Coping, wellbeing, and psychopathology during high-risk pregnancy: A systematic review.

OBJECTIVE: Complications during pregnancy can negatively impact the physical and psychological wellbeing of mothers. This systematic review aimed to synthesise evidence which has examined the impact of high-risk conditions developed during pregnancy on women's coping, wellbeing, and symptoms of psychopathology. DATA SOURCES: Medline (Ovid), PsycINFO (ProQuest), Scopus, Web of science, AMED (Ebsco), CINAHL (Ebsco) and ProQuest databases were searched in May 2021 with no restrictions on publication date. STUDY SELECTION: English-language literature was reviewed to identify 31 articles. DATA EXTRACTION: Fifteen articles examined Gestational Diabetes Mellitus (GDM: 48%), nine examined multiple high-risk pregnancy conditions (29%), four examined Hypertensive disorders of pregnancy, Preeclampsia (PE: 13%), two did not specify the condition examined (7%), and one examined Pregnancy-Related Acute Kidney Injury (PR-AKI: 3%). The most common study design was quantitative, non-randomised, and survey-based. DATA SYNTHESIS: Twenty-seven articles (87%) reported a high-risk pregnancy resulted in decreased wellbeing and ability to cope, and increased symptoms of psychopathology. The remaining four articles (13%) reported no difference in wellbeing or psychopathology outcomes for women experiencing high-risk compared to healthy pregnancies. Moreover, hypertensive disorders and GDM were associated with ineffective submissive or avoidant coping, reduced wellbeing, and quality-of-life, and exacerbated symptoms of anxiety and depression. CONCLUSION: High-risk pregnancy negatively impacts coping, wellbeing, and psychopathology, and preventive and supportive interventions to mitigate this should focus on empowering women to feel optimistic and in control of their pregnancy. A holistic and culturally sensitive approach is recommended, where pregnant women (and their partners or support people) are involved in healthcare decisions, thus promoting wellbeing, coping, satisfaction, and improved treatment outcome.

Survivorship care planning, quality of life, and confidence to transition to survivorship: A randomized controlled trial with women ending treatment for breast cancer.

PURPOSE: The Polaris Oncology Survivorship Transition (POST) system is a computer-based program that integrates information from the electronic health record, oncology team, and the patient to produce a personalized Survivorship Care Plan. The purpose of this study was to compare the POST to treatment as usual on confidence, quality of life, and interest in mental health referrals in women ending treatment for breast cancer. SAMPLE: Two hundred women (100 POST, 100 treatment as usual) ending treatment for breast cancer were enrolled in a randomized controlled trial. DESIGN: Women randomized to the POST condition received a personalized care plan during a baseline/intervention appointment. At enrollment and baseline/intervention, a number of outcomes were examined in this study, including confidence to enter survivorship measured by the Confidence in Survivorship Index (CSI) and Quality of Life (QOL). One, three, and six month follow up assessments were also conducted. FINDINGS: Treatment groups did not differ in terms of QOL scores at any time points. Mean CSI scores were statistically different between POST and treatment as usual at baseline for the total CSI score and both subscales, but only for confidence in knowledge about prevention and treatment at the 1-month follow-up. All significant differences were in favor of the POST intervention as mean CSI scores were higher for participants who received the POST intervention as opposed to treatment as usual. These findings disappeared at the 3 and 6 month follow up assessments. Finally, patients who received the POST intervention were twice as likely to request mental health/social services referrals compared to women who received treatment as usual. IMPLICATIONS: Oncologists may use the POST to build personalized care plans for women ending treatment for cancer, which may enhance patients' confidence in the short term as well as encourage use of mental health resources.

The Effect of Sexual Concerns on Placement Changes and School Transfers for Youth in the Child Welfare System.

Youths in the child welfare system experience high rates of placement changes and school transfers; therefore, prior research focused on variables that may be linked with such disruptions. Indeed, researchers have established that mental health symptoms (e.g., PTSD symptoms) are linked with placement disruptions. However, an important aspect of mental health for youth in the child welfare system has largely been ignored: sexual concerns (e.g., distress, preoccupation). Thus, the present study evaluated whether higher levels of sexual preoccupation and distress among a sample of child welfare-involved youths (N = 124) in a northeastern state predicted placement changes and school transfers above and beyond variables previously linked with these disruptions. Our hypotheses were partially supported such that higher levels of sexual distress were linked with increased odds of experiencing a placement change (OE = 2.60; p <.01). Counter to our hypotheses, higher levels of sexual preoccupation were linked with lower odds of experiencing both placement changes (OE = -2.98; p <.01) and school transfers (OR = 0.18; p < .05). Furthermore, sexual preoccupation and sexual distress were not linked with increased rates of placement changes. The current findings have implications for the assessment of sexual concerns and the prevention of placement changes among youth in the child welfare system.

