Formation and function of bacterial organelles.

Advances in imaging technologies have revealed that many bacteria possess organelles with a proteomically defined lumen and a macromolecular boundary. Some are bound by a lipid bilayer (such as thylakoids, magnetosomes and anammoxosomes), whereas others are defined by a lipid monolayer (such as lipid bodies), a proteinaceous coat (such as carboxysomes) or have a phase-defined boundary (such as nucleolus-like compartments). These diverse organelles have various metabolic and physiological functions, facilitating adaptation to different environments and driving the evolution of cellular complexity. This Review highlights that, despite the diversity of reported organelles, some unifying concepts underlie their formation, structure and function. Bacteria have fundamental mechanisms of organelle formation, through which conserved processes can form distinct organelles in different species depending on the proteins recruited to the luminal space and the boundary of the organelle. These complex subcellular compartments provide evolutionary advantages as well as enabling metabolic specialization, biogeochemical processes and biotechnological advances. Growing evidence suggests that the presence of organelles is the rule, rather than the exception, in bacterial cells.

[Desulfovibrio hontreensis sp. nov., a Sulfate-Reducing Bacterium Isolated from Marine Biofoulings at the South Vietnam Coastal Area].

A Desulfovibrio strain physiologically similar to and phylogeneticall related to "D. caledoniensis" SEBR 7250, D. portus MSL79, and D. dechloracetivorans ATCC 700912 (96.9, 95.9, and 95.8% similarity of the 16S rRNA gen sequences, respectively) was isolated from marine biofouling in the coastal zone of the South China Sae (Nha Trang, South Vietnam). The cells of strain ME were gram-negative motile vibrios (0.4-0.6 x 1.3-2 µm) with a single flagellum. The strain grew at 20 to 39 degrees C (growth optimum at 34-37 degrees C), pH 5.8 to 8.5 (pH optimum at 6.8-7.5), and salinity from 0.08 to 1.1 M Na+ (optimum at 0.2-0.3 M Na+). In the presence of sulfate, the strain grew autotrophically with hydrogen or on lactate, formate, pyruvate, fumarate, and malate. Weak growth occurred on succinate, glycerol, and fructose. In the absence of sulfate, the strain was able to ferment pyruvate, malate (weakly), but not lactate. Sulfate, sulfite, thiosulfate, elemental sulfur, and dimethyl sulfoxide were used as electron acceptors. Vitamins and yeast extract were not required for growth. The G+C content was 52.4 mol %. Predominant fatty acids were C18:0 (13.9%), C16:0 (9.6%), iso-C16:0 (9.5%), C18: 1w7 (8.8%), anteiso-C15:0 (8.1%), and iso-C 17:1 (7.2%). The fatty acid composition was close to that of D. dechloracetivorans BO and has some similarity to that of D. portus. Based on its genotypic and phenotypic characteristics, strain ME maybe considered as a new species, for which the name Desulfovibrio hontrensis sp. nov. is proposed.

'Candidatus Ancillula trichonymphae', a novel lineage of endosymbiotic Actinobacteria in termite gut flagellates of the genus Trichonympha.

Termite gut flagellates are colonized by host-specific lineages of ectosymbiotic and endosymbiotic bacteria. Previous studies have shown that flagellates of the genus Trichonympha may harbour more than one type of symbiont. Using a comprehensive approach that combined cloning of SSU rRNA genes with fluorescence in situ hybridization and electron microscopy, we investigated the phylogeny and subcellular locations of the symbionts in a variety of Trichonympha species from different termites. The flagellates in Trichonympha Cluster I were the only species associated with 'Endomicrobia', which were located in the posterior part of the cell, confirming previous results. Trichonympha species of Cluster II from the termite genus Incisitermes (family Kalotermitidae) lacked 'Endomicrobia' and were associated with endosymbiotic Actinobacteria, which is highly unusual. The endosymbionts, for which we suggest the name 'Candidatus Ancillula trichonymphae', represent a novel, deep-branching lineage in the Micrococcineae that consists exclusively of clones from termite guts. They preferentially colonized the anterior part of the flagellate host and were highly abundant in all species of Trichonympha Cluster II except Trichonympha globulosa. Here, they were outnumbered by a Desulfovibrio species associated with the cytoplasmic lamellae at the anterior cell pole. Such symbionts are present in both Trichonympha clusters, but not in all species. Unlike the intracellular location reported for the Desulfovibrio symbionts of Trichonympha agilis (Cluster I), the Desulfovibrio symbionts of T. globulosa (Cluster II) were situated in deep invaginations of the plasma membrane that were clearly connected to the exterior of the host cell.

