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Sinking particles and particle-associated microbes influence global biogeochemistry through particulate matter export from the surface to the deep ocean. Despite ongoing studies of particle-associated microbes, viruses in these habitats remain largely unexplored. Whether, where, and which viruses might contribute to particle production and export remain open to investigation. In this study, we analyzed 857 virus population genomes associated with sinking particles collected over three years in sediment traps moored at 4000 m in the North Pacific Subtropical Gyre. Particle-associated viruses here were linked to cellular hosts through matches to bacterial and archaeal metagenome-assembled genome (MAG)-encoded prophages or CRISPR spacers, identifying novel viruses infecting presumptive deep-sea bacteria such as Colwellia, Moritella, and Shewanella. We also identified lytic viruses whose abundances correlated with particulate carbon flux and/or were exported from the photic to abyssal ocean, including cyanophages. Our data are consistent with some of the predicted outcomes of the viral shuttle hypothesis, and further suggest that viral lysis of both autotrophic and heterotrophic prokaryotes may play a role in carbon export. Our analyses revealed the diversity and origins of prevalent viruses found on deep-sea sinking particles and identified prospective viral groups for future investigation into processes that govern particle export in the open ocean.

Complex assemblages of microbes in the surface ocean are responsible for approximately half of global carbon fixation. The persistence of high taxonomic diversity despite competition for a small suite of relatively homogeneously distributed nutrients, that is, 'the paradox of the plankton', represents a long-standing challenge for ecological theory. Here we find evidence consistent with temporal niche partitioning of nitrogen assimilation processes over a diel cycle in the North Pacific Subtropical Gyre. We jointly analysed transcript abundances, lipids and metabolites and discovered that a small number of diel archetypes can explain pervasive periodic dynamics. Metabolic pathway analysis of identified diel signals revealed asynchronous timing in the transcription of nitrogen uptake and assimilation genes among different microbial groups-cyanobacteria, heterotrophic bacteria and eukaryotes. This temporal niche partitioning of nitrogen uptake emerged despite synchronous transcription of photosynthesis and central carbon metabolism genes and associated macromolecular abundances. Temporal niche partitioning may be a mechanism by which microorganisms in the open ocean mitigate competition for scarce resources, supporting community coexistence.

Horizontal gene transfer accelerates microbial evolution, promoting diversification and adaptation. The globally abundant marine cyanobacterium Prochlorococcus has a highly streamlined genome with frequent gene exchange reflected in its extensive pangenome. The source of its genomic variability, however, remains elusive since most cells lack the common mechanisms that enable horizontal gene transfer, including conjugation, transformation, plasmids and prophages. Examining 623 genomes, we reveal a diverse system of mobile genetic elements – cargo-carrying transposons we named tycheposons – that shape Prochlorococcus’ genomic plasticity. The excision and integration of tycheposons at seven tRNA genes drive the remodeling of larger genomic islands containing most of Prochlorococcus’ flexible genes. Most tycheposons carry genes important for niche differentiation through nutrient acquisition; others appear similar to phage parasites. Tycheposons are highly enriched in extracellular vesicles and phage particles in ocean samples, suggesting efficient routes for their dispersal, transmission and propagation. Supported by evidence for similar elements in other marine microbes, our work underpins the role of vesicle- and virus-mediated transfer of mobile genetic elements in the diversification and adaptation of microbes in dilute aquatic environments – adding a significant piece to the puzzle of what governs microbial evolution in the planet’s largest habitat.

Phage satellites are mobile genetic elements that propagate by parasitizing bacteriophage replication. We report here the discovery of abundant and diverse phage satellites that were packaged as concatemeric repeats within naturally occurring bacteriophage particles in seawater. These same phage-parasitizing mobile elements were found integrated in the genomes of dominant co-occurring bacterioplankton species. Like known phage satellites, many of the marine phage satellites encoded genes for integration, DNA replication, phage interference, and capsid assembly. Many also contained distinctive gene suites indicative of unique virus hijacking, phage interference and mobilization mechanisms. Marine phage satellite sequences were widespread in local and global oceanic virioplankton populations, reflecting their ubiquity, abundance, and temporal persistence in marine planktonic communities worldwide. Their gene content and putative life cycles suggest they may impact host-cell phage immunity and defense, lateral gene transfer, and bacteriophage-induced cell mortality and host and virus productivity. These previously unrecognized marine phage satellites therefore have potential to impact the ecology and evolution of bacteria and their bacteriophages in the ocean, and similar phage parasites likely thrive in many other microbial habitats as well.Significance statementPhage satellites are mobile genetic elements that parasitize bacteriophage, thereby exerting profound biological and ecological impacts. To date however, phage satellites have been found primarily in Gram-positive cocci and a few Gram-negative bacteria, many of which are human pathogens. Direct inspection of “wild” marine virus particles however, revealed that phage satellites are widely distributed in the sea, and that their genetic diversity, gene repertoires, and host ranges are much greater than previously supposed. Our analyses provide insight into their parasitic life cycles, potential satellite-helper-phage interactions, and reproductive strategies of these newly recognized phage-parasitizing mobile elements. Their properties, diversity and environmental distributions suggest they exert pervasive influence on marine plankton ecology and bacterial and virus evolution in the sea.

