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Faculty & Staff

  • Image of Karthik Anantharaman

    Karthik Anantharaman

    Assistant Professor of Bacteriology

    4550 Microbial Sciences Building
    Office: (608) 265-4537
    Lab: (608) 265-4307

Start and Promotion Dates

  • Assistant Professor: 2018


B.Tech., Civil Engineering, National Institute of Technology, Karnataka, India. 2007
M.S.E., Civil and Environmental Engineering, University of Michigan. 2008
Ph.D., Earth and Environmental Sciences, University of Michigan. 2014
Postdoctoral Research: University of California-Berkeley

Areas of Study

Microbial sulfur metabolism
Microbial community interactions
Coevolution of viruses and their hosts

Research Overview

Microbial metabolism has the potential to impact the evolutionary ecology of a system across various spatial and temporal scales ranging from the scope of a single cell, ecosystem, to the earth as a whole. Understanding how microbial communities function is critical to unraveling how they underpin human health, and predicting global ecosystem dynamics, especially in the context of environmental perturbations like anthropogenic global change. The broad goal of our research is to unravel how (i) fine-scale genomic diversity impacts microbial community structure and function across various environments, (ii) individual microbes function in a community context, and (iii) microbes impact biogeochemical cycling at various spatial and temporal scales.

Evolutionary ecology of microbial sulfur metabolism. Microorganisms control and modulate transformations associated with the element sulfur in natural and engineered systems. Sulfur plays a central role in biochemistry, impacts carbon and nitrogen turnover in various environments, and is critical to maintaining the health of oceans in the future. We use biotic sulfur transformations as a model to study the evolution and ecology of microbial energy metabolism. These processes are abundant across both aerobic (sulfur oxidation) and anaerobic environments (sulfate reduction). We utilize a combination of fieldwork, laboratory experiments, and multi-omics based approaches to investigate the microbiology of sulfureous environments such as deep-sea hydrothermal vents, freshwater ecosystems, and the human gut.

Microbial community interactions. Recent advances in DNA sequencing and bioinformatics approaches have enabled the recovery of thousands of strain-resolved microbial genomes from a single ecosystem thereby providing a window into fine scale microbial interactions and metabolic networks in complex communities. Broadly, we are interested in studying three types of interactions in microbial communities using sulfur transformations as a model – virus-microbe, microbe-microbe, and microbial “metabolic handoffs”. We focus on virus-microbe interactions involving “auxiliary metabolic genes”, which are host-derived genes utilized by viruses in selfishly altering microbial metabolism. We also study microbe-microbe interactions, and “metabolic handoffs” primarily focused on dissimilatory sulfur metabolism. We seek to quantify and predict the impact of such interactions on biogeochemical cycling at cellular, ecosystem, and global scales.

Lab Personnel

Picture of Adams
Alyssa Adams
Picture of Breister
Adam Breister
Research Intern
Picture of Cowley
Elise Cowley
Grad Student
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Jacob Hotvedt
Grad Student
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Kristopher Kieft
Grad Student
Picture of Klier
Katherine Klier
Grad Student
Picture of Langwig
Marguerite Langwig
Grad Student
Picture of Tran
Patricia Tran
Grad Student
Picture of Zhou
Chao (Zhichao) Zhou

Research Papers

  • Tran PQ, Anantharaman K (2021) Biogeochemistry Goes Viral: towards a Multifaceted Approach To Study Viruses and Biogeochemical Cycling. mSystems :e0113821 · Pubmed · DOI

    No abstract available.

  • Kieft K, Breister AM, Huss P, Linz AM, Zanetakos E, Zhou Z, Rahlff J, Esser SP, Probst AJ, Raman S, Roux S, Anantharaman K (2021) Virus-associated organosulfur metabolism in human and environmental systems. Cell reports 36((5)):109471 · Pubmed · DOI

    No abstract available.

  • Kieft K, Zhou Z, Anderson RE, Buchan A, Campbell BJ, Hallam SJ, Hess M, Sullivan MB, Walsh DA, Roux S, Anantharaman K (2021) Ecology of inorganic sulfur auxiliary metabolism in widespread bacteriophages. Nature communications 12((1)):3503 PMC8190135 · Pubmed · DOI

    No abstract available.

  • Mangalea MR, Paez-Espino D, Kieft K, Chatterjee A, Chriswell ME, Seifert JA, Feser ML, Demoruelle MK, Sakatos A, Anantharaman K, Deane KD, Kuhn KA, Holers VM, Duerkop BA (2021) Individuals at risk for rheumatoid arthritis harbor differential intestinal bacteriophage communities with distinct metabolic potential. Cell host & microbe 29((5)):726-739.e5 PMC8186507 · Pubmed · DOI

    No abstract available.

  • Doden HL, Wolf PG, Gaskins HR, Anantharaman K, Alves JMP, Ridlon JM (2021) Completion of the gut microbial epi-bile acid pathway. Gut microbes 13((1)):1-20 PMC8096331 · Pubmed · DOI

    No abstract available.

  • Tran PQ, Bachand SC, McIntyre PB, Kraemer BM, Vadeboncoeur Y, Kimirei IA, Tamatamah R, McMahon KD, Anantharaman K (2021) Depth-discrete metagenomics reveals the roles of microbes in biogeochemical cycling in the tropical freshwater Lake Tanganyika. The ISME journal 15((7)):1971-1986 PMC8245535 · Pubmed · DOI

    No abstract available.

