title: Comparative evaluation of five serological methods and RT-PCR assay for the detection of IPNV in fish.
abstract: In the present study, six diagnostic methods for the detection of infectious pancreatic necrosis virus (IPNV) (indirect immunofluorescence, flow cytometry, immunoperoxidase, immunodot blot, immunostaphylococcus-protein A, and RT-PCR) have been comparatively evaluated using the seroneutralization as the reference assay, and 83 Spanish isolates and 3 reference strains. The most reliable methods were flow cytometry and RT-PCR which could detect virus at titers of 1x10(2) and 1x10(3) TCID50/ml, respectively. At a multiplicity of infection of 50, both assays allowed the earliest detection of IPNV at 4 h post-inoculation. Indirect immunofluorescence and immunoperoxidase assays required at least 6 h post-inoculation to detect viral antigens. The immunodot blot assay possesses low sensitivity and the immunostaphylococcus-protein A test cannot be applied for routine examination of IPNV. Positive reactions were obtained in 100% of the samples tested by seroneutralization and RT-PCR, 90.4% by the flow cytometry, 80.7% by the indirect immunofluorescence assay, 67.5% by the immunoperoxidase, 62.6% by the immunodot blot, and only 27.7% by immunostaphylococcus-protein A test. Therefore, RT-PCR and flow cytometry were the most appropriate and sensitive methods for the routine detection of IPNV from affected fish.
authors.affiliation: Centro de Investigaciones Biológicas, C.S.I.C, Velázquez, 144, 28071-, Madrid, Spain.
authors.forename: S
authors.initials: S
authors.lastname: Rodriguez Saint-Jean
authors.forename: J J
authors.initials: JJ
authors.lastname: Borrego
authors.forename: S I
authors.initials: SI
authors.lastname: Perez-Prieto
mesh_terms: D000818:Animals; D018175:Birnaviridae Infections / Q000175:diagnosis / Q000662:veterinary / Q000821:virology; D002478:Cells, Cultured; D005393:Fish Diseases / Q000175:diagnosis / Q000821:virology; D005434:Flow Cytometry; D018061:Infectious pancreatic necrosis virus / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D012367:RNA, Viral / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012474:Salmon / Q000821:virology*; D012680:Sensitivity and Specificity; D012703:Serotyping
chemical_list: D012367:RNA, Viral