Feasibility and acceptability of a brief suicide intervention for youth involved with the family court.

As efforts to develop models for suicide prevention and intervention in the juvenile justice (JJ) system continue to grow, research to understand the feasibility and acceptability of implementing these models is critical. Examining organizational readiness for implementation, ensuring leadership and staff buy-in for delivering the intervention, and planning for sustainability of staff participation in implementation efforts is essential. The current study involved semi-structured formative evaluation interviews with key JJ stakeholders (n = 10) to determine perspectives on the acceptability (perceived need and fit of the intervention) and feasibility (organizational readiness for change) of a proposed brief safety planning intervention for youth with suicidal ideation delivered by nonclinical staff and integrated into the existing system. Qualitative data revealed stakeholders' perceived need for the intervention in the family court context and their agreement that the aims of the intervention were congruent with the goals of the family court. Some barriers to successful implementation were noted, which, addressed through selection of appropriate implementation strategies, can be overcome in a future test of the safety intervention.

Comparing the efficacy of a brief self-esteem and self-compassion intervention for state body dissatisfaction and self-improvement motivation.

This study explored the efficacy of reflective writing tasks for reducing state body dissatisfaction and improving self-improvement motivation. Participants (N = 153) were exposed to a threatening body image scenario, after which they reported their state body dissatisfaction. Following randomisation to one of three interventions (self-esteem, self-compassion, or positive distraction control), participants completed the state body dissatisfaction measures again along with a measure of self-improvement motivation. At post-intervention, state weight dissatisfaction and appearance dissatisfaction were significantly lower and self-improvement motivation was significantly higher in the self-compassion group than in both the self-esteem and control groups. Trait body dissatisfaction moderated the efficacy of the intervention whereby the benefit of the self-compassion intervention became evident at moderate levels of trait body dissatisfaction, and was most apparent at high levels of body dissatisfaction. Self-compassion was a robust and efficacious method through which to promote immediate improvements to bodily feelings and desire to self-improve.

Ribonucleotide reductases reveal novel viral diversity and predict biological and ecological features of unknown marine viruses.

Virioplankton play a crucial role in aquatic ecosystems as top-down regulators of bacterial populations and agents of horizontal gene transfer and nutrient cycling. However, the biology and ecology of virioplankton populations in the environment remain poorly understood. Ribonucleotide reductases (RNRs) are ancient enzymes that reduce ribonucleotides to deoxyribonucleotides and thus prime DNA synthesis. Composed of three classes according to O2 reactivity, RNRs can be predictive of the physiological conditions surrounding DNA synthesis. RNRs are universal among cellular life, common within viral genomes and virioplankton shotgun metagenomes (viromes), and estimated to occur within >90% of the dsDNA virioplankton sampled in this study. RNRs occur across diverse viral groups, including all three morphological families of tailed phages, making these genes attractive for studies of viral diversity. Differing patterns in virioplankton diversity were clear from RNRs sampled across a broad oceanic transect. The most abundant RNRs belonged to novel lineages of podoviruses infecting α-proteobacteria, a bacterial class critical to oceanic carbon cycling. RNR class was predictive of phage morphology among cyanophages and RNR distribution frequencies among cyanophages were largely consistent with the predictions of the "kill the winner-cost of resistance" model. RNRs were also identified for the first time to our knowledge within ssDNA viromes. These data indicate that RNR polymorphism provides a means of connecting the biological and ecological features of virioplankton populations.

Shotgun metagenomics indicates novel family A DNA polymerases predominate within marine virioplankton.

