Shandong Key Laboratory of Animal Disease Control and Breeding, Institute of Animal Science and Veterinary Medicine, Shandong Academy of Agricultural Sciences, Jinan, China.
National Veterinary Microbiological Drug Resistance Risk Assessment Laboratory, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China.
Foodborne Pathog Dis. 2021 Apr;18(4):290-296. doi: 10.1089/fpd.2020.2868. Epub 2021 Jan 28.
remains a serious threat to the worldwide swine industry and human health. In this study, rapid assays for the detection of three common virulence-related factors (, , and ) were developed, evaluated, and applied. Loop-mediated isothermal amplification (LAMP) primers were designed using Primer Explorer V5 software. The sensitivity and specificity of the LAMP assays were determined based on sample turbidity. For all three genes, LAMP assays were performed at 62°C with a reaction time of 60 min. The detection limit of conventional polymerase chain reaction (PCR) was 1 ng/μL, 10 pg/μL, and 100 fg/μL for the , , and genes, respectively. For the LAMP assays, the detection limits were 10 pg/μL, 10 fg/μL, and 100 fg/μL for , , and , respectively, representing sensitivities 100-1000 times higher than those of the PCR assay. Furthermore, when the LAMP assays were applied to clinical strains, the results were consistent with those of the PCR assay, confirming the LAMP assays as rapid and reliable detection techniques. In conclusion, the LAMP assays described in this study have the potential to become standard methods to detect the virulence factors , , and . To the best of our knowledge, this is the first study to report the application of LAMP to detect the , , and genes.
仍然是全球养猪业和人类健康的严重威胁。在本研究中，开发、评估和应用了三种常见毒力相关因子（、和）的快速检测方法。使用 Primer Explorer V5 软件设计了环介导等温扩增（LAMP）引物。根据样品浊度确定了 LAMP 检测方法的灵敏度和特异性。对于所有三个基因，LAMP 检测在 62°C 下进行，反应时间为 60 分钟。传统聚合酶链反应（PCR）的检测限分别为 1ng/μL、10pg/μL 和 100fg/μL。对于 LAMP 检测，检测限分别为 10pg/μL、10fg/μL 和 100fg/μL，灵敏度比 PCR 检测高 100-1000 倍。此外，当将 LAMP 检测应用于临床菌株时，结果与 PCR 检测一致，证实了 LAMP 检测是一种快速可靠的检测技术。总之，本研究中描述的 LAMP 检测法具有成为检测毒力因子、和的标准方法的潜力。据我们所知，这是首次报道应用 LAMP 检测、和基因的研究。
