A colorimetric microtiter plate PCR system detects respiratory syncytial virus in nasal aspirates and discriminates subtypes A and B
We developed a colorimetric microtiter plate (MTP) PCR system for specific detection of the respiratory syncytial virus (RSV) nucleocapsid gene and differentiation of viral subtypes A and B. Of 47 pediatric nasal aspirate specimens, the sensitivity and specificity were 94.4% (17 of 18) and 100% (15...
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| Veröffentlicht in: | Diagnostic microbiology and infectious disease 1999-08, Vol.34 (4), p.333-337 |
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| container_title | Diagnostic microbiology and infectious disease |
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| creator | Tang, Yi-Wei Heimgartner, Paul J Tollefson, Sharon J Berg, Timothy J Rys, Paul N Li, Haijing Smith, Thomas F Persing, David H Wright, Peter F |
| description | We developed a colorimetric microtiter plate (MTP) PCR system for specific detection of the respiratory syncytial virus (RSV) nucleocapsid gene and differentiation of viral subtypes A and B. Of 47 pediatric nasal aspirate specimens, the sensitivity and specificity were 94.4% (17 of 18) and 100% (15 of 15), respectively, when compared with RSV cell culture isolation in HEp-2 cells. An additional 14 specimens positive for adenoviruses, rhinoviruses, influenza, or parainfluenza viruses did not give positive reactions. PCR testing detected a mean of 0.15 (0.01 to 7.00) plaque-forming units of RSV virions. Inhibition of PCR amplification was detected in 33.3% (6/18) of undiluted specimens and could be avoided by a dilution (1:10) of extracted RNA without decreasing test sensitivity. RSV subtype, as determined by allele-specific probes, was identical to that determined by an immunofluorescence assay. These results indicate that the MTP PCR system provides a sensitive and specific test for clinical laboratory diagnosis and simultaneous subgroup classification of RSV infection. |
| doi_str_mv | 10.1016/S0732-8893(99)00049-8 |
| format | Article |
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Of 47 pediatric nasal aspirate specimens, the sensitivity and specificity were 94.4% (17 of 18) and 100% (15 of 15), respectively, when compared with RSV cell culture isolation in HEp-2 cells. An additional 14 specimens positive for adenoviruses, rhinoviruses, influenza, or parainfluenza viruses did not give positive reactions. PCR testing detected a mean of 0.15 (0.01 to 7.00) plaque-forming units of RSV virions. Inhibition of PCR amplification was detected in 33.3% (6/18) of undiluted specimens and could be avoided by a dilution (1:10) of extracted RNA without decreasing test sensitivity. RSV subtype, as determined by allele-specific probes, was identical to that determined by an immunofluorescence assay. These results indicate that the MTP PCR system provides a sensitive and specific test for clinical laboratory diagnosis and simultaneous subgroup classification of RSV infection.</description><identifier>ISSN: 0732-8893</identifier><identifier>EISSN: 1879-0070</identifier><identifier>DOI: 10.1016/S0732-8893(99)00049-8</identifier><identifier>PMID: 10459487</identifier><identifier>CODEN: DMIDDZ</identifier><language>eng</language><publisher>New York, NY: Elsevier Inc</publisher><subject>Biological and medical sciences ; Cell Culture Techniques - methods ; Child ; Child, Preschool ; Colorimetry - methods ; Efficiency ; Enzyme-Linked Immunosorbent Assay - methods ; Fundamental and applied biological sciences. 