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

抗生素耐药性现在是世界范围内一个有据可查的问题。其他相关问题，如细菌持久性导致抗生素耐受性和逃避宿主免疫防御尚不清楚。表面上对抗生素敏感的微生物的持续存在是导致感染复发和疾病慢性表现的原因。所谓的难治性感染（如生物膜感染、留置生物医学设备感染、囊性纤维化患者肺部感染和动物复发性感染，如牛乳腺炎）涉及不能被抗生素或宿主免疫系统有效杀死的非典型细菌亚群。这种非典型细菌根据其出现的机制被称为小菌落变异（scv）或perister。众所周知的病原体金黄色葡萄球菌可以表现为SCV。与常见的正常细胞相比，scv产生大量的保护性生物膜，对抗生素具有耐受性，并且能够隐藏并持续存在于宿主上皮细胞中。许多scv在所谓的呼吸链中有突变，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的细胞对大多数抗生素都有耐受性，而抗生素通常会杀死生长活跃的细菌。幸运的是，我们的研究小组已经在番茄植物中发现了一种能够杀死金黄色葡萄球菌scv的天然产物（番茄碱）。这种新型抗生素的目标是ATP合酶。这一发现指出了在scv中观察到的残余ATP合酶活性的基本功能。与金黄色葡萄球菌scv不同的是，其他持续性病毒通常不是由突变产生的。例如，有大量证据表明，大肠杆菌持久性细胞是在对抗生素暴露等压力作出反应的积极机制中产生的。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，这种细菌肽破坏了其自身的呼吸链和ATP的产生。这导致产生耐受抗生素作用的休眠细胞，类似于在呼吸缺陷金黄色葡萄球菌scv中观察到的结果。根据我们目前对番茄碱如何作用于金黄色葡萄球菌scv的了解，我们有理由认为，persistent也可能对抑制细菌ATP合酶的药物敏感。本研究的目的是描述导致scv和持续型细菌形成的机制，证明ATP合酶对这些类型的细菌至关重要，并证明我们针对金黄色葡萄球菌scv开发的控制和预防措施可以扩展到其他类型的scv和持续型细菌。scv残留的ATP合成酶活性低，但显然是必需的，这可能同样代表了所有来自革兰氏阳性或革兰氏阴性病原体的持久性细胞的致命弱点。

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.