title: Simultaneous detection of three fish rhabdoviruses using multiplex real-time quantitative RT-PCR assay.
abstract: Spring viremia of carp virus (SVCV), infectious hematopoietic necrosis virus (IHNV) and viral hemorrhagic septicemia virus (VHSV) are three important fish rhabdoviruses, causing serious Office International des Epizooties (OIE) classified diseases in wild and farmed fish. Here, a new multiplex real-time quantitative RT-PCR (mqRT-PCR) assay was developed for simultaneous detection, identification and quantification of these three rhabdoviruses. The sets of primers and probes were targeted to conserved regions of glycoprotein (G) gene of SVCV, nucleoprotein (N) gene of IHNV and G gene of VHSV and used to amplify. The sensitivity, specificity and interference test of mqRT-PCR assay was analyzed. It was shown that the detection levels of 100 copies of SVCV, 220 copies of IHNV and 140 copies of VHSV were achieved, and there was no non-specific amplification and cross-reactivity using RNA of pike fry rhabdovirus (PFRV), infectious pancreatic necrosis virus (IPNV) and grass carp reovirus (GCRV). A total of 80 clinical fish samples were tested using the mqRT-PCR assay and the results were confirmed by antigen-capture ELISA and cell culture assay. This assay has the potential to be used for both research applications and diagnosis.
authors.affiliation: College of Fisheries, Huazhong Agricultural University, Wuhan 430070, PR China; Show-how Bio-tech Institute, Beihai 536000, PR China.
authors.forename: Zongxiao
authors.initials: Z
authors.lastname: Liu
authors.forename: Yong
authors.initials: Y
authors.lastname: Teng
authors.forename: Hong
authors.initials: H
authors.lastname: Liu
authors.forename: Yulin
authors.initials: Y
authors.lastname: Jiang
authors.forename: Xiayang
authors.initials: X
authors.lastname: Xie
authors.forename: Huifang
authors.initials: H
authors.lastname: Li
authors.forename: Jiangqiang
authors.initials: J
authors.lastname: Lv
authors.forename: Longying
authors.initials: L
authors.lastname: Gao
authors.forename: Junqiang
authors.initials: J
authors.lastname: He
authors.forename: Xiujie
authors.initials: X
authors.lastname: Shi
authors.forename: Feiyan
authors.initials: F
authors.lastname: Tian
authors.forename: Jingshun
authors.initials: J
authors.lastname: Yang
authors.forename: Congxin
authors.initials: C
authors.lastname: Xie
mesh_terms: D000818:Animals; D002460:Cell Line; D005393:Fish Diseases / Q000175:diagnosis / Q000821:virology; D005399:Fishes / Q000821:virology; D029142:Infectious hematopoietic necrosis virus / Q000302:isolation & purification*; D029141:Novirhabdovirus / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D018353:Rhabdoviridae Infections / Q000175:diagnosis / Q000662:veterinary / Q000821:virology; D012680:Sensitivity and Specificity; D018116:Vesiculovirus / Q000302:isolation & purification*