The low-temperature germinating spores of the thermophilic Desulfofundulus contribute to an extremely high sulfate reduction in burning coal seams.

Burning coal seams, characterized by massive carbon monoxide (CO) emissions, the presence of secondary sulfates, and high temperatures, represent suitable environments for thermophilic sulfate reduction. The diversity and activity of dissimilatory sulfate reducers in these environments remain unexplored. In this study, using metagenomic approaches, in situ activity measurements with a radioactive tracer, and cultivation we have shown that members of the genus Desulfofundulus are responsible for the extremely high sulfate reduction rate (SRR) in burning lignite seams in the Altai Mountains. The maximum SRR reached 564 ± 21.9 nmol S cm-3 day-1 at 60°C and was of the same order of magnitude for both thermophilic (60°C) and mesophilic (23°C) incubations. The 16S rRNA profiles and the search for dsr gene sequences in the metagenome revealed members of the genus Desulfofundulus as the main sulfate reducers. The thermophilic Desulfofundulus sp. strain Al36 isolated in pure culture, did not grow at temperatures below 50°C, but produced spores that germinated into metabolically active cells at 20 and 15°C. Vegetative cells germinating from spores produced up to 0.738 ± 0.026 mM H2S at 20°C and up to 0.629 ± 0.007 mM H2S at 15°C when CO was used as the sole electron donor. The Al36 strain maintains significant production of H2S from sulfate over a wide temperature range from 15°C to 65°C, which is important in variable temperature biotopes such as lignite burning seams. Burning coal seams producing CO are ubiquitous throughout the world, and biogenic H2S may represent an overlooked significant flux to the atmosphere. The thermophilic spore outgrowth and their metabolic activity at temperatures below the growth minimum may be important for other spore-forming bacteria of environmental, industrial and clinical importance.

Antibiotic-Resistant Desulfovibrio Produces H2S from Supplements for Animal Farming.

Sulphate-reducing bacteria, primarily Desulfovibrio, are responsible for the active generation of H2S in swine production waste. The model species for sulphate reduction studies, Desulfovibrio vulgaris strain L2, was previously isolated from swine manure characterized by high rates of dissimilatory sulphate reduction. The source of electron acceptors in low-sulphate swine waste for the high rate of H2S formation remains uncertain. Here, we demonstrate the ability of the L2 strain to use common animal farming supplements including L-lysine-sulphate, gypsum and gypsum plasterboards as electron acceptors for H2S production. Genome sequencing of strain L2 revealed the presence of two megaplasmids and predicted resistance to various antimicrobials and mercury, which was confirmed in physiological experiments. Most of antibiotic resistance genes (ARG) are carried by two class 1 integrons located on the chromosome and on the plasmid pDsulf-L2-2. These ARGs, predicted to confer resistance to beta-lactams, aminoglycosides, lincosamides, sulphonamides, chloramphenicol and tetracycline, were probably laterally acquired from various Gammaproteobacteria and Firmicutes. Resistance to mercury is likely enabled by two mer operons also located on the chromosome and on pDsulf-L2-2 and acquired via horizontal gene transfer. The second megaplasmid, pDsulf-L2-1, encoded nitrogenase, catalase and type III secretion system suggesting close contact of the strain with intestinal cells in the swine gut. The location of ARGs on mobile elements allows us to consider D. vulgaris strain L2 as a possible vector transferring antimicrobials resistance determinants between the gut microbiote and microbial communities in environmental biotopes.

Active Sulfate-Reducing Bacterial Community in the Camel Gut.

