title: Diagnostic validation of three test methods for detection of cyprinid herpesvirus 3 (CyHV-3).
abstract: Cyprinid herpesvirus 3 (CyHV-3) is the aetiological agent of koi herpesvirus disease in koi and common carp. The disease is notifiable to the World Organisation for Animal Health. Three tests-quantitative polymerase chain reaction (qPCR), conventional PCR (cPCR) and virus isolation by cell culture (VI)-were validated to assess their fitness as diagnostic tools for detection of CyHV-3. Test performance metrics of diagnostic accuracy were sensitivity (DSe) and specificity (DSp). Repeatability and reproducibility were measured to assess diagnostic precision. Estimates of test accuracy, in the absence of a gold standard reference test, were generated using latent class models. Test samples originated from wild common carp naturally exposed to CyHV-3 or domesticated koi either virus free or experimentally infected with the virus. Three laboratories in Canada participated in the precision study. Moderate to high repeatability (81 to 99%) and reproducibility (72 to 97%) were observed for the qPCR and cPCR tests. The lack of agreement observed between some of the PCR test pair results was attributed to cross-contamination of samples with CyHV-3 nucleic acid. Accuracy estimates for the PCR tests were 99% for DSe and 93% for DSp. Poor precision was observed for the VI test (4 to 95%). Accuracy estimates for VI/qPCR were 90% for DSe and 88% for DSp. Collectively, the results show that the CyHV-3 qPCR test is a suitable tool for surveillance, presumptive diagnosis and certification of individuals or populations as CyHV-3 free.
authors.affiliation: Fisheries and Oceans Canada, Freshwater Institute, 501 University Crescent, Winnipeg, Manitoba R3T 2N6, Canada.
authors.forename: Sharon C
authors.initials: SC
authors.lastname: Clouthier
authors.forename: Carol
authors.initials: C
authors.lastname: McClure
authors.forename: Tamara
authors.initials: T
authors.lastname: Schroeder
authors.forename: Megan
authors.initials: M
authors.lastname: Desai
authors.forename: Laura
authors.initials: L
authors.lastname: Hawley
authors.forename: Sunita
authors.initials: S
authors.lastname: Khatkar
authors.forename: Melissa
authors.initials: M
authors.lastname: Lindsay
authors.forename: Geoff
authors.initials: G
authors.lastname: Lowe
authors.forename: Jon
authors.initials: J
authors.lastname: Richard
authors.forename: Eric D
authors.initials: ED
authors.lastname: Anderson
mesh_terms: D000818:Animals; D003530:Cyprinidae; D004279:DNA, Viral / Q000235:genetics; D005393:Fish Diseases / Q000175:diagnosis* / Q000821:virology; D006564:Herpesviridae / Q000145:classification* / Q000302:isolation & purification*; D006566:Herpesviridae Infections / Q000175:diagnosis / Q000662:veterinary* / Q000821:virology; D010957:Plasmids; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D004279:DNA, Viral
keywords: Accuracy; CyHV-3; Diagnostic validation; Precision; Quantitative PCR; Virus isolation