Extracellular Elemental Sulfur Research Articles

Organic Stabilization of Extracellular Elemental Sulfur in a Sulfurovum-Rich Biofilm: A New Role for Extracellular Polymeric Substances?

This work shines light on the role of extracellular polymeric substance (EPS) in the formation and preservation of elemental sulfur biominerals produced by sulfur-oxidizing bacteria. We characterized elemental sulfur particles produced within a Sulfurovum-rich biofilm in the Frasassi Cave System (Italy). The particles adopt spherical and bipyramidal morphologies, and display both stable (α-S8) and metastable (β-S8) crystal structures. Elemental sulfur is embedded within a dense matrix of EPS, and the particles are surrounded by organic envelopes rich in amide and carboxylic groups. Organic encapsulation and the presence of metastable crystal structures are consistent with elemental sulfur organomineralization, i.e., the formation and stabilization of elemental sulfur in the presence of organics, a mechanism that has previously been observed in laboratory studies. This research provides new evidence for the important role of microbial EPS in mineral formation in the environment. We hypothesize that the extracellular organics are used by sulfur-oxidizing bacteria for the stabilization of elemental sulfur minerals outside of the cell wall as a store of chemical energy. The stabilization of energy sources (in the form of a solid electron acceptor) in biofilms is a potential new role for microbial EPS that requires further investigation.

Discovery of a new subgroup of sulfur dioxygenases and characterization of sulfur dioxygenases in the sulfur metabolic network of Acidithiobacillus caldus.

Acidithiobacillus caldus is a chemolithoautotrophic sulfur-oxidizing bacterium that is widely used for bioleaching processes. Acidithiobacillus spp. are suggested to contain sulfur dioxygenases (SDOs) that facilitate sulfur oxidation. In this study, two putative sdo genes (A5904_0421 and A5904_1112) were detected in the genome of A. caldus MTH-04 by BLASTP searching with the previously identified SDO (A5904_0790). We cloned and expressed these genes, and detected the SDO activity of recombinant protein A5904_0421 by a GSH-dependent in vitro assay. Phylogenetic analysis indicated that A5904_0421and its homologous SDOs, mainly found in autotrophic bacteria, were distantly related to known SDOs and were categorized as a new subgroup of SDOs. The potential functions of genes A5904_0421 (termed sdo1) and A5904_0790 (termed sdo2) were investigated by generating three knockout mutants (Δsdo1, Δsdo2 and Δsdo1&2), two sdo overexpression strains (OE-sdo1 and OE-sdo2) and two sdo complemented strains (Δsdo1/sdo1’ and Δsdo2/sdo2’) of A. caldus MTH-04. Deletion or overexpression of the sdo genes did not obviously affect growth of the bacteria on S0, indicating that the SDOs did not play an essential role in the oxidation of extracellular elemental sulfur in A. caldus. The deletion of sdo1 resulted in complete inhibition of growth on tetrathionate, slight inhibition of growth on thiosulfate and increased GSH-dependent sulfur oxidation activity on S0. Transcriptional analysis revealed a strong correlation between sdo1 and the tetrathionate intermediate pathway. The deletion of sdo2 promoted bacterial growth on tetrathionate and thiosulfate, and overexpression of sdo2 altered gene expression patterns of sulfide:quinone oxidoreductase and rhodanese. Taken together, the results suggest that sdo1 is essential for the survival of A. caldus when tetrathionate is used as the sole energy resource, and sdo2 may also play a role in sulfur metabolism.

Mechanisms of extracellular S0 globule production and degradation in Chlorobaculumtepidum via dynamic cell-globule interactions.

