Occurrence of sulfate-reducing bacteria in well water: identification of anaerobic sulfidogenic bacterial enrichment cultures.

Bacteriological studies of well water mainly focus on aerobic and facultative aerobic coliform bacteria. However, the presence of obligate anaerobic bacteria in well water, especially sulfate-reducing bacteria (SRB), possible causative agents of some diseases, is often ignored. In this study, the presence of SRB and coexisting anaerobic bacteria with SRB in sulfate-reducing enrichment cultures obtained from 10 well water samples in Istanbul was investigated. A nested polymerase chain reaction-denaturing gradient gel electrophoresis strategy was performed to characterize the bacterial community structure of the enrichments. The most probable number method was used to determine SRB number. Out of 10, SRB growth was observed in only one (10%) enrichment culture and the SRB number was low (<10 cells/mL). Community members were identified as Desulfolutivibrio sulfodismutans and Anaerosinus sp. The results show that SRB coexist with Anaerosinus sp., and this may indicate poor water quality, posing a risk to public health. Furthermore, Anaerosinus sp., found in the human intestinal tract, may be used as an alternative anaerobic fecal indicator. It is worth noting that the detection of bacteria using molecular analyzes following enrichment culture techniques can bring new perspectives to determine the possible origin and presence of alternative microbial indicators in aquatic environments.

Fusibacter ferrireducens sp. nov., an anaerobic, Fe(â ¢)- and sulphur-reducing bacterium isolated from mangrove sediment.

An anaerobic, alkaliphilic, halotolerant, Gram-stain-positive and rod-shaped bacterium, designated Q10-2T, was isolated from mangrove sediment sampled at the Jiulong river estuary, PR China. The cells of strain Q10-2T were motile and 0.5×2-4 µm in size. Strain Q10-2T grew at 8-45 °C (optimum, 32 °C), at pH 7.0-10.5 (optimum, pH 8.5) and in the presence of 0-6 % (w/v) NaCl (optimum, 3 %). It could use complex organic compounds and carbohydrates including d-fructose, d-galactose, d-glucose, d-mannitol, d-xylose, trehalose, lactose, maltose, sucrose and starch as carbon sources and electron donors. It could reduce sulphate, thiosulphate and elemental sulphur to sulphide, but not sulphite. Fe (Ⅲ) citrate, ferrihydrite, haematite and goethite in the presence of glucose as the electron donor were also reduced. Acetate, butyrate, ethanol, CO2 and H2 were end products of glucose fermentation. The predominant cellular fatty acids were composed of C14 : 0, C16 : 0 and summed features containing C16 : 1 ω7c and/or iso-C15 : 0 2-OH and iso-C17 : 1 and/or anteiso-C17 : 1 B. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the novel strain was most closely related to Fusibacter paucivorans DSM 12116T (95.5 % sequence similarity). The genome size of strain Q10-2T was 5.0 Mb, with a G+C content of 37.4 mol%. The average nucleotide identity and digital DNA-DNA hybridization values between strain Q10-2T and F. paucivorans DSM 12116T were 69.1 and 21.8 %, respectively. The combined genotypic and phenotypic data showed that strain Q10-2T represents a novel species of the genus Fusibacter, for which the name Fusibacter ferrireducens sp. nov. is proposed. The type strain is Q10-2T (=MCCC 1A16257T=KCTC 15906T).

Desulfomarina profundi gen. nov., sp. nov., a novel mesophilic, hydrogen-oxidizing, sulphate-reducing chemolithoautotroph isolated from a deep-sea hydrothermal vent chimney.

