保守和应用三个假设的蛋白质编码基因，用于直接检测痰液标本中的结核分枝杆菌。
The conservation and application of three hypothetical protein coding gene for direct detection of Mycobacterium tuberculosis in sputum specimens.
机构信息
Shanghai Key Laboratory of Tuberculosis, Shanghai Pulmonary Hospital, Tongji University School of Medicine, Shanghai, China.
出版信息
PLoS One. 2013 Sep 13;8(9):e73955. doi: 10.1371/journal.pone.0073955. eCollection 2013.
BACKGROUND
Accurate and early diagnosis of tuberculosis (TB) is of major importance in the control of TB. One of the most important technical advances in diagnosis of tuberculosis is the development of nucleic acid amplification (NAA) tests. However, the choice of the target sequence remains controversial in NAA tests. Recently, interesting alternatives have been found in hypothetical protein coding sequences from mycobacterial genome.
METHODOLOGY/PRINCIPAL FINDINGS: To obtain rational biomarker for TB diagnosis, the conservation of three hypothetical genes was firstly evaluated in 714 mycobacterial strains. The results showed that SCAR1 (Sequenced Characterized Amplified Region) based on Rv0264c coding gene showed the highest conservation (99.8%) and SCAR2 based on Rv1508c gene showed the secondary high conservation (99.7%) in M. tuberculosis (MTB) strains. SCAR3 based on Rv2135c gene (3.2%) and IS6110 (8%) showed relatively high deletion rate in MTB strains. Secondly, three SCAR markers were evaluated in 307 clinical sputum from patients in whom TB was suspected or patients with diseases other than TB. The amplification of IS6110 and 16SrRNA sequences together with both clinical and bacteriological identification was as a protocol to evaluate the efficacy of SCAR markers. The sensitivities and specificities, positive predictive value (PPV) and negative predictive value (NPV) of all NAA tests were higher than those of bacteriological detection. In four NAA tests, IS6110 and SCAR3 showed the highest PPV (100%) and low NPV (70% and 68.8%, respectively), and SCAR1 and SCAR2 showed the relatively high PPV and NPV (97% and 82.6%, 95.6% and 88.8%, respectively).
CONCLUSIONS/SIGNIFICANCE: Our result indicated that SCAR1 and SCAR2 with a high degree of sequence conservation represent efficient and promising alternatives as NAA test targets in identification of MTB. Moreover, the targets developed from this study may provide more alternative targets for the development of a multisite system to effectively detect MTB in samples.
背景
准确和早期诊断结核病（TB）对于结核病控制非常重要。结核病诊断方面最重要的技术进展之一是核酸扩增（NAA）检测的发展。然而，在 NAA 检测中，目标序列的选择仍然存在争议。最近，在分枝杆菌基因组的假设蛋白编码序列中发现了有趣的替代物。
方法/主要发现：为了获得用于 TB 诊断的合理生物标志物，首先评估了 714 株分枝杆菌菌株中三个假设基因的保守性。结果表明，基于 Rv0264c 编码基因的 SCAR1（测序特征扩增区）在结核分枝杆菌（MTB）菌株中显示出最高的保守性（99.8%），基于 Rv1508c 基因的 SCAR2 显示出其次高的保守性（99.7%）。基于 Rv2135c 基因（3.2%）和 IS6110（8%）的 SCAR3 显示出在 MTB 菌株中相对较高的缺失率。其次，在 307 例疑似结核病或非结核病患者的临床痰标本中评估了三个 SCAR 标记物。将 IS6110 和 16SrRNA 序列的扩增与临床和细菌学鉴定相结合，作为评估 SCAR 标记物疗效的方案。所有 NAA 检测的灵敏度和特异性、阳性预测值（PPV）和阴性预测值（NPV）均高于细菌学检测。在四项 NAA 检测中，IS6110 和 SCAR3 显示出最高的 PPV（100%）和低 NPV（70%和 68.8%），SCAR1 和 SCAR2 显示出相对较高的 PPV 和 NPV（97%和 82.6%，95.6%和 88.8%）。
结论/意义：我们的结果表明，具有高度序列保守性的 SCAR1 和 SCAR2 作为 NAA 检测的靶点，代表了鉴定 MTB 的有效和有前途的替代物。此外，本研究开发的靶点可能为开发多站点系统提供更多的替代靶点，以有效检测样本中的 MTB。
Zotero 插件