The diversity and activity of sulfate-reducing bacteria (SRB) in the camel gut remains largely unexplored. An abundant SRB community has been previously revealed in the feces of Bactrian camels (Camelus bactrianus). This study aims to combine the 16S rRNA gene profiling, sulfate reduction rate (SRR) measurement with a radioactive tracer, and targeted cultivation to shed light on SRB activity in the camel gut. Fresh feces of 55 domestic Bactrian camels grazing freely on semi-arid mountain pastures in the Kosh-Agach district of the Russian Altai area were analyzed. Feces were sampled in early winter at an ambient temperature of -15 °C, which prevented possible contamination. SRR values measured with a radioactive tracer in feces were relatively high and ranged from 0.018 to 0.168 nmol S cm-3 day-1. The 16S rRNA gene profiles revealed the presence of Gram-negative Desulfovibrionaceae and spore-forming Desulfotomaculaceae. Targeted isolation allowed us to obtain four pure culture isolates belonging to Desulfovibrio and Desulforamulus. An active SRB community may affect the iron and copper availability in the camel intestine due to metal ions precipitation in the form of sparingly soluble sulfides. The copper-iron sulfide, chalcopyrite (CuFeS2), was detected by X-ray diffraction in 36 out of 55 analyzed camel feces. In semi-arid areas, gypsum, like other evaporite sulfates, can be used as a solid-phase electron acceptor for sulfate reduction in the camel gastrointestinal tract.

Coexistence of Psychrophilic, Mesophilic, and Thermophilic Sulfate-Reducing Bacteria in a Deep Subsurface Aquifer Associated with Coal-Bed Methane Production.

The microbial community of subsurface environments remains understudied due to limited access to deep strata and aquifers. Coal-bed methane (CBM) production is associated with a large number of wells pumping water out of coal seams. CBM wells provide access to deep biotopes associated with coal-bed water. Temperature is one of the key constraints for the distribution and activity of subsurface microorganisms, including sulfate-reducing prokaryotes (SRP). The 16S rRNA gene amplicon sequencing coupled with in situ sulfate reduction rate (SRR) measurements with a radioactive tracer and cultivation at various temperatures revealed that the SRP community of the coal bed water of the Kuzbass coal basin is characterized by an overlapping mesophilic-psychrophilic boundary. The genus Desulfovibrio comprised a significant share of the SRP community. The D. psychrotolerans strain 1203, which has a growth optimum below 20 °C, dominated the cultivated SRP. SRR in coal bed water varied from 0.154 ± 0.07 to 2.04 ± 0.048 nmol S cm-3 day-1. Despite the ambient water temperature of ~ 10-20 °C, an active thermophilic SRP community occurred in the fracture water, which reduced sulfate with the rate of 0.159 ± 0.023 to 0.198 ± 0.007 nmol S cm-3 day-1 at 55 °C. A novel moderately thermophilic "Desulforudis audaxviator"-clade SRP has been isolated in pure culture from the coal-bed water.

Desulfovibrio desulfuricans AY5 Isolated from a Patient with Autism Spectrum Disorder Binds Iron in Low-Soluble Greigite and Pyrite.

The sulphate-reducing bacteria (SRB) of genus Desulfovibrio are a group of prokaryotes associated with autism spectrum disorders (ASD). The connection between the elevated numbers of Desulfovibrio in the gut of children with ASD compared with healthy children remains unresolved. A conceivable consequence of SRB overgrowth in the gut is the conversion of bioavailable iron into low-soluble crystalline iron sulphides, causing iron deficiency in the organism. In this study, we report the draft genome sequence and physiological features of the first cultivable isolate from a patient with ASD, Desulfovibrio desulfuricans strain AY5.The capability of the strain to produce crystalline iron sulphides was studied under different pH conditions. The most notable greigite(Fe3S4) and pyrite (FeS2) formation was revealed at pH 6.0, which suggests that the iron loss due to insoluble sulphide formation may occur in the proximal part of the gastrointestinal tract. Strain AY5 was adapted to grow under nitrogen-limiting conditions by N2 fixation. The urease found in the strain's genome may play a role in resistance to acidic pH.

Desulfosporosinus metallidurans sp. nov., an acidophilic, metal-resistant sulfate-reducing bacterium from acid mine drainage.

