Self-funded PhD opportunity Identification of factors regulating production of biofilm by antibiotic resistant Clostridium difficile using transcriptomics.
This opportunity is now closed.
Subject: Biomedical Sciences
Background to the project
Clostridium difficile is the most common cause of antibiotic-associated bacterial diarrhoea, with significant impact on morbidity and mortality rates of Healthcare-Associated Infection (HAI). Antibiotic associated dysbiosis of the gut microbiome leads to the development of C. difficile infection (CDI) mediated by the toxins (A and B) produced by the organism. C. difficile is a multiply antibiotic resistant pathogen, and increased recurrences of CDI are hypothesised to be due, in part, to decreased efficacy of conventional antibiotic therapies. In recent years the formation of biofilms by C. difficile has been documented. This provides a specialised growth opportunity for the pathogen in which antibiotic resistance might be enhanced thereby contributing to treatment failures. The ability to form biofilm has been increasingly linked to antibiotic resistance, sporulation, and recurrence of CDI. Notably, the extent of biofilm formation by C. difficile strains reported in the literature varies considerably, suggesting that a multiplicity of factors influence biofilm in this organism.
We have recently shown that ClosTron-mediated disruption of the dnaK gene in C. difficile strain 630 erm results in a variety of physiological changes, including significantly increased biofilm formation. More recent data from our lab indicates that a variety of other factors, including antibiotic sensitivity, cell surface structure, motility and response to gut metabolites is also significantly changed in the dnaK mutant.
The overall aim of this project is to dissect more fully, using state of the art molecular biology and bioimaging techniques, the changes in physiology and gene expression associated with growth of C. difficile in biofilms. The dnaK mutant represents an excellent model system in which to examine such changes in an “enhanced biofilm former”, enabling comparison with the wild-type and a variety of antibiotic-resistant clinical C. difficile isolates.
Methods to be used
The project builds upon a series of successful PhD projects within our group and consequently the majority of techniques to be used (RNA extraction, analysis and sequencing, q-RT-PCR, biofilm assays, proteomics, antibiotic resistance and cell-staining assays) are well established in our laboratory. Full training will be provided in these and other relevant techniques to enable the candidate to successfully complete the programme of research. We hypothesise that changes in the phenotype of biofilm grown cells will map to altered gene expression in the transcriptome as well as to changes in key structural proteins.
We will test this hypothesis by growing C. difficile strains planktonically and as biofilms under a range of physiologically relevant conditions. Transcriptomes will be subject to RNAseq to determine global gene expression differences, with validation by qRT-PCR. Proteome changes will be analysed by GeLC-MS or Western blots as appropriate. Of interest will be the effects of low levels of antibiotics, gut metabolites or novel antibacterial peptides on gene/protein expression and biofilm ultrastructure.
Overall, the project seeks to assist with the rational design of new antimicrobials through post identification of genes that are important for biofilm growth, development and survival. Impact The time-line between the increasing frequency of infections caused by multidrug-resistant bacteria and the development of new antibacterial agents is widening. Antibiotic development is no longer considered to be an economically wise investment for the pharmaceutical industry due to reduced economic incentives and challenging regulatory requirements. The effectiveness of currently available antibiotics needs to be optimised and extended. This will only be achieved by a multidisciplinary approach to the problem. Understanding the physiology of antibiotic resistant pathogens, as proposed in this project, will be essential to support initiatives in infection control and antimicrobial stewardship policies (ASP) that have been recently proposed.
This project will be based at Ulster's Nutrition Innovation Centre for Food and Health (NICHE). Applicants should note that £15,000 in bench fees/consumables will be required to fund the project.
- Upper Second Class Honours (2:1) Degree from a UK institution (or overseas award deemed equivalent via UK NARIC)
This is a self-funded PhD opportunity.
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