In silico re-assessment of a diagnostic RT-qPCR assay for universal detection of Influenza A viruses
Identifieur interne : 000F65 ( Ncbi/Merge ); précédent : 000F64; suivant : 000F66In silico re-assessment of a diagnostic RT-qPCR assay for universal detection of Influenza A viruses
Auteurs : Alexander Nagy [République tchèque] ; Tomáš Ji Inec [République tchèque] ; Helena Ji Incová [République tchèque] ; Lenka Erníková [République tchèque] ; Martina Havlí Ková [République tchèque]Source :
- Scientific Reports [ 2045-2322 ] ; 2019.
Abstract
The ongoing evolution of microbial pathogens represents a significant issue in diagnostic PCR/qPCR. Many assays are burdened with false negativity due to mispriming and/or probe-binding failures. Therefore, PCR/qPCR assays used in the laboratory should be periodically re-assessed
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DOI: 10.1038/s41598-018-37869-w
PubMed: 30733500
PubMed Central: 6367508
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tchèque</country><wicri:regionArea>State Veterinary Institute Prague, Prague</wicri:regionArea><placeName><settlement type="city">Prague</settlement><region type="région" nuts="2">Bohême centrale</region></placeName></affiliation><affiliation wicri:level="1"><nlm:aff id="Aff2"><institution-wrap><institution-id institution-id-type="ISNI">0000 0001 2184 1595</institution-id><institution-id institution-id-type="GRID">grid.425485.a</institution-id><institution>National Institute of Public Health, National Reference Laboratory for Influenza,</institution></institution-wrap>Prague, Czech Republic</nlm:aff><country xml:lang="fr">République tchèque</country><wicri:regionArea>Prague</wicri:regionArea></affiliation></author><author><name sortKey="Ji Inec, Tomas" sort="Ji Inec, Tomas" uniqKey="Ji Inec T" first="Tomáš" last="Ji Inec">Tomáš Ji Inec</name><affiliation wicri:level="1"><nlm:aff id="Aff2"><institution-wrap><institution-id institution-id-type="ISNI">0000 0001 2184 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tchèque</country><wicri:regionArea>Prague</wicri:regionArea></affiliation></author><author><name sortKey=" Ernikova, Lenka" sort=" Ernikova, Lenka" uniqKey=" Ernikova L" first="Lenka" last=" Erníková">Lenka Erníková</name><affiliation wicri:level="3"><nlm:aff id="Aff1">State Veterinary Institute Prague, Prague, Czech Republic</nlm:aff><country xml:lang="fr">République tchèque</country><wicri:regionArea>State Veterinary Institute Prague, Prague</wicri:regionArea><placeName><settlement type="city">Prague</settlement><region type="région" nuts="2">Bohême centrale</region></placeName></affiliation></author><author><name sortKey="Havli Kova, Martina" sort="Havli Kova, Martina" uniqKey="Havli Kova M" first="Martina" last="Havlí Ková">Martina Havlí Ková</name><affiliation wicri:level="1"><nlm:aff id="Aff2"><institution-wrap><institution-id institution-id-type="ISNI">0000 0001 2184 1595</institution-id><institution-id institution-id-type="GRID">grid.425485.a</institution-id><institution>National Institute of Public Health, National Reference Laboratory for Influenza,</institution></institution-wrap>Prague, Czech Republic</nlm:aff><country xml:lang="fr">République tchèque</country><wicri:regionArea>Prague</wicri:regionArea></affiliation></author></analytic><series><title level="j">Scientific Reports</title><idno type="eISSN">2045-2322</idno><imprint><date when="2019">2019</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p id="Par1">The ongoing evolution of microbial pathogens represents a significant issue in diagnostic PCR/qPCR. Many assays are burdened with false negativity due to mispriming and/or probe-binding failures. Therefore, PCR/qPCR assays used in the laboratory should be periodically re-assessed <italic>in silico</italic> on public sequences to evaluate the ability to detect actually circulating strains and to infer potentially escaping variants. In the work presented we re-assessed