A novel, spore-forming, acidophilic and metal-resistant sulfate-reducing bacterium, strain OLT, was isolated from a microbial mat in a tailing dam at a gold ore mining site. Cells were slightly curved immotile rods, 0.5 µm in diameter and 2.0-3.0 µm long. Cells were stained Gram-negative, despite the Gram-positive cell structure revealed by electron microscopy of ultrathin layers. OLT grew at pH 4.0-7.0 with an optimum at 5.5. OLT utilised H2, lactate, pyruvate, malate, formate, propionate, ethanol, glycerol, glucose, fructose, sucrose, peptone and tryptone as electron donors for sulfate reduction. Sulfate, sulfite, thiosulfate, nitrate and fumarate were used as electron acceptors in the presence of lactate. Elemental sulfur, iron (III), and arsenate did not serve as electron acceptors. The major cellular fatty acids were C16:1ω7c (39.0 %) and C16 : 0 (12.1 %). The draft genome of OLT was 5.29 Mb in size and contained 4909 protein-coding genes. The 16S rRNA gene sequence placed OLT within the phylum Firmicutes, class Clostridia, family Peptococcaceae, genus Desulfosporosinus. Desulfosporosinus nitroreducens 59.4BT was the closest relative with 97.6 % sequence similarity. On the basis of phenotypic and phylogenetic characteristics, strain OLT represents a novel species within the genus Desulfosporosinus, for which we propose the name Desulfosporosinus metallidurans sp. nov. with the type strain OLT (=DSM 104464T=VKM В-3021T).

Isolation, characterization, and genome insights into an anaerobic sulfidogenic Tissierella bacterium from Cu-bearing coins.

Recent reports on antimicrobial effects of metallic Cu prompted this study of anaerobic microbial communities on copper surfaces. Widely circulating copper-containing coinage was used as a potential source for microorganisms that had had human contact and were tolerant to copper. This study reports on the isolation, characterization, and genome of an anaerobic sulfidogenic Tissierella sp. P1from copper-containing brass coinage. Dissimilatory (bi)sulfite reductase dsrAB present in strain P1 genome and the visible absorbance around 630 nm in the cells suggested the presence of a desulfoviridin-type protein. However, the sulfate reduction rate measurements with 35SO42- did not confirm the dissimilatory sulfate reduction by the strain. The P1 genome lacks APS reductase, sulfate adenylyltransferase, DsrC, and DsrMK necessary for dissimilatory sulfate reduction. The isolate produced up to 0.79 mM H2S during growth, possibly due to cysteine synthase (CysK) and/or cysteine desulfhydrase (CdsH) activities, encoded in the genome. The strain can tolerate up to 2.4 mM Cu2+(150 mg/l) in liquid medium, shows affinity to metallic copper, and can survive on copper-containing coins up to three days under ambient air and dry conditions. The genome sequence of strain P1 contained cutC, encoding a copper resistance protein, which distinguishes it from all other Tissierella strains with published genomes.

Isolation, Characterization, and Metal Response of Novel, Acid-Tolerant Penicillium spp. from Extremely Metal-Rich Waters at a Mining Site in Transbaikal (Siberia, Russia).

The role of fungi in metal cycling in acidic environments has been little explored to date. In this study, two acid-tolerant and metal-resistant Penicillium isolates, strains ShG4B and ShG4C, were isolated from a mine site in the Transbaikal area of Siberia (Russia). Waters at the mine site were characterized by extremely high metal concentrations: up to 18 g l-1 Fe and > 2 g l-1 each of Cu, Zn, Al, and As. Both isolates were identified as Penicillium spp. by phylogenetic analyses and they grew well in Czapek medium acidified to pH 2.5. Resistance to Cu, Cd, Ni, Co, and arsenate was in the range of 1-10 g l-1. Further experiments with Penicillium strain ShG4C demonstrated that growth in Cu-containing media was accompanied by the precipitation of Cu-oxalate (moolooite) and the formation of extracellular vesicles enriched in Cu on the mycelia. Vesicles were greatly reduced in size in Cd-containing media and were not formed in the presence of Ni or Co. Cd-oxalate was detected as a crystalline solid phase in Cd-exposed mycelia. Hydrated Ni-sulfate (retgersite) and Co-sulfate (bieberite) were detected in mycelia grown in the presence of Ni and Co, respectively. The results demonstrated that acid-tolerant and metal-resistant Penicillium constitute a component in extremophilic microbiomes, contributing to organic matter breakdown and formation of secondary solid phases at pH ranges found in acid rock drainage.