Virioplankton have a significant role in marine ecosystems, yet we know little of the predominant biological characteristics of aquatic viruses that influence the flow of nutrients and energy through microbial communities. Family A DNA polymerases, critical to DNA replication and repair in prokaryotes, are found in many tailed bacteriophages. The essential role of DNA polymerase in viral replication makes it a useful target for connecting viral diversity with an important biological feature of viruses. Capturing the full diversity of this polymorphic gene by targeted approaches has been difficult; thus, full-length DNA polymerase genes were assembled out of virioplankton shotgun metagenomic sequence libraries (viromes). Within the viromes novel DNA polymerases were common and found in both double-stranded (ds) DNA and single-stranded (ss) DNA libraries. Finding DNA polymerase genes in ssDNA viral libraries was unexpected, as no such genes have been previously reported from ssDNA phage. Surprisingly, the most common virioplankton DNA polymerases were related to a siphovirus infecting an α-proteobacterial symbiont of a marine sponge and not the podoviral T7-like polymerases seen in many other studies. Amino acids predictive of catalytic efficiency and fidelity linked perfectly to the environmental clades, indicating that most DNA polymerase-carrying virioplankton utilize a lower efficiency, higher fidelity enzyme. Comparisons with previously reported, PCR-amplified DNA polymerase sequences indicated that the most common virioplankton metagenomic DNA polymerases formed a new group that included siphoviruses. These data indicate that slower-replicating, lytic or lysogenic phage populations rather than fast-replicating, highly lytic phages may predominate within the virioplankton.

A metagenomic framework for the study of airborne microbial communities.

Understanding the microbial content of the air has important scientific, health, and economic implications. While studies have primarily characterized the taxonomic content of air samples by sequencing the 16S or 18S ribosomal RNA gene, direct analysis of the genomic content of airborne microorganisms has not been possible due to the extremely low density of biological material in airborne environments. We developed sampling and amplification methods to enable adequate DNA recovery to allow metagenomic profiling of air samples collected from indoor and outdoor environments. Air samples were collected from a large urban building, a medical center, a house, and a pier. Analyses of metagenomic data generated from these samples reveal airborne communities with a high degree of diversity and different genera abundance profiles. The identities of many of the taxonomic groups and protein families also allows for the identification of the likely sources of the sampled airborne bacteria.

Diffuse flow environments within basalt- and sediment-based hydrothermal vent ecosystems harbor specialized microbial communities.

Hydrothermal vents differ both in surface input and subsurface geochemistry. The effects of these differences on their microbial communities are not clear. Here, we investigated both alpha and beta diversity of diffuse flow-associated microbial communities emanating from vents at a basalt-based hydrothermal system along the East Pacific Rise (EPR) and a sediment-based hydrothermal system, Guaymas Basin. Both Bacteria and Archaea were targeted using high throughput 16S rRNA gene pyrosequencing analyses. A unique aspect of this study was the use of a universal set of 16S rRNA gene primers to characterize total and diffuse flow-specific microbial communities from varied deep-sea hydrothermal environments. Both surrounding seawater and diffuse flow water samples contained large numbers of Marine Group I (MGI) Thaumarchaea and Gammaproteobacteria taxa previously observed in deep-sea systems. However, these taxa were geographically distinct and segregated according to type of spreading center. Diffuse flow microbial community profiles were highly differentiated. In particular, EPR dominant diffuse flow taxa were most closely associated with chemolithoautotrophs, and off axis water was dominated by heterotrophic-related taxa, whereas the opposite was true for Guaymas Basin. The diversity and richness of diffuse flow-specific microbial communities were strongly correlated to the relative abundance of Epsilonproteobacteria, proximity to macrofauna, and hydrothermal system type. Archaeal diversity was higher than or equivalent to bacterial diversity in about one third of the samples. Most diffuse flow-specific communities were dominated by OTUs associated with Epsilonproteobacteria, but many of the Guaymas Basin diffuse flow samples were dominated by either OTUs within the Planctomycetes or hyperthermophilic Archaea. This study emphasizes the unique microbial communities associated with geochemically and geographically distinct hydrothermal diffuse flow environments.

Isolation and genome analysis of single virions using 'single virus genomics'.

Whole genome amplification and sequencing of single microbial cells enables genomic characterization without the need of cultivation (1-3). Viruses, which are ubiquitous and the most numerous entities on our planet (4) and important in all environments (5), have yet to be revealed via similar approaches. Here we describe an approach for isolating and characterizing the genomes of single virions called 'Single Virus Genomics' (SVG). SVG utilizes flow cytometry to isolate individual viruses and whole genome amplification to obtain high molecular weight genomic DNA (gDNA) that can be used in subsequent sequencing reactions.

Metagenomic investigation of microbes and viruses in patients with jaw osteonecrosis associated with bisphosphonate therapy.

