Cell Architecture of the Giant Sulfur Bacterium Achromatium oxaliferum: Extra-cytoplasmic Localization of Calcium Carbonate Bodies.

Achromatium oxaliferum is a large sulfur bacterium easily recognized by large intracellular calcium carbonate bodies. Although these bodies often fill major parts of the cells' volume, their role and specific intracellular location are unclear. In this study, we used various microscopy and staining techniques to identify the cell compartment harboring the calcium carbonate bodies. We observed that Achromatium cells often lost their calcium carbonate bodies, either naturally or induced by treatments with diluted acids, ethanol, sodium bicarbonate and UV radiation which did not visibly affect the overall shape and motility of the cells (except for UV radiation). The water-soluble fluorescent dye fluorescein easily diffused into empty cavities remaining after calcium carbonate loss. Membranes (stained with Nile Red) formed a network stretching throughout the cell and surrounding empty or filled calcium carbonate cavities. The cytoplasm (stained with FITC and SYBR Green for nucleic acids) appeared highly condensed and showed spots of dissolved Ca2+ (stained with Fura-2). From our observations, we conclude that the calcium carbonate bodies are located in the periplasm, in extra-cytoplasmic pockets of the cytoplasmic membrane and are thus kept separate from the cell's cytoplasm. This periplasmic localization of the carbonate bodies might explain their dynamic formation and release upon environmental changes.

Community-like genome in single cells of the sulfur bacterium Achromatium oxaliferum.

Polyploid bacteria are common, but the genetic and functional diversity resulting from polyploidy is unknown. Here we use single-cell genomics, metagenomics, single-cell amplicon sequencing, and fluorescence in situ hybridization, to show that individual cells of Achromatium oxaliferum, the world's biggest known freshwater bacterium, harbor genetic diversity typical of whole bacterial communities. The cells contain tens of transposable elements, which likely cause the unprecedented diversity that we observe in the sequence and synteny of genes. Given the high within-cell diversity of the usually conserved 16S ribosomal RNA gene, we suggest that gene conversion occurs in multiple, separated genomic hotspots. The ribosomal RNA distribution inside the cells hints to spatially differential gene expression. We also suggest that intracellular gene transfer may lead to extensive gene reshuffling and increased diversity.The cells of Achromatium bacteria are remarkably large and contain multiple chromosome copies. Here, Ionescu et al. show that chromosome copies within individual cells display high diversity, similar to that of bacterial communities, and contain tens of transposable elements.

[Selection of a community of acidochemolithotrophic microorganisms with a high oxidation rate of pyrrhotite- containing sulphide ore flotatation concentrate].

A community of acidochemolithotrophic microorganisms with a high oxidation rate of pyrrhotite-containing sulphide ore flotation concentrate was selected. The Acidithiobacillus caldus OP-1 and Ferroplasma acidiphilum OP-2 cultures were identified to be dominating members. The presence of the Acidithio- bacillusferrooxidans OP-3, Leptospirillumferriphilum OP-4, and Sulfobacillus thermosulfidooxidans OP-5 cultures in the community's composition was also mentioned. The analysis results of solid residues of the process showed a greater elemental sulfur oxidation level and gold recovery when the initial pH value in tank I was maintained at a level of 1.8-2.0 (90.5%) rather than 1.6-1.8 (86.3%).

Thermobacillus xylanilyticus gen. nov., sp. nov., a new aerobic thermophilic xylan-degrading bacterium isolated from farm soil.

An aerobic, thermophilic, xylanolytic, spore-forming bacterium, XETP (T = type strain; P = patent strain), has been isolated from farm soil situated underneath a manure heap in northern France. Strain XETP, which stained negative in the Gram test, occurs as short rods which sometimes form chains. Its spores are ellipsoidal, central to subterminal and occur in swollen sporangia. It grows at temperatures up to 63 degrees C and in the pH range 6.5-8.5. When grown on glucose in optimal conditions, its doubling time was found to be 33 min. CO2 was observed to have a growth-stimulating effect at the start of the culture. In addition to glucose, the isolate utilizes xylose, arabinose, mannose, cellobiose, galactose, maltose, sucrose, xylan and starch. Growth is inhibited by 5% NaCl. The G+C content of strain XETP is 57.5 mol%. The 16S rDNA sequence analysis indicated that strain XETP falls into the radiation of the Bacillus-Lactobacillus-Streptococcus subdivision of the Gram-positive phylum. Its three closest phylogenetic relatives are 'Bacillus viscosus', Paenibacillus curdlanolyticus and Bacillus popilliae with identity values of 91.15, 90.94 and 90.92%, respectively. The major cellular fatty acids are 14-methyl pentadecanoic acid (16:0 iso), hexadecanoic acid (16:0) and 14-methyl hexadecanoic acid (17:0 anteiso). On the basis of 16S rRNA sequence and chemotaxonomic characteristics, the isolate is different enough for it to be considered as a member of a new genus. It is therefore proposed that this isolate represents a new genus and species: Thermobacillus xylanilyticus. Strain XETP, the type strain of Thermobacillus xylanilyticus, has been deposited in the Collection Nationale de Cultures Microbiennes (CNCM I-1017) as a patent strain.

