Li Renfeng, Cao Wenyan, Yuan Jiakang, Li Linyue, Zhou Yanlin, Wang Fangyu, Wang Ziliang, Tian Xiangqin
College of Animal Science and Veterinary Medicine, Henan Institute of Science and Technology, Xinxiang, China.
Sanquan College of Xinxiang Medical University, Xinxiang, China.
Front Microbiol. 2024 Sep 16;15:1465923. doi: 10.3389/fmicb.2024.1465923. eCollection 2024.
The emergence of porcine deltacoronavirus (PDCoV) presents a significant threat to both human and animal health due to its ability to cause highly contagious enteric diseases. This underscores the crucial need for timely and accurate diagnosis to facilitate effective epidemiological investigation and clinical management. This research aimed to establish a visual detection method based on reverse transcription loop-mediated isothermal amplification (RT-LAMP) for PDCoV testing. In this study, six pairs of primers were designed according to the conserved sequences of PDCoV ORF1a/b genes. The primer sets and parameters that affect LAMP reaction were optimized. The visual RT-LAMP method was developed by incorporating methyl red into the optimized reaction system, it exclusively detected PDCoV without cross-reactivity with other viruses and the detection limits for PDCoV could reach 10 copies/μL. In comparison with RT-PCR for testing 132 clinical samples, the relative specificity and sensitivity of the visual RT-LAMP were found to be 99.2 and 100%, respectively, with a concordance rate of 99.2% and a kappa value of 0.959, indicating that the visual RT-LAMP is a reliable method for the application of PDCoV detection in clinical samples.
猪德尔塔冠状病毒（PDCoV）的出现因其能够引发高度传染性肠道疾病，对人类和动物健康都构成了重大威胁。这突出表明迫切需要及时、准确的诊断，以促进有效的流行病学调查和临床管理。本研究旨在建立一种基于逆转录环介导等温扩增（RT-LAMP）的可视化检测方法用于PDCoV检测。在本研究中，根据PDCoV ORF1a/b基因的保守序列设计了6对引物。对影响LAMP反应的引物组和参数进行了优化。通过将甲基红加入优化后的反应体系中开发出可视化RT-LAMP方法，该方法仅能检测PDCoV，与其他病毒无交叉反应，对PDCoV的检测限可达10拷贝/μL。与用于检测132份临床样本的RT-PCR相比，可视化RT-LAMP的相对特异性和敏感性分别为99.2%和100%，一致性率为99.2%，kappa值为0.959，表明可视化RT-LAMP是一种在临床样本中应用于PDCoV检测的可靠方法。
