Publications

@article{D2017,

title = {Characterization of two cryptic plasmids isolated in Haiti from clinical Vibrio cholerae non-O1/non-O139},

author = {Ceccarelli D and Garriss G and Choi SY and Hasan NA and Stepanauskas R and Pop M and Huq A and Colwell RR},

url = {https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5703827/},

doi = {10.3389/fmicb.2017.02283},

year  = {2017},

date = {2017-11-23},

journal = {Frontiers in Microbiology},

volume = {8},

abstract = {We report the complete sequence of two novel plasmids, pSDH-1 and pSDH-2, isolated from clinical Vibrio cholerae non-O1/non-O139 during the early phase of the 2010 Haitian cholera epidemic. Plasmids were revealed by employing single-cell genomics and their genome content suggests self-mobilization and, for pSDH-2, a toxin-antitoxin (TA) system for plasmid stabilization was identified. The putative origin of replication of pSDH-2 was mapped suggesting it replicates following the ColE1 model of plasmid replication. pSDH-1 and pSDH-2 were widespread among environmental V. cholerae non-O1/non-O139 with variable prevalence in four Haitian Departments. pSDH-2 was the most common element, either alone or with pSDH-1. The two plasmids detection adds to the composite scenario of mobile genetic elements (MGEs) observed in V. cholerae in Haiti. The role these small cryptic plasmids circulating in Vibrio spp. play in bacterial fitness or pathogenicity merits further investigation.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{AK2017,

title = {Diverse Marinimicrobia bacteria may mediate coupled biogeochemical cycles along eco-thermodynamic gradients},

author = {Hawley AK and Nobu MK and Wright JJ and Durno WE and Morgan-Lang C and Sage B and Schwientek P and Swan BK and Rinke C and Torres-Beltr\'{a}n M and Mewis K and Liu WT and Stepanauskas R and Woyke T and Hallam SJ},

url = {https://www.nature.com/articles/s41467-017-01376-9},

doi = {10.1038/s41467-017-01376-9},

year  = {2017},

date = {2017-11-15},

journal = {Nature Communications},

volume = {8},

abstract = {Microbial communities drive biogeochemical cycles through networks of metabolite exchange that are structured along energetic gradients. As energy yields become limiting, these networks favor co-metabolic interactions to maximize energy disequilibria. Here we apply single-cell genomics, metagenomics, and metatranscriptomics to study bacterial populations of the abundant “microbial dark matter” phylum Marinimicrobia along defined energy gradients. We show that evolutionary diversification of major Marinimicrobia clades appears to be closely related to energy yields, with increased co-metabolic interactions in more deeply branching clades. Several of these clades appear to participate in the biogeochemical cycling of sulfur and nitrogen, filling previously unassigned niches in the ocean. Notably, two Marinimicrobia clades, occupying different energetic niches, express nitrous oxide reductase, potentially acting as a global sink for the greenhouse gas nitrous oxide.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{ED2017b,

title = {Genomic comparison of two family-level groups of the uncultivated NAG1 archaeal lineage from chemically and geographically disparate hot springs},

author = {Becraft ED and Dodsworth JA and Murugapiran SK and Thomas SC and Ohlsson JI and Stepanauskas R and Hedlund BP and Swingley WD},

url = {https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5671600/},

doi = {10.3389/fmicb.2017.02082},

year  = {2017},

date = {2017-10-31},

journal = {Frontiers in Microbiology },

volume = {8},

abstract = {Recent progress based on single-cell genomics and metagenomic investigations of archaea in a variety of extreme environments has led to significant advances in our understanding of the diversity, evolution, and metabolic potential of archaea, yet the vast majority of archaeal diversity remains undersampled. In this work, we coordinated single-cell genomics with metagenomics in order to construct a near-complete genome from a deeply branching uncultivated archaeal lineage sampled from Great Boiling Spring (GBS) in the U.S. Great Basin, Nevada. This taxon is distantly related (distinct families) to an archaeal genome, designated “Novel Archaeal Group 1” (NAG1), which was extracted from a metagenome recovered from an acidic iron spring in Yellowstone National Park (YNP). We compared the metabolic predictions of the NAG1 lineage to better understand how these archaea could inhabit such chemically distinct environments. Similar to the NAG1 population previously studied in YNP, the NAG1 population from GBS is predicted to utilize proteins as a primary carbon source, ferment simple carbon sources, and use oxygen as a terminal electron acceptor under oxic conditions. However, GBS NAG1 populations contained distinct genes involved in central carbon metabolism and electron transfer, including nitrite reductase, which could confer the ability to reduce nitrite under anaerobic conditions. Despite inhabiting chemically distinct environments with large variations in pH, GBS NAG1 populations shared many core genomic and metabolic features with the archaeon identified from YNP, yet were able to carve out a distinct niche at GBS.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{J.J2017,

title = {Metabolic Network Analysis and Metatranscriptomics Reveal Auxotrophies and Nutrient Sources of the Cosmopolitan Freshwater Microbial Lineage acI},

author = {Hamilton J.J and Garcia S.L and Brown B.S and Oyserman B.O and Moya-Flores F and Bertilsson S and Malmstrom R.R and Forest K and McMahon K },

url = {https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5574706/pdf/mSystems.00091-17.pdf},

doi = {10.1128/mSystems.00091-17},

year  = {2017},

date = {2017-08-29},

journal = {American Society for Microbiology},

volume = {2},

number = {4},

pages = {e00091-17},

abstract = {An explosion in the number of available genome sequences obtained through metagenomics and single-cell genomics has enabled a new view of the diversity of microbial life, yet we know surprisingly little about how microbes interact with each other or their environment. In fact, the majority of microbial species remain uncultivated, while our perception of their ecological niches is based on reconstruction of their metabolic potential. In this work, we demonstrate how the "seed set framework," which computes the set of compounds that an organism must acquire from its environment (E. Borenstein, M. Kupiec, M. W. Feldman, and E. Ruppin, Proc Natl Acad Sci U S A 105:14482-14487, 2008, https://doi.org/10.1073/pnas.0806162105), enables computational analysis of metabolic reconstructions while providing new insights into a microbe's metabolic capabilities, such as nutrient use and auxotrophies. We apply this framework to members of the ubiquitous freshwater actinobacterial lineage acI, confirming and extending previous experimental and genomic observations implying that acI bacteria are heterotrophs reliant on peptides and saccharides. We also present the first metatranscriptomic study of the acI lineage, revealing high expression of transport proteins and the light-harvesting protein actinorhodopsin. Putative transport proteins complement predictions of nutrients and essential metabolites while providing additional support of the hypothesis that members of the acI are photoheterotrophs. IMPORTANCE The metabolic activity of uncultivated microorganisms contributes to numerous ecosystem processes, ranging from nutrient cycling in the environment to influencing human health and disease. Advances in sequencing technology have enabled the assembly of genomes for these microorganisms, but our ability to generate reference genomes far outstrips our ability to analyze them. Common approaches to analyzing microbial metabolism require reconstructing the entirety of an organism's metabolic pathways or performing targeted searches for genes involved in a specific process. This paper presents a third approach, in which draft metabolic reconstructions are used to identify compounds through which an organism may interact with its environment. These compounds can then guide more-intensive metabolic reconstruction efforts and can also provide new hypotheses about the specific contributions that microbes make to ecosystem-scale metabolic processes.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{JF2017b,

title = {Physiological and ecological implications of an iron- or hydrogen-oxidizing member of the Zetaproteobacteria, Ghiorsea bivora, gen. nov., sp. nov},

author = {Mori JF and Scott JJ and Hager KW and Moyer CL and Kusel K and Emerson D},

url = {https://www.nature.com/articles/ismej2017132},

doi = {10.1038/ismej.2017.132},

year  = {2017},

date = {2017-08-18},

journal = {The ISME Journal},

volume = {11},

pages = {2624-2636},

abstract = {Chemosynthetic Fe-oxidizing communities are common at diffuse-flow hydrothermal vents throughout the world's oceans. The foundational members of these communities are the Zetaproteobacteria, a class of Proteobacteria that is primarily associated with ecosystems fueled by ferrous iron, Fe(II). We report here the discovery of two new isolates of Zetaproteobacteria isolated from the Mid-Atlantic Ridge (TAG-1), and the Mariana back-arc (SV-108), that are unique in that they can utilize either Fe(II) or molecular hydrogen (H2) as sole electron donor and oxygen as terminal electron acceptor for growth. Both strains precipitated Fe-oxyhydroxides as amorphous particulates. The cell doubling time on H2 vs Fe(II) for TAG-1 was 14.1 vs 21.8 h, and for SV-108 it was 16.3 vs 20 h, and it appeared both strains could use either H2 or Fe(II) simultaneously. The strains were close relatives, based on genomic analysis, and both possessed genes for the uptake NiFe-hydrogenase required for growth on H2. These two strains belong to Zetaproteobacteria operational taxonomic unit 9 (ZetaOTU9). A meta-analysis of public databases found ZetaOTU9 was only associated with Fe(II)-rich habitats, and not in other environments where known H2-oxidizers exist. These results expand the metabolic repertoire of the Zetaproteobacteria, yet confirm that Fe(II) metabolism is the primary driver of their physiology and ecology.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{RM2017b,

title = {Minimum information about a single amplified genome (MISAG) and a metagenome-assembled genome (MIMAG) of bacteria and archaea},

author = {Bowers RM and Kyrpides NC and Stepanauskas R and Harmon-Smith M and Doud D and Reddy TBK and Schulz F and Jarett J and Rivers AR and Eloe-Fadrosh EA and Tringe SG and Ivanova NN and Copeland A and Clum A and Becraft ED and Malmstrom RR and Birren B and Podar M and Bork P and Weinstock GM and Garrity GM and Dodsworth JA and Yooseph S and Sutton G and Gl\"{o}ckner FO and Gilbert JA and Nelson WC and Hallam SJ and Jungbluth SP and Ettema TJG and Tighe S and Konstantinidis KT and Liu WT and Baker BJ and Rattei T and Eisen JA and Hedlund B and McMahon KD and Fierer N and Knight R and Finn R and Cochrane G and Karsch-Mizrachi I and Tyson GW and Rinke C and Lapidus A and Meyer F and Yilmaz P and Parks DH and Eren AM and Schriml L and Banfield JF and Hugenholtz P and Woyke T},

url = {https://www.nature.com/articles/nbt.3893},

doi = {10.1038/nbt.3893},

year  = {2017},

date = {2017-08-08},

journal = {Nature Biotechnology},

volume = {35},

pages = {725-731},

abstract = {We present two standards developed by the Genomic Standards Consortium (GSC) for reporting bacterial and archaeal genome sequences. Both are extensions of the Minimum Information about Any (x) Sequence (MIxS). The standards are the Minimum Information about a Single Amplified Genome (MISAG) and the Minimum Information about a Metagenome-Assembled Genome (MIMAG), including, but not limited to, assembly quality, and estimates of genome completeness and contamination. These standards can be used in combination with other GSC checklists, including the Minimum Information about a Genome Sequence (MIGS), Minimum Information about a Metagenomic Sequence (MIMS), and Minimum Information about a Marker Gene Sequence (MIMARKS). Community-wide adoption of MISAG and MIMAG will facilitate more robust comparative genomic analyses of bacterial and archaeal diversity.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{A2017b,

