Performance and cost evaluation of one commercial and six in-house conventional and real-time reverse transcription-PCR assays for detection of severe acute respiratory syndrome coronavirus
Identifieur interne : 005A95 ( Main/Exploration ); précédent : 005A94; suivant : 005A96Performance and cost evaluation of one commercial and six in-house conventional and real-time reverse transcription-PCR assays for detection of severe acute respiratory syndrome coronavirus
Auteurs : James B. Mahony [Canada] ; Astrid Petrich [Canada] ; Lisa Louie [Canada] ; XINYU SONG [Canada] ; Sylvia Chong [Canada] ; Marek Smieja [Canada] ; Max Chernesky [Canada] ; Mark Loeb [Canada] ; Susan Richardson [Canada]Source :
- Journal of clinical microbiology : (Print) [ 0095-1137 ] ; 2004.
Descripteurs français
- KwdFr :
- MESH :
- analyse : ARN viral.
- génétique : Virus du SRAS.
- isolement et purification : Virus du SRAS.
- virologie : Syndrome respiratoire aigu sévère.
- économie : RT-PCR, Trousses de réactifs pour diagnostic.
- Pascal (Inist)
English descriptors
- KwdEn :
- Coronavirus, Costs and Cost Analysis, Detection, Humans, Microbiology, Performance evaluation, RNA, Viral (analysis), Reagent Kits, Diagnostic (economics), Real time, Reverse Transcriptase Polymerase Chain Reaction (economics), Reverse Transcriptase Polymerase Chain Reaction (methods), Reverse transcription polymerase chain reaction, SARS Virus (genetics), SARS Virus (isolation & purification), Sensitivity and Specificity, Severe Acute Respiratory Syndrome (virology), Severe acute respiratory syndrome.
- MESH :
- chemical , analysis : RNA, Viral.
- chemical , economics : Reagent Kits, Diagnostic.
- economics : Reverse Transcriptase Polymerase Chain Reaction.
- genetics : SARS Virus.
- isolation & purification : SARS Virus.
- methods : Reverse Transcriptase Polymerase Chain Reaction.
- virology : Severe Acute Respiratory Syndrome.
- Costs and Cost Analysis, Humans, Sensitivity and Specificity.
Abstract
We evaluated seven reverse transcription-PCR (RT-PCR) assays, including six in-house assays and one commercial assay for the detection of severe acute respiratory syndrome coronavirus (SARS-CoV) RNA in clinical specimens. RT-PCR assays targeted different genomic regions and included three conventional assays (one nested and two non-nested) run on a conventional heat block and four real-time assays performed in a LightCycler (LC; Roche Diagnostics). All in-house assays were optimized for assay parameters, including MgCl2, primer, and probe concentrations. The commercial assay was the RealArt HPA CoV RT-PCR assay (Artus), which was run in the LC. Testing serial dilutions of cultured SARS-CoV showed that the analytical sensitivity of the assays ranged from 10-8 to 10-6, corresponding to 1 and 100 copies of viral RNA, respectively. Significant differences in analytical sensitivities were observed between assays (P < 0.01, probit regression analysis for 50% sensitivity levels for the top two assays versus the others). Testing 68 clinical specimens (including 17 respiratory tract specimens, 29 urine samples, and 22 stools or rectal swabs) demonstrated that six of the seven assays detected at least 17 of 18 positives (defined as positive in at least two assays), and two of the assays had a sensitivity of 100%. There were no significant differences in sensitivity between the assays (P = 0.5 [Cochrance Q test, least sensitive 15 of 18 versus 18 of 18]). The specificities of the assays ranged from 94.0 to 100% without significant differences (P = 0.25 to 0.5 [McNemar test]). The reagent and technologist cost of performing the in-house PCR assays ranged from $5.46 to $9.81 Canadian dollars (CDN) per test. The commercial assay cost was considerably higher at $40.37 per test. The results demonstrated good performance for all assays, providing laboratories that need to do SARS RNA testing with a choice of assay formats.
