Use of Mycobacteriophage Quantitative PCR on MGIT Broths for a Rapid Tuberculosis Antibiogram
Use of Mycobacteriophage Quantitative PCR on MGIT Broths for a Rapid Tuberculosis Antibiogram
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DOI：
10.1128/jcm.03637-13
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发表时间：
2014-05-01
影响因子：
9.4
通讯作者：
Houpt, Eric R.
中科院分区：
文献类型：
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作者：
Foongladda, Suporn;Klayut, Wiphat;Houpt, Eric R.
基于表型培养的结核分枝杆菌药物敏感性测试 (DST) 是一种宝贵的工具，可用于识别个体化耐多药 (MDR) 结核病 (TB) 治疗方案中的四到六种活性药物。然而，目前基于文化的方法进展缓慢；因此，我们评估了一种基于分枝杆菌噬菌体的快速定量 PCR (qPCR) 测定，可直接用于结核分枝杆菌阳性 MGIT 肉汤。我们对 325 个临床结核分枝杆菌 MGIT 肉汤培养物与通过琼脂比例测试的各自传代分离株进行了噬菌体 qPCR 比较，使用简单的 Delta Cq 值截断值 3（其中 Cq 是定量周期，Delta Cq 计算为起始噬菌体的 Cq 减去含药物培养基中结核分离株的 Cq）。测试的 13 种药物/浓度的中位准确度为 98%，其中大多数差异是假抗性结果。对 Lwenstein-Jensen 培养基上生长的 393 个分离株的更多耐药菌株进行噬菌体 qPCR 评估，结果显示相似的结果，中位准确度、敏感性和特异性分别为 97%、90% 和 99%。这种基于快速培养的 DST 方法可用于任何药物，在 TB 阳性 MGIT 肉汤上进行，从样本到抗菌谱的周转时间约为 23.9 天，而在固体培养基上分离株生长需要 58.6 天，然后进行琼脂比例 DST。
Phenotypic culture-based drug susceptibility testing (DST) for Mycobacterium tuberculosis is a valuable tool to identify four to six active drugs for individualized multidrug-resistant (MDR) tuberculosis (TB) regimens. Current culture-based methods are slow, however; therefore, we evaluated a rapid mycobacteriophage-based quantitative PCR (qPCR) assay for use directly on M. tuberculosis-positive MGIT broths. We compared phage qPCRs, using a simple cutoff of 3 for the Delta Cq value (where Cq is quantification cycle, and Delta Cq is calculated as the Cq of starting phage minus the Cq of TB isolates in drug-containing medium), on 325 clinical M. tuberculosis MGIT broth cultures versus the respective subcultured isolates tested by agar proportion. The median accuracy for the 13 drugs/concentrations tested was 98%, with most discrepancies being false-resistant results. Evaluation of phage qPCR on greater numbers of resistant strains of 393 isolates grown on Lwenstein-Jensen medium showed similar findings, with a median accuracy, sensitivity, and specificity of 97%, 90%, and 99%, respectively. This rapid culture-based DST methodology can be performed for any drug on TB-positive MGIT broths, with a specimen-to-antibiogram turnaround time of approximately 23.9 days, compared with waiting 58.6 days for isolate growth on solid medium followed by agar proportion DST.