title = {Viral to metazoan marine plankton nucleotide sequences from the Tara Oceans expedition},

author = {Alberti A and Poulain J and Engelen S and Labadie K and Romac S and Ferrera I and Albini G and Aury JM and Belser C and Bertrand A and Cruaud C and Da Silva C and Dossat C and Gavory F and Gas S and Guy J and Haquelle M and Jacoby E and Jaillon O and Lemainque A and Pelletier E and Samson G and Wessner M and Acinas SG and Royo-Llonch M and Cornejo-Castillo FM and Logares R and Fern\'{a}ndez-G\'{o}mez B and Bowler C and Cochrane G and Amid C and Hoopen PT and De Vargas C and Grimsley N and Desgranges E and Kandels-Lewis S and Ogata H and Poulton N and Sieracki ME and Stepanauskas R and Sullivan MB and Brum JR and Duhaime MB and Poulos BT and Hurwitz BL and Pesant S and Karsenti E and Wincker P and Bazire P and Beluche O and Bertrand L and Besnard-Gonnet M and Bordelais I and Boutard M and Dubois M and Dumont C and Ettedgui E and Fernandez P and Garcia E and Aiach NG and Guerin T and Hamon C and Brun E and Lebled S and Lenoble P and Louesse C and Mahieu E and Mairey B and Martins N and Megret C and Milani C and Muanga J and Orvain C and Payen E and Perroud P and Petit E and Robert D and Ronsin M and Vacherie B and Bork P and Boss E and Follows M and Gorsky G and Hingamp P and Iudicone D and Karp-Boss L and Not F and Raes J and Sardet C and Speich S and Stemmann L and Sunagawa S},

url = {https://www.nature.com/articles/sdata201793},

doi = {10.1038/sdata.2017.93},

year  = {2017},

date = {2017-08-01},

journal = {Scientific Data},

volume = {4},

abstract = {A unique collection of oceanic samples was gathered by the Tara Oceans expeditions (2009\textendash2013), targeting plankton organisms ranging from viruses to metazoans, and providing rich environmental context measurements. Thanks to recent advances in the field of genomics, extensive sequencing has been performed for a deep genomic analysis of this huge collection of samples. A strategy based on different approaches, such as metabarcoding, metagenomics, single-cell genomics and metatranscriptomics, has been chosen for analysis of size-fractionated plankton communities. Here, we provide detailed procedures applied for genomic data generation, from nucleic acids extraction to sequence production, and we describe registries of genomics datasets available at the European Nucleotide Archive (ENA, www.ebi.ac.uk/ena). The association of these metadata to the experimental procedures applied for their generation will help the scientific community to access these data and facilitate their analysis. This paper complements other efforts to provide a full description of experiments and open science resources generated from the Tara Oceans project, further extending their value for the study of the world’s planktonic ecosystems.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{M2017b,

title = {Exploring microdiversity in novel Kordia sp. (Bacteroidetes) with proteorhodopsin from the tropical Indian Ocean via Single Amplified Genomes},

author = {Royo-Llonch M and Ferrera I and Cornejo-Castillo FM and S\'{a}nchez P and Salazar G and Stepanauskas R and Gonz\'{a}lez JM and Sieracki ME and Speich S and Stemmann L and Pedr\'{o}s-Ali\'{o} C and Acinas SG},

url = {https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5525439/},

doi = {10.3389/fmicb.2017.01317},

year  = {2017},

date = {2017-07-25},

journal = {Frontiers in Microbiology},

volume = {8},

abstract = {Marine Bacteroidetes constitute a very abundant bacterioplankton group in the oceans that plays a key role in recycling particulate organic matter and includes several photoheterotrophic members containing proteorhodopsin. Relatively few marine Bacteroidetes species have been described and, moreover, they correspond to cultured isolates, which in most cases do not represent the actual abundant or ecologically relevant microorganisms in the natural environment. In this study, we explored the microdiversity of 98 Single Amplified Genomes (SAGs) retrieved from the surface waters of the underexplored North Indian Ocean, whose most closely related isolate is Kordia algicida OT-1. Using Multi Locus Sequencing Analysis (MLSA) we found no microdiversity in the tested conserved phylogenetic markers (16S rRNA and 23S rRNA genes), the fast-evolving Internal Transcribed Spacer and the functional markers proteorhodopsin and the beta-subunit of RNA polymerase. Furthermore, we carried out a Fragment Recruitment Analysis (FRA) with marine metagenomes to learn about the distribution and dynamics of this microorganism in different locations, depths and size fractions. This analysis indicated that this taxon belongs to the rare biosphere, showing its highest abundance after upwelling-induced phytoplankton blooms and sinking to the deep ocean with large organic matter particles. This uncultured Kordia lineage likely represents a novel Kordia species (Kordia sp. CFSAG39SUR) that contains the proteorhodopsin gene and has a widespread spatial and vertical distribution. The combination of SAGs and MLSA makes a valuable approach to infer putative ecological roles of uncultured abundant microorganisms.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{R2017,

title = {Improved genome recovery and integrated cell-size analyses of individual uncultured microbial cells and viral particles},

author = {Stepanauskas R and Fergusson EA and Brown J and Poulton NJ and Tupper B and Labont\'{e} JM and Becraft ED and Brown JM and Pachiadaki MJ and Povilaitis T and Thompson BP and Mascena CJ and Bellows WK and Lubys A},

url = {https://www.nature.com/articles/s41467-017-00128-z},

doi = {10.1038/s41467-017-00128-z},

year  = {2017},

date = {2017-07-20},

journal = {Nature Communications},

volume = {8},

number = {84},

abstract = {Microbial single-cell genomics can be used to provide insights into the metabolic potential, interactions, and evolution of uncultured microorganisms. Here we present WGA-X, a method based on multiple displacement amplification of DNA that utilizes a thermostable mutant of the phi29 polymerase. WGA-X enhances genome recovery from individual microbial cells and viral particles while maintaining ease of use and scalability. The greatest improvements are observed when amplifying high G+C content templates, such as those belonging to the predominant bacteria in agricultural soils. By integrating WGA-X with calibrated index-cell sorting and high-throughput genomic sequencing, we are able to analyze genomic sequences and cell sizes of hundreds of individual, uncultured bacteria, archaea, protists, and viral particles, obtained directly from marine and soil samples, in a single experiment. This approach may find diverse applications in microbiology and in biomedical and forensic studies of humans and other multicellular organisms.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{A2017,

title = {Unexpected genomic features in widespread intracellular bacteria: evidence for motility of marine chlamydiae},

author = {Collingro A and Köstlbacher S and Mussmann M and Stepanauskas R and Hallam S and Horn M},

url = {https://www.nature.com/ismej/journal/vaop/ncurrent/full/ismej201795a.html},

doi = {10.1038/ismej.2017.95},

year  = {2017},

date = {2017-06-23},

journal = {ISME},

abstract = {Chlamydiae are obligate intracellular bacteria comprising important human pathogens and symbionts of protists. Molecular evidence indicates a tremendous diversity of chlamydiae particularly in marine environments, yet our current knowledge is based mainly on terrestrial representatives. Here we provide first insights into the biology of marine chlamydiae representing three divergent clades. Our analysis of single-cell amplified genomes revealed hallmarks of the chlamydial lifestyle, supporting the ancient origin of their characteristic developmental cycle and major virulence mechanisms. Surprisingly, these chlamydial genomes encode a complete flagellar apparatus, a previously unreported feature. We show that flagella are an ancient trait that was subject to differential gene loss among extant chlamydiae. Together with a chemotaxis system, these marine chlamydiae are likely motile, with flagella potentially playing a role during host cell infection. This study broadens our view on chlamydial biology and indicates a largely underestimated potential to adapt to different hosts and environments.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{Luo2017,

title = {Excess of non-conservative amino acid changes in marine bacterioplankton lineages with reduced genomes},

author = {Haiwei Luo and Yongjie Huang and Ramunas Stepanauskas and Jijun Tang},

url = {https://www.nature.com/articles/nmicrobiol201791},

doi = {10.1007/s11096-011-9488-z},

year  = {2017},

date = {2017-06-12},

journal = {Nature Microbiology},

volume = {2},

number = {17091},

abstract = {Surface ocean waters are dominated by planktonic bacterial lineages with highly reduced genomes. The best examples are the cyanobacterial genus Prochlorococcus, the alphaproteobacterial clade SAR11 and the gammaproteobacterial clade SAR86, which together represent over 50% of the cells in surface oceans. Several studies have identified signatures of selection on these lineages in today's ocean and have postulated selection as the primary force throughout their evolutionary history. However, massive loss of genomic DNA in these lineages often occurred in the distant past, and the selective pressures underlying these ancient events have not been assessed. Here, we probe ancient selective pressures by computing %GC-corrected rates of conservative and radical nonsynonymous nucleotide substitutions. Surprisingly, we found an excess of radical changes in several of these lineages in comparison to their relatives with larger genomes. Furthermore, analyses of allelic genome sequences of several populations within these lineages consistently supported that radical replacements are more likely to be deleterious than conservative changes. Our results suggest coincidence of massive genomic DNA losses and increased power of genetic drift, but we also suggest that additional evidence independent of the nucleotide substitution analyses is needed to support a primary role of genetic drift driving ancient genome reduction of marine bacterioplankton lineages.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{N2017,

title = {Fundamental differences in diversity and genomic population structure between Atlantic and Pacific Prochlorococcus},

author = {Kashtan N and Roggensack SE and Berta-Thompson JW and Grinberg M and Stepanauskas R and Chisholm SW},

url = {http://www.nature.com/ismej/journal/vaop/ncurrent/full/ismej201764a.html},

doi = {10.1038/ismej.2017.64},

year  = {2017},

date = {2017-05-19},

journal = {ISME},

abstract = {The Atlantic and Pacific Oceans represent different biogeochemical regimes in which the abundant marine cyanobacterium Prochlorococcus thrives. We have shown that Prochlorococcus populations in the Atlantic are composed of hundreds of genomically, and likely ecologically, distinct coexisting subpopulations with distinct genomic backbones. Here we ask if differences in the ecology and selection pressures between the Atlantic and Pacific are reflected in the diversity and genomic composition of their indigenous Prochlorococcus populations. We applied large-scale single-cell genomics and compared the cell-by-cell genomic composition of wild populations of co-occurring cells from samples from Station ALOHA off Hawaii, and from Bermuda Atlantic Time Series Station off Bermuda. We reveal fundamental differences in diversity and genomic structure of populations between the sites. The Pacific populations are more diverse than those in the Atlantic, composed of significantly more coexisting subpopulations and lacking dominant subpopulations. Prochlorococcus from the two sites seem to be composed of mostly non-overlapping distinct sets of subpopulations with different genomic backbones\textemdashlikely reflecting different sets of ocean-specific micro-niches. Furthermore, phylogenetically closely related strains carry ocean-associated nutrient acquisition genes likely reflecting differences in major selection pressures between the oceans. This differential selection, along with geographic separation, clearly has a significant role in shaping these populations.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{WH2017,