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Of 47 pediatric nasal aspirate specimens, the sensitivity and specificity were 94.4% (17 of 18) and 100% (15 of 15), respectively, when compared with RSV cell culture isolation in HEp-2 cells. An additional 14 specimens positive for adenoviruses, rhinoviruses, influenza, or parainfluenza viruses did not give positive reactions. PCR testing detected a mean of 0.15 (0.01 to 7.00) plaque-forming units of RSV virions. Inhibition of PCR amplification was detected in 33.3% (6/18) of undiluted specimens and could be avoided by a dilution (1:10) of extracted RNA without decreasing test sensitivity. RSV subtype, as determined by allele-specific probes, was identical to that determined by an immunofluorescence assay. These results indicate that the MTP PCR system provides a sensitive and specific test for clinical laboratory diagnosis and simultaneous subgroup classification of RSV infection.</description><subject>Biological and medical sciences</subject><subject>Cell Culture Techniques - methods</subject><subject>Child</subject><subject>Child, Preschool</subject><subject>Colorimetry - methods</subject><subject>Efficiency</subject><subject>Enzyme-Linked Immunosorbent Assay - methods</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Human viral diseases</subject><subject>Humans</subject><subject>Infant</subject><subject>Infant, Newborn</subject><subject>Infectious diseases</subject><subject>Medical sciences</subject><subject>Microbiology</subject><subject>Nasal Lavage Fluid - virology</subject><subject>Respiratory Syncytial Virus Infections - diagnosis</subject><subject>Respiratory Syncytial Viruses - classification</subject><subject>Respiratory Syncytial Viruses - isolation & purification</subject><subject>Reverse Transcriptase Polymerase Chain Reaction - methods</subject><subject>Reverse Transcriptase Polymerase Chain Reaction - standards</subject><subject>Sensitivity and Specificity</subject><subject>Techniques used in virology</subject><subject>Viral diseases</subject><subject>Viral diseases of the respiratory system and ent viral diseases</subject><subject>Virology</subject><issn>0732-8893</issn><issn>1879-0070</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1999</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkUmPFCEUgInROD2jP0HDwZjxUPooqho4mbbjlkyicTkTCh4JpjaBmqTu_nCZro5688TyvrfwQcgTBi8ZsP2rryB4XUmp-LVSLwCgUZW8R3ZMClUBCLhPdn-QC3KZ0g8AVqsGHpILBk2rGil25NeB2qmfYhgwx2DpEGyccsgY6dybjPTz8QtNa8o4UIcZbU40YppDNHmKawmNds3B9PQ2xCXRMNLRpHI0G4OJmtFRF5ItPcJ4uklLl9e5bA6n4JtH5IE3fcLH5_WKfH_39tvxQ3Xz6f3H4-GmslxBrlroOsGdl1wwwxWXruG1A--5cLyWHoUE5bhF4KorT_TCg-V72HdMeMc6fkWeb3XnOP1cMGU9lLmw782I05L0XinJ6rYtYLuBRUZKEb2ey_QmrpqBvtOvT_r1nVutlD7p17LkPT03WLoB3T9Zm-8CPDsDJlnT-2hGG9JfTrFWclaw1xuGxcZtwKiTDThadCGWL9BuCv-Z5DecgaQA</recordid><startdate>19990801</startdate><enddate>19990801</enddate><creator>Tang, Yi-Wei</creator><creator>Heimgartner, Paul J</creator><creator>Tollefson, Sharon J</creator><creator>Berg, Timothy J</creator><creator>Rys, Paul N</creator><creator>Li, Haijing</creator><creator>Smith, Thomas F</creator><creator>Persing, David H</creator><creator>Wright, Peter F</creator><general>Elsevier Inc</general><general>Elsevier</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope></search><sort><creationdate>19990801</creationdate><title>A colorimetric microtiter plate PCR system detects respiratory syncytial virus in nasal aspirates and discriminates subtypes A and B</title><author>Tang, Yi-Wei ; Heimgartner, Paul J ; Tollefson, Sharon J ; Berg, Timothy J ; Rys, Paul N ; Li, Haijing ; Smith, Thomas F ; Persing, David H ; Wright, Peter F</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c390t-50bb73df8371a3938d432d0ff37d328fe7809d3ce039b104f7f0c3606b17fd1b3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1999</creationdate><topic>Biological and medical sciences</topic><topic>Cell Culture Techniques - methods</topic><topic>Child</topic><topic>Child, Preschool</topic><topic>Colorimetry - methods</topic><topic>Efficiency</topic><topic>Enzyme-Linked Immunosorbent Assay - methods</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Human viral diseases</topic><topic>Humans</topic><topic>Infant</topic><topic>Infant, Newborn</topic><topic>Infectious