title: Isolation of a methanogen from deep marine sediments that contain methane hydrates, and description of Methanoculleus submarinus sp. nov.
abstract: We isolated a methanogen from deep in the sediments of the Nankai Trough off the eastern coast of Japan. At the sampling site, the water was 950 m deep and the sediment core was collected at 247 m below the sediment surface. The isolated methanogen was named Nankai-1. Cells of Nankai-1 were nonmotile and highly irregular coccoids (average diameter, 0.8 to 2 micro m) and grew with hydrogen or formate as a catabolic substrate. Cells required acetate as a carbon source. Yeast extract and peptones were not required but increased the growth rate. The cells were mesophilic, growing most rapidly at 45 degrees C (no growth at </=10 degrees C or >/=55 degrees C). Cells grew with a maximum specific growth rate of 2.43 day(-1) at 45 degrees C. Cells grew at pH values between 5.0 and 8.7 but did not grow at pH 4.7 or 9.0. Strain Nankai-1 grew in a wide range of salinities, from 0.1 to 1.5 M Na(+). The described phenotypic characteristics of this novel isolate were consistent with the in situ environment of the Nankai Trough. This is the first report of a methanogenic isolate from methane hydrate-bearing sediments. Phylogenetic analysis of its 16S rRNA gene sequence indicated that it is most closely related to Methanoculleus marisnigri (99.1% sequence similarity), but DNA hybridization experiments indicated a DNA sequence similarity of only 49%. Strain Nankai-1 was also found to be phenotypically similar to M. marisnigri, but two major phenotypic differences were found: strain Nankai-1 does not require peptones, and it grows fastest at a much higher temperature. We propose a new species, Methanoculleus submarinus, with strain Nankai-1 as the type strain.
authors.affiliation: Biology Department, Portland State University, Portland, Oregon 97207-0751, USA.
authors.forename: Jill A
authors.initials: JA
authors.lastname: Mikucki
authors.forename: Yitai
authors.initials: Y
authors.lastname: Liu
authors.forename: Mark
authors.initials: M
authors.lastname: Delwiche
authors.forename: Frederick S
authors.initials: FS
authors.lastname: Colwell
authors.forename: David R
authors.initials: DR
authors.lastname: Boone
mesh_terms: D003470:Culture Media; D019641:DNA, Archaeal / Q000032:analysis; D004275:DNA, Ribosomal / Q000032:analysis; D019605:Euryarchaeota / Q000145:classification* / Q000235:genetics* / Q000254:growth & development / Q000302:isolation & purification; D019015:Geologic Sediments / Q000382:microbiology*; D008697:Methane / Q000378:metabolism; D019638:Methanomicrobiaceae / Q000145:classification* / Q000235:genetics* / Q000254:growth & development / Q000302:isolation & purification; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012623:Seawater / Q000382:microbiology*; D017422:Sequence Analysis, DNA
chemical_list: D003470:Culture Media; D019641:DNA, Archaeal; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S; D008697:Methane