A novel mesophilic, strictly anaerobic, chemolithoautotrophic sulphate-reducing bacterium, designated strain KT2T, was isolated from a deep-sea hydrothermal vent chimney at the Suiyo Seamount in the Izu-Bonin Arc. Strain KT2T grew at 25-40 °C (optimum 35 °C) and pH 5.5-7.0 (optimum 6.6) in the presence of 25-45 g l-1 NaCl (optimum 30 g l-1). Growth occurred with molecular hydrogen as the electron donor and sulphate, thiosulphate, and sulphite as the electron acceptors. The isolate utilized CO2 as the sole carbon source for chemolithoautotrophic growth on H2. Glycerol, succinate, fumarate, malate, glutamate, or casamino acids could serve as an alternative electron donor in the presence of CO2. Malate, citrate, glutamate, and casamino acids were used as fermentative substrates for weak growth. The G+C content of genomic DNA was 46.1 %. Phylogenetic analysis of the 16S rRNA gene sequence indicated that strain KT2T is a member of the family Desulfobulbaceae, showing a sequence similarity of 94.3 % with Desulforhopalus singaporensis. Phylogenomic analysis based on concatenated 156 single-copy marker genes confirmed the same topology as the 16S rRNA gene phylogeny. The ANI and AAI values between strain KT2T and related genera of the family Desulfobulbaceae were 65.6-68.6 % and 53.1-62.9 %. Based on the genomic, molecular, and physiological characteristics, strain KT2T represents a novel genus and species within the family Desulfobulbaceae, for which the name Desulfomarina profundi gen. nov., sp. nov. is proposed, with KT2T (=JCM 34118T = DSM 111364T) as the type strain.

Detection of Sulfate-Reducing Bacteria as an Indicator for Successful Mitigation of Sulfide Production.

Sulfate-reducing bacteria (SRBs) are one of the main sources of biogenic H2S generation in oil reservoirs. Excess H2S production in these systems leads to oil biosouring, which causes operational risks and health hazards and can increase the cost of refining crude oil. Nitrate salts are often added to the system to suppress sulfidogenesis. Because SRB populations can persist in biofilms even after nitrate treatment, identifying shifts in the sessile community is crucial for successful mitigation. However, sampling the sessile community is hampered by its inaccessibility. Here, we use the results of a long-term (148 days) ex situ experiment to identify particular sessile community members from observations of the sample waste stream. Microbial community structure was determined for 731 samples across 20 bioreactors using 16S rRNA gene sequencing. By associating microbial community structure with specific steps in the mitigation process, we could distinguish between taxa associated with H2S production and mitigation. After initiation of nitrate treatment, certain SRB populations increased in the planktonic community during critical time points, indicating the dissociation of SRBs from the biofilm. Predicted relative abundances of the dissimilatory sulfate reduction pathway also increased during the critical time points. Here, by analyzing the planktonic community structure, we describe a general method that uses high-throughput amplicon sequencing, metabolic inferences, and cell abundance data to identify successful biofilm mitigation. We anticipate that our approach is also applicable to other systems where biofilms must be mitigated but cannot be sampled easily. IMPORTANCE Microbial biofilms are commonly present in many industrial processes and can negatively impact performance and safety. Within the oil industry, subterranean biofilms cause biosouring with implications for oil quality, cost, occupational health, and the environment. Because these biofilms cannot be sampled directly, methods are needed to indirectly assess the success of mitigation measures. This study demonstrates how the planktonic microbial community can be used to assess the dissociation of sulfate-reducing bacterium (SRB)-containing biofilms. We found that an increase in the abundance of a specific SRB population in the effluent after nitrate treatment can be used as a potential indicator for the successful mitigation of biofilm-forming SRBs. Moreover, a method for determining critical time points for detecting potential indicators is suggested. This study expands our knowledge of improving mitigation strategies for biosouring and could have broader implications in other systems where biofilms lead to adverse consequences.

Cupidesulfovibrio liaohensis gen. nov., sp. nov., a novel sulphate-reducing bacterium isolated from an oil reservoir and reclassification of Desulfovibrio oxamicus and Desulfovibrio termitidis as Cupidesulfovibrio oxamicus comb. nov. and Cupidesulfovibrio termitidis comb. nov.

A novel sulphate-reducing, Gram-stain-negative, anaerobic strain, isolate XJ01T, recovered from production fluid at the LiaoHe oilfield, PR China, was the subject of a polyphasic study. The isolate together with Desulfovibrio oxamicus NCIMB 9442T and Desulfovibrio termitidis DSM 5308T formed a distinct, well-supported clade in the Desulfovibrionaceae 16S rRNA gene tree. The taxonomic status of the clade was underscored by complementary phenotypic data. The three isolates comprising the clade formed distinct phyletic branches and were distinguished using a combination of physiological features and by low average nucleotide identity and digital DNA-DNA hybridization values. Consequently, it is proposed that isolate XJ01T represents a novel genus and species for which the name Cupidesulfovibrio liaohensis gen. nov., sp. nov. is proposed with the type strain XJ01T (=CGMCC 1.5227T=DSM 107637T). It is also proposed that D. oxamicus and D. termitidis be reclassified as Cupidesulfovibrio oxamicus comb. nov. and Cupidesulfovibrio termitidis comb. nov., respectively.

