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A novel methane-oxidizing bacterium, strain HTM55T, was isolated from subsurface hot aquifer water from a Japanese gold mine. Strain HTM55T was a Gram-negative, aerobic, motile, coccoid bacterium with a single polar flagellum and the distinctive intracytoplasmic membrane arrangement of a type I methanotroph. Strain HTM55T was a moderately thermophilic, obligate methanotroph that grew on methane and methanol at 37–65 °C (optimum 55–60 °C). The isolate grew at pH 5.2–7.5 (optimum 5.8–6.3) and with 0–1 % NaCl (optimum 0–0.3 %). The ribulose monophosphate pathway was operative for carbon assimilation. The DNA G+C content was 54.4 mol% and the major fatty acids were C16 : 0 (52.0 %) and C18 : 1ω7c (34.8 %). Phylogenetic analysis of the 16S rRNA gene sequence indicated that strain HTM55T was closely related to Methylothermus thermalis MYHTT (99.2 % 16S rRNA gene sequence similarity), which is within the class Gammaproteobacteria. However, DNA–DNA relatedness between strain HTM55T and Methylothermus thermalis MYHTT was ≤39 %. On the basis of distinct phylogenetic, chemotaxonomic and physiological characteristics, strain HTM55T represents a novel species of the genus Methylothermus, for which the name Methylothermus subterraneus sp. nov. is proposed. The type strain is HTM55T ( = JCM 13664T = DSM 19750T).
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