Desulfovibrio magneticus RS-1 contains an iron- and phosphorus-rich organelle distinct from its bullet-shaped magnetosomes.

Intracellular magnetite crystal formation by magnetotactic bacteria has emerged as a powerful model for investigating the cellular and molecular mechanisms of biomineralization, a process common to all branches of life. Although magnetotactic bacteria are phylogenetically diverse and their crystals morphologically diverse, studies to date have focused on a few, closely related species with similar crystal habits. Here, we investigate the process of magnetite biomineralization in Desulfovibrio magneticus sp. RS-1, the only reported species of cultured magnetotactic bacteria that is outside of the alpha-Proteobacteria and that forms bullet-shaped crystals. Using a variety of high-resolution imaging and analytical tools, we show that RS-1 cells form amorphous, noncrystalline granules containing iron and phosphorus before forming magnetite crystals. Using NanoSIMS (dynamic secondary ion mass spectroscopy), we show that the iron-phosphorus granules and the magnetite crystals are likely formed through separate cellular processes. Analysis of the cellular ultrastructure of RS-1 using cryo-ultramicrotomy, cryo-electron tomography, and tomography of ultrathin sections reveals that the magnetite crystals are not surrounded by membranes but that the iron-phosphorus granules are surrounded by membranous compartments. The varied cellular paths for the formation of these two minerals lead us to suggest that the iron-phosphorus granules constitute a distinct bacterial organelle.

Desulfovibrio brasiliensis sp. nov., a moderate halophilic sulfate-reducing bacterium from Lagoa Vermelha (Brazil) mediating dolomite formation.

A novel halotolerant sulfate-reducing bacterium, Desulfovibrio brasiliensis strain LVform1, was isolated from sediments of a dolomite-forming hypersaline coastal lagoon, Lagoa Vermelha, in the state of Rio de Janeiro, Brazil. The cells are vibrio-shaped and 0.30 to 0.45 microm by 1.0 to 3.5 microm in size. These bacteria mediate the precipitation of dolomite [CaMg(CO3)2] in culture experiments. The strain was identified as a member of the genus Desulfovibrio in the delta-subclass of the Proteobacteria on the basis of its 16S rRNA gene sequence, its physiological and morphological properties. Strain LVform1 is obligate sodium-dependent and grows at NaCl concentrations of up to 15%. The 16S rRNA sequence revealed that this strain is closely related to Desulfovibrio halophilus (96.2% similarity) and to Desulfovibrio oxyclinae (96.8% similarity), which were both isolated from Solar Lake, a hypersaline coastal lake in the Sinai, Egypt. Strain LVform1 is barotolerant, growing under pressures of up to 370 bar (37 MPa). We propose strain LVform1 to be the type strain of a novel species of the genus Desulfovibrio, Desulfovibrio brasiliensis (type strain LVform1 = DSMZ No. 15816 and JCM No. 12178). The GenBank/EMBL accession number for the 16S rDNA sequence of strain LVform1 is AJ544687.

Impact of clay minerals on sulfate-reducing activity in aquifers.

