Evolutionary and Functional Significance of Novel Mutations in MDR-XDR TB

描述(申请人提供)：去年报告的活动性肺结核病例超过900万例，死于结核病的病例超过100万例。耐药结核分枝杆菌的发展正在增加，并威胁到结核病控制努力。全球耐药结核病和诊断联合会(GCDD)已经确定了100多个(在419个中)耐药结核分枝杆菌分离株，没有已知的导致耐药性的突变。此外，GCDD还在其四个研究地点(印度、摩尔多瓦、菲律宾和南非)确定了七个不同的区域基因-表型关系 一线和二线药物，表明结核分枝杆菌在每个部位都采取了不同的进化路线来产生耐药性。这也可能意味着结核分枝杆菌在这些研究地点使用不同的产生耐药性的机制，因此可能很快就需要为每个进化路径/研究地点使用一套新的抗结核药物。虽然不同进化路径的出现可能是由于抗结核药物的可获得性不同，但这种特殊性的出现令人震惊，如果持续下去，结核分枝杆菌可能演变为不同的总耐药性状态(TDR-TB)，导致一种流行病，需要针对区域的治疗和公共卫生战略。该项目旨在通过对新发现的与耐药相关的基因和调控元件进行电子功能表征来揭示以前未知的耐药机制。这将解释那些无法解释的抵抗案例。功能表征将有助于识别涉及这些新突变的每种药物的耐药机制，识别可能导致对多种药物产生耐药性的“Keystone”突变，并识别功能相互作用网络中可作为新药物靶点的中心位点。该项目还将对所有400多个GCDD Mtb基因组进行系统发育分析。新的和已知突变的功能表征与耐药性的进化分析相结合，将回答不同的耐药进化路径和机制是否成为现实。如果是这样的话，该模型可以扩展到预测未来的区域演化趋势。试验性重组技术将用于在实验室确认导致耐药性的Keystone突变的重要性。

DESCRIPTION (provided by applicant): Last year over 9 million cases of active TB and 1 million deaths due to TB were reported. The development of drug-resistant Mtb is increasing and threatens TB control efforts. Global Consortium for Drug-resistant TB and Diagnostics (GCDD) has identified over 100 (out of 419) resistant Mtb isolates without known resistance-causing mutations. Additionally, GCDD has identified distinct regional genotypic-phenotypic relationships at its four study sites (India, Moldova, the Philippines, and South Africa) for seven first and second line drugs, suggesting that Mtb is taking a different evolutionary route to resistance at each site. This could also mean that Mtb is using different mechanisms for producing resistance at these study sites and therefore could soon require new set of anti-TB drugs for each evolutionary path/study site. While the emergence of the distinct evolutionary paths are probably due to variable availability of anti-TB drugs, the emergence of this distinctiveness is alarming and if continued, Mtb can evolve into different states of total drug resistance (TDR- TB) causing an epidemic that requires region-specific treatment and public health strategy. This project aims to uncover previously-unknown mechanisms of drug resistance through in silico functional characterization of newly identified gene and regulatory elements associated with drug resistance. This will explain the unexplained cases of resistance. The functional characterization will allow the identification of the mechanism of resistance for each drug involving these novel mutations, identification of "keystone" mutations that can cause resistance to multiple drugs, and identify central loci in the functional interaction network that can serve as new drug targets. This project will also perform a phylogenetic analysis of all 400+ GCDD Mtb genomes. The combination of functional characterization of novel and known mutations and the evolutionary analysis of drug resistance will answer whether distinct evolutionary paths and mechanisms of resistance are a reality or not. If so, the model can be extended to predict future regional evolutionary trends. Experimental recombinant technology will be used to confirm the significant the keystone mutations responsible for resistance in the laboratory.