在抗细菌感染中，细菌因适应性进化导致耐药已显著占据上风。前期发现，抗生素胁迫下，携带耐药质粒的大肠埃希菌连续多次传代后对抗生素的MIC、质粒拷贝数、质粒稳定性和质粒转移能力均显著增高，机制尚不清楚。我们假设，在选择压力胁迫下，细菌与质粒相互适应性进化并导致其耐药能力增强，潜在的分子机制可能涉及细菌和质粒的基因突变及差异表达。本课题拟研究：①抗生素胁迫下，携带耐药质粒的大肠埃希菌连续多次传代后的生物学变化；②高通量测序检测多次传代后细菌全基因组和质粒的基因突变和突变导致的结果；③转录组测序检测多次传代后细菌和质粒的差异表达基因和sRNA；④筛选调节适应性变化的差异表达基因和sRNA，过表达和基因敲除这些基因，明确其对适应性进化的调控作用，阐明抗生素作用下细菌和质粒适应性进化和导致耐药能力增强的分子机制，为阻断耐药基因通过质粒水平转移、为抗菌药物研发提供新靶点和丰富细菌耐药发生的理论提供依据。

Antibiotic resistance is a serious threat to human health and new ways to combat it are urgently needed. The worldwide increase in antibiotic resistance has motivated numerous studies aimed at understanding the phenotypic and genotypic evolution of antibiotic resistance. A key aspect of bacterial survival is the ability to evolve while migrating across spatially varying environmental challenges. In our preliminary study, We found that under the treatment of antibiotic, the copy number of plasmid, plasmid stability with and without antibiotic, plasmid transfer frequency and minimal inhibitory concentration of E. coli SM10λπ (pUCP24T) with continuously subculture had been enhanced significantly. However, little is known about the mechanisms. We speculated that antibiotic selection drove plasmid-host compensator revolution and adaptive evolution, resulting in increased resistance ability, with a potential molecular mechanism relating to genes mutations and differential expression in host and plasmid. In this project, to illuminate the mechanisms of antibiotic resistance strengthened by adaptive evolution of E. coli and plasmid under antibiotic stress, we will: ①obverse the biology changes of E. coli harboring resistance plasmids after continuously subculture under the treatment of antibiotic, the biology changes including minimal inhibitory concentration, the fitness cost imposed by resistance plasmids on E. coli hosts, the copy number of the resistance plasmids, plasmid stability with and without antibiotic, and plasmid transfer frequency; ②explore the adaptive adaptation to this fitness cost by chromosomal and/or plasmid mutations using whole-genome sequencing; ③identify differential expression genes and small non-coding RNA (sRNA) by RNA sequencing; ④confirm the functions of those candidate differential expression genes and sRNA regulating the adaptive evolution by over-expression and knockout technology. We hope to provide rationale strategies to blocking-up horizontal gene transfer, ameliorate antibiotic resistance mediated by plasmids, provide the drug target for novel antibiotics discovery studies and enrich the theory of antibiotic resistance.