  • Murray AE, Freudenstein J, Gribaldo S, Hatzenpichler R, Hugenholtz P, Kämpfer P, Konstantinidis KT, Lane CE, Papke RT, Parks DH, Rossello-Mora R, Stott MB, Sutcliffe IC, Thrash JC, Venter SN, Whitman WB, Acinas SG, Amann RI, Anantharaman K, Armengaud J, Baker BJ, Barco RA, Bode HB, Boyd ES, Brady CL, Carini P, Chain PSG, Colman DR, DeAngelis KM, de Los Rios MA, Estrada-de Los Santos P, Dunlap CA, Eisen JA, Emerson D, Ettema TJG, Eveillard D, Girguis PR, Hentschel U, Hollibaugh JT, Hug LA, Inskeep WP, Ivanova EP, Klenk HP, Li WJ, Lloyd KG, Löffler FE, Makhalanyane TP, Moser DP, Nunoura T, Palmer M, Parro V, Pedrós-Alió C, Probst AJ, Smits THM, Steen AD, Steenkamp ET, Spang A, Stewart FJ, Tiedje JM, Vandamme P, Wagner M, Wang FP, Yarza P, Hedlund BP, Reysenbach AL (2020) Author Correction: Roadmap for naming uncultivated Archaea and Bacteria. Nature microbiology 6((1)):136 PMC7752755 · Pubmed · DOI

    No abstract available.

  • Zhou Z, Liu Y, Pan J, Cron BR, Toner BM, Anantharaman K, Breier JA, Dick GJ, Li M (2020) Gammaproteobacteria mediating utilization of methyl-, sulfur- and petroleum organic compounds in deep ocean hydrothermal plumes. The ISME journal 14((12)):3136-3148 PMC7784996 · Pubmed · DOI

    No abstract available.

  • McDaniel EA, Peterson BD, Stevens SLR, Tran PQ, Anantharaman K, McMahon KD (2020) Expanded Phylogenetic Diversity and Metabolic Flexibility of Mercury-Methylating Microorganisms. mSystems 5((4)): PMC7438021 · Pubmed · DOI

    No abstract available.

  • Kieft K, Zhou Z, Anantharaman K (2020) VIBRANT: automated recovery, annotation and curation of microbial viruses, and evaluation of viral community function from genomic sequences. Microbiome 8((1)):90 PMC7288430 · Pubmed · DOI

    No abstract available.

  • Murray AE, Freudenstein J, Gribaldo S, Hatzenpichler R, Hugenholtz P, Kämpfer P, Konstantinidis KT, Lane CE, Papke RT, Parks DH, Rossello-Mora R, Stott MB, Sutcliffe IC, Thrash JC, Venter SN, Whitman WB, Acinas SG, Amann RI, Anantharaman K, Armengaud J, Baker BJ, Barco RA, Bode HB, Boyd ES, Brady CL, Carini P, Chain PSG, Colman DR, DeAngelis KM, de Los Rios MA, Estrada-de Los Santos P, Dunlap CA, Eisen JA, Emerson D, Ettema TJG, Eveillard D, Girguis PR, Hentschel U, Hollibaugh JT, Hug LA, Inskeep WP, Ivanova EP, Klenk HP, Li WJ, Lloyd KG, Löffler FE, Makhalanyane TP, Moser DP, Nunoura T, Palmer M, Parro V, Pedrós-Alió C, Probst AJ, Smits THM, Steen AD, Steenkamp ET, Spang A, Stewart FJ, Tiedje JM, Vandamme P, Wagner M, Wang FP, Yarza P, Hedlund BP, Reysenbach AL (2020) Roadmap for naming uncultivated Archaea and Bacteria. Nature microbiology 5((8)):987-994 PMC7381421 · Pubmed · DOI

    No abstract available.

  • Zhou Z, Tran PQ, Kieft K, Anantharaman K (2020) Genome diversification in globally distributed novel marine Proteobacteria is linked to environmental adaptation. The ISME journal 14((8)):2060-2077 PMC7367891 · Pubmed · DOI

    No abstract available.

  • Chen LX, Anantharaman K, Shaiber A, Eren AM, Banfield JF (2020) Accurate and complete genomes from metagenomes. Genome research 30((3)):315-333 PMC7111523 · Pubmed · DOI

    No abstract available.

  • Al-Shayeb B, Sachdeva R, Chen LX, Ward F, Munk P, Devoto A, Castelle CJ, Olm MR, Bouma-Gregson K, Amano Y, He C, Méheust R, Brooks B, Thomas A, Lavy A, Matheus-Carnevali P, Sun C, Goltsman DSA, Borton MA, Sharrar A, Jaffe AL, Nelson TC, Kantor R, Keren R, Lane KR, Farag IF, Lei S, Finstad K, Amundson R, Anantharaman K, Zhou J, Probst AJ, Power ME, Tringe SG, Li WJ, Wrighton K, Harrison S, Morowitz M, Relman DA, Doudna JA, Lehours AC, Warren L, Cate JHD, Santini JM, Banfield JF (2020) Clades of huge phages from across Earth's ecosystems. Nature 578((7795)):425-431 PMC7162821 · Pubmed · DOI

    No abstract available.

  • Diamond S, Andeer PF, Li Z, Crits-Christoph A, Burstein D, Anantharaman K, Lane KR, Thomas BC, Pan C, Northen TR, Banfield JF (2019) Mediterranean grassland soil C-N compound turnover is dependent on rainfall and depth, and is mediated by genomically divergent microorganisms. Nature microbiology 4((8)):1356-1367 PMC6784897 · Pubmed · DOI

    Soil microbial activity drives the carbon and nitrogen cycles and is an important determinant of atmospheric trace gas turnover, yet most soils are dominated by microorganisms with unknown metabolic capacities. Even Acidobacteria, among the most abundant bacteria in soil, remain poorly characterized, and functions across groups such as Verrucomicrobia, Gemmatimonadetes, Chloroflexi and Rokubacteria are understudied. Here, we have resolved 60 metagenomic and 20 proteomic data sets from a Mediterranean grassland soil ecosystem and recovered 793 near-complete microbial genomes from 18 phyla, representing around one-third of all microorganisms detected. Importantly, this enabled extensive genomics-based metabolic predictions for these communities. Acidobacteria from multiple previously unstudied classes have genomes that encode large enzyme complements for complex carbohydrate degradation. Alternatively, most microorganisms encode carbohydrate esterases that strip readily accessible methyl and acetyl groups from polymers like pectin and xylan, forming methanol and acetate, the availability of which could explain the high prevalence of C metabolism and acetate utilization in genomes. Microorganism abundances among samples collected at three soil depths and under natural and amended rainfall regimes indicate statistically higher associations of inorganic nitrogen metabolism and carbon degradation in deep and shallow soils, respectively. This partitioning decreased in samples under extended spring rainfall, indicating that long-term climate alteration can affect both carbon and nitrogen cycling. Overall, by leveraging natural and experimental gradients with genome-resolved metabolic profiles, we link microorganisms lacking prior genomic characterization to specific roles in complex carbon, C, nitrate and ammonia transformations, and constrain factors that impact their distributions in soil.

