Molecular Detection of a Novel Human Influenza (H1N1) of Pandemic Potential by Conventional and Real-Time Quantitative RT-PCR Assays
Identifieur interne : 000025 ( PascalFrancis/Checkpoint ); précédent : 000024; suivant : 000026Molecular 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
- Pascal (Inist)
- Wicri :
- topic : Homme, Analyse quantitative, Biochimie.
English descriptors
- KwdEn :
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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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></titleStmt><publicationStmt><idno type="wicri:source">INIST</idno><idno type="inist">09-0358359</idno><date when="2009">2009</date><idno type="stanalyst">PASCAL 09-0358359 INIST</idno><idno type="RBID">Pascal:09-0358359</idno><idno type="wicri:Area/PascalFrancis/Corpus">000025</idno><idno type="wicri:Area/PascalFrancis/Curation">000049</idno><idno type="wicri:Area/PascalFrancis/Checkpoint">000025</idno><idno type="wicri:explorRef" wicri:stream="PascalFrancis" wicri:step="Checkpoint">000025</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en" level="a">Molecular Detection of a Novel Human Influenza (H1N1) of Pandemic Potential by Conventional and Real-Time Quantitative RT-PCR Assays</title><author><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><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>Assay</term><term>Biochemistry</term><term>Clinical biology</term><term>Detection</term><term>Diagnosis</term><term>Human</term><term>Influenza</term><term>Molecular biology</term><term>Pandemic</term><term>Quantitative analysis</term><term>Real time polymerase chain reaction</term></keywords><keywords scheme="Pascal" xml:lang="fr"><term>Diagnostic</term><term>Détection</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>Pandémie</term><term>Réaction chaîne polymérase en temps réel</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><inist><standard h6="B"><pA><fA01 i1="01" i2="1"><s0>0009-9147</s0></fA01><fA02 i1="01"><s0>CLCHAU</s0></fA02><fA03 i2="1"><s0>Clin. chem. : (Baltim. Md.)</s0></fA03><fA05><s2>55</s2></fA05><fA06><s2>8</s2></fA06><fA08 i1="01" i2="1" l="ENG"><s1>Molecular Detection of a Novel Human Influenza (H1N1) of Pandemic Potential by Conventional and Real-Time Quantitative RT-PCR Assays</s1></fA08><fA11 i1="01" i2="1"><s1>POON (Leo L. M.)</s1></fA11><fA11 i1="02" i2="1"><s1>CHAN (K. H.)</s1></fA11><fA11 i1="03" i2="1"><s1>SMITH (G. J.)</s1></fA11><fA11 i1="04" i2="1"><s1>LEUNG (C. S. W.)</s1></fA11><fA11 i1="05" i2="1"><s1>GUAN (Y.)</s1></fA11><fA11 i1="06" i2="1"><s1>YUEN (K. Y.)</s1></fA11><fA11 i1="07" i2="1"><s1>PEIRIS (J. S. M.)</s1></fA11><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></fA14><fA14 i1="02"><s1>Department of Microbiology, Queen Mary Hospital</s1><s3>HKG</s3><sZ>2 aut.</sZ><sZ>6 aut.</sZ></fA14><fA14 i1="03"><s1>HKU-Pasteur Research Centre</s1><s3>HKG</s3><sZ>7 aut.</sZ></fA14><fA20><s1>1555-1558</s1></fA20><fA21><s1>2009</s1></fA21><fA23 i1="01"><s0>ENG</s0></fA23><fA43 i1="01"><s1>INIST</s1><s2>7603</s2><s5>354000170938820160</s5></fA43><fA44><s0>0000</s0><s1>© 2009 INIST-CNRS. All rights reserved.</s1></fA44><fA45><s0>11 ref.