Performance of TaqMan Array Card to detect TB drug resistance on direct specimens

Culture based phenotypic drug susceptibility testing (DST) for Mycobacterium tuberculosis (TB) is time consuming therefore rapid genotypic methods are increasingly being utilized. We previously developed and evaluated on TB isolates a rapid genotypic TaqMan array card (TAC) that detects mutations in several resistance-associated genes using dozens of primer pairs, probes, and high resolution melt analysis, with >96% accuracy versus Sanger sequencing. In this study we examined the performance of TAC on sputum, comparing results between 71 paired sputum and multi drug resistant TB isolates. We also adapted the TAC to include wild-type probes and broadened coverage for rpoB and gyrA mutations. TAC was 89% successful at detecting wild-type or mutations within inhA, katG, rpoB, eis, gyrA, rplC, and pncA on smear positive sputa and 33% successful on smear negative sputa. The overall accuracy of these detections as compared to the TAC results of the paired isolate was 95% ± 7 (average se...

基于培养的表型药物敏感性试验（DST）用于结核分枝杆菌（TB）的检测耗时冗长，因此快速基因型检测方法的应用愈发广泛。本研究团队此前已针对结核分枝杆菌分离株开发并评估了一款快速基因型TaqMan阵列卡（TAC），该检测系统通过数十对引物、探针及高分辨率熔解曲线分析，可检测多种耐药相关基因的突变，与桑格测序相比准确率达96%以上。本研究中，我们考察了TAC应用于痰液样本的检测性能，对比了71组配对痰液与耐多药结核分枝杆菌分离株的检测结果。同时，我们对TAC进行了优化升级，加入了野生型探针，并拓展了针对rpoB与gyrA基因突变的检测覆盖范围。在痰涂片阳性痰液样本中，TAC对inhA、katG、rpoB、eis、gyrA、rplC及pncA基因的野生型序列或相关突变的检出成功率为89%；在痰涂片阴性痰液样本中，该检出成功率为33%。以配对分离株的TAC检测结果作为参照标准，此类检测的整体准确率为95%±7（平均 se...