Digital PCR assay detection of circulating Mycobacterium tuberculosis DNA in pulmonary tuberculosis patient plasma
Digital PCR assay detection of circulating Mycobacterium tuberculosis DNA in pulmonary tuberculosis patient plasma
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DOI：
10.1016/j.tube.2016.04.004
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发表时间：
2016-07-01
期刊：
影响因子：
3.2
通讯作者：
Kaneko, Takeshi
中科院分区：
文献类型：
--
作者：
Ushio, Ryota;Yamamoto, Masaki;Kaneko, Takeshi
核酸扩增试验是肺结核（PTB）的主要诊断工具。最近，数字PCR（dPCR）测定已经提高了用于检测小拷贝数的靶分子的灵敏度。本研究的目的是探讨dPCR检测结核分枝杆菌（MTB）DNA在PTB患者血浆中的应用。从新诊断的痰涂片阳性PTB患者的血浆样品中纯化总DNA。用靶向IS 6110或gyrB的dPCR测定定量样品中MTB特异性基因的拷贝数。共入组33例PTB患者。PTB患者和对照组的两种靶基因拷贝数存在显著差异：IS 6110平均值+/- SD，144.8 +/- 538.3 vs 0.44 +/- 0.49（拷贝/20 μ L，p = 0.004; Manne-Whitney U检验）和gyrB平均值+/- SD分别为359.0 +/- 2116和0.07 +/- 0.28（拷贝/20 μ L，p = 0.011; Manne-Whitney U检验）。该检测的灵敏度为65%或29%，特异性为93%或100%，分别与IS 6110靶向或gyrB靶向检测。dPCR测定成功检测到PTB患者血浆中的MTB DNA。这种微创和准确的方法有可能成为一种替代的诊断选择。(C)2016爱思唯尔有限公司版权所有
Nucleic acid amplification tests are a major diagnostic tool for pulmonary tuberculosis (PTB). Recently, digital PCR (dPCR) assay has improved sensitivity for the detection of small copy numbers of target molecules. The aim of this study is to explore the utility of dPCR for detecting Mycobacterium tuberculosis (MTB) DNA in PTB patient plasma. Total DNA was purified from plasma samples of newly diagnosed sputum smear-positive PTB patients. Copy numbers of MTB-specific genes in the samples were quantified with dPCR assays targeted for IS6110 or gyrB. A total of 33 PTB patients were enrolled. Significant differences between PTB patients and controls were observed in copy numbers of both targets: IS6110 mean +/- SD, 144.8 +/- 538.3 vs 0.44 +/- 0.49 (copies/20 mu L, p = 0.004; Manne-Whitney U test) and gyrB mean +/- SD, 359.0 +/- 2116 vs 0.07 +/- 0.28 (copies/20 mu L, p = 0.011; Manne-Whitney U test), respectively. This test had sensitivities of 65% or 29% and a specificity of 93% or 100% with the IS6110-targeted or gyrB-targeted assays, respectively. A dPCR assay successfully detected MTB DNA in PTB patient plasma. This minimally invasive and accurate method has potential to become an alternative diagnostic option. (C) 2016 Elsevier Ltd. All rights reserved.