Development of a quantitative assay for SARS coronavirus and correlation of GAPDH mRNA with SARS coronavirus in clinical specimens
Identifieur interne : 004C30 ( Main/Exploration ); précédent : 004C29; suivant : 004C31Development of a quantitative assay for SARS coronavirus and correlation of GAPDH mRNA with SARS coronavirus in clinical specimens
Auteurs : S C C. Wong [République populaire de Chine] ; J K C. Chan [République populaire de Chine] ; K C Lee [République populaire de Chine] ; E S F. Lo [République populaire de Chine] ; D N C. Tsang [République populaire de Chine]Source :
- Journal of Clinical Pathology [ 0021-9746 ] ; 2005-03.
Descripteurs français
- KwdFr :
- ARN messager (génétique), ARN viral (génétique), Actines (biosynthèse), Actines (génétique), Adulte, Adulte d'âge moyen, Contrôle de qualité, Faux négatifs, Femelle, Glyceraldehyde 3-phosphate dehydrogenases (biosynthèse), Glyceraldehyde 3-phosphate dehydrogenases (génétique), Humains, Marqueurs biologiques (métabolisme), Mâle, RT-PCR (), Sensibilité et spécificité, Sujet âgé, Syndrome respiratoire aigu sévère (diagnostic), Virus du SRAS (isolement et purification), Études de suivi.
- MESH :
- biosynthèse : Actines, Glyceraldehyde 3-phosphate dehydrogenases.
- diagnostic : Syndrome respiratoire aigu sévère.
- génétique : ARN messager, ARN viral, Actines, Glyceraldehyde 3-phosphate dehydrogenases.
- isolement et purification : Virus du SRAS.
- métabolisme : Marqueurs biologiques.
- Pascal (Inist)
- MESH :
- Wicri :
- topic : Analyse quantitative.
English descriptors
- KwdEn :
- Actins (biosynthesis), Actins (genetics), Adult, Aged, Anatomic pathology, Biomarkers (metabolism), CoV, coronavirus, Coronavirus, Correlation, Development, False Negative Reactions, Female, Follow-Up Studies, GAPDH, glyceraldehyde-3-phosphate dehydrogenase, Glyceraldehyde-3-Phosphate Dehydrogenases (biosynthesis), Glyceraldehyde-3-Phosphate Dehydrogenases (genetics), Humans, Male, Messenger RNA, Middle Aged, Q-RT-PCR, quantitative reverse transcriptase polymerase chain reaction, Quality Control, Quantitative analysis, RNA, Messenger (genetics), RNA, Viral (genetics), RT-PCR, reverse transcriptase polymerase chain reaction, Reverse Transcriptase Polymerase Chain Reaction (methods), SARS Virus (isolation & purification), SARS, severe acute respiratory syndrome, Sensitivity and Specificity, Severe Acute Respiratory Syndrome (diagnosis), Severe acute respiratory syndrome, WHO, World Health Organisation, cDNA, complementary deoxyribonucleic acid.
- MESH :
- chemical , biosynthesis : Actins, Glyceraldehyde-3-Phosphate Dehydrogenases.
- chemical , genetics : Actins, Glyceraldehyde-3-Phosphate Dehydrogenases, RNA, Messenger, RNA, Viral.
- chemical , metabolism : Biomarkers.
- diagnosis : Severe Acute Respiratory Syndrome.
- isolation & purification : SARS Virus.
- methods : Reverse Transcriptase Polymerase Chain Reaction.
- Teeft :
- Actin, Actin mrna, Administrative region, Adult, Aged, Aliquot, Aspirate, Assay, Cdna, Clinical specimens, Coronavirus, Dehydrogenase, Detectable gapdh mrna, False Negative Reactions, Female, Follow-Up Studies, Gapdh, Gapdh mrna, Gapdh mrna concentration, Gapdh mrna concentrations, Gapdh mrna detection, Healthy controls, Healthy subjects, Higher amounts, Hong kong, Humans, Male, Middle Aged, Mrna, Nasopharyngeal, Nasopharyngeal swabs, Normal subjects, Polymerase, Positive controls, Primer, Quality Control, Queen elizabeth hospital, Rectal, Respiratory syndrome, Respiratory syndrome coronavirus, Sampling techniques, Sars, Sars coronavirus, Sars sars, Sensitivity and Specificity, Significant difference, Statistical analysis, Stool samples, Swab, Syndrome, Throat swabs, Transcriptase, Transcriptase polymerase chain reaction, Transcriptase polymerase chain reaction assay, Unrelated diseases, Urine, Urine specimens, World health organisation.