Parasulfuritortus cantonensis gen. nov., sp. nov., a microaerophilic sulfur-oxidizing bacterium isolated from freshwater sediment.

A novel sulfur-oxidizing bacterium, designated strain LSR1T, was enriched and isolated from a freshwater sediment sample collected from the Pearl River in Guangzhou, PR China. The strain was an obligate chemolithoautotroph, using thiosulfate or sulfide as an electron donor and energy source. Growth of strain LSR1T was observed at 15-40 °C, pH 6.0-7.5 and NaCl concentrations of 0-1.5 %. Strain LSR1T was microaerophilic, with growth only at oxygen content less than 10 %. Anaerobic growth was also observed when using nitrate as the sole electron acceptor. The major cellular fatty acids were C16 : 0 and summed feature 3 (comprising C16 : 1 ω7c and/or C16 : 1 ω6c). The DNA G+C content of the draft genome sequence was 67.5 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain LSR1T formed a lineage within the family Thiobacillaceae, showing sequence identities of 92.87, 92.33 and 90.80 % with its closest relative genera Sulfuritortus, Annwoodia and Thiobacillus, respectively. The genome of strain LSR1T contained multiple genes encoding sulfur-oxidizing enzymes that catalyse thiosulfate and sulfide oxidation, and the gene encoding cbb 3-type cytochrome c oxidase and bd-type quinol oxidase, which enables strain LSR1T to perform sulphur oxidation under microaerophilic conditions. On the basis of phenotypic, genotypic and phylogenetic results, strain LSR1T is considered to represent a novel species of a new genus Parasulfuritortus within the family Thiobacillaceae, for which the name Parasulfuritortus cantonensis gen. nov., sp. nov. is proposed. The type strain is LSR1T (=GDMCC 1.1549=JCM 33645).

Thiomicrorhabdus sediminis sp. nov. and Thiomicrorhabdus xiamenensis sp. nov., novel sulfur-oxidizing bacteria isolated from coastal sediments and an emended description of the genus Thiomicrorhabdus.

Two novel Gram-strain-negative and rod-shaped bacteria, designated strain G1T and G2T, were isolated from sediment samples collected from the coast of Xiamen, PR China. The cells were motile by a single polar flagellum. Growth of strain G1T occurred at 10-40 °C (optimum, 30 °C), at pH 6.0-9.0 (optimum, pH 7.5) and with 5-1530 mM NaCl (optimum, 510 mM), while the temperature, pH and NaCl concentration ranges for G2T were 4-45 °C (optimum, 28 °C), pH 5.5-8.0 (optimum, pH 6.5) and 85-1530 mM NaCl (optimum, 340 mM). The two isolates were obligate chemolithoautotrophs capable of using thiosulfate, sulfide, elemental sulphur or tetrathionate as an energy source. Strain G1T used molecular oxygen or nitrite as an electron acceptor, while strain G2T used molecular oxygen as the sole electron acceptor. The dominant fatty acids of G1T and G2T were summed feature 3 (C16:1 ω7c and/or C16:1 ω6c), C16 : 0 and summed feature 8 (C18:1 ω7c and/or C18:1 ω6c). The DNA G+C content of G1T and G2T were 45.1 and 48.3 mol%, respectively. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain G1T and G2T were members of the genus Thiomicrorhabdus, and most closely related to Thiomicrorhabdus hydrogeniphila MAS2T (96.0 %) and Thiomicrorhabdus indica 13-15AT (95.4 %), respectively. The 16S rRNA gene sequence similarity between strains G1T and G2T was 95.8 %. Based on the phylogenetic, genomic and phenotypic data presented here, the isolate strains represent novel species of the genus Thiomicrorhabdus, for which the names Thiomicrorhabdus sediminis sp. nov. (type strain G1T=MCCC 1A14511T=KCTC 15841T) and Thiomicrorhabdus xiamenensis sp. nov. (type strain G2T=MCCC 1A14512T=KCTC 15842T) are proposed.

Sulfurimonas indica sp. nov., a hydrogen- and sulfur-oxidizing chemolithoautotroph isolated from a hydrothermal sulfide chimney in the Northwest Indian Ocean.

