Overcoming antibiotic resistance by studying antibiotic hypersensitivity

通过研究抗生素超敏反应克服抗生素耐药性

• 批准号: BB/J016691/1
• 负责人: Margaret Smith
• 金额: $ 69.6万
• 依托单位: University of York
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2012
• 资助国家: 英国
• 起止时间: 2012 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FJ016691%2F1
• 关键词: Overcoming; antibiotic; resistance; studying; hypersensitivity

Antibiotics have saved millions of lives since their discovery. Antibiotics kill pathogenic bacteria by targeting an essential metabolic process. Pathogenic bacteria can protect themselves from antibiotics by altering or providing a new target that no longer binds the antibiotic, destroying the antibiotic or pumping the antibiotic away from the target. Antibiotic resistance is now a serious problem in treating diseases caused by pathogenic bacteria such that in the EU 25,000 people die annually from untreatable infections. Globally there are 440,000 new cases annually of multidrug resistant tuberculosis resulting in 150,000 deaths. Solutions to the problem of antibiotic resistant bacteria are being sought on several fronts including better control over the use of existing antibiotics, the discovery and development of new antibiotics and antibacterial strategies that do not rely on antibiotics such as phage therapy, bacteriocins, antibacterial peptides and vaccines. A hitherto underexplored but potentially exciting approach is to use combination therapy in which two or more drugs are used simultaneously and act synergistically to kill antibiotic resistant bacteria. This approach has been used for combating HIV and tuberculosis for some time. However combination therapy need not always involve two antibiotics; one of the drugs used may not itself have anti-microbial activity but potentiates the activity of the antibiotic. A well-known potentiator that has been taken by most people is clavulanic acid, an inhibitor of the enzyme beta-lactamase that destroys beta-lactam antibiotics such as penicillin. How many potentiator targets are there and how do we find them? There is evidence that there are hundreds of potentiator targets in bacteria of varying efficacy and that might act against different types of antibiotics. This evidence comes from measuring the antibiotic sensitivity in bacteria that have single gene mutations; those mutants with greater sensitivity to an antibiotic compared to a strain with an intact gene (the parent) indicate that the mutated gene or its consequences on metabolic processes is a potentiator target. We have isolated mutants in a bacterium, Streptomyces coelicolor (a relative of Mycobacterium tuberculosis) that are hypersensitive to a subset of antibiotics including two antibiotics that are so-called 'last resort' antibiotics for some pathogenic bacteria. The mutations lie in enzymes required to modify proteins being localised to the outside of the cell with sugars. Knocking out this modification system may have a variety of consequences on metabolic processes, all unknown at present. We hypothesise that if we understand what these consequences are at the metabolic level, we can identify rational targets for potentiators and, in some cases, undermine their resistance mechanisms. Our first objective is to ask whether mutations in the protein modification system in related bacteria are also hypersensitive to establish whether our observations are general, and to initiate screens for potentiator chemicals in collaboration with NovaBiotics Ltd and the Marine Biodiscovery Centre in Aberdeen. Second we plan to determine what major metabolic changes have occurred in the mutants compared to the parent strain by studying the proteins that might be affected by modification in the cell surface and by measuring changes in gene expression. Third we plan to identify what genetic changes need to happen to the hypersensitive strains to make them resistant again and this will point to both an explanation of the hypersensitivity and how resistance to potentiators might arise. At the end of this project we hope to be in a position where we can start screening for potentiators for use with antibiotics that act against Mycobacterium tuberculosis and some vancomycin resistant pathogens.

