摘要
为建立媒介蚊虫自然感染种群样本携带病原体快速灵敏的检测技术,本研究将实验室经口感染乙脑病毒的致倦库蚊阳性样品和未感染乙脑病毒的阴性样品按一定比例混合成混合样本,分别采用环介导等温扩增技术(Loop Mediated Isothermal Amplification,LAMP)和实时荧光定量聚合酶链反应(Real-time Fluorescent Quantitative PCR,qPCR)检测方法,对感染乙脑病毒的致倦库蚊混合样本进行检测。结果显示:当阳性样本在混合样本中所占比例为0.01%时,LAMP仍然可检测到病原体;定量PCR的最低检出比例是阳性样本在混合样本中所占0.14%以上。通过进一步优化反应体系各组分的浓度、反应时间和温度,建立了灵敏性高的LAMP扩增方法。
In order to establish a rapid and sensitive detection technique for the pathogen of vector mosquito naturally infected population, this study mixed the positive samples of Culex pipiens quinquefasciatus tiretis and negative samples of uninfected encephalitis virus into mixed samples in a certain proportion, using Loop Mediated Isotherm respectively. Loop-mediated isothermal amplification(Loop Mediated Isothermal Amplification, LAMP) and real-time fluorescent quantitative PCR methods were used to detect the mixed samples of Cx pipiens quinquefasciatus infected with JEV. The results showed that when the proportion of positive samples in mixed samples was 0.01%, LAMP could still detect pathogens;The lowest detection ratio of quantitative PCR was 0.14% of the positive samples in the mixed sample. By further optimizing the concentration, reaction time and temperature of each component in the reaction system, a LAMP amplification method with high sensitivity and specificity was established.
作者
郭晓霞
高剑
李春晓
邢丹
董言德
赵彤言
GUO Xiao-Xia;GAO Jian;LI Chun-Xiao;XING Dan;DONG Yan-De;ZHAO Tong-Yan(Beijing Key Laboratory of Vector Borne and Natural Infectious Disease,State Key Laboratory of Pathogens and Biosecurity,Beijing Institute of Microbiology and Epidemiology,Beijing 100071,China)
出处
《寄生虫与医学昆虫学报》
CAS
2019年第1期36-40,共5页
Acta Parasitologica et Medica Entomologica Sinica
基金
国家科技重大专项(2017ZX100303404)
关键词
LAMP
乙脑病毒
混合样本
Culex pipiens quinquefasciatus
Mixed samples
Real-time Fluorescent Quantitative PCR
Loop Mediated Isothermal Amplification