A novel mesophilic, hydrogen- and sulfur-oxidizing bacterium, designated strain NW8NT, was collected from a sulfide chimney at the deep-sea hydrothermal vent on the Carlsberg Ridge of the Northwest Indian Ocean. The cells were Gram-stain-negative, motile, short rods with a single polar flagellum. The temperature, pH and salinity ranges for growth of strain NW8NT were 4-40 °C (optimum, 33 °C), pH 4.5-7.5 (optimum, pH 5.5) and 340-680 mM NaCl (optimum, 510 mM). The isolate was an obligate chemolithoautotroph capable of growth using hydrogen, thiosulfate, sulfide or elemental sulphur as the sole energy source, carbon dioxide as the sole carbon source and molecular oxygen as the sole electron acceptor. The major cellular fatty acids of strain NW8NT were summed feature 3 (C16 : 1 ω7c and/or C16 : 1 ω6c), C16 : 0 and summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c). The total size of its genome was 2 093 492 bp and the genomic DNA G+C content was 36.9 mol%. Phylogenetic analysis based on 16S rRNA gene sequences and core genes showed that the novel isolate belonged to the genus Sulfurimonas and was most closely related to Sulfurimonas paralvinellae GO25T (97.4 % sequence identity). The average nucleotide identity and DNA-DNAhybridization values between strain NW8NT and S. paralvinellae GO25T was 77.8 and 21.1 %, respectively. Based on the phylogenetic, genomic and phenotypic data presented here, strain NW8NT represents a novel species of the genus Sulfurimonas, for which the name Sulfurimonas indica sp. nov. is proposed, with the type strain NW8NT (=MCCC 1A13988T=KTCC 15780T).

Candidatus Sulfurimonas marisnigri sp. nov. and Candidatus Sulfurimonas baltica sp. nov., thiotrophic manganese oxide reducing chemolithoautotrophs of the class Campylobacteria isolated from the pelagic redoxclines of the Black Sea and the Baltic Sea.

Species of the genus Sulfurimonas are reported and isolated from terrestrial habitats and marine sediments and water columns with steep redox gradients. Here we report on the isolation of strains SoZ1 and GD2 from the pelagic redoxcline of the Black Sea and the Baltic Sea, respectively. Both strains are gram-stain-negative and appear as short and slightly curved motile rods. The autecological preferences for growth of strain SoZ1 were 0-25°C (optimum 20°C), pH 6.5-9.0 (optimum pH 7.5-8.0) and salinity 10-40gL-1 (optimum 25gL-1). Preferences for growth of strain GD2 were 0-20°C (optimum 15°C), pH 7.0-8.0 (optimum pH 7.0-7.5) and salinity 5-40gL-1 (optimum 21gL-1). Strain SoZ1 grew chemolithoautotrophically, while strain GD2 also showed heterotrophic growth with short chained fatty acids as carbon source. Both species utilized hydrogen (H2), sulfide (H2S here taken as the sum of H2S, HS- and S2-), elemental sulfur (S0) and thiosulfate (S2O32-) as electron donors and nitrate (NO3-), oxygen (O2) and particulate manganese oxide (MnO2) as electron acceptors. Based on 16S rRNA gene sequence similarity, both strains cluster within the genus Sulfurimonas with Sulfurimonas gotlandica GD1T as the closest cultured relative species with a sequence similarity of 96.74% and 96.41% for strain SoZ1 and strain GD2, respectively. Strains SoZ1 and GD2 share a ribosomal 16S sequence similarity of 99.27% and were demarcated based on average nucleotide identity and average amino acid identity of the whole genome sequence. These calculations have been applied to the whole genus. We propose the names Candidatus Sulfurimonas marisnigri sp. nov. and Candidatus Sulfurimonas baltica sp. nov. for the thiotrophic manganese reducing culture isolates from the Black Sea and Baltic Sea, respectively.

Isachenkonia alkalipeptolytica gen. nov. sp. nov., a new anaerobic, alkaliphilic proteolytic bacterium capable of reducing Fe(III) and sulfur.

