title: Development and use of a multiplex real-time quantitative polymerase chain reaction assay for detection and differentiation of Porcine circovirus-2 genotypes 2a and 2b in an epidemiological survey.
abstract: By the end of 2004, the Canadian swine population had experienced a severe increase in the incidence of Porcine circovirus-associated disease (PCVAD), a problem that was associated with the emergence of a new Porcine circovirus-2 genotype (PCV-2b), previously unrecovered in North America. Thus, it became important to develop a diagnostic tool that could differentiate between the old and new circulating genotypes (PCV-2a and PCV-2b, respectively). Consequently, a multiplex real-time quantitative polymerase chain reaction (mrtqPCR) assay that could sensitively and specifically identify and differentiate PCV-2 genotypes was developed. A retrospective epidemiologic survey that used the mrtqPCR assay was performed to determine if cofactors could affect the risk of PCVAD. From 121 PCV-2-positive cases gathered for this study, 4.13%, 92.56%, and 3.31% were positive for PCV-2a, PCV-2b, and both genotypes, respectively. In a data analysis using univariate logistic regressions, the PCVAD-compatible (PCVAD/c) score was significantly associated with the presence of Porcine reproductive and respiratory syndrome virus (PRRSV), PRRSV viral load, PCV-2 viral load, and PCV-2 immunohistochemistry (IHC) results. Polytomous logistic regression analysis revealed that PCVAD/c score was affected by PCV-2 viral load (P = 0.0161) and IHC (P = 0.0128), but not by the PRRSV variables (P > 0.9), which suggests that mrtqPCR in tissue is a reliable alternative to IHC. Logistic regression analyses revealed that PCV-2 increased the odds ratio of isolating 2 major swine pathogens of the respiratory tract, Actinobacillus pleuropneumoniae and Streptococcus suis serotypes 1/2, 1, 2, 3, 4, and 7, which are serotypes commonly associated with clinical diseases.
authors.affiliation: Faculté de médecine vétérinaire, Université de Montréal, 3200 rue Sicotte, St-Hyacinthe, Québec, Canada. carl.a.gagnon@umontreal.ca
authors.forename: Carl A
authors.initials: CA
authors.lastname: Gagnon
authors.forename: Jérome R E
authors.initials: JR
authors.lastname: del Castillo
authors.forename: Nedzad
authors.initials: N
authors.lastname: Music
authors.forename: Guy
authors.initials: G
authors.lastname: Fontaine
authors.forename: Josée
authors.initials: J
authors.lastname: Harel
authors.forename: Donald
authors.initials: D
authors.lastname: Tremblay
mesh_terms: D000818:Animals; D018173:Circoviridae Infections / Q000662:veterinary; D017925:Circovirus / Q000145:classification / Q000235:genetics*; D017931:DNA Primers; D005838:Genotype; D010957:Plasmids; D016133:Polymerase Chain Reaction / Q000379:methods*; D011792:Quebec; D012680:Sensitivity and Specificity; D013552:Swine; D013553:Swine Diseases / Q000453:epidemiology / Q000821:virology; D019282:Wasting Syndrome / Q000453:epidemiology / Q000662:veterinary / Q000821:virology
chemical_list: D017931:DNA Primers