  • Matheus Carnevali PB, Schulz F, Castelle CJ, Kantor RS, Shih PM, Sharon I, Santini JM, Olm MR, Amano Y, Thomas BC, Anantharaman K, Burstein D, Becraft ED, Stepanauskas R, Woyke T, Banfield JF (2019) Author Correction: Hydrogen-based metabolism as an ancestral trait in lineages sibling to the Cyanobacteria. Nature communications 10((1)):1451 PMC6435703 · Pubmed · DOI

    The original version of this Article contained errors in Fig. 4. In panel a, the labels 'F420-reducing NiFe hydrogenase (group 3a)' and 'Group 2 NiFe hydrogenase' were misplaced. These errors have been corrected in both the PDF and HTML versions of the Article.

  • Bouma-Gregson K, Olm MR, Probst AJ, Anantharaman K, Power ME, Banfield JF (2019) Impacts of microbial assemblage and environmental conditions on the distribution of anatoxin-a producing cyanobacteria within a river network. The ISME journal 13((6)):1618-1634 PMC6776057 · Pubmed · DOI

    Blooms of planktonic cyanobacteria have long been of concern in lakes, but more recently, harmful impacts of riverine benthic cyanobacterial mats been recognized. As yet, we know little about how various benthic cyanobacteria are distributed in river networks, or how environmental conditions or other associated microbes in their consortia affect their biosynthetic capacities. We performed metagenomic sequencing for 22 Oscillatoriales-dominated (Cyanobacteria) microbial mats collected across the Eel River network in Northern California and investigated factors associated with anatoxin-a producing cyanobacteria. All microbial communities were dominated by one or two cyanobacterial species, so the key mat metabolisms involve oxygenic photosynthesis and carbon oxidation. Only a few metabolisms fueled the growth of the mat communities, with little evidence for anaerobic metabolic pathways. We genomically defined four cyanobacterial species, all which shared <96% average nucleotide identity with reference Oscillatoriales genomes and are potentially novel species in the genus Microcoleus. One of the Microcoleus species contained the anatoxin-a biosynthesis genes, and we describe the first anatoxin-a gene cluster from the Microcoleus clade within Oscillatoriales. Occurrence of these four Microcoleus species in the watershed was correlated with total dissolved nitrogen and phosphorus concentrations, and the species that contains the anatoxin-a gene cluster was found in sites with higher nitrogen concentrations. Microbial assemblages in mat samples with the anatoxin-a gene cluster consistently had a lower abundance of Burkholderiales (Betaproteobacteria) species than did mats without the anatoxin-producing genes. The associations of water nutrient concentrations and certain co-occurring microbes with anatoxin-a producing Microcoleus motivate further exploration for their roles as potential controls on the distributions of toxigenic benthic cyanobacteria in river networks.

  • Devoto AE, Santini JM, Olm MR, Anantharaman K, Munk P, Tung J, Archie EA, Turnbaugh PJ, Seed KD, Blekhman R, Aarestrup FM, Thomas BC, Banfield JF (2019) Megaphages infect Prevotella and variants are widespread in gut microbiomes. Nature microbiology 4((4)):693-700 PMC6784885 · Pubmed · DOI

    Bacteriophages (phages) dramatically shape microbial community composition, redistribute nutrients via host lysis and drive evolution through horizontal gene transfer. Despite their importance, much remains to be learned about phages in the human microbiome. We investigated the gut microbiomes of humans from Bangladesh and Tanzania, two African baboon social groups and Danish pigs; many of these microbiomes contain phages belonging to a clade with genomes >540 kilobases in length, the largest yet reported in the human microbiome and close to the maximum size ever reported for phages. We refer to these as Lak phages. CRISPR spacer targeting indicates that Lak phages infect bacteria of the genus Prevotella. We manually curated to completion 15 distinct Lak phage genomes recovered from metagenomes. The genomes display several interesting features, including use of an alternative genetic code, large intergenic regions that are highly expressed and up to 35 putative transfer RNAs, some of which contain enigmatic introns. Different individuals have distinct phage genotypes, and shifts in variant frequencies over consecutive sampling days reflect changes in the relative abundance of phage subpopulations. Recent homologous recombination has resulted in extensive genome admixture of nine baboon Lak phage populations. We infer that Lak phages are widespread in gut communities that contain the Prevotella species, and conclude that megaphages, with fascinating and underexplored biology, may be common but largely overlooked components of human and animal gut microbiomes.

