ISOLATION AND CHARACTERIZATION OF EXTREMELY THERMOPHILIC ARCHAEBACTERIA RELATED TO THE GENUS THERMOCOCCUS FROM DEEP-SEA HYDROTHERMAL GUAYMAS BASIN

During the ''Guaynaut'' oceanographic cruise performed by IFREMER in November 1991, sediment cores were collected from high-temperature and petroleum-rich deposits in an active hydrothermal zone, at the Guaymas basin (Central gulf of California). Those samples were collected by the French deep-sea manned submersible ''Nautile'' at a depth of 2000 meters. Four sediment cores of 20-40-cm length were drilled at the bottom of a block assemblage of active smokers inside sediments whose temperatures were 3.5 degrees C on the top to 105 degrees C at 20 cm depth. They were subsampled in 22 slices of 5-cm thickness and used for isolation of heterotrophic hyperthermophilic microorganisms, after inoculation in sulfur-free SME liquid medium. From those enrichments 18 isolates were obtained, 2 growing at 95 degrees C and 16 at 80 degrees C, and their taxonomic characterization was undertaken. Lipid analysis indicated the presence of diethers and tetraethers in the cell walls and membranes, characteristic of Archaebacteria. Examinations by scanning electron microscopy showed that isolates were cocci of heterogeneous sizes (diameter from 0.1 to 0.5 micrometers) or thick, piled-up discs 0.5 mu m thick and 1 mu m in diameter. Both forms were embedded in a dense fiber network. Physiologically they were found to be anaerobic, heterotrophic, and hyperthermophilic (80 degrees-95 degrees C). Determination of the DNA base composition resulted in G + C mol % values ranging from 36 to 57. Qualitative hybridizations of the 18 isolate DNAs with hyperthermophilic Archaebacteria reference strain DNAs showed that hybridizations occurred neither with the two species of Pyrococcus nor with the two species of Desulfurococcus, nor with Staphylothermus marinus. On the other hand, all isolates hybridized with at least one of the three species of Thermococcus tested (T. celer, T. stetteri, T. litoralis). Restriction polymorphism on a PCR-amplified fragment of the rrn operon showed that 12 isolates had the same profile as T. celer and T. stetteri, 4 isolates had the same profile as T. litoralis, and 2 had new profiles, suggestive that they are new species.