title: Development of real-time PCR assay for differential detection of Bordetella bronchiseptica and Bordetella parapertussis.
abstract: Bordetella parapertussis is a causative agent of whooping cough in humans, and B. bronchiseptica is causing wide variety of respiratory infections in mammals, including humans. Specific diagnostic tests are not currently available. Our first objective was to develop a real-time PCR test for the specific detection of B. bronchiseptica based on the previously described end-point PCR, targeting an intergenomic sequence of the fla gene locus, but it has not been reached. However, there is cross-reactivity between B. parapertussis and B. bronchiseptica. Therefore, the targeted region of several clinical isolates of both species was sequenced, and alignment of the sequences allowed the development of a 2-step real-time PCR assay. The first PCR assay detected the DNA of all clinical isolates of both B. bronchiseptica and B. parapertussis tested. The second PCR assay detected only the DNA of B. parapertussis clinical isolates, thereby allowing discrimination between B. parapertussis and B. bronchiseptica.
authors: Institute of Chemical Technology, Department of Biochemistry and Microbiology, Technicka 5, 166 28 Prague 6, Czech Republic; Institut Pasteur, Unité Prévention et Thérapie Moléculaires des Maladies Humaines, 25 rue du Dr Roux, 75015 Paris, France; Centre National de la Recherche Scientifique (CNRS) -URA 3012, Paris, France.
authors: Anette
authors: A
authors: Tizolova
authors: Institut Pasteur, Unité Prévention et Thérapie Moléculaires des Maladies Humaines, 25 rue du Dr Roux, 75015 Paris, France; Centre National de la Recherche Scientifique (CNRS) -URA 3012, Paris, France.
authors: Delphine
authors: D
authors: Brun
authors: Institut Pasteur, Unité Prévention et Thérapie Moléculaires des Maladies Humaines, 25 rue du Dr Roux, 75015 Paris, France; Centre National de la Recherche Scientifique (CNRS) -URA 3012, Paris, France.
authors: Nicole
authors: N
authors: Guiso
authors: Institut Pasteur, Unité Prévention et Thérapie Moléculaires des Maladies Humaines, 25 rue du Dr Roux, 75015 Paris, France; Centre National de la Recherche Scientifique (CNRS) -URA 3012, Paris, France. Electronic address: sophie.guillot@pasteur.fr.
authors: Sophie
authors: S
authors: Guillot
mesh_terms: D001885:Bordetella Infections / Q000175:diagnosis* / Q000382:microbiology; D016950:Bordetella bronchiseptica / Q000235:genetics / Q000302:isolation & purification*; D042483:Bordetella parapertussis / Q000235:genetics / Q000302:isolation & purification*; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D003937:Diagnosis, Differential; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D008969:Molecular Sequence Data; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D017422:Sequence Analysis, DNA
chemical_list: D004269:DNA, Bacterial
keywords: Bordetella bronchiseptica; Bordetella parapertussis; Diagnosis; Real-time PCR