MAS-PCR Rapid Screening Test for Multidrug-Resistant Tuberculosis (MDR-TB)

描述（由申请人提供）：结核分枝杆菌对用于治疗结核感染的药物的耐药性已成为一个重大问题，并且耐多药结核病（MDR-TB）的存在在全世界范围内不断增加。耐多药结核病被定义为对利福平（RIF）和异烟肼（INH）这两种用于治疗结核病的一线药物产生耐药性。目前用于检测耐多药结核病的传统方法需要在培养基中培养结核菌，然后对用于治疗的药物进行测试。这是费力的，需要数周到数月的时间才能知道结果，这会延迟使用更昂贵的二线药物进行有效治疗。已经确定了与耐多药结核病相关的基因突变，并开发了几种快速聚合酶链反应（PCR）检测方法来筛查耐多药结核病。这些基于聚合酶链反应的检测提供了更快速的结果，但它们昂贵，需要专门的实验室设备，而结核病高负担国家的大多数实验室无法轻易负担得起，这些国家大多是资源有限的发展中国家。拟议研究的主要目的是测试一种简单、廉价的多重等位基因特异性聚合酶链反应（MAS-PCR）技术，该技术可用于在高负担资源有限的国家快速筛查耐多药结核病。MAS-PCR检测已经在Mahidol大学医学院微生物学系耐药结核病（DRTB）实验室和泰国曼谷的单一实验室环境中开发和测试。拟议的研究由两部分组成，旨在确认该测试可用于多个实验室，并可在这些实验室中提供准确和一致的结果。在研究的第一部分，已知对RIF和INH敏感或耐药的结核病模拟样本将被送往泰国和中国的五个实验室。实验室不知道这些样本是敏感的还是耐药的。每个实验室的检测结果将与样本的已知状态进行比较，以及与另一种更昂贵的基于PCR的检测（基因型MTBDR Plus）的结果进行比较。在研究的第二部分，中国和泰国的实验室将接收来自其医院确诊结核病患者的痰液样本，并将用MAS-PCR检测进行筛查，以确定他们是否可能是耐多药结核病病例。将通过在DRTB实验室（位于Alarraj Hospital）使用基因型MTBDR Plus对来自这些标本的DNA样本进行复检来确认当地标本的这些检测结果。如果MAS-PCR筛查测试在泰国和中国的实验室成功实施，它将成为其他高负担资源有限国家进一步测试和实施耐多药结核病常规筛查的候选方法。

DESCRIPTION (provided by applicant): Resistance of Mycobacterium tuberculosis to drugs used for treatment of TB infection has become a significant problem, and the existence of multidrug-resistant tuberculosis (MDR-TB) is increasing throughout the world. MDR-TB is defined as resistance to both rifampicin (RIF) and isoniazid (INH), two of the first-line drugs used for treatment of TB. The conventional method used today to detect MDR-TB requires growing the TB organism in culture and then testing it against the drugs used for treatment. This is laborious and requires weeks to months before the result is known, which delays starting treatment with more expensive second-line drugs that will be effective. Genetic mutations associated with MDR-TB have been identified, and several rapid polymerase chain reaction (PCR) tests have been developed to screen for MDR-TB. These PCR-based tests provide more rapid results, but they are expensive and require specialized lab equipment that cannot be easily afforded by most labs in high-burden TB countries, which are mostly resource-limited developing countries. The primary objective of the proposed research is to test a simple and inexpensive multiplex allele-specific polymerase chain reaction (MAS-PCR) technique that can be used for rapid screening of MDR-TB in high burden resource-limited countries. The MAS-PCR test has already been developed and tested in a single laboratory environment at the Drug-Resistant Tuberculosis (DRTB) Laboratory, Department of Microbiology, Faculty of Medicine, Siriraj Hospital, Mahidol University, and Bangkok, Thailand. The proposed study consists of two parts designed to confirm that this test is feasible for use in multiple labs and can provide accurate and consistent results across these labs. For the first part of the study, simulated samples of TB with known susceptibility or resistance to RIF and INH will be sent to each of five labs in Thailand and China. The labs will not know whether the samples are susceptible or resistant. Test results from each lab will be compared to the known status of the sample, as well as to the result from another more expensive PCR-based test, the Genotype MTBDR Plus. In the second part of the study, the labs in China and Thailand will receive sputum samples from current patients at their hospitals with confirmed TB and will screen them with the MAS-PCR test to determine whether they might be MDR-TB cases. The results from these tests on local specimens will be confirmed by retesting on the DNA samples from these specimens with Genotype MTBDR Plus at the DRTB Laboratory at Siriraj Hospital. If the MAS-PCR screening test is successfully implemented in the labs in Thailand and China, it will be a candidate for further testing in other high-burden resource limited countries and implementation for routine screening for MDR-TB.