title = {Genomic exploration of individual giant ocean viruses},

author = {Wilson WH and Gilg IC and Moniruzzaman M and Field EK and Koren S and LeCleir GR and Martínez Martínez J and Poulton NJ and Swan BK and Stepanauskas R and Wilhelm SW},

url = {https://www.nature.com/ismej/journal/vaop/ncurrent/full/ismej201761a.html},

doi = {10.1038/ismej.2017.61},

year  = {2017},

date = {2017-05-12},

journal = {ISME},

abstract = {Viruses are major pathogens in all biological systems. Virus propagation and downstream analysis remains a challenge, particularly in the ocean where the majority of their microbial hosts remain recalcitrant to current culturing techniques. We used a cultivation-independent approach to isolate and sequence individual viruses. The protocol uses high-speed fluorescence-activated virus sorting flow cytometry, multiple displacement amplification (MDA), and downstream genomic sequencing. We focused on ‘giant viruses’ that are readily distinguishable by flow cytometry. From a single-milliliter sample of seawater collected from off the dock at Boothbay Harbor, ME, USA, we sorted almost 700 single virus particles, and subsequently focused on a detailed genome analysis of 12. A wide diversity of viruses was identified that included Iridoviridae, extended Mimiviridae and even a taxonomically novel (unresolved) giant virus. We discovered a viral metacaspase homolog in one of our sorted virus particles and discussed its implications in rewiring host metabolism to enhance infection. In addition, we demonstrated that viral metacaspases are widespread in the ocean. We also discovered a virus that contains both a reverse transcriptase and a transposase; although highly speculative, we suggest such a genetic complement would potentially allow this virus to exploit a latency propagation mechanism. Application of single virus genomics provides a powerful opportunity to circumvent cultivation of viruses, moving directly to genomic investigation of naturally occurring viruses, with the assurance that the sequence data is virus-specific, non-chimeric and contains no cellular contamination.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{Y2017,

title = {Single-cell genomics reveals pyrrolysine-encoding potential in members of uncultivated archaeal candidate division MSBL1},

author = {Guan Y and Haroon MF and Alam I and Ferry JG and Stingl U },

url = {https://www.ncbi.nlm.nih.gov/pubmed/28493460},

doi = {10.1111/1758-2229.12545},

year  = {2017},

date = {2017-05-10},

volume = {9},

number = {4},

pages = {404-410},

abstract = {Pyrrolysine (Pyl), the 22nd canonical amino acid, is only decoded and synthesized by a limited number of organisms in the domains Archaea and Bacteria. Pyl is encoded by the amber codon UAG, typically a stop codon. To date, all known Pyl-decoding archaea are able to carry out methylotrophic methanogenesis. The functionality of methylamine methyltransferases, an important component of corrinoid-dependent methyltransfer reactions, depends on the presence of Pyl. Here, we present a putative pyl gene cluster obtained from single-cell genomes of the archaeal Mediterranean Sea Brine Lakes group 1 (MSBL1) from the Red Sea. Functional annotation of the MSBL1 single cell amplified genomes (SAGs) also revealed a complete corrinoid-dependent methyl-transfer pathway suggesting that members of MSBL1 may possibly be capable of synthesizing Pyl and metabolizing methylated amines.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{Z2017,

title = {SAR202 Genomes from the Dark Ocean Predict Pathways for the Oxidation of Recalcitrant Dissolved Organic Matter},

author = {Landry Z and Swan BK and Herndl GJ and Stepanauskas R and Giovannoni SJ},

url = {http://mbio.asm.org/content/8/2/e00413-17.full},

doi = {10.1128/mBio.00413-17},

year  = {2017},

date = {2017-04-18},

journal = {mBio},

abstract = {Deep-ocean regions beyond the reach of sunlight contain an estimated 615 Pg of dissolved organic matter (DOM), much of which persists for thousands of years. It is thought that bacteria oxidize DOM until it is too dilute or refractory to support microbial activity. We analyzed five single-amplified genomes (SAGs) from the abundant SAR202 clade of dark-ocean bacterioplankton and found they encode multiple families of paralogous enzymes involved in carbon catabolism, including several families of oxidative enzymes that we hypothesize participate in the degradation of cyclic alkanes. The five partial genomes encoded 152 flavin mononucleotide/F420-dependent monooxygenases (FMNOs), many of which are predicted to be type II Baeyer-Villiger monooxygenases (BVMOs) that catalyze oxygen insertion into semilabile alicyclic alkanes. The large number of oxidative enzymes, as well as other families of enzymes that appear to play complementary roles in catabolic pathways, suggests that SAR202 might catalyze final steps in the biological oxidation of relatively recalcitrant organic compounds to refractory compounds that persist.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{SP2017,

title = {Genomic comparisons of a bacterial lineage that inhabits both marine and terrestrial deep subsurface systems},

author = {Jungbluth SP and del Rio TG and Tringe SG and Stepanauskas R and Rapp\'{e} MS},

url = {https://peerj.com/articles/3134/},

doi = {10.7717/peerj.3134},

year  = {2017},

date = {2017-04-06},

journal = {PeerJ},

abstract = {It is generally accepted that diverse, poorly characterized microorganisms reside deep within Earth’s crust. One such lineage of deep subsurface-dwelling bacteria is an uncultivated member of the Firmicutes phylum that can dominate molecular surveys from both marine and continental rock fracture fluids, sometimes forming the sole member of a single-species microbiome. Here, we reconstructed a genome from basalt-hosted fluids of the deep subseafloor along the eastern Juan de Fuca Ridge flank and used a phylogenomic analysis to show that, despite vast differences in geographic origin and habitat, it forms a monophyletic clade with the terrestrial deep subsurface genome of “Candidatus Desulforudis audaxviator” MP104C. While a limited number of differences were observed between the marine genome of “Candidatus Desulfopertinax cowenii” modA32 and its terrestrial relative that may be of potential adaptive importance, here it is revealed that the two are remarkably similar thermophiles possessing the genetic capacity for motility, sporulation, hydrogenotrophy, chemoorganotrophy, dissimilatory sulfate reduction, and the ability to fix inorganic carbon via the Wood-Ljungdahl pathway for chemoautotrophic growth. Our results provide insights into the genetic repertoire within marine and terrestrial members of a bacterial lineage that is widespread in the global deep subsurface biosphere, and provides a natural means to investigate adaptations specific to these two environments.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{P2017c,

title = {Microbial community assembly and evolution in subseafloor sediment},

author = {Starnawski P and Bataillon T and Ettema T.J.G and Jochum L.M and Schreiber L and Chen X and Lever M, Ploz M, J\orgensen B and Schramm A and Kjeldsen K },

url = {http://www.pnas.org/content/114/11/2940},

doi = {https://doi.org/10.1073/pnas.1614190114},

year  = {2017},

date = {2017-03-14},

journal = {PNAS},

volume = {114},

number = {11},

pages = {2940-2945},

abstract = {Bacterial and archaeal communities inhabiting the subsurface seabed live under strong energy limitation and have growth rates that are orders of magnitude slower than laboratory-grown cultures. It is not understood how subsurface microbial communities are assembled and whether populations undergo adaptive evolution or accumulate mutations as a result of impaired DNA repair under such energy-limited conditions. Here we use amplicon sequencing to explore changes of microbial communities during burial and isolation from the surface to the >5,000-y-old subsurface of marine sediment and identify a small core set of mostly uncultured bacteria and archaea that is present throughout the sediment column. These persisting populations constitute a small fraction of the entire community at the surface but become predominant in the subsurface. We followed patterns of genome diversity with depth in four dominant lineages of the persisting populations by mapping metagenomic sequence reads onto single-cell genomes. Nucleotide sequence diversity was uniformly low and did not change with age and depth of the sediment. Likewise, there was no detectable change in mutation rates and efficacy of selection. Our results indicate that subsurface microbial communities predominantly assemble by selective survival of taxa able to persist under extreme energy limitation.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{JF2017,

title = {Accessing the genomic information of unculturable oceanic picoeukaryotes by combining multiple single cells},

author = {Mangot JF and Logares R and Sanchez P and Latorre F and Seeleuthner Y and Mondy S and Sieracki ME and Jaillon O and Wincker P and Vargas C and Massana R},

url = {https://www.nature.com/articles/srep41498},

doi = {doi:10.1038/srep41498},

year  = {2017},

date = {2017-01-27},

journal = {Scientific Reports},

volume = {7},

number = {41498},

abstract = {Pico-sized eukaryotes play key roles in the functioning of marine ecosystems, but we still have a limited knowledge on their ecology and evolution. The MAST-4 lineage is of particular interest, since it is widespread in surface oceans, presents ecotypic differentiation and has defied culturing efforts so far. Single cell genomics (SCG) are promising tools to retrieve genomic information from these uncultured organisms. However, SCG are based on whole genome amplification, which normally introduces amplification biases that limit the amount of genomic data retrieved from a single cell. Here, we increase the recovery of genomic information from two MAST-4 lineages by co-assembling short reads from multiple Single Amplified Genomes (SAGs) belonging to evolutionary closely related cells. We found that complementary genomic information is retrieved from different SAGs, generating co-assembly that features >74% of genome recovery, against about 20% when assembled individually. Even though this approach is not aimed at generating high-quality draft genomes, it allows accessing to the genomic information of microbes that would otherwise remain unreachable. Since most of the picoeukaryotes still remain uncultured, our work serves as a proof-of-concept that can be applied to other taxa in order to extract genomic data and address new ecological and evolutionary questions.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{MJ2017,

title = {Endozoicomonas genomes reveal functional adaptation and plasticity in bacterial strains symbiotically associated with diverse marine hosts},

author = {Neave MJ and Michell CT and Apprill A and Voolstra CR},

url = {https://www.nature.com/articles/srep40579?WT.feed_name=subjects_environmental-microbiology},

