Multiplexed reverse transcriptase PCR assay for identification of viral respiratory pathogens at the point of care

Sonia E. Létant, Josue I. Ortiz, Lance F Bentley Tammero, James M. Birch, Robert W. Derlet, Stuart H Cohen, Dannelle Manning, Mary T. McBride

Research output: Contribution to journalArticle

41 Citations (Scopus)

Abstract

We have developed a nucleic acid-based assay that is rapid, sensitive, and specific and can be used for the simultaneous detection of five common human respiratory pathogens, including influenza virus A, influenza virus B, parainfluenza virus types 1 and 3, respiratory syncytial virus (RSV), and adenovirus groups B, C, and E. Typically, diagnosis on an unextracted clinical sample can be provided in less than 3 h, including sample collection, preparation, and processing, as well as data analysis. Such a multiplexed panel would enable rapid broad-spectrum pathogen testing on nasal swabs and therefore allow implementation of infection control measures and the timely administration of antiviral therapies. We present here a summary of the assay performance in terms of sensitivity and specificity. The limits of detection are provided for each targeted respiratory pathogen, and result comparisons were performed on clinical samples, our goal being to compare the sensitivity and specificity of the multiplexed assay to the combination of immunofluorescence and shell vial culture currently implemented at the University of California-Davis Medical Center hospital. Overall, the use of the multiplexed reverse transcription-PCR assay reduced the rate of false-negative results by 4% and reduced the rate of false-positive results by up to 10%. The assay correctly identified 99.3% of the clinical negatives and 97% of the adenovirus, 95% of the RSV, 92% of the influenza virus B, and 77% of the influenza virus A samples without any extraction performed on the clinical samples. The data also showed that extraction will be needed for parainfluenza virus, which was only identified correctly 24% of the time on unextracted samples.

Original languageEnglish (US)
Pages (from-to)3498-3505
Number of pages8
JournalJournal of Clinical Microbiology
Volume45
Issue number11
DOIs
StatePublished - Nov 2007

Fingerprint

Point-of-Care Systems
Influenza A virus
Reverse Transcriptase Polymerase Chain Reaction
Respiratory Syncytial Viruses
Adenoviridae
Human parainfluenza virus 1
Human parainfluenza virus 3
Influenza B virus
Cercopithecine Herpesvirus 1
Paramyxoviridae Infections
Sensitivity and Specificity
Infection Control
Nose
Nucleic Acids
Reverse Transcription
Antiviral Agents
Fluorescent Antibody Technique
Limit of Detection
Viruses
Polymerase Chain Reaction

ASJC Scopus subject areas

  • Microbiology (medical)
  • Microbiology

Cite this

Multiplexed reverse transcriptase PCR assay for identification of viral respiratory pathogens at the point of care. / Létant, Sonia E.; Ortiz, Josue I.; Tammero, Lance F Bentley; Birch, James M.; Derlet, Robert W.; Cohen, Stuart H; Manning, Dannelle; McBride, Mary T.

In: Journal of Clinical Microbiology, Vol. 45, No. 11, 11.2007, p. 3498-3505.

Research output: Contribution to journalArticle

Létant, SE, Ortiz, JI, Tammero, LFB, Birch, JM, Derlet, RW, Cohen, SH, Manning, D & McBride, MT 2007, 'Multiplexed reverse transcriptase PCR assay for identification of viral respiratory pathogens at the point of care', Journal of Clinical Microbiology, vol. 45, no. 11, pp. 3498-3505. https://doi.org/10.1128/JCM.01712-07
Létant, Sonia E. ; Ortiz, Josue I. ; Tammero, Lance F Bentley ; Birch, James M. ; Derlet, Robert W. ; Cohen, Stuart H ; Manning, Dannelle ; McBride, Mary T. / Multiplexed reverse transcriptase PCR assay for identification of viral respiratory pathogens at the point of care. In: Journal of Clinical Microbiology. 2007 ; Vol. 45, No. 11. pp. 3498-3505.
@article{80d24cd1e1664bfd9ec4950e80ac48a3,
title = "Multiplexed reverse transcriptase PCR assay for identification of viral respiratory pathogens at the point of care",
abstract = "We have developed a nucleic acid-based assay that is rapid, sensitive, and specific and can be used for the simultaneous detection of five common human respiratory pathogens, including influenza virus A, influenza virus B, parainfluenza virus types 1 and 3, respiratory syncytial virus (RSV), and adenovirus groups B, C, and E. Typically, diagnosis on an unextracted clinical sample can be provided in less than 3 h, including sample collection, preparation, and processing, as well as data analysis. Such a multiplexed panel would enable rapid broad-spectrum pathogen testing on nasal swabs and therefore allow implementation of infection control measures and the timely administration of antiviral therapies. We present here a summary of the assay performance in terms of sensitivity and specificity. The limits of detection are provided for each targeted respiratory pathogen, and result comparisons were performed on clinical samples, our goal being to compare the sensitivity and specificity of the multiplexed assay to the combination of immunofluorescence and shell vial culture currently implemented at the University of California-Davis Medical Center hospital. Overall, the use of the multiplexed reverse transcription-PCR assay reduced the rate of false-negative results by 4{\%} and reduced the rate of false-positive results by up to 10{\%}. The assay correctly identified 99.3{\%} of the clinical negatives and 97{\%} of the adenovirus, 95{\%} of the RSV, 92{\%} of the influenza virus B, and 77{\%} of the influenza virus A samples without any extraction performed on the clinical samples. The data also showed that extraction will be needed for parainfluenza virus, which was only identified correctly 24{\%} of the time on unextracted samples.",
author = "L{\'e}tant, {Sonia E.} and Ortiz, {Josue I.} and Tammero, {Lance F Bentley} and Birch, {James M.} and Derlet, {Robert W.} and Cohen, {Stuart H} and Dannelle Manning and McBride, {Mary T.}",
year = "2007",
month = "11",
doi = "10.1128/JCM.01712-07",
language = "English (US)",
volume = "45",
pages = "3498--3505",
journal = "Journal of Clinical Microbiology",
issn = "0095-1137",
publisher = "American Society for Microbiology",
number = "11",

