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Open accessJournal ArticleDOI: 10.3390/MICROORGANISMS9030524

Biases in Viral Metagenomics-Based Detection, Cataloguing and Quantification of Bacteriophage Genomes in Human Faeces, a Review

04 Mar 2021-Vol. 9, Iss: 3, pp 524
Abstract: The human gut is colonised by a vast array of microbes that include bacteria, viruses, fungi, and archaea. While interest in these microbial entities has largely focused on the bacterial constituents, recently the viral component has attracted more attention. Metagenomic advances, compared to classical isolation procedures, have greatly enhanced our understanding of the composition, diversity, and function of viruses in the human microbiome (virome). We highlight that viral extraction methodologies are crucial in terms of identifying and characterising communities of viruses infecting eukaryotes and bacteria. Different viral extraction protocols, including those used in some of the most significant human virome publications to date, have introduced biases affecting their a overall conclusions. It is important that protocol variations should be clearly highlighted across studies, with the ultimate goal of identifying and acknowledging biases associated with different protocols and, perhaps, the generation of an unbiased and standardised method for examining this portion of the human microbiome.

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Topics: Human virome (66%), Viral metagenomics (61%), Metagenomics (55%) ... show more
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5 results found


Journal ArticleDOI: 10.1038/S41579-021-00602-Y
Abstract: We commonly acknowledge that bacterial viruses (phages) shape the composition and evolution of bacterial communities in nature and therefore have important roles in ecosystem functioning. This view stems from studies in the 1990s to the first decade of the twenty-first century that revealed high viral abundance, high viral diversity and virus-induced microbial death in aquatic ecosystems as well as an association between collapses in bacterial density and peaks in phage abundance. The recent surge in metagenomic analyses has provided deeper insight into the abundance, genomic diversity and spatio-temporal dynamics of phages in a wide variety of ecosystems, ranging from deep oceans to soil and the mammalian digestive tract. However, the causes and consequences of variations in phage community compositions remain poorly understood. In this Review, we explore current knowledge of the composition and evolution of phage communities, as well as their roles in controlling the population and evolutionary dynamics of bacterial communities. We discuss the need for greater ecological realism in laboratory studies to capture the complexity of microbial communities that thrive in natural environments.

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Topics: Bacterial virus (60%), Population (51%)

3 Citations


Open accessPosted ContentDOI: 10.21203/RS.3.RS-803286/V1
01 Sep 2021-
Abstract: In this study we report the first comprehensive metagenomic analysis of the prokaryotic and eukaryotic virome occupying luminal and mucosa-associated habitats along the entire length of the gastrointestinal tract (GIT) in two animal species, the domestic pig and rhesus macaque. The highest loads and diversity of bacteriophages are found in the lumen of the large intestine in both mammals. Mucosal samples contain much lower viral loads but a higher proportion of eukaryotic viruses. Parenchymal organs contained significant amounts of bacteriophages of gut origin, in addition to some eukaryotic viruses. GIT virome composition is both region- and species-specific with a strong separation between upper and lower gut. Nonetheless, certain viral and phage species are found ubiquitously from the oral cavity to the distal colon. Correlations between individual phages and their potential microbial hosts in the GIT are overwhelmingly positive, which confirms earlier concepts of the temperate-like life cycles of the majority of gut phages and a prevalence of the “piggyback-the-winner” ecological dynamics.

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Topics: Gastrointestinal tract (57%)

1 Citations


Journal ArticleDOI: 10.1016/J.COVIRO.2021.05.008
Sean Benler1, Eugene V. Koonin1Institutions (1)
Abstract: Metagenomics and metatranscriptomics have become the principal approaches for discovery of novel bacteriophages and preliminary characterization of their ecology and biology. Metagenomic sequencing dramatically expanded the known diversity of tailed and non-tailed phages with double-stranded DNA genomes and those with single-stranded DNA genomes, whereas metatranscriptomics led to the discovery of thousands of new single-stranded RNA phages. Apart from expanding phage diversity, metagenomics studies discover major novel groups of phages with unique features of genome organization, expression strategy and virus-host interaction, such as the putative order 'crAssvirales', which includes the most abundant human-associated viruses. The continued success of metagenomics hinges on the combination of the most powerful computational methods for phage genome assembly and analysis including harnessing CRISPR spacers for the discovery of novel phages and host assignment. Together, these approaches could make a comprehensive characterization of the earth phageome a realistic goal.

