Antibiotic tolerance, bacterial persistence and ATP synthase / Tolérance aux antibiotiques, persistance bactérienne et ATP synthétase

抗生素耐药性现在是一个有据可查的全球问题。其他相关问题，如导致抗生素耐受性的细菌持久性和逃避宿主免疫防御等，人们了解较少。表面上对抗生素敏感的微生物的顽固性是感染、复发和疾病慢性表现的原因。所谓难以治疗的感染(例如，生物被膜、留置生物医学设备的感染、囊性纤维化患者的肺部感染以及牛乳腺炎等动物的反复感染)涉及的非典型细菌亚群不能被抗生素或宿主免疫系统有效地杀死。这种非典型细菌被称为小菌落变种(SCV)或Peristers，这取决于它们出现的机制。 众所周知的病原体金黄色葡萄球菌可以出现为SCV。与通常发现的正常对应物相比，SCV产生大量的保护性生物膜，对抗生素耐受，并能够隐藏和持续在宿主上皮细胞内。许多SCV在所谓的呼吸链上存在突变，这种突变会影响负责产生细菌生长所需能量的酶(ATP合成酶)。生长缓慢的细菌或休眠细胞对大多数抗生素都有耐受性，这些抗生素通常会杀死活跃生长的细菌。幸运的是，我们的研究小组已经确定了在番茄植物中发现的一种能够杀死金黄色葡萄球菌SCV的天然产品(番茄碱)。这种新型抗生素的靶标是三磷酸腺苷合成酶。这一发现指出了在SCV中观察到的残留ATP合成酶活性的一个重要功能。 与金黄色葡萄球菌SCV不同，其他持久者通常不是从突变中产生的。有大量证据表明，例如，大肠埃希氏菌的持久细胞来自于对压力的反应机制，如抗生素暴露引起的压力。导致大肠杆菌持久细胞的机制的一个例子涉及细菌肽的产生，这种细菌肽破坏了它自己的呼吸链和ATP的产生。这会导致对抗生素耐受的休眠细胞的产生，这一结果类似于在呼吸缺陷的金黄色葡萄球菌SCV中观察到的结果。根据我们目前对托马替丁如何作用于金黄色葡萄球菌SCV的了解，有理由认为持久者也可能对抑制细菌ATP合成酶的药物过敏。 这项拟议研究的目的是描述导致SCV和持久者形成的机制，证明ATP合酶对这些类型的细菌是必不可少的，并证明我们开发的针对金黄色葡萄球菌SCV的控制和预防措施可以扩展到其他类型的SCV和持久者。SCV的低残留但明显必需的ATP合成酶活性可能同样是革兰氏阳性或革兰氏阴性病原体的所有持久细胞的致命弱点。

Antibiotic resistance is now a well-documented problem worldwide. Other related issues such as bacterial persistence leading to antibiotic tolerance and evasion of the host immune defenses are less understood. Persistence by seemingly antibiotic-susceptible microorganisms is responsible for infection relapse and chronic manifestation of diseases. So-called difficult-to-treat infections (e.g., biofilms, infections of indwelling biomedical devices, lung infections in cystic fibrosis patients and recurrent infections in animals such as bovine mastitis) involve subpopulations of atypical bacteria that are not efficiently killed by antibiotics or the host immune system. Such atypical bacteria are called small-colony variants (SCVs) or Peristers depending on the mechanism by which they appear. The well-known pathogen Staphylococcus aureus can appear as a SCV. Compared to their commonly found normal counterparts, SCVs produce high amounts of protective biofilms, are tolerant to antibiotics and are capable of hiding and persisting within host epithelial cells. Many SCVs have mutations in the so-called respiratory chain that affect the enzyme (ATP synthase) responsible for producing the energy necessary for growth of the bacteria. Slow growing bacteria or dormant cells are tolerant to most antibiotics that usually kill actively growing bacteria. Fortunately, our research group has identified a natural product found in the tomato plant (tomatidine) that is able to kill S. aureus SCVs. The target of that new class of antibiotics is the ATP synthase. This discovery points to an essential function for the residual ATP synthase activity observed in SCVs. Unlike the S. aureus SCVs, other Persisters do not usually arise from mutations. There is abundant evidence that for example Escherichia coli persister cells arise from active mechanisms in response to a stress such as that caused by antibiotic exposure. An example of the mechanisms leading to E. coli persister cells involves the production of a bacterial peptide that sabotages its own respiratory chain and ATP production. This results in the generation of dormant cells tolerant to antibiotic action, a consequence similar to that observed in respiratory-deficient S. aureus SCVs. Based on our current understanding on how tomatidine acts on S. aureus SCVs, it is reasonable to suggest that Persisters may also be hypersusceptible to drugs inhibiting bacterial ATP synthase. The objectives of the proposed research is to characterize the mechanisms leading to the formation of SCVs and Persisters, to demonstrate that the ATP synthase is essential for these types of bacteria, and to demonstrate that the control and prevention measures we developed for S. aureus SCVs can be extended to other types of SCVs and Persisters. The low residual but apparently essential ATP synthase activity of SCVs might likewise represents the Achilles' heel of all persister cells from Gram positive or Gram negative pathogens.