OBJECTIVE: The goal of this preliminary study was to use metagenomic approaches to investigate the taxonomic diversity of microorganisms in patients with bisphosphonate-related osteonecrosis of the jaw (BRONJ). STUDY DESIGN: Samples of saliva for planktonic microbial analysis and biofilm cultivation were collected from 10 patients (5 with BRONJ and 5 non-BRONJ control subjects) who met all ascertainment criteria. Prophage induction experiments-16S rRNA polymerase chain reaction and 454 pyrosequencing-and epifluorescent microscopy were performed for characterization and enumeration of microbes and viruses. RESULTS: Three phyla of microbes-Proteobacteria (70%), Firmicutes (26.9%), and Actinobacteria (1.95%)-dominated all BRONJ samples and accounted for almost 99% of the total data. Viral abundance was ∼1 order of magnitude greater than microbial cell abundance and comprised mainly phage viruses. CONCLUSIONS: Individuals with jaw osteonecrosis harbored different microbial assemblages than nonaffected patients, and in general viral abundance and prophage induction increased with biofilm formation, suggesting that biofilm formation encouraged lysogenic interactions between viruses and microbial hosts and may contribute to pathogenicity.

Metagenomic exploration of viruses throughout the Indian Ocean.

The characterization of global marine microbial taxonomic and functional diversity is a primary goal of the Global Ocean Sampling Expedition. As part of this study, 19 water samples were collected aboard the Sorcerer II sailing vessel from the southern Indian Ocean in an effort to more thoroughly understand the lifestyle strategies of the microbial inhabitants of this ultra-oligotrophic region. No investigations of whole virioplankton assemblages have been conducted on waters collected from the Indian Ocean or across multiple size fractions thus far. Therefore, the goals of this study were to examine the effect of size fractionation on viral consortia structure and function and understand the diversity and functional potential of the Indian Ocean virome. Five samples were selected for comprehensive metagenomic exploration; and sequencing was performed on the microbes captured on 3.0-, 0.8- and 0.1 µm membrane filters as well as the viral fraction (<0.1 µm). Phylogenetic approaches were also used to identify predicted proteins of viral origin in the larger fractions of data from all Indian Ocean samples, which were included in subsequent metagenomic analyses. Taxonomic profiling of viral sequences suggested that size fractionation of marine microbial communities enriches for specific groups of viruses within the different size classes and functional characterization further substantiated this observation. Functional analyses also revealed a relative enrichment for metabolic proteins of viral origin that potentially reflect the physiological condition of host cells in the Indian Ocean including those involved in nitrogen metabolism and oxidative phosphorylation. A novel classification method, MGTAXA, was used to assess virus-host relationships in the Indian Ocean by predicting the taxonomy of putative host genera, with Prochlorococcus, Acanthochlois and members of the SAR86 cluster comprising the most abundant predictions. This is the first study to holistically explore virioplankton dynamics across multiple size classes and provides unprecedented insight into virus diversity, metabolic potential and virus-host interactions.

Groundtruthing next-gen sequencing for microbial ecology-biases and errors in community structure estimates from PCR amplicon pyrosequencing.

Analysis of microbial communities by high-throughput pyrosequencing of SSU rRNA gene PCR amplicons has transformed microbial ecology research and led to the observation that many communities contain a diverse assortment of rare taxa-a phenomenon termed the Rare Biosphere. Multiple studies have investigated the effect of pyrosequencing read quality on operational taxonomic unit (OTU) richness for contrived communities, yet there is limited information on the fidelity of community structure estimates obtained through this approach. Given that PCR biases are widely recognized, and further unknown biases may arise from the sequencing process itself, a priori assumptions about the neutrality of the data generation process are at best unvalidated. Furthermore, post-sequencing quality control algorithms have not been explicitly evaluated for the accuracy of recovered representative sequences and its impact on downstream analyses, reducing useful discussion on pyrosequencing reads to their diversity and abundances. Here we report on community structures and sequences recovered for in vitro-simulated communities consisting of twenty 16S rRNA gene clones tiered at known proportions. PCR amplicon libraries of the V3-V4 and V6 hypervariable regions from the in vitro-simulated communities were sequenced using the Roche 454 GS FLX Titanium platform. Commonly used quality control protocols resulted in the formation of OTUs with >1% abundance composed entirely of erroneous sequences, while over-aggressive clustering approaches obfuscated real, expected OTUs. The pyrosequencing process itself did not appear to impose significant biases on overall community structure estimates, although the detection limit for rare taxa may be affected by PCR amplicon size and quality control approach employed. Meanwhile, PCR biases associated with the initial amplicon generation may impose greater distortions in the observed community structure.

Influence of nutrients and currents on the genomic composition of microbes across an upwelling mosaic.