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Mahony</name><affiliation wicri:level="4"><inist:fA14 i1="01"><s1>McMaster University</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ><sZ>8 aut.</sZ></inist:fA14><country>Canada</country><placeName><settlement type="city">Hamilton (Ontario)</settlement><region type="state">Ontario</region></placeName><orgName type="university">Université McMaster</orgName></affiliation><affiliation wicri:level="1"><inist:fA14 i1="02"><s1>Father Sean O'Sullivan Research Centre, St. Joseph's Healthcare</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>4 aut.</sZ><sZ>5 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ></inist:fA14><country>Canada</country><wicri:noRegion>Hamilton</wicri:noRegion></affiliation></author><author><name sortKey="Petrich, Astrid" sort="Petrich, Astrid" uniqKey="Petrich A" first="Astrid" last="Petrich">Astrid Petrich</name><affiliation wicri:level="4"><inist:fA14 i1="01"><s1>McMaster 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aut.</sZ></inist:fA14><country>Canada</country><wicri:noRegion>Hamilton</wicri:noRegion></affiliation></author><author><name sortKey="Smieja, Marek" sort="Smieja, Marek" uniqKey="Smieja M" first="Marek" last="Smieja">Marek Smieja</name><affiliation wicri:level="4"><inist:fA14 i1="01"><s1>McMaster University</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ><sZ>8 aut.</sZ></inist:fA14><country>Canada</country><placeName><settlement type="city">Hamilton (Ontario)</settlement><region type="state">Ontario</region></placeName><orgName type="university">Université McMaster</orgName></affiliation><affiliation wicri:level="1"><inist:fA14 i1="02"><s1>Father Sean O'Sullivan Research Centre, St. Joseph's Healthcare</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>4 aut.</sZ><sZ>5 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ></inist:fA14><country>Canada</country><wicri:noRegion>Hamilton</wicri:noRegion></affiliation></author><author><name 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Mahony</name><affiliation wicri:level="4"><inist:fA14 i1="01"><s1>McMaster University</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ><sZ>8 aut.</sZ></inist:fA14><country>Canada</country><placeName><settlement type="city">Hamilton (Ontario)</settlement><region type="state">Ontario</region></placeName><orgName type="university">Université McMaster</orgName></affiliation><affiliation wicri:level="1"><inist:fA14 i1="02"><s1>Father Sean O'Sullivan Research Centre, St. Joseph's Healthcare</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>4 aut.</sZ><sZ>5 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ></inist:fA14><country>Canada</country><wicri:noRegion>Hamilton</wicri:noRegion></affiliation></author><author><name sortKey="Petrich, Astrid" sort="Petrich, Astrid" uniqKey="Petrich A" first="Astrid" last="Petrich">Astrid Petrich</name><affiliation wicri:level="4"><inist:fA14 i1="01"><s1>McMaster University</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ><sZ>8 aut.</sZ></inist:fA14><country>Canada</country><placeName><settlement type="city">Hamilton (Ontario)</settlement><region type="state">Ontario</region></placeName><orgName type="university">Université McMaster</orgName></affiliation><affiliation wicri:level="1"><inist:fA14 i1="02"><s1>Father Sean O'Sullivan Research Centre, St. Joseph's Healthcare</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>4 aut.</sZ><sZ>5 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ></inist:fA14><country>Canada</country><wicri:noRegion>Hamilton</wicri:noRegion></affiliation></author><author><name sortKey="Louie, Lisa" sort="Louie, Lisa" uniqKey="Louie L" first="Lisa" last="Louie">Lisa Louie</name><affiliation wicri:level="1"><inist:fA14 i1="03"><s1>Sunnybrook and Women's College Health Sciences Centre</s1><s2>Toronto, Ontario L8N 4A6</s2><s3>CAN</s3><sZ>3 aut.</sZ></inist:fA14><country>Canada</country><wicri:noRegion>Sunnybrook and Women's College Health Sciences Centre</wicri:noRegion></affiliation></author><author><name sortKey="Xinyu Song" sort="Xinyu Song" uniqKey="Xinyu Song" last="Xinyu Song">XINYU SONG</name><affiliation wicri:level="1"><inist:fA14 i1="02"><s1>Father Sean O'Sullivan Research Centre, St. Joseph's Healthcare</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>4 aut.</sZ><sZ>5 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ></inist:fA14><country>Canada</country><wicri:noRegion>Hamilton</wicri:noRegion></affiliation></author><author><name sortKey="Chong, Sylvia" sort="Chong, Sylvia" uniqKey="Chong S" first="Sylvia" last="Chong">Sylvia Chong</name><affiliation wicri:level="1"><inist:fA14 i1="02"><s1>Father Sean O'Sullivan Research Centre, St. Joseph's Healthcare</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>4 aut.