An obligately alkaliphilic, anaerobic, proteolytic bacterium was isolated from a sample of Tanatar III soda lake sediment (Altai region, Russia) and designated as strain Z-1701T. Cells of strain Z-1701T were short, straight, motile Gram-stain-positive rods. Growth of Z-1701T obligately depended on the presence of sodium carbonate. Strain Z-1701T could utilize various peptides mixtures, such as beef and yeast extracts, peptone, soytone, trypticase and tryptone, as well as such proteins as albumin, gelatin and sodium caseinate. It was able to grow oligotrophically with 0.02 g l-1 yeast extract as the sole energy and carbon source. Carbohydrates did not support the growth of strain Z-1701T. The main products released during the growth of strain Z-1701T on tryptone were formate, acetate and ammonium. Strain Z-1701T was able to reduce ferrihydrite, Fe(III)-EDTA, anthraquinone-2,6-disulfonate and elemental sulfur, using proteinaceous substrates as electron donors. In all cases the presence of the electron acceptor in the medium stimulated growth. The main cellular fatty acids were iso-C15 : 0, iso-C15 : 0 aldehyde, iso-C15 : 1 ω6, C16 : 0, iso-C17 : 0 aldehyde, C16 : 0 aldehyde and C14 : 0. The DNA G+C content of the isolate was 43.9 mol%. Phylogenetic analysis based on the concatenated alignment of 120 protein-marker sequences revealed that strain Z-1701T falls into a cluster with the genus Tindallia, family Clostridiaceae. 16S rRNA gene sequence identity between strain Z-1701T and Tindallia species were 88.3-89.75 %. On the basis of its phenotypic characteristics and phylogenetic position, the novel isolate is considered to be a representative of a novel genus and species for which the name Isachenkonia alkalipeptolytica gen. nov., sp. nov. is proposed, with Z-1701T (=JCM 32929Т=DSM 109060Т=VKM B-3261Т) as its type strain.

Analytical validation of a modified turbidimetric assay to screen sulphur oxidizing bacteria.

Conventional turbidimetric assay for sulphate determination was modified to 100 times lesser reaction volume on a convenient format using microtitre plate based platform, targeting routine microbiological applications to screen sulphur oxidizing bacteria (SOB) cultures. The modified assay was linear up to 1500 mg/L of sulphate concentration, which is about 37.5 times more than that of conventional assay. Upon regression analysis, linear equation y = 1.243× + 0.011 was obtained having R2 value of 0.998. The modified assay was fully validated in terms of precision, limit of detection (LOD), limit of quantification (LOQ), sensitivity, selectivity and robustness to assure the reliability during final applications. LOD and LOQ were found as 7.4 mg/L and 24.8 mg/L of sulphate concentration respectively. Further, accuracy of the assay over routine SOB screening media components was tested, and proved as reliable and suitable for the intended application.

Sulfuriferula nivalis sp. nov., a sulfur oxidizer isolated from snow and emended description of Sulfuriferula plumbiphila.

A chemolithoautotrophic sulfur-oxidizing bacterium, strain SGTMT was isolated from snow collected in Japan. As electron donors for growth, SGTMT oxidized thiosulfate, tetrathionate and elemental sulfur. Heterotrophic growth was not observed. Growth of the novel isolate was observed at a temperature range of 5-28 °C, with optimum growth at 18 °C. SGTMT grew at a pH range of 4.3-7.4, with optimum growth at pH 6.1-7.1. Major components in the cellular fatty acid profile were summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c) and C16 : 0. The complete genome of SGTMT consisted of a circular chromosome of approximately 3.4 Mbp and two plasmids. Phylogenetic analysis based on the 16S rRNA gene indicated that SGTMT represented a member of the genus Sulfuriferula, and its closest relative is Sulfuriferula thiophila mst6T with a sequence identity of 98 %. A comparative genome analysis showed dissimilarity between the genomes of SGTMT and S. thiophila mst6T, as low values of average nucleotide identity (74.9 %) and digital DNA-DNA hybridization (20.4%). On the basis of its genomic and phenotypic properties, SGTMT (=DSM 109609T=BCRC 81185T) is proposed as the type strain of a novel species, Sulfuriferula nivalis sp. nov. Some characteristics of another species in the same genus, Sulfuriferula plumbiphila, were also investigated to revise and supplement its description. The type strain of S. plumbiphila can grow on thiosulfate, tetrathionate and elemental sulfur. The strain showed optimum growth at pH 6.3-7.0 and shared major cellular fatty acids with the other species of the genus Sulfuriferula.

Desulfobotulus mexicanus sp. nov., a novel sulfate-reducing bacterium isolated from the sediment of an alkaline crater lake in Mexico.

