非洲猪瘟病毒核酸可视化检测方法的建立与评价

Establishment and assessment of a visual detection method for nucleic acid of African swine fever virus

非洲猪瘟(African swine fever,ASF)是由非洲猪瘟病毒(African swine fever virus,ASFV)引起猪的一种高度传染性病毒性疾病。本病传染性强、致病性高,自2018年以来在我国迅速蔓延流行,对我国生猪养殖业造成了巨大的影响。为实现ASFV的快速检测,本研究将环介导等温扩增技术(loop-mediated isothermal amplification,LAMP)与封闭式核酸可视化试纸相结合,建立了一种检测ASFV B646L基因的核酸可视化方法。该方法在65℃条件下,50 min内可检出1.16 copies/μL的重组质粒,且该方法与猪瘟病毒、猪繁殖和呼吸综合征病毒、猪细小病毒、猪传染性胃肠炎病毒无交叉反应。此外该方法无需特殊设备,操作简便,能有效避免传统等温扩增气溶胶污染造成假阳性的缺点。本研究为ASF疑似感染病例的筛选与早期诊断提供了一种快速、便捷、灵敏、可靠的候选方法。

African swine fever(ASF),caused by the African swine fever virus(ASFV),is a highly contagious infectious disease of pigs.This disease has been spread rapidly in China since 2018,posing a huge threat to China's pig farming industry.To rapid detect the ASFV,a loop-mediated isothermal amplification(LAMP)combined with the disposable nucleic acid visualization test strip was established for visual detection of the nucleic acid of ASFV B646L gene.The method was easy to operate without special instruments and equipment,while it effectively avoided the disadvantage of false positives caused by aerosol contamination.The method was able to detect 1.16 copies/μL of the recombinant plasmid in 50 min at 65℃.In addition,the method was specific with no cross-reaction with classical swine fever virus,porcine reproductive and respiratory syndrome virus,porcine parvovirus,transmissible gastroenteritis virus.The results in this study provides a rapid,convenient,sensitive and reliable method for early diagnosis and screening for ASFV suspected infection cases.