doi = {doi:10.1038/srep40579},

year  = {2017},

date = {2017-01-17},

journal = {Scientific Reports},

volume = {7},

number = {40579},

abstract = {Endozoicomonas bacteria are globally distributed and often abundantly associated with diverse marine hosts including reef-building corals, yet their function remains unknown. In this study we generated novel Endozoicomonas genomes from single cells and metagenomes obtained directly from the corals Stylophora pistillata, Pocillopora verrucosa, and Acropora humilis. We then compared these culture-independent genomes to existing genomes of bacterial isolates acquired from a sponge, sea slug, and coral to examine the functional landscape of this enigmatic genus. Sequencing and analysis of single cells and metagenomes resulted in four novel genomes with 60\textendash76% and 81\textendash90% genome completeness, respectively. These data also confirmed that Endozoicomonas genomes are large and are not streamlined for an obligate endosymbiotic lifestyle, implying that they have free-living stages. All genomes show an enrichment of genes associated with carbon sugar transport and utilization and protein secretion, potentially indicating that Endozoicomonas contribute to the cycling of carbohydrates and the provision of proteins to their respective hosts. Importantly, besides these commonalities, the genomes showed evidence for differential functional specificity and diversification, including genes for the production of amino acids. Given this metabolic diversity of Endozoicomonas we propose that different genotypes play disparate roles and have diversified in concert with their hosts.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{J2016,

title = {Strategies to improve reference databases for soil microbiomes},

author = {Choi J and Yang F and Stepanauskas R and Cardenas E and Garoutte A and Williams R and Flater J and Tiedje JM and Hofmockel KS and 6 and Gelder B and Howe A},

url = {https://www.nature.com/ismej/journal/v11/n4/full/ismej2016168a.html},

doi = { doi:10.1038/ismej.2016.168},

year  = {2016},

date = {2016-12-09},

journal = {ISME},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{T2016,

title = {Survey of the green picoalga Bathycoccus genomes in the global ocean},

author = {Vannier T and Leconte J and Seeleuthner Y and Mondy S and Pelletier E and Aury JM and De Vargas C and Sieracki M and Iudicone D and Vaulot D and Wincker P and Jaillon O},

url = {https://www.nature.com/articles/srep37900},

doi = {10.1038/srep37900},

year  = {2016},

date = {2016-11-30},

journal = {Scientific Reports},

volume = {6},

abstract = {Bathycoccus is a cosmopolitan green micro-alga belonging to the Mamiellophyceae, a class of picophytoplankton that contains important contributors to oceanic primary production. A single species of Bathycoccus has been described while the existence of two ecotypes has been proposed based on metagenomic data. A genome is available for one strain corresponding to the described phenotype. We report a second genome assembly obtained by a single cell genomics approach corresponding to the second ecotype. The two Bathycoccus genomes are divergent enough to be unambiguously distinguishable in whole DNA metagenomic data although they possess identical sequence of the 18S rRNA gene including in the V9 region. Analysis of 122 global ocean whole DNA metagenome samples from the Tara-Oceans expedition reveals that populations of Bathycoccus that were previously identified by 18S rRNA V9 metabarcodes are only composed of these two genomes. Bathycoccus is relatively abundant and widely distributed in nutrient rich waters. The two genomes rarely co-occur and occupy distinct oceanic niches in particular with respect to depth. Metatranscriptomic data provide evidence for gain or loss of highly expressed genes in some samples, suggesting that the gene repertoire is modulated by environmental conditions.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{MJ2016,

title = {Differential specificity between closely related corals and abundant Endozoicomonas endosymbionts across global scales},

author = {Neave MJ and Rachmawati R and Xun L and Michell CT and Bourne DG and Apprill A and Voolstra CR},

url = {https://www.nature.com/ismej/journal/v11/n1/full/ismej201695a.html},

doi = {doi:10.1038/ismej.2016.95},

year  = {2016},

date = {2016-07-08},

journal = {ISME},

abstract = {Reef-building corals are well regarded not only for their obligate association with endosymbiotic algae, but also with prokaryotic symbionts, the specificity of which remains elusive. To identify the central microbial symbionts of corals, their specificity across species and conservation over geographic regions, we sequenced partial SSU ribosomal RNA genes of Bacteria and Archaea from the common corals Stylophora pistillata and Pocillopora verrucosa across 28 reefs within seven major geographical regions. We demonstrate that both corals harbor Endozoicomonas bacteria as their prevalent symbiont. Importantly, catalyzed reporter deposition\textendashfluorescence in situ hybridization (CARD\textendashFISH) with Endozoicomonas-specific probes confirmed their residence as large aggregations deep within coral tissues. Using fine-scale genotyping techniques and single-cell genomics, we demonstrate that P. verrucosa harbors the same Endozoicomonas, whereas S. pistillata associates with geographically distinct genotypes. This specificity may be shaped by the different reproductive strategies of the hosts, potentially uncovering a pattern of symbiont selection that is linked to life history. Spawning corals such as P. verrucosa acquire prokaryotes from the environment. In contrast, brooding corals such as S. pistillata release symbiont-packed planula larvae, which may explain a strong regional signature in their microbiome. Our work contributes to the factors underlying microbiome specificity and adds detail to coral holobiont functioning.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{MJ2016b,

title = {Differential specificity between closely related corals and abundant Endozoicomonas endosymbionts across global scales},

author = {Neave MJ and Rachmawati R and Xun L and Michell CT and Bourne DG and Apprill A and Voolstra CR},

url = {https://www.nature.com/articles/ismej201695},

doi = {10.1038/ismej.2016.95},

year  = {2016},

date = {2016-07-08},

journal = {The ISME Journal},

volume = {11},

pages = {186-200},

abstract = {Reef-building corals are well regarded not only for their obligate association with endosymbiotic algae, but also with prokaryotic symbionts, the specificity of which remains elusive. To identify the central microbial symbionts of corals, their specificity across species and conservation over geographic regions, we sequenced partial SSU ribosomal RNA genes of Bacteria and Archaea from the common corals Stylophora pistillata and Pocillopora verrucosa across 28 reefs within seven major geographical regions. We demonstrate that both corals harbor Endozoicomonas bacteria as their prevalent symbiont. Importantly, catalyzed reporter deposition\textendashfluorescence in situ hybridization (CARD\textendashFISH) with Endozoicomonas-specific probes confirmed their residence as large aggregations deep within coral tissues. Using fine-scale genotyping techniques and single-cell genomics, we demonstrate that P. verrucosa harbors the same Endozoicomonas, whereas S. pistillata associates with geographically distinct genotypes. This specificity may be shaped by the different reproductive strategies of the hosts, potentially uncovering a pattern of symbiont selection that is linked to life history. Spawning corals such as P. verrucosa acquire prokaryotes from the environment. In contrast, brooding corals such as S. pistillata release symbiont-packed planula larvae, which may explain a strong regional signature in their microbiome. Our work contributes to the factors underlying microbiome specificity and adds detail to coral holobiont functioning.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{K2016,

title = {Single-Cell Genome and Group-Specific dsrAB Sequencing Implicate Marine Members of the Class Dehalococcoidia (Phylum Chloroflexi) in Sulfur Cycling},

author = {Wasmund K and Cooper M and Schreiber L and Lloyd KG and Baker BJ and Petersen DJ and J\orgensen BB and Stepanauskas R and Reinhardt R and Schramm A and Loy A and Adrian L},

url = {http://mbio.asm.org/content/7/3/e00266-16.full},

doi = {10.1128/mBio.00266-16},

year  = {2016},

date = {2016-05-03},

journal = {mBio},

volume = {7},

number = {3},

abstract = {

The marine subsurface sediment biosphere is widely inhabited by bacteria affiliated with the class Dehalococcoidia (DEH), phylum Chloroflexi, and yet little is known regarding their metabolisms. In this report, genomic content from a single DEH cell (DEH-C11) with a 16S rRNA gene that was affiliated with a diverse cluster of 16S rRNA gene sequences prevalent in marine sediments was obtained from sediments of Aarhus Bay, Denmark. The distinctive gene content of this cell suggests metabolic characteristics that differ from those of known DEH and Chloroflexi. The presence of genes encoding dissimilatory sulfite reductase (Dsr) suggests that DEH could respire oxidized sulfur compounds, although Chloroflexi have never been implicated in this mode of sulfur cycling. Using long-range PCR assays targeting DEH dsr loci, dsrAB genes were amplified and sequenced from various marine sediments. Many of the amplified dsrAB sequences were affiliated with the DEH Dsr clade, which we propose equates to a family-level clade. This provides supporting evidence for the potential for sulfite reduction by diverse DEH species. DEH-C11 also harbored genes encoding reductases for arsenate, dimethyl sulfoxide, and halogenated organics. The reductive dehalogenase homolog (RdhA) forms a monophyletic clade along with RdhA sequences from various DEH-derived contigs retrieved from available metagenomes. Multiple facts indicate that this RdhA may not be a terminal reductase. The presence of other genes indicated that nutrients and energy may be derived from the oxidation of substituted homocyclic and heterocyclic aromatic compounds. Together, these results suggest that marine DEH play a previously unrecognized role in sulfur cycling and reveal the potential for expanded catabolic and respiratory functions among subsurface DEH.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{H2016,

title = {Comparative Single-Cell Genomics of Chloroflexi from the Okinawa Trough Deep Subsurface Biosphere},

author = {Fullerton H and Moyer CL},

url = {http://aem.asm.org/content/early/2016/03/07/AEM.00624-16},

doi = {10.1128/AEM.00624-16},

year  = {2016},

date = {2016-03-11},

journal = {Applied and Environmental Microbiology},

volume = {82},

pages = {3000-3008},

abstract = {Chloroflexi SSU rRNA gene sequences are frequently recovered from subseafloor environments, but the metabolic potential of this phylum is poorly understood. The phylum Chloroflexi is represented by isolates with diverse metabolic strategies including, anoxic phototrophy, fermentation and reductive dehalogenation; therefore, function cannot be attributed to these organisms based solely on phylogeny. Single cell genomics can provide metabolic insights into uncultured organisms, like the deep-subsurface Chloroflexi. Nine SSU rRNA gene sequences were identified from single-cell sorts of whole-round core material collected as part of IODP Expedition 331 (Deep Hot Biosphere) from the Okinawa Trough at Iheya North hydrothermal field. Previous studies of subsurface Chloroflexi single amplified genomes (SAGs) suggest heterotrophic or lithotrophic metabolisms and provide no evidence for growth by reductive dehalogenation. Our nine Chloroflexi SAGs (of which seven are Anaerolineales) indicate that in addition to encoding genes for the Wood-Ljungdahl pathway, exogenous carbon sources can be actively transported into the cells. At least one subunit for the pyruvate ferredoxin oxidoreductase was found in four of the Chloroflexi SAGs. This protein can provide a link between the Wood-Ljungdahl pathway and other carbon anabolic pathways. Finally, one of the seven Anaerolineales SAGs contains a distinct reductive dehalogenase homologous (rdhA) gene; suggesting reductive dehalogenation is not limited to the Dehalococcoidia class of Chloroflexi.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{BJ2016,

title = {Genomic inference of the metabolism of cosmopolitan subsurface Archaea, Hadesarchaea},

author = {Baker BJ and Saw JH and Lind AE and Lazar CS and Hinrichs KU and Teske AP and Ettema TJ},

