title: Development of a real-time duplex TaqMan-PCR for the detection of Equine rhinitis A and B viruses in clinical specimens.
abstract: Equine rhinitis A and B viruses (ERAV and ERBV) are respiratory viruses of horses belonging to the family Picornaviridae. Although these viruses are considered to cause respiratory disease in horses and are potentially infectious for humans, little is known about their prevalence and pathogenesis. Virus isolation is often unsuccessful due to their inefficient growth and lack of cytopathic effect in cell cultures. Therefore, molecular assays should be considered as the method of choice to detect infection in symptomatic or apparently healthy horses. In the present study, a novel real-time duplex PCR was developed for the detection and differentiation of both ERAV and ERBV. The method was evaluated for its ability to detect viral RNA in cell culture supernatants and nasal swabs, and lung and urine spiked with known quantities of virus. The assay demonstrated high analytical specificity, sensitivity and good reproducibility, with coefficients of variation (CV%) ranging from 1% to 7.4% and from 1.2% to 12% for intra- and inter-assay variability respectively. The assay detected ERBV in 14 of 86 nasal swabs collected from horses with respiratory disease. The real-time duplex PCR is a useful new diagnostic method for the rapid detection and differentiation of ERAV and ERBV.
authors.affiliation: Istituto Zooprofilattico Sperimentale delle Venezie, Research & Development Department, Viale dell'Universita' 10, 35020 Legnaro, Padova, Italy.
authors.forename: Antonio
authors.initials: A
authors.lastname: Mori
authors.forename: Paola
authors.initials: P
authors.lastname: De Benedictis
authors.forename: Sabrina
authors.initials: S
authors.lastname: Marciano
authors.forename: Bianca
authors.initials: B
authors.lastname: Zecchin
authors.forename: Antonio
authors.initials: A
authors.lastname: Zuin
authors.forename: Barbara
authors.initials: B
authors.lastname: Zecchin
authors.forename: Ilaria
authors.initials: I
authors.lastname: Capua
authors.forename: Giovanni
authors.initials: G
authors.lastname: Cattoli
mesh_terms: D000818:Animals; D017931:DNA Primers; D052698:Erbovirus / Q000235:genetics / Q000302:isolation & purification*; D006734:Horse Diseases / Q000175:diagnosis / Q000821:virology; D006736:Horses; D010849:Picornaviridae / Q000235:genetics / Q000302:isolation & purification*; D010850:Picornaviridae Infections / Q000175:diagnosis / Q000662:veterinary* / Q000821:virology; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012220:Rhinitis / Q000662:veterinary* / Q000821:virology; D012680:Sensitivity and Specificity; D019914:Taq Polymerase
chemical_list: D017931:DNA Primers; D019914:Taq Polymerase