Kublanov, Ilya V.

A novel aerobic moderately thermophilic bacterium, strain 3753OT, was isolated from a Chukotka hot spring (Arctic, Russia) using the newly developed technology of laser engineering of microbial systems. Сells were regular short rods, 0.4×0.8–2.0 µm in size, with a monoderm-type envelope and a single flagellum. The temperature and pH ranges for growth were 42–60 °C and pH 6.5–8.5, the optima being 50–54 °C and pH 7.3. Strain 3753OT grew chemoorganoheterotrophically on a number of carbohydrates or peptidic substrates and volatile fatty acids, and chemolithoautotrophically with siderite (FeCO3) as the electron donor. The major cellular fatty acid was branched C19 : 0. Phosphatidylethanolamine, phosphatidylglycerol and two unidentified phospholipids as well as two yellow carotenoid-type pigments were detected in the polar lipid extract. Strain 3753OT was inhibited by chloramphenicol, polymyxin B, vancomycin, streptomycin, neomycin and kanamycin, but resistant to the action of novobiocin and ampicillin. The DNA G+C content was 69.9 mol%. The 16S rRNA gene as well as 51 conservative protein sequence-based phylogenetic analyses placed strain 3753OT within the previously uncultivated lineage OLB14 in the phylum Chloroflexi . Taking into account the phylogenetic position as well as phenotypic properties of the novel isolate, the novel genus and species Tepidiforma bonchosmolovskayae gen. nov., sp. nov., within the Tepidiformaceae fam. nov., the Tepidiformales ord. nov. and the Tepidiformia classis nov. are proposed. The type strain of Tepidiforma bonchosmolovskayae is 3753OT (=VKM B-3389T=KTCT 72284T).

Summary A novel moderately thermophilic, facultatively anaerobic chemoorganotrophic bacterium strain P3M ‐2 T was isolated from a microbial mat developing on the wooden surface of a chute under the flow of hot water (46° C ) coming out of a 2775‐m‐deep oil exploration well ( T omsk region, R ussia). Strain P3M ‐2 T is a moderate thermophile and facultative anaerobe growing on mono‐, di‐ or polysaccharides by aerobic respiration, fermentation or by reducing diverse electron acceptors [nitrite, F e( III ), As ( V )]. Its closest cultivated relative (90.8% rRNA gene sequence identity) is I gnavibacterium album , the only chemoorganotrophic member of the phylum C hlorobi . New genus and species M elioribacter roseus are proposed for isolate P3M ‐2 T . Together with I . album , the new organism represents the class I gnavibacteria assigned to the phylum Chlorobi . The revealed group includes a variety of uncultured environmental clones, the 16S rRNA gene sequences of some of which have been previously attributed to the candidate division ZB1 . Phylogenetic analysis of M . roseus and I . album based on their 23S rRNA and RecA sequences confirmed that these two organisms could represent an even deeper, phylum‐level lineage. Hence, we propose a new phylum Ignavibacteriae within the Bacteroidetes – C hlorobi group with a sole class I gnavibacteria , two families I gnavibacteriaceae and M elioribacteraceae and two species I . album and M . roseus . This proposal correlates with chemotaxonomic data and phenotypic differences of both organisms from other cultured representatives of C hlorobi . The most essential differences, supported by the analyses of complete genomes of both organisms, are motility, facultatively anaerobic and obligately organotrophic mode of life, the absence of chlorosomes and the apparent inability to grow phototrophically.

Two novel thermophilic and slightly acidophilic strains, Kam940Tand Kam1507b, which shared 99 % 16S rRNA gene sequence identity, were isolated from terrestrial hot springs of the Uzon caldera on the Kamchatka peninsula. Cells of both strains were non-motile, regular cocci. Growth was observed between 55 and 85 °C, with an optimum at 65–70 °C (doubling time, 6.1 h), and at pH 4.5–7.5, with optimum growth at pH 5.5–6.0. The isolates were strictly anaerobic organotrophs and grew on a narrow spectrum of energy-rich substrates, such as beef extract, gelatin, peptone, pyruvate, sucrose and yeast extract, with yields above 107cells ml−1. Sulfate, sulfite, thiosulfate and nitrate added as potential electron acceptors did not stimulate growth when tested with peptone. H2at 100 % in the gas phase inhibited growth on peptone. Glycerol dibiphytanyl glycerol tetraethers (GDGTs) with zero to four cyclopentyl rings were present in the lipid fraction of isolate Kam940T. The G+C content of the genomic DNA of strain Kam940Twas 37 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the isolates were archaea of the phylumCrenarchaeota, only distantly related to the cultured members of the classThermoprotei(no more than 89 % identity), and formed an independent lineage adjacent to the ordersDesulfurococcalesandAcidilobalesand clustering only with uncultured clones from hot springs of Yellowstone National Park and Iceland as the closest relatives. On the basis of their phylogenetic position and novel phenotypic features, isolates Kam940Tand Kam1507b are proposed to be assigned to a new genus and species,Fervidicoccus fontisgen. nov., sp. nov. The type strain ofFervidicoccus fontisis strain Kam940T(=DSM 19380T=VKM B-2539T). The phylogenetic data as well as phenotypic properties suggest that the novel crenarchaeotes form the basis of a new family,Fervidicoccaceaefam. nov., and order,Fervidicoccalesord. nov., within the classThermoprotei.