Viruses are the most abundant biological entities on Earth and play key roles in host ecology, evolution, and horizontal gene transfer. Despite recent progress in viral metagenomics, the inherent genetic complexity of virus populations still poses technical difficulties for recovering complete virus genomes from natural assemblages. To address these challenges, we developed an assembly-free, single-molecule nanopore sequencing approach, enabling direct recovery of complete virus genome sequences from environmental samples. Our method yielded thousands of full-length, high-quality draft virus genome sequences that were not recovered using standard short-read assembly approaches. Additionally, our analyses discriminated between populations whose genomes had identical direct terminal repeats versus those with circularly permuted repeats at their termini, thus providing new insight into native virus reproduction and genome packaging. Novel DNA sequences were discovered, whose repeat structures, gene contents, and concatemer lengths suggest they are phage-inducible chromosomal islands, which are packaged as concatemers in phage particles, with lengths that match the size ranges of co-occurring phage genomes. Our new virus sequencing strategy can provide previously unavailable information about the genome structures, population biology, and ecology of naturally occurring viruses and viral parasites.

Microbial communities are critical to ecosystem dynamics and biogeochemical cycling in the open oceans. Viruses are essential elements of these communities, influencing the productivity, diversity, and evolution of cellular hosts. To further explore the natural history and ecology of open-ocean viruses, we surveyed the spatiotemporal dynamics of double-stranded DNA (dsDNA) viruses in both virioplankton and bacterioplankton size fractions in the North Pacific Subtropical Gyre, one of the largest biomes on the planet. Assembly and clustering of viral genomes revealed a peak in virioplankton diversity at the base of the euphotic zone, where virus populations and host species richness both reached their maxima. Simultaneous characterization of both extracellular and intracellular viruses suggested depth-specific reproductive strategies. In particular, analyses indicated elevated lytic interactions in the mixed layer, more temporally variable temperate phage interactions at the base of the euphotic zone, and increased lysogeny in the mesopelagic ocean. Furthermore, the depth variability of auxiliary metabolic genes suggested habitat-specific strategies for viral influence on light-energy, nitrogen, and phosphorus acquisition during host infection. Most virus populations were temporally persistent over several years in this environment at the 95% nucleic acid identity level. In total, our analyses revealed variable distributional patterns and diverse reproductive and metabolic strategies of virus populations in the open-ocean water column.

With advances in metagenomic sequencing, the microbial composition of diverse environmental systems has been investigated, providing new perspectives on potential ecological dynamics and dimensions for experimental investigations. Here, we characterized exocellular free DNA via metagenomics, using a newly developed method that separates free DNA from cells, viruses, and vesicles, and facilitated the independent characterization of each fraction.

Sinking particles and particle-associated microbes influence global biogeochemistry through particulate matter export from the surface to the deep ocean. Despite ongoing studies of particle-associated microbes, viruses in these habitats remain largely unexplored. Whether, where, and which viruses might contribute to particle production and export remain open to investigation. In this study, we analyzed 857 virus population genomes associated with sinking particles collected over three years in sediment traps moored at 4000 m in the North Pacific Subtropical Gyre. Particle-associated viruses here were linked to cellular hosts through matches to bacterial and archaeal metagenome-assembled genome (MAG)-encoded prophages or CRISPR spacers, identifying novel viruses infecting presumptive deep-sea bacteria such as Colwellia, Moritella, and Shewanella. We also identified lytic viruses whose abundances correlated with particulate carbon flux and/or were exported from the photic to abyssal ocean, including cyanophages. Our data are consistent with some of the predicted outcomes of the viral shuttle hypothesis, and further suggest that viral lysis of both autotrophic and heterotrophic prokaryotes may play a role in carbon export. Our analyses revealed the diversity and origins of prevalent viruses found on deep-sea sinking particles and identified prospective viral groups for future investigation into processes that govern particle export in the open ocean.

Long-read sequencing technologies hold big promises for the genomic analysis of complex samples such as microbial communities. Yet, despite improving accuracy, basic gene prediction on long-read data is still often impaired by frameshifts resulting from small indels. Consensus polishing using either complementary short reads or to a lesser extent the long reads themselves can mitigate this effect but requires universally high sequencing depth, which is difficult to achieve in complex samples where the majority of community members are rare. Here we present proovframe, a software implementing an alternative approach to overcome frameshift errors in long-read assemblies and raw long reads. We utilize protein-to-nucleotide alignments against reference databases to pinpoint indels in contigs or reads and correct them by deleting or inserting 1-2 bases, thereby conservatively restoring reading-frame fidelity in aligned regions. Using simulated and real-world benchmark data we show that proovframe performs comparably to short-read-based polishing on assembled data, works well with remote protein homologs, and can even be applied to raw reads directly. Together, our results demonstrate that protein-guided frameshift correction significantly improves the analyzability of long-read data both in combination with and as an alternative to common polishing strategies. Proovframe is available from https://github.com/thackl/proovframe.

