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Diseases of Aquatic Organisms

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DAO 139:131-137 (2020)  -  DOI: https://doi.org/10.3354/dao03484

A sensitive and specific SYBR Green-based qPCR assay for detecting scale drop disease virus (SDDV) in Asian sea bass

Sukhontip Sriisan1,2, Chuenchit Boonchird1, Siripong Thitamadee1,2, Molruedee Sonthi3,4, Ha Thanh Dong5, Saengchan Senapin1,2,6,*

1Department of Biotechnology, Faculty of Science, Mahidol University, Bangkok 10400, Thailand
2Center of Excellence for Shrimp Molecular Biology and Biotechnology (Centex Shrimp), Faculty of Science, Mahidol University, Bangkok 10400, Thailand
3Faculty of Marine Technology, Burapha University Chanthaburi Campus, Chanthaburi 22170, Thailand
4Aquatic Animal Disease Diagnostics and Immunology Research Unit, Burapha University Chanthaburi Campus, Chanthaburi 22170, Thailand
5Faculty of Science and Technology, Suan Sunandha Rajabhat University, Bangkok 10300, Thailand
6National Center for Genetic Engineering and Biotechnology (BIOTEC), National Science and Technology Development Agency (NSTDA), Pathum Thani 12120, Thailand
*Corresponding author:

ABSTRACT: Scale drop disease virus (SDDV) is a megalocytivirus known to cause disease in Asian sea bass in Southeast Asia. To support SDDV diagnosis and surveillance, we report on a sensitive and specific SYBR Green qPCR assay. The qPCR primers were designed to target a 135 bp fragment of the SDDV ATPase gene. The optimized SDDV qPCR assay reliably detected 2 copies of a plasmid dsDNA control and did not cross-amplify DNA to any of 12 viral or bacterial pathogens commonly found in aquatic animals. When assessed with 86 field samples, the assay detected SDDV in DNA extracted from each of 34 scale drop disease-affected fish collected from 5 affected farms. The qPCR also detected SDDV in DNA from 30 of 52 overtly healthy fish collected from 9 farms where SDDV had not been detected previously, using a semi-nested conventional PCR. The higher sensitivity of our SDDV qPCR assay can thus be useful in detecting fish with subclinical/chronic infections. However, the qPCR showed that SDDV DNA loads varied from 8.0 × 102 to 6.8 × 104 viral DNA copies per 200 ng DNA template among the 8 organ tissue types sampled from 3 diseased fish. In circumstances requiring SDDV to be detected unequivocally in subclinical carriers with lower-level infection, qPCR testing of more than one type of tissue is advisable.


KEY WORDS: ATPase · Scale drop disease · Lates calcarifer · SDDV · qPCR


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Cite this article as: Sriisan S, Boonchird C, Thitamadee S, Sonthi M, Thanh Dong H, Senapin S (2020) A sensitive and specific SYBR Green-based qPCR assay for detecting scale drop disease virus (SDDV) in Asian sea bass. Dis Aquat Org 139:131-137. https://doi.org/10.3354/dao03484

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