Leptospirillum gen. nov. (ex Markosyan 1972), nom. rev., including Leptospirillum ferrooxidans sp. nov. (ex Markosyan 1972), nom. rev. and Leptospirillum thermoferrooxidans sp. nov. (Golovacheva et al. 1992).

The name Leptospirillum ferrooxidans is not in the Approved Lists of Bacterial Names (1980), nor has it been subsequently validly published. In accordance with the rules of the International Code of Nomenclature of Bacteria, the name Leptospirillum for the genus (gen. nov., nom. rev.) and Leptospirillum ferrooxidans for the species (sp. nov., nom. rev.) is revived here. The type species is Leptospirillum ferrooxidans strain L15T (= DSM 2705T). The second species in the genus is Leptospirillum thermoferrooxidans (Golovacheva et al. 1992) (type strain L-88T; Institute of Microbiology, INMI, Moscow, Russia).

Idiomarina gen. nov., comprising novel indigenous deep-sea bacteria from the Pacific Ocean, including descriptions of two species, Idiomarina abyssalis sp. nov. and Idiomarina zobellii sp. nov.

Two bacterial strains, KMM 227T and 231T, were isolated from seawater samples collected from the north-western Pacific Ocean at a depth of 4000-5000 m and were characterized using polyphasic taxonomy. Both were Gram-negative, psychrotolerant, heterotrophic, aerobic and required NaCl for growth (0.6-15.0%). The temperature for growth was 4-30 degrees C. Both strains were rod-shaped, with a single flagellum. However, strain KMM 231T revealed a single long fimbrium. Cellular fatty acids detected in the isolates were predominantly odd-numbered and iso-branched, with 15 and 17 carbons (ca. 70%). Also present were saturated and monounsaturated straight-chain fatty acids. Results of phylogenetic analyses, employing three tree-making methods, strongly indicated that the two strains formed a distinct lineage within a clade containing the genera Alteromonas, Colwellia and Pseudoalteromonas, in the gamma-Proteobacteria. The two strains shared 16S rDNA sequence similarity of 96.9% and genomic DNA relatedness of 27%; the latter was determined by dot-blot hybridization. The strains were differentiated by the presence of fimbria, production of chitinase, ability to grow on 15% NaCl and BIOLOG profiles. Given the polyphasic evidence accumulated in this study, it is proposed that the two deep-sea isolates be classified in the genus Idiomarina gen. nov., as Idiomarina abyssalis sp. nov. (type strain is KMM 227T) and Idiomarina zobellii sp. nov. (type strain is KMM 231T).

Substrate uptake by uncultured bacteria from the genus Achromatium determined by microautoradiography.

Microautoradiography was used to investigate substrate uptake by natural communities of uncultured bacteria from the genus Achromatium. Studies of the uptake of (14)C-labelled substrates demonstrated that Achromatium cells from freshwater sediments were able to assimilate (14)C from bicarbonate, acetate, and protein hydrolysate; however, (14)C-labelled glucose was not assimilated. The pattern of substrate uptake by Achromatium spp. was therefore similar to those of a number of other freshwater and marine sulfur-oxidizing bacteria. Different patterns of radiolabelled bicarbonate uptake were noted for Achromatium communities from different geographical locations and indicated that one community (Rydal Water) possessed autotrophic potential, while the other (Hell Kettles) did not. Furthermore, the patterns of organic substrate uptake within a single population suggested that physiological diversity existed in natural communities of Achromatium. These observations are consistent with and may relate to the phylogenetic diversity observed in Achromatium communities. Incubation of Achromatium-bearing sediment cores from Rydal Water with (35)S-labelled sulfate in the presence and absence of sodium molybdate demonstrated that this bacterial population was capable of oxidizing sulfide to intracellular elemental sulfur. This finding supported the role of Achromatium in the oxidative component of a tightly coupled sulfur cycle in Rydal Water sediment. The oxidation of sulfide to sulfur and ultimately to sulfate by Achromatium cells from Rydal Water sediment is consistent with an ability to conserve energy from sulfide oxidation.

Description of Pseudaminobacter gen. nov. with two new species, Pseudaminobacter salicylatoxidans sp. nov. and Pseudaminobacter defluvii sp. nov.

