Real-Time Sequence-Validated Loop-Mediated Isothermal Amplification Assays for Detection of Middle East Respiratory Syndrome Coronavirus (MERS-CoV)
Identifieur interne : 001087 ( Ncbi/Merge ); précédent : 001086; suivant : 001088Real-Time Sequence-Validated Loop-Mediated Isothermal Amplification Assays for Detection of Middle East Respiratory Syndrome Coronavirus (MERS-CoV)
Auteurs : Sanchita Bhadra [États-Unis] ; Yu Sherry Jiang [États-Unis] ; Mia R. Kumar [États-Unis] ; Reed F. Johnson [États-Unis] ; Lisa E. Hensley [États-Unis] ; Andrew D. Ellington [États-Unis]Source :
- PLoS ONE [ 1932-6203 ] ; 2015.
Descripteurs français
- KwdFr :
- Coronavirus du syndrome respiratoire du Moyen-Orient (génétique), Coronavirus du syndrome respiratoire du Moyen-Orient (isolement et purification), Coronavirus du syndrome respiratoire du Moyen-Orient (pathogénicité), Génome viral, Humains, Infections à coronavirus (diagnostic), Infections à coronavirus (génétique), Infections à coronavirus (virologie), Protéines virales (génétique), Réaction de polymérisation en chaine en temps réel ().
- MESH :
- diagnostic : Infections à coronavirus.
- génétique : Coronavirus du syndrome respiratoire du Moyen-Orient, Infections à coronavirus, Protéines virales.
- isolement et purification : Coronavirus du syndrome respiratoire du Moyen-Orient.
- pathogénicité : Coronavirus du syndrome respiratoire du Moyen-Orient.
- virologie : Infections à coronavirus.
- Génome viral, Humains, Réaction de polymérisation en chaine en temps réel.
English descriptors
- KwdEn :
- Coronavirus Infections (diagnosis), Coronavirus Infections (genetics), Coronavirus Infections (virology), Genome, Viral, Humans, Middle East Respiratory Syndrome Coronavirus (genetics), Middle East Respiratory Syndrome Coronavirus (isolation & purification), Middle East Respiratory Syndrome Coronavirus (pathogenicity), Real-Time Polymerase Chain Reaction (methods), Viral Proteins (genetics).
- MESH :
- chemical , genetics : Viral Proteins.
- diagnosis : Coronavirus Infections.
- genetics : Coronavirus Infections, Middle East Respiratory Syndrome Coronavirus.
- isolation & purification : Middle East Respiratory Syndrome Coronavirus.
- methods : Real-Time Polymerase Chain Reaction.
- pathogenicity : Middle East Respiratory Syndrome Coronavirus.
- virology : Coronavirus Infections.
- Genome, Viral, Humans.
Abstract
The Middle East respiratory syndrome coronavirus (MERS-CoV), an emerging human coronavirus, causes severe acute respiratory illness with a 35% mortality rate. In light of the recent surge in reported infections we have developed asymmetric five-primer reverse transcription loop-mediated isothermal amplification (RT-LAMP) assays for detection of MERS-CoV. Isothermal amplification assays will facilitate the development of portable point-of-care diagnostics that are crucial for management of emerging infections. The RT-LAMP assays are designed to amplify MERS-CoV genomic loci located within the open reading frame (ORF)1a and ORF1b genes and upstream of the E gene. Additionally we applied one-step strand displacement probes (OSD) for real-time sequence-specific verification of LAMP amplicons. Asymmetric amplification effected by incorporating a single loop primer in each assay accelerated the time-to-result of the OSD-RT-LAMP assays. The resulting assays could detect 0.02 to 0.2 plaque forming units (PFU) (5 to 50 PFU/ml) of MERS-CoV in infected cell culture supernatants within 30 to 50 min and did not cross-react with common human respiratory pathogens.