</s0></fA45><fA47 i1="01" i2="1"><s0>09-0358359</s0></fA47><fA60><s1>P</s1><s3>CC</s3></fA60><fA61><s0>A</s0></fA61><fA64 i1="01" i2="1"><s0>Clinical chemistry : (Baltimore, Md.)</s0></fA64><fA66 i1="01"><s0>USA</s0></fA66><fC01 i1="01" l="ENG"><s0>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.</s0></fC01><fC02 i1="01" i2="X"><s0>002B24</s0></fC02><fC02 i1="02" i2="X"><s0>002A02</s0></fC02><fC02 i1="03" i2="X"><s0>002A03</s0></fC02><fC03 i1="01" i2="X" l="FRE"><s0>Diagnostic</s0><s5>02</s5></fC03><fC03 i1="01" i2="X" l="ENG"><s0>Diagnosis</s0><s5>02</s5></fC03><fC03 i1="01" i2="X" l="SPA"><s0>Diagnóstico</s0><s5>02</s5></fC03><fC03 i1="02" i2="X" l="FRE"><s0>Détection</s0><s5>03</s5></fC03><fC03 i1="02" i2="X" l="ENG"><s0>Detection</s0><s5>03</s5></fC03><fC03 i1="02" i2="X" l="SPA"><s0>Detección</s0><s5>03</s5></fC03><fC03 i1="03" i2="X" l="FRE"><s0>Homme</s0><s5>05</s5></fC03><fC03 i1="03" i2="X" l="ENG"><s0>Human</s0><s5>05</s5></fC03><fC03 i1="03" i2="X" l="SPA"><s0>Hombre</s0><s5>05</s5></fC03><fC03 i1="04" i2="X" l="FRE"><s0>Grippe</s0><s5>06</s5></fC03><fC03 i1="04" i2="X" l="ENG"><s0>Influenza</s0><s5>06</s5></fC03><fC03 i1="04" i2="X" l="SPA"><s0>Gripe</s0><s5>06</s5></fC03><fC03 i1="05" i2="X" l="FRE"><s0>Analyse quantitative</s0><s5>08</s5></fC03><fC03 i1="05" i2="X" l="ENG"><s0>Quantitative analysis</s0><s5>08</s5></fC03><fC03 i1="05" i2="X" l="SPA"><s0>Análisis cuantitativo</s0><s5>08</s5></fC03><fC03 i1="06" i2="X" l="FRE"><s0>Dosage</s0><s5>09</s5></fC03><fC03 i1="06" i2="X" l="ENG"><s0>Assay</s0><s5>09</s5></fC03><fC03 i1="06" i2="X" l="SPA"><s0>Dosificación</s0><s5>09</s5></fC03><fC03 i1="07" i2="X" l="FRE"><s0>Biochimie</s0><s5>11</s5></fC03><fC03 i1="07" i2="X" l="ENG"><s0>Biochemistry</s0><s5>11</s5></fC03><fC03 i1="07" i2="X" l="SPA"><s0>Bioquímica</s0><s5>11</s5></fC03><fC03 i1="08" i2="X" l="FRE"><s0>Biologie clinique</s0><s5>12</s5></fC03><fC03 i1="08" i2="X" l="ENG"><s0>Clinical biology</s0><s5>12</s5></fC03><fC03 i1="08" i2="X" l="SPA"><s0>Biología clínica</s0><s5>12</s5></fC03><fC03 i1="09" i2="X" l="FRE"><s0>Biologie moléculaire</s0><s5>17</s5></fC03><fC03 i1="09" i2="X" l="ENG"><s0>Molecular biology</s0><s5>17</s5></fC03><fC03 i1="09" i2="X" l="SPA"><s0>Biología molecular</s0><s5>17</s5></fC03><fC03 i1="10" i2="X" l="FRE"><s0>Pandémie</s0><s4>CD</s4><s5>96</s5></fC03><fC03 i1="10" i2="X" l="ENG"><s0>Pandemic</s0><s4>CD</s4><s5>96</s5></fC03><fC03 i1="11" i2="X" l="FRE"><s0>Réaction chaîne polymérase en temps réel</s0><s4>CD</s4><s5>97</s5></fC03><fC03 i1="11" i2="X" l="ENG"><s0>Real time polymerase chain reaction</s0><s4>CD</s4><s5>97</s5></fC03><fC03 i1="11" i2="X" l="SPA"><s0>Reacción en cadena de la polimerasa a tiempo real</s0><s4>CD</s4><s5>97</s5></fC03><fC07 i1="01" i2="X" l="FRE"><s0>Virose</s0></fC07><fC07 i1="01" i2="X" l="ENG"><s0>Viral disease</s0></fC07><fC07 i1="01" i2="X" l="SPA"><s0>Virosis</s0></fC07><fC07 i1="02" i2="X" l="FRE"><s0>Infection</s0></fC07><fC07 i1="02" i2="X" l="ENG"><s0>Infection</s0></fC07><fC07 i1="02" i2="X" l="SPA"><s0>Infección</s0></fC07><fN21><s1>257</s1></fN21><fN44 i1="01"><s1>OTO</s1></fN44><fN82><s1>OTO</s1></fN82></pA></standard></inist><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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