A novel Gram-negative, non-spore-forming, vibrio-shaped, anaerobic, alkaliphilic, sulfate-reducing bacterium, designated strain PAR22NT, was isolated from sediment samples collected at an alkaline crater lake in Guanajuato (Mexico). Strain PAR22NT grew at temperatures between 15 and 37 °C (optimum, 32 °C), at pH between pH 8.3 and 10.1 (optimum, pH 9.0-9.6), and in the presence of NaCl up to 10 %. Pyruvate, 2-methylbutyrate and fatty acids (4-18 carbon atoms) were used as electron donors in the presence of sulfate as a terminal electron acceptor and were incompletely oxidized to acetate and CO2. Besides sulfate, both sulfite and elemental sulfur were also used as terminal electron acceptors and were reduced to sulfide. The predominant fatty acids were summed feature 10 (C18 : 1 ω7c and/or C18 : 1 ω9t and/or C18 : 1 ω12t), C18 : 1 ω9c and C16 : 0. The genome size of strain PAR22NT was 3.8 Mb including 3391 predicted genes. The genomic DNA G+C content was 49.0 mol%. Phylogenetic analysis based on 16S rRNA gene sequences showed that it belongs to the genus Desulfobotulus within the class Deltaproteobacteria. Its closest phylogenetic relatives are Desulfobotulus alkaliphilus (98.4 % similarity) and Desulfobotulus sapovorans (97.9 % similarity). Based on phylogenetic, phenotypic and chemotaxonomic characteristics, we propose that the isolate represents a novel species of the genus Desulfobotulus with the name Desulfobotulus mexicanus sp. nov. The type strain is PAR22NT (=DSM 105758T=JCM 32146T).

Sulfurimonas crateris sp. nov., a facultative anaerobic sulfur-oxidizing chemolithoautotrophic bacterium isolated from a terrestrial mud volcano.

A novel mesophilic facultative anaerobic bacterium, strain SN118T, was isolated from a terrestrial mud volcano in Taman Peninsula, Russia. The cells were Gram-negative, motile, short, straight or curved rods with a single polar flagellum. Growth was observed at 5-40 °C (optimum, 30 °C) and pH 5.5-9.5 (optimum, pH 8.0). Growth of strain SN118T was observed in NaCl concentrations ranging from 0.5 to 8.0 % (w/v) with an optimum at 2.0-3.0 % (w/v). The isolate grew chemolithoautotrophically with sulfide, elemental sulfur or thiosulfate as electron donor, oxygen, nitrate or nitrite as an electron acceptor and CO2/HCO3 - as a carbon source. Molecular hydrogen or organic substances did not support growth. Nitrate was reduced to N2. The dominant fatty acids were C16 : 1ω7c, C16 : 0 and C18  :  1ω7c. The total size of the genome of the novel isolate was 2 209 279 bp and the genomic DNA G+C content was 38.8 mol%. Results of phylogenetic analysis based on 16S rRNA gene sequences indicated that the novel isolate belonged to the genus Sulfurimonas and was most closely related to Sulfurimonas denitrificans DSM 1251T (96.74 %). Based on its physiological properties and results from phylogenetic analyses, including average nucleotide identity and in silico DNA-DNA hybridization values, the isolate is considered to represent a novel species of the genus Sulfurimonas, for which the name Sulfurimonas crateris sp. nov. is proposed. The type strain is SN118T (=DSM 109248T=VKM B-3378T).

Development of a process for microbial sulfate reduction in cold mining waters - Cold acclimation of bacterial consortia from an Arctic mining district.

Biological sulfate removal is challenging in cold climates due to the slower metabolism of mesophilic bacteria; however, cold conditions also offer the possibility to isolate bacteria that have adapted to low temperatures. The present research focused on the cold acclimation and characterization of sulfate-reducing bacterial (SRB) consortia enriched from an Arctic sediment sample from northern Finland. Based on 16S rDNA analysis, the most common sulfate-reducing bacterium in all enriched consortia was Desulfobulbus, which belongs to the δ-Proteobacteria. The majority of the cultivated consortia were able to reduce sulfate at temperatures as low as 6 °C with succinic acid as a carbon source. The sulfate reduction rates at 6 °C varied from 13 to 42 mg/L/d. The cultivation medium used in this research was a Postgate medium supplemented with lactate, ethanol or succinic acid. The obtained consortia were able to grow with lactate and succinic acid but surprisingly not with ethanol. Enriched SRB consortia are useful for the biological treatment of sulfate-containing industrial wastewaters in cold conditions.

