An extremely thermophilic Methanococcus from a deep sea hydrothermal vent and its plasmid

An extremely thermophilic methanogen was isolated from a hydrothermal vent core sample from Guaymas Basin, Gulf of California, at a depth of 2003 m. The isolate, designated strain AG86, was a coccoid autotroph using H₂-CO₂ as energy and carbon source with a growth temperature range of 48 to 92°C, optimum, 85°C. AG86 required NaCl and Mg²⁺ and trace amounts of selenite and tungstate. Vitamins were not required. However, yeast extract, Casamino acids and Trypticase stimulated growth significantly. When grown in the presence of these stimulants and at the optimal growth temperature and pH 6.5, the minimum doubling time was 20 min. Cells were fragile and readily lysed by detergents. The mol% G+C was 33%. These results and partial 16S rRNA sequencing indicated that AG86 belonged to the genus Methanococcus and closely resembled Methanococcus jannaschii. Tests for extrachromosomal DNA revealed a plasmid in AG86 and two plasmids in M. jannaschii. Different patterns were obtained from restriction endonuclease digestion of the three plasmids, and no homology was observed with DNA-DNA hybridization.Abbreviations CCC DNA covalently close circular DNA - DM defined marine medium - G+C Guanine plus cytosine - MPN most probable number