Significance Sinking particles composed of both organic and inorganic material feed the deep-sea ecosystem and contribute centrally to ocean carbon sequestration. Despite their importance, little is known about the biological composition of sinking particles reaching the deep sea. Time-series analyses of sinking particles unexpectedly revealed bacterial assemblages that were simple and homogeneous over time. Particle-associated eukaryote assemblages, however, were more variable and complex. Several modes of export were observed, including summer inputs from the surface, more stochastic export of surface-derived protists and animals, and contributions from midwater animals and deep-sea bacteria. In summary, sinking particles exhibited temporally variable, heterogeneous biological sources and activities that reflected their important roles in the downward transport and transformation of organic matter in the deep sea., Sinking particles are a critical conduit for the export of organic material from surface waters to the deep ocean. Despite their importance in oceanic carbon cycling and export, little is known about the biotic composition, origins, and variability of sinking particles reaching abyssal depths. Here, we analyzed particle-associated nucleic acids captured and preserved in sediment traps at 4,000-m depth in the North Pacific Subtropical Gyre. Over the 9-month time-series, Bacteria dominated both the rRNA-gene and rRNA pools, followed by eukaryotes (protists and animals) and trace amounts of Archaea. Deep-sea piezophile-like Gammaproteobacteria, along with Epsilonproteobacteria, comprised >80% of the bacterial inventory. Protists (mostly Rhizaria, Syndinales, and ciliates) and metazoa (predominantly pelagic mollusks and cnidarians) were the most common sinking particle-associated eukaryotes. Some near-surface water-derived eukaryotes, especially Foraminifera, Radiolaria, and pteropods, varied greatly in their abundance patterns, presumably due to sporadic export events. The dominance of piezophile-like Gammaproteobacteria and Epsilonproteobacteria, along with the prevalence of their nitrogen cycling-associated gene transcripts, suggested a central role for these bacteria in the mineralization and biogeochemical transformation of sinking particulate organic matter in the deep ocean. Our data also reflected several different modes of particle export dynamics, including summer export, more stochastic inputs from the upper water column by protists and pteropods, and contributions from sinking mid- and deep-water organisms. In total, our observations revealed the variable and heterogeneous biological origins and microbial activities of sinking particles that connect their downward transport, transformation, and degradation to deep-sea biogeochemical processes.

1.AbstractMesoscale eddies have been shown to support elevated dinitrogen (N2) fixation rates (NFRs) and abundances of N2-fixing microorganisms (diazotrophs), but the mechanisms underlying these observations are not well understood. We explored relationships among NFRs and cyanobacterial diazotroph abundances in eddy pairs of opposite polarity sampled in the North Pacific Subtropical Gyre and compared our observations to seasonal trends from the Hawaii Ocean Time-series (HOT) program. Consistent with previous reports, we found that NFRs were anomalously high for this region (up to 3.7-fold above previous monthly HOT observations) in the centers of the sampled anticyclones, coinciding with elevated abundances of Crocosphaera in the summertime. We then coupled our field-based observations, together with transcriptomic analyses of nutrient stress marker genes and ecological models, to evaluate potential mechanisms controlling diazotroph abundance and activity at the mesoscale. Specifically, we evaluated the role of biological (via estimates of growth and grazing rates) and physical controls on populations of Crocosphaera, Trichodesmium, and diatom symbionts. Our results suggest that increased Crocosphaera abundances associated with summertime anticyclones resulted from the alleviation of phosphate limitation, allowing cells to grow at rates exceeding grazing losses. In contrast, distributions of larger, buoyant taxa (Trichodesmium and diatom symbionts) appeared less affected by eddy-driven biological controls. Instead, they appeared driven by physical dynamics along frontal boundaries that separate cyclonic and anticyclonic eddies. Together, the interplay of eddy-specific changes in bottom-up control, top-down control, and the physical accumulation of cells likely explains the elevated diazotroph abundances and NFRs associated with anticyclones and eddy fronts.