Previous studies have shown that sulfate-reduction activity occurs in a heterogeneous manner throughout the terrestrial subsurface. Low-activity regions are often observed in the presence of clay minerals. Here we report that clays inhibit sulfate reduction activity in sediments and in a pure culture of Desulfovibrio vulgaris. Clay minerals including bentonite and kaolinite inhibited sulfate reduction by 70-90% in sediments. Intact clays and clay colloids or soluble components, capable of passing through a 0.2-microm filter, were also inhibitory to sulfate-reducing bacteria. Other adsorbent materials, including anion or cation exchangers and a zeolite, did not inhibit sulfate reduction in sediments, suggesting that the effect of clays was not due to their cation-exchange capacity. We observed a strong correlation between the Al2O3 content of clays and their relative ability to inhibit sulfate reduction in sediments (r2 = 0.82). This suggested that inhibition might be a direct effect of Al3+ (aq) on the bacteria. We then tested pure aluminum oxide (Al2O3) and showed it to act in a similar manner to clay. As dissolved aluminum is known to be toxic to a variety of organisms at low concentrations, our results suggest that the effects of clay on sulfate-reducing bacteria may be directly due to aluminum. Thus, our experiments provide an explanation for the lack of sulfate-reduction activity in clay-rich regions and presents a mechanism for the effect.

Toxicity of Al to Desulfovibrio desulfuricans.

The toxicity of Al to Desulfovibrio desulfuricans G20 was assessed over a period of 8 weeks in a modified lactate C medium buffered at four initial pHs (5.0, 6.5, 7.2, and 8.3) and treated with five levels of added Al (0, 0.01, 0.1, 1.0, and 10 mM). At pH 5, cell population densities decreased significantly and any effect of Al was negligible compared to that of the pH. At pHs 6.5 and 7.2, the cell population densities increased by 30-fold during the first few days and then remained stable for soluble-Al concentrations of <5 x 10(-5) M. In treatments having total-Al concentrations of > or =1 mM, soluble-Al concentrations exceeded 5 x 10(-5) M and limited cell population growth substantially and proportionally. At pH 8.3, soluble-Al concentrations were below the 5 x 10(-5) M toxicity threshold and cell population density increases of 20- to 40-fold were observed. An apparent cell population response to added Al at pH 8.3 was attributed to the presence of large, spirilloidal bacteria (accounting for as much as 80% of the cells at the 10 mM added Al level). Calculations of soluble-Al speciation for the pH 6.5 and 7.2 treatments that showed Al toxicity suggested the possible presence of the Al(13)O(4)(OH)(24)(H(2)O)(12)(7+) "tridecamer" cation and an inverse correlation of the tridecamer concentration and the cell population density. Analysis by (27)Al nuclear magnetic resonance spectroscopy, however, yielded no evidence of this species in freshly prepared samples or those taken 800 days after inoculation. Exclusion of the tridecamer species from the aqueous speciation calculations at pHs 6.5 and 7.2 yielded inverse correlations of the neutral Al(OH)(3) and anionic Al(OH)(4)(-) monomeric species with cell population density, suggesting that one or both of these ions bear primary responsibility for the toxicity observed.

Cadmium recovery by a sulfate-reducing magnetotactic bacterium, Desulfovibrio magneticus RS-1, using magnetic separation.

Cadmium recovery by a sulfate-reducing magnetotactic bacterium, Desulfovibrio magneticus strain RS-1, was investigated. D. magneticus precipitated >95% of cadmium at an initial concentration of 1.3 ppm in the growth medium. Electron microscopic analysis revealed that D. magneticus formed electron-dense particles on its surface when cultivated in the presence of cadmium ions (Cd2+). Sulfide was also found in the precipitate, and the composition ratio of sulfide/cadmium was 0.7. Sixty percent of viable RS-1 cells was recovered by a simple magnetic separation revealing the removal of 58% cadmium from the culture medium.

Activity of sulfate-reducing bacteria in human periodontal pocket.

Samples of subgingival dental tissues were examined for the presence of sulfate-reducing bacteria (SRB). Using enrichment cultures, SRBs were detected in 9 of 17 individuals. A pure culture of SRB was obtained from one sample collected from a patient with type IV periodontal disease. The characterization of this isolate showed that it belongs to the genus Desulfovibrio. The isolate used pyruvate, lactate, glucose, fructose, and ethanol as the sole source of carbon. However, the isolate was unable to use acetate and methanol as a carbon source, indicating it as an incomplete oxidizer unable to carry out the terminal oxidation of substrates. Apart from using sulfate as electron acceptor, the isolate also used thiosulfate and nitrate as an electron acceptor. It has the ability to use a variety of nitrogen sources, including ammonium chloride, nitrate, and glutamate. The optimum growth temperature of the isolate was 37 degrees C and the optimum pH for growth was 6.8. The SRB isolate contained the electron carrier desulfoviridin. The numbers of SRB in the mouth are assumed to be limited by sulfate. Potential sources of sulfate in the subgingival area include free sulfate in pocket fluid and glycosaminoglycans and sulfur-containing amino acids from periodontal tissues.