  • Matheus Carnevali PB, Schulz F, Castelle CJ, Kantor RS, Shih PM, Sharon I, Santini JM, Olm MR, Amano Y, Thomas BC, Anantharaman K, Burstein D, Becraft ED, Stepanauskas R, Woyke T, Banfield JF (2019) Hydrogen-based metabolism as an ancestral trait in lineages sibling to the Cyanobacteria. Nature communications 10((1)):463 PMC6349859 · Pubmed · DOI

    The evolution of aerobic respiration was likely linked to the origins of oxygenic Cyanobacteria. Close phylogenetic neighbors to Cyanobacteria, such as Margulisbacteria (RBX-1 and ZB3), Saganbacteria (WOR-1), Melainabacteria and Sericytochromatia, may constrain the metabolic platform in which aerobic respiration arose. Here, we analyze genomic sequences and predict that sediment-associated Margulisbacteria have a fermentation-based metabolism featuring a variety of hydrogenases, a streamlined nitrogenase, and electron bifurcating complexes involved in cycling of reducing equivalents. The genomes of ocean-associated Margulisbacteria encode an electron transport chain that may support aerobic growth. Some Saganbacteria genomes encode various hydrogenases, and others may be able to use O under certain conditions via a putative novel type of heme copper O reductase. Similarly, Melainabacteria have diverse energy metabolisms and are capable of fermentation and aerobic or anaerobic respiration. The ancestor of all these groups may have been an anaerobe in which fermentation and H metabolism were central metabolic features. The ability to use O as a terminal electron acceptor must have been subsequently acquired by these lineages.

  • Linz AM, He S, Stevens SLR, Anantharaman K, Rohwer RR, Malmstrom RR, Bertilsson S, McMahon KD (2018) Freshwater carbon and nutrient cycles revealed through reconstructed population genomes. PeerJ 6:e6075 (PMC6292386) · Pubmed · DOI

    Although microbes mediate much of the biogeochemical cycling in freshwater, the categories of carbon and nutrients currently used in models of freshwater biogeochemical cycling are too broad to be relevant on a microbial scale. One way to improve these models is to incorporate microbial data. Here, we analyze both genes and genomes from three metagenomic time series and propose specific roles for microbial taxa in freshwater biogeochemical cycles. Our metagenomic time series span multiple years and originate from a eutrophic lake (Lake Mendota) and a humic lake (Trout Bog Lake) with contrasting water chemistry. Our analysis highlights the role of polyamines in the nitrogen cycle, the diversity of diazotrophs between lake types, the balance of assimilatory vs. dissimilatory sulfate reduction in freshwater, the various associations between types of phototrophy and carbon fixation, and the density and diversity of glycoside hydrolases in freshwater microbes. We also investigated aspects of central metabolism such as hydrogen metabolism, oxidative phosphorylation, methylotrophy, and sugar degradation. Finally, by analyzing the dynamics over time in nitrogen fixation genes and genomes, we show that the potential for nitrogen fixation is linked to specific populations in Lake Mendota. This work represents an important step towards incorporating microbial data into ecosystem models and provides a better understanding of how microbes may participate in freshwater biogeochemical cycling.

  • Castelle CJ, Brown CT, Anantharaman K, Probst AJ, Huang RH, Banfield JF (2018) Biosynthetic capacity, metabolic variety and unusual biology in the CPR and DPANN radiations. Nat. Rev. Microbiol. 16(10):629-645 · Pubmed · DOI

    Candidate phyla radiation (CPR) bacteria and DPANN (an acronym of the names of the first included phyla) archaea are massive radiations of organisms that are widely distributed across Earth's environments, yet we know little about them. Initial indications are that they are consistently distinct from essentially all other bacteria and archaea owing to their small cell and genome sizes, limited metabolic capacities and often episymbiotic associations with other bacteria and archaea. In this Analysis, we investigate their biology and variations in metabolic capacities by analysis of approximately 1,000 genomes reconstructed from several metagenomics-based studies. We find that they are not monolithic in terms of metabolism but rather harbour a diversity of capacities consistent with a range of lifestyles and degrees of dependence on other organisms. Notably, however, certain CPR and DPANN groups seem to have exceedingly minimal biosynthetic capacities, whereas others could potentially be free living. Understanding of these microorganisms is important from the perspective of evolutionary studies and because their interactions with other organisms are likely to shape natural microbiome function.

  • Anantharaman K, Hausmann B, Jungbluth SP, Kantor RS, Lavy A, Warren LA, Rappé MS, Pester M, Loy A, Thomas BC, Banfield JF (2018) Expanded diversity of microbial groups that shape the dissimilatory sulfur cycle. ISME J 12(7):1715-1728 (PMC6018805) · Pubmed · DOI

    A critical step in the biogeochemical cycle of sulfur on Earth is microbial sulfate reduction, yet organisms from relatively few lineages have been implicated in this process. Previous studies using functional marker genes have detected abundant, novel dissimilatory sulfite reductases (DsrAB) that could confer the capacity for microbial sulfite/sulfate reduction but were not affiliated with known organisms. Thus, the identity of a significant fraction of sulfate/sulfite-reducing microbes has remained elusive. Here we report the discovery of the capacity for sulfate/sulfite reduction in the genomes of organisms from 13 bacterial and archaeal phyla, thereby more than doubling the number of microbial phyla associated with this process. Eight of the 13 newly identified groups are candidate phyla that lack isolated representatives, a finding only possible given genomes from metagenomes. Organisms from Verrucomicrobia and two candidate phyla, Candidatus Rokubacteria and Candidatus Hydrothermarchaeota, contain some of the earliest evolved dsrAB genes. The capacity for sulfite reduction has been laterally transferred in multiple events within some phyla, and a key gene potentially capable of modulating sulfur metabolism in associated cells has been acquired by putatively symbiotic bacteria. We conclude that current functional predictions based on phylogeny significantly underestimate the extent of sulfate/sulfite reduction across Earth's ecosystems. Understanding the prevalence of this capacity is integral to interpreting the carbon cycle because sulfate reduction is often coupled to turnover of buried organic carbon. Our findings expand the diversity of microbial groups associated with sulfur transformations in the environment and motivate revision of biogeochemical process models based on microbial community composition.