Selection for novel, acid-tolerant Desulfovibrio spp. from a closed Transbaikal mine site in a temporal pH-gradient bioreactor.

Almost all the known isolates of acidophilic or acid-tolerant sulphate-reducing bacteria (SRB) belong to the spore-forming genus Desulfosporosinus in the Firmicutes. The objective of this study was to isolate acidophilic/acid-tolerant members of the genus Desulfovibrio belonging to deltaproteobacterial SRB. The sample material originated from microbial mat biomass submerged in mine water and was enriched for sulphate reducers by cultivation in anaerobic medium with lactate as an electron donor. A stirred tank bioreactor with the same medium composition was inoculated with the sulphidogenic enrichment. The bioreactor was operated with a temporal pH gradient, changing daily, from an initial pH of 7.3 to a final pH of 3.7. Among the bacteria in the bioreactor culture, Desulfovibrio was the only SRB group retrieved from the bioreactor consortium as observed by 16S rRNA-targeted denaturing gradient gel electrophoresis. Moderately acidophilic/acid-tolerant isolates belonged to Desulfovibrio aerotolerans-Desulfovibrio carbinophilus-Desulfovibrio magneticus and Desulfovibrio idahonensis-Desulfovibrio mexicanus clades within the genus Desulfovibrio. A moderately acidophilic strain, Desulfovibrio sp. VK (pH optimum 5.7) and acid-tolerant Desulfovibrio sp. ED (pH optimum 6.6) dominated in the bioreactor consortium at different time points and were isolated in pure culture.

Genome sequence of the acid-tolerant Desulfovibrio sp. DV isolated from the sediments of a Pb-Zn mine tailings dam in the Chita region, Russia.

Here we report the draft genome sequence of the acid-tolerant Desulfovibrio sp. DV isolated from the sediments of a Pb-Zn mine tailings dam in the Chita region, Russia. The draft genome has a size of 4.9 Mb and encodes multiple K+-transporters and proton-consuming decarboxylases. The phylogenetic analysis based on concatenated ribosomal proteins revealed that strain DV clusters together with the acid-tolerant Desulfovibrio sp. TomC and Desulfovibrio magneticus. The draft genome sequence and annotation have been deposited at GenBank under the accession number MLBG00000000.

Genome sequence of the copper resistant and acid-tolerant Desulfosporosinus sp. BG isolated from the tailings of a molybdenum-tungsten mine in the Transbaikal area.

Here, we report on the draft genome of a copper-resistant and acidophilic Desulfosporosinus sp. BG, isolated from the tailings of a molybdenum-tungsten mine in Transbaikal area. The draft genome has a size of 4.52 Mb and encodes transporters of heavy metals. The phylogenetic analysis based on concatenated ribosomal proteins revealed that strain BG clusters together with the other acidophilic copper-resistant strains Desulfosporosinus sp. OT and Desulfosporosinus sp. I2. The K+-ATPase, Na+/H+ antiporter and amino acid decarboxylases may participate in enabling growth at low pH. The draft genome sequence and annotation have been deposited at GenBank under the accession number NZ_MASS00000000.

Genomic insights into a new acidophilic, copper-resistant Desulfosporosinus isolate from the oxidized tailings area of an abandoned gold mine.

