A novel acidiphilic, hydrogenotrophic methanogen, designated strain 6A8T, was isolated from an acidic (pH 4.0–4.5) and ombrotrophic (rain-fed) bog located near Ithaca, NY, USA. Cultures were dimorphic, containing thin rods (0.2–0.3 μm in diameter and 0.8–3.0 μm long) and irregular cocci (0.2–0.8 μm in diameter). The culture utilized H2/CO2 to produce methane but did not utilize formate, acetate, methanol, ethanol, 2-propanol, butanol or trimethylamine. Optimal growth conditions were near pH 5.1 and 35 °C. The culture grew in basal medium containing as little as 0.43 mM Na+ and growth was inhibited completely by 50 mM NaCl. To our knowledge, strain 6A8T is one of the most acidiphilic (lowest pH optimum) and salt-sensitive methanogens in pure culture. Acetate, coenzyme M, vitamins and yeast extract were required for growth. It is proposed that a new genus and species be established for this organism, Methanoregula boonei gen. nov., sp. nov. The type strain of Methanoregula boonei is 6A8T (=DSM 21154T =JCM 14090T).
A novel methane-producing archaeon, strain SMSPT, was isolated from an anaerobic, propionate-degrading enrichment culture that was originally obtained from granular sludge in a mesophilic upflow anaerobic sludge blanket (UASB) reactor used to treat a beer brewery effluent. Cells were non-motile, blunt-ended, straight rods, 1.0–2.6 μm long by 0.5 μm wide; cells were sometimes up to 7 μm long. Asymmetrical cell division was observed in rod-shaped cells. Coccoid cells (0.5–1.0 μm in diameter) were also observed in mid- to late-exponential phase cultures. Growth was observed between 10 and 40 °C (optimum, 30–33 °C) and pH 7.0 and 7.6 (optimum, pH 7.4). The G+C content of the genomic DNA was 56.2 mol%. The strain utilized formate and hydrogen for growth and methane production. Based on comparative sequence analyses of the 16S rRNA and mcrA (encoding the alpha subunit of methyl-coenzyme M reductase, a key enzyme in the methane-producing pathway) genes, strain SMSPT was affiliated with group E1/E2 within the order Methanomicrobiales. The closest relative based on both 16S rRNA and mcrA gene sequences was Methanoregula boonei 6A8T (96.3 % 16S rRNA gene sequence similarity, 85.4 % deduced McrA amino acid sequence similarity). The percentage of 16S rRNA gene sequence similarity indicates that strain SMSPT and Methanoregula boonei 6A8T represent different species within the same genus. This is supported by our findings of shared phenotypic properties, including cell morphology and growth temperature range, and phenotypic differences in substrate usage and pH range. Based on these genetic and phenotypic properties, we propose that strain SMSPT represents a novel species of the genus Methanoregula, for which we propose the name Methanoregula formicica sp. nov., with the type strain SMSPT (=NBRC 105244T =DSM 22288T).
Two methane-producing archaea, designated Mic5c12T and Mic6c05T, were isolated from sludge deposited in a crude oil storage tank and a tubercle on the interior of a pipe transporting natural gas-containing brine, respectively. The isolates were Gram-staining-variable, non-motile rods and grew only on H2/CO2. Strain Mic6c05T produced methane from some alcohols without showing any growth; strain Mic5c12T did not utilize alcohols. The optimum growth conditions for strain Mic5c12T were 35 °C, pH 6.5 and 0–0.68 M NaCl and for strain Mic6c05T were 40 °C, pH 6.0–7.5 and 0.34 M NaCl. Strain Mic5c12T was halotolerant and strain Mic6c05T was halophilic. Comparative 16S rRNA gene sequence analysis revealed that strains Mic5c12T and Mic6c05T belonged to the genus Methanobacterium and their closest relative was Methanobacterium subterraneum A8pT (97.3 and 97.9 % 16S rRNA gene sequence similarity, respectively). The findings from the 16S rRNA gene sequence analyses were supported by analysis of McrA, the alpha subunit of methyl-coenzyme M reductase. On the basis of phylogenetic analyses and phenotypic characteristics, two novel species are proposed, Methanobacterium petrolearium sp. nov. and Methanobacterium ferruginis sp. nov., with type strains Mic5c12T (=NBRC 105198T =DSM 22353T) and Mic6c05T (=NBRC 105197T =DSM 21974T), respectively.
A mesophilic, non-motile, hydrogenotrophic, rod-shaped methanogen, designated M2T, was isolated from Holocene permafrost sediments of the Kolyma lowland in the Russian Arctic. Cells were 3–6 μm long and 0.45–0.5 μm wide. Strain M2T grew on H2/CO2 and formate. Optimum conditions for growth were 37 °C, pH 6.8–7.2 and 0.1 M NaCl. The DNA G+C content was 38.1 mol%. On the basis of 16S rRNA gene sequence comparison with known methanogens, strain M2T was affiliated with the genus Methanobacterium and was most closely related to Methanobacterium veterum MK4T and Methanobacterium bryantii DSM 863T (both 99 % 16S rRNA gene sequence similarity). However, no significant DNA–DNA relatedness was observed between strain M2T and these type strains. We propose that strain M2T represents a novel species, with the name Methanobacterium arcticum sp. nov., with type strain M2T (=DSM 19844T =VKM B-2371T).