The Chlorobiales are anoxygenic phototrophs that produce solid, extracellular elemental sulfur globules as an intermediate step in the oxidation of sulfide to sulfate. These organisms must export sulfur while preventing cell encrustation during S0 globule formation; during globule degradation they must find and mobilize the sulfur for intracellular oxidation to sulfate. To understand how the Chlorobiales address these challenges, we characterized the spatial relationships and physical dynamics of Chlorobaculum tepidum cells and S0 globules by light and electron microscopy. Cba. tepidum commonly formed globules at a distance from cells. Soluble polysulfides detected during globule production may allow for remote nucleation of globules. Polysulfides were also detected during globule degradation, probably produced as an intermediate of sulfur oxidation by attached cells. Polysulfides could feed unattached cells, which made up over 80% of the population and had comparable growth rates to attached cells. Given that S0 is formed remotely from cells, there is a question as to how cells are able to move toward S0 in order to attach. Time-lapse microscopy shows that Cba. tepidum is in fact capable of twitching motility, a finding supported by the presence of genes encoding type IV pili. Our results show how Cba. tepidum is able to avoid mineral encrustation and benefit from globule degradation even when not attached. In the environment, Cba. tepidum may also benefit from soluble sulfur species produced by other sulfur-oxidizing or sulfur-reducing bacteria as these organisms interact with its biogenic S0 globules.

Thiosulfate oxidation by Thiomicrospira thermophila: metabolic flexibility in response to ambient geochemistry.

Previous studies of the stoichiometry of thiosulfate oxidation by colorless sulfur bacteria have failed to demonstrate mass balance of sulfur, indicating that unidentified oxidized products must be present. Here the reaction stoichiometry and kinetics under variable pH conditions during the growth of Thiomicrospira thermophila strain EPR85, isolated from diffuse hydrothermal fluids at the East Pacific Rise, is presented. At pH 8.0, thiosulfate was stoichiometrically converted to sulfate. At lower pH, the products of thiosulfate oxidation were extracellular elemental sulfur and sulfate. We were able to replicate previous experiments and identify the missing sulfur as tetrathionate, consistent with previous reports of the activity of thiosulfate dehydrogenase. Tetrathionate was formed under slightly acidic conditions. Genomic DNA from T. thermophila strain EPR85 contains genes homologous to those in the Sox pathway (soxAXYZBCDL), as well as rhodanese and thiosulfate dehydrogenase. No other sulfur oxidizing bacteria containing sox(CD)2 genes have been reported to produce extracellular elemental sulfur. If the apparent modified Sox pathway we observed in T. thermophila is present in marine Thiobacillus and Thiomicrospira species, production of extracellular elemental sulfur may be biogeochemically important in marine sulfur cycling.

Chlorobaculum tepidum growth on biogenic S(0) as the sole photosynthetic electron donor.

The green sulfur bacteria, the Chlorobi, are phototrophic bacteria that oxidize sulfide and deposit extracellular elemental sulfur globules [S(0)]. These are subsequently consumed after sulfide is exhausted. S(0) globules from a Chlorobaculum tepidum mutant strain were purified and used to show that the wild-type strain of Cba. tepidum can grow on biogenic S(0) globules as the sole photosynthetic electron donor, i.e. in medium with no other source of reducing power. Growth yields and rates on biogenic S(0) are comparable with those previously determined for Cba. tepidum grown on sulfide as the sole electron donor. Contact between cells and S(0) was required for growth. However, only a fraction of the cell population was firmly attached to S(0) globules. Microscopic examination of cultures growing on S(0) demonstrated cell-S(0) attachment and allowed for the direct observation of S(0) globule degradation. Bulk chemical analysis, scanning electron microscopy, secondary ion mass spectrometry and SDS-PAGE indicate that Cba. tepidum biogenic S(0) globules contain carbon, oxygen and nitrogen besides S and may be associated with specific proteins. These observations suggest that current models of S(0) oxidation in the Chlorobi need to be revised to take into account the role of cell-S(0) interactions in promoting S(0) degradation.

