铜绿假单胞菌核酸酶MutL介导的DNA损伤导致持留菌形成从而暂时性抵抗双链RNA噬菌体的机制研究

细菌对噬菌体抵抗机制的研究不仅丰富了我们对噬菌体-宿主相互作用的认识，还诞生了CRISPR-Cas等分子生物学工具。目前，普遍认为细菌抵抗（resistant）噬菌体的机制是通过遗传元件或遗传变异介导的，还没有研究报道细菌可通过改变自身状态而暂时耐受（tolerate）噬菌体感染。基于前期铜绿假单胞菌抵抗噬菌体PaoP5的机制研究，我们设计了一组包含dsRNA噬菌体，且能充分抑制抵抗菌出现的噬菌体鸡尾酒。然而意外发现，随着培养时间延长，平板上长出少数小菌落，且这些菌落再次培养时对噬菌体依旧敏感。据此，我们提出“耐受噬菌体的持留菌”的概念。通过预实验，我们发现其形成依赖于MutL介导的DNA双链断裂，且仅能暂时耐受dsRNA噬菌体。本课题拟采用分子生物学、基因组学和单细胞成像技术，研究耐受dsRNA噬菌体的持留菌的形成机制，揭示一种全新的细菌状态改变介导的噬菌体耐受机制。

The study of phage resistance mechanism not only enriches our understanding of phage host interaction, but also leads to the discovery of CRISPR-Cas and other biological tools. However, at present, all studies have reported the mechanism of bacterial resistance to phage through genetic elements or genetic variation, but we don’t know whether bacteria can temporarily tolerate phage infection through changing bacterial state. Based on the phage resistance mechanism of Pseudomonas aeruginosa, we designed a group of phage cocktails containing PaoP5, phiYY（dsRNA phage） and PaoP5-m1, which can fully inhibit the emergence of phage resistant mutant. However, after prolongation of culture time, a few small colonies formed. These colonies were slow growing, sensitive to dsRNA phage and heritable. Therefore, we propose the concept of "phage resistant persister" to describes these bacteria. In this project, molecular genetics, genomics and single cell imaging technology will be combined to study the formation and tolerance mechanism of dsRNA phage tolerant persisters. This study will reveal a new dsRNA phage tolerance mechanism mediated by DNA damage induced persisters.