url = {https://www.nature.com/articles/nmicrobiol20162},

doi = {10.1038/nmicrobiol.2016.2},

year  = {2016},

date = {2016-02-15},

journal = {Nature Microbiology},

volume = {1},

number = {16002},

abstract = {The subsurface biosphere is largely unexplored and contains a broad diversity of uncultured microbes1. Despite being one of the few prokaryotic lineages that is cosmopolitan in both the terrestrial and marine subsurface2,​3,​4, the physiological and ecological roles of SAGMEG (South-African Gold Mine Miscellaneous Euryarchaeal Group) Archaea are unknown. Here, we report the metabolic capabilities of this enigmatic group as inferred from genomic reconstructions. Four high-quality (63\textendash90% complete) genomes were obtained from White Oak River estuary and Yellowstone National Park hot spring sediment metagenomes. Phylogenomic analyses place SAGMEG Archaea as a deeply rooting sister clade of the Thermococci, leading us to propose the name Hadesarchaea for this new Archaeal class. With an estimated genome size of around 1.5 Mbp, the genomes of Hadesarchaea are distinctly streamlined, yet metabolically versatile. They share several physiological mechanisms with strict anaerobic Euryarchaeota. Several metabolic characteristics make them successful in the subsurface, including genes involved in CO and H2 oxidation (or H2 production), with potential coupling to nitrite reduction to ammonia (DNRA). This first glimpse into the metabolic capabilities of these cosmopolitan Archaea suggests they are mediating key geochemical processes and are specialized for survival in the subsurface biosphere.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{S2016,

title = {Ubiquitous Gammaproteobacteria dominate dark carbon fixation in coastal sediments},

author = {Dyksma S and Bischof K and Fuchs BM and Hoffmann K and Meier D and Meyerdierks A and Pjevac P and Probandt D and Richter M and Stepanauskas R and Mu\ssmann M},

url = {https://www.nature.com/articles/ismej2015257},

doi = {doi:10.1038/ismej.2015.257},

year  = {2016},

date = {2016-02-12},

journal = {ISME},

volume = {10},

pages = {1939\textendash1953},

abstract = {Marine sediments are the largest carbon sink on earth. Nearly half of dark carbon fixation in the oceans occurs in coastal sediments, but the microorganisms responsible are largely unknown. By integrating the 16S rRNA approach, single-cell genomics, metagenomics and transcriptomics with 14C-carbon assimilation experiments, we show that uncultured Gammaproteobacteria account for 70\textendash86% of dark carbon fixation in coastal sediments. First, we surveyed the bacterial 16S rRNA gene diversity of 13 tidal and sublittoral sediments across Europe and Australia to identify ubiquitous core groups of Gammaproteobacteria mainly affiliating with sulfur-oxidizing bacteria. These also accounted for a substantial fraction of the microbial community in anoxic, 490-cm-deep subsurface sediments. We then quantified dark carbon fixation by scintillography of specific microbial populations extracted and flow-sorted from sediments that were short-term incubated with 14C-bicarbonate. We identified three distinct gammaproteobacterial clades covering diversity ranges on family to order level (the Acidiferrobacter, JTB255 and SSr clades) that made up >50% of dark carbon fixation in a tidal sediment. Consistent with these activity measurements, environmental transcripts of sulfur oxidation and carbon fixation genes mainly affiliated with those of sulfur-oxidizing Gammaproteobacteria. The co-localization of key genes of sulfur and hydrogen oxidation pathways and their expression in genomes of uncultured Gammaproteobacteria illustrates an unknown metabolic plasticity for sulfur oxidizers in marine sediments. Given their global distribution and high abundance, we propose that a stable assemblage of metabolically flexible Gammaproteobacteria drives important parts of marine carbon and sulfur cycles.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{Y2016,

title = {Ecological Genomics of the Uncultivated Marine Roseobacter Lineage CHAB-I-5},

author = {Zhang Y and Sun Y and Jiao N and Stepanauskas R and Luo H},

url = {http://aem.asm.org/content/82/7/2100.long},

doi = {10.1128/AEM.03678-15},

year  = {2016},

date = {2016-01-29},

journal = {Applied and Environmental Microbiology},

volume = {82},

number = {7},

abstract = {Members of the marine Roseobacter clade are major participants in global carbon and sulfur cycles. While roseobacters are well represented in cultures, several abundant pelagic lineages, including SAG-O19, DC5-80-3, and NAC11-7, remain largely uncultivated and show evidence of genome streamlining. Here, we analyzed the partial genomes of three single cells affiliated with CHAB-I-5, another abundant but exclusively uncultivated Roseobacter lineage. Members of this lineage encode several metabolic potentials that are absent in streamlined genomes. Examples are quorum sensing and type VI secretion systems, which enable them to effectively interact with host and other bacteria. Further analysis of the CHAB-I-5 single-cell amplified genomes (SAGs) predicted that this lineage comprises members with relatively large genomes (4.1 to 4.4 Mbp) and a high fraction of noncoding DNA (10 to 12%), which is similar to what is observed in many cultured, nonstreamlined Roseobacter lineages. The four uncultured lineages, while exhibiting highly variable geographic distributions, together represent >60% of the global pelagic roseobacters. They are consistently enriched in genes encoding the capabilities of light harvesting, oxidation of “energy-rich” reduced sulfur compounds and methylated amines, uptake and catabolism of various carbohydrates and osmolytes, and consumption of abundant exudates from phytoplankton. These traits may define the global prevalence of the four lineages among marine bacterioplankton.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{A2016,

title = {Tuning fresh: radiation through rewiring of central metabolism in streamlined bacteria},

author = {Eiler A and Mondav R and Sinclair L and Fernandez-Vidal L and Scofield DG and Schwientek P and Martinez-Garcia M and Torrents D and McMahon KD and Andersson SGE and Stepanauskas R and Woyke T and Bertilsson S},

doi = {doi:10.1038/ismej.2015.260},

year  = {2016},

date = {2016-01-19},

journal = {ISME},

volume = {10},

pages = {1902\textendash1914},

abstract = {Most free-living planktonic cells are streamlined and in spite of their limitations in functional flexibility, their vast populations have radiated into a wide range of aquatic habitats. Here we compared the metabolic potential of subgroups in the Alphaproteobacteria lineage SAR11 adapted to marine and freshwater habitats. Our results suggest that the successful leap from marine to freshwaters in SAR11 was accompanied by a loss of several carbon degradation pathways and a rewiring of the central metabolism. Examples for these are C1 and methylated compounds degradation pathways, the Entner\textendashDoudouroff pathway, the glyoxylate shunt and anapleuretic carbon fixation being absent from the freshwater genomes. Evolutionary reconstructions further suggest that the metabolic modules making up these important freshwater metabolic traits were already present in the gene pool of ancestral marine SAR11 populations. The loss of the glyoxylate shunt had already occurred in the common ancestor of the freshwater subgroup and its closest marine relatives, suggesting that the adaptation to freshwater was a gradual process. Furthermore, our results indicate rapid evolution of TRAP transporters in the freshwater clade involved in the uptake of low molecular weight carboxylic acids. We propose that such gradual tuning of metabolic pathways and transporters toward locally available organic substrates is linked to the formation of subgroups within the SAR11 clade and that this process was critical for the freshwater clade to find and fix an adaptive phenotype.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{A2015,

title = {De novo synthesis and functional analysis of the phosphatase-encoding gene acI-B of uncultured Actinobacteria from Lake Stechlin (NE Germany)},

author = {Srivastava A and McMahon KD and Stepanauskas R and Grossart HP},

url = {http://dx.doi.org/10.2436/20.1501.01.262},

doi = {10.2436/20.1501.01.262},

year  = {2015},

date = {2015-12-18},

journal = {International Microbiology},

volume = {19},

number = {39-47},

abstract = {The National Center for Biotechnology Information [http://www.ncbi.nlm.nih.gov/guide/taxonomy/] database enlists more than 15,500 bacterial species. But this also includes a plethora of uncultured bacterial representations. Owing to their metabolism, they directly influence biogeochemical cycles, which underscores the the important status of bacteria on our planet. To study the function of a gene from an uncultured bacterium, we have undertaken a de novo gene synthesis approach. Actinobacteria of the acI-B subcluster are important but yet uncultured members of the bacterioplankton in temperate lakes of the northern hemisphere such as oligotrophic Lake Stechlin (NE Germany). This lake is relatively poor in phosphate (P) and harbors on average ~1.3 x 10 6 bacterial cells/ml, whereby Actinobacteria of the ac-I lineage can contribute to almost half of the entire bacterial community depending on seasonal variability. Single cell genome analysis of Actinobacterium SCGC AB141-P03, a member of the acI-B tribe in Lake Stechlin has revealed several phosphate-metabolizing genes. The genome of acI-B Actinobacteria indicates potential to degrade polyphosphate compound. To test for this genetic potential, we targeted the exoP-annotated gene potentially encoding polyphosphatase and synthesized it artificially to examine its biochemical role. Heterologous overexpression of the gene in Escherichia coli and protein purification revealed phosphatase activity. Comparative genome analysis suggested that homologs of this gene should be also present in other Actinobacteria of the acI lineages. This strategic retention of specialized genes in their genome provides a metabolic advantage over other members of the aquatic food web in a P-limited ecosystem.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{DK2015,

title = {Diversification and niche adaptations of Nitrospina-like bacteria in the polyextreme interfaces of Red Sea brines},

author = {Ngugi DK and Blom J and Stepanauskas R and Stingl U},

url = {https://www.nature.com/articles/ismej2015214},

doi = {10.1038/ismej.2015.214},

year  = {2015},

date = {2015-12-15},

journal = {ISME},

volume = {10},

pages = {1383\textendash1399},

abstract = {Nitrite-oxidizing bacteria (NOB) of the genus Nitrospina have exclusively been found in marine environments. In the brine\textendashseawater interface layer of Atlantis II Deep (Red Sea), Nitrospina-like bacteria constitute up to one-third of the bacterial 16S ribosomal RNA (rRNA) gene sequences. This is much higher compared with that reported in other marine habitats (~10% of all bacteria), and was unexpected because no NOB culture has been observed to grow above 4.0% salinity, presumably due to the low net energy gained from their metabolism that is insufficient for both growth and osmoregulation. Using phylogenetics, single-cell genomics and metagenomic fragment recruitment approaches, we document here that these Nitrospina-like bacteria, designated as Candidatus Nitromaritima RS, are not only highly diverged from the type species Nitrospina gracilis (pairwise genome identity of 69%) but are also ubiquitous in the deeper, highly saline interface layers (up to 11.2% salinity) with temperatures of up to 52 °C. Comparative pan-genome analyses revealed that less than half of the predicted proteome of Ca. Nitromaritima RS is shared with N. gracilis. Interestingly, the capacity for nitrite oxidation is also conserved in both genomes. Although both lack acidic proteomes synonymous with extreme halophiles, the pangenome of Ca. Nitromaritima RS specifically encodes enzymes with osmoregulatory and thermoprotective roles (i.e., ectoine/hydroxyectoine biosynthesis) and of thermodynamic importance (i.e., nitrate and nitrite reductases). Ca. Nitromaritima RS also possesses many hallmark traits of microaerophiles and high-affinity NOB. The abundance of the uncultured Ca. Nitromaritima lineage in marine oxyclines suggests their unrecognized ecological significance in deoxygenated areas of the global ocean.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{JH2015,