Significance The ocean’s “biological pump” exports sinking particles containing carbon, nutrients, and energy to the deep sea, contributing centrally to the global carbon cycle. Here, we identify key organisms and biological processes associated with elevated carbon flux to the abyss. Our analyses reveal that, during summer export, specific populations of photosynthetic algae, heterotrophic protists, and bacteria reach the abyss on sinking particles. Deep-sea bacteria respond rapidly to this elevated nutrient delivery to the abyss in summer. During other seasons, different organisms and processes appear responsible for particle export to the deep sea. Our analyses reveal key biota and biological processes that interconnect surface productivity, particle export, and the deep-sea ecosystem, thereby influencing the function and efficiency of the ocean’s biological pump., In the open ocean, elevated carbon flux (ECF) events increase the delivery of particulate carbon from surface waters to the seafloor by severalfold compared to other times of year. Since microbes play central roles in primary production and sinking particle formation, they contribute greatly to carbon export to the deep sea. Few studies, however, have quantitatively linked ECF events with the specific microbial assemblages that drive them. Here, we identify key microbial taxa and functional traits on deep-sea sinking particles that correlate positively with ECF events. Microbes enriched on sinking particles in summer ECF events included symbiotic and free-living diazotrophic cyanobacteria, rhizosolenid diatoms, phototrophic and heterotrophic protists, and photoheterotrophic and copiotrophic bacteria. Particle-attached bacteria reaching the abyss during summer ECF events encoded metabolic pathways reflecting their surface water origins, including oxygenic and aerobic anoxygenic photosynthesis, nitrogen fixation, and proteorhodopsin-based photoheterotrophy. The abundances of some deep-sea bacteria also correlated positively with summer ECF events, suggesting rapid bathypelagic responses to elevated organic matter inputs. Biota enriched on sinking particles during a spring ECF event were distinct from those found in summer, and included rhizaria, copepods, fungi, and different bacterial taxa. At other times over our 3-y study, mid- and deep-water particle colonization, predation, degradation, and repackaging (by deep-sea bacteria, protists, and animals) appeared to shape the biotic composition of particles reaching the abyss. Our analyses reveal key microbial players and biological processes involved in particle formation, rapid export, and consumption, that may influence the ocean’s biological pump and help sustain deep-sea ecosystems.

Big data abound in microbiology, but the workflows designed to enable researchers to interpret data can constrain the biological questions that can be asked. Five years after anvi'o was first published, this community-led multi-omics platform is maturing into an open software ecosystem that reduces constraints in 'omics data analyses. Non

Horizontal gene transfer accelerates microbial evolution, promoting diversification and adaptation. The globally abundant marine cyanobacteriumProchlorococcushas a highly streamlined genome with frequent gene exchange reflected in its extensive pangenome. The source of its genomic variability, however, remains elusive since most cells lack the common mechanisms that enable horizontal gene transfer, including conjugation, transformation, plasmids and prophages. Examining 623 genomes, we reveal a diverse system of mobile genetic elements – cargo-carrying transposons we named tycheposons – that shapeProchlorococcus’ genomic plasticity. The excision and integration of tycheposons at seven tRNA genes drive the remodeling of larger genomic islands containing most ofProchlorococcus’ flexible genes. Most tycheposons carry genes important for niche differentiation through nutrient acquisition; others appear similar to phage parasites. Tycheposons are highly enriched in extracellular vesicles and phage particles in ocean samples, suggesting efficient routes for their dispersal, transmission and propagation. Supported by evidence for similar elements in other marine microbes, our work underpins the role of vesicle- and virus-mediated transfer of mobile genetic elements in the diversification and adaptation of microbes in dilute aquatic environments – adding a significant piece to the puzzle of what governs microbial evolution in the planet’s largest habitat.

Sunlight drives daily rhythms of photosynthesis, growth, and division of photoautotrophs throughout the surface oceans. However, the cascading impacts of oscillatory light input on diverse microbial communities and community-scale metabolism remains unclear. Here we use an unsupervised machine learning approach to show that a small number of diel archetypes can explain pervasive periodic dynamics amongst more than 65,000 distinct time series, including transcriptional activity, macromolecules, lipids, and metabolites from the North Pacific Subtropical Gyre. Overall, we find evidence for synchronous timing of carbon-cycle gene expression that underlie daily oscillations in the concentrations of particulate organic carbon. In contrast, we find evidence of asynchronous timing in gene transcription related to nitrogen metabolism and related metabolic processes consistent with temporal niche partitioning amongst microorganisms in the bacterial and eukaryotic domains.

Viruses are the most abundant biological entities on Earth, and play key roles in host ecology, evolution, and horizontal gene transfer. Despite recent progress in viral metagenomics, the inherent genetic complexity of virus populations still poses technical difficulties for recovering complete virus genomes from natural assemblages. To address these challenges, we developed an assembly-free, single-molecule nanopore sequencing approach enabling direct recovery of high-quality viral genome sequences from environmental samples. Our method yielded over a thousand high quality, full-length draft virus genome sequences that could not be fully recovered using short read assembly approaches applied to the same samples. Additionally, novel DNA sequences were discovered whose repeat structures, gene contents and concatemer lengths suggested that they represent phage-inducible chromosomal islands that were packaged as concatemers within phage particles. Our new approach provided novel insight into genome structures, population biology, and ecology of naturally occurring viruses and viral parasites.

