基于酶切探针等温扩增技术快速检测汉坦病毒S4亚型方法的建立

RAPID DETECTION METHOD OF HANTAVIRUS SEOV-S4 BASED ON ENZYMATIC PROBE ISOTHERMAL AMPLIFICATION

汉坦病毒(hantavirus,HV)属汉城病毒S4亚型(Seoul virus-S4 subtype,SEOV-4)是东南沿海地区的主要HV流行亚型,目前尚缺乏快速诊断检测方法。本研究基于酶切探针等温扩增技术(Enzymatic Probe Isothermal Amplification,EPIA),以SEOV-S4较为保守的S序列为靶序列设计引物和探针,建立了一种快速检测SEOV-S4的方法。用靶基因合成质粒梯度稀释进行灵敏度和重复性验证,并用人源其他亚型HV进行特异性验证,结果显示,基于EPIA的SEOV-S4检测方法灵敏度可以达到10 copies/μL,且与人源其他亚型HV无交叉反应。结果表明,本研究建立的检测方法具有良好的敏感性、特异性和重复性,可用于相关样本的诊断与筛查。

Hantavirus(HV)belongs to Seoul virus-S4 subtype(SEOV-4),which is the main epidemic subtype of HV in southeast coastal area.In this study,a rapid detection method for SEOV-S4 was established based on Enzymatic Probe Isothermal Amplification(EPIA)and primers and probes were designed to target the relatively conserved S sequence of SEOV-S4.The sensitivity and repeatability were verified by gradient dilution of target gene synthetic plasmid,and the specificity was verified by other human subtype HV.The results showed that the sensitivity of SEOV-S4 method based on EPIA could reach 10 copies/μL.There was no cross-reaction with other human subtypes of HV.The method established in this study has good sensitivity,specificity and repeatability,and can be used for the diagnosis and screening of relevant samples.