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

• Main Authors: Valentina Kolodkina, Vladimir Martinov, Andrey Babenko
• Format: Article
• Language: English
• Published: Tehran University of Medical Sciences 2014-06-01
• Series: Iranian Journal of Microbiology
• Subjects: Bordetella parapertussis; Bordetella pertussis; Multiplex real-time PCR; diagnosis
• Online Access: https://ijm.tums.ac.ir/index.php/ijm/article/view/429
• ISSN: 2008-3289; 2008-4447

Background and Objective: Rapid diagnosis of pertussis is important for the timely isolation of the infection source and early prevention measures among the contact persons, especially among non-vaccinated infants for whom pertussis is life- threatening. Materials and Methods: Targets IS481, IS1001, BP0026 and human GAPDH gene were used to develop a multiplex real- time PCR assay based on the TaqMan technology for detection and identification of Bordetella pertussis and Bordetella parapertussis in clinical samples. A total of 121 human clinical specimens obtained within 2012-2013 were used to evaluate the multiplex real-time PCR assay. Clinical specimens were also tested for culture and conventional PCR. Sensitivity and specificity for culture, conventional PCR, and multiplex real-time PCR were measured in comparison with a clinical standard for B. pertussis infection. Results: The lower limit of detection (LLOD) of the multiplex assay was similar to the LLOD of each target in an individual assay format, which was approximately 1 genomic equivalent per reaction for IS481, IS1001 and 10 genomic equivalents per reaction for BP0026 target. When the B. pertussis assays were compared with a clinical standard for B. pertussis infection, sensitivity was 5, 59 and 89% the specificity was 100, 100 and 100% for culture, conventional PCR, and multiplex real-time PCR, respectively. Conclusions: Developed multiplex real-time PCR offers a fast tool with high sensitivity and specificity for the diagnosis of B. pertussis and B. parapertussis infections which is suitable for implementation in a routine laboratory diagnostics.