Multimodal metagenomic analysis reveals microbial single nucleotide variants as superior biomarkers for early detection of colorectal cancer.

Microbial signatures show remarkable potentials in predicting colorectal cancer (CRC). This study aimed to evaluate the diagnostic powers of multimodal microbial signatures, multi-kingdom species, genes, and single-nucleotide variants (SNVs) for detecting precancerous adenomas. We performed cross-cohort analyses on whole metagenome sequencing data of 750 samples via xMarkerFinder to identify adenoma-associated microbial multimodal signatures. Our data revealed that fungal species outperformed species from other kingdoms with an area under the ROC curve (AUC) of 0.71 in distinguishing adenomas from controls. The microbial SNVs, including dark SNVs with synonymous mutations, displayed the strongest diagnostic capability with an AUC value of 0.89, sensitivity of 0.79, specificity of 0.85, and Matthews correlation coefficient (MCC) of 0.74. SNV biomarkers also exhibited outstanding performances in three independent validation cohorts (AUCs = 0.83, 0.82, 0.76; sensitivity = 1.0, 0.72, 0.93; specificity = 0.67, 0.81, 0.67, MCCs = 0.69, 0.83, 0.72) with high disease specificity for adenoma. In further support of the above results, functional analyses revealed more frequent inter-kingdom associations between bacteria and fungi, and abnormalities in quorum sensing, purine and butanoate metabolism in adenoma, which were validated in a newly recruited cohort via qRT-PCR. Therefore, these data extend our understanding of adenoma-associated multimodal alterations in the gut microbiome and provide a rationale of microbial SNVs for the early detection of CRC.

Pathways of Iron and Sulfur Acquisition, Cofactor Assembly, Destination, and Storage in Diverse Archaeal Methanogens and Alkanotrophs.

Archaeal methanogens, methanotrophs, and alkanotrophs have a high demand for iron (Fe) and sulfur (S); however, little is known of how they acquire, traffic, deploy, and store these elements. Here, we examined the distribution of homologs of proteins mediating key steps in Fe/S metabolism in model microorganisms, including iron(II) sensing/uptake (FeoAB), sulfide extraction from cysteine (SufS), and the biosynthesis of iron-sulfur [Fe-S] clusters (SufBCDE), siroheme (Pch2 dehydrogenase), protoheme (AhbABCD), cytochrome c (Cyt c) (CcmCF), and iron storage/detoxification (Bfr, FtrA, and IssA), among 326 publicly available, complete or metagenome-assembled genomes of archaeal methanogens/methanotrophs/alkanotrophs. The results indicate several prevalent but nonuniversal features, including FeoB, SufBC, and the biosynthetic apparatus for the basic tetrapyrrole scaffold, as well as its siroheme (and F430) derivatives. However, several early-diverging genomes lacked SufS and pathways to synthesize and deploy heme. Genomes encoding complete versus incomplete heme biosynthetic pathways exhibited equivalent prevalences of [Fe-S] cluster binding proteins, suggesting an expansion of catalytic capabilities rather than substitution of heme for [Fe-S] in the former group. Several strains with heme binding proteins lacked heme biosynthesis capabilities, while other strains with siroheme biosynthesis capability lacked homologs of known siroheme binding proteins, indicating heme auxotrophy and unknown siroheme biochemistry, respectively. While ferritin proteins involved in ferric oxide storage were widespread, those involved in storing Fe as thioferrate were unevenly distributed. Collectively, the results suggest that differences in the mechanisms of Fe and S acquisition, deployment, and storage have accompanied the diversification of methanogens/methanotrophs/alkanotrophs, possibly in response to differential availability of these elements as these organisms evolved. IMPORTANCE Archaeal methanogens, methanotrophs, and alkanotrophs, argued to be among the most ancient forms of life, have a high demand for iron (Fe) and sulfur (S) for cofactor biosynthesis, among other uses. Here, using comparative bioinformatic approaches applied to 326 genomes, we show that major differences in Fe/S acquisition, trafficking, deployment, and storage exist in this group. Variation in these characters was generally congruent with the phylogenetic placement of these genomes, indicating that variation in Fe/S usage and deployment has contributed to the diversification and ecology of these organisms. However, incongruency was observed among the distribution of cofactor biosynthesis pathways and known protein destinations for those cofactors, suggesting auxotrophy or yet-to-be-discovered pathways for cofactor biosynthesis.