a RT-qPCR assay for the universal detection of influenza A (IA) viruses currently recommended by the European Union Reference Laboratory for Avian Influenza. To this end, the primers and probe sequences were challenged against more than 99,000 M-segment sequences in five data pools. To streamline this process, we developed a simple algorithm called the SequenceTracer designed for alignment stratification, compression, and personal sequence subset selection and also demonstrated its utility. The re-assessment confirmed the high inclusivity of the assay for the detection of avian, swine and human pandemic H1N1 IA viruses. On the other hand, the analysis identified human H3N2 strains with a critical probe-interfering mutation circulating since 2010, albeit with a significantly fluctuating proportion. Minor variations located in the forward and reverse primers identified in the avian and swine data were also considered.</p></div></front><back><div1 type="bibliography"><listBibl><biblStruct><analytic><author><name sortKey="Holland, Pm" uniqKey="Holland P">PM Holland</name></author><author><name sortKey="Abramson, Rd" uniqKey="Abramson R">RD Abramson</name></author><author><name sortKey="Watson, R" uniqKey="Watson R">R Watson</name></author><author><name sortKey="Gelfand, Dh" uniqKey="Gelfand D">DH Gelfand</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Lemmon, Gh" uniqKey="Lemmon G">GH Lemmon</name></author><author><name sortKey="Gardner, Sn" uniqKey="Gardner S">SN Gardner</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Stevenson, J" uniqKey="Stevenson J">J Stevenson</name></author><author><name sortKey="Hymas, W" uniqKey="Hymas W">W Hymas</name></author><author><name sortKey="Hillyard, D" uniqKey="Hillyard D">D 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uniqKey="Larsson A">A Larsson</name></author></analytic></biblStruct><biblStruct></biblStruct></listBibl></div1></back></TEI><pmc article-type="research-article"><pmc-dir>properties open_access</pmc-dir>
<front><journal-meta><journal-id journal-id-type="nlm-ta">Sci Rep</journal-id><journal-id journal-id-type="iso-abbrev">Sci Rep</journal-id><journal-title-group><journal-title>Scientific Reports</journal-title></journal-title-group><issn pub-type="epub">2045-2322</issn><publisher><publisher-name>Nature Publishing Group UK</publisher-name><publisher-loc>London</publisher-loc></publisher></journal-meta><article-meta><article-id pub-id-type="pmid">30733500</article-id><article-id pub-id-type="pmc">6367508</article-id><article-id pub-id-type="publisher-id">37869</article-id><article-id pub-id-type="doi">10.1038/s41598-018-37869-w</article-id><article-categories><subj-group subj-group-type="heading"><subject>Article</subject></subj-group></article-categories><title-group><article-title><italic>In silico</italic> re-assessment of a diagnostic RT-qPCR assay for universal detection of Influenza A viruses</article-title></title-group><contrib-group><contrib contrib-type="author" corresp="yes"><name><surname>Nagy</surname><given-names>Alexander</given-names></name><address><email>alexander.nagy@svupraha.cz</email></address><xref ref-type="aff" rid="Aff1">1</xref><xref ref-type="aff" rid="Aff2">2</xref></contrib><contrib contrib-type="author"><name><surname>Jiřinec</surname><given-names>Tomáš</given-names></name><xref ref-type="aff" rid="Aff2">2</xref></contrib><contrib contrib-type="author"><name><surname>Jiřincová</surname><given-names>Helena</given-names></name><xref ref-type="aff" rid="Aff2">2</xref></contrib><contrib contrib-type="author"><name><surname>Černíková</surname><given-names>Lenka</given-names></name><xref ref-type="aff" rid="Aff1">1</xref></contrib><contrib contrib-type="author"><name><surname>Havlíčková</surname><given-names>Martina</given-names></name><xref ref-type="aff" rid="Aff2">2</xref></contrib><aff id="Aff1"><label>1</label>State Veterinary Institute Prague, Prague, Czech Republic</aff><aff