  • Probst AJ, Ladd B, Jarett JK, Geller-McGrath DE, Sieber CMK, Emerson JB, Anantharaman K, Thomas BC, Malmstrom RR, Stieglmeier M, Klingl A, Woyke T, Ryan MC, Banfield JF (2018) Differential depth distribution of microbial function and putative symbionts through sediment-hosted aquifers in the deep terrestrial subsurface. Nat Microbiol 3(3):328-336 (PMC6792436) · Pubmed · DOI

    An enormous diversity of previously unknown bacteria and archaea has been discovered recently, yet their functional capacities and distributions in the terrestrial subsurface remain uncertain. Here, we continually sampled a CO-driven geyser (Colorado Plateau, Utah, USA) over its 5-day eruption cycle to test the hypothesis that stratified, sandstone-hosted aquifers sampled over three phases of the eruption cycle have microbial communities that differ both in membership and function. Genome-resolved metagenomics, single-cell genomics and geochemical analyses confirmed this hypothesis and linked microorganisms to groundwater compositions from different depths. Autotrophic Candidatus "Altiarchaeum sp." and phylogenetically deep-branching nanoarchaea dominate the deepest groundwater. A nanoarchaeon with limited metabolic capacity is inferred to be a potential symbiont of the Ca. "Altiarchaeum". Candidate Phyla Radiation bacteria are also present in the deepest groundwater and they are relatively abundant in water from intermediate depths. During the recovery phase of the geyser, microaerophilic Fe- and S-oxidizers have high in situ genome replication rates. Autotrophic Sulfurimonas sustained by aerobic sulfide oxidation and with the capacity for N fixation dominate the shallow aquifer. Overall, 104 different phylum-level lineages are present in water from these subsurface environments, with uncultivated archaea and bacteria partitioned to the deeper subsurface.

  • Zecchin S, Mueller RC, Seifert J, Stingl U, Anantharaman K, von Bergen M, Cavalca L, Pester M (2017) Rice Paddy Nitrospirae Carry and Express Genes Related to Sulfate Respiration: Proposal of the New Genus "Candidatus Sulfobium". Appl. Environ. Microbiol. 84(5): (PMC5812927) · Pubmed · DOI

    spp. distantly related to thermophilic, sulfate-reducing species are regularly observed in environmental surveys of anoxic marine and freshwater habitats. Here we present a metaproteogenomic analysis of bacterium Nbg-4 as a representative of this clade. Its genome was assembled from replicated metagenomes of rice paddy soil that was used to grow rice in the presence and absence of gypsum (CaSO·2HO). Nbg-4 encoded the full pathway of dissimilatory sulfate reduction and showed expression of this pathway in gypsum-amended anoxic bulk soil as revealed by parallel metaproteomics. In addition, Nbg-4 encoded the full pathway of dissimilatory nitrate reduction to ammonia (DNRA), with expression of its first step being detected in bulk soil without gypsum amendment. The relative abundances of Nbg-4 were similar under both treatments, indicating that Nbg-4 maintained stable populations while shifting its energy metabolism. Whether Nbg-4 is a strict sulfate reducer or can couple sulfur oxidation to DNRA by operating the pathway of dissimilatory sulfate reduction in reverse could not be resolved. Further genome reconstruction revealed the potential to utilize butyrate, formate, H, or acetate as an electron donor; the Wood-Ljungdahl pathway was expressed under both treatments. Comparison to publicly available genome bins revealed the pathway for dissimilatory sulfate reduction also in related recovered from groundwater. Subsequent phylogenomics showed that such microorganisms form a novel genus within the , with Nbg-4 as a representative species. Based on the widespread occurrence of this novel genus, we propose for Nbg-4 the name " Sulfobium mesophilum," gen. nov., sp. nov. Rice paddies are indispensable for the food supply but are a major source of the greenhouse gas methane. If it were not counterbalanced by cryptic sulfur cycling, methane emission from rice paddy fields would be even higher. However, the microorganisms involved in this sulfur cycling are little understood. By using an environmental systems biology approach with Italian rice paddy soil, we could retrieve the population genome of a novel member of the phylum This microorganism encoded the full pathway of dissimilatory sulfate reduction and expressed it in anoxic paddy soil under sulfate-enriched conditions. Phylogenomics and comparison to the results of environmental surveys showed that such microorganisms are actually widespread in freshwater and marine environments. At the same time, they represent an undiscovered genus within the little-explored phylum Our results will be important for the design of enrichment strategies and postgenomic studies to further understanding of the contribution of these novel spp. to the global sulfur cycle.

  • Banfield JF, Anantharaman K, Williams KH, Thomas BC (2017) Complete 4.55-Megabase-Pair Genome of " Fluviicola riflensis," Curated from Short-Read Metagenomic Sequences. Genome Announc 5(47): (PMC5701471) · Pubmed · DOI

    We report the 4.55-Mbp genome of " Fluviicola riflensis" () that was manually curated to completion from Illumina data. " Fluviicola riflensis" is a facultative anaerobe. Its ability to grow over a range of O levels may favor its proliferation in an aquifer adjacent to the Colorado River in the United States.

  • Hernsdorf AW, Amano Y, Miyakawa K, Ise K, Suzuki Y, Anantharaman K, Probst A, Burstein D, Thomas BC, Banfield JF (2017) Potential for microbial H and metal transformations associated with novel bacteria and archaea in deep terrestrial subsurface sediments. ISME J 11(8):1915-1929 (PMC5520028) · Pubmed · DOI

    Geological sequestration in deep underground repositories is the prevailing proposed route for radioactive waste disposal. After the disposal of radioactive waste in the subsurface, H may be produced by corrosion of steel and, ultimately, radionuclides will be exposed to the surrounding environment. To evaluate the potential for microbial activities to impact disposal systems, we explored the microbial community structure and metabolic functions of a sediment-hosted ecosystem at the Horonobe Underground Research Laboratory, Hokkaido, Japan. Overall, we found that the ecosystem hosted organisms from diverse lineages, including many from the phyla that lack isolated representatives. The majority of organisms can metabolize H, often via oxidative [NiFe] hydrogenases or electron-bifurcating [FeFe] hydrogenases that enable ferredoxin-based pathways, including the ion motive Rnf complex. Many organisms implicated in H metabolism are also predicted to catalyze carbon, nitrogen, iron and sulfur transformations. Notably, iron-based metabolism is predicted in a novel lineage of Actinobacteria and in a putative methane-oxidizing ANME-2d archaeon. We infer an ecological model that links microorganisms to sediment-derived resources and predict potential impacts of microbial activity on H consumption and retardation of radionuclide migration.