Few microbial time-series studies have been conducted in open ocean habitats having low seasonal variability such as the North Pacific Subtropical Gyre (NPSG), where surface waters experience comparatively mild seasonal variation. To better describe microbial seasonal variability in this habitat, we analyzed rRNA amplicon and shotgun metagenomic data over two years at the Hawaii Ocean Time-series Station ALOHA. We postulated that this relatively stable habitat might reveal different environmental factors that influence planktonic microbial community diversity than those previously observed in more seasonally dynamic habitats. Unexpectedly, the data showed that microbial diversity at 25 m was positively correlated with average wind speed 3 to 10 days prior to sampling. In addition, microbial community composition at 25 m exhibited significant correlations with solar irradiance. Many bacterial groups whose relative abundances varied with solar radiation corresponded to taxa known to exhibit strong seasonality in other oceanic regions. Network co-correlation analysis of 25 m communities showed seasonal transitions in composition, and distinct successional cohorts of co-occurring phylogenetic groups. Similar network analyses of metagenomic data also indicated distinct seasonality in genes originating from cyanophage, and several bacterial clades including SAR116 and SAR324. At 500 m, microbial community diversity and composition did not vary significantly with any measured environmental parameters. The minimal seasonal variability in the NPSG facilitated detection of more subtle environmental influences, such as episodic wind variation, on surface water microbial diversity. Community composition in NPSG surface waters varied in response to solar irradiance, but less dramatically than reported in other ocean provinces., Gordon and Betty Moore Foundation (Grant 492.01), Gordon and Betty Moore Foundation (Grant 3777), United States. Environmental Protection Agency (STAR Fellowship)

We present Bioconda (https://bioconda.github.io), a distribution of bioinformatics software for the lightweight, multiplatform and language-agnostic package manager Conda. Currently, Bioconda offers a collection of over 3000 software packages, which is continuously maintained, updated, and extended by a growing global community of more than 200 contributors. Bioconda improves analysis reproducibility by allowing users to define isolated environments with defined software versions, all of which are easily installed and managed without administrative privileges.

The temporal dynamics of phytoplankton growth and activity have large impacts on fluxes of matter and energy, yet obtaining in situ metabolic measurements of sufficient resolution for even dominant microorganisms remains a considerable challenge. We performed Lagrangian diel sampling with synoptic measurements of population abundances, dinitrogen (N2) fixation, mortality, productivity, export and transcription in a bloom of Crocosphaera over eight days in the North Pacific Subtropical Gyre (NPSG). Quantitative transcriptomic analyses revealed clear diel oscillations in transcript abundances for 34% of Crocosphaera genes identified, reflecting a systematic progression of gene expression in diverse metabolic pathways. Significant time-lagged correspondence was evident between nifH transcript abundance and maximal N2 fixation, as well as sepF transcript abundance and cell division, demonstrating the utility of transcriptomics to predict the occurrence and timing of physiological and biogeochemical processes in natural populations. Indirect estimates of carbon fixation by Crocosphaera were equivalent to 11% of net community production, suggesting that under bloom conditions this diazotroph has a considerable impact on the wider carbon cycle. Our cross-scale synthesis of molecular, population and community-wide data underscores the tightly coordinated in situ metabolism of the keystone N2-fixing cyanobacterium Crocosphaera, as well as the broader ecosystem-wide implications of its activities.

The core properties of microbial genomes, including GC content and genome size, are known to vary widely among different bacteria and archaea 1,2 . Several hypotheses have been proposed to explain this genomic variability, but the fundamental drivers that shape bacterial and archaeal genomic properties remain uncertain 3–7 . Here, we report the existence of a sharp genomic transition zone below the photic zone, where bacterial and archaeal genomes and proteomes undergo a community-wide punctuated shift. Across a narrow range of increasing depth of just tens of metres, diverse microbial clades trend towards larger genome size, higher genomic GC content, and proteins with higher nitrogen but lower carbon content. These community-wide changes in genome features appear to be driven by gradients in the surrounding environmental energy and nutrient fields. Collectively, our data support hypotheses invoking nutrient limitation as a central driver in the evolution of core bacterial and archaeal genomic and proteomic properties. Metagenomic analyses reveal that microbial genomes undergo a community-wide transition in size and GC content across a narrow depth range, indicating that nutrient limitation is a major driver in marine microbial genomic and proteomic evolution.