Abstract
Aims: To develop a quantitative reverse transcriptase polymerase chain reaction (Q-RT-PCR) for severe acute respiratory syndrome coronavirus (SARS-CoV) detection and explore the potential of using glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA as an internal control to exclude false negative results. Methods: SARS-CoV and GAPDH mRNA were both measured in 26 specimens from 16 patients with SARS, 40 follow up specimens from the same batch of patients, and appropriate control subjects. The relation between SARS positivity and GAPDH mRNA concentration was investigated using the χ2 test. Increasing the sensitivity for SARS-CoV and GAPDH mRNA detection was investigated in follow up specimens in which SARS-CoV and GAPDH mRNA were not detected initially. Results: Varying amounts of SARS-CoV were found in the 26 SARS-CoV positive specimens and SARS-CoV was not detected in the 40 follow up specimens and controls. In addition, concentrations of GAPDH mRNA were significantly different between the patients with SARS, follow up specimens, and healthy controls (Kruskal-Wallis test, p<0.05). Moreover, GAPDH mRNA concentrations were highly correlated with SARS-CoV positivity (χ2 = 5.43; p<0.05). Finally, SARS-CoV and GAPDH mRNA were both detected in three follow up urine specimens that were initially negative when the amount of cDNA used was increased from 5 μl to 10 and 15 μl. Conclusions: This Q-RT-PCR assay can be used to detect SARS-CoV. Moreover, GAPDH mRNA may be useful to rule out false negative results in SARS-CoV detection, and the current extraction method for urine may not be sensitive enough to detect low titres of SARS-CoV.
Url:
DOI: 10.1136/jcp.2004.016592
Affiliations:
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Le document en format XML
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Wong</name><affiliation wicri:level="4"><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Department of Clinical Oncology, Prince of Wales Hospital, The Chinese University of Hong Kong, Hong Kong Special Administrative Region</wicri:regionArea><orgName type="university">Université chinoise de Hong Kong</orgName><placeName><settlement type="city">Sha Tin</settlement></placeName></affiliation></author><author><name sortKey="Chan, J K C" sort="Chan, J K C" uniqKey="Chan J" first="J K C" last="Chan">J K C. Chan</name><affiliation wicri:level="1"><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Department of Pathology, Queen Elizabeth Hospital, Hong Kong Special Administrative Region</wicri:regionArea><wicri:noRegion>Hong Kong Special Administrative Region</wicri:noRegion></affiliation></author><author><name sortKey="Lee, K C" sort="Lee, K C" uniqKey="Lee K" first="K C" last="Lee">K C Lee</name><affiliation wicri:level="1"><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Department of Pathology, Queen Elizabeth Hospital, Hong Kong Special Administrative Region</wicri:regionArea><wicri:noRegion>Hong Kong Special Administrative Region</wicri:noRegion></affiliation></author><author><name sortKey="Lo, E S F" sort="Lo, E S F" uniqKey="Lo E" first="E S F" last="Lo">E S F. Lo</name><affiliation wicri:level="1"><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Department of Pathology, Queen Elizabeth Hospital, Hong Kong Special Administrative Region</wicri:regionArea><wicri:noRegion>Hong Kong Special Administrative Region</wicri:noRegion></affiliation></author><author><name sortKey="Tsang, D N C" sort="Tsang, D N C" uniqKey="Tsang D" first="D N C" last="Tsang">D N C. Tsang</name><affiliation wicri:level="1"><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Department of Pathology, Queen Elizabeth Hospital, Hong Kong Special Administrative Region</wicri:regionArea><wicri:noRegion>Hong Kong Special Administrative Region</wicri:noRegion></affiliation></author></analytic><monogr></monogr><series><title level="j">Journal of Clinical Pathology</title><title level="j" type="abbrev">J Clin Pathol</title><idno type="ISSN">0021-9746</idno><idno type="eISSN">1472-4146</idno><imprint><publisher>BMJ Publishing Group Ltd and Association of Clinical Pathologists</publisher><date type="published" when="2005-03">2005-03</date><biblScope unit="volume">58</biblScope><biblScope unit="issue">3</biblScope><biblScope unit="page" from="276">276</biblScope></imprint><idno