Conexivisphaera calida gen. nov., sp. nov., a thermophilic sulfur- and iron-reducing archaeon, and proposal of Conexivisphaeraceae fam. nov., Conexivisphaerales ord. nov., and Conexivisphaeria class. nov. in the phylum Thaumarchaeota.

A thermoacidophilic, anaerobic, and iron- and sulfur-reducing archaeon, strain NAS-02T, was isolated from a terrestrial hot spring in Japan, as previously reported. This organism is the first non-ammonia-oxidizing isolate in the phylum Thaumarchaeota. Here, we propose Conexivisphaera calida gen. nov., sp. nov. to accommodate this strain. The type strain of the type species is NAS-02T (=JCM 31663T=DSM 105898T). The values of 16S rRNA gene similarity and average amino acid identity between NAS-02T and its closest relatives are <86 and <42 %, respectively. Based on the phylogeny and physiology, we propose the family Conexivisphaeraceae fam. nov., the order Conexivisphaerales ord. nov. and the class Conexivisphaeria class. nov. to accommodate the novel genus.

Young «oil site¼ of the Uzon Caldera as a habitat for unique microbial life.

BACKGROUND: The Uzon Caldera is one of the places on our planet with unique geological, ecological, and microbiological characteristics. Uzon oil is the youngest on Earth. Uzon oil has unique composition, with low proportion of heavy fractions and relatively high content of saturated hydrocarbons. Microbial communities of the «oil site¼ have a diverse composition and live at high temperatures (up to 97 °C), significant oscillations of Eh and pH, and high content of sulfur, sulfides, arsenic, antimony, and mercury in water and rocks. RESULTS: The study analyzed the composition, structure and unique genetics characteristics of the microbial communities of the oil site, analyzed the metabolic pathways in the communities. Metabolic pathways of hydrocarbon degradation by microorganisms have been found. The study found statistically significant relationships between geochemical parameters, taxonomic composition and the completeness of metabolic pathways. It was demonstrated that geochemical parameters determine the structure and metabolic potential of microbial communities. CONCLUSIONS: There were statistically significant relationships between geochemical parameters, taxonomic composition, and the completeness of metabolic pathways. It was demonstrated that geochemical parameters define the structure and metabolic potential of microbial communities. Metabolic pathways of hydrocarbon oxidation was found to prevail in the studied communities, which corroborates the hypothesis on abiogenic synthesis of Uzon hydrothermal petroleum.

Altered Gut Archaea Composition and Interaction With Bacteria Are Associated With Colorectal Cancer.

BACKGROUND & AIMS: Changes in the intestinal microbiota have been associated with development and progression of colorectal cancer (CRC). Archaea are stable components of the microbiota, but little is known about their composition or contribution to colorectal carcinogenesis. We analyzed archaea in fecal microbiomes of 2 large cohorts of patients with CRC. METHODS: We performed shotgun metagenomic analyses of fecal samples from 585 participants (184 patients with CRC, 197 patients with adenomas, and 204 healthy individuals) from discovery (165 individuals) and validation (420 individuals) cohorts. Assignment of taxonomies was performed by exact k-mer alignment against an integrated microbial reference genome database. RESULTS: Principal component analysis of archaeomes showed distinct clusters in fecal samples from patients with CRC, patients with adenomas, and control individuals (P < .001), indicating an alteration in the composition of enteric archaea during tumorigenesis. Fecal samples from patients with CRC had significant enrichment of halophilic and depletion of methanogenic archaea. The halophilic Natrinema sp. J7-2 increased progressively in samples from control individuals, to patients with adenomas, to patients with CRC. Abundances of 9 archaea species that were enriched in fecal samples from patients with CRC distinguished them from control individuals with areas under the receiver operating characteristic curve of 0.82 in the discovery cohort and 0.83 in the validation cohort. An association between archaea and bacteria diversities was observed in fecal samples from control individuals but not from patients with CRC. Archaea that were enriched in fecal samples from patients with CRC had an extensive mutual association with bacteria that were enriched in the same samples and exclusivity with bacteria that were lost from these samples. CONCLUSIONS: Archaeomes of fecal samples from patients with CRC are characterized by enrichment of halophiles and depletion of methanogens. Studies are needed to determine whether associations between specific archaea and bacteria species in samples from patients with CRC contribute to or are a response to colorectal tumorigenesis.

