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目的建立1种快速、特异、灵敏的发热伴血小板减少综合征病毒(SFTSV)的检测方法。方法针对SFTSV M基因保守区域的核酸序列,设计逆转录-环介导等温扩增(RT-LAMP)特异性引物,优化反应体系和条件。用毛细管电泳法和横向流动试纸条(LFD)法检测扩增产物,建立RT-LAMP检测方法。进行敏感性、特异性检验,并与Real-time RT-PCR法进行比较。结果以LFD法和毛细管电泳法检测RT-LAMP扩增产物具有相同的敏感性和特异性;RT-LAMP-LFD检测SFTSV的敏感性为10copies RNA分子/反应,且与其他病毒无交叉反应。RT-LAMP-LFD和Real-time RT-PCR检测临床标本阳性率差异无统计学意义(P>0.05),2种方法一致性好(Kappa=0.918)。结论建立的RT-LAMP方法快速、简单、操作简单,具有较高的敏感性和特异性,适合应用于基层单位和现场SFTSV的快速检测。
Objective To establish a rapid, specific and sensitive detection of fever with thrombocytopenic syndrome virus (SFTSV). Methods According to the nucleotide sequence of the conserved region of SFTSV M gene, RT-LAMP-specific primers were designed and the reaction system and conditions were optimized. The amplification products were detected by capillary electrophoresis and lateral flow test strips (LFD), and RT-LAMP assay was established. Sensitivity and specificity tests were performed and compared with Real-time RT-PCR. Results The detection of RT-LAMP by LFD and capillary electrophoresis showed the same sensitivity and specificity. The sensitivity of RT-LAMP-LFD for detection of SFTSV was 10copies RNA molecules / reaction and no cross-reaction with other viruses. The positive rates of RT-LAMP-LFD and Real-time RT-PCR in detecting clinical specimens showed no significant difference (P> 0.05). The two methods were consistent (Kappa = 0.918). Conclusion The established RT-LAMP method is rapid, simple, simple to operate and has high sensitivity and specificity. It is suitable for rapid detection of grass-root units and on-site SFTSV.