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Diseases of Aquatic Organisms

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DAO 31:181-186 (1997)  -  doi:10.3354/dao031181

Detection of yellow-head virus (YHV) of Penaeus monodon by RT-PCR amplification

Chainarong Wongteerasupaya1, Wansika Tongchuea2, Vichai Boonsaeng3,*, Sakol Panyim3, Anchalee Tassanakajon4, Boonsirm Withyachumnarnkul5, T. W. Flegel6

1Dept Biochemistry, Faculty of Medicine, Srinakharinwirot University, Soi 23 Sukhumvit Road, Bangkok 10110, Thailand
2National Center for Genetic Engineering and Biotechnology, Rama 6 Road, Bangkok 10400, Thailand
3Dept Biochemistry, 5Dept Anatomy and 6Dept Biotechnology, Faculty of Science, Mahidol University, Rama 6 Road, Bangkok 10400, Thailand
4Dept Biochemistry, Faculty of Science, Chulalongkorn University, Phya Thai Road, Bangkok 10400, Thailand
*Addressee for correspondence. E-mail:

A nucleic acid probe was developed using cDNA prepared from ssRNA extracted from yellow-head virus (YHV), a serious pathogen of the black tiger prawn Penaeus monodon. The specificity and sensitivity of this probe was established using dot-blot hybridization with nucleic acid extracts from YHV and from shrimp, bacteria and other viruses. Based on the sequence of this cloned YHV cDNA fragment, a YHV specific primer set for reverse transcription polymerase chain reaction (RT-PCR) of a 135 base pair (bp) sub-fragment was designed for detection of YHV infections in penaeid shrimp. When applied in RT-PCR with templates derived from experimentally or naturally YHV-infected shrimp and with purified YHV or YHV nucleic acid, the expected 135 bp amplification product was obtained. By contrast, nucleic acids extracted from tissue samples of healthy shrimp and from other shrimp pathogens gave no such fragment. This confirmed the specificity of the designed YHV RNA specific primers. RT-PCR based detection demonstrated high sensitivity, in that it could detect 0.01 pg of purified YHV-RNA. In a time course study of an experimental YHV infection, the RT-PCR detection showed evidence of infection at 6 to 12 h post exposure to the virus. However, histopathology typical of YHV infection [i.e. karyorhexis and pycnosis of haemocytes in haematoxylin and eosin (H&E) stained haemolymph smears] was not visible until 42 to 48 h post exposure. The results suggested that RT-PCR might be useful to shrimp aquaculturists for early detection of YHV outbreaks or for detection of asymptomatic carriers.

Yellow-head virus (YHV) · Polymerase chain reaction (PCR) · Penaeus monodon

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