Anna-Louise Reysenbach

Thermoterrabacterium ferrireducens gen. nov., sp. nov., a thermophilic anaerobic dissimilatory Fe(III)-reducing bacterium from a continental hot spring.

A strain of a thermophilic, anaerobic, dissimilatory, Fe(III)-reducing bacterium, Thermoterrabacterium ferrireducens gen. nov., sp. nov. (type strain JW/AS-Y7T; DSM 11255), was isolated from hot springs in Yellowstone National Park and New Zealand. The gram-positive-staining cells occurred singly or in pairs as straight to slightly curved rods, 0.3 to 0.4 by 1.6 to 2.7 microns, with rounded ends and exhibited a tumbling motility. Spores were not observed. The temperature range for growth was 50 to 74 degrees C with an optimum at 65 degrees C. The pH range for growth at 65 degrees C was from 5.5 to 7.6, with an optimum at 6.0 to 6.2. The organism coupled the oxidation of glycerol to reduction of amorphous Fe(III) oxide or Fe(III) citrate as an electron acceptor. In the presence as well as in the absence of Fe(III) and in the presence of CO2, glycerol was metabolized by incomplete oxidation to acetate as the only organic metabolic product; no H2 was produced during growth. The organism utilized glycerol, lactate, 1,2-propanediol, glycerate, pyruvate, glucose, fructose, mannose, and yeast extract as substrates. In the presence of Fe(III) the bacterium utilized molecular hydrogen. The organism reduced 9,10-anthraquinone-2,6-disulfonic acid, fumarate (to succinate), and thiosulfate (to elemental sulfur) but did not reduce MnO2, nitrate, sulfate, sulfite, or elemental sulfur. The G + C content of the DNA was 41 mol% (as determined by high-performance liquid chromatography). The 16S ribosomal DNA sequence analysis placed the isolated strain as a member of a new genus within the gram-type-positive Bacillus-Clostridium subphylum.

Evidence for the microbial basis of a chemoautotrophic invertebrate community at a whale fall on the deep seafloor: Bone-colonizing bacteria and invertebrate endosymbionts

To explore the microbial basis for a remarkable macrofaunal community at the site of a whale skeleton on the seafloor of the Santa Catalina Basin, we obtained samples of whale bone, bone‐colonizing invertebrates, microbial mats, and the dominant fauna in the adjacent sulfide‐rich sediments during Alvin expeditions in 1988 and 1991. Invertebrate tissues were examined by transmission electron microscopy (TEM) and mats and bone‐penetrating bacteria by epifluorescence microscopy (EM). Tissues from the dominant bivalve Vesicomya c.f. gigas, the mytilid mussel Idasola washingtonia, and selected gastropods and limpets were also assayed chemically for enzymes diagnostic of sulfur‐ and methane‐based chemoautotrophy and for stable carbon isotopic composition. Results of all analyses were consistent with dominant sulfur‐based endosymbioses in the clam and mussel (the first record of endosymbiosis in the genus Idasola) and the general absence of methane symbioses at the site, strengthening the analogy of the whale‐skeleton faunal community to those known from distant Pacific hydrothermal vent sites. Examples of minor endosymbionts, either nitrifying or methanotrophic cells according to internal membrane structures by TEM, raised the possibility of a supplemental mode of nutrition to the clam, or means to remove ammonia in the gill tissue, in the event of significant changes in the chemical environment. Microsc. Res. Tech. 37:162–170, 1997.

Isolation and characterization of the homoacetogenic thermophilic bacterium Moorella glycerini sp. nov.

A thermophilic, anaerobic, spore-forming bacterium (strain JW/AS-Y6T) was isolated from a mixed sediment-water sample from a hot spring (Calcite Spring area) at Yellowstone National Park. The vegetative cells of this organism were straight rods, 0.4 to 0.6 by 3.0 to 6.5 microns. Cells occurred singly and exhibited a slight tumbling motility. They formed round refractile endospores in terminal swollen sporangia. Cells stained gram positive. The temperature range for growth at pH 6.8 was 43 to 65 degrees C, with optimum growth at 58 degrees C. The range for growth at 60 degrees C (pH60C; with the pH meter calibrated at 60 degrees C) was 5.9 to 7.8, with an optimum pH60C of 6.3 to 6.5. The substrates utilized included glycerol, glucose, fructose, mannose, galactose, xylose, lactate, glycerate, pyruvate, and yeast extract. In the presence of CO2, acetate was the only organic product from glycerol and carbohydrate fermentation. No H2 was produced during growth. The strain was not able to grow chemolithotrophically at the expense of H2-CO2; however, suspensions of cells in the exponential growth phase consumed H2. The bacterium reduced fumarate to succinate and thiosulfate to elemental sulfur. Growth was inhibited by ampicillin, chloramphenicol, erythromycin, rifampin, and tetracycline, but not by streptomycin. The G+C content of the DNA was 54.5 mol% (as determined by high-performance liquid chromatography). The 16S ribosomal DNA sequence analysis placed the isolate in the Gram type-positive Bacillus-Clostridium subphylum. On the basis of physiological properties and phylogenetic analysis we propose that the isolated strain constitutes a new species, Moorella glycerini; the type strain is JW/AS-Y6 (= DSM 11254 = ATCC 700316).