  • Kantor RS, Huddy RJ, Iyer R, Thomas BC, Brown CT, Anantharaman K, Tringe S, Hettich RL, Harrison ST, Banfield JF (2017) Genome-Resolved Meta-Omics Ties Microbial Dynamics to Process Performance in Biotechnology for Thiocyanate Degradation. Environ. Sci. Technol. 51(5):2944-2953 · Pubmed · DOI

    Remediation of industrial wastewater is important for preventing environmental contamination and enabling water reuse. Biological treatment for one industrial contaminant, thiocyanate (SCN), relies upon microbial hydrolysis, but this process is sensitive to high loadings. To examine the activity and stability of a microbial community over increasing SCN loadings, we established and operated a continuous-flow bioreactor fed increasing loadings of SCN. A second reactor was fed ammonium sulfate to mimic breakdown products of SCN. Biomass was sampled from both reactors for metagenomics and metaproteomics, yielding a set of genomes for 144 bacteria and one rotifer that constituted the abundant community in both reactors. We analyzed the metabolic potential and temporal dynamics of these organisms across the increasing loadings. In the SCN reactor, Thiobacillus strains capable of SCN degradation were highly abundant, whereas the ammonium sulfate reactor contained nitrifiers and heterotrophs capable of nitrate reduction. Key organisms in the SCN reactor expressed proteins involved in SCN degradation, sulfur oxidation, carbon fixation, and nitrogen removal. Lower performance at higher loadings was linked to changes in microbial community composition. This work provides an example of how meta-omics can increase our understanding of industrial wastewater treatment and inform iterative process design and development.

  • Zaremba-Niedzwiedzka K, Caceres EF, Saw JH, Bäckström D, Juzokaite L, Vancaester E, Seitz KW, Anantharaman K, Starnawski P, Kjeldsen KU, Stott MB, Nunoura T, Banfield JF, Schramm A, Baker BJ, Spang A, Ettema TJ (2017) Asgard archaea illuminate the origin of eukaryotic cellular complexity. Nature 541(7637):353-358 · Pubmed · DOI

    The origin and cellular complexity of eukaryotes represent a major enigma in biology. Current data support scenarios in which an archaeal host cell and an alphaproteobacterial (mitochondrial) endosymbiont merged together, resulting in the first eukaryotic cell. The host cell is related to Lokiarchaeota, an archaeal phylum with many eukaryotic features. The emergence of the structural complexity that characterizes eukaryotic cells remains unclear. Here we describe the 'Asgard' superphylum, a group of uncultivated archaea that, as well as Lokiarchaeota, includes Thor-, Odin- and Heimdallarchaeota. Asgard archaea affiliate with eukaryotes in phylogenomic analyses, and their genomes are enriched for proteins formerly considered specific to eukaryotes. Notably, thorarchaeal genomes encode several homologues of eukaryotic membrane-trafficking machinery components, including Sec23/24 and TRAPP domains. Furthermore, we identify thorarchaeal proteins with similar features to eukaryotic coat proteins involved in vesicle biogenesis. Our results expand the known repertoire of 'eukaryote-specific' proteins in Archaea, indicating that the archaeal host cell already contained many key components that govern eukaryotic cellular complexity.

  • Burstein D, Harrington LB, Strutt SC, Probst AJ, Anantharaman K, Thomas BC, Doudna JA, Banfield JF (2016) New CRISPR-Cas systems from uncultivated microbes. Nature 542(7640):237-241 (PMC5300952) · Pubmed · DOI

    CRISPR-Cas systems provide microbes with adaptive immunity by employing short DNA sequences, termed spacers, that guide Cas proteins to cleave foreign DNA. Class 2 CRISPR-Cas systems are streamlined versions, in which a single RNA-bound Cas protein recognizes and cleaves target sequences. The programmable nature of these minimal systems has enabled researchers to repurpose them into a versatile technology that is broadly revolutionizing biological and clinical research. However, current CRISPR-Cas technologies are based solely on systems from isolated bacteria, leaving the vast majority of enzymes from organisms that have not been cultured untapped. Metagenomics, the sequencing of DNA extracted directly from natural microbial communities, provides access to the genetic material of a huge array of uncultivated organisms. Here, using genome-resolved metagenomics, we identify a number of CRISPR-Cas systems, including the first reported Cas9 in the archaeal domain of life, to our knowledge. This divergent Cas9 protein was found in little-studied nanoarchaea as part of an active CRISPR-Cas system. In bacteria, we discovered two previously unknown systems, CRISPR-CasX and CRISPR-CasY, which are among the most compact systems yet discovered. Notably, all required functional components were identified by metagenomics, enabling validation of robust in vivo RNA-guided DNA interference activity in Escherichia coli. Interrogation of environmental microbial communities combined with in vivo experiments allows us to access an unprecedented diversity of genomes, the content of which will expand the repertoire of microbe-based biotechnologies.

