A new ecological adaptation to high sulfide by a Hydrogenobacter sp. growing on sulfur compounds but not on hydrogen

Thermophilic bacteria were isolated from a sulfide-rich, neutral hot spring in Iceland on gelrite minimal medium with 16 mM thiosulfate. The isolates were aerobic, obligate chemolithoautotrophs and used thiosulfate and sulfur as electron donors, producing sulfate from both substrates. No growth was observed with hydrogen as the sole electron donor, and no hydrogenase activity was detected. The cells were gram-negative and usually single, 4—5 μm long and 0.7 μm in diameter and formed sulfur globules after a few days of incubation. By SSU rRNA sequence comparisons, the bacterium was placed in the genus Hydrogenobacter with the closest relative to be Calderobacterium hydrogenophilum with 98.3% sequence similarity. This novel bacterium shows an ecological adaptation to high sulfide springs and is differentiated from its closest known relatives by lack of H₂ oxidation, deposition of sulfur and lower growth temperature.     

Use of reduced sulfur compounds by Beggiatoa sp.

A strain of Beggiatoa cf. leptomitiformis (OH-75-B, clone 2a) was isolated which is unique among reported strains in its ability to deposit internal sulfur granules from thiosulfate. It also deposited these characteristic granules (as all BEggiatoa species do) from sulfide. In cultures where growth was limited by exhaustion of organic substrates, these granules generally comprised about 20% of the total cell weight. With medium containing acetate and thiosulfate, no measurable utilization of thiosulfate or deposition of elemental sulfur (S0) took place until after the exponential growth phase. Neither sulfide nor thiosulfate added an increment to heterotrophic growth yield except for the weight of the deposited S0. The deposition of S0 from thiosulfate was probably a disproportionation in which S0 and sulfate were produced in a 1:1 ratio. Some of the S0 was further oxidized to sulfate. No autotrophic or mixotrophic growth was demonstrated for this strain. When inoculated in small, well-dispersed quantities into yeast extract medium, this strain grew only after long lags. Addition of the enzyme catalase eliminated initial lags and increased growth rates slightly. In contrast, catalase had no influence on growth rate when added to mineral medium containing acetate. In yeast extract medium, the inhibition of growth rate was presumably because of peroxides. Addition of thiosulfate was almost as effective as catalase in eliminating this inhibition. The S0 granules which, in this case, were deposited during the exponential growth phase, appeared to be partly responsible for this relief. This strain of Beggiatoa sp. remained active for at least 5 days under strictly anaerobic conditions, and under those conditions, it increased its dry weight by about 2.5-fold. Anaerobic "growth" and maintenance required the presence of an energy source, such as acetate. When cells containing much internal S0 were transferred to an organic anaerobic medium, a substantial portion of the internal S0 was eventually converted to sulfide.     

Rhodopseudomonas adriatica sp. nov., a new species of the Rhodospirillaceae,dependent on reduced sulfur compounds

A new purple nonsulfur bacterium was isolated from enrichment cultures of a sulfide-containing marine lagoon. The bacterium is similar to Rhodopseudomonas capsulata and is described as a new species of the genus Rhodopseudomonas: Rhodopseudomonas adriatica. Cells are non-motile, 0.5–0.8 m by 1.3–1.8 m, and multiply by binary fission. Intracytoplasmic membranes are of the vesicular type. The photosynthetic pigments are bacteriochlorophyll a and carotenoids of the spheroidene group. Growth is possible anaerobically in the light and at low pO₂ in the dark. Biotin and thiamine are required as growth factors. A wide variety of organic compounds, as well as sulfide and thiosulfate, are used as photosynthetic electron donors. Sulfide is oxidized to elemental sulfur, which is subsequently converted to sulfate, whereas thiosulfate oxidation occurs without measurable intermediate. Rhodopseudomonas adriatica is unable to assimilate sulfate, growth is only possible in the presence of a reduced sulfur compound.     