type="ISSN">0021-9746</idno></series></biblStruct></sourceDesc><seriesStmt><idno type="ISSN">0021-9746</idno></seriesStmt></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Actins (biosynthesis)</term><term>Actins (genetics)</term><term>Adult</term><term>Aged</term><term>Anatomic pathology</term><term>Biomarkers (metabolism)</term><term>CoV, coronavirus</term><term>Coronavirus</term><term>Correlation</term><term>Development</term><term>False Negative Reactions</term><term>Female</term><term>Follow-Up Studies</term><term>GAPDH, glyceraldehyde-3-phosphate dehydrogenase</term><term>Glyceraldehyde-3-Phosphate Dehydrogenases (biosynthesis)</term><term>Glyceraldehyde-3-Phosphate Dehydrogenases (genetics)</term><term>Humans</term><term>Male</term><term>Messenger RNA</term><term>Middle Aged</term><term>Q-RT-PCR, quantitative reverse transcriptase polymerase chain reaction</term><term>Quality Control</term><term>Quantitative analysis</term><term>RNA, Messenger (genetics)</term><term>RNA, Viral (genetics)</term><term>RT-PCR, reverse transcriptase polymerase chain reaction</term><term>Reverse Transcriptase Polymerase Chain Reaction (methods)</term><term>SARS Virus (isolation & purification)</term><term>SARS, severe acute respiratory syndrome</term><term>Sensitivity and Specificity</term><term>Severe Acute Respiratory Syndrome (diagnosis)</term><term>Severe acute respiratory syndrome</term><term>WHO, World Health Organisation</term><term>cDNA, complementary deoxyribonucleic acid</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>ARN messager (génétique)</term><term>ARN viral (génétique)</term><term>Actines (biosynthèse)</term><term>Actines (génétique)</term><term>Adulte</term><term>Adulte d'âge moyen</term><term>Contrôle de qualité</term><term>Faux négatifs</term><term>Femelle</term><term>Glyceraldehyde 3-phosphate dehydrogenases (biosynthèse)</term><term>Glyceraldehyde 3-phosphate dehydrogenases (génétique)</term><term>Humains</term><term>Marqueurs biologiques (métabolisme)</term><term>Mâle</term><term>RT-PCR ()</term><term>Sensibilité et spécificité</term><term>Sujet âgé</term><term>Syndrome respiratoire aigu sévère (diagnostic)</term><term>Virus du SRAS (isolement et purification)</term><term>Études de suivi</term></keywords><keywords scheme="MESH" type="chemical" qualifier="biosynthesis" xml:lang="en"><term>Actins</term><term>Glyceraldehyde-3-Phosphate Dehydrogenases</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Actins</term><term>Glyceraldehyde-3-Phosphate Dehydrogenases</term><term>RNA, Messenger</term><term>RNA, Viral</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>Biomarkers</term></keywords><keywords scheme="MESH" qualifier="biosynthèse" xml:lang="fr"><term>Actines</term><term>Glyceraldehyde 3-phosphate dehydrogenases</term></keywords><keywords scheme="MESH" qualifier="diagnosis" xml:lang="en"><term>Severe Acute Respiratory Syndrome</term></keywords><keywords scheme="MESH" qualifier="diagnostic" xml:lang="fr"><term>Syndrome respiratoire aigu sévère</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>ARN messager</term><term>ARN viral</term><term>Actines</term><term>Glyceraldehyde 3-phosphate dehydrogenases</term></keywords><keywords scheme="MESH" qualifier="isolation & purification" xml:lang="en"><term>SARS Virus</term></keywords><keywords scheme="MESH" qualifier="isolement et purification" xml:lang="fr"><term>Virus du SRAS</term></keywords><keywords scheme="MESH" qualifier="methods" xml:lang="en"><term>Reverse Transcriptase Polymerase Chain Reaction</term></keywords><keywords scheme="MESH" qualifier="métabolisme" xml:lang="fr"><term>Marqueurs biologiques</term></keywords><keywords scheme="Pascal" xml:lang="fr"><term>Analyse quantitative</term><term>Anatomopathologie</term><term>Coronavirus</term><term>Corrélation</term><term>Développement</term><term>RNA messager</term><term>Syndrome respiratoire aigu sévère</term></keywords><keywords scheme="Teeft" xml:lang="en"><term>Actin</term><term>Actin mrna</term><term>Administrative region</term><term>Adult</term><term>Aged</term><term>Aliquot</term><term>Aspirate</term><term>Assay</term><term>Cdna</term><term>Clinical specimens</term><term>Coronavirus</term><term>Dehydrogenase</term><term>Detectable gapdh mrna</term><term>False Negative Reactions</term><term>Female</term><term>Follow-Up