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.

中文翻译：

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一种灵敏且基于SYBR Green的qPCR分析方法，用于检测亚洲鲈鱼中的鳞屑病病毒（SDDV）。

摘要：鳞屑病病毒（SDDV）是一种巨轮病毒，已知会引起东南亚亚洲鲈鱼的疾病。为了支持SDDV的诊断和监测，我们报告了一种灵敏且特异的SYBR Green qPCR检测方法。设计qPCR引物以靶向SDDV ATPase的135 bp片段基因。优化的SDDV qPCR分析能够可靠地检测到质粒dsDNA对照的2个拷贝，并且不会将DNA交叉扩增到水生动物中常见的12种病毒或细菌病原体中的任何一种。当对86个田间样品进行评估时，该检测方法从从5个受影响的农场收集的34种受疾病影响的受鳞鳞下降的鱼类的每条鱼中提取的DNA中检测到SDDV。qPCR还使用半巢式常规PCR检测了9个养殖场收集的52条健康鱼的DNA中的SDDV，这些鱼以前从未检测到SDDV。因此，我们SDDV qPCR分析的更高灵敏度可用于检测亚临床/慢性感染的鱼类。然而，qPCR显示SDDV DNA的负载范围从8.0×10 ²到6.8×10 ⁴从3种患病鱼中取样的8种器官组织类型中，每200 ng DNA模板的病毒DNA拷贝数。在需要在感染较轻的亚临床载体中明确检测SDDV的情况下，建议对一种以上类型的组织进行qPCR检测。