Metagenomic data sets were generated from samples collected along a coastal to open ocean transect between Southern California Bight and California Current waters during a seasonal upwelling event, providing an opportunity to examine the impact of episodic pulses of cold nutrient-rich water into surface ocean microbial communities. The data set consists of ~5.8 million predicted proteins across seven sites, from three different size classes: 0.1-0.8, 0.8-3.0 and 3.0-200.0 µm. Taxonomic and metabolic analyses suggest that sequences from the 0.1-0.8 µm size class correlated with their position along the upwelling mosaic. However, taxonomic profiles of bacteria from the larger size classes (0.8-200 µm) were less constrained by habitat and characterized by an increase in Cyanobacteria, Bacteroidetes, Flavobacteria and double-stranded DNA viral sequences. Functional annotation of transmembrane proteins indicate that sites comprised of organisms with small genomes have an enrichment of transporters with substrate specificities for amino acids, iron and cadmium, whereas organisms with larger genomes have a higher percentage of transporters for ammonium and potassium. Eukaryotic-type glutamine synthetase (GS) II proteins were identified and taxonomically classified as viral, most closely related to the GSII in Mimivirus, suggesting that marine Mimivirus-like particles may have played a role in the transfer of GSII gene functions. Additionally, a Planctomycete bloom was sampled from one upwelling site providing a rare opportunity to assess the genomic composition of a marine Planctomycete population. The significant correlations observed between genomic properties, community structure and nutrient availability provide insights into habitat-driven dynamics among oligotrophic versus upwelled marine waters adjoining each other spatially.

From bacterial to microbial ecosystems (metagenomics).

Metagenomics is revolutionizing the field of microbial ecology through techniques that eliminate the prerequisite of culturing. Metagenomic studies of microbial populations in different environments reveal the incredible diversity and adaptive capabilities of these organisms. With the advent of cheaper, high-throughput sequencing technologies, these studies are also producing vast amounts of sequence data. Here, we discuss the different components of a metagenomic study including sample collection, DNA extraction, sequencing, and informatics. We highlight their issues and challenges, and review the solutions that are currently in use. We conclude with examples of metagenomic studies conducted on environments of varying complexities.

Separation of single-stranded DNA, double-stranded DNA and RNA from an environmental viral community using hydroxyapatite chromatography.

Viruses, particularly bacteriophages (phages), are the most numerous biological entities on Earth. Viruses modulate host cell abundance and diversity, contribute to the cycling of nutrients, alter host cell phenotype, and influence the evolution of both host cell and viral communities through the lateral transfer of genes. Numerous studies have highlighted the staggering genetic diversity of viruses and their functional potential in a variety of natural environments. Metagenomic techniques have been used to study the taxonomic diversity and functional potential of complex viral assemblages whose members contain single-stranded DNA (ssDNA), double-stranded DNA (dsDNA) and RNA genotypes. Current library construction protocols used to study environmental DNA-containing or RNA-containing viruses require an initial nuclease treatment in order to remove nontargeted templates. However, a comprehensive understanding of the collective gene complement of the virus community and virus diversity requires knowledge of all members regardless of genome composition. Fractionation of purified nucleic acid subtypes provides an effective mechanism by which to study viral assemblages without sacrificing a subset of the community's genetic signature. Hydroxyapatite, a crystalline form of calcium phosphate, has been employed in the separation of nucleic acids, as well as proteins and microbes, since the 1960s. By exploiting the charge interaction between the positively-charged Ca(2+) ions of the hydroxyapatite and the negatively charged phosphate backbone of the nucleic acid subtypes, it is possible to preferentially elute each nucleic acid subtype independent of the others. We recently employed this strategy to independently fractionate the genomes of ssDNA, dsDNA and RNA-containing viruses in preparation of DNA sequencing. Here, we present a method for the fractionation and recovery of ssDNA, dsDNA and RNA viral nucleic acids from mixed viral assemblages using hydroxyapatite chromatography.

TheViral MetaGenome Annotation Pipeline(VMGAP):an automated tool for the functional annotation of viral Metagenomic shotgun sequencing data.

