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首页> 外文期刊>Journal of Clinical Microbiology >Development of Multiplex Assay for Rapid Characterization of Mycobacterium tuberculosis
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Development of Multiplex Assay for Rapid Characterization of Mycobacterium tuberculosis

机译：快速鉴定结核分枝杆菌的多重检测方法的建立

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We have developed a multiplex assay, based on multiplex ligation-dependent probe amplification (MLPA), that allows simultaneous detection of multiple drug resistance mutations and genotype-specific mutations at any location in the Mycobacterium tuberculosis genome. The assay was validated on a reference panel of well-characterized strains, and the results show that M. tuberculosis can be accurately characterized by our assay. Eighteen discriminatory markers identifying drug resistance (rpoB, katG, inhA, embB), members of the M. tuberculosis complex (16S rRNA, IS6110, TbD1), the principal genotypic group (katG, gyrA), and Haarlem and Beijing strains (ogt, mutT2, mutT4) were targeted. A sequence specificity of 100% was reached for 16 of the 18 selected genetic targets. In addition, a panel of 47 clinical M. tuberculosis isolates was tested by MLPA in order to determine the correlation between phenotypic drug resistance and MLPA and between spoligotyping and MLPA. Again, all mutations present in these isolates that were targeted by the 16 functional probes were identified. Resistance-associated mutations were detected by MLPA in 71% of the identified rifampin-resistant strains and in 80% of the phenotypically isoniazid-resistant strains. Furthermore, there was a perfect correlation between MLPA results and spoligotypes. When MLPA is used on confirmed M. tuberculosis clinical specimens, it can be a useful and informative instrument to aid in the detection of drug resistance, especially in laboratories where drug susceptibility testing is not common practice and where the rates of multidrug-resistant and extensively drug resistant tuberculosis are high. The flexibility and specificity of MLPA, along with the ability to simultaneously genotype and detect drug resistance mutations, make MLPA a promising tool for pathogen characterization.
机译：我们已经开发出一种基于多重连接依赖探针扩增（MLPA）的多重检测方法，该方法可以同时检测结核分枝杆菌基因组中任何位置的多个耐药突变和基因型特异性突变。该方法在特征明确的菌株参考面板上得到验证，结果显示为 M。我们的检测方法可以准确地鉴定结核病。识别耐药性的18种歧视性标记（ rpoB ， katG ， inhA ， embB ），成员M.结核病复合物（16S rRNA，IS 6110 ，TbD1），主要基因型组（ katG ， gyrA ），哈莱姆和以北京菌株（ ogt ， mutT2 ， mutT4 ）为研究对象。对于18个选定的遗传靶标中的16个，序列特异性达到100％。此外，一组47位临床 M。 MLPA检测结核病分离株，以确定表型耐药与MLPA之间的相关性，以及Spoligotyping与MLPA之间的相关性。再次，鉴定了这些分离物中存在的所有被16个功能探针靶向的突变。 MLPA在71％的已确定的利福平耐药菌株和80％的表型异烟肼耐药菌株中检测到了耐药相关突变。此外，MLPA结果与spoligotypes之间存在完美的关联。当在已确认的 M上使用MLPA时。结核病临床标本，它可以作为一种有用的信息工具，有助于检测耐药性，尤其是在实验室中，药物敏感性测试并不常见，且耐多药和广泛耐药结核病的比率很高高。 MLPA的灵活性和特异性，以及同时进行基因分型和检测耐药性突变的能力，使MLPA成为病原体鉴定的有前途的工具。

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