Division of Biomedical Research, Sri Narayani Hospital and Research Centre, Thirumalaikodi, Sripuram, Vellore, Tamil Nadu, India.
Mol Diagn Ther. 2010 Aug 1;14(4):223-7. doi: 10.1007/BF03256377.
With 1.8 million new cases each year, India carries 20% of the global burden of tuberculosis, a situation that is now further exacerbated with the emergence of drug resistance. The current diagnostic technique suggested by the Government of India's Revised National Tuberculosis Control Programme is Ziehl-Neelsen staining of a sputum smear. This technique is known to be inadequate.
The aim of this study was to evaluate nested PCR (nPCR) in the detection of pulmonary tuberculosis in sputum samples in comparison with conventional smear findings, in an effort to improve detection rates from those obtained by the smear-alone approach.
Patients attending a tertiary-care hospital (situated in a rural area of Vellore district) with clinical suspicion of pulmonary tuberculosis were prospectively recruited from mid-April 2009 to mid-December 2009 and investigated. The sputum samples were stained by Ziehl-Neelsen staining for smear examination. DNA extracted from concentrated sputum was tested by nPCR, targeting the IS6110 sequence in the Mycobacterium tuberculosis genome.
Among 84 patients tested (median age 45.5 years), 80.95% were from the rural community and 19.05% were from the peri-urban community. Seventeen patients (20.24%; mid-p 95% CI 31.5, 52.4) tested positive by the smear examination and 35 (41.67%; mid-p 95% CI 12.7, 29.8) tested positive by nPCR. The difference in detection rates was statistically significant (chi(2) = 9.02; p = 0.002). The kappa coefficient between smear findings and nPCR findings was 0.47, which was a statistically significant agreement (Z = 4.91; p < 0.0001).
This report describes the molecular detection of M. tuberculosis in patients' sputum samples tested by the nPCR format, using IS6110 as a target sequence. A high prevalence of pulmonary tuberculosis was identified by the nPCR assay, which was shown to have a significantly higher detection rate than conventional smear staining.
印度每年新增结核病病例 180 万例，占全球结核病负担的 20%，而随着耐药性的出现，这一情况现在更加恶化。印度修订后的国家结核病控制规划建议的当前诊断技术是痰液涂片的萋-尼氏染色。已知该技术不够充分。
本研究的目的是评估巢式 PCR（nPCR）在检测痰液样本中的肺结核，与常规涂片结果进行比较，以提高仅通过涂片获得的检测率。
2009 年 4 月中旬至 12 月中旬，从一个位于维洛尔地区农村地区的三级保健医院（农村地区），对具有肺结核临床疑似症状的患者进行前瞻性招募和调查。痰液样本用萋-尼氏染色进行涂片检查。从浓缩痰液中提取的 DNA 通过 nPCR 检测，针对结核分枝杆菌基因组中的 IS6110 序列。
在 84 名接受测试的患者中（中位年龄 45.5 岁），80.95%来自农村社区，19.05%来自城市周边社区。17 名患者（20.24%；中值 95%CI 31.5，52.4）经涂片检查阳性，35 名患者（41.67%；中值 95%CI 12.7，29.8）经 nPCR 检查阳性。检测率差异具有统计学意义（卡方=9.02；p=0.002）。涂片检查结果与 nPCR 检查结果之间的 Kappa 系数为 0.47，具有统计学显著一致性（Z=4.91；p<0.0001）。
本报告描述了使用 IS6110 作为靶序列，通过 nPCR 格式对患者痰液样本中的结核分枝杆菌进行分子检测。nPCR 检测法确定了高患病率的肺结核，其检测率明显高于常规涂片染色。
