摘要
目的应用多重实时荧光PCR技术对北京地区有免疫史麻疹患者临床样本进行核酸检测，鉴别麻疹野病毒和疫苗病毒。方法于2011年7月至2012年2月在北京采集16例8～9月龄初免后疑似麻疹患者的10份咽拭子和15份尿液标本。先用麻疹疫苗病毒、野病毒和其他呼吸道病毒检测多重实时荧光PCR方法的特异性，后对麻疹疫苗株和野病毒滴定后稀释，检测方法的敏感性，并与PCR—RFLP法比较。同时，检测麻疹患者的临床样本，并测序确认。结果Fam、Joe及Cy5检测通道引物与探针对麻疹病毒有很好的检测特异性，对疫苗病毒和野病毒有很好的区分效果。此方法检测麻疹疫苗病毒的灵敏度达到0．1CCID50／0．1ml；检测麻疹野病毒的灵敏度可达0．006CCID50／0．1ml，敏感性高于PCR—RFLP法。16例接种过首剂麻疹疫苗的麻疹患者中，3例阴性，其余13例均为麻疹病毒A基因型。经测序验证均为疫苗株。结论多重实时荧光PCR鉴别麻疹疫苗株和野毒株的方法准确、快速、灵敏，可以为筛查麻疹患者、鉴定疫苗相关病例提供帮助。
Objective To identify wild measles virus and vaccine virus by detection nucleic acid of clinical samples from measles patients with immunization history circulating in Beijing through muhiplex real-time fluorescent PCR technology. Methods From July 2011 to February 2012, 10 throat swabs and 15 urine specimens were collected from 16 suspected measles patients who were 8 -9 months old infants with immunization history in Beijing. The specificity of multiplex real-time fluorescent PCR was firstly tested by measles vaccine virus, wild virus and other respiratory virus. Then the vaccine virus and wild virus were titrated and diluted to test the sensitivity of the PCR method. The result was then eompared with it analyzed by PCR-RFLP method. Meanwhile, the clinical sample of the measles patients were tested and confirmed by sequencing method. Results The primer-probe sets of Faro, Joe and Cy5 showed great specificity of measles virus, and could distinguish the measles vaccine virus and wild virus well. The sensitivity of this method to detect measles vaccine virus reached 0. 1 CCID50/0. 1 ml; and the sensitivity to wild virus reached 0. 006 CCID50/0. 1 ml; which were both higher than the sensitivity of PCR-RFLP method. Out of the 16 measles patients with vaccination history, 3 were negative and the other 13 all belonged to measles virus genotype A, and were confirmed to be vaccine virus by sequencing method. Conclusion Multiplex real-time PCR method is accurate, rapid and sensitive to identify measles vaccine virus and wild virus. The method could greatly help us to find measles patients and to identify the vaccine-related cases.
出处
《中华预防医学杂志》
CAS
CSCD
北大核心
2012年第10期942-945,共4页
Chinese Journal of Preventive Medicine
关键词
聚合酶链反应
麻疹疫苗
麻疹病毒
Polymerase chain reaction
Measles vaccine
Measles virus