}

TY - JOUR

T1 - Multiplexed reverse transcriptase PCR assay for identification of viral respiratory pathogens at the point of care

AU - Létant, Sonia E.

AU - Ortiz, Josue I.

AU - Tammero, Lance F Bentley

AU - Birch, James M.

AU - Derlet, Robert W.

AU - Cohen, Stuart H

AU - Manning, Dannelle

AU - McBride, Mary T.

PY - 2007/11

Y1 - 2007/11

N2 - We have developed a nucleic acid-based assay that is rapid, sensitive, and specific and can be used for the simultaneous detection of five common human respiratory pathogens, including influenza virus A, influenza virus B, parainfluenza virus types 1 and 3, respiratory syncytial virus (RSV), and adenovirus groups B, C, and E. Typically, diagnosis on an unextracted clinical sample can be provided in less than 3 h, including sample collection, preparation, and processing, as well as data analysis. Such a multiplexed panel would enable rapid broad-spectrum pathogen testing on nasal swabs and therefore allow implementation of infection control measures and the timely administration of antiviral therapies. We present here a summary of the assay performance in terms of sensitivity and specificity. The limits of detection are provided for each targeted respiratory pathogen, and result comparisons were performed on clinical samples, our goal being to compare the sensitivity and specificity of the multiplexed assay to the combination of immunofluorescence and shell vial culture currently implemented at the University of California-Davis Medical Center hospital. Overall, the use of the multiplexed reverse transcription-PCR assay reduced the rate of false-negative results by 4% and reduced the rate of false-positive results by up to 10%. The assay correctly identified 99.3% of the clinical negatives and 97% of the adenovirus, 95% of the RSV, 92% of the influenza virus B, and 77% of the influenza virus A samples without any extraction performed on the clinical samples. The data also showed that extraction will be needed for parainfluenza virus, which was only identified correctly 24% of the time on unextracted samples.

AB - We have developed a nucleic acid-based assay that is rapid, sensitive, and specific and can be used for the simultaneous detection of five common human respiratory pathogens, including influenza virus A, influenza virus B, parainfluenza virus types 1 and 3, respiratory syncytial virus (RSV), and adenovirus groups B, C, and E. Typically, diagnosis on an unextracted clinical sample can be provided in less than 3 h, including sample collection, preparation, and processing, as well as data analysis. Such a multiplexed panel would enable rapid broad-spectrum pathogen testing on nasal swabs and therefore allow implementation of infection control measures and the timely administration of antiviral therapies. We present here a summary of the assay performance in terms of sensitivity and specificity. The limits of detection are provided for each targeted respiratory pathogen, and result comparisons were performed on clinical samples, our goal being to compare the sensitivity and specificity of the multiplexed assay to the combination of immunofluorescence and shell vial culture currently implemented at the University of California-Davis Medical Center hospital. Overall, the use of the multiplexed reverse transcription-PCR assay reduced the rate of false-negative results by 4% and reduced the rate of false-positive results by up to 10%. The assay correctly identified 99.3% of the clinical negatives and 97% of the adenovirus, 95% of the RSV, 92% of the influenza virus B, and 77% of the influenza virus A samples without any extraction performed on the clinical samples. The data also showed that extraction will be needed for parainfluenza virus, which was only identified correctly 24% of the time on unextracted samples.

UR - http://www.scopus.com/inward/record.url?scp=36348949815&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=36348949815&partnerID=8YFLogxK

U2 - 10.1128/JCM.01712-07

DO - 10.1128/JCM.01712-07

M3 - Article

C2 - 17855573

AN - SCOPUS:36348949815

VL - 45

SP - 3498

EP - 3505

JO - Journal of Clinical Microbiology

JF - Journal of Clinical Microbiology

SN - 0095-1137

IS - 11

ER -