Microbial community structure in deep natural gas-bearing aquifers subjected to sulfate-containing fluid injection.

In the natural gas field located in central Japan, high concentrations of natural gases and iodide ions are dissolved in formation water and commercially produced in deep aquifers. In the iodine recovery process, the produced formation water is amended with sulfate, and this fluid is injected into gas-bearing aquifers, which may lead to infrastructure corrosion by hydrogen sulfide. In this study, we examined the microbial community in aquifers subjected to sulfate-containing fluid injection. Formation water samples were collected from production wells located at different distances from the injection wells. The chemical analysis showed that the injection fluid contained oxygen, nitrate, nitrite and sulfate, in contrast to the formation water, which had previously been shown to be depleted in these components. Sulfur isotopic analysis indicated that sulfate derived from the injection fluid was present in the sample collected from near the injection wells. Quantitative and sequencing analysis of dissimilatory sulfite reductase and 16S rRNA genes revealed that sulfate-reducing bacteria (SRB), sulfur-oxidizing bacteria, and anaerobic methanotrophic archaea (ANME) in the wells located near injection wells were more abundant than those in wells located far from the injection wells, suggesting that fluid injection stimulated these microorganisms through the addition of oxygen, nitrate, nitrite and sulfate to the methane-rich aquifers. The predominant taxa were assigned to the ANME-2 group, its sulfate-reducing partner SEEP-SRB1 cluster and sulfur-oxidizing Epsilonproteobacteria. These results provide important insights for future studies to support the development of natural gas and iodine resources in Japan.

Petroleum hydrocarbon rich oil refinery sludge of North-East India harbours anaerobic, fermentative, sulfate-reducing, syntrophic and methanogenic microbial populations.

BACKGROUND: Sustainable management of voluminous and hazardous oily sludge produced by petroleum refineries remains a challenging problem worldwide. Characterization of microbial communities of petroleum contaminated sites has been considered as the essential prerequisite for implementation of suitable bioremediation strategies. Three petroleum refinery sludge samples from North Eastern India were analyzed using next-generation sequencing technology to explore the diversity and functional potential of inhabitant microorganisms and scope for their on-site bioremediation. RESULTS: All sludge samples were hydrocarbon rich, anaerobic and reduced with sulfate as major anion and several heavy metals. High throughput sequencing of V3-16S rRNA genes from sludge metagenomes revealed dominance of strictly anaerobic, fermentative, thermophilic, sulfate-reducing bacteria affiliated to Coprothermobacter, Fervidobacterium, Treponema, Syntrophus, Thermodesulfovibrio, Anaerolinea, Syntrophobacter, Anaerostipes, Anaerobaculum, etc., which have been well known for hydrocarbon degradation. Relatively higher proportions of archaea were detected by qPCR. Archaeal 16S rRNA gene sequences showed presence of methanogenic Methanobacterium, Methanosaeta, Thermoplasmatales, etc. Detection of known hydrocarbon utilizing aerobic/facultative anaerobic (Mycobacterium, Pseudomonas, Longilinea, Geobacter, etc.), nitrate reducing (Gordonia, Novosphigobium, etc.) and nitrogen fixing (Azovibrio, Rhodobacter, etc.) bacteria suggested niche specific guilds with aerobic, facultative anaerobic and strict anaerobic populations. Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt) predicted putative genetic repertoire of sludge microbiomes and their potential for hydrocarbon degradation; lipid-, nitrogen-, sulfur- and methane- metabolism. Methyl coenzyme M reductase A (mcrA) and dissimilatory sulfite reductase beta-subunit (dsrB) genes phylogeny confirmed methanogenic and sulfate-reducing activities within sludge environment endowed by hydrogenotrophic methanogens and sulfate-reducing Deltaproteobacteria and Firmicutes members. CONCLUSION: Refinery sludge microbiomes were comprised of hydrocarbon degrading, fermentative, sulfate-reducing, syntrophic, nitrogen fixing and methanogenic microorganisms, which were in accordance with the prevailing physicochemical nature of the samples. Analysis of functional biomarker genes ascertained the activities of methanogenic and sulfate-reducing organisms within sludge environment. Overall data provided better insights on microbial diversity and activity in oil contaminated environment, which could be exploited suitably for in situ bioremediation of refinery sludge.

