利用发光自显影技术快速筛选结核杆菌的利福平耐药性

Rapid detection of rifampicin susceptibility of Mycobacterium tuberculosis by bioluminescence based on film

目的 利用发光自显影技术建立一种快速、简便的结核分支杆菌利福平耐药性筛选方法。方法 利用重组的分支杆菌噬菌体可在活菌内增殖的原理 ,来检测结核杆菌在含利福平液体培养基中的生长状况 ;采用发光自显影方法检测发光强度并确定活菌数 ,对 4 3份结核杆菌样本进行快速检测 ,并与L J培养基检测结果进行对比。结果 发光自显影方法与L J培养法相比 ,阳性率和真阴性率分别为 95.7%、90 % ,最佳检测的细菌浓度为 10 7～ 10 8CFU/ml,最佳药敏试验的利福平浓度为 1mg/ml ,利福平与结核分支杆菌孵育 4 8小时后即可进行发光自显影分析。结论 生物发光方法可用于快速筛选结核分支杆菌的利福平耐药性 ,是一种简单、快速、灵敏、实用的利福平药敏筛选方法。

Objective To develop a rapid method for determining rifampicin susceptibility of Mycobacterium tuberculosis by luminescence based on Polaroid T677 film.Methods The recombinant mycobacteriophage which can express luciferase during it propagated in related bacteria was used to detect the vitality of M.tuberculosis in liquid medium containing rifampicin and luminescence was recorded by an Polaroid T677 film.The susceptibility of 43 strains of tuberculosis mycobacterium to rifampicin was determined by luminescent photography of Polaroid T 677 film.Results Compared with method of L J slant,the sensitivity and specificity of bioluminescent photography were 95.7% and 90% respectively.The optimal bacteria concentration for luminescence detection was 10 7～10 8 CFU/ml bacteria;the optimal reaction time for rifampicin reaction was 48 hours;and the optimal rifampicin concentration for drug susceptibility test is 1mg/ml.The detection time was within 72 hours and can be detected by naked eyes.Conclusion Luminescent photography assay was a rapid,simple,sensitive and selective method for detection rifampicin susceptibility of Mycobacterium tuberculosis strains.