Multicenter comparison of nucleic acid extraction methods for detection of severe acute respiratory syndrome coronavirus RNA in stool specimens.
Identifieur interne : 003F90 ( Main/Merge ); précédent : 003F89; suivant : 003F91Multicenter comparison of nucleic acid extraction methods for detection of severe acute respiratory syndrome coronavirus RNA in stool specimens.
Auteurs : A. Petrich [Canada] ; J. Mahony ; S. Chong ; G. Broukhanski ; F. Gharabaghi ; G. Johnson ; L. Louie ; K. Luinstra ; B. Willey ; P. Akhaven ; L. Chui ; F. Jamieson ; M. Louie ; T. Mazzulli ; R. Tellier ; M. Smieja ; W. Cai ; M. Chernesky ; S E RichardsonSource :
- Journal of clinical microbiology [ 0095-1137 ] ; 2006.
Descripteurs français
- KwdFr :
- ARN viral (analyse), ARN viral (génétique), ARN viral (isolement et purification), Fèces (virologie), Humains, RT-PCR, Sensibilité et spécificité, Syndrome respiratoire aigu sévère (diagnostic), Syndrome respiratoire aigu sévère (virologie), Techniques de diagnostic moléculaire, Virus du SRAS (génétique), Virus du SRAS (isolement et purification).
- MESH :
- analyse : ARN viral.
- diagnostic : Syndrome respiratoire aigu sévère.
- génétique : ARN viral, Virus du SRAS.
- isolement et purification : ARN viral, Virus du SRAS.
- virologie : Fèces, Syndrome respiratoire aigu sévère.
- Humains, RT-PCR, Sensibilité et spécificité, Techniques de diagnostic moléculaire.
English descriptors
- KwdEn :
- Feces (virology), Humans, Molecular Diagnostic Techniques, RNA, Viral (analysis), RNA, Viral (genetics), RNA, Viral (isolation & purification), Reverse Transcriptase Polymerase Chain Reaction, SARS Virus (genetics), SARS Virus (isolation & purification), Sensitivity and Specificity, Severe Acute Respiratory Syndrome (diagnosis), Severe Acute Respiratory Syndrome (virology).
- MESH :
- chemical , analysis : RNA, Viral.
- chemical , genetics : RNA, Viral.
- chemical , isolation & purification : RNA, Viral.
- diagnosis : Severe Acute Respiratory Syndrome.
- genetics : SARS Virus.
- isolation & purification : SARS Virus.
- virology : Feces, Severe Acute Respiratory Syndrome.
- Humans, Molecular Diagnostic Techniques, Reverse Transcriptase Polymerase Chain Reaction, Sensitivity and Specificity.
Abstract
The emergence of a novel coronavirus (CoV) as the cause of severe acute respiratory syndrome (SARS) catalyzed the development of rapid diagnostic tests. Stool samples have been shown to be appropriate for diagnostic testing for SARS CoV, although it has been recognized to be a heterogeneous and difficult sample that contains amplification inhibitors. Limited information on the efficiency of extraction methods for the purification and concentration of SARS CoV RNA from stool samples is available. Our study objectives were to determine the optimal extraction method for SARS CoV RNA detection and to examine the effect of increased specimen volume for the detection of SARS CoV RNA in stool specimens. We conducted a multicenter evaluation of four automated and four manual extraction methods using dilutions of viral lysate in replicate mock stool samples, followed by quantitation of SARS CoV RNA using real-time reverse transcriptase PCR. The sensitivities of the manual methods ranged from 50% to 100%, with the Cortex Biochem Magazorb method, a magnetic bead isolation method, allowing detection of all 12 positive samples. The sensitivities of the automated methods ranged from 75% to 100%. The bioMérieux NucliSens automated extractor and miniMag extraction methods each had a sensitivity of 100%. Examination of the copy numbers detected and the generation of 10-fold dilutions of the extracted material indicated that a number of extraction methods retained inhibitory substances that prevented optimal amplification. Increasing the volume of sample input did improve detection. This information could be useful for the extraction of other RNA viruses from stool samples and demonstrates the need to evaluate extraction methods for different specimen types.