自从发现抗生素以来，它们已经拯救了数百万人的生命。抗生素通过针对一个重要的代谢过程来杀死病原菌。病原菌可以通过改变或提供一个不再与抗生素结合的新靶点、销毁抗生素或将抗生素抽离目标来保护自己免受抗生素的伤害。抗生素耐药性现在是治疗由病原菌引起的疾病的一个严重问题，在欧盟，每年有2.5万人死于无法治疗的感染。全球每年新增44万例耐多药结核病病例，导致15万人死亡。目前正在几个方面寻求解决耐药细菌问题的办法，包括更好地控制现有抗生素的使用，发现和开发新的抗生素，以及不依赖抗生素的抗菌策略，如噬菌体疗法、细菌素、抗菌肽和疫苗。到目前为止，一种尚未被探索但可能令人兴奋的方法是使用联合疗法，即两种或两种以上药物同时使用，并协同作用以杀死抗生素耐药性细菌。这种方法用于抗击艾滋病毒和结核病已有一段时间了。然而，联合疗法并不总是需要两种抗生素；所使用的其中一种药物本身可能没有抗微生物活性，但会增强抗生素的活性。大多数人服用的一种众所周知的增效剂是克拉维酸，它是一种酶β-内酰胺酶的抑制剂，可以破坏青霉素等β-内酰胺类抗生素。有多少增效剂靶标？我们如何找到它们？有证据表明，细菌中有数百个不同效力的增效剂靶标，它们可能对不同类型的抗生素起作用。这一证据来自于测量具有单基因突变的细菌的抗生素敏感性；与具有完整基因的菌株(亲本)相比，对抗生素具有更高敏感性的突变株表明，突变的基因或其对代谢过程的影响是增强剂的靶标。我们在一种名为天蓝色链霉菌(结核分枝杆菌的近亲)的细菌中分离出了突变株，它们对一系列抗生素过敏，其中包括两种抗生素，这两种抗生素对一些致病菌来说是所谓的“最后手段”抗生素。突变存在于修饰蛋白质所需的酶中，这些酶定位于含有糖的细胞外。敲除这种修饰系统可能会对新陈代谢过程产生各种后果，目前都是未知的。我们假设，如果我们了解代谢水平上的这些后果，我们就可以确定增强剂的合理靶点，在某些情况下，破坏它们的耐药机制。我们的第一个目标是询问相关细菌中蛋白质修饰系统的突变是否也是超敏的，以确定我们的观察结果是否具有一般性，并与NovaBiotics有限公司和位于阿伯丁的海洋生物发现中心合作启动对增强剂化学物质的筛选。其次，我们计划通过研究可能受到细胞表面修饰影响的蛋白质，以及通过测量基因表达的变化，来确定与亲本菌株相比，突变体中发生了哪些主要的代谢变化。第三，我们计划确定超敏菌株需要发生哪些基因变化才能使它们再次产生抗药性，这将同时解释超敏反应和对增强剂的抗药性可能产生的原因。在这个项目结束时，我们希望能够开始筛选与抗结核分枝杆菌和一些万古霉素耐药病原体作用的抗生素一起使用的增效剂。

项目成果

An operon encoding enzymes for synthesis of a putative extracellular carbohydrate attenuates acquired vancomycin resistance in Streptomyces coelicolor.

• DOI: 10.1099/mic.0.000763
• 发表时间: 2019-01
• 期刊: Microbiology
• 影响因子: 1.5
• 作者: N. Read;R. Howlett;Margaret C. M. Smith

Streptomyces coelicolor strains lacking polyprenol phosphate mannose synthase and protein O-mannosyl transferase are hyper-susceptible to multiple antibiotics.

• DOI: 10.1099/mic.0.000605
• 发表时间: 2018-03
• 期刊: Microbiology (Reading, England)
• 作者: Howlett R;Read N;Varghese A;Kershaw C;Hancock Y;Smith MCM
• 通讯作者: Smith MCM

Disruption of the GDP-mannose synthesis pathway in Streptomyces coelicolor results in antibiotic hyper-susceptible phenotypes.

• DOI: 10.1099/mic.0.000636
• 发表时间: 2018-04
• 期刊: Microbiology (Reading, England)
• 作者: Howlett R;Anttonen K;Read N;Smith MCM
• 通讯作者: Smith MCM

The glycoproteome in Streptomyces coelicolor includes enzymes required for cell wall biogenesis

天蓝色链霉菌中的糖蛋白质组包括细胞壁生物合成所需的酶

• 发表时间: 2019
• 期刊: Submitted
• 作者: Keenan, T.

Evolutionary Relationships among Actinophages and a Putative Adaptation for Growth in Streptomyces spp.

• DOI: 10.1128/jb.00618-13
• 发表时间: 2013-11-01
• 期刊: JOURNAL OF BACTERIOLOGY
• 影响因子: 3.2
• 作者: Smith, Margaret C. M.;Hendrix, Roger W.;Hatfull, Graham F.
• 通讯作者: Hatfull, Graham F.