id="Aff2"><label>2</label><institution-wrap><institution-id institution-id-type="ISNI">0000 0001 2184 1595</institution-id><institution-id institution-id-type="GRID">grid.425485.a</institution-id><institution>National Institute of Public Health, National Reference Laboratory for Influenza,</institution></institution-wrap>Prague, Czech Republic</aff></contrib-group><pub-date pub-type="epub"><day>7</day><month>2</month><year>2019</year></pub-date><pub-date pub-type="pmc-release"><day>7</day><month>2</month><year>2019</year></pub-date><pub-date pub-type="collection"><year>2019</year></pub-date><volume>9</volume><elocation-id>1630</elocation-id><history><date date-type="received"><day>31</day><month>8</month><year>2017</year></date><date date-type="accepted"><day>10</day><month>12</month><year>2018</year></date></history><permissions><copyright-statement>© The Author(s) 2019</copyright-statement><license license-type="OpenAccess"><license-p><bold>Open Access</bold> This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit <ext-link ext-link-type="uri" xlink:href="http://creativecommons.org/licenses/by/4.0/">http://creativecommons.org/licenses/by/4.0/</ext-link>.</license-p></license></permissions><abstract id="Abs1"><p id="Par1">The ongoing evolution of microbial pathogens represents a significant issue in diagnostic PCR/qPCR. Many assays are burdened with false negativity due to mispriming and/or probe-binding failures. Therefore, PCR/qPCR assays used in the laboratory should be periodically re-assessed <italic>in silico</italic> on public sequences to evaluate the ability to detect actually circulating strains and to infer potentially escaping variants. In the work presented we re-assessed a RT-qPCR assay for the universal detection of influenza A (IA) viruses currently recommended by the European Union Reference Laboratory for Avian Influenza. To this end, the primers and probe sequences were challenged against more than 99,000 M-segment sequences in five data pools. To streamline this process, we developed a simple algorithm called the SequenceTracer designed for alignment stratification, compression, and personal sequence subset selection and also demonstrated its utility. The re-assessment confirmed the high inclusivity of the assay for the detection of avian, swine and human pandemic H1N1 IA viruses. On the other hand, the analysis identified human H3N2 strains with a critical probe-interfering mutation circulating since 2010, albeit with a significantly fluctuating proportion. Minor variations located in the forward and reverse primers identified in the avian and swine data were also considered.</p></abstract><custom-meta-group><custom-meta><meta-name>issue-copyright-statement</meta-name><meta-value>© The Author(s) 2019</meta-value></custom-meta></custom-meta-group></article-meta></front></pmc><affiliations><list><country><li>République tchèque</li></country><region><li>Bohême centrale</li></region><settlement><li>Prague</li></settlement></list><tree><country name="République tchèque"><region name="Bohême centrale"><name sortKey="Nagy, Alexander" sort="Nagy, Alexander" uniqKey="Nagy A" first="Alexander" last="Nagy">Alexander Nagy</name></region><name sortKey=" Ernikova, Lenka" sort=" Ernikova, Lenka" uniqKey=" Ernikova L" first="Lenka" last=" Erníková">Lenka Erníková</name><name sortKey="Havli Kova, Martina" sort="Havli Kova, Martina" uniqKey="Havli Kova M" first="Martina" last="Havlí Ková">Martina Havlí Ková</name><name sortKey="Ji Incova, Helena" sort="Ji Incova, Helena" uniqKey="Ji Incova H" first="Helena" last="Ji Incová">Helena Ji Incová</name><name sortKey="Ji Inec, Tomas" sort="Ji Inec, Tomas" uniqKey="Ji Inec T" first="Tomáš" last="Ji Inec">Tomáš Ji Inec</name><name sortKey="Nagy, Alexander" sort="Nagy, Alexander" uniqKey="Nagy A" first="Alexander" last="Nagy">Alexander Nagy</name></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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