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目的探讨Xpert MTB/RIF检测痰标本中结核分枝杆菌(MTB)的最佳实验条件并进行方法学评价。方法应用Xpert MTB/RIF检测MBT标准株H37Rv、H37Ra和牛分枝杆菌及20种常见非结核分枝杆菌(NTM)、18种非分枝杆菌病原体和临床分离株,分析Xpert MTB/RIF的敏感性、特异性和重复性,比较不同前处理方法对实验结果的影响,针对仪器出现的报警、错误及无效结果进行相应的处置和总结。结果 MTB标准菌株H37Rv、H37Ra和牛分枝杆菌Xpert MTB/RIF检测阳性,敏感度为1×102 CFU/ml(在痰标本中的检测极限为106 CFU/ml);20种常见NTM和18种非分枝杆菌病原体检测阴性;标本中含有1%浓度的红细胞、白细胞、血清未对103/ml的H37Rv产生干扰作用;重复试验表明本法批间、批内变异系数均小于15%。100株MTB(其中50株Hain线性探针试验确认存在rpoB基因突变)和20株NTM临床分离株加入阴性临床痰标本制备成为含菌浓度为2×102条/ml的模拟痰标本经Xpert MTB/RIF检测均相符。在SR处理液加入半胱氨酸的改良直接法前处理痰液Xpert MTB/RIF检测出现“错误+无效”的比例显著低于沉淀处理法和直接法前处理的痰标本(χ2值分别为4.19和6.74,P<0.01)。结论 Xpert MTB/RIF灵敏、特异、快速、简便,可用于MTB和利福平耐药MTB的快速检测,但仍存在错误和无效检测而影响实验结果,应采取有效措施予以避免。
Objective To investigate the best experimental conditions for Xpert MTB / RIF detection of Mycobacterium tuberculosis (MTB) in sputum and to evaluate the methodology. Methods Xpert MTB / RIF was used to detect the sensitivity of Xpert MTB / RIF to H37Rv, H37Ra and Mycobacterium bovis, 20 common non-mycobacterium tuberculosis (NTM), 18 non-mycobacterial pathogens and clinical isolates. , Specificity and repeatability. The effects of different pretreatment methods on the experimental results were compared. Corresponding disposal and summary were made for the alarms, errors and invalid results of the instrument. Results MTB standard strains H37Rv, H37Ra and M. bovis Xpert MTB / RIF were positive with a sensitivity of 1 × 102 CFU / ml (detection limit of 106 CFU / ml in sputum samples); 20 common NTMs and 18 non-specific The mycobacterial pathogen test negative; the specimen contains 1% concentration of red blood cells, white blood cells, serum did not interfere with 103 / ml of H37Rv; repeated tests showed that the method of inter-batch, intra-batch variation coefficients were less than 15%. 100 MTB (of which 50 Hain linear probe test confirmed the existence of rpoB gene mutation) and 20 strains of NTM clinical isolates were added to the negative clinical sputum specimens to prepare a simulated sputum specimen with a bacterial concentration of 2 × 102 / ml by Xpert MTB / RIF test are consistent. The percentage of “wrong + ineffective” in the Xpert MTB / RIF assay for the sputum treated with modified direct method with cysteine added to the SR treatment was significantly lower than that of the sputum treated with the sedimentation and direct pretreatment 4.19 and 6.74, P <0.01). Conclusion Xpert MTB / RIF is sensitive, specific, rapid and simple and can be used for the rapid detection of MTB and rifampicin-resistant MTB. However, there are still errors and invalid tests that affect the experimental results. Effective measures should be taken to avoid this.