Yiran Dong

Halomonas sulfidaeris-dominated microbial community inhabits a 1.8 km-deep subsurface Cambrian sandstone reservoir

A low-diversity microbial community, dominated by the γ-proteobacterium Halomonas sulfidaeris, was detected in samples of warm saline formation porewater collected from the Cambrian Mt. Simon Sandstone in the Illinois Basin of the North American Midcontinent (1.8u2009km/5872u2009ft burial depth, 50°C, pHu20098, 181 bars pressure). These highly porous and permeable quartz arenite sandstones are directly analogous to reservoirs around the world targeted for large-scale hydrocarbon extraction, as well as subsurface gas and carbon storage. A new downhole low-contamination subsurface sampling probe was used to collect in situ formation water samples for microbial environmental metagenomic analyses. Multiple lines of evidence suggest that this H.u2009sulfidaeris-dominated subsurface microbial community is indigenous and not derived from drilling mud microbial contamination. Data to support this includes V1-V3 pyrosequencing of formation water and drilling mud, as well as comparison with previously published microbial analyses of drilling muds in other sites. Metabolic pathway reconstruction, constrained by the geology, geochemistry and present-day environmental conditions of the Mt. Simon Sandstone, implies that H.u2009sulfidaeris-dominated subsurface microbial community may utilize iron and nitrogen metabolisms and extensively recycle indigenous nutrients and substrates. The presence of aromatic compound metabolic pathways suggests this microbial community can readily adapt to and survive subsurface hydrocarbon migration.

Fe-oxide grain coatings support bacterial Fe-reducing metabolisms in 1.7-2.0 km-deep subsurface quartz arenite sandstone reservoirs of the Illinois Basin (USA).

The Cambrian-age Mt. Simon Sandstone, deeply buried within the Illinois Basin of the midcontinent of North America, contains quartz sand grains ubiquitously encrusted with iron-oxide cements and dissolved ferrous iron in pore-water. Although microbial iron reduction has previously been documented in the deep terrestrial subsurface, the potential for diagenetic mineral cementation to drive microbial activity has not been well studied. In this study, two subsurface formation water samples were collected at 1.72 and 2.02 km, respectively, from the Mt. Simon Sandstone in Decatur, Illinois. Low-diversity microbial communities were detected from both horizons and were dominated by Halanaerobiales of Phylum Firmicutes. Iron-reducing enrichment cultures fed with ferric citrate were successfully established using the formation water. Phylogenetic classification identified the enriched species to be related to Vulcanibacillus from the 1.72 km depth sample, while Orenia dominated the communities at 2.02 km of burial depth. Species-specific quantitative analyses of the enriched organisms in the microbial communities suggest that they are indigenous to the Mt. Simon Sandstone. Optimal iron reduction by the 1.72 km enrichment culture occurred at a temperature of 40°C (range 20–60°C) and a salinity of 25 parts per thousand (range 25–75 ppt). This culture also mediated fermentation and nitrate reduction. In contrast, the 2.02 km enrichment culture exclusively utilized hydrogen and pyruvate as the electron donors for iron reduction, tolerated a wider range of salinities (25–200 ppt), and exhibited only minimal nitrate- and sulfate-reduction. In addition, the 2.02 km depth community actively reduces the more crystalline ferric iron minerals goethite and hematite. The results suggest evolutionary adaptation of the autochthonous microbial communities to the Mt. Simon Sandstone and carries potentially important implications for future utilization of this reservoir for CO2 injection.

Response of Soil Microbial Communities to Elevated Antimony and Arsenic Contamination Indicates the Relationship between the Innate Microbiota and Contaminant Fractions

Mining of sulfide ore deposits containing metalloids, such as antimony and arsenic, has introduced serious soil contamination around the world, posing severe threats to food safety and human health. Hence, it is important to understand the behavior and composition of the microbial communities that control the mobilization or sequestration of these metal(loid)s. Here, we selected two sites in Southwest China with different levels of Sb and As contamination to study interactions among various Sb and As fractions and the soil microbiota, with a focus on the microbial response to metalloid contamination. Comprehensive geochemical analyses and 16S rRNA gene amplicon sequencing demonstrated distinct soil taxonomic inventories depending on Sb and As contamination levels. Stochastic gradient boosting indicated that citric acid extractable Sb(V) and As(V) contributed 5% and 15%, respectively, to influencing the community diversity. Random forest predicted that low concentrations of Sb(V) and As(V) could enhance the community diversity but generally, the Sb and As contamination impairs microbial diversity. Co-occurrence network analysis indicated a strong correlation between the indigenous microbial communities and various Sb and As fractions. A number of taxa were identified as core genera due to their elevated abundances and positive correlation with contaminant fractions (total Sb and As concentrations, bioavailable Sb and As extractable fractions, and Sb and As redox species). Shotgun metagenomics indicated that Sb and As biogeochemical redox reactions may exist in contaminated soils. All these observations suggest the potential for bioremediation of Sb- and As-contaminated soils.