Rhodovulum tesquicola sp. nov., a haloalkaliphilic purple non-sulfur bacterium from brackish steppe soda lakes

Two strains of purple non-sulfur bacteria (A-36s(T) and A-51s) were isolated from brackish steppe soda lakes of southern Siberia. Genetically, the isolates were related most closely to the type strains of Rhodovulum steppense and Rhodovulum strictum, from which they differed at the species level (98.5% 16S rRNA gene sequence similarity, 40-53% DNA-DNA relatedness). Cells of the two strains were ovoid to rod-shaped, 0.4-0.8 µm wide and 1.0-2.5 µm long, and motile by means of a polar flagellum. They contained internal photosynthetic membranes of vesicular type and photosynthetic pigments (bacteriochlorophyll a and carotenoids of the spheroidene series). The strains were obligate haloalkaliphiles, growing over wide ranges of salinity (0.3-10.0% NaCl) and pH (7.5-10.0), with growth optima at 1.0-3.0% NaCl and pH 8.5-9.0. Photoheterotrophic and chemoheterotrophic growth occurred with a number of organic compounds and biotin, p-aminobenzoate, thiamine and niacin as growth factors. No anaerobic respiration on nitrite, nitrate or fumarate and no fermentation were demonstrated. The strains grew photolithoautotrophically and chemolithoautotrophically with sulfide, sulfur and thiosulfate, oxidizing them to sulfate. Sulfide was oxidized via deposition of extracellular elemental sulfur. No growth with H(2) as the electron donor was observed. The major fatty acid was C(18:1) (78%). The major quinone was ubiquinone Q-10. The DNA G+C content of strain A-36s(T) was 65.4 mol% (T(m)). According to genotypic and phenotypic characteristics, the investigated strains were assigned to a novel species of the genus Rhodovulum, for which the name Rhodovulum tesquicola sp. nov. is proposed. The type strain is A-36s(T) ( = VKM B-2491(T) = ATCC BAA-1573(T)), which was isolated from steppe soda lake Sul'fatnoe (Zabaikal'skii Krai, southern Siberia, Russia).

FORMATION OF SULPHUR COMPOUNDS WITH DIFFERENT OXIDATION BACTERIAL CELLS CHLOROBIUM LIMICOLA IМВ K-8

Green sulfur bacteria C. limicola IМВ K-8 during cultivation in medium with H2 S accumulate extracellular elemental sulfur. After full use of hydrogen sulfide, sulfur content decreases, and in the medium it is observed the formation of sulfate ion, the maximum amount of which is piled up in 13 days. In experiments with washed cells C. limicola IМВ K-8 it is shown that the concentration of sulfate ion reaches maximum, in twenty hours of incubation both in the light and in darkness. It is established that priority substrate oxidation in the absence of hydrogen sulfide is endogenous glucose and glycogen. The sources of energy under stress conditions are used in the following sequence: glucose – glycogen – elemental sulfur.

Complete genome sequence of Rhodospirillum rubrum type strain (S1).

Rhodospirillum rubrum (Esmarch 1887) Molisch 1907 is the type species of the genus Rhodospirillum, which is the type genus of the family Rhodospirillaceae in the class Alphaproteobacteria. The species is of special interest because it is an anoxygenic phototroph that produces extracellular elemental sulfur (instead of oxygen) while harvesting light. It contains one of the most simple photosynthetic systems currently known, lacking light harvesting complex 2. Strain S1T can grow on carbon monoxide as sole energy source. With currently over 1,750 PubMed entries, R. rubrum is one of the most intensively studied microbial species, in particular for physiological and genetic studies. Next to R. centenum strain SW, the genome sequence of strain S1T is only the second genome of a member of the genus Rhodospirillum to be published, but the first type strain genome from the genus. The 4,352,825 bp long chromosome and 53,732 bp plasmid with a total of 3,850 protein-coding and 83 RNA genes were sequenced as part of the DOE Joint Genome Institute Program DOEM 2002.

