain required NaCl for growth (1 % w/v), and tolerated high NaCl concentration (up to 12 % w/v) with an optimum of 3 % (w/v). Sulfate, thiosulfate and sulfite served as terminal electron acceptors, but not elemental sulfur, fumarate, nitrate and nitrite. Strain P1BSRT utilized lactate, pyruvate, formate, d-fructose and glycerol as carbon and energy sources. The main cellular fatty acid was C16 : 0 (50.8 %). The genomic DNA G+C content was 47.7 mol%. Phylogenetic analysis of 16S rRNA gene sequence similarity indicated that strain P1BSRT was affiliated to the genus Desulfovibrio, with the type strains Desulfovibrio salexigens (96.51 %), Desulfovibrio zosterae (95.68 %), Desulfovibrio hydrothermalis (94.81 %) and Desulfovibrio ferrireducens (94.73 %) as its closest phylogenetic relatives. On the basis of genotypic, phenotypic and phylogenetic characteristics, it is proposed to assign strain P1BSRT to a novel species of the genus Desulfovibrio, Desulfovibrio salinus sp. nov. The type strain is P1BSRT (=DSM 101510T=JCM 31065T).

Transfer of Pseudomonas pictorum Gray and Thornton 1928 to genus Stenotrophomonas as Stenotrophomonas pictorum comb. nov., and emended description of the genus Stenotrophomonas.

A polyphasic taxonomic approach including analysis of phenotypic, physiological and genotypic characteristics, 16S rRNA gene sequence and DNA-DNA hybridization analysis was used to determine the most consistent affiliation of Pseudomonas pictorum. Pseudomonas pictorum ATCC 23328T exhibited phenotypic traits of members of the genus Stenotrophomonas including cellular fatty acid composition, quinone and limited range of substrates that could be used. Antibiotic susceptibility and physiological characteristics were determined. The DNA G+C content was 65.7 mol%. Phylogenetic analysis revealed that the type strains of Stenotrophomonas terrae, Stenotrophomonashumi, Stenotrophomonasnitritireducens and Stenotrophomonasacidaminiphila were the nearest relatives (16S rRNA gene sequence similarity of 98.0 to 98.8 %). All the other type strains of species of the genus Stenotrophomonas showed high 16S rRNA gene sequence similarities (96.8 to 97.2 %). DNA-DNA hybridizations revealed 31.0, 32.0, 43.3 and 43.6 % reassociation between Pseudomonas pictorum ATCC 23328T and the type strains of S. terrae, S. humi, S. nitritireducens and S. acidaminiphila, respectively. Our overall results indicate that Pseudomonas pictorum should be transferred to the genus Stenotrophomonas as a novel species of this genus, Stenotrophomonas pictorum comb. nov. Since the original description of the genus Stenotrophomonaswas made with only one species (Stenotrophomonasmaltophilia), an emendation of the genus description is proposed in order to match better with the characteristics of the eleven novel species assigned to this genus since then.

Characterization of the first cultured representative of a Bacteroidetes clade specialized on the scavenging of cyanobacteria.

The anaerobic, mesophilic and moderately halophilic strain L21-Spi-D4T was recently isolated from the suboxic zone of a hypersaline cyanobacterial mat using protein-rich extracts of Arthrospira (formerly Spirulina) platensis as substrate. Phylogenetic analyses based on 16S rRNA genes indicated an affiliation of the novel strain with the Bacteroidetes clade MgMjR-022, which is widely distributed and abundant in hypersaline microbial mats and heretofore comprised only sequences of uncultured bacteria. Analyses of the complete genome sequence of strain L21-Spi-D4T revealed a possible specialization on the degradation of cyanobacterial biomass. Besides genes for enzymes degrading specific cyanobacterial proteins a conspicuous transport complex for the polypeptide cyanophycin could be identified that is homologous to typical polysaccharide utilization loci of Bacteroidetes. A distinct and reproducible co-occurrence pattern of environmental 16S rRNA gene sequences of the MgMjR-022 clade and cyanobacteria in the suboxic zone of hypersaline mats points to a specific dependence of members of this clade on decaying cyanobacteria. Based on a comparative analysis of phenotypic, genomic and ecological characteristics we propose to establish the novel taxa Salinivirga cyanobacteriivorans gen. nov., sp. nov., represented by the type strain L21-Spi-D4T , and Salinivirgaceae fam. nov., comprising sequences of the MgMjR-022 clade.

Serpentinicella alkaliphila gen. nov., sp. nov., a novel alkaliphilic anaerobic bacterium isolated from the serpentinite-hosted Prony hydrothermal field, New Caledonia.

