Fermentation, Hydrogen, and Sulfur Metabolism in Multiple Uncultivated Bacterial Phyla

Wrighton, Kelly; Thomas, Brian C.; Sharon, Itai; Miller, Christopher S.; Castelle, Cindy J.; VerBerkmoes, Nathan C.; Wilkins, Michael J.; Hettich, Robert L.; Lipton, Mary; Williams, Kenneth H.; Long, Philip E.; Banfield, Jillian F.

doi:10.1126/science.1224041

Cited by 607 publications

(766 citation statements)

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“…Until recently, recovering genomes from metagenomic data was restricted to samples with low microbial diversity 13 , but improved sequencing throughput and advances in computational techniques now allow metagenome-assembled genomes (MAGs) to be recovered from high diversity environments 14,15 . MAGs are obtained by grouping or 'binning' together assembled contigs with similar sequence composition, depth of coverage across one or more related samples and taxonomic affiliations 16,17 .…”

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confidence: 99%

Recovery of nearly 8,000 metagenome-assembled genomes substantially expands the tree of life

et al. 2017

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Challenges in cultivating microorganisms have limited the phylogenetic diversity of currently available microbial genomes. This is being addressed by advances in sequencing throughput and computational techniques that allow for the cultivation-independent recovery of genomes from metagenomes. Here, we report the reconstruction of 7,903 bacterial and archaeal genomes from > 1,500 public metagenomes. All genomes are estimated to be ≥ 50% complete and nearly half are ≥ 90% complete with ≤ 5% contamination. These genomes increase the phylogenetic diversity of bacterial and archaeal genome trees by > 30% and provide the first representatives of 17 bacterial and three archaeal candidate phyla. We also recovered 245 genomes from the Patescibacteria superphylum (also known as the Candidate Phyla Radiation) and find that the relative diversity of this group varies substantially with different protein marker sets. The scale and quality of this data set demonstrate that recovering genomes from metagenomes provides an expedient path forward to exploring microbial dark matter. Articles NATuRe MiCRobiologyform the UBA data set as they met our filtering criteria of having an estimated quality ≥ 50 (defined as the estimated completeness of a genome minus five times its estimated contamination) and consisting of ≤ 500 scaffolds with an N50 ≥ 10 kb ( Fig. 1 and Supplementary Table 2). Over 93% of the 7,903 UBA genomes have an average coverage of ≥ 10× (5th percentile, 9.2× , 95th percentile, 268× ) and 95.8% have > 5× coverage over 90% of bases, providing assurance of high-quality base-calling across the genomes 3,36 . Among the UBA genomes is a subset of 3,438 near-complete genomes (3,225 bacterial and 213 archaeal) estimated to be ≥ 90% complete with ≤ 5% contamination (Fig. 1a). These genomes consist of ≤ 100 scaffolds in 70.2% of cases (≤ 200 scaffolds in 92.0% genomes) and have an average N50 of 136 kb. Comparison of near-complete UBA genomes that are conspecific strains of complete isolate genomes also suggest that the recovered MAGs have no systematic loss of genomic content, with the exception of extrachromosomal elements such as plasmids (Supplementary Note 1).The UBA data set was also assessed relative to the criteria used by the Human Microbiome Project (HMP) for defining high-quality draft genomes 3,37 . Of the 3,438 UBA genomes we have defined as near complete, 3,201 (93.1%) pass all of the HMP criteria, with the only substantial exception being 4.8% of the genomes having scaffolds with an N50 of < 20 kb (Supplementary Table 3). Nearly half of the remaining 4,465 UBA genomes also pass the HMP criteria for being high quality except that they are estimated to be < 90% complete.The presence of tRNAs for the standard 20 amino acids was examined as a secondary measure of genome quality (Fig. 1c). The 3,438 near-complete UBA genomes have tRNAs that encode for an average of 17.3 ± 2.2 of the 20 amino acids and ≥ 15 amino acids in 90.3% of the genomes. The correlation between estimated genome completeness and identified tRN...

