Plasmid-Bacteria Coevolution Promotes the Spread of Antibiotic Resistance

项目摘要/摘要 许多领先的人类健康组织，如世界卫生组织和疾病中心 美国疾病控制和预防中心(CDC)宣布，增加的细菌病原体是 对多种抗生素的抗药性是一个重大的人类健康危机。这些多重耐药的出现 (MDR)病原菌很大程度上是由于质粒介导的水平基因共享抗性基因 调职。细菌质粒是一种可移动的遗传元件，可以对多种抗生素产生耐药性， 包括那些被认为是“最后手段的毒品”。我们的长期目标是帮助发展 可以通过洞察共同进化来减缓抗生素耐药性传播的策略 允许细菌提高新获得的多药耐药质粒的持久性的过程。新收购的 在缺乏抗生素的情况下，耐药质粒通常不会持续存在，但我们和其他人已经证明 细菌宿主、质粒或两者的单一突变可以迅速提高这种持久性。我们和其他人 还确定了染色体编码的辅助解旋酶的关键突变。质粒-解旋酶相互作用 因此，细菌中的多药耐药可能是细菌病原体保留新获得的多药耐药质粒能力的关键。 不幸的是，解释这些突变对质粒的积极影响的分子机制 坚持不懈是未知的。重要的是，我们还首次表明，这些突变预先适应了 细菌与后来获得的其他多药耐药质粒结合，从而增强其持久性(参考 到增加的质粒透过率)。这表明，具有更强的耐受性的细菌可以 作为多个多药耐药质粒的稳定储存库，最终产生具有扩展的 抗性基因。这种可能性从未被测试过。利用分子技术，实验进化 和数学模型，我们建议检验以下假设：(I)染色体突变可以预激 使细菌适应其他质粒，导致更大的质粒透过性；(Ii)质粒透过性可以 在细菌物种中扩大抗生素耐药性特征的范围；以及(Iii)辅助解旋酶是 与新获得的多药耐药质粒在广泛范围的细菌病原体中的持久性有关。这 将通过实现以下具体目标来完成：(1)检测(I)增加的质粒的通用性 宿主/质粒共同进化后的通透性，以及(Ii)作为宿主机制的解旋酶突变 适应新的多药耐药质粒；(2)确定质粒持久性和 对出现扩张性耐药的容忍性；(3)测定分子 辅助解旋酶突变导致的质粒成本改善的机制。如果我们的 假设得到了我们的数据的支持，稳定一个质粒的突变可能会导致持久性的提高 并扩大同一细胞中的抗性基因库。我们的发现将有助于 开发新的疗法，旨在减缓抗生素耐药性在细菌病原体中的传播。

PROJECT SUMMARY/ABSTRACT Many leading human health organizations such as the World Health Organization and the Centers for Disease Control and Prevention (CDC) have declared that the increased prevalence of bacterial pathogens that are resistant to multiple antibiotics is a significant human health crisis. The emergence of these multi-drug resistant (MDR) pathogens is largely due to the sharing of resistance genes by plasmid mediated horizontal gene transfer. Bacterial plasmids are mobile genetic elements that can confer resistance to a variety of antibiotics, including those that are considered to be “drugs of last resort”. Our long-term goal is to aid the development of strategies that can slow the spread of antibiotic resistance by gaining insight into the co-evolutionary processes that allow bacteria to improve the persistence of newly acquired MDR plasmids. Newly acquired resistance plasmids often do not persist in the absence of antibiotics, but we and others have shown that single mutations in the bacterial host, the plasmid, or both can rapidly improve this persistence. We and others also identified critical mutations in chromosomally encoded accessory helicases. Plasmid-helicase interactions in bacteria may therefore be key to the ability of bacterial pathogens to retain newly acquired MDR plasmids. Unfortunately, the molecular mechanisms that explain the positive effects of these mutations on plasmid persistence are unknown. Importantly, we also showed for the first time that these mutations pre-adapt the bacteria to other MDR plasmids that they acquire later in time, leading to their enhanced persistence (referred to as increased plasmid permissiveness). This suggests that bacteria with increased permissiveness can serve as stable repositories for multiple MDR plasmids, eventually generating strains with an expanded arsenal of resistance genes. This possibility has never been tested. Using molecular techniques, experimental evolution and mathematical modeling, we propose to test the following hypotheses: (i) chromosomal mutations can pre- adapt bacteria to other plasmids, leading to greater plasmid permissiveness; (ii) plasmid permissiveness can expand the spectrum of antibiotic resistance traits within a bacterial species; and (iii) accessory helicases are linked to the persistence of newly acquired MDR plasmids across a wide spectrum of bacterial pathogens. This will be done through achieving the following Specific Aims: (1) Test the generality of (i) increased plasmid permissiveness after host/plasmid coevolution, and (ii) helicase mutations as a mechanism of host adaptation to novel MDR plasmids.; (2) determine the effects of plasmid persistence and permissiveness on the emergence of expanded drug resistance; (3) determine the molecular mechanism of plasmid cost amelioration resulting from mutations in accessory helicases. If our hypotheses are supported by our data, mutations that stabilize one plasmid could lead to improved persistence of other plasmids, and expand the arsenal of resistance genes in the same cell. Our findings will aid the development of new therapies aimed at slowing down the spread of antibiotic resistance in bacterial pathogens.!