Isolation of Desulfovibrio intestinalis sp. nov. from the hindgut' of the lower termite Mastotermes darwiniensis.

A Gram-negative, anaerobic sulfate-reducing bacterium was isolated from hindgut contents of the lower termite Mastotermes darwiniensis Froggatt (strain KMS2). Strain KMS2 is motile by a single polar flagellum. The isolate possesses desulfoviridin and catalase activity. The G+C content of its DNA is in the range of 54.5-55.5 mol% (strain KMS2). It respires hydrogen and different low molecular weight organic compounds in the presence of sulfate, thiosulfate, and sulfite, and also oxygen. The isolated strain ferments pyruvate. Fastest growth with a doubling time of 12.5 h was obtained at 37 degrees C and not at 28 degrees C, the temperature at which the termites were grown. The isolate showed a 16S rDNA sequence homology of 95.9% to Desulfovibrio desulfuricans ATCC 27774 and a DNA-DNA homology of 44.6% to D. desulfuricans Essex 6 (type strain). Based on its biochemical properties and 16S rDNA sequence, the isolate was assigned to a new species named Desulfovibrio intestinalis.

Reduction of technetium by Desulfovibrio desulfuricans: biocatalyst characterization and use in a flowthrough bioreactor.

Resting cells of Desulfovibrio desulfuricans coupled the oxidation of a range of electron donors to Tc(VII) reduction. The reduced technetium was precipitated as an insoluble low-valence oxide. The optimum electron donor for the biotransformation was hydrogen, although rapid rates of reduction were also supported when formate or pyruvate was supplied to the cells. Technetium reduction was less efficient when the growth substrates lactate and ethanol were supplied as electron donors, while glycerol, succinate, acetate, and methanol supported negligible reduction. Enzyme activity was stable for several weeks and was insensitive to oxygen. Transmission electron microscopy showed that the radionuclide was precipitated at the periphery of the cell. Cells poisoned with Cu(II), which is selective for periplasmic but not cytoplasmic hydrogenases, were unable to reduce Tc(VII), a result consistent with the involvement of a periplasmic hydrogenase in Tc(VII) reduction. Resting cells, immobilized in a flowthrough membrane bioreactor and supplied with Tc(VII)-supplemented solution, accumulated substantial quantities of the radionuclide when formate was supplied as the electron donor, indicating the potential of this organism as a biocatalyst to treat Tc-contaminated wastewaters.

Psychrotolerant sulfate-reducing bacteria from an oxic freshwater sediment, description of Desulfovibrio cuneatus sp. nov. and Desulfovibrio litoralis sp. nov.

The most abundant culturable sulfate-reducing bacteria were isolated from the littoral sediment of the oligotrophic Lake Stechlin. The strains STL1 and STL4 were obtained from the oxic uppermost layer, while strain STL6 was isolated from the anoxic zone in 20 to 30 mm depth. The isolates showed a striking morphological feature in tapering off at one end of the cell. Physiological characteristics related them to the genus Desulfovibrio. They contained desulfoviridin. H2, formate, pyruvate, lactate, and fumarate were utilized with sulfate, sulfite, thiosulfate, or elemental sulfur as electron acceptors. All isolates were able to reduce oxygen and survived 120 h of aeration. However, aerobic growth was not observed. The isolates were psychrotolerant, and grew with rates of up to 0.29 d-1 at 4 degrees C. Analysis of the 16S rDNA confirmed that the strains belong to the genus Desulfovibrio. However, they were not closely related to any known member of this genus and formed a new cluster with at least two new species. Strain STL1 and STL4, exhibiting 99.7% sequence similarity in 16S rRNA, are proposed as the new species Desulfovibrio cuneatus sp. nov., while strain STL6 is assigned to the new species Desulfovibrio litoralis sp. nov.

