NDLI: Multiplex LightCycler PCR Assay for Detection and Differentiation of Bordetella pertussis and Bordetella parapertussis in Nasopharyngeal Specimens

Content Provider	PubMed Central
Author	Sloan, Lynne M. Hopkins, Marlene K. Mitchell, P. Shawn Vetter, Emily A. Rosenblatt, Jon E. Harmsen, W. Scott Cockerill, Franklin R. Patel, Robin
Copyright Year	2002
Abstract	A rapid real-time multiplex PCR assay for detecting and differentiating Bordetella pertussis and Bordetella parapertussis in nasopharyngeal swabs was developed. This assay (LC-PCR-IS) targets the insertion sequences IS481 and IS1001 of B. pertussis and B. parapertussis, respectively, and is performed using the LightCycler (Roche Molecular Biochemicals, Indianapolis, Ind.). The analytical sensitivity is less than one organism per reaction. Results for Bordetella culture and/or direct fluorescent antibody testing and a second LightCycler PCR assay (target, pertussis toxin gene) were compared to results of the LC-PCR-IS assay for 111 nasopharyngeal swabs submitted for pertussis testing. Of the specimens, 12 were positive (9 B. pertussis and 3 B. parapertussis) and 68 specimens were negative by all methods. Three other specimens were positive for B. pertussis by at least two of the methods (including the LC-PCR-IS assay), and another 28 specimens were positive for B. pertussis by the LC-PCR-IS assay only. No specimens were negative by the LC-PCR-IS assay and positive by the other methods. A conventional PCR method (target, IS481) was also compared to the LC-PCR-IS assay for a different group of nasopharyngeal swab specimens (n = 96): 44 specimens were positive and 41 specimens were negative for B. pertussis with both PCR methods. Nine specimens were positive for B. pertussis by the LC-PCR-IS assay and negative by the conventional PCR assay, and two specimens were positive for B. pertussis by the conventional PCR assay and negative by the LC-PCR-IS assay. Positivity of the two assays was not significantly different (P = 0.0654). The insertion sequence IS481 is also present in Bordetella holmesii; specimens containing B. holmesii may yield false-positive results. The LC-PCR-IS assay takes approximately 45 min to complete post-nucleic acid extraction, compared to 24 h for the conventional PCR assay previously used in our laboratory. The LC-PCR-IS assay is easier to perform than the conventional PCR assay, and the closed system decreases the chance of contamination. All of these characteristics represent a significant improvement in the detection of B. pertussis and B. parapertussis in nasopharyngeal specimens.
Related Links	http://dx.doi.org/10.1128/jcm.40.1.96-100.2002
Ending Page	100
Page Count	5
Starting Page	96
File Format	PDF
ISSN	00951137
e-ISSN	1098660X
Journal	Journal of Clinical Microbiology
Issue Number	1
Volume Number	40
Language	English
Publisher	American Society for Microbiology
Publisher Date	2002-01-01
Access Restriction	Open
Rights Holder	American Society for Microbiology
Subject Keyword	Microbiology (medical) Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Microbiology (medical)

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Multiplex real-time PCR assay for detection and differentiation of Bordetella pertussis and Bordetella parapertussis

Multiplex PCR-based assay for detection of Bordetella pertussis in nasopharyngeal swab specimens.

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Multiplex LightCycler PCR Assay for Detection and Differentiation of Bordetella pertussis and Bordetella parapertussis in Nasopharyngeal Specimens

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