Degradation of ogranic sulfur compounds and the reduction of dibenzothiophene to biphenyl and hydrogen sulfide byDesulfovibrio desulfuricans M6

The microbial degradation of organic sulfur compounds was studied in the anaerobic conditions usingDesulfovibrio desulfuricans M6, a sulfate-reducing bacterium isolated from soil. Biphenyl was the major dibenzothiophene degradation product.     

A newly isolated heterotrophic bacterium,Xanthomonas sp. DY44, to oxidize hydrogen sulfide to polysulfide

Summary A newly isolated heterotrophic bacterium,Xanthomonas sp. DY44, was found to be capable of oxidizing hydrogen sulfide (H₂S). Cells made non-viable by heat treatment (120°C, 20 min) did not show H₂S oxidation. However, both cells sterilized by -rays irradiation and cell-free extract oxidized H₂S, suggesting the existence of the heat-labile intracellular enzymatic system for H₂S oxidation. AsXanthomonas sp. DY44 exhibited no autotrophic growth with H₂S in basal mineral medium, the H₂S oxidation was judged not to be a consequence of chemolithotrophic activity. Using X-rays photoelectron spectroscopy (XPS), the metabolic product of H₂S oxidation was assessed to be polysulfide.     

Degradation of hydrogen sulfide by Xanthomonas sp. strain DY44 isolated from peat.

Xanthomonas sp. strain DY44, capable of degrading H2S, was isolated from dimethyl disulfide-acclimated peat. This bacterium removed H2S either as a single gas or in the presence of the sulfur-containing compounds methanethiol, dimethyl sulfide, and dimethyl disulfide. The maximum specific H2S removal rate, obtained in the late stationary phase, was 3.92 mmol g of dry cells-1 h-1 (6.7 x 10(-16) mol cell-1 h-1) at pH 7 and 30 degrees C through a batch experiment in a basal mineral medium. Since Xanthomonas sp. strain DY44 exhibited no autotrophic growth with H2S, the H2S removal was judged not to be a consequence of chemolithotrophic activity. By using X-ray photoelectron spectroscopy, the metabolic product of H2S oxidation was determined to be polysulfide, which has properties very similar to those of elemental sulfur. Autoclaved cells (120 degrees C, 20 min) did not show H2S degradation, but cells killed by gamma-irradiation and cell extracts both oxidized H2S, suggesting the existence of a heat-labile intracellular enzymatic system for H2S oxidation. When Xanthomonas sp. strain DY44 was inoculated into fibrous peat, this strain degraded H2S without lag time, suggesting that it will be a good candidate for maintaining high H2S removability in the treatment of exhaust gases.     

Degradation of hydrogen sulfide by Xanthomonas sp. strain DY44 isolated from peat.

Xanthomonas sp. strain DY44, capable of degrading H2S, was isolated from dimethyl disulfide-acclimated peat. This bacterium removed H2S either as a single gas or in the presence of the sulfur-containing compounds methanethiol, dimethyl sulfide, and dimethyl disulfide. The maximum specific H2S removal rate, obtained in the late stationary phase, was 3.92 mmol g of dry cells-1 h-1 (6.7 x 10(-16) mol cell-1 h-1) at pH 7 and 30 degrees C through a batch experiment in a basal mineral medium. Since Xanthomonas sp. strain DY44 exhibited no autotrophic growth with H2S, the H2S removal was judged not to be a consequence of chemolithotrophic activity. By using X-ray photoelectron spectroscopy, the metabolic product of H2S oxidation was determined to be polysulfide, which has properties very similar to those of elemental sulfur. Autoclaved cells (120 degrees C, 20 min) did not show H2S degradation, but cells killed by gamma-irradiation and cell extracts both oxidized H2S, suggesting the existence of a heat-labile intracellular enzymatic system for H2S oxidation. When Xanthomonas sp. strain DY44 was inoculated into fibrous peat, this strain degraded H2S without lag time, suggesting that it will be a good candidate for maintaining high H2S removability in the treatment of exhaust gases.     