</sZ><sZ>5 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ></inist:fA14><country>Canada</country><wicri:noRegion>Hamilton</wicri:noRegion></affiliation></author><author><name sortKey="Smieja, Marek" sort="Smieja, Marek" uniqKey="Smieja M" first="Marek" last="Smieja">Marek Smieja</name><affiliation wicri:level="4"><inist:fA14 i1="01"><s1>McMaster University</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ><sZ>8 aut.</sZ></inist:fA14><country>Canada</country><placeName><settlement type="city">Hamilton (Ontario)</settlement><region type="state">Ontario</region></placeName><orgName type="university">Université McMaster</orgName></affiliation><affiliation wicri:level="1"><inist:fA14 i1="02"><s1>Father Sean O'Sullivan Research Centre, St. Joseph's Healthcare</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>4 aut.</sZ><sZ>5 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ></inist:fA14><country>Canada</country><wicri:noRegion>Hamilton</wicri:noRegion></affiliation></author><author><name sortKey="Chernesky, Max" sort="Chernesky, Max" uniqKey="Chernesky M" first="Max" last="Chernesky">Max Chernesky</name><affiliation wicri:level="4"><inist:fA14 i1="01"><s1>McMaster University</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ><sZ>8 aut.</sZ></inist:fA14><country>Canada</country><placeName><settlement type="city">Hamilton (Ontario)</settlement><region type="state">Ontario</region></placeName><orgName type="university">Université McMaster</orgName></affiliation><affiliation wicri:level="1"><inist:fA14 i1="02"><s1>Father Sean O'Sullivan Research Centre, St. Joseph's Healthcare</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>4 aut.</sZ><sZ>5 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ></inist:fA14><country>Canada</country><wicri:noRegion>Hamilton</wicri:noRegion></affiliation></author><author><name sortKey="Loeb, Mark" sort="Loeb, Mark" uniqKey="Loeb M" first="Mark" last="Loeb">Mark Loeb</name><affiliation wicri:level="4"><inist:fA14 i1="01"><s1>McMaster University</s1><s2>Hamilton</s2><s3>CAN</s3><sZ>1 aut.</sZ><sZ>2 aut.</sZ><sZ>6 aut.</sZ><sZ>7 aut.</sZ><sZ>8 aut.</sZ></inist:fA14><country>Canada</country><placeName><settlement type="city">Hamilton (Ontario)</settlement><region type="state">Ontario</region></placeName><orgName type="university">Université McMaster</orgName></affiliation></author><author><name sortKey="Richardson, Susan" sort="Richardson, Susan" uniqKey="Richardson S" first="Susan" last="Richardson">Susan Richardson</name><affiliation wicri:level="1"><inist:fA14 i1="04"><s1>University of Toronto and Heathcare Network, The Hospital for Sick Children</s1><s2>Toronto, Ontario L8N 4A6</s2><s3>CAN</s3><sZ>9 aut.</sZ></inist:fA14><country>Canada</country><wicri:noRegion>Toronto, Ontario L8N 4A6</wicri:noRegion></affiliation></author></analytic><series><title level="j" type="main">Journal of clinical microbiology : (Print)</title><title level="j" type="abbreviated">J. clin. microbiol. : (Print)</title><idno type="ISSN">0095-1137</idno><imprint><date when="2004">2004</date></imprint></series></biblStruct></sourceDesc><seriesStmt><title level="j" type="main">Journal of clinical microbiology : (Print)</title><title level="j" type="abbreviated">J. clin. microbiol. : (Print)</title><idno type="ISSN">0095-1137</idno></seriesStmt></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Coronavirus</term><term>Costs and Cost Analysis</term><term>Detection</term><term>Humans</term><term>Microbiology</term><term>Performance evaluation</term><term>RNA, Viral (analysis)</term><term>Reagent Kits, Diagnostic (economics)</term><term>Real time</term><term>Reverse Transcriptase Polymerase Chain Reaction (economics)</term><term>Reverse Transcriptase Polymerase Chain Reaction (methods)</term><term>Reverse transcription polymerase chain reaction</term><term>SARS Virus (genetics)</term><term>SARS Virus (isolation & purification)</term><term>Sensitivity and Specificity</term><term>Severe Acute Respiratory Syndrome (virology)</term><term>Severe acute respiratory syndrome</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>ARN viral (analyse)</term><term>Coûts et analyse des coûts</term><term>Humains</term><term>RT-PCR ()</term><term>RT-PCR (économie)</term><term>Sensibilité et spécificité</term><term>Syndrome respiratoire aigu sévère (virologie)</term><term>Trousses de réactifs pour diagnostic (économie)</term><term>Virus du SRAS (génétique)</term><term>Virus du SRAS (isolement et purification)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="analysis" xml:lang="en"><term>RNA, Viral</term></keywords><keywords scheme="MESH" type="chemical" qualifier="economics" xml:lang="en"><term>Reagent Kits, Diagnostic</term></keywords><keywords scheme="MESH" qualifier="analyse" xml:lang="fr"><term>ARN viral</term></keywords><keywords scheme="MESH" qualifier="economics" xml:lang="en"><term>Reverse Transcriptase Polymerase Chain Reaction</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>SARS