Developement of the Isolation System and Higher Sensitivive Diagnosis Method for Koi Herpesvirus
Date Issued
2009
Date
2009
Author(s)
Liu, Hsin-Chun
Abstract
Koi herpesvirus (KHV) was categorized as a member of the Alloherpesviridae under the species name Cyprinid herpesvirus 3 (CyHV3). It causes a highly contagious disease which leads up to 80 – 100 % mortality in koi (Cyprinus carpio koi) and common carp (Cyprinus carpio carpio). KHV was found firstly in 1998, and the first case in Taiwan has been identified in December 2002. Util now, there has been no suitable cells developed for KHV virus isolation in Taiwan, which becomes an obstacle for the research. Many previous researchers suggest the possibility in latent infection of KHV; however, we lack a diagnosis method providing higher sensitivity to distinguish the healthy and the latent infected fish. n our study, the KHV isolation, we inoculated the tissue homogenates of infected fish to tested cells, observed the cytopathic effect, and then detected the KHV DNA by polymerase chain reaction. Our results revealed that only Taiwan koi fin (TKF1) cell, established by our laboratory, showed the CPE and propagatde KHV successfully. Other cells derived from koi, goldfish, fathead minnow, grouper, eel, tilapia and soft-shell turtle were not susceptible for the KHV. Moreover, we detected the ORF 81 protein of KHV by indirect immunoflourescene assay, the positive signal appeared after 12 hours post infection.or the more sensitive diagnostic method, we developed a nested polymerase chain reaction with good specificity, and it was able to detect as less as 17 virions in the sample. We have been applied the diagnosis method to detect the fish from one koi farm, which was suspected to be the KHV endemic farm. According to the history of this farm and the results of nested PCR, we suggest the high sensitivity diagnosis method has the potential to apply to detect latent infection cases.
Subjects
KHV
Nested PCR
Virus isolation
Type
thesis
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