Rhodovulum steppense sp. nov., an obligately haloalkaliphilic purple nonsulfur bacterium widespread in saline soda lakes of Central Asia

Seven strains of purple nonsulfur bacteria isolated from the shallow-water steppe soda lakes of the cryoarid zone of Central Asia formed a genetically homogeneous group within the genus Rhodovulum. The isolates were most closely related to Rhodovulum strictum, from which they differed at the species level (99.5 % 16S rRNA gene identity and 42-44 % DNA-DNA hybridization level). According to genotypic and phenotypic characteristics, the strains were assigned to a new species of the genus Rhodovulum, for which the name Rhodovulum steppense sp. nov. is proposed. Cells of all strains were ovoid to rod-shaped, 0.3-0.8 microm wide and 1-2.5 microm long, and motile by means of polar flagella. They contained internal photosynthetic membranes of the vesicular type and photosynthetic pigments (bacteriochlorophyll a and carotenoids of the spheroidene series). All strains were obligate haloalkaliphiles, growing within a wide range of salinity (0.3-10 %) and pH (7.5-10), with growth optima at 1-5 % NaCl and pH 8.5. Photo- and chemoheterotrophic growth occurred with a number of organic compounds and biotin, thiamine and niacin as growth factors. No anaerobic respiration on nitrite, nitrate or fumarate and no fermentation was demonstrated. Bacteria grew photo- and chemolithoautotrophically with sulfide, sulfur and thiosulfate, oxidizing them to sulfate. Sulfide was oxidized via deposition of extracellular elemental sulfur. No growth with H(2) as electron donor was demonstrated. The major fatty acid was 18 : 1 (81.0 %). The major quinone was Q-10. The DNA G+C content was 66.1 mol% (T(m)). The type strain, A-20s(T) (=VKM B-2489(T) =DSM 21153(T)), was isolated from soda lake Khilganta (Zabaikal'skii Krai, southern Siberia, Russia).

Production of elemental sulfur by green and purple sulfur bacteria

The utilization of sulfide by phototrophic sulfur bacteria temporarily results in the accumulation of elemental sulfur. In the green sulfur bacteria (Chlorobiaceae), the sulfur is deposited outside the cells, whereas in the purple sulfur bacteria (Chromatiaceae) sulfur is found intracellularly. Consequently, in the latter case, sulfur is unattainable for other individuals. Attempts were made to analyze the impact of the formation of extracellular elemental sulfur compared to the deposition of intracellular sulfur.

Growth of thermophilic, obligatorily chemolithoautotrophic hydrogen-oxidizing bacteria related toHydrogenobacterwith thiosulfate and elemental sulfur as electron and energy source

Strains related to Hydrogenobacter, a genus of thermophilic, obligatorily chemolithoautotrophic bacteria, were able to utilize elemental sulfur or thiosulfate, as well as molecular hydrogen, as sole electron and energy source. Extracellular elemental sulfur was produced as an intermediate during oxidation of thiosulfate. Growth with thiosulfate alone was strongly microaerophilic, whereas no hydrogenase activity was detected. Mixolithotrophic growth with both hydrogen and thiosulfate was faster than with hydrogen alone, and the cells harbored a hydrogenase activity comparable to that of cells grown under hydrogen without thiosulfate.

Growth of thermophilic, obligatorily chemolithoautotrophic hydrogen-oxidizing bacteria related to Hydrogenobacter with thiosulfate and elemental sulfur as electron and energy source

Strains related to Hydrogenobacter, a genus of thermophilic, obligatorily chemolithoautotrophic bacteria, were able to utilize elemental sulfur or thiosulfate, as well as molecular hydrogen, as sole electron and energy source. Extracellular elemental sulfur was produced as an intermediate during oxidation of thiosulfate. Growth with thiosulfate alone was strongly microaerophilic, whereas no hydrogenase activity was detected. Mixolithotrophic growth with both hydrogen and thiosulfate was faster than with hydrogen alone, and the cells harbored a hydrogenase activity comparable to that of cells grown under hydrogen without thiosulfate.

Coexistence of Chlorobium and Chromatium in a sulfide-limited continuous culture

Competition experiments between Chromatium vinosum and Chlorobium limicola in sulfide-limited continuous culture under photolithoautotrophic conditions resulted in the coexistence of both organisms. The ratio between the two bacteria was dilution-rate as well as pH dependent. The observed coexistence can be explained as a hitherto not reported form of dual substrate limitation. The two substrates involved are the electron donors sulfide (growth-limiting substrate in the reservoir vessel) and extracellular elemental sulfur (formed by Chlorobium as a result of sulfide oxidation). It is argued that, although Chlorobium may have the better affinity for both substrates involved, Chromatium can compete successfully on the basis of its intracellular storage of sulfur. Ecological implication of the observed coexistence with respect to natural blooms are discussed.