Orenia metallireducens sp. nov. Strain Z6, a Novel Metal-Reducing Member of the Phylum Firmicutes from the Deep Subsurface

ABSTRACT A novel halophilic and metal-reducing bacterium, Orenia metallireducens strain Z6, was isolated from briny groundwater extracted from a 2.02 km-deep borehole in the Illinois Basin, IL. This organism shared 96% 16S rRNA gene similarity with Orenia marismortui but demonstrated physiological properties previously unknown for this genus. In addition to exhibiting a fermentative metabolism typical of the genus Orenia, strain Z6 reduces various metal oxides [Fe(III), Mn(IV), Co(III), and Cr(VI)], using H2 as the electron donor. Strain Z6 actively reduced ferrihydrite over broad ranges of pH (6 to 9.6), salinity (0.4 to 3.5 M NaCl), and temperature (20 to 60°C). At pH 6.5, strain Z6 also reduced more crystalline iron oxides, such as lepidocrocite (γ-FeOOH), goethite (α-FeOOH), and hematite (α-Fe2O3). Analysis of X-ray absorption fine structure (XAFS) following Fe(III) reduction by strain Z6 revealed spectra from ferrous secondary mineral phases consistent with the precipitation of vivianite [Fe3(PO4)2] and siderite (FeCO3). The draft genome assembled for strain Z6 is 3.47 Mb in size and contains 3,269 protein-coding genes. Unlike the well-understood iron-reducing Shewanella and Geobacter species, this organism lacks the c-type cytochromes for typical Fe(III) reduction. Strain Z6 represents the first bacterial species in the genus Orenia (order Halanaerobiales) reported to reduce ferric iron minerals and other metal oxides. This microbe expands both the phylogenetic and physiological scopes of iron-reducing microorganisms known to inhabit the deep subsurface and suggests new mechanisms for microbial iron reduction. These distinctions from other Orenia spp. support the designation of strain Z6 as a new species, Orenia metallireducens sp. nov. IMPORTANCE A novel iron-reducing species, Orenia metallireducens sp. nov., strain Z6, was isolated from groundwater collected from a geological formation located 2.02 km below land surface in the Illinois Basin, USA. Phylogenetic, physiologic, and genomic analyses of strain Z6 found it to have unique properties for iron reducers, including (i) active microbial iron-reducing capacity under broad ranges of temperatures (20 to 60°C), pHs (6 to 9.6), and salinities (0.4 to 3.5 M NaCl), (ii) lack of c-type cytochromes typically affiliated with iron reduction in Geobacter and Shewanella species, and (iii) being the only member of the Halanaerobiales capable of reducing crystalline goethite and hematite. This study expands the scope of phylogenetic affiliations, metabolic capacities, and catalytic mechanisms for iron-reducing microbes.

A Combination of Stable Isotope Probing, Illumina Sequencing, and Co-occurrence Network to Investigate Thermophilic Acetate- and Lactate-Utilizing Bacteria

Anaerobic digestion is a complicated microbiological process that involves a wide diversity of microorganisms. Acetate is one of the most important intermediates, and interactions between acetate-oxidizing bacteria and archaea could play an important role in the formation of methane in anoxic environments. Anaerobic digestion at thermophilic temperatures is known to increase methane production, but the effects on the microbial community are largely unknown. In the current study, stable isotope probing was used to characterize acetate- and lactate-oxidizing bacteria in thermophilic anaerobic digestion. In microcosms fed 13C-acetate, bacteria related to members of Clostridium, Hydrogenophaga, Fervidobacterium, Spirochaeta, Limnohabitans, and Rhodococcus demonstrated elevated abundances of 13C-DNA fractions, suggesting their activities in acetate oxidation. In the treatments fed 13C-lactate, Anaeromyxobacter, Desulfobulbus, Syntrophus, Cystobacterineae, and Azospira were found to be the potential thermophilic lactate utilizers. PICRUSt predicted that enzymes related to nitrate and nitrite reduction would be enriched in 13C-DNA fractions, suggesting that the acetate and lactate oxidation may be coupled with nitrate and/or nitrite reduction. Co-occurrence network analysis indicated bacterial taxa not enriched in 13C-DNA fractions that may also play a critical role in thermophilic anaerobic digestion.

