Towards phage therapy against Clostridium difficile

Clostridium difficile is a serious public health threat, imposing a considerable burden on health services worldwide. In 2016/17, 97.4% of the 12,840 cases of C. difficile infection (CDI) in England and Wales were linked to mortality (Public Health England, 2017) and cost the NHS over £120 million. In the USA, the cost of managing the 500,000 plus cases of CDI is estimated at $6.3 billion. With the emergence of increasingly antimicrobial resistant C. difficile isolates, and dependence on a very limited selection of antibiotics for treatment, alternative therapies are required. The use of bacteriophages to both treat and prevent CDI offers considerable attractions, not least because phage are highly specific and would, compared to the use of antibiotics, reduce the impact of treatment on the natural gut microbiota. Classic therapy is, however, reliant on obligate lytic phage. We, like others, have only been able to isolate lysogenic phage. The aim of this project is to explore the potential of C. difficile phage in either the (i) lytic or (ii) lysogenic mode for treating CDI. (i) The former may be explored through the creation of synthetic, lytic phage through a combination of DNA synthesis and/or CRISPR-Cas9 genome editing in which rational mutations are made that prevent the phage from entering lysogeny. (ii) Rational modification of a lysogenic phage will enable the production of lethal gene products (eg., CRISPR-Cas9 or SASP, Small Acid Soluble Proteins) that kill the lysogenic host The PhD will be underpinned by the isolation of bacteriophage from anaerobic digesters and their thorough characterisation in terms of morphology (TEM), host range, genome (Illumina Paired-end), regulation (transcriptomics) and gene function (CRISPR-Cas9 editing). These phage will be compared and contrasted to those already isolated at Nottingham, before choosing candidate(s) for the proposed studies