Isolation and evaluation of susceptibility to sulphasalazine of Desulfovibrio desulfuricans strains from the human digestive tract.

Various genera of sulphate reducing bacteria (SRB) have been found in the human digestive tract. It is suggested that some of SRB species may be responsible for the development of the clinical symptoms of ulcerative colitis and other disease of large intestine. Sulphasalazine (salicyl-azo-sulphapyridine, SAS) is commonly used to treat patients with ulcerative colitis and Crohn disease. Above 30 samples of faeces or biopsy specimens from 25 patients (age 45 +/- 14 years; M/F, 13/12) suffering from gastrointestinal disorders were used for isolation of Desulfovibrio desulfuricans species. The morphological, physiological and biochemical characteristics of isolated strains and also their susceptibility to SAS was determined. D. desulfuricans isolates were obtained from 5 amongst all patients assayed. Some abnormal, cigar-shaped cells were detected as accompanying the cells represented by rods, curved rods and vibrios. After strains purification, two types of colonies were present on the solid Postgate's medium B (containing lactate as a carbon source and sulphate for energy conservation): the black colonies growing in bulk of agar medium and the transparent, surface-growing mucous colonies. These two types of D. desulfuricans colonies may be a result of different iron availability for bacterial cells. High metabolic activity of strain was not always accompanied by the presence of H2S gas lock in the test tube, although the H2S odor was perceptible. All tested strains multiplied inconsiderably slowly in the presence of SAS at concentrations 10, 20, 40 and 60 mg/cm3. The growing concentrations of SAS did not cause a proportional decrease of the bacterial cells number. Taking into account the positive results of using SAS to treat patients with some colonic diseases and the indicated resistance to SAS of intestinal D. desulfuricans strains, it appears probable, that this SRB species isn't responsible for the development of mentioned diseases.

Electron-dense granules in Desulfovibrio gigas do not consist of inorganic triphosphate but of a glucose pentakis(diphosphate).

Under certain growth conditions the sulfate-reducing bacterium Desulfovibrio gigas forms electron-dense granules in the cells which had been claimed to consist of a magnesium triphosphate). We observed granules after cultivation in media with a low Fe2+ or NH4+ concentration and reinvestigated the nature of the electron-dense bodies. Energy-dispersive X-ray analysis of the granules in the cells showed that they contain large amounts of P, Mg, and K. Gel electrophoresis and chromatographic analyses of isolated granules which had been dissolved in 20 mM EDTA, however, revealed discrepancies with commercially available polyphosphates. 31P-NMR spectra also lacked the peaks in the -22-ppm region which are characteristic for inner phosphates of polyphosphates confirming that the phosphocompound as isolated from the electron-dense bodies of D. gigas did not consist of polyphosphates. Using multinuclear NMR spectroscopy we showed that the electron-dense bodies of D. gigas contained a novel metabolite which was identified as alpha-glucose 1,2,3,4,6-pentakis(diphosphate).

Desulfovibrio gabonensis sp. nov., a new moderately halophilic sulfate-reducing bacterium isolated from an oil pipeline.

Two moderately halophilic sulfate-reducing bacteria were isolated from an African oil pipeline and designated strains SEBR 3640 and SEBR 2840T (T = type strain). Both of these strains possess traits that define the genus Desulfovibrio. The cells of both isolates were motile curved rods that had a single polar flagellum and contained desulfoviridin, and both isolates utilized lactate, pyruvate, malate, fumarate, succinate, and ethanol in the presence of sulfate. Sulfite, thiosulfate, and elemental sulfur were also used as an electron acceptors in the presence of lactate. However, both strains tolerated higher concentrations of NaCl (up to 17%) than all other Desulfovibrio species except Desulfovibrio halophilus, which tolerated a similar level of NaCl. The results of a 16S rRNA gene sequence analysis also placed the designated type strain, strain SEBR 2840, in the genus Desulfovibrio but revealed that this organism was significantly different from D. halophilus and all other validly described Desulfovibrio species. On the basis of our results, we propose that strain SEBR 2840T is a member of a new species of the genus Desulfovibrio, Desulfovibrio gabonensis. The type strain of D. gabonensis is strain SEBR 2840 (= DSM 10636).