Strain HIMB100 is a planktonic marine bacterium in the class Alphaproteobacteria. This strain is of interest because it is one of the first known isolates from a globally ubiquitous clade of marine bacteria known as SAR116 within the family Rhodospirillaceae. Here we describe preliminary features of the organism, together with the draft genome sequence and annotation. This is the second genome sequence of a member of the SAR116 clade. The 2,458,945 bp genome contains 2,334 protein-coding and 42 RNA genes., Gordon and Betty Moore Foundation, National Science Foundation (U.S.) (Science and Technology Center (award No. EF0424599)), Center for Microbial Oceanography: Research and Education, University of Hawaii at Manoa. School of Ocean and Earth Science and Technology

To investigate the temporal, spatial and phylogenetic resolution of marine microbial community structure and variability, we designed and expanded a genome proxy array (an oligonucleotide microarray targeting marine microbial genome fragments and genomes), evaluated it against metagenomic sequencing, and applied it to time-series samples from the Monterey Bay. The expanded array targeted 268 microbial genotypes across much of the known diversity of cultured and uncultured marine microbes. The target abundances measured by the array were highly correlated to pyrosequence-based abundances (linear regression R(2) = 0.85-0.91, P < 0.0001). Fifty-seven samples from ∼4 years in Monterey Bay were examined with the array, spanning the photic zone (0 m), the base of the surface mixed layer (30 m) and the subphotic zone (200 m). A significant portion of the expanded genome proxy array's targets showed signal (95 out of 268 targets present in ≥ 1 sample). The multi-year community survey showed the consistent presence of a core group of common and abundant targeted taxa at each depth in Monterey Bay, higher variability among shallow than deep samples, and episodic occurrences of more transient marine genotypes. The abundance of the most dominant genotypes peaked after strong episodic upwelling events. The genome-proxy array's ability to track populations of closely related genotypes indicated population shifts within several abundant target taxa, with specific populations in some cases clustering by depth or oceanographic season. Although 51 cultivated organisms were targeted (representing 19% of the array) the majority of targets detected and of total target signal (85% and ∼92% respectively) were from uncultivated genotypes, often those derived from Monterey Bay. The array provided a relatively cost-effective approach (∼$15 per array) for surveying the natural history of uncultivated lineages.

Speciation as the result of barriers to genetic exchange is the foundation for the general biological species concept. However, the relevance of genetic exchange for defining microbial species is uncertain. In fact, the extent to which microbial populations comprise discrete clusters of evolutionarily related organisms is generally unclear. Metagenomic data from an acidophilic microbial community enabled a genomewide, comprehensive investigation of variation in individuals from two coexisting natural archaeal populations. Individuals are clustered into species-like groups in which cohesion appears to be maintained by homologous recombination. We quantified the dependence of recombination frequency on sequence similarity genomewide and found a decline in recombination with increasing evolutionary distance. Both inter- and intralineage recombination frequencies have a log-linear dependence on sequence divergence. In the declining phase of interspecies genetic exchange, recombination events cluster near the origin of replication and are localized by tRNAs and short regions of unusually high sequence similarity. The breakdown of genetic exchange with increasing sequence divergence could contribute to, or explain, the establishment and preservation of the observed population clusters in a manner consistent with the biological species concept.

Planktonic microbial communities in the ocean are typically dominated by several cosmopolitan clades of Bacteria, Archaea, and Eukarya characterized by their ribosomal RNA gene phylogenies and genomic features. Although the environments these communities inhabit range from coastal to open ocean waters, how the biological dynamics vary between such disparate habitats is not well known. To gain insight into the differential activities of microbial populations inhabiting different oceanic provinces we compared the daily metatranscriptome profiles of related microbial populations inhabiting surface waters of both a coastal California upwelling region (CC) as well as the oligotrophic North Pacific Subtropical Gyre (NPSG). Transcriptional networks revealed that the dominant photoautotrophic microbes in each environment (Ostreococcus in CC, Prochlorococcus in NPSG) were central determinants of overall community transcriptome dynamics. Furthermore, heterotrophic bacterial clades common to both ecosystems (SAR11, SAR116, SAR86, SAR406, and Roseobacter) displayed conserved, genome-wide inter- and intrataxon transcriptional patterns and diel cycles. Populations of SAR11 and SAR86 clades in particular exhibited tightly coordinated transcriptional patterns in both coastal and pelagic ecosystems, suggesting that specific biological interactions between these groups are widespread in nature. Our results identify common diurnally oscillating behaviors among diverse planktonic microbial species regardless of habitat, suggesting that highly conserved temporally phased biotic interactions are ubiquitous among planktonic microbial communities worldwide.

Schouten et al. (1), referencing Lincoln et al. (2), question whether Marine Group II Euryarchaeota (MG-II) contribute significantly to glycerol dibiphytanyl glycerol tetraether (GDGT) lipids in the ocean. We respond below to what we believe are some mistaken claims and interpretations.

