Function and inhibition of multidrug efflux systems in Campylobacter

 描述（由申请方提供）：空肠弯曲菌是全球腹泻的主要细菌原因，对临床重要抗生素的耐药性越来越强。在弯曲杆菌中涉及抗生素耐药性的已知机制中，多药外排系统CmeABC（一种RND型外排转运蛋白）是一个重要的参与者，并赋予对结构多样的抗生素和有毒化合物的耐药性。此外，CmeABC在胆汁抗性中起关键作用，并且对于肠道中的弯曲杆菌定殖是必需的。最近发现，作为氧化应激反应调节剂的CosR可作为cmeABC的阻遏物并调节cmeABC在氧化应激反应中的表达。这些结果有力地表明，氧化应激是一个以前未确定的生理信号调节CmeABC的功能和CosR介导的反应，氧化应激可能是重要的适应C。空肠，一种在传播和感染宿主期间经历氧化应激的微需氧生物。尽管有强有力的初步证据，氧化应激和CosR-CmeABC途径之间相互作用的详细机制以及这种相互作用在抗生素和氧化应激抗性中的作用仍然未知。为了控制耐药弯曲杆菌，我们最近开发了一种新的策略，利用抗cmeABC肽核酸（PNA）来抑制cmeABC的表达。发现这种方法在使弯曲杆菌对抗生素敏感方面是有效的，这表明靶向cmeABC的反义PNA是对抗抗生素抗性弯曲杆菌的有希望的方法。本申请基于这些令人兴奋的发现，并将追求两个特定目的：i）确定氧化应激与CosR-CmeABC途径相互作用的机制，并定义这种相互作用在弯曲杆菌适应抗生素和氧化应激中的作用，和ii）确定抗-cmeABC PNA在动物模型中预防抗生素抗性突变体的出现和增强抗生素对弯曲杆菌的作用。该提案在概念上和技术上都是创新的，因为它解决了氧化应激反应和抗生素外排系统之间界面的一个新兴主题，并通过靶向CmeABC开发了一种新的抗弯曲杆菌方法。拟议的工作将显着推进氧化应激传感和抗性是细菌病原体中抗生素外排系统的常见生理功能的概念，并将开发一种有效的手段来扩展现有抗生素对耐药弯曲杆菌的效用。此外，该项目建立的技术平台可能适用于控制其他抗药性病原体。

 DESCRIPTION (provided by applicant): Campylobacter jejuni is a leading bacterial cause of diarrhea worldwide and is increasingly resistant to clinically important antibiotics. Among the known mechanisms involved in antibiotic resistance in Campylobacter, the multidrug efflux system CmeABC (an RND-type efflux transporter) is a significant player and confers resistance to structurally diverse antibiotics and toxic compounds. Additionally, CmeABC plays a critical role in bile resistance and is essential for Campylobacter colonization in the intestinal tract. Mot recently it was found that CosR, a response regulator for oxidative stress, serves as a repressor for cmeABC and modulates the expression of cmeABC in response to oxidative stress. These results strongly suggest that oxidative stress is a previously unidentified physiological signal modulating the function of CmeABC and the CosR-mediated response to oxidative stress may be important for the adaptation of C. jejuni, a microaerobic organism that experiences oxidative stress during transmission and infection of a host. Despite strong preliminary evidence, the detailed mechanisms of the interplay between oxidative stress and the CosR- CmeABC pathway and the role of this interaction in antibiotic and oxidative stress resistance are still unknown. To control antibiotic-resistant Campylobacter, we recently developed a novel strategy that utilizes anti-cmeABC peptide nucleic acid (PNA) to inhibit the expression of cmeABC. This approach was found to be effective in sensitizing Campylobacter to antibiotics, suggesting that antisense PNA targeting cmeABC is a promising approach for combating antibiotic-resistant Campylobacter. This application is based on these exciting findings and will pursue two Specific Aims i) to determine the mechanisms by which oxidative stress interacts with the CosR-CmeABC pathway and define the role of this interaction in Campylobacter adaptation to antibiotic and oxidative stresses, and ii) to determine the efficiency of anti-cmeABC PNA in preventing the emergence of antibiotic resistant mutants and potentiating antibiotics against Campylobacter in animal models. The proposal is innovative both conceptually and technically as it addresses an emerging theme at the interface between oxidative stress response and antibiotic efflux systems and develops a novel anti-Campylobacter approach by targeting CmeABC. The proposed work will significantly advance the concept that oxidative stress sensing and resistance is a common physiological function of antibiotic efflux systems in bacterial pathogens and will develop an effective mean to extend the utility of existing antibiotic against drug-resistant Campylobacter. Furthermore, the technical platform established in this project can be potentially adapted for the control of other antibiotic-resistant pathogens.