Understanding multiple antibiotic resistance in Gram-negative bacteria

Antibiotics underpin modern medical, veterinary and farming practices worldwide. However, the efficacy of antibiotics is decreasing. A simple genetic system called mar, which can provide protection against multiple antibiotics, will be the focus of our project. Briefly, mutations in this system result in triggering of bacterial defences. Consequently, the bacterial cell is able to protect itself from many different toxic agents. We seek to identify and understand all of the cellular pathways that link this defence system to antibiotic resistance. By understanding pathways to drug resistance, we will provide new opportunities to control bacterial infections. Importantly, the pathways we will investigate are similar in many microbes. As a result, our findings will be applicable to many bacterial infections and so of broad interest.

Antibiotics are often used with no knowledge of the infecting microorganism. Hence, widespread use of antibiotics favours the emergence of antibiotic resistant bacteria. Our experiments will be done using a bacterium called E. coli. This organism is commonly found in people and animals, including pigs. Furthermore, pigs are among the most common recipients of antibiotics in the EU and the number of serious E. coli infections in people is increasing. Indeed, the type of E. coli we will investigate has acquired mutations in the defence system we will study. Therefore, we will address a major health issue relevant to the UK animal food production industry as well as animal and human health. Importantly, this application comes at a time when there are global calls for a better understanding of antibiotic resistance.

Antibiotics are a cornerstone of modern medicine and used extensively by doctors, veterinarians and farmers in the UK. Worryingly, many bacteria are now resistant to some, or all, antibiotics. The multiple antibiotic resistance (mar) system, or its homologs (e.g. ram and sox) in other species, play a key role in facilitating drug resistance. Consequently, mar will be the focus of our work. We will analyse the mar system of Escherichia coli because it is easily accessible using genetic, biochemical and genomic tools. Furthermore, we will focus on Enterotoxigenic E. coli (ETEC) as drug resistant ETEC are frequently isolated from farm animals. Briefly, the mar system encodes two transcription factors; a repressor called MarR and an activator called MarA. Usually, MarR prevents MarA expression. However, in multidrug resistant bacteria, the marR gene and integenic region often contain mutations, which render MarR poorly active. Consequently, MarA is constitutively over-expressed. Once expressed, MarA switches on bacterial defence systems that can protect cells against many antimicrobial compounds. Surprisingly, our preliminary data indicates that most genes targeted by MarA are undefined. This is a significant omission; the new MarA targets include hitherto undescribed determinants for antibiotic resistance and potential novel therapeutic targets. Thus, the aim of this project is to identify all targets of the mar system and define the underlying mechanisms of antibiotic resistance. We expect these resistance mechanisms to be broadly conserved in Gram-negative bacteria.

This project will provide a significant advance in understanding of antibiotic resistance and provide potential new targets for which inhibitors can be sought. Thus, our results will be applicable to the development of novel treatment strategies providing societal and economic benefit for human and veterinary medicine.

The end users that will benefit from this research are:

1. Association of the British Pharmaceutical Industry (ABPI) and its members
2. Agriculture & Horticulture Development Board (AHDB) and its members
3. Policy makers and government departments (Department of Health, Public Health England, NHS Trusts, EU, WHO)
4. The wider public (schools, students, general public)
5. Academia (biologists, medics and bioinformaticians internationally)

We have expanded on how these different groups will benefit below.

THE PRIVATE SECTOR (ABPI and AHBD; Immediate to long term impact): The ABPI represents most small, medium and large research-based pharmaceutical companies in the UK. Members of the ABPI supply 90 % of all medicines used by the NHS and are responsible for over two-thirds of current drug development. One role of the ABPI is to support its members by facilitating collaboration and innovation via interactions with academia. Hence, the ABPI and their members will benefit from our identification and characterisation of potential new drug targets. Antibiotic resistance is also a major issue for the farming industry. In a report to the House of Commons science and technology committee farmers stated that, without antibiotics, pig production in the UK would be "pretty much impossible". The AHDB is funded by levy payers (farmers, veterinarians and others in the supply chain). This generates an annual income in excess of £60 million. This is used to equip levy payers with independent, evidence-based information and tools to i) become more competitive and sustainable and ii) prevent and manage disease. Thus, AHDB and their levy payers will benefit from a better understanding of antibiotic resistance to the most commonly used drugs in animals.

POLICY MAKERS AND GOVERNMENT DEPARTMENTS (Department of Health, Public Health England, NHS Trusts, EU, WHO; Immediate to medium term impact): These government departments will benefit from knowledge gained about antibiotic resistance mechanisms to help formulate evidence based policy in relation to national and global infection challenges.

THE WIDER PUBLIC (SCHOOLS, STUDENTS, GENERAL PUBLIC; Immediate to long term impact): Our work will raise general awareness of antibiotic resistance and how it affects people and animals. This will generate impact by improving public understanding of drug-resistance and highlighting how individuals can make a difference.

ACADEMIA (Academics and the PDRA; Immediate to long term impact): Academics will benefit from i) new approaches to study bacterial systems ii) the availability of genomic datasets that can be mined in order to understand other bacterial systems. The PDRA will benefit from training to underpin future career success. In the long term this will contribute to the scientific advancement and economic prosperity of the UK.