The Microbiome in Cystic Fibrosis Pulmonary Disease.

Cystic fibrosis (CF) is a genetic disease with mutational changes leading to profound dysbiosis, both pulmonary and intestinal, from a very young age. This dysbiosis plays an important role in clinical manifestations, particularly in the lungs, affected by chronic infection. The range of microbiological tools has recently been enriched by metagenomics based on next-generation sequencing (NGS). Currently applied essentially in a gene-targeted manner, metagenomics has enabled very exhaustive description of bacterial communities in the CF lung niche and, to a lesser extent, the fungi. Aided by progress in bioinformatics, this now makes it possible to envisage shotgun sequencing and opens the door to other areas of the microbial world, the virome, and the archaeome, for which almost everything remains to be described in cystic fibrosis. Paradoxically, applying NGS in microbiology has seen a rebirth of bacterial culture, but in an extended manner (culturomics), which has proved to be a perfectly complementary approach to NGS. Animal models have also proved indispensable for validating microbiome pathophysiological hypotheses. Description of pathological microbiomes and correlation with clinical status and therapeutics (antibiotic therapy, cystic fibrosis transmembrane conductance regulator (CFTR) modulators) revealed the richness of microbiome data, enabling description of predictive and follow-up biomarkers. Although monogenic, CF is a multifactorial disease, and both genotype and microbiome profiles are crucial interconnected factors in disease progression. Microbiome-genome interactions are thus important to decipher.

Anaerobic and hydrogenogenic carbon monoxide-oxidizing prokaryotes: Versatile microbial conversion of a toxic gas into an available energy.

Carbon monoxide (CO) is a gas that is toxic to various organisms including humans and even microbes; however, it has low redox potential, which can fuel certain microbes, namely, CO oxidizers. Hydrogenogenic CO oxidizers utilize an energy conservation system via a CO dehydrogenase/energy-converting hydrogenase complex to produce hydrogen gas, a zero emission fuel, by CO oxidation coupled with proton reduction. Biochemical and molecular biological studies using a few model organisms have revealed their enzymatic reactions and transcriptional response mechanisms using CO. Biotechnological studies for CO-dependent hydrogen production have also been carried out with these model organisms. In this chapter, we review recent advances in the studies of these microbes, which reveal their unique and versatile metabolic profiles and provides future perspectives on ecological roles and biotechnological applications. Over the past decade, the number of isolates has doubled (37 isolates in 5 phyla, 20 genera, and 32 species). Some of the recently isolated ones show broad specificity to electron acceptors. Moreover, accumulating genomic information predicts their unique physiologies and reveals their phylogenomic relationships with novel potential hydrogenogenic CO oxidizers. Combined with genomic database surveys, a molecular ecological study has unveiled the wide distribution and low abundance of these microbes. Finally, recent biotechnological applications of hydrogenogenic CO oxidizers have been achieved via diverse approaches (e.g., metabolic engineering and co-cultivation), and the identification of thermophilic facultative anaerobic CO oxidizers will promote industrial applications as oxygen-tolerant biocatalysts for efficient hydrogen production by genomic engineering.

Densely Populated Water Droplets in Heavy-Oil Seeps.

