PhD Study : Identification of factors regulating production of biofilm by antibiotic resistant Clostridium difficile using transcriptomics.

Summary

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).

Essential criteria

Applicants should hold, or expect to obtain, a First or Upper Second Class Honours Degree in a subject relevant to the proposed area of study.

We may also consider applications from those who hold equivalent qualifications, for example, a Lower Second Class Honours Degree plus a Master’s Degree with Distinction.

In exceptional circumstances, the University may consider a portfolio of evidence from applicants who have appropriate professional experience which is equivalent to the learning outcomes of an Honours degree in lieu of academic qualifications.

• Sound understanding of subject area as evidenced by a comprehensive research proposal

Desirable Criteria

If the University receives a large number of applicants for the project, the following desirable criteria may be applied to shortlist applicants for interview.

• First Class Honours (1st) Degree
• Masters at 65%
• Research project completion within taught Masters degree or MRES
• Practice-based research experience and/or dissemination
• Experience using research methods or other approaches relevant to the subject domain
• Work experience relevant to the proposed project
• Publications - peer-reviewed
• Experience of presentation of research findings
• A comprehensive and articulate personal statement
• Relevant professional qualification and/or a Degree in a Health or Health related area

Funding and eligibility

The University offers the following levels of support:

Vice Chancellors Research Studentship (VCRS)

The following scholarship options are available to applicants worldwide:

• Full Award: (full-time tuition fees + £19,000 (tbc))
• Part Award: (full-time tuition fees + £9,500)
• Fees Only Award: (full-time tuition fees)

These scholarships will cover full-time PhD tuition fees for three years (subject to satisfactory academic performance) and will provide a £900 per annum research training support grant (RTSG) to help support the PhD researcher.

Applicants who already hold a doctoral degree or who have been registered on a programme of research leading to the award of a doctoral degree on a full-time basis for more than one year (or part-time equivalent) are NOT eligible to apply for an award.

Please note: you will automatically be entered into the competition for the Full Award, unless you state otherwise in your application.

Department for the Economy (DFE)

The scholarship will cover tuition fees at the Home rate and a maintenance allowance of £19,000 (tbc) per annum for three years (subject to satisfactory academic performance).

This scholarship also comes with £900 per annum for three years as a research training support grant (RTSG) allocation to help support the PhD researcher.

• Candidates with pre-settled or settled status under the EU Settlement Scheme, who also satisfy a three year residency requirement in the UK prior to the start of the course for which a Studentship is held MAY receive a Studentship covering fees and maintenance.
• Republic of Ireland (ROI) nationals who satisfy three years’ residency in the UK prior to the start of the course MAY receive a Studentship covering fees and maintenance (ROI nationals don’t need to have pre-settled or settled status under the EU Settlement Scheme to qualify).
• Other non-ROI EU applicants are ‘International’ are not eligible for this source of funding.
• Applicants who already hold a doctoral degree or who have been registered on a programme of research leading to the award of a doctoral degree on a full-time basis for more than one year (or part-time equivalent) are NOT eligible to apply for an award.

Due consideration should be given to financing your studies. Further information on cost of living