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目的建立噬菌体生物扩增法(PhaB)快速测定吡嗪酰胺耐药性,并探讨其在结核分枝杆菌吡嗪酰胺耐药性测定中的应用价值。方法应用建立的 PhaB 测定108株结核分枝杆菌临床分离株的吡嗪酰胺耐药性,并与绝对浓度法的药敏结果进行比较,对不符合的菌株进行最低抑菌浓度(MIC)测定和序列测定分析。结果 PhaB 检测吡嗪酰胺耐药性的最佳测定条件为 pH 值5.5、药物浓度200μg/ml、37℃作用48 h。用绝对浓度法检测108株结核分枝杆菌临床分离株,其中吡嗪酰胺敏感33株、耐药75株;用 PhaB 检测该108株结核分枝杆菌临床分离株,若以噬菌斑减少95%为判断标准,则吡嗪酰胺敏感32株,耐药76株。2种方法检测均为敏感的为28株,均为耐药的为71株,符合率为91.7%;2种方法检测结果不符的为9株,其中5株的 PhaB 结果与 MIC 结果相符,4株的结果不符,测序结果表明9株中有7株的 PhaB 结果与测序结果相符。如以绝对浓度法药敏结果为判断标准,PhaB 检测吡嗪酰胺耐药性的敏感性为94.7%,特异性为84.8%,阳性预测值为93.4%,阴性预测值为87.5%,准确性为91.7%。结论 PhaB 测定吡嗪酰胺耐药性简便、快速,3 d 即可获得药敏结果,可作为吡嗪酰胺耐药性的快速筛选方法。
Objective To establish a phage amplification assay (PhaB) for the rapid determination of pyrazinamide resistance and to explore its value in the determination of mycobacterium tuberculosis pyrazinamide resistance. Methods PhaB was used to determine the drug resistance of 108 clinical isolates of Mycobacterium tuberculosis and compared with the susceptibility results of the absolute concentration method. The minimum inhibitory concentration (MIC) Sequence analysis. Results The best assay conditions of Phab for detecting pyrazinamide resistance were pH 5.5, drug concentration 200μg / ml and 37 ℃ for 48 h. The clinical isolates of 108 strains of Mycobacterium tuberculosis were detected by absolute concentration method, of which 33 were pyrazinamide-sensitive and 75 were drug-resistant. The clinical isolates of 108 strains of Mycobacterium tuberculosis were detected by PhaB. If plaque reduction was 95% To determine the standard, then pyrazinamide sensitive 32, resistant 76. 28 strains were sensitive to both methods, 71 were resistant, the coincidence rate was 91.7%; 9 strains were out of line with the two methods, of which 5 were consistent with the results of MIC, 4 The results of the strains did not match. The sequencing results showed that PhaB results of 7 of 9 strains were consistent with the sequencing results. For example, the sensitivities of PhaB to pyrazinamide resistance were 94.7%, 84.8%, 93.4%, and 87.5%, respectively. The accuracy of PhaB in detecting pyrazinamide resistance was 91.7%. Conclusion PhaB determination of pyrazinamide resistance is simple, rapid, 3 d to obtain drug susceptibility results, as a rapid screening method for pyrazinamide resistance.