Url:
DOI: 10.1371/journal.pone.0123126
PubMed: 25856093
PubMed Central: 4391951
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PMC:4391951Le document en format XML
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<front><div type="abstract" xml:lang="en"><p>The Middle East respiratory syndrome coronavirus (MERS-CoV), an emerging human coronavirus, causes severe acute respiratory illness with a 35% mortality rate. In light of the recent surge in reported infections we have developed asymmetric five-primer reverse transcription loop-mediated isothermal amplification (RT-LAMP) assays for detection of MERS-CoV. Isothermal amplification assays will facilitate the development of portable point-of-care diagnostics that are crucial for management of emerging infections. The RT-LAMP assays are designed to amplify MERS-CoV genomic loci located within the open reading frame (ORF)1a and ORF1b genes and upstream of the E gene. Additionally we applied one-step strand displacement probes (OSD) for real-time sequence-specific verification of LAMP amplicons. Asymmetric amplification effected by incorporating a single loop primer in each assay accelerated the time-to-result of the OSD-RT-LAMP assays. The resulting assays could detect 0.02 to 0.2 plaque forming units (PFU) (5 to 50 PFU/ml) of MERS-CoV in infected cell culture supernatants within 30 to 50 min and did not cross-react with common human respiratory pathogens.</p>
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<author><name sortKey="Bhadra, Sanchita" sort="Bhadra, Sanchita" uniqKey="Bhadra S" first="Sanchita" last="Bhadra">Sanchita Bhadra</name>
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</nlm:aff>
<country xml:lang="fr">États-Unis</country>
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<placeName><region type="state">Texas</region>
<settlement type="city">Austin (Texas)</settlement>
</placeName>
<orgName type="university">Université du Texas à Austin</orgName>
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</author>
<author><name sortKey="Jiang, Yu Sherry" sort="Jiang, Yu Sherry" uniqKey="Jiang Y" first="Yu Sherry" last="Jiang">Yu Sherry Jiang</name>
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</nlm:aff>
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<settlement type="city">Austin (Texas)</settlement>
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<author><name sortKey="Kumar, Mia R" sort="Kumar, Mia R" uniqKey="Kumar M" first="Mia R." last="Kumar">Mia R. Kumar</name>
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</nlm:aff>
<country xml:lang="fr">États-Unis</country>
<wicri:regionArea>Emerging Viral Pathogens Section, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Maryland</wicri:regionArea>
<placeName><region type="state">Maryland</region>
</placeName>
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</author>
<author><name sortKey="Johnson, Reed F" sort="Johnson, Reed F" uniqKey="Johnson R" first="Reed F." last="Johnson">Reed F. Johnson</name>
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</nlm:aff>
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<placeName><region type="state">Maryland</region>
</placeName>
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</author>
<author><name sortKey="Hensley, Lisa E" sort="Hensley, Lisa E" uniqKey="Hensley L" first="Lisa E." last="Hensley">Lisa E. Hensley</name>
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</nlm:aff>
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<wicri:regionArea>Integrated Research Facility, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Fort Detrick, Maryland</wicri:regionArea>
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</placeName>
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</author>
<author><name sortKey="Ellington, Andrew D" sort="Ellington, Andrew D" uniqKey="Ellington A" first="Andrew D." last="Ellington">Andrew D. Ellington</name>
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</nlm:aff>
<country xml:lang="fr">États-Unis</country>
<wicri:regionArea>Institute for Cellular and Molecular Biology, Center for Systems and Synthetic Biology, Department of Chemistry and Biochemistry, University of Texas at Austin, Austin, Texas</wicri:regionArea>
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<series><title level="j">PLoS ONE</title>
<idno type="eISSN">1932-6203</idno>
<imprint><date when="2015">2015</date>
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<front><div type="abstract" xml:lang="en"><p>The Middle East respiratory syndrome coronavirus (MERS-CoV), an emerging human coronavirus, causes severe acute respiratory illness with a 35% mortality rate. In light of the recent surge in reported infections we have developed asymmetric five-primer reverse transcription loop-mediated isothermal amplification (RT-LAMP) assays for detection of MERS-CoV. Isothermal amplification assays will facilitate the development of portable point-of-care diagnostics that are crucial for management of emerging infections. The RT-LAMP assays are designed to amplify MERS-CoV genomic loci located within the open reading frame (ORF)1a and ORF1b genes and upstream of the E gene. Additionally we applied one-step strand displacement probes (OSD) for real-time sequence-specific verification of LAMP amplicons. Asymmetric amplification effected by incorporating a single loop primer in each assay accelerated the time-to-result of the OSD-RT-LAMP assays. The resulting assays could detect 0.02 to 0.2 plaque forming units (PFU) (5 to 50 PFU/ml) of MERS-CoV in infected cell culture supernatants within 30 to 50 min and did not cross-react with common human respiratory pathogens.</p>
</div>
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<sourceDesc><biblStruct><analytic><title xml:lang="en">Real-time sequence-validated loop-mediated isothermal amplification assays for detection of Middle East respiratory syndrome coronavirus (MERS-CoV).</title>