Chlorobaculum macestae sp. nov., a new green sulfur bacterium

The investigated green sulfur bacterium, strain M, was isolated from a sulfidic spring on the Black Sea Coast of the Caucasus. The cells of strain M are straight or curved rods 0.6-0.9 x 1.8-4.2 microm in size. According to the cell wall structure, the bacteria are gram-negative. Chlorosomes are located along the cell periphery. Strain M is an obligate anaerobe capable of photoautotrophic growth on sulfide, thiosulfate, and H2. It utilizes ammonium, urea, casein hydrolysate, and N2 as nitrogen sources and sulfide, thiosulfate, and elemental sulfur as sulfur sources. Bacteriochlorophyll c and the carotenoid chlorobactene are the main pigments. The optimal growth temperature is 25-28 degrees C; the optimal pH is 6.8. The strain does not require NaCl. Vitamin B12 stimulates growth. The content of the G+C base pairs in the DNA of strain M is 58.3 mol %. In the phylogenetic tree constructed on the basis of analysis of nucleotide sequences of 16S rRNA genes, strain M forms a separate branch, which occupies an intermediate position between the phylogenetic cluster containing representatives of the genus Chlorobaculum (94.9-96.8%) and the cluster containing species of the genus Chlorobium (94.1-96.5%). According to the results of analysis of the amino acid sequence corresponding to the fmo gene, strain M represents a branch which, unlike that in the "ribosomal" tree, falls into the cluster of the genus Chlorobaculum (95.8-97.2%). Phylogenetic analysis of the amino acid sequence corresponding to the nifH gene placed species of the genera Chlorobaculum and Chlorobium into a single cluster, whereas strain M formed a separate branch. The results obtained allow us to describe strain M as a new species of the genus Chlorobaculum. Chlorobaculum macestae sp. nov.     

Chemolithotrophic growth of Desulfovibrio sulfodismutans sp. nov. by disproportionation of inorganic sulfur compounds

A new Desulfovibrio strain ThAc01 was isolated from freshwater mud; the strain conserved energy for growth under strictly anaerobic conditions by disproportionation of thiosulfate or sulfite to sulfate and sulfide according to the following reactions: $$\begin{gathered} S_2 O_3^{2 - } + H_2 O \to SO_4^{2 - } + HS^ - + H^ + \hfill \\ 4SO_3^{2 - } + H^ + {\text{ }} \to 3SO_4^{2 - } + HS^ - \hfill \\ \end{gathered}$$ Strain ThAc01 required acetate as a carbon source, but was unable to utilize acetate as an oxidizable energy source. In a defined medium with acetate and bicarbonate as carbon sources, the growth yields per mol of substrate disproportionated were 2.1 g or 3.2 g dry cell mass on thiosulfate or sulfite, respectively. Strain ThAc01 was also able to grow by dissimilatory sulfate reduction with lactate, ethanol, propanol, or butanol as electron donors and carbon sources which were incompletely oxidized to the corresponding fatty acids. However, growth by sulfate reduction was slower than by disproportionation. Elemental sulfur, nitrate, fumarate, or malate did not serve as electron acceptors. Strain ThAc01 contained desulfoviridin and cytochromes; it required panthothenate and biotin as growth factors and had a DNA base ratio of 64.1 mol% G+C. Disproportionating bacteria similar to strain ThAc01 were enriched with either thiosulfate or sulfite from various freshwater, brackish or marine mud samples. Most probable number enumeration indicated that 2×10⁶ thiosulfate-disproportionating bacteria were present per ml freshwater mud. Of various other sulfate-reducing bacteria tested, only Desulfobacter curvatus (strain AcRM3) was able to disproportionate thiosulfate or sulfite. Desulfovibrio vulgaris (strain Marburg) slowly disproportionated sulfite, but effected only a slight increase in cell density. Strain ThAc01 is proposed as the type strain of a new species, Desulfovibrio sulfodismutans.     

