Feng Zhuo, Shi Kaichuang, Yin Yanwen, Shi Yuwen, Feng Shuping, Long Feng, Wei Zuzhang, Si Hongbin
College of Animal Science and Technology, Guangxi University, Nanning 530005, China.
Guangxi Center for Animal Disease Control and Prevention, Nanning 530001, China.
Animals (Basel). 2024 Dec 9;14(23):3551. doi: 10.3390/ani14233551.
African swine fever virus (ASFV), classical swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV), and porcine pseudorabies virus (PRV) induce similar clinical signs in infected pigs, including hyperthermia, anorexia, hemorrhage, respiratory distress, neurological symptoms, and/or abortions in pregnant sows. The differential diagnosis of these diseases relies on laboratory examinations. In this study, a quadruplex RT-qPCR was established using four pairs of specific primers and probes aimed at the (p72) gene of ASFV, the 5' untranslated region () of CSFV, the gene of PRRSV, and the gene of PRV for the detection and differentiation of ASFV, CSFV, PRRSV, and PRV. The assay exhibited great sensitivity with limits of detection (LODs) of 134.585, 139.831, 147.076, and 142.331 copies/reaction for ASFV, CSFV, PRRSV, and PRV, respectively. The assay exclusively identified ASFV, CSFV, PRRSV, and PRV, yielding negative results for the other control swine viruses used in this study. The intra-assay and inter-assay coefficients of variation (CVs) were not higher than 1.12%, indicating good reproducibility of the assay. The quadruplex RT-qPCR assay was used to analyze 3116 clinical tissue samples from pigs in Guangxi province, China, from April 2023 to September 2024. ASFV, CSFV, PRRSV, and PRV had positivity rates of 10.84% (338/3116), 0.80% (25/3116), 14.92% (465/3116), and 1.38% (43/3116), respectively, demonstrating a coincidence rate of ≥99.45% with the previously described RT-qPCR assays, which were also used to test these same samples. The established assay was rapid, sensitive, and accurate in detecting and differentiating ASFV, CSFV, PRRSV, and PRV.
非洲猪瘟病毒（ASFV）、经典猪瘟病毒（CSFV）、猪繁殖与呼吸综合征病毒（PRRSV）和猪伪狂犬病病毒（PRV）在感染猪中引发相似的临床症状，包括高热、厌食、出血、呼吸窘迫、神经症状和/或怀孕母猪流产。这些疾病的鉴别诊断依赖实验室检查。在本研究中，使用四对针对ASFV的（p72）基因、CSFV的5'非翻译区（）、PRRSV的基因和PRV的基因的特异性引物和探针建立了四重RT-qPCR，用于检测和区分ASFV、CSFV、PRRSV和PRV。该检测方法具有很高的灵敏度，ASFV、CSFV、PRRSV和PRV的检测限（LOD）分别为134.585、139.831、147.076和142.331拷贝/反应。该检测方法仅能鉴定出ASFV、CSFV、PRRSV和PRV，对本研究中使用的其他对照猪病毒产生阴性结果。批内和批间变异系数（CV）均不高于1.12%，表明该检测方法具有良好的重复性。使用该四重RT-qPCR检测方法对2023年4月至2024年9月来自中国广西省猪的3116份临床组织样本进行分析。ASFV、CSFV、PRRSV和PRV的阳性率分别为10.84%（338/3116）、0.80%（25/3116）、14.92%（465/3116）和1.38%（43/3116），与之前用于检测这些相同样本的RT-qPCR检测方法的符合率≥ 99.45%。所建立的检测方法在检测和区分ASFV、CSFV、PRRSV和PRV方面快速、灵敏且准确。