Localization of membrane-associated (NiFe) and (NiFeSe) hydrogenases of Desulfovibrio vulgaris using immunoelectron microscopic procedures.

The intracellular location of membrane-associated (NiFe) and (NiFeSe) hydrogenases of Desulfovibrio vulgaris was determined using pre-embedding and post-embedding immunoelectron microscopic procedures. Polyclonal antisera directed against the purified (NiFe) and (NiFeSe) hydrogenases were raised in rabbits. One-day-old cultures of D. vulgaris, grown on a lactate/sulfate medium, were used for all experiments in these studies. For post-embedding labeling studies cells were fixed with 0.2% glutaraldehyde and 0.3% formaldehyde, dehydrated with methanol, and embedded in the low-temperature resin Lowicryl K4M. Our post-embedding studies using antibody-gold or protein-A-gold as electron-dense markers revealed the location of the two hydrogenases exclusively at the cell periphery; the precise membrane location was then demonstrated by pre-embedding labeling. Spheroplasts were incubated with the polyclonal antisera against (NiFe) and (NiFeSe) hydrogenase followed by ferritin-linked secondary antibodies prior to embedding and sectioning. The observed labeling pattern unequivocally revealed that the antigenic reactive sites of the (NiFe) hydrogenase are located in the near vicinity of the cytoplasmic membrane facing into the periplasmic space, whereas the (NiFeSe) hydrogenase is associated with the cytoplasmic side of the cytoplasmic membrane.

Putative signal peptide on the small subunit of the periplasmic hydrogenase from Desulfovibrio vulgaris.

We sequenced the NH2 terminus of the large and small subunits of the periplasmic hydrogenase from the sulfate-reducing bacterium Desulfovibrio vulgaris (Hildenborough) and found that the small subunit lacks a region of 34 NH4-terminal amino acids coded by the gene for the small subunit (G. Voordouw and S. Brenner, Eur. J. Biochem. 148:515-520, 1985). We suggest that this region constitutes a signal peptide based on comparison with known procaryotic signal peptides.

[Cellular study of a lysogenic Desulfovibrio desulfuricans culture after mitomycin C induction]. / Izuchenie kletok lizogennoi kul'tury Desulfovibrio desulfuricans posle induktsii mitomitsinom C.

Ultrathin sections through the cells of Desulfovibrio desulfuricans, strain B, were examined using electron microscopy after inducing the formation of phages with mitomycin C at a concentration of 4 micrograms/ml. Changes were found in the size of the cells, their cell wall and nucleoids. The cells contained phage particles of different maturity. The electron microscopic study of the induced phage has shown that it does not differ in morphology from the phage of this culture discovered by us earlier.

Localization of dehydrogenases, reductases, and electron transfer components in the sulfate-reducing bacterium Desulfovibrio gigas.

Various dehydrogenases, reductases, and electron transfer proteins involved in respiratory sulfate reduction by Desulfovibrio gigas have been localized with respect to the periplasmic space, membrane, and cytoplasm. This species was grown on a lactate-sulfate medium, and the distribution of enzyme activities and concentrations of electron transfer components were determined in intact cells, cell fractions prepared with a French press, and lysozyme spheroplasts. A significant fraction of formate dehydrogenase was demonstrated to be localized in the periplasmic space in addition to hydrogenase and some c-type cytochrome. Cytochrome b, menaquinone, fumarate reductase, and nitrite reductase were largely localized on the cytoplasmic membrane. Fumarate reductase was situated on the inner aspect on the membrane, and the nitrite reductase appeared to be transmembraneous. Adenylylsulfate reductase, bisulfite reductase (desulfoviridin), pyruvate dehydrogenase, and succinate dehydrogenase activities were localized in the cytoplasm. Significant amounts of hydrogenase and c-type cytochromes were also detected in the cytoplasm. Growth of D. gigas on a formate-sulfate medium containing acetate resulted in a 10-fold increase in membrane-bound formate dehydrogenase and a doubling of c-type cytochromes. Growth on fumarate with formate resulted in an additional increase in b-type cytochrome compared with lactate-sulfate-grown cells.