Sekiguchi, Yuji

Summary Oxidation of nitrite to nitrate is an important process in the global nitrogen cycle. Recent molecular biology‐based studies have revealed that the widespread nitrite‐oxidizing bacteria (NOB) belonging to the genus ‘ Candidatus Nitrotoga’ may be highly important for the environment. However, the insufficient availability of pure Nitrotoga cultures has limited our understanding of their physiological and genomic characteristics. Here, we isolated the ‘ Ca . Nitrotoga’ sp. strain AM1P, from a previously enriched Nitrotoga culture, using an improved isolation strategy. Although ‘ Ca . Nitrotoga’ have been recognized as cold‐adapted NOB, the strain AM1P had a slightly higher optimum growth temperature at 23°C. Strain AM1P showed a pH optimum of 8.3 and was not inhibited even at high nitrite concentrations (20 mM). We obtained the complete genome of the strain and compared the genome profile to five previously sequenced ‘ Ca . Nitrotoga’ strains. Comparative genomics suggested that lactate dehydrogenase may be only encoded in the strain AM1P and closely related genomes. While the growth yield of AM1P did not change, we observed faster growth in the presence of lactate in comparison to purely chemolithoautotrophic growth. The characterization of the new strain AM1P sheds light on the physiological adaptation of this environmentally important, but understudied genus ‘ Ca . Nitrotoga’.

One thermophilic (strain IMO-1T) and two mesophilic (strains KIBI-1Tand YMTK-2T) non-spore-forming, non-motile, Gram-negative, multicellular filamentous micro-organisms, which were previously isolated as members of the tentatively named class ‘Anaerolineae’ of the phylumChloroflexi, were characterized. All isolates were strictly anaerobic micro-organisms. The length of the three filamentous isolates was greater than 100 μm and the width was 0.3–0.4 μm for strain IMO-1T, 0.4–0.5 μm for strain KIBI-1Tand thinner than 0.2 μm for strain YMTK-2T. Strain IMO-1Tcould grow at pH 6.0–7.5 (optimum growth at pH 7.0). The optimal temperature for growth of strain IMO-1Twas around 50 °C (growth occurred between 42 and 55 °C). Growth of the mesophilic strains KIBI-1Tand YMTK-2Toccurred at pH 6.0–7.2 with optimal growth at pH 7.0. Both of the mesophilic strains were able to grow in a temperature range of 25–50 °C with optimal growth at around 37 °C. Yeast extract was required for growth of all three strains. All the strains could grow with a number of carbohydrates in the presence of yeast extract. The G+C contents of the DNA of strains IMO-1T, KIBI-1Tand YMTK-2Twere respectively 53.3, 59.5 and 48.2 mol%. Major fatty acids for thermophilic strain IMO-1Twere anteiso-C17 : 0, iso-C15 : 0, C16 : 0and anteiso-C15 : 0, whereas those for mesophilic strains KIBI-1Tand YMTK-2Twere branched C14 : 0, iso-C15 : 0, C16 : 0and branched C17 : 0, and branched C17 : 0, C16 : 0, C14 : 0and C17 : 0, respectively. Detailed phylogenetic analyses based on their 16S rRNA gene sequences indicated that the isolates belong to the class-level taxon ‘Anaerolineae’ of the bacterial phylumChloroflexi, which for a long time had been considered as a typical uncultured clone cluster. Their morphological, physiological, chemotaxonomic and genetic traits strongly support the conclusion that these strains should be described as three novel independent taxa in the phylumChloroflexi. Here,Anaerolinea thermolimosasp. nov. (type strain IMO-1T=JCM 12577T=DSM 16554T),Levilinea saccharolyticagen. nov., sp. nov. (type strain KIBI-1T=JCM 12578T=DSM 16555T) andLeptolinea tardivitalisgen. nov., sp. nov. (type strain YMTK-2T=JCM 12579T=DSM 16556T) are proposed. In addition, we formally propose to subdivide the tentative class-level taxon ‘Anaerolineae’ intoAnaerolineaeclassis nov. andCaldilineaeclassis nov. We also propose the subordinate taxaAnaerolinealesord. nov.,Caldilinealesord. nov.,Anaerolineaceaefam. nov. andCaldilineaceaefam. nov.