Most of the microbial degradation in oil reservoirs is believed to take place at the oil-water transition zone (OWTZ). However, a recent study indicates that there is microbial life enclosed in microliter-sized water droplets dispersed in heavy oil of Pitch Lake in Trinidad and Tobago. This life in oil suggests that microbial degradation of oil also takes place in water pockets in the oil-bearing rock of an oil leg independent of the OWTZ. However, it is unknown whether microbial life in water droplets dispersed in oil is a generic property of oil reservoirs rather than an exotic exception. Hence, we took samples from three heavy-oil seeps, Pitch Lake (Trinidad and Tobago), the La Brea Tar Pits (California, USA), and an oil seep on the McKittrick oil field (California, USA). All three tested oil seeps contained dispersed water droplets. Larger droplets between 1 and 10 µl revealed high cell densities of up to 109 cells ml-1 Testing for ATP content and LIVE/DEAD staining showed that these populations consist of active and viable microbial cells with an average of 60% membrane-intact cells and ATP concentrations comparable to those of other subsurface ecosystems. Microbial community analyses based on 16S rRNA gene amplicon sequencing revealed the presence of known anaerobic oil-degrading microorganisms. Surprisingly, the community analyses showed similarities between all three oil seeps, revealing common OTUs, although the sampling sites were thousands of kilometers apart. Our results indicate that small water inclusions are densely populated microhabitats in heavy oil and possibly a generic trait of degraded-oil reservoirs.IMPORTANCE Our results confirmed that small water droplets in oil are densely populated microhabitats containing active microbial communities. Since these microhabitats occurred in three tested oil seeps which are located thousands of kilometers away from each other, such populated water droplets might be a generic trait of biodegraded oil reservoirs and might be involved in the overall oil degradation process. Microbial degradation might thus also take place in water pockets in the oil-bearing oil legs of the reservoir rock rather than only at the oil-water transition zone.

Bacterial and archeal dynamics of a labscale HYBRID gas fermentation bioreactor fed with CO2 and H2.

16s rDNA-based methods were used in order to identify the dynamics of microbial profiles in a HYBRID gas fermentation bio-methanization reactor. The effects of various H2 and CO2 ratios on microbial community were investigated. The HYBRID gas fermentation reactor was composed of granular anaerobic seed and the system fed with only H2 and CO2 gases. No additional organic material and trace element was fed during the throughout the experiments; thus, the microbial diversity was directly related to production of methane. The dynamics of the microbial communities were investigated with DGGE and real-time PCR analysis. The results showed that Methanobacteriales members were more dominated than Methanosarcinales and Methanomicrobiales members in the system. DGGE results indicated that Methanosaeta concilii, Methanoculleus sp., Methanosphaerula palustris, Methanofollis formosanus, Methanolinea sp., and Methanobacterium palustre were the most prominent methanogens depending on different H2/CO2 ratios. DGGE profiles suggested that hydrogenotrophic and acetoclastic species were responsible for the production of methane. The survival of syntrophic bacteria and acetoclastic methanogens was attributed to their utilization of organic materials provided by lysis. To the best of our knowledge, this is the first microbial profile detection study in a hybrid bioreactor system operated with only pure hydrogen and carbon dioxide.

Expansion of Thaumarchaeota habitat range is correlated with horizontal transfer of ATPase operons.

Thaumarchaeota are responsible for a significant fraction of ammonia oxidation in the oceans and in soils that range from alkaline to acidic. However, the adaptive mechanisms underpinning their habitat expansion remain poorly understood. Here we show that expansion into acidic soils and the high pressures of the hadopelagic zone of the oceans is tightly linked to the acquisition of a variant of the energy-yielding ATPases via horizontal transfer. Whereas the ATPase genealogy of neutrophilic Thaumarchaeota is congruent with their organismal genealogy inferred from concatenated conserved proteins, a common clade of V-type ATPases unites phylogenetically distinct clades of acidophilic/acid-tolerant and piezophilic/piezotolerant species. A presumptive function of pumping cytoplasmic protons at low pH is consistent with the experimentally observed increased expression of the V-ATPase in an acid-tolerant thaumarchaeote at low pH. Consistently, heterologous expression of the thaumarchaeotal V-ATPase significantly increased the growth rate of E. coli at low pH. Its adaptive significance to growth in ocean trenches may relate to pressure-related changes in membrane structure in which this complex molecular machine must function. Together, our findings reveal that the habitat expansion of Thaumarchaeota is tightly correlated with extensive horizontal transfer of atp operons.

