title: A multiplex real-time PCR assay for the detection and differentiation of Francisella tularensis subspecies.
abstract: Francisella tularensis is the aetiological agent of tularaemia, a zoonotic disease with worldwide prevalence. F. tularensis is a highly pathogenic organism and has been designated a category A biothreat agent by the Centers for Disease Control and Prevention. Tularaemia is endemic in much of the USA, Europe and parts of Asia. It is transmitted by numerous vectors and vehicles such as deer flies, ticks and rabbits. Currently, there are four recognized subspecies of F. tularensis: tularensis (type A), holarctica (type B), mediasiatica and novicida. Within the type A classification there are two subclassifications, type A.I and A.II, each with a specific geographical distribution across the USA. F. tularensis subsp. holartica (type B) is found in both the USA and Europe. Because of virulence differences among subtypes, it is important that health departments, hospitals and other government agencies are able to quickly identify each subtype. The purpose of this study was to develop a multiplex real-time PCR assay for the identification and discrimination of type A.I, type A.II, type B and novicida subspecies of F. tularensis. The assay was validated using 119 isolates of F. tularensis, three of its nearest neighbours and 14 other bacterial pathogens. This assay proved to be ~98 % successful at identifying the known subspecies of F. tularensis and could prove to be a useful tool in the characterization of this important pathogen.
authors.affiliation: Microbiology Branch, Life Sciences Division, Dugway Proving Ground, Dugway, UT 84022, USA.|Department of Microbiology and Molecular Biology, Brigham Young University, Provo, UT 84602, USA.
authors.forename: Mark K
authors.initials: MK
authors.lastname: Gunnell
authors.affiliation: Department of Microbiology and Molecular Biology, Brigham Young University, Provo, UT 84602, USA.
authors.forename: Charity D
authors.initials: CD
authors.lastname: Lovelace
authors.affiliation: Department of Microbiology and Molecular Biology, Brigham Young University, Provo, UT 84602, USA.
authors.forename: Benjamin A
authors.initials: BA
authors.lastname: Satterfield
authors.affiliation: Department of Microbiology and Molecular Biology, Brigham Young University, Provo, UT 84602, USA.
authors.forename: Emily A
authors.initials: EA
authors.lastname: Moore
authors.affiliation: Department of Microbiology and Molecular Biology, Brigham Young University, Provo, UT 84602, USA.
authors.forename: Kim L
authors.initials: KL
authors.lastname: O'Neill
authors.affiliation: Department of Microbiology and Molecular Biology, Brigham Young University, Provo, UT 84602, USA.
authors.forename: Richard A
authors.initials: RA
authors.lastname: Robison
mesh_terms: D000818:Animals; D001431:Bacteriological Techniques; D023482:Bioterrorism; D004269:DNA, Bacterial / Q000235:genetics; D019353:Endemic Diseases; D005604:Francisella tularensis / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification* / Q000472:pathogenicity; D005819:Genetic Markers; D016680:Genome, Bacterial; D006801:Humans; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013045:Species Specificity; D014406:Tularemia / Q000453:epidemiology / Q000382:microbiology*; D014774:Virulence
chemical_list: D004269:DNA, Bacterial; D005819:Genetic Markers