手足口病病原体EV71的检测方法学比较

Comparison of detection methods of pathogen EV71 in hand-foot and mouth disease

目的比较4种不同的方法对手足口病(hand-foot and mouth disease,HFMD)病原体肠病毒71型(enterovirus type 71,EV71)感染的检测效果。方法非洲绿猴肾细胞(Vero)培养至对数生长期后,加入EV71病毒株感染。分别采用细胞病变效应法(cytopathic effect,CPE)、实时荧光定量聚合酶链式反应(real time polymerase chain reaction,Real-Time PCR)、Western blot和免疫荧光法检测病毒感染情况,并对4种检测结果进行比较。结果 4种方法均可检测到EV71的感染,其中CPE最经济实惠,但是检测周期长,需要至少96 h,并且对病毒活力要求高;Western blot和免疫荧光检测相近,72 h即可完成实验,但灵敏度低,对病毒量有一定要求;Real-Time PCR检测既快速灵敏性又高,48 h即可完成实验。结论不同的方法各有其优缺点,需要根据不同的需要进行选择,达到快速准确诊断病原体的目的。

Objective To compare the effects of four different methods in the detection of pathogen enterovirus type 71（EV71） of hand-foot and mouth disease（HFMD）,and to provide reference for clinical detection. Methods African green monkey cells（Vero） were cultured until the logarithmic growth phase before they were infected with the EV71 strain. The cytopathic effect（CPE） method,real-time polymerase chain reaction（Real-Time PCR）,Western blot and immunofluorescence were used respectively to detect the infection of EV71. The results of the four tests were compared. Results All the four methods were able to detect the infection of EV71. Among them,CPE was the most economical method,but its detection cycle was so long as to take at least 96 hours,and it required good vitality of the virus. The effect of detection of Western blot and immunofluorescence was similar,and the experiment could be completed in 72 hours,but their sensitivity was low and a relatively large amount of virus was required. Real-Time PCR detection was both quick and highly sensitive,and the experiment could be completed in 48 hours. Conclusions Each method has its own advantages and disadvantages. A method should be chosen according to different needs so as to make quick and accurate diagnosis of pathogens.