Oxidation of dimethylsulfide to tetrathionate by Methylophaga thiooxidans sp. nov.: a new link in the sulfur cycle

A new pathway of dimethylsulfide (DMS) metabolism was identified in a novel species of Gammaproteobacteria, Methylophaga thiooxidans sp. nov., in which tetrathionate (S₄O₆^2?) was the end‐product of DMS oxidation. Inhibitor evidence indicated that DMS degradation was initiated by demethylation, catalysed by a corrinoid demethylase. Thiosulfate was an intermediate, which was oxidized to tetrathionate by a cytochrome‐linked thiosulfate dehydrogenase. Thiosulfate oxidation was coupled to ATP synthesis, and M. thiooxidans could also use exogenous thiosulfate as an energy source during chemolithoheterotrophic growth on DMS or methanol. Cultures grown on a variety of substrates oxidized thiosulfate, indicating that thiosulfate oxidation was constitutive. The observations have relevance to interactions among sulfur‐metabolizing bacteria in the marine environment. The production of tetrathionate from an organosulfur precursor is previously undocumented and represents a potential step in the biogeochemical sulfur cycle, providing a ‘shunt’ across the cycle.     

Utilization of dimethyl sulfide as a sulfur source with the aid of light by Marinobacterium sp. strain DMS-S1

Strain DMS-S1 isolated from seawater was able to utilize dimethyl sulfide (DMS) as a sulfur source only in the presence of light in a sulfur-lacking medium. Phylogenetic analysis based on 16S ribosomal DNA genes indicated that the strain was closely related to Marinobacterium georgiense. The strain produced dimethyl sulfoxide (DMSO), which was a main metabolite, and small amounts of formate and formaldehyde when grown on DMS as the sole sulfur source. The cells of the strain grown with succinate as a carbon source were able to use methyl mercaptan or methanesulfonate besides DMS but not DMSO or dimethyl sulfone as a sole sulfur source. DMS was transformed to DMSO primarily at wavelengths between 380 and 480 nm by heat-stable photosensitizers released by the strain. DMS was also degraded to formaldehyde in the presence of light by unidentified heat-stable factors released by the strain, and it appeared that strain DMS-S1 used the degradation products, which should be sulfite, sulfate, or methanesulfonate, as sulfur sources.     

Effect of biologically produced sulfur on gas absorption in a biotechnological hydrogen sulfide removal process

Absorption of hydrogen sulfide in aqueous suspensions of biologically produced sulfur particles was studied in a batch stirred cell reactor, and in a continuous set-up, consisting of a lab-scale gas absorber column and a bioreactor. Presence of biosulfur particles was found to enhance the absorption rate of H(2)S gas in the mildly alkaline liquid. The mechanism for this enhancement was however found to depend on the type of particles used. In the gently stirred cell reactor only small hydrophilic particles were present (d(p) < 3 microm) and the enhancement of the H(2)S absorption rate can be explained from the heterogeneous reaction between dissolved H(2)S and solid elemental sulfur to polysulfide ions, S(x) (2-). Conditions favoring enhanced H(2)S absorption for these hydrophilic particles are: low liquid side mass transfer (k(L)), high sulfur content, and presence of polysulfide ions. In the set-up of gas absorber column and bioreactor, both small hydrophilic particles and larger, more hydrophobic particles were continuously produced (d(p) up to 20 microm). Here, observed enhancement could not be explained by the heterogeneous reaction between sulfide and sulfur, due to the relatively low specific particle surface area, high k(L), and low [S(x) (2-)]. A more likely explanation for enhancement here is the more hydrophobic behavior of the larger particles. A local increase of the hydrophobic sulfur particle concentration near the gas/liquid interface and specific adsorption of H(2)S at the particle surface can result in an increase in the H(2)S absorption rate.     

A novel membrane-bound flavocytochrome c sulfide dehydrogenase from the colourless sulfur bacterium Thiobacillus sp. W5

