H7N9禽流感病毒实时荧光RT-PCR检测方法的建立与评价

Development and evaluation of real time RT-PCR for avian influenza (H7N9) virus

目的应用Taq Man探针实时荧光RT-PCR技术,建立一种快速检测H7N9禽流感病毒的方法。方法针对H7N9禽流感病毒的HA和NA基因分别设计特异性引物和Taq Man-LNA探针,建立H7N9禽流感病毒特异性实时荧光RT-PCR快速检测方法。结果本研究建立的方法对检测H7N9禽流感病毒株的灵敏度达到10 PFU/ml。该方法特异性强,与其他亚型流感病毒株及呼吸道病原体无交叉反应。与对照方法相比,本研究方法的检测阳性符合率为99.07%,阴性符合率为100%,Kappa值为0.993。结论建立的检测试剂盒具有快速灵敏、结果准确可靠等优点,适用于H7N9禽流感病毒的分子生物学检测和监测。

Objective To establish a method of rapid diagnosis detection of influenza A（H7N9） virus with Taq Man probe real-time fluorescence RT-PCR technology. Methods A Taq Man real-time RT-PCR test based on the NA and HA gene of avian influenza（H7N9） virus was developed in the present study. Results The evaluation results indicated that the sensitivity of the RT-PCR was 10 PFU / ml and no cross reaction with other influenza virus or respiratory pathogens. The result of clinical trials showed that the RT-PCR was a specific, sensitive and rapid assay. Conclusion The H7N9 RT-PCR is an promising method for routine detection of avian influenza（H7N9） virus and could be used to monitor and survey avian influenza（H7N9）virus.