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confidence: 99%

Recovery of nearly 8,000 metagenome-assembled genomes substantially expands the tree of life

et al. 2017

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“…Scaffold coverage was calculated by mapping reads back to the assemblies using Bowtie2 (ref. 37). Given the dominance and high strain variation in some samples, highly abundant genomes (>400×) often failed to assemble.…”

Section: Methodsmentioning

confidence: 99%

“…All scaffolds ≥5 kb (≥1 kb for subassemblies, Methanohalophilus-1, and the GB enrichment culture) were included when binning genomes from the metagenomic assembly. Scaffolds were annotated as described previously 36,37 by predicting open reading frames using MetaProdigal 39 . Sequences were compared using USEARCH 40 to KEGG, UniRef90 and InterproScan 41 with single and reverse best hit (RBH) matches greater than 60 bits reported.…”

Section: Methodsmentioning

confidence: 99%

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Microbial metabolisms in a 2.5-km-deep ecosystem created by hydraulic fracturing in shales

et al. 2016

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Hydraulic fracturing is the industry standard for extracting hydrocarbons from shale formations. Attention has been paid to the economic benefits and environmental impacts of this process, yet the biogeochemical changes induced in the deep subsurface are poorly understood. Recent single-gene investigations revealed that halotolerant microbial communities were enriched after hydraulic fracturing. Here, the reconstruction of 31 unique genomes coupled to metabolite data from the Marcellus and Utica shales revealed that many of the persisting organisms play roles in methylamine cycling, ultimately supporting methanogenesis in the deep biosphere. Fermentation of injected chemical additives also sustains long-term microbial persistence, while thiosulfate reduction could produce sulfide, contributing to reservoir souring and infrastructure corrosion. Extensive links between viruses and microbial hosts demonstrate active viral predation, which may contribute to the release of labile cellular constituents into the extracellular environment. Our analyses show that hydraulic fracturing provides the organismal and chemical inputs for colonization and persistence in the deep terrestrial subsurface. S hale gas accounts for one-third of natural gas energy resources worldwide. It has been estimated that shale gas will provide half of the natural gas in the USA, annually, by 2040, with the Marcellus shale in the Appalachian Basin projected to produce three times more than any other formation 1 . Recovery of these hydrocarbons is dependent on hydraulic fracturing technologies, where the high-pressure injection of water and chemical additives generates extensive fractures in the shale matrix. Hydrocarbons trapped in tiny pore spaces are subsequently released and collected at the wellpad surface, together with a portion of the injected fluids that have reacted with the shale formation. The mixture of injected fluids and hydrocarbons collected is referred to as 'produced fluids'.Microbial metabolism and growth in hydrocarbon reservoirs has both positive and negative impacts on energy recovery. Whereas stimulation of methanogens in coal beds enhances energy recovery 2 , bacterial hydrogen sulfide production ('reservoir souring') decreases profits and contributes to corrosion and the risk of environmental contamination 3 . Additionally, biomass accumulation within newly generated fractures may reduce their permeability, decreasing natural gas recovery. Despite these potential microbial impacts, little is known about the function and activity of microorganisms in hydraulically fractured shale.Initial work by our group and others 4-9 used single marker gene analyses to identify microorganisms from several geographically distinct shale formations. These analyses showed similar halotolerant taxa in produced fluids several months after hydraulic fracturing. To assign functional roles to these organisms, we conducted metagenomic and metabolite analyses on input and produced fluids up to a year after hydraulic fracturing (HF) from two Appala...

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“…This bias partly results from our inability to cultivate most microbes 2 , which is a necessary step for traditional whole-genome sequencing. Through cultivationindependent approaches, namely the assembly and binning of metagenome shotgun sequencing data 3,4 and single-cell genomics (SCG) 5,6 , one can now access the genetic makeup of uncultivated microbes. Both methods bypass the conventional cultivation step, and they can be applied directly to environmental samples.…”

Section: Introductionmentioning

confidence: 99%

Obtaining genomes from uncultivated environmental microorganisms using FACS–based single-cell genomics

et al. 2014

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single-cell genomics is a powerful tool for exploring the genetic makeup of environmental microorganisms, the vast majority of which are difficult, if not impossible, to cultivate with current approaches. Here we present a comprehensive protocol for obtaining genomes from uncultivated environmental microbes via high-throughput single-cell isolation by Facs. the protocol encompasses the preservation and pretreatment of differing environmental samples, followed by the physical separation, lysis, whole-genome amplification and 16s rrna-based identification of individual bacterial and archaeal cells. the described procedure can be performed with standard molecular biology equipment and a Facs machine. It takes <12 h of bench time over a 4-d time period, and it generates up to 1 mg of genomic Dna from an individual microbial cell, which is suitable for downstream applications such as pcr amplification and shotgun sequencing. the completeness of the recovered genomes varies, with an average of ~50%.

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Fermentation, Hydrogen, and Sulfur Metabolism in Multiple Uncultivated Bacterial Phyla

Cited by 607 publications

References 58 publications

Recovery of nearly 8,000 metagenome-assembled genomes substantially expands the tree of life

Recovery of nearly 8,000 metagenome-assembled genomes substantially expands the tree of life

Microbial metabolisms in a 2.5-km-deep ecosystem created by hydraulic fracturing in shales

Obtaining genomes from uncultivated environmental microorganisms using FACS–based single-cell genomics

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