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Topics: RNA Phages (62%), Metagenomics (56%), Genome (51%)

1 Citations



Open accessDOI: 10.1099/MGEN.0.000686
01 Nov 2021-
Abstract: The vast majority of described prokaryotic viruses have double-stranded or single-stranded DNA or double-stranded RNA genomes. Until 2020, a mere four prokaryotic single-stranded, positive-sense RNA viruses have been classified in two genera (Riboviria; Lenarviricota; Allassoviricetes; Leviviridae). Several recent metagenomic and metatranscriptomic studies revealed a vastly greater diversity of these viruses in prokaryotic soil communities than ever anticipated. Phylogenetic analysis of these newly discovered viruses prompted the reorganization of class Allassoviricetes, now renamed Leviviricetes, to include two orders, Norzivirales and Timlovirales, and a total of six families, 428 genera and 882 species. Here we outline the new taxonomy of Leviviricetes, approved and ratified in 2021 by the International Committee on Taxonomy of Viruses, and describe open-access hidden Markov models to accommodate the anticipated identification and future classification of hundreds, if not thousands, of additional class members into this new taxonomic framework.

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Topics: Virus classification (59%)
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80 results found


Open accessJournal ArticleDOI: 10.1073/PNAS.082089499
Abstract: Fundamental to most genetic analysis is availability of genomic DNA of adequate quality and quantity. Because DNA yield from human samples is frequently limiting, much effort has been invested in developing methods for whole genome amplification (WGA) by random or degenerate oligonucleotide-primed PCR. However, existing WGA methods like degenerate oligonucleotide-primed PCR suffer from incomplete coverage and inadequate average DNA size. We describe a method, termed multiple displacement amplification (MDA), which provides a highly uniform representation across the genome. Amplification bias among eight chromosomal loci was less than 3-fold in contrast to 4–6 orders of magnitude for PCR-based WGA methods. Average product length was >10 kb. MDA is an isothermal, strand-displacing amplification yielding about 20–30 μg product from as few as 1–10 copies of human genomic DNA. Amplification can be carried out directly from biological samples including crude whole blood and tissue culture cells. MDA-amplified human DNA is useful for several common methods of genetic analysis, including genotyping of single nucleotide polymorphisms, chromosome painting, Southern blotting and restriction fragment length polymorphism analysis, subcloning, and DNA sequencing. MDA-based WGA is a simple and reliable method that could have significant implications for genetic studies, forensics, diagnostics, and long-term sample storage.

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1,466 Citations


Open accessJournal ArticleDOI: 10.1038/NATURE09199
Alejandro Reyes1, Matthew Haynes2, Nicole Hanson2, Florent E. Angly3  +4 moreInstitutions (3)
15 Jul 2010-Nature
Abstract: Viral diversity and life cycles are poorly understood in the human gut and other body habitats. Phages and their encoded functions may provide informative signatures of a human microbiota and of microbial community responses to various disturbances, and may indicate whether community health or dysfunction is manifest after apparent recovery from a disease or therapeutic intervention. Here we report sequencing of the viromes (metagenomes) of virus-like particles isolated from faecal samples collected from healthy adult female monozygotic twins and their mothers at three time points over a one-year period. We compared these data sets with data sets of sequenced bacterial 16S ribosomal RNA genes and total-faecal-community DNA. Co-twins and their mothers share a significantly greater degree of similarity in their faecal bacterial communities than do unrelated individuals. In contrast, viromes are unique to individuals regardless of their degree of genetic relatedness. Despite remarkable interpersonal variations in viromes and their encoded functions, intrapersonal diversity is very low, with >95% of virotypes retained over the period surveyed, and with viromes dominated by a few temperate phages that exhibit remarkable genetic stability. These results indicate that a predatory viral-microbial dynamic, manifest in a number of other characterized environmental ecosystems, is notably absent in the very distal intestine.

