DST/NRF Centre of Excellence for Biomedical TB Research, School of Pathology, Faculty of Health Sciences, University of the Witwatersrand and the National Health Laboratory Service, Johannesburg, South Africa.
DST/NRF Centre of Excellence for Biomedical TB Research, School of Pathology, Faculty of Health Sciences, University of the Witwatersrand and the National Health Laboratory Service, Johannesburg, South Africa
J Clin Microbiol. 2017 Dec;55(12):3384-3394. doi: 10.1128/JCM.01111-17. Epub 2017 Sep 20.
Molecular diagnostics have revolutionized the management of health care through enhanced detection of disease or infection and effective enrollment into treatment. In recognition of this, the World Health Organization approved the rollout of nucleic acid amplification technologies for identification of using platforms such as GeneXpert MTB/RIF, the GenoType MTBDR line probe assay, and, more recently, GeneXpert MTB/RIF Ultra. These assays can simultaneously detect tuberculosis infection and assess rifampin resistance. However, their widespread use in health systems requires verification and quality assurance programs. To enable development of these, we report the construction of genetically modified strains of that mimic the profile of on both the GeneXpert MTB/RIF and the MTBDR line probe diagnostic tests. Using site-specific gene editing, we also created derivatives that faithfully mimic the diagnostic result of rifampin-resistant , with mutations at positions 513, 516, 526, 531, and 533 in the rifampin resistance-determining region of the gene. Next, we extended this approach to other diseases and demonstrated that a gene sequence can be introduced into to generate a positive response for the SCC probe in the GeneXpert SA Nasal Complete molecular diagnostic cartridge, designed for identification of methicillin-resistant These biomimetic strains are cost-effective, have low biohazard content, accurately mimic drug resistance, and can be produced with relative ease, thus illustrating their potential for widespread use as verification standards for diagnosis of a variety of diseases.
分子诊断通过提高疾病或感染的检测能力以及有效地进行治疗干预，彻底改变了医疗保健的管理方式。有鉴于此，世界卫生组织批准了核酸扩增技术的推广，以使用 GeneXpert MTB/RIF、GenoType MTBDR 线探针检测等平台来识别结核分枝杆菌，最近还批准了 GeneXpert MTB/RIF Ultra。这些检测方法可同时检测结核感染并评估利福平耐药性。然而，要在卫生系统中广泛使用这些方法，需要进行验证和质量保证计划。为了能够开展这些工作，我们报告了构建与 GeneXpert MTB/RIF 和 MTBDR 线探针诊断检测结果一致的结核分枝杆菌基因修饰株。通过定点基因编辑，我们还构建了突变位于利福平耐药决定区 513、516、526、531 和 533 位的衍生菌株，可忠实地模拟利福平耐药结核分枝杆菌的诊断结果。接下来，我们将这种方法扩展到其他疾病，并证明可以将 基因序列引入 中，以在用于鉴定耐甲氧西林金黄色葡萄球菌的 GeneXpert SA Nasal Complete 分子诊断试剂盒中产生 SCC 探针的阳性反应。这些生物模拟菌株具有成本效益、低生物危害含量、可准确模拟耐药性，并且相对容易生产，因此展示了它们作为各种疾病诊断验证标准的广泛应用潜力。