Oscillating diurnal rhythms of gene transcription, metabolic activity, and behavior are found in all three domains of life. However, diel cycles in naturally occurring heterotrophic bacteria and archaea have rarely been observed. Here, we report time-resolved whole-genome transcriptome profiles of multiple, naturally occurring oceanic bacterial populations sampled in situ over 3 days. As anticipated, the cyanobacterial transcriptome exhibited pronounced diel periodicity. Unexpectedly, several different heterotrophic bacterioplankton groups also displayed diel cycling in many of their gene transcripts. Furthermore, diel oscillations in different heterotrophic bacterial groups suggested population-specific timing of peak transcript expression in a variety of metabolic gene suites. These staggered multispecies waves of diel gene transcription may influence both the tempo and the mode of matter and energy transformation in the sea.

Archaea are ubiquitous in marine plankton, and fossil forms of archaeal tetraether membrane lipids in sedimentary rocks document their participation in marine biogeochemical cycles for >100 million years. Ribosomal RNA surveys have identified four major clades of planktonic archaea but, to date, tetraether lipids have been characterized in only one, the Marine Group I Thaumarchaeota. The membrane lipid composition of the other planktonic archaeal groups—all uncultured Euryarchaeota—is currently unknown. Using integrated nucleic acid and lipid analyses, we found that Marine Group II Euryarchaeota (MG-II) contributed significantly to the tetraether lipid pool in the North Pacific Subtropical Gyre at shallow to intermediate depths. Our data strongly suggested that MG-II also synthesize crenarchaeol, a tetraether lipid previously considered to be a unique biomarker for Thaumarchaeota. Metagenomic datasets spanning 5 y indicated that depth stratification of planktonic archaeal groups was a stable feature in the North Pacific Subtropical Gyre. The consistent prevalence of MG-II at depths where the bulk of exported organic matter originates, together with their ubiquitous distribution over diverse oceanic provinces, suggests that this clade is a significant source of tetraether lipids to marine sediments. Our results are relevant to archaeal lipid biomarker applications in the modern oceans and the interpretation of these compounds in the geologic record.

Strain HIMB11 is a planktonic marine bacterium isolated from coastal seawater in Kaneohe Bay, Oahu, Hawaii belonging to the ubiquitous and versatile Roseobacter clade of the alphaproteobacterial family Rhodobacteraceae. Here we describe the preliminary characteristics of strain HIMB11, including annotation of the draft genome sequence and comparative genomic analysis with other members of the Roseobacter lineage. The 3,098,747 bp draft genome is arranged in 34 contigs and contains 3,183 protein-coding genes and 54 RNA genes. Phylogenomic and 16S rRNA gene analyses indicate that HIMB11 represents a unique sublineage within the Roseobacter clade. Comparison with other publicly available genome sequences from members of the Roseobacter lineage reveals that strain HIMB11 has the genomic potential to utilize a wide variety of energy sources (e.g. organic matter, reduced inorganic sulfur, light, carbon monoxide), while possessing a reduced number of substrate transporters., Gordon and Betty Moore Foundation, Agouron Institute, University of Hawaii and Manoa School of Ocean and Earth Science and Technology, National Science Foundation (U.S.). Science and Technology Center (Award EF0424599)

Oscillating diurnal rhythms of gene transcription, metabolic activity, and behavior are found in all three domains of life. However, diel cycles in naturally occurring heterotrophic bacteria and archaea have rarely been observed. Here, we report time-resolved whole-genome transcriptome profiles of multiple, naturally occurring oceanic bacterial populations sampled in situ over 3 days. As anticipated, the cyanobacterial transcriptome exhibited pronounced diel periodicity. Unexpectedly, several different heterotrophic bacterioplankton groups also displayed diel cycling in many of their gene transcripts. Furthermore, diel oscillations in different heterotrophic bacterial groups suggested population-specific timing of peak transcript expression in a variety of metabolic gene suites. These staggered multispecies waves of diel gene transcription may influence both the tempo and the mode of matter and energy transformation in the sea., Gordon and Betty Moore Foundation (GBMF 492.01), Gordon and Betty Moore Foundation (GBMF 3777), National Science Foundation (U.S.) (Grant EF0424599), David & Lucile Packard Foundation

Planktonic marine microbes live in dynamic habitats that demand rapid sensing and response to periodic as well as stochastic environmental change. The kinetics, regularity, and specificity of microbial responses in situ, however, are not well-described. We report here simultaneous multitaxon genome-wide transcriptome profiling in a naturally occurring picoplankton community. An in situ robotic sampler using a Lagrangian sampling strategy enabled continuous tracking and repeated sampling of coherent microbial populations over 2 d. Subsequent RNA sequencing analyses yielded genome-wide transcriptome profiles of eukaryotic (Ostreococcus) and bacterial (Synechococcus) photosynthetic picoplankton as well as proteorhodopsin-containing heterotrophs, including Pelagibacter, SAR86-cluster Gammaproteobacteria, and marine Euryarchaea. The photosynthetic picoplankton exhibited strong diel rhythms over thousands of gene transcripts that were remarkably consistent with diel cycling observed in laboratory pure cultures. In contrast, the heterotrophs did not cycle diurnally. Instead, heterotrophic picoplankton populations exhibited cross-species synchronous, tightly regulated, temporally variable patterns of gene expression for many genes, particularly those genes associated with growth and nutrient acquisition. This multitaxon, population-wide gene regulation seemed to reflect sporadic, short-term, reversible responses to high-frequency environmental variability. Although the timing of the environmental responses among different heterotrophic species seemed synchronous, the specific metabolic genes that were expressed varied from taxon to taxon. In aggregate, these results provide insights into the kinetics, diversity, and functional patterns of microbial community response to environmental change. Our results also suggest a means by which complex multispecies metabolic processes could be coordinated, facilitating the regulation of matter and energy processing in a dynamically changing environment.