  • Marcus DN, Pinto A, Anantharaman K, Ruberg SA, Kramer EL, Raskin L, Dick GJ (2016) Diverse manganese(II)-oxidizing bacteria are prevalent in drinking water systems. Environ Microbiol Rep 9(2):120-128 · Pubmed · DOI

    Manganese (Mn) oxides are highly reactive minerals that influence the speciation, mobility, bioavailability and toxicity of a wide variety of organic and inorganic compounds. Although Mn(II)-oxidizing bacteria are known to catalyze the formation of Mn oxides, little is known about the organisms responsible for Mn oxidation in situ, especially in engineered environments. Mn(II)-oxidizing bacteria are important in drinking water systems, including in biofiltration and water distribution systems. Here, we used cultivation dependent and independent approaches to investigate Mn(II)-oxidizing bacteria in drinking water sources, a treatment plant and associated distribution system. We isolated 29 strains of Mn(II)-oxidizing bacteria and found that highly similar 16S rRNA gene sequences were present in all culture-independent datasets and dominant in the studied drinking water treatment plant. These results highlight a potentially important role for Mn(II)-oxidizing bacteria in drinking water systems, where biogenic Mn oxides may affect water quality in terms of aesthetic appearance, speciation of metals and oxidation of organic and inorganic compounds. Deciphering the ecology of these organisms and the factors that regulate their Mn(II)-oxidizing activity could yield important insights into how microbial communities influence the quality of drinking water.

  • Anantharaman K, Brown CT, Hug LA, Sharon I, Castelle CJ, Probst AJ, Thomas BC, Singh A, Wilkins MJ, Karaoz U, Brodie EL, Williams KH, Hubbard SS, Banfield JF (2016) Thousands of microbial genomes shed light on interconnected biogeochemical processes in an aquifer system. Nat Commun 7:13219 (PMC5079060) · Pubmed

    The subterranean world hosts up to one-fifth of all biomass, including microbial communities that drive transformations central to Earth's biogeochemical cycles. However, little is known about how complex microbial communities in such environments are structured, and how inter-organism interactions shape ecosystem function. Here we apply terabase-scale cultivation-independent metagenomics to aquifer sediments and groundwater, and reconstruct 2,540 draft-quality, near-complete and complete strain-resolved genomes that represent the majority of known bacterial phyla as well as 47 newly discovered phylum-level lineages. Metabolic analyses spanning this vast phylogenetic diversity and representing up to 36% of organisms detected in the system are used to document the distribution of pathways in coexisting organisms. Consistent with prior findings indicating metabolic handoffs in simple consortia, we find that few organisms within the community can conduct multiple sequential redox transformations. As environmental conditions change, different assemblages of organisms are selected for, altering linkages among the major biogeochemical cycles.

  • Hug LA, Baker BJ, Anantharaman K, Brown CT, Probst AJ, Castelle CJ, Butterfield CN, Hernsdorf AW, Amano Y, Ise K, Suzuki Y, Dudek N, Relman DA, Finstad KM, Amundson R, Thomas BC, Banfield JF (2016) A new view of the tree of life. Nat Microbiol 1:16048 · Pubmed

    The tree of life is one of the most important organizing principles in biology(1). Gene surveys suggest the existence of an enormous number of branches(2), but even an approximation of the full scale of the tree has remained elusive. Recent depictions of the tree of life have focused either on the nature of deep evolutionary relationships(3-5) or on the known, well-classified diversity of life with an emphasis on eukaryotes(6). These approaches overlook the dramatic change in our understanding of life's diversity resulting from genomic sampling of previously unexamined environments. New methods to generate genome sequences illuminate the identity of organisms and their metabolic capacities, placing them in community and ecosystem contexts(7,8). Here, we use new genomic data from over 1,000 uncultivated and little known organisms, together with published sequences, to infer a dramatically expanded version of the tree of life, with Bacteria, Archaea and Eukarya included. The depiction is both a global overview and a snapshot of the diversity within each major lineage. The results reveal the dominance of bacterial diversification and underline the importance of organisms lacking isolated representatives, with substantial evolution concentrated in a major radiation of such organisms. This tree highlights major lineages currently underrepresented in biogeochemical models and identifies radiations that are probably important for future evolutionary analyses.

  • Probst AJ, Castelle CJ, Singh A, Brown CT, Anantharaman K, Sharon I, Hug LA, Burstein D, Emerson JB, Thomas BC, Banfield JF (2016) Genomic resolution of a cold subsurface aquifer community provides metabolic insights for novel microbes adapted to high CO concentrations. Environ. Microbiol. 19(2):459-474 · Pubmed · DOI

    As in many deep underground environments, the microbial communities in subsurface high-CO ecosystems remain relatively unexplored. Recent investigations based on single-gene assays revealed a remarkable variety of organisms from little studied phyla in Crystal Geyser (Utah, USA), a site where deeply sourced CO -saturated fluids are erupted at the surface. To provide genomic resolution of the metabolisms of these organisms, we used a novel metagenomic approach to recover 227 high-quality genomes from 150 microbial species affiliated with 46 different phylum-level lineages. Bacteria from two novel phylum-level lineages have the capacity for CO fixation. Analyses of carbon fixation pathways in all studied organisms revealed that the Wood-Ljungdahl pathway and the Calvin-Benson-Bassham Cycle occurred with the highest frequency, whereas the reverse TCA cycle was little used. We infer that this, and selection for form II RuBisCOs, are adaptions to high CO -concentrations. However, many autotrophs can also grow mixotrophically, a strategy that confers metabolic versatility. The assignment of 156 hydrogenases to 90 different organisms suggests that H is an important inter-species energy currency even under gaseous CO -saturation. Overall, metabolic analyses at the organism level provided insight into the biochemical cycles that support subsurface life under the extreme condition of CO saturation.

