Molecular Detection of a Novel Human Influenza (H1N1) of Pandemic Potential by Conventional and Real-Time Quantitative RT-PCR Assays
Identifieur interne : 001201 ( Main/Exploration ); précédent : 001200; suivant : 001202Molecular Detection of a Novel Human Influenza (H1N1) of Pandemic Potential by Conventional and Real-Time Quantitative RT-PCR Assays
Auteurs : Leo L. M. Poon [Hong Kong] ; K. H. Chan [Hong Kong] ; G. J. Smith [Hong Kong] ; C. S. W. Leung [Hong Kong] ; Y. Guan [Hong Kong] ; K. Y. Yuen [Hong Kong] ; J. S. M. Peiris [Hong Kong]Source :
- Clinical chemistry : (Baltimore, Md.) [ 0009-9147 ] ; 2009.
Descripteurs français
- KwdFr :
- ADN viral (analyse), ADN viral (génétique), ARN viral (analyse), ARN viral (génétique), ARN viral (isolement et purification), Animaux, Facteurs temps, Grippe humaine (diagnostic), Grippe humaine (virologie), Humains, Infections à Orthomyxoviridae (diagnostic), Infections à Orthomyxoviridae (virologie), RT-PCR (), RT-PCR (économie), Sensibilité et spécificité, Sous-type H1N1 du virus de la grippe A (), Sous-type H1N1 du virus de la grippe A (génétique), Sous-type H1N1 du virus de la grippe A (isolement et purification), Sous-type H2N2 du virus de la grippe A (), Sous-type H2N2 du virus de la grippe A (génétique), Sous-type H2N2 du virus de la grippe A (isolement et purification), Sous-type H3N2 du virus de la grippe A (), Sous-type H3N2 du virus de la grippe A (génétique), Sous-type H3N2 du virus de la grippe A (isolement et purification), Sous-type H5N1 du virus de la grippe A (), Sous-type H5N1 du virus de la grippe A (génétique), Sous-type H5N1 du virus de la grippe A (isolement et purification), Suidae (virologie), Séquence nucléotidique, Techniques de diagnostic moléculaire (), Techniques de diagnostic moléculaire (économie).
- MESH :
- analyse : ADN viral, ARN viral.
- diagnostic : Grippe humaine, Infections à Orthomyxoviridae.
- génétique : ADN viral, ARN viral, Sous-type H1N1 du virus de la grippe A, Sous-type H2N2 du virus de la grippe A, Sous-type H3N2 du virus de la grippe A, Sous-type H5N1 du virus de la grippe A.
- isolement et purification : ARN viral, Sous-type H1N1 du virus de la grippe A, Sous-type H2N2 du virus de la grippe A, Sous-type H3N2 du virus de la grippe A, Sous-type H5N1 du virus de la grippe A.
- virologie : Grippe humaine, Infections à Orthomyxoviridae, Suidae.
- économie : RT-PCR, Techniques de diagnostic moléculaire.
- Pascal (Inist)
- Animaux, Diagnostic, Détection, Facteurs temps, Homme, Grippe, Analyse quantitative, Dosage, Biochimie, Biologie clinique, Biologie moléculaire, Humains, Pandémie, RT-PCR, Réaction chaîne polymérase en temps réel, Sensibilité et spécificité, Sous-type H1N1 du virus de la grippe A, Sous-type H2N2 du virus de la grippe A, Sous-type H3N2 du virus de la grippe A, Sous-type H5N1 du virus de la grippe A, Séquence nucléotidique, Techniques de diagnostic moléculaire.
- Wicri :
- topic : Homme, Analyse quantitative, Biochimie.