Ectothiorhodospira halochloris sp. nov., a new extremely halophilic phototrophic bacterium containing bacteriochlorophyll b

A new bacteriochlorophyll b containing phototrophic bacterium was isolated from extremely saline and alkaline soda lakes in Egypt. Enrichment and isolation were performed using a synthetic medium with high contents of sodium carbonate, sodium sulfate and sodium chloride. Photoautotrophic growth occurred with hydrogen sulfide as photosynthetic electron donor. During oxidation of sulfide to sulfate extracellular elemental sulfur globules appeared in the medium. Cells were also capable to grow under photoheterotrophic conditions with acetate, propionate, pyruvate, succinate, fumarate or malate as carbon sources and electron donors. Under these conditions sulfate was assimilated. Optimal growth under the applied experimental conditions occurred at a total salinity of 14–27%, a pH-range between 8.1 and 9.1 and a temperature between 47°C and 50°C. The cells were 0.5–0.6 μm wide and, depending on cultural conditions, 2.5–8.0 μm long; they were spiral shaped, multiplied by binary fission and were motile by means of bipolar flagella. Intercytoplasmic photosynthetic membranes were present as stacks. Bacteriochlorophyll b was the main photosynthetic pigment; small amounts of carotenoids were mainly present as glucosides of rhodopin and its methoxy derivative. The new organism is described as Ectothiorhodospira halochloris.

Applications of Infrared Spectroscopy in Biochemistry, Biology, and Medicine.Frank S. Parker

Previous articleNext article No AccessNew Biological BooksApplications of Infrared Spectroscopy in Biochemistry, Biology, and Medicine. Frank S. Parker George J. Thomas, Jr.George J. Thomas, Jr. Search for more articles by this author PDFPDF PLUS Add to favoritesDownload CitationTrack CitationsPermissionsReprints Share onFacebookTwitterLinkedInRedditEmail SectionsMoreDetailsFiguresReferencesCited by The Quarterly Review of Biology Volume 47, Number 4Dec., 1972 Published in association with Stony Brook University Article DOIhttps://doi.org/10.1086/407481 Views: 4Total views on this site Citations: 1Citations are reported from Crossref PDF download Crossref reports the following articles citing this article:Brandi Cron, Jennifer L. Macalady, Julie Cosmidis Organic Stabilization of Extracellular Elemental Sulfur in a Sulfurovum-Rich Biofilm: A New Role for Extracellular Polymeric Substances?, Frontiers in Microbiology 12 (Aug 2021).https://doi.org/10.3389/fmicb.2021.720101

Sulfide utilization by purple nonsulfur bacteria

The purple nonsulfur bacteria Rhodospirillum rubrum SMG 107, Rhodopseudomonas capsulata SMG 155, Rps. sphaeroides SMG 158 and Rps. palustris SMG 124 were tested for a possible utilization of sulfide. The first three strains were found to oxidize sulfide to extracellular elemental sulfur only, whereas Rps. palustris SMG 124 converted sulfide into sulfate without intermediate accumulation of elemental sulfur. Growth ceased at lower sulfide concentrations than usually found with purple sulfur bacteria. In consequence of the low sulfide tolerance information on the specific growth rates obtainable with sulfide as photosynthetic electron donor could not be provided by cultivation in batch cultures. Sulfide-limited chemostat cultures of Rps. capsulata SMG 155 showed that the maximum specific growth rate was close to 0.14 h-1 (doubling time 5 h). Sulfide was converted into extracellular elemental sulfur at all dilution rates tested. The maximum specific growth rate of Rps. palustris SMG 124 was found to be much lower (less than 0.03 h-1). Sulfate was the only product of the conversion of sulfide.