Phylum All. Euryarchaeota phy. nov.

The phylum currently consists of seven classes: the Methanobacteria, the Methanococci, the Halobacteria, the Thermoplasmata, the Thermococci, the Archaeoglobi, and the Methanopyri. With the sole exception of the Methanococci, which is subdivided into three orders, each class contains a single order. The Euryarchaeota are morphologically diverse and occur as rods, cocci, irregular cocci, lancet-shaped, spiral-shaped, disk-shaped, triangular, or square cells. Cells stain Gram-positive or Gram-negative based on the presence or absence of pseudomurein in cell walls. In some classes, cell walls consist entirely of protein or may be completely absent (Thermoplasmata). Five major physiological groups have been described previously: the methanogenic Archaea, the extremely halophilic Archaea, Archaea lacking a cell wall, sulfate reducing Archaea, and the extremely thermophilic S0 metabolizers.

Isolation and characterization of Thermococcus barossii, sp. nov., a hyperthermophilic archaeon isolated from a hydrothermal vent flange formation.

A new hyperthermophilic microorganism, Thermococcus barossii, was isolated from rock fragments of a hydrothermal vent flange formation, located along the East Pacific Rise of the Juan de Fuca Ridge. This organism is obligately anaerobic and grows over a temperature range of at least 60-92 degrees C in artificial seawater-based media, containing elemental sulfur, tryptone and yeast extract. The addition of a maltooligosaccharide mixture and tungsten to this medium improved growth to some extent. At the Topt for growth (82.5 degrees C), cell densities as high as 4 x 10(8) cells/ml could be obtained in 18-liter batch fermentations, with a doubling time of approximately 40 minutes, if culture access to elemental sulfur was sufficient. In continuous culture at the same temperature, comparable cell densities could be obtained but only at slower growth rates. Morphologically, T. barossii is coccoid-shaped, forming irregularly-shaped spheres; under optimal conditions, these coccoids become more regular and smaller, a characteristic of other hyperthermophilic archaea. Negatively-stained preparations showed no pili or flagella associated with the cell surface. 16S rRNA sequencing reveals that T. barossii is most similar to Thermococcus celer (99.7%). Yet, further comparisons with T. celer showed that T. barossii is a new Thermococcus species: different growth temperature optimum (82.5 degrees C vs. 88 degrees C), obligate requirement for sulfur, higher G + C content (60% vs. 56.7%) and 47.7% DNA-DNA hybridization. The nucleotide and translated amino acid sequence for the gene encoding a DNA polymerase from T. barossii was compared to sequences of related genes from other Thermacoccales. The polymerase phylogenies were congruent with those obtained from the 16S rRNA phylogenetic analyses. Based on the high degree of similarity among members of the genus Termococcus for the criteria used thus far, aspects of enzymology may be an important mechanism of differenting one species from another.

Phylum BII. Thermotogae phy. nov.

Extremely thermophilic rod-shaped bacteria, non-sporeforming; Gram negative with an outer sheath-like envelope of “toga”. The members are all anaerobic and fermentative. The phylum is represented by a single class and a single order.

Phylum Al. Crenarchaeota phy. nov.

The phylum consists of a single class, the Thermoprotei, which is well supported by 16S rDNA sequence data. It is subdivided into three orders: the Thermoproteales, the Desulfurococcales, and the Sulfolobales. Morphologically diverse, including rods, cocci, filamentous forms, and disk-shaped cells. Stain Gram-negative. Motility observed in some genera. Obligately thermophilic, with growth occurring at temperatures ranging from 70 to 113°C. Acidophilic. Aerobic, facultatively anaerobic, or strictly anaerobic-chemolithoautotrophs or chemoheterotrophs. Most metabolize S0. Chemoheterotrophs, may grow by sulfur respiration. RNA polymerase of the BAC type.

Community Structure along a Thermal Gradient in a Stream Near Obsidian Pool, Yellowstone National Park

The study of the diversity of life on earth has traditionally been one of the cornerstones of biological science. Examination of variations among plant and animal species became the foundation for Charles Darwin’s theory of evolution by natural selection. More recently, ecological studies have demonstrated that community diversity is an important determinant in ecosystem processes and stability (Tilman et al., 1994; Tilman et al., 1997). The study of the evolution and diversity of microorganisms, however, has lagged behind that of other life forms, largely because of the difficulties inherent in assessing microbial diversity in nature. Microorganisms lack the complex morphologies used to classify higher organisms, and shared physiological traits have also proven unreliable in inferring evolutionary relationships (Fox et al., 1980). Traditional culture techniques often lead to underestimation of diversity by selecting for those organisms best suited to the growth media (Brock, 1987).