English descriptors
- KwdEn :
- Animals, Assay, Base Sequence, Biochemistry, Clinical biology, DNA, Viral (analysis), DNA, Viral (genetics), Detection, Diagnosis, Human, Humans, Influenza, Influenza A Virus, H1N1 Subtype (classification), Influenza A Virus, H1N1 Subtype (genetics), Influenza A Virus, H1N1 Subtype (isolation & purification), Influenza A Virus, H2N2 Subtype (classification), Influenza A Virus, H2N2 Subtype (genetics), Influenza A Virus, H2N2 Subtype (isolation & purification), Influenza A Virus, H3N2 Subtype (classification), Influenza A Virus, H3N2 Subtype (genetics), Influenza A Virus, H3N2 Subtype (isolation & purification), Influenza A Virus, H5N1 Subtype (classification), Influenza A Virus, H5N1 Subtype (genetics), Influenza A Virus, H5N1 Subtype (isolation & purification), Influenza, Human (diagnosis), Influenza, Human (virology), Molecular Diagnostic Techniques (economics), Molecular Diagnostic Techniques (methods), Molecular biology, Orthomyxoviridae Infections (diagnosis), Orthomyxoviridae Infections (virology), Pandemic, Quantitative analysis, RNA, Viral (analysis), RNA, Viral (genetics), RNA, Viral (isolation & purification), Real time polymerase chain reaction, Reverse Transcriptase Polymerase Chain Reaction (economics), Reverse Transcriptase Polymerase Chain Reaction (methods), Sensitivity and Specificity, Swine (virology), Time Factors.
- MESH :
- chemical , analysis : DNA, Viral, RNA, Viral.
- chemical , genetics : DNA, Viral, RNA, Viral.
- classification : Influenza A Virus, H1N1 Subtype, Influenza A Virus, H2N2 Subtype, Influenza A Virus, H3N2 Subtype, Influenza A Virus, H5N1 Subtype.
- diagnosis : Influenza, Human, Orthomyxoviridae Infections.
- economics : Molecular Diagnostic Techniques, Reverse Transcriptase Polymerase Chain Reaction.
- genetics : Influenza A Virus, H1N1 Subtype, Influenza A Virus, H2N2 Subtype, Influenza A Virus, H3N2 Subtype, Influenza A Virus, H5N1 Subtype.
- isolation & purification : Influenza A Virus, H1N1 Subtype, Influenza A Virus, H2N2 Subtype, Influenza A Virus, H3N2 Subtype, Influenza A Virus, H5N1 Subtype, RNA, Viral.
- methods : Molecular Diagnostic Techniques, Reverse Transcriptase Polymerase Chain Reaction.
- virology : Influenza, Human, Orthomyxoviridae Infections, Swine.
- Animals, Base Sequence, Humans, Sensitivity and Specificity, Time Factors.
Abstract
BACKGROUND: Influenza A viruses are medically important viral pathogens that cause significant mortality and morbidity throughout the world. The recent emergence of a novel human influenza A virus (H1N1) poses a serious health threat. Molecular tests for rapid detection of this virus are urgently needed. METHODS: We developed a conventional 1-step RT-PCR assay and a 1-step quantitative real-time RT-PCR assay to detect the novel H1N1 virus, but not the seasonal H1N1 viruses. We also developed an additional real-time RT-PCR that can discriminate the novel H1N1 from other swine and human H1 subtype viruses. RESULTS: All of the assays had detection limits for the positive control in the range of 1.0 × 10-4 to 2.0 × 10-3 of the median tissue culture infective dose. Assay specificities were high, and for the conventional and real-time assays, all negative control samples were negative, including 7 human seasonal H1N1 viruses, 1 human H2N2 virus, 2 human seasonal H3N2 viruses, 1 human H5N1 virus, 7 avian influenza viruses (HA subtypes 4, 5, 7, 8, 9, and 10), and 48 nasopharyngeal aspirates (NPAs) from patients with noninfluenza respiratory diseases; for the assay that discriminates the novel H1N1 from other swine and human H1 subtype viruses, all negative controls were also negative, including 20 control NPAs, 2 seasonal human H1N1 viruses, 2 seasonal human H3N2 viruses, and 2 human H5N1 viruses. CONCLUSIONS: These assays appear useful for the rapid diagnosis of cases with the novel H1N1 virus, thereby allowing better pandemic preparedness.
