数字聚合酶链反应
肺结核
结核分枝杆菌
DNA提取
实时聚合酶链反应
结核分枝杆菌复合物
DNA
聚合酶链反应
核酸
医学
生物
病理
遗传学
基因
作者
Sun-Mi Cho,Saeam Shin,Yoonjung Kim,Wonkeun Song,Seong Geun Hong,Seok Hoon Jeong,Minhee Kang,Kyung A. Lee
标识
DOI:10.1016/j.cmi.2019.11.012
摘要
ObjectivesThe rapid diagnosis of tuberculosis (TB) is important for patient treatment and infection control. Current molecular diagnostic techniques for TB have insufficient sensitivity to detect samples with low bacterial loads. The sensitivity of molecular testing depends on not only the performance of the assay technique but also the nucleic acid extraction method. Here, we present a novel approach using exosomal DNA (exoDNA) and droplet digital PCR (ddPCR) platforms to detect Mycobacterium tuberculosis DNA in clinical samples.MethodsThe ddPCR platform targeting IS6110 was evaluated in parallel using total DNA and exoDNA. The clinical performance of ddPCR method was assessed with 190 respiratory samples from patients with suspected pulmonary TB.ResultsCompared with mycobacterial culture, sensitivity and specificity of ddPCR were 61.5% (95% CI 44.6–76.6%) and 98.0% (95% CI 94.3–99.6%) using total DNA, and 76.9% (95% CI 60.7–88.9%) and 98.0% (95% CI 94.3–99.6%) using exoDNA, respectively. Among 15 culture-positive specimens with low concentrations of target molecules (2~99 positive droplets with exoDNA), only 53.3% (8/15), 46.7% (7/15), and 26.7% (4/15) of cases were detected using ddPCR with total DNA, real-time PCR with exoDNA, and real-time PCR with total DNA, respectively.DiscussionOur platform using ddPCR and exoDNA has the potential to provide sensitive and accurate methodology for TB diagnosis.
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