Isolation and characterization of a thermophilic sulfur- and iron-reducing thaumarchaeote from a terrestrial acidic hot spring.

A deep-branching clade of Thaumarchaeota, conventionally called Terrestrial hot spring creanarchaeotic group (THSCG), is a missing link between thaumarchaeotic ammonia oxidizers and the deeper-branching non-ammonia oxidizers, such as Crenarchaeota and Candidatus Korarchaeota. Here, we report isolation of the first cultivated representative from the THSCG, named as NAS-02. Physiological characterization demonstrated that the isolate was a thermoacidophilic, sulfur- and iron-reducing organoheterotroph, which was supported by gene contents encoded in its complete genome. There was no evidence for ammonia oxidation by the isolate. Members in THSCG are likely thermophiles, and may play roles in degrading cell debris as a scavenger and in biogeochemical cycling of sulfur and iron in the hot environments, as suggested by the physiological characteristics of the isolate and the geographical distribution of the 16S rRNA gene sequences of THSCG in terrestrial hot springs and marine hydrothermal fields. Phylogenetic analysis suggests that the THSCG lineage represented by NAS-02 has gained the ability of sulfur reduction via horizontal gene transfer. Based on the phylogeny and physiology, we propose the name Conexivisphaera calidus gen. nov., sp. nov. to accommodate the isolate.

Iron requirements and uptake strategies of the globally abundant marine ammonia-oxidising archaeon, Nitrosopumilus maritimus SCM1.

Ammonia-oxidising archaea (AOA) mediate the rate-limiting step of nitrification, the central component of the marine nitrogen cycle that converts ammonia to nitrite then nitrate. Competition with phytoplankton for ammonium and light inhibition are considered to restrict AOA activity to below the photic zone, but observations of surface nitrification now demand a further understanding of the factors driving AOA distribution and activity. Pico- to nanomolar concentrations of iron (Fe) limit the growth of microorganisms in a significant portion of the world's surface oceans, yet there is no examination of the role of Fe in AOA growth despite the process of ammonia oxidation being considered to rely on the micronutrient. Here we investigate the Fe requirements and Fe uptake strategies of the Nitrosopumilus maritimus strain SCM1, a strain representative of globally abundant marine AOA. Using trace metal clean culturing techniques, we found that N. maritimus growth is determined by Fe availability, displaying a free inorganic Fe (Fe') half saturation constant 1-2 orders of magnitude greater for cell growth than numerous marine phytoplankton and heterotrophic bacterial species driven by a reduced affinity for Fe'. In addition, we discovered that whilst unable to produce siderophores to enhance access to Fe, N. maritimus is able to use the exogenous siderophore desferrioxamine B (DFB), likely through a reductive uptake pathway analogous to that demonstrated in phytoplankton. Our work suggests AOA growth in surface waters may be Fe limited and advances our understanding of AOA physiology on the cellular and mechanistic levels with implications for ecosystem dynamics and the biogeochemical N-cycle.

Dynamics of Microbial Populations Responsible for Biodegradation during the Full-Scale Treatment of Palm Oil Mill Effluent.