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M." last="Poon">Leo L. M. Poon</name><affiliation wicri:level="1"><inist:fA14 i1="01"><s1>Department of Microbiology, The University of Hong Kong</s1><s3>HKG</s3><sZ>1 aut.</sZ><sZ>3 aut.</sZ><sZ>4 aut.</sZ><sZ>5 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ></inist:fA14><country>Hong Kong</country><wicri:noRegion>Department of Microbiology, The University of Hong Kong</wicri:noRegion></affiliation></author><author><name sortKey="Chan, K H" sort="Chan, K H" uniqKey="Chan K" first="K. H." last="Chan">K. H. Chan</name><affiliation wicri:level="1"><inist:fA14 i1="02"><s1>Department of Microbiology, Queen Mary Hospital</s1><s3>HKG</s3><sZ>2 aut.</sZ><sZ>6 aut.</sZ></inist:fA14><country>Hong Kong</country><wicri:noRegion>Department of Microbiology, Queen Mary Hospital</wicri:noRegion></affiliation></author><author><name sortKey="Smith, G J" sort="Smith, G J" uniqKey="Smith G" first="G. J." last="Smith">G. J. Smith</name><affiliation wicri:level="1"><inist:fA14 i1="01"><s1>Department of Microbiology, The University of Hong Kong</s1><s3>HKG</s3><sZ>1 aut.</sZ><sZ>3 aut.</sZ><sZ>4 aut.</sZ><sZ>5 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ></inist:fA14><country>Hong Kong</country><wicri:noRegion>Department of Microbiology, The University of Hong Kong</wicri:noRegion></affiliation></author><author><name sortKey="Leung, C S W" sort="Leung, C S W" uniqKey="Leung C" first="C. S. W." last="Leung">C. S. W. Leung</name><affiliation wicri:level="1"><inist:fA14 i1="01"><s1>Department of Microbiology, The University of Hong Kong</s1><s3>HKG</s3><sZ>1 aut.</sZ><sZ>3 aut.</sZ><sZ>4 aut.</sZ><sZ>5 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ></inist:fA14><country>Hong Kong</country><wicri:noRegion>Department of Microbiology, The University of Hong Kong</wicri:noRegion></affiliation></author><author><name sortKey="Guan, Y" sort="Guan, Y" uniqKey="Guan Y" first="Y." last="Guan">Y. Guan</name><affiliation wicri:level="1"><inist:fA14 i1="01"><s1>Department of Microbiology, The University of Hong Kong</s1><s3>HKG</s3><sZ>1 aut.</sZ><sZ>3 aut.</sZ><sZ>4 aut.</sZ><sZ>5 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ></inist:fA14><country>Hong Kong</country><wicri:noRegion>Department of Microbiology, The University of Hong Kong</wicri:noRegion></affiliation></author><author><name sortKey="Yuen, K Y" sort="Yuen, K Y" uniqKey="Yuen K" first="K. Y." last="Yuen">K. Y. Yuen</name><affiliation wicri:level="1"><inist:fA14 i1="01"><s1>Department of Microbiology, The University of Hong Kong</s1><s3>HKG</s3><sZ>1 aut.</sZ><sZ>3 aut.</sZ><sZ>4 aut.</sZ><sZ>5 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ></inist:fA14><country>Hong Kong</country><wicri:noRegion>Department of Microbiology, The University of Hong Kong</wicri:noRegion></affiliation><affiliation wicri:level="1"><inist:fA14 i1="02"><s1>Department of Microbiology, Queen Mary Hospital</s1><s3>HKG</s3><sZ>2 aut.</sZ><sZ>6 aut.</sZ></inist:fA14><country>Hong Kong</country><wicri:noRegion>Department of Microbiology, Queen Mary Hospital</wicri:noRegion></affiliation></author><author><name sortKey="Peiris, J S M" sort="Peiris, J S M" uniqKey="Peiris J" first="J. S. M." last="Peiris">J. S. M. Peiris</name><affiliation wicri:level="1"><inist:fA14 i1="01"><s1>Department of Microbiology, The University of Hong Kong</s1><s3>HKG</s3><sZ>1 aut.</sZ><sZ>3 aut.</sZ><sZ>4 aut.</sZ><sZ>5 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ></inist:fA14><country>Hong Kong</country><wicri:noRegion>Department of Microbiology, The University of Hong Kong</wicri:noRegion></affiliation><affiliation wicri:level="1"><inist:fA14 i1="03"><s1>HKU-Pasteur Research Centre</s1><s3>HKG</s3><sZ>7 aut.