Disruption of cytoplasmic membrane-associated functions in Staphylococcus aureus by epicatechin gallate

金黄色葡萄球菌通常是皮肤和粘膜的定植菌，但当细菌有机会进入人体时，会引起一系列感染。它还会产生一系列导致食物中毒的毒素：如果食品工人携带这种细菌，它就会进入食物链，并且由于其耐盐能力，即使在含盐的食物中也能存活。葡萄球菌通过获得耐药机制显示出对常用抗生素的显著抵抗能力。特别是，它们可以获得mecA基因，其产物青霉素结合蛋白（PBP） 2a使其能够在高浓度青霉素（如甲氧西林和苯氧西林）存在下存活。这种耐甲氧西林金黄色葡萄球菌（MRSA）对进入医院的患者构成威胁，特别是大手术患者，并且越来越多地在社区获得。这些感染曾一度局限于人类，但近年来在伴侣动物，特别是狗和猫中也发现了这些感染；这些细菌极有可能最初是从人类身上获得的。动物和人类MRSA感染难以治疗，迫切需要新的治疗和控制方法，特别是那些降低耐药性出现率的方法。我们已经确定，绿茶的主要成分之一，一种相对不稳定的物质，被称为表儿茶素没食子酸酯，或ECg，以一种对动物和人类有益的方式改变金黄色葡萄球菌的特性。尽管ECg对MRSA的生长速度没有影响，但它可以将其对青霉素的耐药性降低500倍，这增加了它可以与传统抗生素一起使用来治疗感染的可能性。由于ECg修饰而不是杀死目标细菌，它不会像目前使用的“杀菌”药物那样施加极端的选择压力，并可能减缓对治疗组合的耐药性的出现。此外，我们发现心电图可以阻止毒素和酶的分泌，这些毒素和酶会使生物体造成组织损伤和感染，并导致食物中毒的症状。ECg还可以防止生物膜的形成：这些细菌的复杂组合在界面上形成，是在组织入侵之前宿主表面和导管和假体等设备定植的必要先决条件。不幸的是，由于其不稳定的性质和适度的活性，天然存在的化学物质不适合用于治疗目的，我们正在制造具有更有吸引力的心电图衍生物。这些尝试由于对ECg如何发挥其抗葡萄球菌活性的不完全理解而受到阻碍。我们开始了解到它具有如此广泛的特性，因为它插入到细菌膜中，而细菌膜是许多基本酶细胞功能的家园。我们知道，这种化合物会改变膜的性质，从而影响嵌入其中的蛋白质。例如，青霉素耐药性是由于两个PBPs 2和2a之间的合作，使细菌能够在药物存在的情况下继续制造细胞壁。我们将确定ECg是否在细胞分裂位点破坏了该复合体，或者是否只是以一种允许在“错误”位点继续合成的方式分离该复合体。越来越明显的是，细胞分裂区域脂质环境的精细细节对确定细菌的有效复制至关重要；我们将研究心电图改变这个参数的方式。最后，ECg增加葡萄球菌盐敏感性的能力意味着它可以成为保存食物和防止与食物中毒相关的毒素细化的有效剂。细菌利用膜上的蛋白质泵出钠离子，我们将检查ECg的能力来破坏这一机制。

Staphylococcus aureus is commonly encountered as a coloniser of skin and mucosa but gives rise to a range of infections when there is an opportunity for the bacteria to enter the body. It also makes a range of toxins that are responsible for food poisoning: if food workers carry the bacterium, it can enter the food chain and will survive even in salty foods due to its ability to tolerate salt. Staphylococci show a remarkable capacity to resist the action of commonly used antibiotics through the acquisition of resistance mechanisms. In particular, they can acquire the mecA gene, whose product penicillin binding protein (PBP) 2a allows it to survive in the presence of high concentrations of penicillins such as methicillin and oxacillin. Such methicillin-resistant S. aureus (MRSA) are a threat to patients entering hospitals, particularly for major surgery, and are increasingly acquired in the community. At one time these infections were confined to humans but in recent years have been encountered in companion animals, especially dogs and cats; in all likelihood the bacteria were initially acquired from humans. Animal and human MRSA infections are difficult to treat and new approaches to therapy and control are urgently needed, in particular those that reduce the rate of emergence of drug resistance. We have established that one of the major components of green tea, a relatively labile substance termed epicatechin gallate, or ECg, alters the properties of S. aureus in a way that should be beneficial to animals and humans. Although ECg has no effect on the growth rate of MRSA, it reduces their resistance to penicillins by a much as five hundredfold, raising the possibility that it can be used alongside conventional antibiotics to treat infections. As ECg modifies rather than kills the target bacteria, it will not apply the same degree of extreme selective pressure as currently used 'bactericidal' drugs and may slow the emergence of resistance to therapeutic combinations. Additionally, we have discovered that ECg prevents the secretion of toxins and enzymes that enable the organism to cause tissue damage and infection and that are responsible for the symptoms of food poisoning. ECg also prevents the formation of biofilms: these complex assemblages of bacteria form at interfaces and are a necessary prerequisite for colonisation of host surfaces and devices such as catheters and prostheses prior to tissue invasion. Unfortunately, the naturally occurring chemical is not ideal for therapeutic purposes due to its labile nature and moderate activity and we are making derivatives of ECg with a more attractive profile. These attempts are hampered by an incomplete understanding of how ECg exerts its anti-staphylococcal activities. We are beginning to learn that it possesses such wide-ranging properties because i inserts into the bacterial membrane, a structure that is home to many essential enzymatic cellular functions. We know that the compound changes the properties of the membrane in a way that will affect the proteins embedded in it. For example, penicillin resistance is due to cooperation between two PBPs, 2 and 2a, enabling the bacteria to continue to make cell wall in the presence of the drug. We will determine if ECg disrupts this complex at the site of cell division or if it simply detaches the complex in a way that allows continued synthesis at 'wrong' sites. It is becoming apparent that the fine detail of the lipid environment in the region of cell division is critical to the determination of efficient bacterial replication; we will examine the way in which ECg alters this parameter. Finally, the capacity of ECg to increase salt sensitivity of staphylococci means that it could become an effective agent for preserving food and preventing the elaboration of toxins associated with food poisoning. The bacteria pump out sodium ions using membrane-located proteins and we will examine the capacity of ECg to compromise this mechanism.