The organism described in this paper, strain ST90T (T = type strain), is a thermophilic, spore-forming, rod-shaped sulfate reducer that was isolated from North Sea oil reservoir formation water. In cultivation the following substances were used as electron donors and carbon sources: H,-CO,, lactate, pyruvate, ethanol, propanol, butanol, and C, to C,, and C,, to C,, carboxylic acids. Sulfate was used as the electron acceptor in these reactions. Lactate was incompletely oxidized. Sulfite and thiosulfate were also used as electron acceptors. In the absence of an electron acceptor, the organism grew syntrophically on propionate together with a hydrogenothrophic methanogen. The optimum conditions for growth on Iactate and sulfate were 62"C, pH 6.7, and 50 to 200 mM NaCl. The G+C content was 56 mol%, as determined by high-performance liquid chromatography and 57 mol% as determined by thermal denaturation. Spore formation was observed when the organism was grown on butyrate or propanol as a substrate and at low pH values. On the basis of differences in G+C content and phenotypic and immunological characteristics when the organism was compared with other thermophilic Desulfotomaculum species, we propose that strain ST90T is a member of a new species, Desulfotomaculum thermocisternum. D. thermocisternum can be quickly identified and distinguished from closely related Desulfotomaculum species by immunoblotting.The genus Desulfotomaculum comprises a heterogeneous group of gram-positive, spore-forming sulfate reducers that includes both mesophilic and thermophilic species. The following seven thermophilic species have been validly described previously: Desulfotomaculum nigYiJicans, which was isolated from canned food (52) and produced oil field water (1, 30); Desulfotomaculum geothermicum, Desulfotomaculum australicum, and Desulfotomaculum kuznetsovii, which were isolated from geothermal groundwater (13, 26, 29) (D. kuznetsovii has also been isolated from cold marine sediment [ 181); Desulfotomaculum thermoacetoxidans and Desulfotomaculum thermobenzoicum, which were isolated from thermophilic fermentation reactors (28, 48); and Desulfotomaculum thermosapovorans, which was isolated from compost (16). Workers have also described several thermophilic Desulfotomaculum strains whose phylogenetic positions within the genus have not been determined (20,39,47).North Sea oil field reservoirs are hot marine habitats that are 1.2 to 6 km below the seafloor; the pressures in these habitats range from 50 to 80 MPa, and the temperatures range from 60 to 200°C. The concentration of sulfate is usually between 0 and 0.6 mM and varies from one reservoir to another. Aliphatic carboxylic acids are the most abundant organic acids in petroleum reservoirs. In North Sea formation water, acetic acid is found at concentrations up to 20 mM, with decreasing concentrations of higher homologs up to octanoic acid (2, 5). These acids are potential electron donors for sulfate reduction. Formic acid is usually not detected (24). During offshore oil producti...
A hyperthermophilic sulfate reducer, strain 7324, was isolated from hot (75°C) oil field waters from an oil production platform in the Norwegian sector of the North Sea. It was enriched on a complex medium and isolated on lactate with sulfate. The cells were nonmotile, irregular coccoid to disc shaped, and 0.3 to 1.0 ,um wide. The temperature for growth was between 60 and 85°C with an optimum of 76°C. Lactate, pyruvate, and valerate plus H2 were utilized as carbon and energy sources with sulfate as electron acceptor. Lactate was completely oxidized to CO2. The cells contained an active carbon mopoxide dehydrogenase but no 2-oxoglutarate dehydrogenase activity, indicating that lactate was oxidized to CO2 via the acetyl coenzyme A/carbon monoxide dehydrogenase pathway. The cells produced small amounts of methane simultaneously with sulfate reduction. F420 was detected in the cells which showed a blue-green fluorescence at 420 nm. On the basis of morphological, physiological, and serological features, the isolate was classified as an Archaeoglobus sp. Strain 7324 showed 100%/ DNA-DNA homology with A. fulgidus Z, indicating that it belongs to the species A. fulgidus.
The distribution of thermophilic marine sulfate reducers in produced oil reservoir waters from the Gullfaks oil field in the Norwegian sector of the North Sea was investigated by using enrichment cultures and genusspecific fluorescent antibodies produced against the genera Archaeoglobus, Desulfotomaculum, and Thermodesulforhabdus. The thermophilic marine sulfate reducers in this environment could mainly be classified as species belonging to the genera Archaeoglobus and Thermodesulforhabdus. In addition, some unidentified sulfate reducers were present. Culturable thermophilic Desulfotomaculum strains were not detected. Specific strains of thermophilic sulfate reducers inhabited different parts of the oil reservoir. No correlation between the duration of seawater injection and the numbers of thermophilic sulfate reducers in the produced waters was observed. Neither was there any correlation between the concentration of hydrogen sulfide and the numbers of thermophilic sulfate reducers. The results indicate that thermophilic and hyperthermophilic sulfate reducers are indigenous to North Sea oil field reservoirs and that they belong to a deep subterranean biosphere.
Methanococcus thermolithotrophicus ST22 was isolated from produced water of a North Sea oil field, on mineral medium with H 2-CO 2 as the sole source of carbon and energy. The isolate grew at 17 to 62؇C, with an optimum at 60؇C. The pH range was 4.9 to 9.8, with optimal growth at pH 5.1 to 5.9; these characteristics reflected its habitat. Strain ST22 was quickly identified and distinguished from the type strain by immunoblotting.
An infectious pancreatic necrosis virus (IPNV) belonging to the Sp serotype, isolated from scallops Pecten maximus, was propagated and suspended in sterile water and cell culture medium with different salinities and incubated at temperatures ranging from -80 to 40°C. Virus stability was examined by measunng vlrus titers under dfferent storage conditions. Virus titers were also measured after repeated freezing and thawing, and in incubated sterile filtered scallop hepatopancreas, haemolymph and crystalline style samples, and salmon Salmo salar kidney homogenate. The virus was stable under most storage conditions. Temperatures rangng from -80 to +20°C. as well as salinities from 0 to 40%0. did not seem to influence the stability of the virus. A reduction was observed above 20°C. Each freezing and thawing procedure resulted in a reduction of the virus titer. This reduction was larger at -80 than at -20°C. The IPNV persisted for a long period in sterile filtered scallop haemolymph, dissolved crystalline style and hepatopancreas. In ludney homogenate from IPNV-infected salmon the virus titers were reduced at least 10-fold during the first day of incubation at all temperatures tested. When virus was propagated in cell culture and subsequently mixed with non-infected salmon kidney homogenate, the virus proved more persistent. Our results illustrate the importance of rapid and standardised laboratory processing of potentially vlrus-containing tissue samples, and are relevant when considering laboratory storage of samples containing IPNV.
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