A novel membrane-bound sulfide-oxidizing enzyme was purified 102-fold from the neutrophilic, obligately chemolithoautotrophic Thiobacillus sp. W5 by means of a six-step procedure. Spectral analysis revealed that the enzyme contains haem c and flavin. SDS-PAGE showed the presence of two types of subunit with molecular masses of 40 and 11 kDa. The smaller subunit contains covalently bound haem c, as was shown by haem staining. A combination of spectral analysis and the pyridine haemochrome test indicated that the sulfide-oxidizing heterodimer contains one molecule of haem c and one molecule of flavin. It appeared that the sulfide-oxidizing enzyme is a member of a small class of redox proteins, the flavocytochromes c, and is structurally most related to the flavocytochrome c sulfide dehydrogenase of the green sulfur bacterium Chlorobium limicola. The pH optimum of the enzyme is 8.6. At pH 9, the V _max was 2.1 ± 0.1 μmol cytochrome c (mg protein)^–1 min^–1, and the K _m values for sulfide and cytochrome c were 1.7 ± 0.4 μM and 3.8 ± 0.8 μM, respectively. Cyanide inhibited the enzyme by the formation of an N-5 adduct with the flavin moiety of the protein. On the basis of electron transfer stoichiometry, it seems likely that sulfur is the oxidation product. Received: 15 October 1996 / Accepted: 7 January 1997     

Thiocapsa halophila sp. nov., a new halophilic phototrophic purple sulfur bacterium

A new phototrophic sulfur bacterium has been isolated from a red layer in a laminated mat occurring underneath a gypsum crust in the mediterranean salterns of Salin-de-Giraud (Camargue, France). Single cells were coccus-shaped, non motile, without gas vacuoles and contained sulfur globules. Bacteriochlorophyll a and okenone were present as major photosynthetic pigments. These properties and the G+C content of DNA (65.9–66.6 mol% G+C) are typical characteristics of the genus Thiocapsa. However, the new isolate differs from known species in the genus, particularly in NaCl requirement (optimum, 7% NaCl; range, 3–20% NaCl) and some physiological characteristics. Therefore, a new species is proposed, Thiocapsa halophila, sp. nov.Dedicated to Prof. Dr. Norbert Pfennig in occasion of his 65th birthday     

Thermococcus acidaminovorans sp. nov., a new hyperthermophilic alkalophilic archaeon growing on amino acids

From a shallow marine hydrothermal system at Vulcano (Italy), a new hyperthermophilic member of the Archaea was isolated. The cells are coccoid – shaped and possess up to five flagella. They grow between 56° and 93°C (optimum 85°C) and pH 5.0–9.5 (optimum 9.0). The organism is strictly anaerobic and grows heterotrophically on defined amino acids and complex organic substrates such as casamino acids, yeast extract, peptone, meat extract, tryptone, and casein. Polysulfide and elemental sulfur are reduced to H₂S. In the absence of polysulfide or elemental sulfur, the isolate grows at a significantly reduced rate. Growth is not influenced by the presence of H₂. DNA–DNA hybridization and 16S rRNA partial sequences indicated that the new isolate belongs to the genus Thermococcus, and represents a new species, Thermococcus acidaminovorans. The type strain is isolate AEDII10 (DSM 11906). Received: September 24, 1997 / Accepted: January 1, 1998     

Characterization of a new platelet-forming purple sulfur bacterium,Amoebobacter pedioformis sp. nov.

The dominant purple sulfur bacterium of a reddish-colored waste water pond near Taichung, Taiwan, was isolated in pure culture, strain CML2. Individual cells were nearly spherical, nonmotile, and contained in their peripheral parts was vacuoles that appeared like elongated, curved tubes. Four to sixteen cells formed platelet-like aggregates reminiscent of Thiopedia rosea. The intracellular photosynthetic membrane system of the cells was of vesicular type; the photosynthetic pigments consisted of bacteriochlorophyll a and spirilloxanthin as the major carotenoid. The color of cell suspensions was pink to rosered. Under anaerobic conditions photolithoautotrophic growth occurred with sulfide, elemental sulfur or thiosulfate; sulfur globules were stored as an intermediary oxidation product. In the presence of sulfide, acetate, lactate and pyruvate were photoassimilated; strain CML2 lacked assimilatory sulfate reduction. Fastest photoautotrophic growth (11 h doubling time) was obtained at pH 7.5, 35°C and a light intensity of about 1000 lux (tungsten lamp). Chemolithoautotrophic growth in the dark was possible under reduced oxygen partial pressure with reduced sulfur compounds as respiratory substrates. The DNA base composition of strain CML2 was 65.5 mol% G+C. Strain CML2 is described as type strain of a new species, Amoebobacter pedioformis sp. nov., in the family Chromatiaceae.