<author><name sortKey="Bhadra, Sanchita" sort="Bhadra, Sanchita" uniqKey="Bhadra S" first="Sanchita" last="Bhadra">Sanchita Bhadra</name>
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<country xml:lang="fr">États-Unis</country>
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<author><name sortKey="Jiang, Yu Sherry" sort="Jiang, Yu Sherry" uniqKey="Jiang Y" first="Yu Sherry" last="Jiang">Yu Sherry Jiang</name>
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<author><name sortKey="Kumar, Mia R" sort="Kumar, Mia R" uniqKey="Kumar M" first="Mia R" last="Kumar">Mia R. Kumar</name>
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<author><name sortKey="Johnson, Reed F" sort="Johnson, Reed F" uniqKey="Johnson R" first="Reed F" last="Johnson">Reed F. Johnson</name>
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<author><name sortKey="Hensley, Lisa E" sort="Hensley, Lisa E" uniqKey="Hensley L" first="Lisa E" last="Hensley">Lisa E. Hensley</name>
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<author><name sortKey="Ellington, Andrew D" sort="Ellington, Andrew D" uniqKey="Ellington A" first="Andrew D" last="Ellington">Andrew D. Ellington</name>
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<country xml:lang="fr">États-Unis</country>
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</placeName>
<orgName type="university">Université du Texas à Austin</orgName>
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<series><title level="j">PloS one</title>
<idno type="eISSN">1932-6203</idno>
<imprint><date when="2015" type="published">2015</date>
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<profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Coronavirus Infections (diagnosis)</term>
<term>Coronavirus Infections (genetics)</term>
<term>Coronavirus Infections (virology)</term>
<term>Genome, Viral</term>
<term>Humans</term>
<term>Middle East Respiratory Syndrome Coronavirus (genetics)</term>
<term>Middle East Respiratory Syndrome Coronavirus (isolation & purification)</term>
<term>Middle East Respiratory Syndrome Coronavirus (pathogenicity)</term>
<term>Real-Time Polymerase Chain Reaction (methods)</term>
<term>Viral Proteins (genetics)</term>
</keywords>
<keywords scheme="KwdFr" xml:lang="fr"><term>Coronavirus du syndrome respiratoire du Moyen-Orient (génétique)</term>
<term>Coronavirus du syndrome respiratoire du Moyen-Orient (isolement et purification)</term>
<term>Coronavirus du syndrome respiratoire du Moyen-Orient (pathogénicité)</term>
<term>Génome viral</term>
<term>Humains</term>
<term>Infections à coronavirus (diagnostic)</term>
<term>Infections à coronavirus (génétique)</term>
<term>Infections à coronavirus (virologie)</term>
<term>Protéines virales (génétique)</term>
<term>Réaction de polymérisation en chaine en temps réel ()</term>
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<keywords scheme="MESH" qualifier="diagnosis" xml:lang="en"><term>Coronavirus Infections</term>
</keywords>
<keywords scheme="MESH" qualifier="diagnostic" xml:lang="fr"><term>Infections à coronavirus</term>
</keywords>
<keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Coronavirus Infections</term>
<term>Middle East Respiratory Syndrome Coronavirus</term>
</keywords>
<keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>Coronavirus du syndrome respiratoire du Moyen-Orient</term>
<term>Infections à coronavirus</term>
<term>Protéines virales</term>
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<keywords scheme="MESH" qualifier="isolation & purification" xml:lang="en"><term>Middle East Respiratory Syndrome Coronavirus</term>
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<keywords scheme="MESH" qualifier="isolement et purification" xml:lang="fr"><term>Coronavirus du syndrome respiratoire du Moyen-Orient</term>
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<keywords scheme="MESH" qualifier="methods" xml:lang="en"><term>Real-Time Polymerase Chain Reaction</term>
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<keywords scheme="MESH" qualifier="pathogenicity" xml:lang="en"><term>Middle East Respiratory Syndrome Coronavirus</term>
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</keywords>
<keywords scheme="MESH" qualifier="virology" xml:lang="en"><term>Coronavirus Infections</term>
</keywords>
<keywords scheme="MESH" xml:lang="en"><term>Genome, Viral</term>
<term>Humans</term>
</keywords>
<keywords scheme="MESH" xml:lang="fr"><term>Génome viral</term>
<term>Humains</term>
<term>Réaction de polymérisation en chaine en temps réel</term>
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<front><div type="abstract" xml:lang="en">The Middle East respiratory syndrome coronavirus (MERS-CoV), an emerging human coronavirus, causes severe acute respiratory illness with a 35% mortality rate. In light of the recent surge in reported infections we have developed asymmetric five-primer reverse transcription loop-mediated isothermal amplification (RT-LAMP) assays for detection of MERS-CoV. Isothermal amplification assays will facilitate the development of portable point-of-care diagnostics that are crucial for management of emerging infections. The RT-LAMP assays are designed to amplify MERS-CoV genomic loci located within the open reading frame (ORF)1a and ORF1b genes and upstream of the E gene. Additionally we applied one-step strand displacement probes (OSD) for real-time sequence-specific verification of LAMP amplicons. Asymmetric amplification effected by incorporating a single loop primer in each assay accelerated the time-to-result of the OSD-RT-LAMP assays. The resulting assays could detect 0.02 to 0.2 plaque forming units (PFU) (5 to 50 PFU/ml) of MERS-CoV in infected cell culture supernatants within 30 to 50 min and did not cross-react with common human respiratory pathogens. </div>
</front>
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