title = {Nanoarchaeota, Their Sulfolobales Host, and Nanoarchaeota Virus Distribution across Yellowstone National Park Hot Springs},

author = {Munson-McGee JH and Field EK and Bateson M and Rooney C and Stepanauskas R and Young MJ},

url = {http://aem.asm.org/content/81/22/7860.short},

doi = {10.1128/AEM.01539-15},

year  = {2015},

date = {2015-09-04},

journal = {Applied and Environmental Microbiology},

abstract = {Nanoarchaeota are obligate symbionts with reduced genomes first described from marine thermal vent environments. Here, both community metagenomics and single-cell analysis revealed the presence of Nanoarchaeota in high-temperature (∼90°C), acidic (pH ≈ 2.5 to 3.0) hot springs in Yellowstone National Park (YNP) (United States). Single-cell genome analysis of two cells resulted in two nearly identical genomes, with an estimated full length of 650 kbp. Genome comparison showed that these two cells are more closely related to the recently proposed Nanobsidianus stetteri from a more neutral YNP hot spring than to the marine Nanoarchaeum equitans. Single-cell and catalyzed reporter deposition-fluorescence in situ hybridization (CARD-FISH) analysis of environmental hot spring samples identified the host of the YNP Nanoarchaeota as a Sulfolobales species known to inhabit the hot springs. Furthermore, we demonstrate that Nanoarchaeota are widespread in acidic to near neutral hot springs in YNP. An integrated viral sequence was also found within one Nanoarchaeota single-cell genome and further analysis of the purified viral fraction from environmental samples indicates that this is likely a virus replicating within the YNP Nanoarchaeota.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{SA2015,

title = {Abundant Atribacteria in deep marine sediment from the Ad\'{e}lie Basin, Antarctica},

author = {Carr SA and Orcutt BN and Mandernack KW and Spear JR},

url = {https://www.frontiersin.org/articles/10.3389/fmicb.2015.00872/full},

doi = {10.3389/fmicb.2015.00872},

year  = {2015},

date = {2015-08-26},

journal = {Frontiers in Microbiology},

abstract = {Bacteria belonging to the newly classified candidate phylum “Atribacteria” (formerly referred to as “OP9” and “JS1”) are common in anoxic methane-rich sediments. However, the metabolic functions and biogeochemical role of these microorganisms in the subsurface remains unrealized due to the lack of pure culture representatives. In this study of deep sediment from Antarctica’s Ad\'{e}lie Basin, collected during Expedition 318 of the Integrated Ocean Drilling Program (IODP), Atribacteria-related sequences of the 16S rRNA gene were abundant (up to 51% of the sequences) and steadily increased in relative abundance with depth throughout the methane-rich zones. To better understand the metabolic potential of Atribacteria within this environment, and to compare with phylogenetically distinct Atribacteria from non-deep-sea environments, individual cells were sorted for single cell genomics from sediment collected from 97.41 m below the seafloor from IODP Hole U1357C. As observed for non-marine Atribacteria, a partial single cell genome suggests a heterotrophic metabolism, with Atribacteria potentially producing fermentation products such as acetate, ethanol, and CO2. These products may in turn support methanogens within the sediment microbial community and explain the frequent occurrence of Atribacteria in anoxic methane-rich sediments. This first report of a single cell genome from deep sediment broadens the known diversity within the Atribacteria phylum and highlights the potential role of Atribacteria in carbon cycling in deep sediment.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{SA2015b,

title = {Abundant Atribacteria in deep marine sediment from the Ad\'{e}lie Basin, Antarctica},

author = {Carr SA and Orcutt BN and Mandernack KW and Spear JR},

url = {https://www.frontiersin.org/articles/10.3389/fmicb.2015.00872/full},

doi = {10.3389/fmicb.2015.00872},

year  = {2015},

date = {2015-08-25},

journal = {Frontiers in Microbiology},

volume = {6},

abstract = {Bacteria belonging to the newly classified candidate phylum "Atribacteria" (formerly referred to as "OP9" and "JS1") are common in anoxic methane-rich sediments. However, the metabolic functions and biogeochemical role of these microorganisms in the subsurface remains unrealized due to the lack of pure culture representatives. In this study of deep sediment from Antarctica's Ad\'{e}lie Basin, collected during Expedition 318 of the Integrated Ocean Drilling Program (IODP), Atribacteria-related sequences of the 16S rRNA gene were abundant (up to 51% of the sequences) and steadily increased in relative abundance with depth throughout the methane-rich zones. To better understand the metabolic potential of Atribacteria within this environment, and to compare with phylogenetically distinct Atribacteria from non-deep-sea environments, individual cells were sorted for single cell genomics from sediment collected from 97.41 m below the seafloor from IODP Hole U1357C. As observed for non-marine Atribacteria, a partial single cell genome suggests a heterotrophic metabolism, with Atribacteria potentially producing fermentation products such as acetate, ethanol, and CO2. These products may in turn support methanogens within the sediment microbial community and explain the frequent occurrence of Atribacteria in anoxic methane-rich sediments. This first report of a single cell genome from deep sediment broadens the known diversity within the Atribacteria phylum and highlights the potential role of Atribacteria in carbon cycling in deep sediment.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{JM2015b,

title = {Single cell genomics indicates horizontal gene transfer and viral infections in a deep subsurface Firmicutes population},

author = {Labont\'{e} JM and Field EK and Lau M and Chivian D and van Heerden E and Wommack KE and Kieft TL and Onstott TC and Stepanauskas R},

url = {https://www.frontiersin.org/articles/10.3389/fmicb.2015.00349/full},

doi = {10.3389/fmicb.2015.00349},

year  = {2015},

date = {2015-04-22},

journal = {Frontiers in Microbiology},

abstract = {A major fraction of Earth's prokaryotic biomass dwells in the deep subsurface, where cellular abundances per volume of sample are lower, metabolism is slower, and generation times are longer than those in surface terrestrial and marine environments. How these conditions impact biotic interactions and evolutionary processes is largely unknown. Here we employed single cell genomics to analyze cell-to-cell genome content variability and signatures of horizontal gene transfer (HGT) and viral infections in five cells of Candidatus Desulforudis audaxviator, which were collected from a 3 km-deep fracture water in the 2.9 Ga-old Witwatersrand Basin of South Africa. Between 0 and 32% of genes recovered from single cells were not present in the original, metagenomic assembly of Desulforudis, which was obtained from a neighboring subsurface fracture. We found a transposable prophage, a retron, multiple clustered regularly interspaced short palindromic repeats (CRISPRs) and restriction-modification systems, and an unusually high frequency of transposases in the analyzed single cell genomes. This indicates that recombination, HGT and viral infections are prevalent evolutionary events in the studied population of microorganisms inhabiting a highly stable deep subsurface environment.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{JM2015,

title = {Single-cell genomics-based analysis of virus\textendashhost interactions in marine surface bacterioplankton},

author = {Labont\'{e} JM and Swan BK and Poulos B and Luo H and Koren S and Hallam SJ and Sullivan MB and Woyke T and Wommack KE and Stepanauskas R},

url = {https://www.nature.com/articles/ismej201548},

doi = {10.1038/ismej.2015.48},

year  = {2015},

date = {2015-04-17},

journal = {ISME},

abstract = {Viral infections dynamically alter the composition and metabolic potential of marine microbial communities and the evolutionary trajectories of host populations with resulting feedback on biogeochemical cycles. It is quite possible that all microbial populations in the ocean are impacted by viral infections. Our knowledge of virus\textendashhost relationships, however, has been limited to a minute fraction of cultivated host groups. Here, we utilized single-cell sequencing to obtain genomic blueprints of viruses inside or attached to individual bacterial and archaeal cells captured in their native environment, circumventing the need for host and virus cultivation. A combination of comparative genomics, metagenomic fragment recruitment, sequence anomalies and irregularities in sequence coverage depth and genome recovery were utilized to detect viruses and to decipher modes of virus\textendashhost interactions. Members of all three tailed phage families were identified in 20 out of 58 phylogenetically and geographically diverse single amplified genomes (SAGs) of marine bacteria and archaea. At least four phage\textendashhost interactions had the characteristics of late lytic infections, all of which were found in metabolically active cells. One virus had genetic potential for lysogeny. Our findings include first known viruses of Thaumarchaeota, Marinimicrobia, Verrucomicrobia and Gammaproteobacteria clusters SAR86 and SAR92. Viruses were also found in SAGs of Alphaproteobacteria and Bacteroidetes. A high fragment recruitment of viral metagenomic reads confirmed that most of the SAG-associated viruses are abundant in the ocean. Our study demonstrates that single-cell genomics, in conjunction with sequence-based computational tools, enable in situ, cultivation-independent insights into host\textendashvirus interactions in complex microbial communities.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{R2015,

title = {Wiretapping into microbial interactions by single cell genomics},

author = {Stepanauskas R},

url = {https://www.frontiersin.org/articles/10.3389/fmicb.2015.00258/full},

doi = {10.3389/fmicb.2015.00258},

year  = {2015},

date = {2015-04-08},

journal = {Frontiers in Microbiology},

abstract = {Over the past decade, single cell genomics (SCG) made a swift transition from science fiction to a handy new tool in the biologist's toolset. The power of this technology lies in its ability to retrieve information-rich genomic blueprints from the most fundamental units of biological organization\textemdashindividual cells. This is particularly significant in the case of bacteria, archaea, and protists, where single cells constitute complete organisms. Such unicellular individuals comprise the vast majority of biological diversity and biomass of our planet, yet only a small fraction of microbial diversity has been discovered and studied. Together with other modern research tools, SCG has been increasingly instrumental in deciphering the genomic composition, metabolic potential and evolutionary histories of the “microbial dark matter.” While cultivation-free recovery of discrete genomes was impossible in 2004, by 2009 it became a routine procedure that is accessible to the broad research community through open-access SCG core facilities (e.g., scgc.bigelow.org). This rapid development has enabled genomic studies of many previously unexplored branches of the tree of life (Marcy et al., 2007; Rinke et al., 2013) and findings of hitherto unrecognized biogeochemical processes and ecological patterns (Swan et al., 2011, 2013; Mason et al., 2012), paving the way for a new wave of discovery in microbiology and biotechnology.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{J2015,

