Reverse Transcription Recombinase Polymerase Amplification Assay for the Detection of Middle East Respiratory Syndrome Coronavirus
Identifieur interne : 001183 ( Pmc/Checkpoint ); précédent : 001182; suivant : 001184Reverse Transcription Recombinase Polymerase Amplification Assay for the Detection of Middle East Respiratory Syndrome Coronavirus
Auteurs : Ahmed Abd El Wahed ; Pranav Patel ; Doris Heidenreich ; Frank T. Hufert ; Manfred WeidmannSource :
- PLoS Currents [ 2157-3999 ] ; 2013.
Abstract
The emergence of Middle East Respiratory Syndrome Coronavirus (MERS-CoV) in the eastern Mediterranean and imported cases to Europe has alerted public health authorities. Currently, detection of MERS-CoV in patient samples is done by real-time RT-PCR. Samples collected from suspected cases are sent to highly-equipped centralized laboratories for screening. A rapid point-of-care test is needed to allow more widespread mobile detection of the virus directly from patient material. In this study, we describe the development of a reverse transcription isothermal Recombinase Polymerase Amplification (RT-RPA) assay for the identification of MERS-CoV. A partial nucleocapsid gene RNA molecular standard of MERS-coronavirus was used to determine the assay sensitivity. The isothermal (42°C) MERS-CoV RT-RPA was as sensitive as real-time RT-PCR (10 RNA molecules), rapid (3-7 minutes) and mobile (using tubescanner weighing 1kg). The MERS-CoV RT-RPA showed cross-detection neither of any of the RNAs of several coronaviruses and respiratory viruses affecting humans nor of the human genome. The developed isothermal real-time RT-RPA is ideal for rapid mobile molecular MERS-CoV monitoring in acute patients and may also facilitate the search for the animal reservoir of MERS-CoV.
Url:
DOI: 10.1371/currents.outbreaks.62df1c7c75ffc96cd59034531e2e8364
PubMed: 24459611
PubMed Central: 3871419
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<front><journal-meta><journal-id journal-id-type="nlm-ta">PLoS Curr</journal-id><journal-id journal-id-type="iso-abbrev">PLoS Curr</journal-id><journal-id journal-id-type="publisher-id">PLoS Curr</journal-id><journal-id journal-id-type="publisher-id">PLoS Curr</journal-id><journal-id journal-id-type="publisher-id">plos</journal-id><journal-title-group><journal-title>PLoS Currents</journal-title></journal-title-group><issn pub-type="epub">2157-3999</issn><publisher><publisher-name>Public Library of Science</publisher-name><publisher-loc>San Francisco, USA</publisher-loc></publisher></journal-meta><article-meta><article-id pub-id-type="pmid">24459611</article-id><article-id pub-id-type="pmc">3871419</article-id><article-id pub-id-type="doi">10.1371/currents.outbreaks.62df1c7c75ffc96cd59034531e2e8364</article-id><article-categories><subj-group subj-group-type="heading"><subject>Research</subject></subj-group></article-categories><title-group><article-title>Reverse Transcription Recombinase Polymerase Amplification Assay for the Detection of Middle East Respiratory Syndrome Coronavirus</article-title></title-group><contrib-group><contrib contrib-type="author"><name><surname>Abd El Wahed</surname><given-names>Ahmed</given-names></name><aff>Department of Virology, University Medical Centre, Goettingen, Germany; Department of Virology, Faculty of Veterinary Medicine, Mansoura University, Mansoura, Egypt</aff><ext-link ext-link-type="uri" xlink:href="http://www.virologie.uni-goettingen.de">http://www.virologie.uni-goettingen.de</ext-link><xref ref-type="bio" rid="d35e36">*</xref></contrib><contrib contrib-type="author"><name><surname>Patel</surname><given-names>Pranav</given-names></name><aff>Centre for Biological Threats and Special Pathogens, Robert Koch Institute, Berlin, Germany</aff></contrib><contrib contrib-type="author"><name><surname>Heidenreich</surname><given-names>Doris</given-names></name><aff>Department of Virology, University Medical Centre, Goettingen, Germany</aff></contrib><contrib contrib-type="author"><name><surname>Hufert</surname><given-names>Frank T.</given-names></name><aff>Department of Virology, University Medical Centre, Goettingen, Germany</aff></contrib><contrib contrib-type="author"><name><surname>Weidmann</surname><given-names>Manfred</given-names></name><aff>Department of Virology, University Medical Centre, Goettingen, Germany</aff></contrib></contrib-group><pub-date pub-type="epub"><day>12</day><month>12</month><year>2013</year></pub-date><volume>5</volume><elocation-id>ecurrents.outbreaks.62df1c7c75ffc96cd59034531e2e8364</elocation-id><permissions><license xlink:href="http://creativecommons.org/licenses/by/4.0/"><license-p>This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.</license-p></license></permissions><abstract><p>The emergence of Middle East Respiratory Syndrome Coronavirus (MERS-CoV) in the eastern Mediterranean and imported cases to Europe has alerted public health authorities. Currently, detection of MERS-CoV in patient samples is done by real-time RT-PCR. Samples collected from suspected cases are sent to highly-equipped centralized laboratories for screening. A rapid point-of-care test is needed to allow more widespread mobile detection of the virus directly from patient material. In this study, we describe the development of a reverse transcription isothermal Recombinase Polymerase Amplification (RT-RPA) assay for the identification of MERS-CoV. A partial nucleocapsid gene RNA molecular standard of MERS-coronavirus was used to determine the assay sensitivity. The isothermal (42°C) MERS-CoV RT-RPA was as sensitive as real-time RT-PCR (10 RNA molecules), rapid (3-7 minutes) and mobile (using tubescanner weighing 1kg). The MERS-CoV RT-RPA showed cross-detection neither of any of the RNAs of several coronaviruses and respiratory viruses affecting humans nor of the human genome. The developed isothermal real-time RT-RPA is ideal for rapid mobile molecular MERS-CoV monitoring in acute patients and may also facilitate the search for the animal reservoir of MERS-CoV. </p></abstract><kwd-group kwd-group-type="simple"><kwd>MERS-coronavirus</kwd><kwd>point-of-care assay</kwd><kwd>RPA</kwd></kwd-group><funding-group><funding-statement>The study was funded by the Federal Ministry of Education and Research (BMBF) (project name: RESCheck, ID: FKN:16SV5436). </funding-statement></funding-group></article-meta></front></pmc><affiliations><list></list><tree><noCountry><name sortKey="Abd El Wahed, Ahmed" sort="Abd El Wahed, Ahmed" uniqKey="Abd El Wahed A" first="Ahmed" last="Abd El Wahed">Ahmed Abd El Wahed</name><name sortKey="Heidenreich, Doris" sort="Heidenreich, Doris" uniqKey="Heidenreich D" first="Doris" last="Heidenreich">Doris Heidenreich</name><name sortKey="Hufert, Frank T" sort="Hufert, Frank T" uniqKey="Hufert F" first="Frank T." last="Hufert">Frank T. Hufert</name><name sortKey="Patel, Pranav" sort="Patel, Pranav" uniqKey="Patel P" first="Pranav" last="Patel">Pranav Patel</name><name sortKey="Weidmann, Manfred" sort="Weidmann, Manfred" uniqKey="Weidmann M" first="Manfred" last="Weidmann">Manfred Weidmann</name></noCountry></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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