</sZ></inist:fA14><country>Hong Kong</country><wicri:noRegion>HKU-Pasteur Research Centre</wicri:noRegion></affiliation></author></analytic><series><title level="j" type="main">Clinical chemistry : (Baltimore, Md.)</title><title level="j" type="abbreviated">Clin. chem. : (Baltim. Md.)</title><idno type="ISSN">0009-9147</idno><imprint><date when="2009">2009</date></imprint></series></biblStruct></sourceDesc><seriesStmt><title level="j" type="main">Clinical chemistry : (Baltimore, Md.)</title><title level="j" type="abbreviated">Clin. chem. : (Baltim. Md.)</title><idno type="ISSN">0009-9147</idno></seriesStmt></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Animals</term><term>Assay</term><term>Base Sequence</term><term>Biochemistry</term><term>Clinical biology</term><term>DNA, Viral (analysis)</term><term>DNA, Viral (genetics)</term><term>Detection</term><term>Diagnosis</term><term>Human</term><term>Humans</term><term>Influenza</term><term>Influenza A Virus, H1N1 Subtype (classification)</term><term>Influenza A Virus, H1N1 Subtype (genetics)</term><term>Influenza A Virus, H1N1 Subtype (isolation & purification)</term><term>Influenza A Virus, H2N2 Subtype (classification)</term><term>Influenza A Virus, H2N2 Subtype (genetics)</term><term>Influenza A Virus, H2N2 Subtype (isolation & purification)</term><term>Influenza A Virus, H3N2 Subtype (classification)</term><term>Influenza A Virus, H3N2 Subtype (genetics)</term><term>Influenza A Virus, H3N2 Subtype (isolation & purification)</term><term>Influenza A Virus, H5N1 Subtype (classification)</term><term>Influenza A Virus, H5N1 Subtype (genetics)</term><term>Influenza A Virus, H5N1 Subtype (isolation & purification)</term><term>Influenza, Human (diagnosis)</term><term>Influenza, Human (virology)</term><term>Molecular Diagnostic Techniques (economics)</term><term>Molecular Diagnostic Techniques (methods)</term><term>Molecular biology</term><term>Orthomyxoviridae Infections (diagnosis)</term><term>Orthomyxoviridae Infections (virology)</term><term>Pandemic</term><term>Quantitative analysis</term><term>RNA, Viral (analysis)</term><term>RNA, Viral (genetics)</term><term>RNA, Viral (isolation & purification)</term><term>Real time polymerase chain reaction</term><term>Reverse Transcriptase Polymerase Chain Reaction (economics)</term><term>Reverse Transcriptase Polymerase Chain Reaction (methods)</term><term>Sensitivity and Specificity</term><term>Swine (virology)</term><term>Time Factors</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>ADN viral (analyse)</term><term>ADN viral (génétique)</term><term>ARN viral (analyse)</term><term>ARN viral (génétique)</term><term>ARN viral (isolement et purification)</term><term>Animaux</term><term>Facteurs temps</term><term>Grippe humaine (diagnostic)</term><term>Grippe humaine (virologie)</term><term>Humains</term><term>Infections à Orthomyxoviridae (diagnostic)</term><term>Infections à Orthomyxoviridae (virologie)</term><term>RT-PCR ()</term><term>RT-PCR (économie)</term><term>Sensibilité et spécificité</term><term>Sous-type H1N1 du virus de la grippe A ()</term><term>Sous-type H1N1 du virus de la grippe A (génétique)</term><term>Sous-type H1N1 du virus de la grippe A (isolement et purification)</term><term>Sous-type H2N2 du virus de la grippe A ()</term><term>Sous-type H2N2 du virus de la grippe A (génétique)</term><term>Sous-type H2N2 du virus de la grippe A (isolement et purification)</term><term>Sous-type