结核分枝杆菌毒素-抗毒素系统对新疆结核分枝杆菌流行株持留生存的调控

The recent researches have found that the bacterial toxin-antitoxin system involved in the bacteria growth regulation under adverse conditions, made the bacteria into persister under environmental stress.By the MazF homologous protein identification to 7 assumed toxin-antitoxin system mazEF of Mycobacterium tuberculosis in Mycobacterium smegmatis, the results showed that the 4 MazF homologous protein of them are toxin - antitoxin system,the 3 of the 4 MazF homologous protein appear a bacteriostatic or bactericidal activity, and may be involved in the growth regulation of Mycobacterium tuberculosis under nutritional deficiencies and anaerobic conditions. Therefore,this research will take the dominating popular strains of Mycobacterium tuberculosis in Xin jiang and the established toxins - antitoxin system mazEF deletion mutant strains of Mycobacterium tuberculosis as the research objects, investigate their main biological characteristics，including bacteria morphology, the dyeing property, the genetic stability of the mazEF deletion mutant strains, the culture characteristics, the bacterial survival rate under Low oxygen conditions，etc. This research will also determine the strains intake and the apoptosis of infected macrophages after Mycobacterium tuberculosis and the mazEF deletion mutant strains infect macrophages. This research is to analyse and discuss the regulation of toxin - antitoxin system mazEF system of the major epidemic Mycobacterium tuberculosis strains for the persistent situation, and to provide a theoretical basis for clarifying the survival mechanism of persistent Mycobacterium tuberculosis, as well as to have important scientific value and social significance for effective prevention, control and eradication of tuberculosis.

近年研究发现，细菌的毒素-抗毒素系统参与不良条件下细菌生长调控，使细菌在应激条件下进入休眠状态。通过在耻垢分枝杆菌中对7个假定的结核分枝杆菌毒素-抗毒素系统mazEF中MazF同源蛋白鉴定，结果显示4个是毒素-抗毒素系统，其中的3个具有抑菌或杀菌活性，可能参与结核分枝杆菌在营养匮乏、低氧等条件下的生长调控。因此，该项目研究将对新疆主要流行结核分枝杆菌菌株、构建的结核分枝杆菌毒素-抗毒素系统 mazEF 系统缺失突变株为研究对象，研究它们的主要生物学特性，包括细菌的形态、染色、缺失突变株的遗传稳定性、培养特性、低氧情况下的细菌存活率；测定结核分枝杆菌、结核分枝杆菌mazEF 系统缺失突变株感染巨噬细胞后，巨噬细胞对菌株摄入量以及受感染巨噬细胞的凋亡率。比较分析和探讨研究结核分枝杆菌毒素-抗毒素系统 mazEF 系统对新疆主要流行的结核分枝杆菌菌株持留生存的调控作用，为阐明结核分枝杆菌持留生存机制提供理论依据，对有效防治控制和根除结核病具有重要的科学价值和社会价值。

该项目研究以新疆主要流行结核分枝杆菌菌株、以及构建相应的结核分枝杆菌毒素-抗毒素系统 mazEF 系统缺失突变株作为研究对象，检测缺失突变株的遗传稳定性、mazEF基因缺失株、野生株结核分枝杆菌主要生物学特性,包括形态、染色特性、培养特性、低氧情况下的细菌存活率等；检测mazEF 系统缺失株、野生株感染巨噬细胞后，巨噬细胞凋亡率。研究结果显示，缺失株与野生株相比，生长速度缓慢且细菌形态短小；缺失毒素-抗毒素mazEF6基因后存活率下降；mazEF 系统缺失突变株及野生株与对照组相比，凋亡率显著高，差异具有统计学意义（P < 0.05）；并且缺失株凋亡率高于野生株，差异具有统计学意义（P < 0.05）。上述结果表明MTB毒素-抗毒素系统 mazEF 对新疆主要流行株“北京/W系” MTB持留生存有调控作用，推测与增强MTB在巨噬细胞内外的存活率、及降低巨噬细胞凋亡率有关。