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Topics: Monozygotic twin (55%)

916 Citations


Journal ArticleDOI: 10.1038/NATURE20167
Mang Shi1, Xian-Dan Lin2, Jun-Hua Tian2, Liangjun Chen1  +12 moreInstitutions (4)
22 Dec 2016-Nature
Abstract: Current knowledge of RNA virus biodiversity is both biased and fragmentary, reflecting a focus on culturable or disease-causing agents. Here we profile the transcriptomes of over 220 invertebrate species sampled across nine animal phyla and report the discovery of 1,445 RNA viruses, including some that are sufficiently divergent to comprise new families. The identified viruses fill major gaps in the RNA virus phylogeny and reveal an evolutionary history that is characterized by both host switching and co-divergence. The invertebrate virome also reveals remarkable genomic flexibility that includes frequent recombination, lateral gene transfer among viruses and hosts, gene gain and loss, and complex genomic rearrangements. Together, these data present a view of the RNA virosphere that is more phylogenetically and genomically diverse than that depicted in current classification schemes and provide a more solid foundation for studies in virus ecology and evolution.

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Topics: Human virome (58%), Viral evolution (57%), RNA virus (57%) ... show more

840 Citations


Open accessJournal ArticleDOI: 10.1101/GR.122705.111
Samuel S. Minot1, Rohini Sinha1, Jun Chen, Hongzhe Li1  +4 moreInstitutions (1)
01 Oct 2011-Genome Research
Abstract: Immense populations of viruses are present in the human gut and other body sites. Understanding the role of these populations (the human "virome") in health and disease requires a much deeper understanding of their composition and dynamics in the face of environmental perturbation. Here, we investigate viromes from human subjects on a controlled feeding regimen. Longitudinal fecal samples were analyzed by metagenomic sequencing of DNA from virus-like particles (VLP) and total microbial communities. Assembly of 336 Mb of VLP sequence yielded 7175 contigs, many identifiable as complete or partial bacteriophage genomes. Contigs were rich in viral functions required in lytic and lysogenic growth, as well as unexpected functions such as viral CRISPR arrays and genes for antibiotic resistance. The largest source of variance among virome samples was interpersonal variation. Parallel deep-sequencing analysis of bacterial populations showed covaration of the virome with the larger microbiome. The dietary intervention was associated with a change in the virome community to a new state, in which individuals on the same diet converged. Thus these data provide an overview of the composition of the human gut virome and associate virome structure with diet.

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Topics: Human virome (69%), Microbiome (52%), Metagenomics (51%)

715 Citations


Open accessJournal ArticleDOI: 10.1101/GR.131383.111
01 Mar 2012-Genome Research
Abstract: New sequencing technology has dramatically altered the landscape of whole-genome sequencing, allowing scientists to initiate numerous projects to decode the genomes of previously unsequenced organisms. The lowest-cost technology can generate deep coverage of most species, including mammals, in just a few days. The sequence data generated by one of these projects consist of millions or billions of short DNA sequences (reads) that range from 50 to 150 nt in length. These sequences must then be assembled de novo before most genome analyses can begin. Unfortunately, genome assembly remains a very difficult problem, made more difficult by shorter reads and unreliable long-range linking information. In this study, we evaluated several of the leading de novo assembly algorithms on four different short-read data sets, all generated by Illumina sequencers. Our results describe the relative performance of the different assemblers as well as other significant differences in assembly difficulty that appear to be inherent in the genomes themselves. Three overarching conclusions are apparent: first, that data quality, rather than the assembler itself, has a dramatic effect on the quality of an assembled genome; second, that the degree of contiguity of an assembly varies enormously among different assemblers and different genomes; and third, that the correctness of an assembly also varies widely and is not well correlated with statistics on contiguity. To enable others to replicate our results, all of our data and methods are freely available, as are all assemblers used in this study.

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Topics: Hybrid genome assembly (65%), Sequence assembly (59%), Genome project (55%) ... show more

712 Citations


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