Despite efforts to address the composition of the microbial community during the anaerobic treatment of palm oil mill effluent (POME), its composition in relation to biodegradation in the full-scale treatment system has not yet been extensively examined. Therefore, a thorough analysis of bacterial and archaeal communities was performed in the present study using MiSeq sequencing at the different stages of the POME treatment, which comprised anaerobic as well as facultative anaerobic and aerobic processes, including the mixed raw effluent (MRE), mixing pond, holding tank, and final discharge phases. Based on the results obtained, the following biodegradation processes were suggested to occur at the different treatment stages: (1) Lactobacillaceae (35.9%) dominated the first stage, which contributed to high lactic acid production; (2) the higher population of Clostridiaceae in the mixing pond (47.7%) and Prevotellaceae in the holding tank (49.7%) promoted acetic acid production; (3) the aceticlastic methanogen Methanosaetaceae (0.6-0.8%) played a role in acetic acid degradation in the open digester and closed reactor for methane generation; (4) Syntrophomonas (21.5-29.2%) appeared to be involved in the degradation of fatty acids and acetic acid by syntrophic cooperation with the hydrogenotrophic methanogen, Methanobacteriaceae (0.6-1.3%); and (5) the phenols and alcohols detected in the early phases, but not in the final discharge phase, indicated the successful degradation of lignocellulosic materials. The present results contribute to a better understanding of the biodegradation mechanisms involved in the different stages of the full-scale treatment of POME.

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.

Outstanding impact of soil tillage on the abundance of soil hydrolases revealed by a metagenomic approach

ABSTRACT The soil represents the main source of novel biocatalysts and biomolecules of industrial relevance. We searched for hydrolases in silico in four shotgun metagenomes (4,079,223 sequences) obtained in a 13-year field trial carried out in southern Brazil, under the no-tillage (NT), or conventional tillage (CT) managements, with crop succession (CS, soybean/wheat), or crop rotation (CR, soybean/maize/wheat/lupine/oat). We identified 42,631 hydrolases belonging to five classes by comparing with the KEGG database, and 44,928 sequences by comparing with the NCBI-NR database. The abundance followed the order: lipases > laccases > cellulases > proteases > amylases > pectinases. Statistically significant differences were attributed to the tillage system, with the NT showing about five times more hydrolases than the CT system. The outstanding differences can be attributed to the management of crop residues, left on the soil surface in the NT, and mechanically broken and incorporated into the soil in the CT. Differences between the CS and the CR were slighter, 10% higher for the CS, but not statistically different. Most of the sequences belonged to fungi (Verticillium, and Colletotrichum for lipases and laccases, and Aspergillus for proteases), and to the archaea Sulfolobus acidocaldarius for amylases. Our results indicate that agricultural soils under conservative managements may represent a hotspot for bioprospection of hydrolases.

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.

Archaea diversity in vegetation gradients from the Brazilian Cerrado

Abstract We used 16S rRNA sequencing to assess the archaeal communities across a gradient of Cerrado. The archaeal communities differed across the gradient. Crenarcheota was the most abundant phyla, with Nitrosphaerales and NRPJ as the predominant classes. Euryachaeota was also found across the Cerrado gradient, including the classes Metanocellales and Methanomassiliicoccaceae.

Microbial community composition of a hydrocarbon reservoir 40 years after a CO2 enhanced oil recovery flood.

Injecting CO2 into depleted oil reservoirs to extract additional crude oil is a common enhanced oil recovery (CO2-EOR) technique. However, little is known about how in situ microbial communities may be impacted by CO2 flooding, or if any permanent microbiological changes occur after flooding has ceased. Formation water was collected from an oil field that was flooded for CO2-EOR in the 1980s, including samples from areas affected by or outside of the flood region, to determine the impacts of CO2-EOR on reservoir microbial communities. Archaea, specifically methanogens, were more abundant than bacteria in all samples, while identified bacteria exhibited much greater diversity than the archaea. Microbial communities in CO2-impacted and non-impacted samples did not significantly differ (ANOSIM: Statistic R = -0.2597, significance = 0.769). However, several low abundance bacteria were found to be significantly associated with the CO2-affected group; very few of these species are known to metabolize CO2 or are associated with CO2-rich habitats. Although this study had limitations, on a broad scale, either the CO2 flood did not impact the microbial community composition of the target formation, or microbial communities in affected wells may have reverted back to pre-injection conditions over the ca. 40 years since the CO2-EOR.

