青蒿琥酯和EDTA协同逆转沙门菌对黏菌素耐药的分子机制

沙门菌耐药给养禽业带来严重经济损失。黏菌素被认为是治疗G-菌感染的最后一道防线，但随着细菌对黏菌素耐药日益增加，而新型抗菌药物的研发相对滞后，促使联合用药作为一种新的用药策略来逆转沙门菌的黏菌素耐药性成为研究热点。本研究的前期将抗疟疾药物青蒿琥酯、金属离子螯合剂EDTA和黏菌素联用，发现三药联用后可显著逆转沙门菌对黏菌素的耐药性（MIC下降32-2000倍），但其具体分子机制尚不清晰。因此，本研究将首先用棋盘法测定联合用药的分级抑菌浓度FIC和最小逆转浓度MRC，并对菌株的表型变化进行研究。接着将分别对类脂A的修饰情况、外膜通透性、黏菌素的胞内蓄积、内膜通透性、转录组、代谢组、ATP活性、胞内ROS的蓄积进行分析，从而确定联合用药的外膜靶点、内膜靶点和胞内靶点。上述研究将为寻找一种新的、低剂量、高活性、低毒副作用的控制耐药菌感染的联合用药策略提供理论依据。

Antibiotic resistance of Salmonella have caused serious economic losses in the poultry industry. Colistin has been re-emerged as a last-resort therapeutic option to treat G- bacteria infections. The rapid emergence of colistin-resistant bacterial pathogens, and the fact that the development of new antimicrobial drugs is still lagging far behind the rising demand have made the new therapeutic strategy of drug combinations become a hot topic. Thus, drug combinations of artesunate, EDTA and colistin were previously investigated against Salmonella, and we found that artesunate and EDTA could synergistically reverse the colistin resistance of Salmonella with a markedly decreased MIC of colistin (32-2000 fold) after three-drug combination. To further elucidate the molecular mechanisms of drug combination, the microdilution methods will be firstly used to determine the FIC and MRC, and the phenotypic changes will be investigeted in this study. Then, the outer membrane target, the inner membrane target and intracellular target will be further explored through the analysis of modification of lipid A, permeability of outer membrane, intracellular accumulation of colistin, permeability of inner membrane, transcriptome, metabolome, ATP activity, accumulation of ROS, etc. Our findings will provide a theoretical basis for a new, low-toxic and side-effect medication strategy of drug combination, which is less dosage but higher antimicrobial activity.