Evaluation of methods to concentrate and purify ocean virus communities through comparative, replicated metagenomics
Identifieur interne : 000C38 ( Pmc/Curation ); précédent : 000C37; suivant : 000C39Evaluation of methods to concentrate and purify ocean virus communities through comparative, replicated metagenomics
Auteurs : Bonnie L. Hurwitz ; Li Deng ; Bonnie T. Poulos ; Matthew B. SullivanSource :
- Environmental Microbiology [ 1462-2912 ] ; 2013.
Abstract
Viruses have global impact through mortality, nutrient cycling and horizontal gene transfer, yet their study is limited by complex methodologies with little validation. Here, we use triplicate metagenomes to compare common aquatic viral concentration and purification methods across four combinations as follows: (i) tangential flow filtration (TFF) and DNase + CsCl, (ii) FeCl3 precipitation and DNase, (iii) FeCl3 precipitation and DNase + CsCl and (iv) FeCl3 precipitation and DNase + sucrose. Taxonomic data (30% of reads) suggested that purification methods were statistically indistinguishable at any taxonomic level while concentration methods were significantly different at family and genus levels. Specifically, TFF-concentrated viral metagenomes had significantly fewer abundant viral types (
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DOI: 10.1111/j.1462-2920.2012.02836.x
PubMed: 22845467
PubMed Central: 3655615
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<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Evaluation of methods to concentrate and purify ocean virus communities through comparative, replicated metagenomics</title><author><name sortKey="Hurwitz, Bonnie L" sort="Hurwitz, Bonnie L" uniqKey="Hurwitz B" first="Bonnie L" last="Hurwitz">Bonnie L. Hurwitz</name></author><author><name sortKey="Deng, Li" sort="Deng, Li" uniqKey="Deng L" first="Li" last="Deng">Li Deng</name></author><author><name sortKey="Poulos, Bonnie T" sort="Poulos, Bonnie T" uniqKey="Poulos B" first="Bonnie T" last="Poulos">Bonnie T. Poulos</name></author><author><name sortKey="Sullivan, Matthew B" sort="Sullivan, Matthew B" uniqKey="Sullivan M" first="Matthew B" last="Sullivan">Matthew B. Sullivan</name></author></titleStmt><publicationStmt><idno type="wicri:source">PMC</idno><idno type="pmid">22845467</idno><idno type="pmc">3655615</idno><idno type="url">http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3655615</idno><idno type="RBID">PMC:3655615</idno><idno type="doi">10.1111/j.1462-2920.2012.02836.x</idno><date when="2013">2013</date><idno type="wicri:Area/Pmc/Corpus">000C38</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Corpus" wicri:corpus="PMC">000C38</idno><idno type="wicri:Area/Pmc/Curation">000C38</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Curation">000C38</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en" level="a" type="main">Evaluation of methods to concentrate and purify ocean virus communities through comparative, replicated metagenomics</title><author><name sortKey="Hurwitz, Bonnie L" sort="Hurwitz, Bonnie L" uniqKey="Hurwitz B" first="Bonnie L" last="Hurwitz">Bonnie L. Hurwitz</name></author><author><name sortKey="Deng, Li" sort="Deng, Li" uniqKey="Deng L" first="Li" last="Deng">Li Deng</name></author><author><name sortKey="Poulos, Bonnie T" sort="Poulos, Bonnie T" uniqKey="Poulos B" first="Bonnie T" last="Poulos">Bonnie T. Poulos</name></author><author><name sortKey="Sullivan, Matthew B" sort="Sullivan, Matthew B" uniqKey="Sullivan M" first="Matthew B" last="Sullivan">Matthew B. Sullivan</name></author></analytic><series><title level="j">Environmental Microbiology</title><idno type="ISSN">1462-2912</idno><idno type="eISSN">1462-2920</idno><imprint><date when="2013">2013</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p>Viruses have global impact through mortality, nutrient cycling and horizontal gene transfer, yet their study is limited by complex methodologies with little validation. Here, we use triplicate metagenomes to compare common aquatic viral concentration and purification methods across four combinations as follows: (i) tangential flow filtration (TFF) and DNase + CsCl, (ii) FeCl<sub>3</sub> precipitation and DNase, (iii) FeCl<sub>3</sub> precipitation and DNase + CsCl and (iv) FeCl<sub>3</sub> precipitation and DNase + sucrose. Taxonomic data (30% of reads) suggested that purification methods were statistically indistinguishable at any taxonomic level while concentration methods were significantly different at family and genus levels. Specifically, TFF-concentrated viral metagenomes had significantly fewer abundant viral types (<italic>Podoviridae</italic> and <italic>Phycodnaviridae</italic>) and more variability among <italic>Myoviridae</italic> than FeCl<sub>3</sub>-precipitated viral metagenomes. More comprehensive analyses using protein clusters (66% of reads) and k-mers (100% of reads) showed 50–53% of these data were common to all four methods, and revealed trace bacterial DNA contamination in TFF-concentrated metagenomes and one of three replicates concentrated using FeCl<sub>3</sub> and purified by DNase alone. Shared k-mer analyses also revealed that polymerases used in amplification impact the resulting metagenomes, with TaKaRa enriching for ‘rare’ reads relative to PfuTurbo. 