Microbial sulfate reduction in acid mine drainage is still considered to be confined to anoxic conditions, although several reports have shown that sulfate-reducing bacteria occur under microaerophilic or aerobic conditions. We have measured sulfate reduction rates of up to 60 nmol S cm(-3) day(-1) in oxidized layers of gold mine tailings in Kuzbass (SW Siberia). A novel, acidophilic, copper-tolerant Desulfosporosinus sp. I2 was isolated from the same sample and its genome was sequenced. The genomic analysis and physiological data indicate the involvement of transporters and additional mechanisms to tolerate metals, such as sequestration by polyphosphates. Desulfosporinus sp. I2 encodes systems for a metabolically versatile life style. The genome possessed a complete Embden-Meyerhof pathway for glycolysis and gluconeogenesis. Complete oxidation of organic substrates could be enabled by the complete TCA cycle. Genomic analysis found all major components of the electron transfer chain necessary for energy generation via oxidative phosphorylation. Autotrophic CO2 fixation could be performed through the Wood-Ljungdahl pathway. Multiple oxygen detoxification systems were identified in the genome. Taking into account the metabolic activity and genomic analysis, the traits of the novel isolate broaden our understanding of active sulfate reduction and associated metabolism beyond strictly anaerobic niches.

Characterization and Genome Analysis of the First Facultatively Alkaliphilic Thermodesulfovibrio Isolated from the Deep Terrestrial Subsurface.

Members of the genus Thermodesulfovibrio belong to the Nitrospirae phylum and all isolates characterized to date are neutrophiles. They have been isolated from terrestrial hot springs and thermophilic methanogenic anaerobic sludges. Their molecular signatures have, however, also been detected in deep subsurface. The purpose of this study was to characterize and analyze the genome of a newly isolated, facultatively alkaliphilic Thermodesulfovibrio from a 2 km deep aquifer system in Western Siberia, Russia. The new isolate, designated N1, grows optimally at pH 8.5 and at 65°C. It is able to reduce sulfate, thiosulfate or sulfite with a limited range of electron donors, such as formate, pyruvate, and lactate. Analysis of the 1.93 Mb draft genome of strain N1 revealed that it contains a set of genes for dissimilatory sulfate reduction, including sulfate adenyltransferase, adenosine-5'-phosphosulfate reductase AprAB, membrane-bound electron transfer complex QmoABC, dissimilatory sulfite reductase DsrABC, and sulfite reductase-associated electron transfer complex DsrMKJOP. Hydrogen turnover is enabled by soluble cytoplasmic, membrane-linked, and soluble periplasmic hydrogenases. The use of thiosulfate as an electron acceptor is enabled by a membrane-linked molybdopterin oxidoreductase. The N1 requirement for organic carbon sources corresponds to the lack of the autotrophic C1-fixation pathways. Comparative analysis of the genomes of Thermodesulfovibrio (T. yellowstonii, T. islandicus, T. àggregans, T. thiophilus, and strain N1) revealed a low overall genetic diversity and several adaptive traits. Consistent with an alkaliphilic lifestyle, a multisubunit Na+/H+ antiporter of the Mnh family is encoded in the Thermodesulfovibrio strain N1 genome. Nitrogenase genes were found in T. yellowstonii, T. aggregans, and T. islandicus, nitrate reductase in T. islandicus, and cellulose synthetase in T. aggregans and strain N1. Overall, our results provide genomic insights into metabolism of the Thermodesulfovibrio lineage in microbial communities of the deep subsurface biosphere.

Draft genome sequence of the first acid-tolerant sulfate-reducing deltaproteobacterium Desulfovibrio sp. TomC having potential for minewater treatment.

The sulfidogenic bacterium Desulfovibrio sp. TomC was isolated from acidic waste at the abandoned gold ore mining site in the Martaiga gold ore belt, Western Siberia. This bacterium, being the first reported acid-tolerant gram-negative sulfate-reducer of the order Deltaproteobacteria, is able to grow at pH as low as 2.5 and is resistant to high concentrations of metals. The draft 5.3 Mb genome sequence of Desulfovibrio sp. TomC has been established and provides the genetic basis for application of this microorganism in bioreactors and other bioremediation schemes for the treatment of metal-containing wastewater.