Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China.
Key Laboratory of Protection & Utilization of Biological Resources in Tarim Basin, College of Life Sciences, Tarim University, Alar 843399, China.
Viruses. 2023 May 25;15(6):1238. doi: 10.3390/v15061238.
Porcine viral diarrhea is very common in clinical practice and has caused huge losses to the pig industry. Porcine epidemic diarrhea virus (PEDV), porcine rotavirus (PoRV), and porcine deltacoronavirus (PDCoV) are important pathogens of porcine viral diarrhea. Co-infection situations among these three viruses in clinics are common, which increases the difficulty of differential diagnosis. Currently, polymerase chain reaction (PCR) is commonly used to detect pathogens. TaqMan real-time PCR is more sensitive than conventional PCR and has better specificity and accuracy. In this study, a triplex real-time RT-PCR assay based on TaqMan probes was developed for differential detection of PEDV, PoRV, and PDCoV. The triplex real-time RT-PCR assay developed in this study could not detect unrelated pathogens and showed satisfactory specificity, sensitivity, repeatability, and reproducibility with a limit of detection (LOD) of 6.0 × 10 copies/μL. Sixteen clinical samples were used to compare the results of the commercial RT-PCR kit and the triplex RT-PCR for PEDV, PoRV, and PDCoV detection, and the results were completely consistent. A total of 112 piglet diarrhea samples collected from Jiangsu province were next used to study the local prevalence of PEDV, PoRV, and PDCoV. The positive rates of PEDV, PoRV, and PDCoV detected by the triplex real-time RT-PCR were 51.79% (58/112), 59.82% (67/112), and 2.68% (3/112), respectively. The co-infections of PEDV and PoRV were frequent (26/112, 23.21%), followed by the co-infections of PDCoV and PoRV (2/112, 1.79%). This study established a useful tool for simultaneous differentiation of PEDV, PoRV, and PDCoV in practice and provided valuable information on the prevalence of these diarrhea viral pathogens in Jiangsu province.
猪病毒性腹泻在临床实践中非常常见，给养猪业造成了巨大损失。猪流行性腹泻病毒（PEDV）、猪轮状病毒（PoRV）和猪德尔塔冠状病毒（PDCoV）是猪病毒性腹泻的重要病原体。这三种病毒在临床上常发生混合感染，增加了鉴别诊断的难度。目前，聚合酶链反应（PCR）常用于检测病原体。TaqMan 实时 PCR 比常规 PCR 更敏感，具有更好的特异性和准确性。本研究建立了一种基于 TaqMan 探针的三重实时 RT-PCR 检测方法，用于 PEDV、PoRV 和 PDCoV 的鉴别检测。本研究建立的三重实时 RT-PCR 检测方法不能检测无关病原体，具有满意的特异性、灵敏度、重复性和再现性，检测限（LOD）为 6.0×10 拷贝/μL。用商业 RT-PCR 试剂盒和三重 RT-PCR 检测 16 份临床样本检测 PEDV、PoRV 和 PDCoV 的结果完全一致。随后，用该方法检测了江苏省采集的 112 份仔猪腹泻样本，研究了 PEDV、PoRV 和 PDCoV 的当地流行情况。三重实时 RT-PCR 检测 PEDV、PoRV 和 PDCoV 的阳性率分别为 51.79%（58/112）、59.82%（67/112）和 2.68%（3/112）。PEDV 和 PoRV 的混合感染较为常见（26/112，23.21%），其次是 PDCoV 和 PoRV 的混合感染（2/112，1.79%）。本研究建立了一种用于 PEDV、PoRV 和 PDCoV 同时鉴别诊断的实用工具，并为江苏省这些腹泻病毒病原体的流行情况提供了有价值的信息。
