Inhibition of bacterial Type III secretion by salicylanilides

大多数细菌无害地存在于环境中；数以百万计的细菌在我们的身体上或体内，但少数物种能够引起严重的疾病。直到上半个世纪抗生素的出现，人类才获得了这种优势。抗生素是一种能消灭细菌或抑制细菌生长的化合物。有时，由于基因突变或获得新基因等遗传变化，细菌会对所使用的抗生素产生自然耐药性。遗憾的是，我们过度依赖抗生素，滥用抗生素导致了一个重大问题：一些细菌已经对几乎所有已知的抗生素产生了耐药性。未来很有可能遇到更多无法治愈的细菌感染。食物中毒病原体肠沙门氏菌可以感染人类和动物，引起胃痛和腹泻。受感染的动物可以通过直接接触或通过受污染的肉、奶或蛋将疾病传染给人类。被摄入的沙门氏菌到达肠道后，必须与寄居在肠道内的数百万“有益”细菌竞争。沙门氏菌通过开启几种“毒力因子”侵入肠壁。这些因素在环境中不是日常生存所必需的，但在宿主中引起疾病却是绝对必要的。其中最引人注目的是iii型分泌系统。这是一个类似注射器的结构，用于将细菌分子直接“注射”到宿主细胞中。然后这些分子劫持宿主细胞的机制，使沙门氏菌进入细胞。从这里沙门氏菌可以通过组织深入体内。当身体检测到这种攻击并试图将沙门氏菌排出体外时，这种入侵会引发疼痛和腹泻。有可能鉴定出能够特异性地使细菌毒力因子失效的新型抗菌化合物，从而预防疾病。理想情况下，这些化合物不会影响细菌的生存或生长，因此细菌对化合物的耐药性应该不太可能出现。拟研究摘要。我们开发了实验室检测方法来测量III型沙门氏菌的分泌和功能。我们将使用这些检测来筛选一些抑制III型分泌和/或表达的化合物。最佳候选药物将用于研究这些化合物如何起作用，并观察它们是否能减少沙门氏菌对培养细胞和肠道组织的侵袭。我们还将测量这些化合物是否有效地减少液体分泌量和肠道炎症，这些炎症是对入侵的反应。由于沙门氏菌实际上有两个TTSS，第二个TTSS在疾病的后期阶段是最必要的，因此针对iii型分泌系统可能是非常有益的，因为一个成功的抗菌化合物可能作用于两个系统来抑制疾病。此外，已知其他致病微生物也使用与沙门氏菌非常相似的III型分泌系统，因此一些化合物可能提供广泛的抗病原体治疗，而对共生的“好”细菌的影响最小。最后，抑制III型分泌系统的化合物可用于研究细菌如何与宿主细胞相互作用并导致疾病的过程，因为它们允许III型分泌系统在感染过程的任何阶段被“关闭”。所有这些研究都将引起科学家和非科学家的兴趣，因为针对毒力因子的抗菌药物有可能彻底改变人类和兽医学。这些化合物可用于预防有感染危险的个体的疾病，或替代地用于治疗已确定的疾病。

Background Most bacteria exist harmlessly in the environment; many millions actually on or within our own bodies, but a few species are capable of causing severe disease. It is only with the advent of antibiotics in the last half century that humanity has gained the advantage. Antibiotics are compounds which destroy bacteria or inhibit their growth. Sometimes bacteria can become naturally resistant to the antibiotic used due to a genetic change such as a mutation in a gene or acquisition of a new gene. Regrettably, we came to rely too heavily on antibiotics and their misuse has lead to a major problem: some bacteria have become resistant to almost all known antibiotics. There is a high possibility of encountering more untreatable bacterial infections in the future. The food-poisoning pathogen, Salmonella enterica, can infect both humans and animals causing stomach pain and diarrhoea. Infected animals can pass the disease to humans by direct contact or through contaminated meat, milk or eggs. Ingested Salmonella reach the intestines and must then compete with the millions of 'good' bacteria that inhabit the gut. Salmonella invades the intestine wall by switching on several 'virulence factors'. These are factors which are not necessary for day-to-day existence in the environment but are absolutely required to cause disease in the host. One of the most striking is the Type-III secretion system. This is a syringe-like structure used to 'inject' bacterial molecules directly into the host cell. These molecules then hijack the machinery of the host cell and enable Salmonella to enter the cell. From here Salmonella can move through the tissues deeper into the body. This invasion triggers pain and diarrhoea as the body detects the attack and tries to flush out the Salmonella. It may be possible to identify new types of antibacterial compound that will specifically disable bacterial virulence factors, thus preventing disease. Ideally, these compounds will not affect survival or growth of the bacteria so resistance of bacteria to the compounds should be less likely to emerge. Summary of proposed research. We have developed laboratory assays to measure Salmonella Type III secretion and functions. We will use these assays to screen a number of compounds for inhibition of Type III secretion and/or expression. The best candidates will be used in assays to investigate how the compounds work and to see whether they can reduce Salmonella invasion of cultured cells and gut tissues. We will also measure if these compounds are effective in reducing the amount of fluid secretion and gut inflammation that occurs in response to the invasion. As Salmonella actually has two TTSS, the second being mostly necessary for later stages of the disease, targeting the Type-III secretion system could therefore be very beneficial because a successful antibacterial compound may act on two systems to inhibit disease. Furthermore, other pathogenic microbes are also known to use a Type III secretion systems that are very similar to that found in Salmonella, so it is possible that some compounds may provide broad-range anti-pathogen treatments with minimal effects on commensal 'good' bacteria. Finally, compounds inhibiting Type III secretion systems could be used to study the process of how bacteria interact with host cells and cause diseases as they allow the type III secretion system to be 'switched off' at any stage of infectious process. All these studies will be of interest to scientists and non-scientists because antimicrobial agents that target virulence factors have the potential to revolutionise human and veterinary medicine. Such compounds may be used to prevent disease in individuals at risk of infection or alternatively used to treat established disease.