摘要

       背景：
       基因型MTBDRplus是来自德国Hain Lifescience的一种商业探针分析（LPA）试剂盒，得到印度RNTCP项目的认可，用于诊断痰涂片阳性患者中的DRTB病例。尽管LPA已经在多个实验室进行了研究，但全球现有的结核分枝杆菌菌株存在着广泛的变异，而且由于在不同环境中存在独特的基因突变，可能会出现错误的结果。
       目标和目的：
       尝试用LPA进行带型分析，并观察MDR菌株中的罕见突变。

       材料和方法：
       从多药耐药嫌疑人身上采集痰样，并运至新德里结核病中心的中间参考实验室（IRL）。痰净化、DNA提取、扩增、杂交和线探针分析条带分析均按照制造商的说明进行。
       结果：
       在3000份具有可解释LPA条带的样品中，600份样品中观察到利福平对异烟肼和非异烟肼的耐药性。LPA检测rpoB基因最常见的突变是Ser516Leu（29.0%）。本研究报告的新突变包括密码子517处的CAG（Gin）到CAT（His）、密码子512处的AGC（Ser）-AGG（Arg）、密码子526处的ACA（Thr）到GCA（Ala）、密码子524处的TTG（Leu）-CTG（Leu）s。
       结论：
       LPA在rpoB基因中发现了高频率的罕见突变，突出了silico检测到的突变可能不会进一步导致表型抗性。

BACKGROUND:

The GenoType MTBDRplus, a commercial Line Probe Assay (LPA) kit from Hain Lifescience, Germany, is endorsed by India's RNTCP Program for diagnosis of DRTB cases among smear-positive sputum samples. Although the LPA has been studied in several laboratories, there is a wide variation in existing M. tuberculosis strains across the globe, and false results can occur due to the presence of unique genetic mutations in different settings.

AIM AND OBJECTIVE:

An attempt was made to carry out band pattern analysis using LPA and also to observe uncommon mutations in MDR strains.

MATERIALS AND METHODS:

Sputum samples were collected from MDR suspects and transported to intermediate reference laboratory (IRL) at New Delhi Tuberculosis Centre in Delhi. Sputum decontamination, DNA extraction, amplification, hybridization, and band pattern analysis of Line Probe assay strips was performed as per manufacturer's instructions.

RESULTS:

Among the 3000 samples with interpretable LPA strips, rifampicin drug resistance with or without isoniazid was observed in 600 samples. The most common mutation detected by LPA in the rpoB gene was Ser516Leu (29.0%). Novel mutations reported in this study include mutation from CAG (Gin) to CAT (His) at codon 517, AGC (Ser)-AGG (Arg) at codon 512, ACA (Thr) to GCA (Ala) at codon 526, TTG (Leu)-CTG (Leu)^s at codon 524.

CONCLUSION:

High frequencies of uncommon mutations in rpoB gene by LPA were observed, highlighting possibility of those in-silico detected mutations that may not impart phenotypic resistance further.

        https://www.sciencedirect.com/science/article/abs/pii/S0019570716302414?via%3Dihub