title = {Single-cell genomics of a rare environmental alphaproteobacterium provides unique insights into Rickettsiaceae evolution},

author = {Martijn J and Schullz F and Zaremba-Niedzwiedzka K and Viklund J and Stepanauskas R and Andersson SGE and Horn M and Guy L and Ettema TJG},

url = {https://www.nature.com/articles/ismej201546},

doi = {10.1038/ismej.2015.46},

year  = {2015},

date = {2015-04-07},

journal = {ISME},

abstract = {The bacterial family Rickettsiaceae includes a group of well-known etiological agents of many human and vertebrate diseases, including epidemic typhus-causing pathogen Rickettsia prowazekii. Owing to their medical relevance, rickettsiae have attracted a great deal of attention and their host-pathogen interactions have been thoroughly investigated. All known members display obligate intracellular lifestyles, and the best-studied genera, Rickettsia and Orientia, include species that are hosted by terrestrial arthropods. Their obligate intracellular lifestyle and host adaptation is reflected in the small size of their genomes, a general feature shared with all other families of the Rickettsiales. Yet, despite that the Rickettsiaceae and other Rickettsiales families have been extensively studied for decades, many details of the origin and evolution of their obligate host-association remain elusive. Here we report the discovery and single-cell sequencing of ‘Candidatus Arcanobacter lacustris’, a rare environmental alphaproteobacterium that was sampled from Damariscotta Lake that represents a deeply rooting sister lineage of the Rickettsiaceae. Intriguingly, phylogenomic and comparative analysis of the partial ‘Candidatus Arcanobacter lacustris’ genome revealed the presence chemotaxis genes and vertically inherited flagellar genes, a novelty in sequenced Rickettsiaceae, as well as several host-associated features. This finding suggests that the ancestor of the Rickettsiaceae might have had a facultative intracellular lifestyle. Our study underlines the efficacy of single-cell genomics for studying microbial diversity and evolution in general, and for rare microbial cells in particular.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{BG2015,

title = {Targeted diversity generation by intraterrestrial archaea and archaeal viruses},

author = {Paul BG, Bagby SC, Czornyj E, Arambula D, Handa S, Sczyrba A, Ghosh P, Miller JF, Valentine DL},

url = {https://www.nature.com/articles/ncomms7585},

doi = {10.1038/ncomms7585},

year  = {2015},

date = {2015-03-23},

journal = {Nature Communications},

abstract = {In the evolutionary arms race between microbes, their parasites, and their neighbours, the capacity for rapid protein diversification is a potent weapon. Diversity-generating retroelements (DGRs) use mutagenic reverse transcription and retrohoming to generate myriad variants of a target gene. Originally discovered in pathogens, these retroelements have been identified in bacteria and their viruses, but never in archaea. Here we report the discovery of intact DGRs in two distinct intraterrestrial archaeal systems: a novel virus that appears to infect archaea in the marine subsurface, and, separately, two uncultivated nanoarchaea from the terrestrial subsurface. The viral DGR system targets putative tail fibre ligand-binding domains, potentially generating >1018 protein variants. The two single-cell nanoarchaeal genomes each possess ≥4 distinct DGRs. Against an expected background of low genome-wide mutation rates, these results demonstrate a previously unsuspected potential for rapid, targeted sequence diversification in intraterrestrial archaea and their viruses.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{R2015b,

title = {Crystal Ball: Re-defining microbial diversity from its single-celled building blocks},

author = {Stepanauskas R},

url = {https://onlinelibrary.wiley.com/doi/abs/10.1111/1758-2229.12226},

doi = {10.1111/1758-2229.12226},

year  = {2015},

date = {2015-02-26},

journal = {Environmental Microbiology Reports},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{EK2014,

title = {Genomic insights into the uncultivated marine Zetaproteobacteria at Loihi Seamount},

author = {Field EK and Sczyrba A and Lyman AE and Harris CC and Woyke T and Stepanauskas R and Emerson D},

url = {https://www.nature.com/articles/ismej2014183},

doi = {10.1038/ismej.2014.183},

year  = {2014},

date = {2014-10-10},

journal = {ISME},

abstract = {The Zetaproteobacteria are a candidate class of marine iron-oxidizing bacteria that are typically found in high iron environments such as hydrothermal vent sites. As much remains unknown about these organisms due to difficulties in cultivation, single-cell genomics was used to learn more about this elusive group at Loihi Seamount. Comparative genomics of 23 phylogenetically diverse single amplified genomes (SAGs) and two isolates indicate niche specialization among the Zetaproteobacteria may be largely due to oxygen tolerance and nitrogen transformation capabilities. Only Form II ribulose 1,5-bisphosphate carboxylase (RubisCO) genes were found in the SAGs, suggesting that some of the uncultivated Zetaproteobacteria may be adapted to low oxygen and/or high carbon dioxide concentrations. There is also genomic evidence of oxygen-tolerant cytochrome c oxidases and oxidative stress-related genes, indicating that others may be exposed to higher oxygen conditions. The Zetaproteobacteria also have the genomic potential for acquiring nitrogen from numerous sources including ammonium, nitrate, organic compounds, and nitrogen gas. Two types of molybdopterin oxidoreductase genes were found in the SAGs, indicating that those found in the isolates, thought to be involved in iron oxidation, are not consistent among all the Zetaproteobacteria. However, a novel cluster of redox-related genes was found to be conserved in 10 SAGs as well as in the isolates warranting further investigation. These results were used to isolate a novel iron-oxidizing Zetaproteobacteria. Physiological studies and genomic analysis of this isolate were able to support many of the findings from SAG analyses demonstrating the value of these data for designing future enrichment strategies.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{M2014,

title = {Unveiling viral\textendashhost interactions within the ‘microbial dark matter’},

author = {Mart\'{i}nez-Garc\'{i}a M and Santos F and Moreno-Paz M and Parro V and Ant\'{o}n J},

url = {https://www.nature.com/articles/ncomms5542},

doi = {10.1038/ncomms5542},

year  = {2014},

date = {2014-08-14},

journal = {Nature Communications},

abstract = {Viruses control natural microbial communities. Identification of virus\textendashhost pairs relies either on their cultivation or on metagenomics and tentative assignment based on genomic signatures. Both approaches have severe drawbacks when aiming to target such pairs within the uncultured majority. Here we present an unambiguous way to assign viruses to hosts that does not rely on any previous information about either of them nor requires their cultivation. First, genomic contents of individual cells present in an environmental sample are retrieved by means of single-cell genomic technologies. Then, individual cell genomes are hybridized against a set of individual viral genomes from the same sample, previously immobilized on a microarray. Infected cells will yield positive hybridization as they carry viral genomes, which can be then sequenced and characterized. Using this method, we pinpoint viruses infecting the ubiquitous hyperhalophilic Nanohaloarchaeota, included in the so-called ‘microbial dark matter’ (the uncultured fraction of the microbial world).},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{DK2014,

title = {Comparative genomics reveals adaptations of a halotolerant thaumarchaeon in the interfaces of brine pools in the Red Sea},

author = {Ngugi DK and Blom J and Alam I and Rashid M and Ba-Alawi W and Zhang G and Hikmawan T and Guan Y and Antunes A and Siam R and El Dorry H and Bajic V and Stingl U},

url = {https://www.nature.com/articles/ismej2014137},

doi = {10.1038/ismej.2014.137},

year  = {2014},

date = {2014-08-08},

journal = {ISME},

abstract = {The bottom of the Red Sea harbors over 25 deep hypersaline anoxic basins that are geochemically distinct and characterized by vertical gradients of extreme physicochemical conditions. Because of strong changes in density, particulate and microbial debris get entrapped in the brine-seawater interface (BSI), resulting in increased dissolved organic carbon, reduced dissolved oxygen toward the brines and enhanced microbial activities in the BSI. These features coupled with the deep-sea prevalence of ammonia-oxidizing archaea (AOA) in the global ocean make the BSI a suitable environment for studying the osmotic adaptations and ecology of these important players in the marine nitrogen cycle. Using phylogenomic-based approaches, we show that the local archaeal community of five different BSI habitats (with up to 18.2% salinity) is composed mostly of a single, highly abundant Nitrosopumilus-like phylotype that is phylogenetically distinct from the bathypelagic thaumarchaea; ammonia-oxidizing bacteria were absent. The composite genome of this novel Nitrosopumilus-like subpopulation (RSA3) co-assembled from multiple single-cell amplified genomes (SAGs) from one such BSI habitat further revealed that it shares ∼54% of its predicted genomic inventory with sequenced Nitrosopumilus species. RSA3 also carries several, albeit variable gene sets that further illuminate the phylogenetic diversity and metabolic plasticity of this genus. Specifically, it encodes for a putative proline-glutamate ‘switch’ with a potential role in osmotolerance and indirect impact on carbon and energy flows. Metagenomic fragment recruitment analyses against the composite RSA3 genome, Nitrosopumilus maritimus, and SAGs of mesopelagic thaumarchaea also reiterate the divergence of the BSI genotypes from other AOA.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{NH2014,

title = {Insights into the metabolism, lifestyle and putative evolutionary history of the novel archaeal phylum ‘Diapherotrites’},

author = {Youssef NH and Rinke C and Stepanauskas R and Farag I and Woyke T and Elshahed MS},

url = {https://www.nature.com/articles/ismej2014141},

doi = {10.1038/ismej.2014.141},

year  = {2014},

date = {2014-08-01},

journal = {ISME},

abstract = {The archaeal phylum ‘Diapherotrites’ was recently proposed based on phylogenomic analysis of genomes recovered from an underground water seep in an abandoned gold mine (Homestake mine in Lead, SD, USA). Here we present a detailed analysis of the metabolic capabilities and genomic features of three single amplified genomes (SAGs) belonging to the ‘Diapherotrites’. The most complete of the SAGs, Candidatus ‘Iainarchaeum andersonii’ (Cand. IA), had a small genome (∼1.24 Mb), short average gene length (822 bp), one ribosomal RNA operon, high coding density (∼90.4%), high percentage of overlapping genes (27.6%) and low incidence of gene duplication (2.16%). Cand. IA genome possesses limited catabolic capacities that, nevertheless, could theoretically support a free-living lifestyle by channeling a narrow range of substrates such as ribose, polyhydroxybutyrate and several amino acids to acetyl-coenzyme A. On the other hand, Cand. IA possesses relatively well-developed anabolic capabilities, although it remains auxotrophic for several amino acids and cofactors. Phylogenetic analysis suggests that the majority of Cand. IA anabolic genes were acquired from bacterial donors via horizontal gene transfer. We thus propose that members of the ‘Diapherotrites’ have evolved from an obligate symbiotic ancestor by acquiring anabolic genes from bacteria that enabled independent biosynthesis of biological molecules previously acquired from symbiotic hosts. ‘Diapherotrites’ 16S rRNA genes exhibit multiple mismatches with the majority of archaeal 16S rRNA primers, a fact that could be responsible for their observed rarity in amplicon-generated data sets. The limited substrate range, complex growth requirements and slow growth rate predicted could be responsible for its refraction to isolation.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{M2014b,

