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Diseases of Aquatic Organisms

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DAO 100:11-18 (2012)  -  DOI:

Development of mRNA-specific RT-PCR for the detection of koi herpesvirus (KHV) replication stage

Kei Yuasa1,*, Jun Kurita1, Morihiko Kawana2, Ikunari Kiryu1, Norihisa Oseko1, Motohiko Sano

1National Research Institute of Aquaculture, Fisheries Research Agency, Minamiise, Mie 516-0193, Japan
2Hokkaido National Fisheries Research Institute, Fisheries Research Agency, Sapporo, Hokkaido 062-0922, Japan
3National Research Institute of Fisheries Science, Fisheries Research Agency, Yokohama, Kanagawa 236-8648, Japan

ABSTRACT: An mRNA-specific reverse transcription (RT)-PCR primer set spanning the exon junction of a spliced putative terminase gene in the koi herpesvirus (KHV) was developed to detect the replicating stage of the virus. The proposed RT-PCR amplified a target gene from the RNA template, but not from a DNA template extracted from common carp brain (CCB) cells infected with KHV. In addition, the RT-PCR did not amplify the target gene of templates extracted from specific cell lines infected with either CyHV-1 or CyHV-2. RT-PCR detected mRNA from the scales of koi experimentally infected with KHV at 24 h post exposure (hpe). However, unlike conventional PCR, RT-PCR could not detect KHV DNA in fish at 0 hpe. The results indicate that the RT-PCR developed in this study is mRNA-specific and that the assay can detect the replicating stage of KHV from both fish and cultured cells infected with the virus.

KEY WORDS: Koi herpesvirus · Cyprinid herpesvirus 3 · Terminase · mRNA · RT-PCR

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Cite this article as: Yuasa K, Kurita J, Kawana M, Kiryu I, Ohseko N, Sano M (2012) Development of mRNA-specific RT-PCR for the detection of koi herpesvirus (KHV) replication stage. Dis Aquat Org 100:11-18.

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