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<front><journal-meta><journal-id journal-id-type="nlm-ta">Environ Microbiol</journal-id><journal-id journal-id-type="iso-abbrev">Environ. Microbiol</journal-id><journal-id journal-id-type="publisher-id">emi</journal-id><journal-title-group><journal-title>Environmental Microbiology</journal-title></journal-title-group><issn pub-type="ppub">1462-2912</issn><issn pub-type="epub">1462-2920</issn><publisher><publisher-name>Blackwell Publishing Ltd</publisher-name><publisher-loc>Oxford, UK</publisher-loc></publisher></journal-meta><article-meta><article-id pub-id-type="pmid">22845467</article-id><article-id pub-id-type="pmc">3655615</article-id><article-id pub-id-type="doi">10.1111/j.1462-2920.2012.02836.x</article-id><article-categories><subj-group subj-group-type="heading"><subject>Research Articles</subject></subj-group></article-categories><title-group><article-title>Evaluation of methods to concentrate and purify ocean virus communities through comparative, replicated metagenomics</article-title></title-group><contrib-group><contrib contrib-type="author"><name><surname>Hurwitz</surname><given-names>Bonnie L</given-names></name></contrib><contrib contrib-type="author"><name><surname>Deng</surname><given-names>Li</given-names></name><xref ref-type="author-notes" rid="fn1">†</xref></contrib><contrib contrib-type="author"><name><surname>Poulos</surname><given-names>Bonnie T</given-names></name></contrib><contrib contrib-type="author"><name><surname>Sullivan</surname><given-names>Matthew B</given-names></name><xref ref-type="corresp" rid="cor1">*</xref></contrib><aff><institution>Ecology and Evolutionary Biology, University of Arizona</institution><addr-line>Tucson, AZ 85721, USA</addr-line></aff></contrib-group><author-notes><corresp id="cor1">*For correspondence. E-mail <email>mbsulli@email.arizona.edu</email>; Tel. (+1) 520 626 9100; Fax (+1) 520 621 9190.</corresp><fn id="fn1"><label>†</label><p>Current address: Helmholtz Zentrum München-German Research Center for Environmental Health, Institute of Groundwater Ecology, Neuherberg, Germany.</p></fn><fn><p>Re-use of this article is permitted in accordance with the Terms and Conditions set out at <ext-link ext-link-type="uri" xlink:href="http://wileyonlinelibrary.com/onlineopen#OnlineOpen_Terms">http://wileyonlinelibrary.com/onlineopen#OnlineOpen_Terms</ext-link></p></fn></author-notes><pub-date pub-type="ppub"><month>5</month><year>2013</year></pub-date><volume>15</volume><issue>5</issue><fpage>1428</fpage><lpage>1440</lpage><history><date date-type="received"><day>30</day><month>11</month><year>2011</year></date><date date-type="rev-recd"><day>29</day><month>6</month><year>2012</year></date><date date-type="accepted"><day>30</day><month>6</month><year>2012</year></date></history><permissions><copyright-statement>Copyright © 2013 Society for Applied Microbiology and Blackwell Publishing Ltd</copyright-statement><copyright-year>2013</copyright-year><license license-type="open-access" xlink:href="http://creativecommons.org/licenses/by/2.5/"><license-p>Re-use of this article is permitted in accordance with the Creative Commons Deed, Attribution 2.5, which does not permit commercial exploitation.</license-p></license></permissions><abstract><p>Viruses have global impact through mortality, nutrient cycling and horizontal gene transfer, yet their study is limited by complex methodologies with little validation. Here, we use triplicate metagenomes to compare common aquatic viral concentration and purification methods across four combinations as follows: (i) tangential flow filtration (TFF) and DNase + CsCl, (ii) FeCl<sub>3</sub> precipitation and DNase, (iii) FeCl<sub>3</sub> precipitation and DNase + CsCl and (iv) FeCl<sub>3</sub> precipitation and DNase + sucrose. Taxonomic data (30% of reads) suggested that purification methods were statistically indistinguishable at any taxonomic level while concentration methods were significantly different at family and genus levels. Specifically, TFF-concentrated viral metagenomes had significantly fewer abundant viral types (<italic>Podoviridae</italic> and <italic>Phycodnaviridae</italic>) and more variability among <italic>Myoviridae</italic> than FeCl<sub>3</sub>-precipitated viral metagenomes. More comprehensive analyses using protein clusters (66% of reads) and k-mers (100% of reads) showed 50–53% of these data were common to all four methods, and revealed trace bacterial DNA contamination in TFF-concentrated metagenomes and one of three replicates concentrated using FeCl<sub>3</sub> and purified by DNase alone. Shared k-mer analyses also revealed that polymerases used in amplification impact the resulting metagenomes, with TaKaRa enriching for ‘rare’ reads relative to PfuTurbo. Together these results provide empirical data for making experimental design decisions in culture-independent viral ecology studies.</p></abstract></article-meta></front></pmc></record>Pour manipuler ce document sous Unix (Dilib)
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