diseases</topic><topic>Medical sciences</topic><topic>Microbiology</topic><topic>Nasal Lavage Fluid - virology</topic><topic>Respiratory Syncytial Virus Infections - diagnosis</topic><topic>Respiratory Syncytial Viruses - classification</topic><topic>Respiratory Syncytial Viruses - isolation & purification</topic><topic>Reverse Transcriptase Polymerase Chain Reaction - methods</topic><topic>Reverse Transcriptase Polymerase Chain Reaction - standards</topic><topic>Sensitivity and Specificity</topic><topic>Techniques used in virology</topic><topic>Viral diseases</topic><topic>Viral diseases of the respiratory system and ent viral diseases</topic><topic>Virology</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Tang, Yi-Wei</creatorcontrib><creatorcontrib>Heimgartner, Paul J</creatorcontrib><creatorcontrib>Tollefson, Sharon J</creatorcontrib><creatorcontrib>Berg, Timothy J</creatorcontrib><creatorcontrib>Rys, Paul N</creatorcontrib><creatorcontrib>Li, Haijing</creatorcontrib><creatorcontrib>Smith, Thomas F</creatorcontrib><creatorcontrib>Persing, David H</creatorcontrib><creatorcontrib>Wright, Peter F</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>Diagnostic microbiology and infectious disease</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Tang, Yi-Wei</au><au>Heimgartner, Paul J</au><au>Tollefson, Sharon J</au><au>Berg, Timothy J</au><au>Rys, Paul N</au><au>Li, Haijing</au><au>Smith, Thomas F</au><au>Persing, David H</au><au>Wright, Peter F</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>A colorimetric microtiter plate PCR system detects respiratory syncytial virus in nasal aspirates and discriminates subtypes A and B</atitle><jtitle>Diagnostic microbiology and infectious disease</jtitle><addtitle>Diagn Microbiol Infect Dis</addtitle><date>1999-08-01</date><risdate>1999</risdate><volume>34</volume><issue>4</issue><spage>333</spage><epage>337</epage><pages>333-337</pages><issn>0732-8893</issn><eissn>1879-0070</eissn><coden>DMIDDZ</coden><abstract>We developed a colorimetric microtiter plate (MTP) PCR system for specific detection of the respiratory syncytial virus (RSV) nucleocapsid gene and differentiation of viral subtypes A and B. Of 47 pediatric nasal aspirate specimens, the sensitivity and specificity were 94.4% (17 of 18) and 100% (15 of 15), respectively, when compared with RSV cell culture isolation in HEp-2 cells. An additional 14 specimens positive for adenoviruses, rhinoviruses, influenza, or parainfluenza viruses did not give positive reactions. PCR testing detected a mean of 0.15 (0.01 to 7.00) plaque-forming units of RSV virions. Inhibition of PCR amplification was detected in 33.3% (6/18) of undiluted specimens and could be avoided by a dilution (1:10) of extracted RNA without decreasing test sensitivity. RSV subtype, as determined by allele-specific probes, was identical to that determined by an immunofluorescence assay. 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| subjects | Biological and medical sciences Cell Culture Techniques - methods Child Child, Preschool Colorimetry - methods Efficiency Enzyme-Linked Immunosorbent Assay - methods Fundamental and applied biological sciences. Psychology Human viral diseases Humans Infant Infant, Newborn Infectious diseases Medical sciences Microbiology Nasal Lavage Fluid - virology Respiratory Syncytial Virus Infections - diagnosis Respiratory Syncytial Viruses - classification Respiratory Syncytial Viruses - isolation & purification Reverse Transcriptase Polymerase Chain Reaction - methods Reverse Transcriptase Polymerase Chain Reaction - standards Sensitivity and Specificity Techniques used in virology Viral diseases Viral diseases of the respiratory system and ent viral diseases Virology |
| title | A colorimetric microtiter plate PCR system detects respiratory syncytial virus in nasal aspirates and discriminates subtypes A and B |
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