title = {From community approaches to single-cell genomics: the discovery of ubiquitous hyperhalophilic Bacteroidetes generalists},

author = {Gomariz M and Martinez-Garcia M and Santos F and Rodriguez F and Capella-Gutierrez S and Gabaldon T and Rosello-Mora R},

url = {https://www.nature.com/articles/ismej201495},

doi = {10.1038/ismej.2014.95},

year  = {2014},

date = {2014-06-13},

journal = {ISME},

abstract = {The microbiota of multi-pond solar salterns around the world has been analyzed using a variety of culture-dependent and molecular techniques. However, studies addressing the dynamic nature of these systems are very scarce. Here we have characterized the temporal variation during 1 year of the microbiota of five ponds with increasing salinity (from 18% to >40%), by means of CARD-FISH and DGGE. Microbial community structure was statistically correlated with several environmental parameters, including ionic composition and meteorological factors, indicating that the microbial community was dynamic as specific phylotypes appeared only at certain times of the year. In addition to total salinity, microbial composition was strongly influenced by temperature and specific ionic composition. Remarkably, DGGE analyses unveiled the presence of most phylotypes previously detected in hypersaline systems using metagenomics and other molecular techniques, such as the very abundant Haloquadratum and Salinibacter representatives or the recently described low GC Actinobacteria and Nanohaloarchaeota. In addition, an uncultured group of Bacteroidetes was present along the whole range of salinity. Database searches indicated a previously unrecognized widespread distribution of this phylotype. Single-cell genome analysis of five members of this group suggested a set of metabolic characteristics that could provide competitive advantages in hypersaline environments, such as polymer degradation capabilities, the presence of retinal-binding light-activated proton pumps and arsenate reduction potential. In addition, the fairly high metagenomic fragment recruitment obtained for these single cells in both the intermediate and hypersaline ponds further confirm the DGGE data and point to the generalist lifestyle of this new Bacteroidetes group.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{Kashtan12014,

title = {Single-Cell Genomics Reveals Hundreds of Coexisting Subpopulations in Wild Prochlorococcus},

author = {Nadav Kashtan and Sara E Roggensack and S\'{e}bastien Rodrigue and Jessie W Thompson and Steven J Biller and Allison Coe and Huiming Ding and Pekka Marttinen and Rex R Malmstrom and Roman Stocker and Michael J Follows and Ramunas Stepanauskas and Sallie W Chisholm},

url = {http://science.sciencemag.org/content/344/6182/416},

doi = {10.1126/science.1248575},

year  = {2014},

date = {2014-04-25},

journal = {Science},

volume = {344},

number = {6182},

pages = {416-420},

abstract = {Extensive genomic diversity within coexisting members of a microbial species has been revealed through selected cultured isolates and metagenomic assemblies. Yet, the cell-by-cell genomic composition of wild uncultured populations of co-occurring cells is largely unknown. In this work, we applied large-scale single-cell genomics to study populations of the globally abundant marine cyanobacterium Prochlorococcus. We show that they are composed of hundreds of subpopulations with distinct “genomic backbones,” each backbone consisting of a different set of core gene alleles linked to a small distinctive set of flexible genes. These subpopulations are estimated to have diverged at least a few million years ago, suggesting ancient, stable niche partitioning. Such a large set of coexisting subpopulations may be a general feature of free-living bacterial species with huge populations in highly mixed habitats.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{LJ2014,

title = {Exploring bacteria\textendashdiatom associations using single-cell whole genome amplification},

author = {Baker LJ and Kemp PF},

url = {http://www.int-res.com/abstracts/ame/v72/n1/p73-88/},

doi = {10.3354/ame01686},

year  = {2014},

date = {2014-04-04},

journal = {Aquatic Microbial Ecology},

volume = {72},

pages = {73-88},

abstract = {Diatoms are responsible for a large fraction of oceanic and freshwater biomass production and are critically important for sequestration of carbon to the deep ocean. As with most surfaces present in aquatic systems, bacteria colonize the exterior of diatom cells, and they interact with the diatom and each other. The ecology of diatoms may be better explained by conceptualizing them as composite organisms consisting of the host cell and its bacterial associates. Such associations could have collective properties that are not predictable from the properties of the host cell alone. Past studies of these associations have employed culture-based, whole-population methods. In contrast, we examined the composition and variability of bacterial assemblages attached to individual diatoms. Samples were collected in an oligotrophic system (Station ALOHA, 22°45’N, 158°00’W) at the deep chlorophyll maximum. Forty eukaryotic host cells were isolated by flow cytometry followed by multiple displacement amplification, including 26 Thalassiosira spp., other diatoms, dinoflagellates, coccolithophorids, and flagellates. Bacteria were identified by amplifying, cloning, and sequencing 16S rDNA using primers that select against chloroplast 16S rDNA. Bacterial sequences were recovered from 32 of 40 host cells, and from parallel samples of the free-living and particle-associated bacteria. Bacterial assemblages varied substantially even among closely related host cells. Host cells and the free-living and particle-associated samples can be placed into distinct groups based on the phylogenetic relatedness of their associated bacteria, rather than the identity of the host cell. As yet, the functional implications of these groups are unknown.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{K2013,

title = {Single cell genomics reveal low recombination frequencies in freshwater bacteria of the SAR11 clade},

author = {Zaremba-Niedzwiedzka K and Viklund J and Zhao W and Ast J and Sczyrba A and Woyke T and McMahon KD and Bertilsson S and Stepanauskas R and Andersson SGE},

issn = {1474-760X},

year  = {2013},

date = {2013-11-28},

journal = {Genome Biology},

volume = {14},

number = {R130},

abstract = {Background

The SAR11 group of Alphaproteobacteria is highly abundant in the oceans. It contains a recently diverged freshwater clade, which offers the opportunity to compare adaptations to salt- and freshwaters in a monophyletic bacterial group. However, there are no cultivated members of the freshwater SAR11 group and no genomes have been sequenced yet.



Results

We isolated ten single SAR11 cells from three freshwater lakes and sequenced and assembled their genomes. A phylogeny based on 57 proteins indicates that the cells are organized into distinct microclusters. We show that the freshwater genomes have evolved primarily by the accumulation of nucleotide substitutions and that they have among the lowest ratio of recombination to mutation estimated for bacteria. In contrast, members of the marine SAR11 clade have one of the highest ratios. Additional metagenome reads from six lakes confirm low recombination frequencies for the genome overall and reveal lake-specific variations in microcluster abundances. We identify hypervariable regions with gene contents broadly similar to those in the hypervariable regions of the marine isolates, containing genes putatively coding for cell surface molecules.



Conclusions

We conclude that recombination rates differ dramatically in phylogenetic sister groups of the SAR11 clade adapted to freshwater and marine ecosystems. The results suggest that the transition from marine to freshwater systems has purged diversity and resulted in reduced opportunities for recombination with divergent members of the clade. The low recombination frequencies of the LD12 clade resemble the low genetic divergence of host-restricted pathogens that have recently shifted to a new host.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{S2013,

title = {Assembling single-cell genomes and mini-metagenomes from chimeric MDA products},

author = {Nurk S and Bankevich A and Antipov D and Gurevich AA and Korobeynikov A and Lapidus A and Prjibelski AD and Pyshkin A and Sirotkin A and Sirotkin Y and Stepanauskas R and Clingenpeel SR and Woyke T and McLean JS and Lasken R and Tesler G and Alekseyev MA and Pevzner PA},

url = {https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3791033/},

doi = {10.1089/cmb.2013.0084},

year  = {2013},

date = {2013-10-20},

journal = {Journal of Computational Biology},

volume = {20},

pages = {714-737},

abstract = {Recent advances in single-cell genomics provide an alternative to largely gene-centric metagenomics studies, enabling whole-genome sequencing of uncultivated bacteria. However, single-cell assembly projects are challenging due to (i) the highly nonuniform read coverage and (ii) a greatly elevated number of chimeric reads and read pairs. While recently developed single-cell assemblers have addressed the former challenge, methods for assembling highly chimeric reads remain poorly explored. We present algorithms for identifying chimeric edges and resolving complex bulges in de Bruijn graphs, which significantly improve single-cell assemblies. We further describe applications of the single-cell assembler SPAdes to a new approach for capturing and sequencing “microbial dark matter” that forms small pools of randomly selected single cells (called a mini-metagenome) and further sequences all genomes from the mini-metagenome at once. On single-cell bacterial datasets, SPAdes improves on the recently developed E+V-SC and IDBA-UD assemblers specifically designed for single-cell sequencing. For standard (cultivated monostrain) datasets, SPAdes also improves on A5, ABySS, CLC, EULER-SR, Ray, SOAPdenovo, and Velvet. Thus, recently developed single-cell assemblers not only enable single-cell sequencing, but also improve on conventional assemblers on their own turf. SPAdes is available for free online download under a GPLv2 license.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{GM2013,

title = {Prokaryotic super program advisory committee DOE joint genome institute, Walnut Creek, CA, March 27, 2012},

author = {Garrity GM and Banfield J and Eisen J van der Lelie N and McMahon T and Rusch D and Delong E and Moran MA and Currie C and Furhman J and Hallam S and Hugenholtz P and Moran N and Nelson K and Roberts R and Stepanauskas R},

url = {https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3910701/},

doi = {10.4056/sigs.4638348},

year  = {2013},

date = {2013-07-30},

journal = {Standards in Genomic Sciences},

volume = {8},

number = {3},

pages = {561-570},

abstract = {The Prokaryotic Super Program Advisory Committee met on March 27, 2013 for their annual review the Prokaryotic Super Program at the DOE Joint Genome Institute. As is the case with any site visit or program review, the objective is to evaluate progress in meeting organizational objectives, provide feedback to from the user-community and to assist the JGI in formulating plans for the coming year. The advisors want to commend the JGI for its central role in developing new technologies and capabilities, and for catalyzing the formation of new collaborative user communities. Highlights of the post-meeting exchanges among the advisors focused on the importance of programmatic initiatives including:



• GEBA, which serves as a phylogenetic “base-map” on which our knowledge of functional diversity can be layered.



• FEBA, which promises to provide new insights into the physiological capabilities of prokaryotes under highly standardized conditions.



• Single-cell genomics technology, which is seen to significantly enhance our ability to interpret genomic and metagenomic data and broaden the scope of the GEBA program to encompass at least a part of the microbial “dark-matter”.



• IMG, which is seen to play a central role in JGI programs and is viewed as a strategically important asset in the JGI portfolio.



On this latter point, the committee encourages the formation of a strategic relationship between IMG and the Kbase to ensure that the intelligence, deep knowledge and experience captured in the former is not lost. The committee strongly urges the DOE to continue its support for maintaining this critical resource.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}