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Nine thermophilic strains of aerobic, non-sporulating, heterotrophic bacteria were isolated after enrichment of chimney material sampled from a deep-sea hydrothermal field at a depth of 2634 m on the East-Pacific Rise (1 °N). The bacteria stained Gram-negative. They were rod-shaped and measured approximately 0.5 μm in width and 1.5–3.5 μm in length. They grew at 55–80 °C, pH 6–8 and 1–6 % NaCl. Optimal growth was observed at 70–75 °C, pH 7.0 and 1–3 % NaCl. The organisms were identified as members of the genus Rhodothermus, having a 16S rRNA gene similarity of 98.1 % with Rhodothermus marinus DSM 4252T. The novel isolates differed morphologically, physiologically and chemotaxonomically from R. marinus, e.g. in lack of pigmentation, response to hydrostatic pressure, maximum growth temperature and DNA G+C content. DNA–DNA hybridization revealed a reassociation value of 37.2 % between strain PRI 2902T and R. marinus DSM 4252T, which strongly suggested that they represent different species. Furthermore, AFLP fingerprinting separated the novel strains from R. marinus reference strains. It is therefore concluded that the strains described here should be classified as representatives of a novel species for which the name Rhodothermus profundi sp. nov. is proposed; the type strain is PRI 2902T (=DSM 22212T =JCM 15944T).

A thermophilic, anaerobic, piezophilic, chemo-organotrophic sulfur-reducing bacterium, designated as KA3T, was isolated from a deep-sea hydrothermal chimney sample collected at a depth of 2630 m on the East-Pacific Rise (13 degrees N). When grown under elevated hydrostatic pressure, the cells are rod-shaped with a sheath-like outer structure, motile, have a mean length of 1-1.5 microm and stain Gram-negative. They appear singly or in short chains. When grown at lower, or atmospheric, pressures, the cells elongate and become twisted. Growth is enhanced by hydrostatic pressure; the optimal pressure for growth is 40 MPa (26 MPa pressure at sampling site). The temperature range for growth is 45-70 degrees C, the optimum being around 65 degrees C (doubling time is approximately 20 min at 40 MPa). Growth is observed from pH 5 to pH 8, the optimum being at pH 6. The salinity range for growth is 10-50 g NaCl l(-1), the optimum being at 30 g l(-1). The isolate is able to grow on a broad spectrum of carbohydrates or complex proteinaceous substrates, and growth is stimulated by L-cystine and elemental sulfur. The G+C content of the genomic DNA is 29 +/- 1 mol%. According to phylogenetic analysis of the 16S rDNA gene, the strain is placed within the order Thermotogales, in the bacterial domain. On the basis of 16S rDNA sequence comparisons and morphological, physiological and genotypic characteristics, it is proposed that the isolate be described as a novel species of the genus Marinitoga, with Marinitoga piezophila sp. nov. as the type species. The type strain is KA3T (= DSM 14283T = JCM 11233T).

A novel barophilic, hyperthermophilic, anaerobic sulfur-metabolizing archaeon, strain MPT (T = type strain), was isolated from a hydrothermal vent site (Snakepit) on the Mid-Atlantic Ridge (depth, 3550 m). Enrichments and isolation were done under 40 MPa hydrostatic pressure at 95 degrees C. Strain MPT was barophilic at 75, 80, 85, 90, 95 and 98 degrees C, and was an obligate barophile between 95 and 100 degrees C (Tmax). For growth above 95 degrees C, a pressure of 15.0-17.5 MPa was required. The strain grew at 48-95 degrees C under atmospheric pressure. The optimal temperature for growth was 85 degrees C at both high (40 MPa) and low (0.3 MPa) pressures. The growth rate was twofold higher at 85 degrees C under in situ hydrostatic pressure compared to at low pressure. Strain MPT cells were motile, coccoid, 0.8-2.0 microns in diameter and covered by a hexagonal S-layer lattice. The optimum pH and NaCl concentration for growth at low pressure were 7.0 and 20-30 g l-1, respectively. The new isolate was an obligate heterotroph and utilized yeast extract, beef extract and peptone for growth. Growth was optimal in the presence of elemental sulfur. Rifampicin and chloramphenicol inhibited growth. The core lipids consisted of a major archaeol and a complex lipid pattern consisting of a major phospholipid. The DNA G + C content was 37.1 mol%. Sequencing of the 16S rRNA gene revealed that strain MPT belonged to the genus Thermococcus and it is proposed that this isolate should be designated as a new species, Thermococcus barophilus.

We performed phenotypic and physiological studies with 20 hyperthermophilic microorganisms isolated from hydrothermal vents located in the North Fiji Basin (southwestern Pacific Ocean) at a depth of 2,000 m. These isolates were strict anaerobes that were regular to irregular coccoids and used elemental sulfur in their metabolism. Growth was observed at temperatures ranging from 50 to 101°C. The DNA base compositions varied from 43 to 60 mol%. All of these organisms were heterotrophs and fermented peptides to acetate, isovalerate, isobutyrate, and propionate. They contained both diether and tetraether lipids in their membranes, which indicates that they belong to the domain Archaea. DNA-DNA hybridization experiments revealed that there were two distinct homology groups, which correlated well with results obtained from sodium dodecyl sulfate-polyacrylamide gel electrophoresis of soluble whole-cell proteins, and these groups corresponded to the genera Pyrococcus and Thermococcus. Five isolates exhibited levels of DNA-DNA relatedness with Pyrococcus abyssi ranging from 71 to 100% and produced almost identical protein patterns. The remaining isolates formed a weakly homogeneous group based on DNA-DNA similarity data and protein patterns; the results of unweighted pair group cluster analyses suggested that these isolates were members of five new species of the genus Thermococcus.