H3N2 du virus de la grippe A ()</term><term>Sous-type H3N2 du virus de la grippe A (génétique)</term><term>Sous-type H3N2 du virus de la grippe A (isolement et purification)</term><term>Sous-type H5N1 du virus de la grippe A ()</term><term>Sous-type H5N1 du virus de la grippe A (génétique)</term><term>Sous-type H5N1 du virus de la grippe A (isolement et purification)</term><term>Suidae (virologie)</term><term>Séquence nucléotidique</term><term>Techniques de diagnostic moléculaire ()</term><term>Techniques de diagnostic moléculaire (économie)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="analysis" xml:lang="en"><term>DNA, Viral</term><term>RNA, Viral</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>DNA, Viral</term><term>RNA, Viral</term></keywords><keywords scheme="MESH" qualifier="analyse" xml:lang="fr"><term>ADN viral</term><term>ARN viral</term></keywords><keywords scheme="MESH" qualifier="classification" xml:lang="en"><term>Influenza A Virus, H1N1 Subtype</term><term>Influenza A Virus, H2N2 Subtype</term><term>Influenza A Virus, H3N2 Subtype</term><term>Influenza A Virus, H5N1 Subtype</term></keywords><keywords scheme="MESH" qualifier="diagnosis" xml:lang="en"><term>Influenza, Human</term><term>Orthomyxoviridae Infections</term></keywords><keywords scheme="MESH" qualifier="diagnostic" xml:lang="fr"><term>Grippe humaine</term><term>Infections à Orthomyxoviridae</term></keywords><keywords scheme="MESH" qualifier="economics" xml:lang="en"><term>Molecular Diagnostic Techniques</term><term>Reverse Transcriptase Polymerase Chain Reaction</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Influenza A Virus, H1N1 Subtype</term><term>Influenza A Virus, H2N2 Subtype</term><term>Influenza A Virus, H3N2 Subtype</term><term>Influenza A Virus, H5N1 Subtype</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>ADN viral</term><term>ARN viral</term><term>Sous-type H1N1 du virus de la grippe A</term><term>Sous-type H2N2 du virus de la grippe A</term><term>Sous-type H3N2 du virus de la grippe A</term><term>Sous-type H5N1 du virus de la grippe A</term></keywords><keywords scheme="MESH" qualifier="isolation & purification" xml:lang="en"><term>Influenza A Virus, H1N1 Subtype</term><term>Influenza A Virus, H2N2 Subtype</term><term>Influenza A Virus, H3N2 Subtype</term><term>Influenza A Virus, H5N1 Subtype</term><term>RNA, Viral</term></keywords><keywords scheme="MESH" qualifier="isolement et purification" xml:lang="fr"><term>ARN viral</term><term>Sous-type H1N1 du virus de la grippe A</term><term>Sous-type H2N2 du virus de la grippe A</term><term>Sous-type H3N2 du virus de la grippe A</term><term>Sous-type H5N1 du virus de la grippe A</term></keywords><keywords scheme="MESH" qualifier="methods" xml:lang="en"><term>Molecular Diagnostic Techniques</term><term>Reverse Transcriptase Polymerase Chain Reaction</term></keywords><keywords scheme="MESH" qualifier="virologie" xml:lang="fr"><term>Grippe humaine</term><term>Infections à Orthomyxoviridae</term><term>Suidae</term></keywords><keywords scheme="MESH" qualifier="virology" xml:lang="en"><term>Influenza, Human</term><term>Orthomyxoviridae Infections</term><term>Swine</term></keywords><keywords scheme="MESH" qualifier="économie" xml:lang="fr"><term>RT-PCR</term><term>Techniques de diagnostic moléculaire</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Animals</term><term>Base Sequence</term><term>Humans</term><term>Sensitivity and Specificity</term><term>Time Factors</term></keywords><keywords scheme="Pascal" xml:lang="fr"><term>Animaux</term><term>Diagnostic</term><term>Détection</term><term>Facteurs