In the past few years, the field of metagenomics has been growing at an accelerated pace, particularly in response to advancements in new sequencing technologies. The large volume of sequence data from novel organisms generated by metagenomic projects has triggered the development of specialized databases and tools focused on particular groups of organisms or data types. Here we describe a pipeline for the functional annotation of viral metagenomic sequence data. The Viral MetaGenome Annotation Pipeline (VMGAP) pipeline takes advantage of a number of specialized databases, such as collections of mobile genetic elements and environmental metagenomes to improve the classification and functional prediction of viral gene products. The pipeline assigns a functional term to each predicted protein sequence following a suite of comprehensive analyses whose results are ranked according to a priority rules hierarchy. Additional annotation is provided in the form of enzyme commission (EC) numbers, GO/MeGO terms and Hidden Markov Models together with supporting evidence.

Repeating patterns of virioplankton production within an estuarine ecosystem.

The Chesapeake Bay, a seasonally variable temperate estuary, provides a natural laboratory for examining the fluctuations and impacts of viral lysis on aquatic microorganisms. Viral abundance (VA) and viral production (VP) were monitored in the Chesapeake Bay over 4 1/2 annual cycles, producing a unique, long-term, interannual study of virioplankton production. High and dynamic VP rates, averaging 7.9 × 10(6) viruses per mL per h, indicate that viral lysis impacts a significant fraction of microorganisms in the Chesapeake. Viral-mediated bacterial mortality, VA, VP, and organic carbon release all displayed similar interannual and seasonal trends with higher values in 2003 and 2006 than in 2004 and 2005 and peaks in early spring and summer. Surprisingly, higher rates of viral lysis occurred in winter, resulting in a magnified effect of viral lysis on bacterioplankton during times of reduced productivity. Viral lysis directly impacted the organic carbon pool, contributing on average 76 µg of C per L per d, an amount capable of sustaining ∼55% of Chesapeake Bay bacterial production. The observed repeating interannual patterns of VP and lysis are likely interlinked with seasonal cycles of host abundance and diversity, which are in turn driven by annual cycles in environmental conditions, emphasizing the complex interplay of seasonality and microbial ecology in the Chesapeake Bay.

Going deeper: metagenome of a hadopelagic microbial community.

The paucity of sequence data from pelagic deep-ocean microbial assemblages has severely restricted molecular exploration of the largest biome on Earth. In this study, an analysis is presented of a large-scale 454-pyrosequencing metagenomic dataset from a hadopelagic environment from 6,000 m depth within the Puerto Rico Trench (PRT). A total of 145 Mbp of assembled sequence data was generated and compared to two pelagic deep ocean metagenomes and two representative surface seawater datasets from the Sargasso Sea. In a number of instances, all three deep metagenomes displayed similar trends, but were most magnified in the PRT, including enrichment in functions for two-component signal transduction mechanisms and transcriptional regulation. Overrepresented transporters in the PRT metagenome included outer membrane porins, diverse cation transporters, and di- and tri-carboxylate transporters that matched well with the prevailing catabolic processes such as butanoate, glyoxylate and dicarboxylate metabolism. A surprisingly high abundance of sulfatases for the degradation of sulfated polysaccharides were also present in the PRT. The most dramatic adaptational feature of the PRT microbes appears to be heavy metal resistance, as reflected in the large numbers of transporters present for their removal. As a complement to the metagenome approach, single-cell genomic techniques were utilized to generate partial whole-genome sequence data from four uncultivated cells from members of the dominant phyla within the PRT, Alphaproteobacteria, Gammaproteobacteria, Bacteroidetes and Planctomycetes. The single-cell sequence data provided genomic context for many of the highly abundant functional attributes identified from the PRT metagenome, as well as recruiting heavily the PRT metagenomic sequence data compared to 172 available reference marine genomes. Through these multifaceted sequence approaches, new insights have been provided into the unique functional attributes present in microbes residing in a deeper layer of the ocean far removed from the more productive sun-drenched zones above.

Single virus genomics: a new tool for virus discovery.

Whole genome amplification and sequencing of single microbial cells has significantly influenced genomics and microbial ecology by facilitating direct recovery of reference genome data. However, viral genomics continues to suffer due to difficulties related to the isolation and characterization of uncultivated viruses. We report here on a new approach called 'Single Virus Genomics', which enabled the isolation and complete genome sequencing of the first single virus particle. A mixed assemblage comprised of two known viruses; E. coli bacteriophages lambda and T4, were sorted using flow cytometric methods and subsequently immobilized in an agarose matrix. Genome amplification was then achieved in situ via multiple displacement amplification (MDA). The complete lambda phage genome was recovered with an average depth of coverage of approximately 437X. The isolation and genome sequencing of uncultivated viruses using Single Virus Genomics approaches will enable researchers to address questions about viral diversity, evolution, adaptation and ecology that were previously unattainable.