Hematite Reduction Buffers Acid Generation and Enhances Nutrient Uptake by a Fermentative Iron Reducing Bacterium, Orenia metallireducens Strain Z6

Fermentative iron-reducing organisms have been identified in a variety of environments. Instead of coupling iron reduction to respiration, they have been consistently observed to use ferric iron minerals as an electron sink for fermentation. In the present study, a fermentative iron reducer, Orenia metallireducens strain Z6, was shown to use iron reduction to enhance fermentation not only by consuming electron equivalents, but also by generating alkalinity that effectively buffers the pH. Fermentation of glucose by this organism in the presence of a ferric oxide mineral, hematite (Fe2O3), resulted in enhanced glucose decomposition compared with fermentation in the absence of an iron source. Parallel evidence (i.e., genomic reconstruction, metabolomics, thermodynamic analyses, and calculation of electron transfer) suggested hematite reduction as a proton-consuming reaction effectively consumed acid produced by fermentation. The buffering effect of hematite was further supported by a greater extent of glucose utilization by strain Z6 in media with increasing buffer capacity. Such maintenance of a stable pH through hematite reduction for enhanced glucose fermentation complements the thermodynamic interpretation of interactions between microbial iron reduction and other biogeochemical processes. This newly discovered feature of iron reducer metabolism also has significant implications for groundwater management and contaminant remediation by providing microbially mediated buffering systems for the associated microbial and/or chemical reactions.

Tepidibacillus decaturensis sp. nov., a microaerophilic, moderately thermophilic iron-reducing bacterium isolated from 1.7 km depth groundwater.

A Gram-stain-negative, microaerophilic rod-shaped organism designated as strain Z9T was isolated from groundwater of 1.7 km depth from the Mt. Simon Sandstone of the Illinois Basin, Illinois, USA. Cells of strain Z9T were rod shaped with dimensions of 0.3×(1-10) µm and stained Gram-negative. Strain Z9T grew within the temperature range 20-60u2009°C (optimum at 30-40u2009°C), between pH 5 and 8 (optimum 5.2-5.8) and under salt concentrations of 1-5u2009% (w/v) NaCl (optimum 2.5u2009% NaCl). In addition to growth by fermentation and nitrate reduction, this strain was able to reduce Fe(III), Mn(IV), Co(III) and Cr(VI) when H2 or organic carbon was available as the electron donor, but did not actively reduce oxidized sulfur compounds (e.g. sulfate, thiosulfate or S0). The G+C content of the DNA from strain Z9T was 36.1u2009mol%. Phylogenetic analysis of the 16S rRNA gene from strain Z9T showed that it belongs to the class Bacilli and shares 97u2009% sequence similarity with the only currently characterized member of the genus Tepidibacillus, T.fermentans. Based on the physiological distinctness and phylogenetic information, strain Z9T represents a novel species within the genus Tepidibacillus, for which the name Tepidibacillus decaturensis sp. nov. is proposed. The type strain is Z9T (=ATCC BAA-2644T=DSM 103037T).

Draft Genome Sequence of Tepidibacillus decaturensis Strain Z9, an Anaerobic, Moderately Thermophilic, and Heterotrophic Bacterium from the Deep Subsurface of the Illinois Basin, USA

ABSTRACT The genome of the moderately thermophilic and halotolerant bacterium Tepidibacillus decaturensis strain Z9 was sequenced. The draft genome comprises three scaffolds, for a total of 2.95 Mb. As the first sequenced genome within the genus Tepidibacillus, 2,895 protein-coding genes, 52 tRNA genes, and 3 rRNA operons were predicted.

Bacterial survival strategies in an alkaline tailing site and the physiological mechanisms of dominant phylotypes as revealed by metagenomic analyses

Microorganisms inhabiting mine tailings require specific metabolic strategies to survive, which may hold potential for pollution clean up. Effective in situ bioremediation will rely on an in-depth understanding of the function of the bacterial communities, especially the abundant and metabolically active phylotypes. In this study, the bacterial communities collected from an alkaline tailing site were profiled by 16S rRNA gene amplicon sequencing as well as shotgun metagenomic analysis. Our results indicated that potentials for carbon and nitrogen fixation as well as metal resistance and transformation were widespread among the bacterial community members, especially in highly enriched phylotypes, such as members of Thiobacillus and Meiothermus. Important functional microbial guilds including carbon and nitrogen fixers may contribute to phytoremediation by providing nutrients for hyperaccumulator plants. In addition, metal-metabolizing bacteria may influence metal speciation and solubility. This discovery provides an understanding for microbial survival strategies in the tailings and lays the foundation for future potential manipulation of the tailing microbiome for in situ bioremediation.