temps</term><term>Homme</term><term>Grippe</term><term>Analyse quantitative</term><term>Dosage</term><term>Biochimie</term><term>Biologie clinique</term><term>Biologie moléculaire</term><term>Humains</term><term>Pandémie</term><term>RT-PCR</term><term>Réaction chaîne polymérase en temps réel</term><term>Sensibilité et spécificité</term><term>Sous-type H1N1 du virus de la grippe A</term><term>Sous-type H2N2 du virus de la grippe A</term><term>Sous-type H3N2 du virus de la grippe A</term><term>Sous-type H5N1 du virus de la grippe A</term><term>Séquence nucléotidique</term><term>Techniques de diagnostic moléculaire</term></keywords><keywords scheme="Wicri" type="topic" xml:lang="fr"><term>Homme</term><term>Analyse quantitative</term><term>Biochimie</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">BACKGROUND: Influenza A viruses are medically important viral pathogens that cause significant mortality and morbidity throughout the world. The recent emergence of a novel human influenza A virus (H1N1) poses a serious health threat. Molecular tests for rapid detection of this virus are urgently needed. METHODS: We developed a conventional 1-step RT-PCR assay and a 1-step quantitative real-time RT-PCR assay to detect the novel H1N1 virus, but not the seasonal H1N1 viruses. We also developed an additional real-time RT-PCR that can discriminate the novel H1N1 from other swine and human H1 subtype viruses. RESULTS: All of the assays had detection limits for the positive control in the range of 1.0 × 10<sup>-4</sup> to 2.0 × 10<sup>-3</sup> of the median tissue culture infective dose. Assay specificities were high, and for the conventional and real-time assays, all negative control samples were negative, including 7 human seasonal H1N1 viruses, 1 human H2N2 virus, 2 human seasonal H3N2 viruses, 1 human H5N1 virus, 7 avian influenza viruses (HA subtypes 4, 5, 7, 8, 9, and 10), and 48 nasopharyngeal aspirates (NPAs) from patients with noninfluenza respiratory diseases; for the assay that discriminates the novel H1N1 from other swine and human H1 subtype viruses, all negative controls were also negative, including 20 control NPAs, 2 seasonal human H1N1 viruses, 2 seasonal human H3N2 viruses, and 2 human H5N1 viruses. CONCLUSIONS: These assays appear useful for the rapid diagnosis of cases with the novel H1N1 virus, thereby allowing better pandemic preparedness.</div></front></TEI><affiliations><list><country><li>Hong Kong</li></country></list><tree><country name="Hong Kong"><noRegion><name sortKey="Poon, Leo L M" sort="Poon, Leo L M" uniqKey="Poon L" first="Leo L. M." last="Poon">Leo L. M. Poon</name></noRegion><name sortKey="Chan, K H" sort="Chan, K H" uniqKey="Chan K" first="K. H." last="Chan">K. H. Chan</name><name sortKey="Guan, Y" sort="Guan, Y" uniqKey="Guan Y" first="Y." last="Guan">Y. Guan</name><name sortKey="Leung, C S W" sort="Leung, C S W" uniqKey="Leung C" first="C. S. W." last="Leung">C. S. W. Leung</name><name sortKey="Peiris, J S M" sort="Peiris, J S M" uniqKey="Peiris J" first="J. S. M." last="Peiris">J. S. M. Peiris</name><name sortKey="Peiris, J S M" sort="Peiris, J S M" uniqKey="Peiris J" first="J. S. M." last="Peiris">J. S. M. Peiris</name><name sortKey="Smith, G J" sort="Smith, G J" uniqKey="Smith G" first="G. J." last="Smith">G. J. Smith</name><name sortKey="Yuen, K Y" sort="Yuen, K Y" uniqKey="Yuen K" first="K. Y." last="Yuen">K. Y. Yuen</name><name sortKey="Yuen, K Y" sort="Yuen, K Y" uniqKey="Yuen K" first="K. Y." last="Yuen">K. Y. 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