DOI: 10.1128/JCM.02460-05
PubMed: 16891478
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pubmed:16891478Le document en format XML
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Petrich</name><affiliation wicri:level="1"><nlm:affiliation>St. Joseph's Healthcare, L424, Microbiology, 50 Charlton Ave. East, Hamilton, ON L8N 4A6, Canada. petricha@mcmaster.ca</nlm:affiliation><country xml:lang="fr">Canada</country><wicri:regionArea>St. Joseph's Healthcare, L424, Microbiology, 50 Charlton Ave. East, Hamilton, ON L8N 4A6</wicri:regionArea><wicri:noRegion>ON L8N 4A6</wicri:noRegion></affiliation></author><author><name sortKey="Mahony, J" sort="Mahony, J" uniqKey="Mahony J" first="J" last="Mahony">J. Mahony</name></author><author><name sortKey="Chong, S" sort="Chong, S" uniqKey="Chong S" first="S" last="Chong">S. Chong</name></author><author><name sortKey="Broukhanski, G" sort="Broukhanski, G" uniqKey="Broukhanski G" first="G" last="Broukhanski">G. Broukhanski</name></author><author><name sortKey="Gharabaghi, F" sort="Gharabaghi, F" uniqKey="Gharabaghi F" first="F" last="Gharabaghi">F. Gharabaghi</name></author><author><name sortKey="Johnson, G" sort="Johnson, G" uniqKey="Johnson G" first="G" last="Johnson">G. Johnson</name></author><author><name sortKey="Louie, L" sort="Louie, L" uniqKey="Louie L" first="L" last="Louie">L. Louie</name></author><author><name sortKey="Luinstra, K" sort="Luinstra, K" uniqKey="Luinstra K" first="K" last="Luinstra">K. Luinstra</name></author><author><name sortKey="Willey, B" sort="Willey, B" uniqKey="Willey B" first="B" last="Willey">B. Willey</name></author><author><name sortKey="Akhaven, P" sort="Akhaven, P" uniqKey="Akhaven P" first="P" last="Akhaven">P. Akhaven</name></author><author><name sortKey="Chui, L" sort="Chui, L" uniqKey="Chui L" first="L" last="Chui">L. Chui</name></author><author><name sortKey="Jamieson, F" sort="Jamieson, F" uniqKey="Jamieson F" first="F" last="Jamieson">F. Jamieson</name></author><author><name sortKey="Louie, M" sort="Louie, M" uniqKey="Louie M" first="M" last="Louie">M. Louie</name></author><author><name sortKey="Mazzulli, T" sort="Mazzulli, T" uniqKey="Mazzulli T" first="T" last="Mazzulli">T. Mazzulli</name></author><author><name sortKey="Tellier, R" sort="Tellier, R" uniqKey="Tellier R" first="R" last="Tellier">R. Tellier</name></author><author><name sortKey="Smieja, M" sort="Smieja, M" uniqKey="Smieja M" first="M" last="Smieja">M. Smieja</name></author><author><name sortKey="Cai, W" sort="Cai, W" uniqKey="Cai W" first="W" last="Cai">W. Cai</name></author><author><name sortKey="Chernesky, M" sort="Chernesky, M" uniqKey="Chernesky M" first="M" last="Chernesky">M. Chernesky</name></author><author><name sortKey="Richardson, S E" sort="Richardson, S E" uniqKey="Richardson S" first="S E" last="Richardson">S E Richardson</name></author></analytic><series><title level="j">Journal of clinical microbiology</title><idno type="ISSN">0095-1137</idno><imprint><date when="2006" type="published">2006</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Feces (virology)</term><term>Humans</term><term>Molecular Diagnostic Techniques</term><term>RNA, Viral (analysis)</term><term>RNA, Viral (genetics)</term><term>RNA, Viral (isolation & purification)</term><term>Reverse Transcriptase 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 (diagnosis)</term><term>Severe Acute Respiratory Syndrome (virology)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>ARN viral (analyse)</term><term>ARN viral (génétique)</term><term>ARN viral (isolement et purification)</term><term>Fèces (virologie)</term><term>Humains</term><term>RT-PCR</term><term>Sensibilité et spécificité</term><term>Syndrome respiratoire aigu sévère (diagnostic)</term><term>Syndrome respiratoire aigu sévère (virologie)</term><term>Techniques de diagnostic moléculaire</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="genetics" xml:lang="en"><term>RNA, Viral</term></keywords><keywords scheme="MESH" type="chemical" qualifier="isolation & purification" xml:lang="en"><term>RNA, Viral</term></keywords><keywords scheme="MESH" qualifier="analyse" xml:lang="fr"><term>ARN viral</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="genetics" xml:lang="en"><term>SARS Virus</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>ARN viral</term><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>ARN viral</term><term>Virus du SRAS</term></keywords><keywords scheme="MESH" qualifier="virologie" xml:lang="fr"><term>Fèces</term><term>Syndrome respiratoire aigu sévère</term></keywords><keywords scheme="MESH" qualifier="virology" xml:lang="en"><term>Feces</term><term>Severe Acute Respiratory Syndrome</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Humans</term><term>Molecular Diagnostic Techniques</term><term>Reverse Transcriptase Polymerase Chain Reaction</term><term>Sensitivity and Specificity</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Humains</term><term>RT-PCR</term><term>Sensibilité et spécificité</term><term>Techniques de diagnostic moléculaire</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">The emergence of a novel coronavirus (CoV) as the cause of severe acute respiratory syndrome (SARS) catalyzed the development of rapid diagnostic tests. Stool samples have been shown to be appropriate for diagnostic testing for SARS CoV, although it has been recognized to be a heterogeneous and difficult sample that contains amplification inhibitors. Limited information on the efficiency of extraction methods for the purification and concentration of SARS CoV RNA from stool samples is available. Our study objectives were to determine the optimal extraction method for SARS CoV RNA detection and to examine the effect of increased specimen volume for the detection of SARS CoV RNA in stool specimens. We conducted a multicenter evaluation of four automated and four manual extraction methods using dilutions of viral lysate in replicate mock stool samples, followed by quantitation of SARS CoV RNA using real-time reverse transcriptase PCR. The sensitivities of the manual methods ranged from 50% to 100%, with the Cortex Biochem Magazorb method, a magnetic bead isolation method, allowing detection of all 12 positive samples. The sensitivities of the automated methods ranged from 75% to 100%. The bioMérieux NucliSens automated extractor and miniMag extraction methods each had a sensitivity of 100%. Examination of the copy numbers detected and the generation of 10-fold dilutions of the extracted material indicated that a number of extraction methods retained inhibitory substances that prevented optimal amplification. Increasing the volume of sample input did improve detection. This information could be useful for the extraction of other RNA viruses from stool samples and demonstrates the need to evaluate extraction methods for different specimen types.</div></front></TEI></record>Pour manipuler ce document sous Unix (Dilib)
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