Virus</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>Virus du SRAS</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="virologie" xml:lang="fr"><term>Syndrome respiratoire aigu sévère</term></keywords><keywords scheme="MESH" qualifier="virology" xml:lang="en"><term>Severe Acute Respiratory Syndrome</term></keywords><keywords scheme="MESH" qualifier="économie" xml:lang="fr"><term>RT-PCR</term><term>Trousses de réactifs pour diagnostic</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Costs and Cost Analysis</term><term>Humans</term><term>Sensitivity and Specificity</term></keywords><keywords scheme="Pascal" xml:lang="fr"><term>Coronavirus</term><term>Coûts et analyse des coûts</term><term>Evaluation performance</term><term>Humains</term><term>RT-PCR</term><term>Sensibilité et spécificité</term><term>Temps réel</term><term>Réaction chaîne polymérase RT</term><term>Détection</term><term>Microbiologie</term><term>Syndrome respiratoire aigu sévère</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">We evaluated seven reverse transcription-PCR (RT-PCR) assays, including six in-house assays and one commercial assay for the detection of severe acute respiratory syndrome coronavirus (SARS-CoV) RNA in clinical specimens. RT-PCR assays targeted different genomic regions and included three conventional assays (one nested and two non-nested) run on a conventional heat block and four real-time assays performed in a LightCycler (LC; Roche Diagnostics). All in-house assays were optimized for assay parameters, including MgCl<sub>2</sub>, primer, and probe concentrations. The commercial assay was the RealArt HPA CoV RT-PCR assay (Artus), which was run in the LC. Testing serial dilutions of cultured SARS-CoV showed that the analytical sensitivity of the assays ranged from 10<sup>-8</sup> to 10<sup>-6</sup>, corresponding to 1 and 100 copies of viral RNA, respectively. Significant differences in analytical sensitivities were observed between assays (P < 0.01, probit regression analysis for 50% sensitivity levels for the top two assays versus the others). Testing 68 clinical specimens (including 17 respiratory tract specimens, 29 urine samples, and 22 stools or rectal swabs) demonstrated that six of the seven assays detected at least 17 of 18 positives (defined as positive in at least two assays), and two of the assays had a sensitivity of 100%. There were no significant differences in sensitivity between the assays (P = 0.5 [Cochrance Q test, least sensitive 15 of 18 versus 18 of 18]). The specificities of the assays ranged from 94.0 to 100% without significant differences (P = 0.25 to 0.5 [McNemar test]). The reagent and technologist cost of performing the in-house PCR assays ranged from $5.46 to $9.81 Canadian dollars (CDN) per test. The commercial assay cost was considerably higher at $40.37 per test. The results demonstrated good performance for all assays, providing laboratories that need to do SARS RNA testing with a choice of assay formats.</div></front></TEI><affiliations><list><country><li>Canada</li></country><region><li>Ontario</li></region><settlement><li>Hamilton (Ontario)</li></settlement><orgName><li>Université McMaster</li></orgName></list><tree><country name="Canada"><region name="Ontario"><name sortKey="Mahony, James B" sort="Mahony, James B" uniqKey="Mahony J" first="James B." last="Mahony">James B. Mahony</name></region><name sortKey="Chernesky, Max" sort="Chernesky, Max" uniqKey="Chernesky M" first="Max" last="Chernesky">Max Chernesky</name><name sortKey="Chernesky, Max" sort="Chernesky, Max" uniqKey="Chernesky M" first="Max" last="Chernesky">Max Chernesky</name><name sortKey="Chong, Sylvia" sort="Chong, Sylvia" uniqKey="Chong S" first="Sylvia" last="Chong">Sylvia Chong</name><name sortKey="Loeb, Mark" sort="Loeb, Mark" uniqKey="Loeb M" first="Mark" last="Loeb">Mark Loeb</name><name sortKey="Louie, Lisa" sort="Louie, Lisa" uniqKey="Louie L" first="Lisa" last="Louie">Lisa Louie</name><name sortKey="Mahony, James B" sort="Mahony, James B" uniqKey="Mahony J" first="James B." last="Mahony">James B. Mahony</name><name sortKey="Petrich, Astrid" sort="Petrich, Astrid" uniqKey="Petrich A" first="Astrid" last="Petrich">Astrid Petrich</name><name sortKey="Petrich, Astrid" sort="Petrich, Astrid" uniqKey="Petrich A" first="Astrid" last="Petrich">Astrid Petrich</name><name sortKey="Richardson, Susan" sort="Richardson, Susan" uniqKey="Richardson S" first="Susan" last="Richardson">Susan Richardson</name><name sortKey="Smieja, Marek" sort="Smieja, Marek" uniqKey="Smieja M" first="Marek" last="Smieja">Marek Smieja</name><name sortKey="Smieja, Marek" sort="Smieja, Marek" uniqKey="Smieja M" first="Marek" last="Smieja">Marek Smieja</name><name sortKey="Xinyu Song" sort="Xinyu Song" uniqKey="Xinyu Song" last="Xinyu Song">XINYU SONG</name></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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