Methylothermus thermalis gen. nov., sp nov., a novel moderately thermophilic obligate methanotroph from a hot spring in Japan

A novel moderately thermophilic methanotroph, strain MYHT, was isolated from a hot spring in Japan. The isolate grew on methane or methanol at 37-67 degrees C, and optimally at 57-59 degrees C. It was found to be a Gram-negative aerobe, with colourless colonies of non-motile coccoid cells, possessing type I intracytoplasmic membranes and regularly arranged surface layers of linear (p2) symmetry. Strain MYHTT expressed only the particulate methane monooxygenase and employed the ribulose monophosphate pathway for formaldehyde assimilation. It is a neutrophilic and halotolerant organism capable of growth at pH 6(.)5-7(.)5 (optimum pH 6(.)8) and in up to 3% NaCl (optimum 0(.)5-1 % NaCl). Phylogenetic analysis based on 16S rRNA gene sequence analysis indicated that strain MYHT T is Most Closely related to the thermophilic undescribed methanotroph 'Methylothermus' HB (91 % identity) and the novel halophilic methanotroph Methylohalobius crimeensis 10Ki(T) (90% identity). Comparative sequence analysis of particulate methane monooxygenase (pmoA) genes also confirmed the clustering of strain MYHTT with 'Methylothermus' HB and Methylohalobius crimeensis 10Ki(T) (98 and 92 % derived amino acid sequence identity, respectively). The DNA G + C content was 62(.)5 mol%. The major cellular fatty acids were C-16:0 (37(.)2%) and C-18:1 omega 9c (35(.)2%) and the major polar lipids were phosphatidylethanolamine and phosphatidylglycerol. The major ubiquinone was Q-8. On the basis of comparative phenotypic and genotypic characteristics, a new genus and species, Methylothermus thermalis gen. nov., sp. nov., is proposed, with MYHTT as the type strain (=VKM B-2345(T)=IPOD FERM P-19714(T)).