Tu Fei, Zhang Yongning, Xu Shengkui, Yang Xintan, Zhou Lei, Ge Xinna, Han Jun, Guo Xin, Yang Hanchun
Key Laboratory of Animal Epidemiology of the Ministry of Agriculture, College of Veterinary Medicine, China Agricultural University, Beijing, P. R. China.
Transbound Emerg Dis. 2022 Jul;69(4):2266-2274. doi: 10.1111/tbed.14241. Epub 2021 Jul 27.
Pseudorabies (PR) is an acute infectious disease of pigs caused by pseudorabies virus (PRV), which has caused great economic losses to the pig industry worldwide. Reliable and timely diagnose is crucial for the surveillance, control and eradication of PR. Here, a real-time fluorescent recombinase-aided amplification (real-time RAA) assay was established to detect PRV. Primers and probes were designed based on the conserved regions of the PRV gE gene. The assay was specific for the detection of wild-type PRV, showing no cross-reactivity with other important porcine viruses (including PRV gE-deleted vaccine strains). Analytical sensitivity of the assay was three 50% tissue culture infectious doses (TCID ) of PRV DNA per reaction with 95% reliability, which is comparable to that of a PRV-specific real-time PCR (qPCR) assay. In diagnosis of 206 clinical tissue samples, the diagnose accordance rate between the real-time RAA assay and qPCR assay was 97.57% (201/206). Interestingly, the amplified products of real-time RAA could be visualized under a portable blue light instrument, making it possible for the rapid detection of PRV in resource-limited settings and on-site screening. Therefore, our developed real-time RAA assay is a diagnostic method for the rapid detection of PRV in the field.
伪狂犬病（PR）是由伪狂犬病病毒（PRV）引起的猪的一种急性传染病，给全球养猪业造成了巨大经济损失。可靠且及时的诊断对于伪狂犬病的监测、控制和根除至关重要。在此，建立了一种实时荧光重组酶辅助扩增（实时RAA）检测方法来检测PRV。基于PRV gE基因的保守区域设计了引物和探针。该检测方法对野生型PRV具有特异性，与其他重要的猪病毒（包括缺失PRV gE的疫苗株）无交叉反应。该检测方法的分析灵敏度为每个反应3个50%组织培养感染剂量（TCID）的PRV DNA，可靠性为95%，与PRV特异性实时荧光定量聚合酶链反应（qPCR）检测方法相当。在对206份临床组织样本的诊断中，实时RAA检测方法与qPCR检测方法的诊断符合率为97.57%（201/206）。有趣的是，实时RAA的扩增产物可以在便携式蓝光仪器下可视化，这使得在资源有限的环境中快速检测PRV和现场筛查成为可能。因此，我们开发的实时RAA检测方法是一种用于现场快速检测PRV的诊断方法。