An aerobic bacterium, strain BN12T, which degrades substituted naphthalenesulfonates and substituted salicylates was isolated from a 6-aminonaphthalene-2-sulfonate-degrading microbial consortium originating from the River Elbe, Germany. Chemotaxonomic investigations of quinones, polyamines and polar lipids allowed allocation of this strain to the alpha-subclass of the Proteobacteria and revealed similarity to species of the genera Aminobacter, Chelatobacter and Mesorhizobium. This was confirmed by typing with 16S rRNA-targeted oligonucleotide probes and 16S rDNA sequencing and phylogenetic analysis, indicating that BN12T clusters most closely with a strain 'Thiobacillus' THI 051T and with the above genera but comprising a separate branch. DNA-DNA hybridizations demonstrated that strain BN12T is different from all species of Aminobacter currently described and recognized. The fatty acid patterns, substrate utilization profile and biochemical characteristics displayed no obvious similarity to the characteristics of Aminobacter and Chelatobacter species. 'Thiobacillus' THI 051T, however, revealed phenotypic similarities to BN12T. Furthermore, 16S rRNA sequences of Chelatobacter heintzii showed a high similarity to the 16S rRNA sequences of all currently recognized Aminobacter species. On the basis of these and previously published results, the new genus Pseudaminobacter is proposed, harbouring the two new species Pseudaminobacter salicylatoxidans sp. nov. and Pseudaminobacter defluvii sp. nov. The type strains are BN12T (= DSM 6986T) and THI 051T (= IFO 14570T), respectively.

Ultrastructure and chemical composition of the sheath of Leptothrix discophora SP-6.

Light microscopy and transmission electron microscopy of thin sections and metal-shadowed specimens showed that the sheath of Leptothrix discophora SP-6 (ATCC 51168) is a tube-like extracellular polymeric structure consisting of a condensed fabric of 6.5-nm-diameter fibrils underlying a more diffuse outer capsular layer. In thin sections, outer membrane bridges seen to contact the inner sheath layer suggested that the sheath fabric was attached to the outer layer of the gram-negative cell wall. The capsular polymers showed an affinity for cationic colloidal iron and polycationic ferritin, indicating that they carry a negative charge. Cell-free sheaths were isolated by treatment with a mixture of lysozyme, EDTA, and N-lauroylsarcosine (Sarkosyl) or sodium dodecyl sulfate (SDS). Both Sarkosyl- and SDS-isolated sheaths were indistinguishable in microscopic appearance. However, the Mn-oxidizing activity of Sarkosyl-isolated sheaths was more stable than that of SDS-isolated sheaths. The Sarkosyl-isolated sheaths also contained more 2-keto-3-deoxyoctanoic acid and more outer membrane protein than SDS-isolated sheaths. The oven-dried mass of detergent-isolated sheaths represented approximately 9% of the total oven-dried biomass of SP-6 cultures; the oven-dried sheaths contained 38% C, 6.9% N, 6% H, and 2.1% S and approximately 34 to 35% carbohydrate (polysaccharide), 23 to 25% protein, 8% lipid, and 4% inorganic ash. Gas-liquid chromatography showed that the polysaccharide was an approximately 1:1 mixture of uronic acids (glucuronic, galacturonic, and mannuronic acids and at least one other unidentified uronic acid) and an amino sugar (galactosamine). Neutral sugars were not detected. Amino acid analysis showed that sheath proteins were enriched in cysteine (6 mol%). The cysteine residues in the sheath proteins probably provide sulfhydryls for disulfide bonds that play an important role in maintaining the structural integrity of the sheath (D. Emerson and W.C. Ghiorse, J. Bacteriol. 175:7819-7827, 1993).

Marinobacter hydrocarbonoclasticus gen. nov., sp. nov., a new, extremely halotolerant, hydrocarbon-degrading marine bacterium.

On the basis of phenotypical characteristics and analysis of 16S rRNA sequence, a new species belonging to a new genus is described, and the name Marinobacter hydrocarbonoclasticus is proposed. This organism, isolated from Mediterranean seawater near a petroleum refinery, is a gram-negative, aerobic, rod-shaped bacterium. It grows at NaCl concentrations of 0.08 to 3.5 M and uses various hydrocarbons as the sole source of carbon and energy. Its DNA has a guanine-plus-cytosine content of 52.7 mol%. The 16S rRNA analysis shows a clear affiliation between M. hydrocarbonoclasticus and the gamma group of the phylum Proteobacteria. A close phylogenetic relationship appears among the species Marinomonas vaga, Oceanospirillum linum, Halomonas elongata, and Pseudomonas aeruginosa. Because of the impossibility of finding a single most closely related species, we suggest that this bacterium be assigned to a new genus, at least temporarily. The possibility of a revision of this status when new data appear is, however, not excluded. The type strain is M. hydrocarbonoclasticus SP.17 (= ATCC 49840).

Flocculation in Azospirillum brasilense and Azospirillum lipoferum: exopolysaccharides and cyst formation.