  • Anantharaman K, Brown CT, Burstein D, Castelle CJ, Probst AJ, Thomas BC, Williams KH, Banfield JF (2016) Analysis of five complete genome sequences for members of the class Peribacteria in the recently recognized Peregrinibacteria bacterial phylum. PeerJ 4:e1607 (PMC4736985) · Pubmed

    Five closely related populations of bacteria from the Candidate Phylum (CP) Peregrinibacteria, part of the bacterial Candidate Phyla Radiation (CPR), were sampled from filtered groundwater obtained from an aquifer adjacent to the Colorado River near the town of Rifle, CO, USA. Here, we present the first complete genome sequences for organisms from this phylum. These bacteria have small genomes and, unlike most organisms from other lineages in the CPR, have the capacity for nucleotide synthesis. They invest significantly in biosynthesis of cell wall and cell envelope components, including peptidoglycan, isoprenoids via the mevalonate pathway, and a variety of amino sugars including perosamine and rhamnose. The genomes encode an intriguing set of large extracellular proteins, some of which are very cysteine-rich and may function in attachment, possibly to other cells. Strain variation in these proteins is an important source of genotypic variety. Overall, the cell envelope features, combined with the lack of biosynthesis capacities for many required cofactors, fatty acids, and most amino acids point to a symbiotic lifestyle. Phylogenetic analyses indicate that these bacteria likely represent a new class within the Peregrinibacteria phylum, although they ultimately may be recognized as members of a separate phylum. We propose the provisional taxonomic assignment as 'Candidatus Peribacter riflensis', Genus Peribacter, Family Peribacteraceae, Order Peribacterales, Class Peribacteria in the phylum Peregrinibacteria.

  • Burstein D, Sun CL, Brown CT, Sharon I, Anantharaman K, Probst AJ, Thomas BC, Banfield JF (2016) Major bacterial lineages are essentially devoid of CRISPR-Cas viral defence systems. Nat Commun 7:10613 (PMC4742961) · Pubmed

    Current understanding of microorganism-virus interactions, which shape the evolution and functioning of Earth's ecosystems, is based primarily on cultivated organisms. Here we investigate thousands of viral and microbial genomes recovered using a cultivation-independent approach to study the frequency, variety and taxonomic distribution of viral defence mechanisms. CRISPR-Cas systems that confer microorganisms with immunity to viruses are present in only 10% of 1,724 sampled microorganisms, compared with previous reports of 40% occurrence in bacteria and 81% in archaea. We attribute this large difference to the lack of CRISPR-Cas systems across major bacterial lineages that have no cultivated representatives. We correlate absence of CRISPR-Cas with lack of nucleotide biosynthesis capacity and a symbiotic lifestyle. Restriction systems are well represented in these lineages and might provide both non-specific viral defence and access to nucleotides.

  • Varaljay VA, Satagopan S, North JA, Witte B, Dourado MN, Anantharaman K, Arbing MA, Hoeft McCann S, Oremland RS, Banfield JF, Wrighton KC, Tabita FR (2015) Functional metagenomic selection of ribulose 1, 5-bisphosphate carboxylase/oxygenase from uncultivated bacteria. Environ. Microbiol. 18(4):1187-99 · Pubmed

    Ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) is a critical yet severely inefficient enzyme that catalyses the fixation of virtually all of the carbon found on Earth. Here, we report a functional metagenomic selection that recovers physiologically active RubisCO molecules directly from uncultivated and largely unknown members of natural microbial communities. Selection is based on CO2 -dependent growth in a host strain capable of expressing environmental deoxyribonucleic acid (DNA), precluding the need for pure cultures or screening of recombinant clones for enzymatic activity. Seventeen functional RubisCO-encoded sequences were selected using DNA extracted from soil and river autotrophic enrichments, a photosynthetic biofilm and a subsurface groundwater aquifer. Notably, three related form II RubisCOs were recovered which share high sequence similarity with metagenomic scaffolds from uncultivated members of the Gallionellaceae family. One of the Gallionellaceae RubisCOs was purified and shown to possess CO2 /O2 specificity typical of form II enzymes. X-ray crystallography determined that this enzyme is a hexamer, only the second form II multimer ever solved and the first RubisCO structure obtained from an uncultivated bacterium. Functional metagenomic selection leverages natural biological diversity and billions of years of evolution inherent in environmental communities, providing a new window into the discovery of CO2 -fixing enzymes not previously characterized.

  • Anantharaman K, Breier JA, Dick GJ (2015) Metagenomic resolution of microbial functions in deep-sea hydrothermal plumes across the Eastern Lau Spreading Center. ISME J 10(1):225-39 (PMC4681857) · Pubmed

    Microbial processes within deep-sea hydrothermal plumes affect ocean biogeochemistry on global scales. In rising hydrothermal plumes, a combination of microbial metabolism and particle formation processes initiate the transformation of reduced chemicals like hydrogen sulfide, hydrogen, methane, iron, manganese and ammonia that are abundant in hydrothermal vent fluids. Despite the biogeochemical importance of this rising portion of plumes, it is understudied in comparison to neutrally buoyant plumes. Here we use metagenomics and bioenergetic modeling to describe the abundance and genetic potential of microorganisms in relation to available electron donors in five different hydrothermal plumes and three associated background deep-sea waters from the Eastern Lau Spreading Center located in the Western Pacific Ocean. Three hundred and thirty one distinct genomic 'bins' were identified, comprising an estimated 951 genomes of archaea, bacteria, eukarya and viruses. A significant proportion of these genomes is from novel microorganisms and thus reveals insights into the energy metabolism of heretofore unknown microbial groups. Community-wide analyses of genes encoding enzymes that oxidize inorganic energy sources showed that sulfur oxidation was the most abundant and diverse chemolithotrophic microbial metabolism in the community. Genes for sulfur oxidation were commonly present in genomic bins that also contained genes for oxidation of hydrogen and methane, suggesting metabolic versatility in these microbial groups. The relative diversity and abundance of genes encoding hydrogen oxidation was moderate, whereas that of genes for methane and ammonia oxidation was low in comparison to sulfur oxidation. Bioenergetic-thermodynamic modeling supports the metagenomic analyses, showing that oxidation of elemental sulfur with oxygen is the most dominant catabolic reaction in the hydrothermal plumes. We conclude that the energy metabolism of microbial communities inhabiting rising hydrothermal plumes is dictated by the underlying plume chemistry, with a dominant role for sulfur-based chemolithoautotrophy.