[A Thermotolerant and Halotolerant Sulfate-reducing Bacterium in Produced Water from an Offshore High-temperature Oilfield in Bohai Bay, China: Isolation, Phenotypic Characterization, and Inhibition].

The growth and activity of sulfate-reducing prokaryotes (SRP) in oilfield environments could produce large amounts of H2S, leading to multifaceted problems, including oilfield souring and microbially-influenced corrosion, yet knowledge about the diversity and physiology of SRP therein was quite limited. To further understand the phenotypic characteristics of SRP residing in an offshore high-temperature oilfield at Bohai Bay, China, and to explore the potential methods for control of SRP-mediated problems, we isolated, using Hungate techniques, a thermotolerant, halotolerant SRP strain, designated BQ1, from the produced water of a high-temperature. We also presented the phenotypic features of BQ1, and investigated the efficacy of five biocides, or metabolic inhibitors, in suppressing the sulfidogenic activity of BQ1. Cells of BQ1 were motile, short rod-shaped, 1.2-2.5 µm in length and 0.5-0.8 µm in width. Although BQ1 shared 99% 16S rRNA gene sequence similarity with Desulfovibrio vulgaris Hildenborough, distinct phenotypic traits between them were observed. Isolated BQ1 could grow at 14-70℃(optimum at 30℃) and pH 6.0-9.0 (optimum pH 7.0), and in the presence of 0%-10% NaCl. Isolated BQ1 utilized a wide range of carbon substrates, including sodium formate, sodium lactate, and acetate. Sulfate, sulfite, thiosulfate, and sulfur were utilized as electron acceptors, but not nitrate or nitrite. Sodium hypochlorite (600 mg·L-1), Benzyltrimethylammonium chloride (300 mg·L-1), or nitrate (800 mg·L-1) failed to inhibit H2S production by BQ1. By contrast, glutaraldehyde (50 mg·L-1), bronopol (30 mg·L-1), chlorine dioxide (50 mg·L-1), and nitrite (70 mg·L-1) inhibited H2S production by BQ1 for at least 30 d, indicating that these compounds may be suitable for the mitigation of microbial souring in this specific, high-temperature, offshore oilfield at Bohai Bay, China.

Sulfurovum denitrificans sp. nov., an obligately chemolithoautotrophic sulfur-oxidizing epsilonproteobacterium isolated from a hydrothermal field.

A novel marine sulfur-oxidizing bacterium, designated strain eps51T, was isolated from a surface rock sample collected from the hydrothermal field of Suiyo Seamount on the Izu-Bonin Arc in the Western Pacific Ocean. This bacterium was Gram-staining-negative, non-motile and rod-shaped. Strain eps51T grew chemolithoautotrophically, by sulfur-oxidizing respiration with elemental sulfur and thiosulfate as electron donors and used only carbon dioxide as a carbon source. Oxygen and nitrate were used as its electron acceptors. The isolate grew optimally at 30 °C, at pH 7.0 and with 3 % NaCl. The predominant fatty acids were C16 : 1ω7c, C18 : 1ω7c and C16 : 0. The respiratory quinone was menaquinone-6 and the genomic DNA G+C content was 40.0 mol%. Phylogenetic analysis based on 16S rRNA gene sequence revealed that eps51T represented a member of the genus Sulfurovum and the closest relative was Sulfurovum aggregans (96.7 %). Based on its phylogenetic position along with its physiological and chemotaxonomic characteristics, the name Sulfurovum denitrificans sp. nov. is proposed, with the type strain eps51T (=NBRC 102602T=DSM 19611T).

The diversity of sulfate-reducing bacteria in the seven bioreactors.

Anaerobic technology has a wide scope of application in different areas such as manufacturing, food industry, and agriculture. Nowadays, it is mainly used to produce electrical and thermal energy from crop processing, solid waste treatment or wastewater treatment. More intensively, trend nowadays is usage of this technology biodegradable and biomass waste processing and biomethane or hydrogen production. In this paper, the diversities of sulfate-reducing bacteria (SRB) under different imputed raw material to the bioreactors were characterized. These diversities at the beginning of sampling and after cultivation were compared. Desulfovibrio, Desulfobulbus, and Desulfomicrobium genus as dominant among sulfate reducers in the bioreactors were detected. The Desulfobulbus species were dominant among other SRB genera before cultivation, but these bacteria were detected only in three out of the seven bioreactors after cultivation dominant.