The phenomena of flocculation and floc formation by Azospirillum brasilense Sp7 (ATCC 29145) and Azospirillum lipoferum Sp59b (ATCC 29707) were studied in aerobic liquid cultures. Carbon sources representative of various entry pathways in combination with various nitrogen sources induced flocculation in both species of azospirilla. Noticeably, the combination of fructose and nitrate was the most effective in terms of floc yields. Phase-contrast microscopic observations revealed a transition in cell morphology from freely motile, vibrioid cells to nonmotile, highly refractile encysting forms during the formation of flocs. The nonmotile forms in flocs appeared to be entangled within a fibrillar matrix, and the cells were highly resistant to desiccation. Dried flocs kept for almost 6 months still maintained the highly refractile encysting forms, and their viability was confirmed by pellicle formation and acetylene reduction in semisolid malate medium. Electron microscopic observations of the desiccated flocs revealed the presence of cell forms containing abundant poly beta-hydroxybutyrate granules within a central body and surrounded by a thick layer of exopolysaccharides. The latter were characterized by alkali and acid digestion, crude cellulase hydrolysis, and calcofluor staining. It was concluded that the overproduction of exocellular polymers induces the flocculent growth and is associated with the concomitant transformation of vegetative cells to the desiccation-resistant encysting forms under limiting cultural conditions.

Mechanism of adhesion of Alysiella bovis to glass surfaces.

Alysiella bovis adheres to surfaces by means of short, ruthenium red-staining, rod-like fimbriae. The fimbriae remain associated with the cell envelope of A. bovis, even when sonicated or exposed sequentially to toluene, Triton X-100, lysozyme, ribonuclease, and deoxyribonuclease. Adhesion of outer membrane-derived cell wall ghosts of A. bovis to glass was inhibited by IO4-, sodium dodecyl sulfate, urea, pronase, and trypsin. Protease treatment digested the fimbriae from the distal end, and exposure to sodium dodecyl sulfate depolymerized the fimbriae. Exposure of ghosts to 1% sodium dodecyl sulfate preferentially solubilized a 16,500-dalton protein which was subsequently purified by gel filtration and demonstrated to be a glycoprotein (ca. 17% carbohydrate). Antibodies raised against the 16,500-dalton glycoprotein agglutinated whole cells and inhibited adhesion of ghosts to glass.

Dorsal-ventral differentiation in Simonsiella and other aspects of its morphology and ultrastructure.

The morphology and ultrastructure of the aerobic, Gram-negative multicellular-filamentous bacteria of the genus simonsiella were investigated by scanning and transmission electron microscopy. The flat, ribbon-shaped, multicellular filaments show dorsal-ventral differentiation with respect to their orientations to solid substrata. The dorsal surface, orientated away from the substrate, is convex and possesses an unstructured capsule. The ventral surface, on which the organisms adhere and glide, is concave and has an extracellular layer with fibrils extending at right angles from the cell wall. The cytoplasm in the ventral region contains a proliferation of intracytoplasmic membranes and few ribosomes in comparison to the cytoplasm in other parts of the cell. Centripetal cell wall formation is asymmetrical and commences preferentially in the ventral region. Quantitative differences in morphology and cytology exist among selected Simonsiella strains. Functional aspects of this dorsal-ventral differentiation are discussed with respect to the colonization and adherence of Simonsiella to mucosal squamous epithelial cells in its ecological habitat, the oral cavities of warm-blooded vertebrates.

[Thermophilic and thermotolerant bacteria that assimilate methane]. / Termofil'nye i termotolerantnye bakterii, assimiliruiushchie metan

Microorganisms assimilating methane at temperatures above 40 degrees C were isolated from various natural sources: ooze, mud, waste water of coal pits. The bacteria are obligate methylotrophs and are represented by two groups: (a) thermotolerant, growing at 37 to 45 degrees C; and (b) thermophilic, growing at 50 to 62 degrees C. The selective factor used to isolate various physiological forms of methylotrophs is corresponding temperatures of growth which allow to isolate from the same substrate meso-, thermotolerant, and thermophilic forms. Morphological and physiological properties of the strains are described. The thermotolerant cultures of methylotrophs are similar to Methylobacter vinelandii, though differ from it by some characteristics. The thermophilic microorganisms should be classed as a separate species Methylococcus thermophilus.

[Thermus ruber obligate thermophilic bacteria in the thermal springs of Kamchatka]. / Obligatno-termofil'nye bakterii Thermus ruber v gidrotermakh Kamchatki

The new species of the obligate-thermophilic bacterium Thermus ruber is widely distributed in hot springs of Kamchatka with the temperature of water from 57 to 90 degrees C. The bacterium does not oxidize sulphur. It contains a red carotenoid pigment similar to neuro-sporaxanthine and retrodehydro-gamma-carotene.