A novel anaerobic, alkaliphilic, Gram-stain-positive, spore-forming bacterium was isolated from a carbonaceous hydrothermal chimney in Prony Bay, New Caledonia. This bacterium, designated strain 3bT, grew at temperatures from 30 to 43 °C (optimum 37 °C) and at pH between 7.8 and 10.1 (optimum 9.5). Added NaCl was not required for growth (optimum 0-0.2 %, w/v), but was tolerated at up to 4 %. Yeast extract was required for growth. Strain 3bT utilized crotonate, lactate and pyruvate, but not sugars. Crotonate was dismutated to acetate and butyrate. Lactate was disproportionated to acetate and propionate. Pyruvate was degraded to acetate plus trace amounts of hydrogen. Growth on lactate was improved by the addition of fumarate, which was used as an electron acceptor and converted to succinate. Sulfate, thiosulfate, elemental sulfur, sulfite, nitrate, nitrite, FeCl3, Fe(III)-citrate, Fe(III)-EDTA, chromate, arsenate, selenate and DMSO were not used as terminal electron acceptors. The G+C content of the genomic DNA was 33.2 mol%. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that the isolate is a member of the family Clostridiaceae, order Clostridiales within the phylum Firmicutes. Strain 3bT was most closely related to 'Alkaliphilus hydrothermalis' FatMR1T (92.2 % 16S rRNA gene sequence similarity), and was positioned approximately equidistantly between the genera Alkaliphilus, Anaerovirgula and Natronincola. On the basis of phylogenetic, genetic, chemotaxonomic and physiological properties, strain 3bT is proposed to represent a novel species of a new genus, for which the name Serpentinicella alkaliphila gen. nov., sp. nov. is proposed. The type strain of Serpentinicella alkaliphila is 3bT (=DSM 100013T=JCM 30645T).

Desulfotomaculum aquiferis sp. nov. and Desulfotomaculum profundi sp. nov., isolated from a deep natural gas storage aquifer.

Two novel strictly anaerobic bacteria, strains Bs105T and Bs107T, were isolated from a deep aquifer-derived hydrocarbonoclastic community. The cells were rod-shaped, not motile and had terminal spores. Phylogenetic affiliation and physiological properties revealed that these isolates belong to two novel species of the genus Desulfotomaculum. Optimal growth temperatures for strains Bs105T and Bs107T were 42 and 45 °C, respectively. The estimated G+C content of the genomic DNA was 42.9 and 48.7 mol%. For both strains, the major cellular fatty acid was palmitate (C16 : 0). Specific carbon fatty acid signatures of Gram-positive bacteria (iso-C17 : 0) and sulfate-reducing bacteria (C17 : 0cyc) were also detected. An insertion was revealed in one of the two 16S rRNA gene copies harboured by strain Bs107T. Similar insertions have previously been highlighted among moderately thermophilic species of the genus Desulfotomaculum. Both strains shared the ability to oxidize aromatic acids (Bs105T: hydroquinone, acetophenone, para-toluic acid, 2-phenylethanol, trans-cinnamic acid, 4-hydroxybenzaldehyde, benzyl alcohol, benzoic acid 4-hydroxybutyl ester; Bs107T: ortho-toluic acid, benzoic acid 4-hydroxybutyl ester). The names Desulfotomaculum aquiferis sp. nov. and Desulfotomaculum profundi sp. nov. are proposed for the type strains Bs105T (=DSM 24088T=JCM 31386T) and Bs107T (=DSM 24093T=JCM 31387T).

Mobilisporobacter senegalensis gen. nov., sp. nov., an anaerobic bacterium isolated from tropical shea cake.

A Gram-stain positive, endospore-forming, strictly anaerobic bacterium, designated strain Gal1T, was isolated from shea cake, a waste material from the production of shea butter, originating from Saraya, Senegal. The cells were rod-shaped, slightly curved, and motile with peritrichous flagella. The strain was oxidase-negative and catalase-negative. Growth was observed at temperatures ranging from 15 to 45 °C (optimum 30 °C) and at pH 6.5-9.3 (optimum pH 7.8). The salinity range for growth was 0-3.5 % NaCl (optimum 1 %). Yeast extract was required for growth. Strain Gal1T fermented various carbohydrates such as mannose, mannitol, arabinose, cellobiose, fructose, glucose, maltose, sucrose, trehalose and lactose and the major end-products were ethanol and acetate. The only major cellular fatty acid was C16 : 0 (19.6 %). The DNA base G+C content of strain Gal1T was 33.8 mol%. Analysis of the 16S rRNA gene sequence of the isolate indicated that this strain was related to Mobilitalea sibirica DSM 26468T with 94.27 % similarity, Clostridium populeti ATTC 35295T with 93.94 % similarity, and Clostridium aminovalericum DSM 1283T and Anaerosporobacter mobilis DSM 15930T with 93.63 % similarity. On the basis of phenotypic characteristics, phylogenetic analysis and the results of biochemical and physiological tests, strain Gal1T was clearly distinguished from closely related genera, and strain Gal1T can be assigned to a novel species of a new genus for which the name Mobilisporobacter senegalensis gen. nov., sp. nov. is proposed. The type strain is Gal1T ( = DSM 26537T = JCM 18753T).