Oxygen minimum zones (OMZs) are natural physical features of the world's oceans. They create steep physiochemical gradients in the water column, which most notably include a dramatic draw down in oxygen concentrations over small vertical distances (

As part of an ongoing survey of microbial community gene expression in the ocean, we sequenced and compared ∼38 Mbp of community transcriptomes and ∼157 Mbp of community genomes from four bacterioplankton samples, along a defined depth profile at Station ALOHA in North Pacific subtropical gyre (NPSG). Taxonomic analysis suggested that the samples were dominated by three taxa: Prochlorales, Consistiales and Cenarchaeales, which comprised 36–69% and 29–63% of the annotated sequences in the four DNA and four cDNA libraries, respectively. The relative abundance of these taxonomic groups was sometimes very different in the DNA and cDNA libraries, suggesting differential relative transcriptional activities per cell. For example, the 125 m sample genomic library was dominated by Pelagibacter (∼36% of sequence reads), which contributed fewer sequences to the community transcriptome (∼11%). Functional characterization of highly expressed genes suggested taxon-specific contributions to specific biogeochemical processes. Examples included Roseobacter relatives involved in aerobic anoxygenic phototrophy at 75 m, and an unexpected contribution of low abundance Crenarchaea to ammonia oxidation at 125 m. Read recruitment using reference microbial genomes indicated depth-specific partitioning of coexisting microbial populations, highlighted by a transcriptionally active high-light-like Prochlorococcus population in the bottom of the photic zone. Additionally, nutrient-uptake genes dominated Pelagibacter transcripts, with apparent enrichment for certain transporter types (for example, the C4-dicarboxylate transport system) over others (for example, phosphate transporters). In total, the data support the utility of coupled DNA and cDNA analyses for describing taxonomic and functional attributes of microbial communities in their natural habitats.

Background Metagenomic analyses of microbial communities that are comprehensive enough to provide multiple samples of most loci in the genomes of the dominant organism types will also reveal patterns of genetic variation within natural populations. New bioinformatic tools will enable visualization and comprehensive analysis of this sequence variation and inference of recent evolutionary and ecological processes. Results We have developed a software package for analysis and visualization of genetic variation in populations and reconstruction of strain variants from otherwise co-assembled sequences. Sequencing reads can be clustered by matching patterns of single nucleotide polymorphisms to generate predicted gene and protein variant sequences, identify conserved intergenic regulatory sequences, and determine the quantity and distribution of recombination events. Conclusion The Strainer software, a first generation metagenomic bioinformatics tool, facilitates comprehension and analysis of heterogeneity intrinsic in natural communities. The program reveals the degree of clustering among closely related sequence variants and provides a rapid means to generate gene and protein sequences for functional, ecological, and evolutionary analyses.

Background Combined metagenomic and metatranscriptomic datasets make it possible to study the molecular evolution of diverse microbial species recovered from their native habitats. The link between gene expression level and sequence conservation was examined using shotgun pyrosequencing of microbial community DNA and RNA from diverse marine environments, and from forest soil. Results Across all samples, expressed genes with transcripts in the RNA sample were significantly more conserved than non-expressed gene sets relative to best matches in reference databases. This discrepancy, observed for many diverse individual genomes and across entire communities, coincided with a shift in amino acid usage between these gene fractions. Expressed genes trended toward GC-enriched amino acids, consistent with a hypothesis of higher levels of functional constraint in this gene pool. Highly expressed genes were significantly more likely to fall within an orthologous gene set shared between closely related taxa (core genes). However, non-core genes, when expressed above the level of detection, were, on average, significantly more highly expressed than core genes based on transcript abundance normalized to gene abundance. Finally, expressed genes showed broad similarities in function across samples, being relatively enriched in genes of energy metabolism and underrepresented by genes of cell growth. Conclusions These patterns support the hypothesis, predicated on studies of model organisms, that gene expression level is a primary correlate of evolutionary rate across diverse microbial taxa from natural environments. Despite their complexity, meta-omic datasets can reveal broad evolutionary patterns across taxonomically, functionally, and environmentally diverse communities., Gordon and Betty Moore Foundation, Agouron Institute, National Science Foundation (U.S.), Center for Microbial Oceanography: Research and Education