Isolation and Characterization of Methanothermobacter crinale sp nov., a Novel Hydrogenotrophic Methanogen from the Shengli Oil Field

Syntrophic acetate oxidation coupled with hydrogenotrophic methanogenesis is an alternative methanogenic pathway in certain thermophilic anaerobic environments such as high-temperature oil reservoirs and thermophilic biogas reactors. In these environments, the dominant thermophilic methanogens were generally related to uncultured organisms of the genus Methanothermobacter. Here we isolated two representative strains, Tm2(T) and HMD, from the oil sands and oil production water in the Shengli oil field in the People's Republic of China. The type strain, Tm2(T), was nonmotile and stained Gram positive. The cells were straight to slightly curved rods (0.3 mu m in width and 2.2 to 5.9 mu m in length), but some of them possessed a coccal shape connecting with the rods at the ends. Strain Tm2(T) grew with H-2-CO2, but acetate is required. Optimum growth of strain Tm2(T) occurred in the presence of 0.025 g/liter NaCl at pH 6.9 and a temperature of 65 degrees C. The G+C content of the genomic DNA was 40.1 mol% +/- 1.3 mol% (by the thermal denaturation method) or 41.1 mol% (by high-performance liquid chromatography). Analysis of the 16S rRNA gene sequence indicated that Tm2(T) was most closely related to Methanothermobacter thermautotrophicus Delta H-T and Methanothermobacter wolfeii VKM B-1829(T) (both with a sequence similarity of 96.4%). Based on these phenotypic and phylogenic characteristics, a novel species was proposed and named Methanothermobacter crinale sp. nov. The type strain is Tm2(T) (ACCC 00699(T) = JCM 17393(T)).