Studies</term><term>Gapdh</term><term>Gapdh mrna</term><term>Gapdh mrna concentration</term><term>Gapdh mrna concentrations</term><term>Gapdh mrna detection</term><term>Healthy controls</term><term>Healthy subjects</term><term>Higher amounts</term><term>Hong kong</term><term>Humans</term><term>Male</term><term>Middle Aged</term><term>Mrna</term><term>Nasopharyngeal</term><term>Nasopharyngeal swabs</term><term>Normal subjects</term><term>Polymerase</term><term>Positive controls</term><term>Primer</term><term>Quality Control</term><term>Queen elizabeth hospital</term><term>Rectal</term><term>Respiratory syndrome</term><term>Respiratory syndrome coronavirus</term><term>Sampling techniques</term><term>Sars</term><term>Sars coronavirus</term><term>Sars sars</term><term>Sensitivity and Specificity</term><term>Significant difference</term><term>Statistical analysis</term><term>Stool samples</term><term>Swab</term><term>Syndrome</term><term>Throat swabs</term><term>Transcriptase</term><term>Transcriptase polymerase chain reaction</term><term>Transcriptase polymerase chain reaction assay</term><term>Unrelated diseases</term><term>Urine</term><term>Urine specimens</term><term>World health organisation</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Adulte</term><term>Adulte d'âge moyen</term><term>Contrôle de qualité</term><term>Faux négatifs</term><term>Femelle</term><term>Humains</term><term>Mâle</term><term>RT-PCR</term><term>Sensibilité et spécificité</term><term>Sujet âgé</term><term>Études de suivi</term></keywords><keywords scheme="Wicri" type="topic" xml:lang="fr"><term>Analyse quantitative</term></keywords></textClass><langUsage><language ident="en">en</language></langUsage></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Aims: To develop a quantitative reverse transcriptase polymerase chain reaction (Q-RT-PCR) for severe acute respiratory syndrome coronavirus (SARS-CoV) detection and explore the potential of using glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA as an internal control to exclude false negative results. Methods: SARS-CoV and GAPDH mRNA were both measured in 26 specimens from 16 patients with SARS, 40 follow up specimens from the same batch of patients, and appropriate control subjects. The relation between SARS positivity and GAPDH mRNA concentration was investigated using the χ2 test. Increasing the sensitivity for SARS-CoV and GAPDH mRNA detection was investigated in follow up specimens in which SARS-CoV and GAPDH mRNA were not detected initially. Results: Varying amounts of SARS-CoV were found in the 26 SARS-CoV positive specimens and SARS-CoV was not detected in the 40 follow up specimens and controls. In addition, concentrations of GAPDH mRNA were significantly different between the patients with SARS, follow up specimens, and healthy controls (Kruskal-Wallis test, p<0.05). Moreover, GAPDH mRNA concentrations were highly correlated with SARS-CoV positivity (χ2 = 5.43; p<0.05). Finally, SARS-CoV and GAPDH mRNA were both detected in three follow up urine specimens that were initially negative when the amount of cDNA used was increased from 5 μl to 10 and 15 μl. Conclusions: This Q-RT-PCR assay can be used to detect SARS-CoV. Moreover, GAPDH mRNA may be useful to rule out false negative results in SARS-CoV detection, and the current extraction method for urine may not be sensitive enough to detect low titres of SARS-CoV.</div></front></TEI><affiliations><list><country><li>République populaire de Chine</li></country><settlement><li>Sha Tin</li></settlement><orgName><li>Université chinoise de Hong Kong</li></orgName></list><tree><country name="République populaire de Chine"><noRegion><name sortKey="Wong, S C C" sort="Wong, S C C" uniqKey="Wong S" first="S C C" last="Wong">S C C. Wong</name></noRegion><name sortKey="Chan, J K C" sort="Chan, J K C" uniqKey="Chan J" first="J K C" last="Chan">J K C. Chan</name><name sortKey="Lee, K C" sort="Lee, K C" uniqKey="Lee K" first="K C" last="Lee">K C Lee</name><name sortKey="Lo, E S F" sort="Lo, E S F" uniqKey="Lo E" first="E S F" last="Lo">E S F. Lo</name><name sortKey="Tsang, D N C" sort="Tsang, D N C" uniqKey="Tsang D" first="D N C" last="Tsang">D N C. Tsang</name></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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