Harnessing a methane-fueled, sediment-free mixed microbial community for utilization of distributed sources of natural gas.

Harnessing the metabolic potential of uncultured microbial communities is a compelling opportunity for the biotechnology industry, an approach that would vastly expand the portfolio of usable feedstocks. Methane is particularly promising because it is abundant and energy-rich, yet the most efficient methane-activating metabolic pathways involve mixed communities of anaerobic methanotrophic archaea and sulfate reducing bacteria. These communities oxidize methane at high catabolic efficiency and produce chemically reduced by-products at a comparable rate and in near-stoichiometric proportion to methane consumption. These reduced compounds can be used for feedstock and downstream chemical production, and at the production rates observed in situ they are an appealing, cost-effective prospect. Notably, the microbial constituents responsible for this bioconversion are most prominent in select deep-sea sediments, and while they can be kept active at surface pressures, they have not yet been cultured in the lab. In an industrial capacity, deep-sea sediments could be periodically recovered and replenished, but the associated technical challenges and substantial costs make this an untenable approach for full-scale operations. In this study, we present a novel method for incorporating methanotrophic communities into bioindustrial processes through abstraction onto low mass, easily transportable carbon cloth artificial substrates. Using Gulf of Mexico methane seep sediment as inoculum, optimal physicochemical parameters were established for methane-oxidizing, sulfide-generating mesocosm incubations. Metabolic activity required >∼40% seawater salinity, peaking at 100% salinity and 35 °C. Microbial communities were successfully transferred to a carbon cloth substrate, and rates of methane-dependent sulfide production increased more than threefold per unit volume. Phylogenetic analyses indicated that carbon cloth-based communities were substantially streamlined and were dominated by Desulfotomaculum geothermicum. Fluorescence in situ hybridization microscopy with carbon cloth fibers revealed a novel spatial arrangement of anaerobic methanotrophs and sulfate reducing bacteria suggestive of an electronic coupling enabled by the artificial substrate. This system: 1) enables a more targeted manipulation of methane-activating microbial communities using a low-mass and sediment-free substrate; 2) holds promise for the simultaneous consumption of a strong greenhouse gas and the generation of usable downstream products; and 3) furthers the broader adoption of uncultured, mixed microbial communities for biotechnological use.

Metabolic capability and in situ activity of microorganisms in an oil reservoir.

BACKGROUND: Microorganisms have long been associated with oxic and anoxic degradation of hydrocarbons in oil reservoirs and oil production facilities. While we can readily determine the abundance of microorganisms in the reservoir and study their activity in the laboratory, it has been challenging to resolve what microbes are actively participating in crude oil degradation in situ and to gain insight into what metabolic pathways they deploy. RESULTS: Here, we describe the metabolic potential and in situ activity of microbial communities obtained from the Jiangsu Oil Reservoir (China) by an integrated metagenomics and metatranscriptomics approach. Almost complete genome sequences obtained by differential binning highlight the distinct capability of different community members to degrade hydrocarbons under oxic or anoxic condition. Transcriptomic data delineate active members of the community and give insights that Acinetobacter species completely oxidize alkanes into carbon dioxide with the involvement of oxygen, and Archaeoglobus species mainly ferment alkanes to generate acetate which could be consumed by Methanosaeta species. Furthermore, nutritional requirements based on amino acid and vitamin auxotrophies suggest a complex network of interactions and dependencies among active community members that go beyond classical syntrophic exchanges; this network defines community composition and microbial ecology in oil reservoirs undergoing secondary recovery. CONCLUSION: Our data expand current knowledge of the metabolic potential and role in hydrocarbon metabolism of individual members of thermophilic microbial communities from an oil reservoir. The study also reveals potential metabolic exchanges based on vitamin and amino acid auxotrophies indicating the presence of complex network of interactions between microbial taxa within the community.