实时荧光定量逆转录-聚合酶链反应鉴定脊髓灰质炎病毒方法的评估

Evaluation of Real Time Fluorescent Quantitative Reverse Transcription Polymerase Chain Reaction for Detection of Poliovirus

目的在中国脊髓灰质炎(脊灰)实验室网络[Poliovirus(PV)Laboratory Network,PLN]中,首次应用实时荧光定量逆转录-聚合酶链反应(Real Time Fluorescent Quantitative Reverse Transcription-Polymerase Chain Reaction,rRT-PCR)对PV进行鉴定,并对该方法进行应用评估。方法采用世界卫生组织(WHO)推荐的、美国疾病控制与预防中心研发的rRT-PCR法,对中国PLN既往分离的10株PV进行型内鉴定(Intratypic Differentiation,ITD)和疫苗衍生PV(Vaccine-derived PV,VDPVs)筛选,并将检测结果与毒株的VP1编码区核苷酸序列测定结果进行比较分析。结果10株PV rRT-PCR的结果与VP1编码区核苷酸序列测定结果不能完全相符,rRT-PCR无法鉴别10株PV中的5株Pre(前)-VDPVs,并且漏检了山西省2007年发现的1株Ⅰ型VDPV。结论作为WHO推荐使用的新型ITD方法,rRT-PCR法是否适用于中国PLN,还有待大样本PV rRT-PCR法的回顾性和前瞻性研究。

Objective Application of Real Time Fluorescent quantitative reverse transcription polymerase chani reaction（rRT-PCR）to poliovirus identification in Chinese Poliomyelitis Laboratory Network and evaluation of the assay.Methods According to Real-time RT-PCR recommended by WHO and developed by USA Centers for Disease Control and Prevention,10 poliovirus isolates from laboratories of Chinese poliomyelitis network were tested for intratypic differentiation（ITD）and vaccine derived polioviruses（VDPVs）screening.The results of Real-time RT-PCR for 10 isolates were compared with those of VP1 region sequencing.Result The Real-time RT-PCR results for 10 isolates did not completely consist with those of VP1 region sequencing.5 Pre-VDPVs can not be identified by Real-time RT-PCR and typeⅠVDPV from Shanxi province in 2009 was missed by the assay.Conclusion The Real-time RT-PCR retrospective and prospective researches for large scale of polioviruses well be conducted to determine if the assay is applicable to Chinese Poliomyelitis Laboratory Network.