Bacteriocins from environmental samples that kill clostridia

Background: Clostridium difficile is a cause of neonatal enteritis in livestock such as pigs, cattle and horses. Affected animals develop respiratory problems, abdominal distention and bloody diarrhea leading to weight loss and in extreme cases mortality. This puts a financial burden on the farming industry, and a potential risk to healthcare if infected animals are not treated with appropriate antibiotics. C. difficile is also a huge burden in our hospitals, killing 3x as many people in teh UK as MRSA. Being an anaerobic spore former, C. difficile is resistant to many antibiotics and treatment is currently limited to only a few (metronidazole, vancomycin, fidaxomicin). Resistance to metronidazole has been observed already, exemplifying the urgent need for new therapies. One alternative option for successful treatment of CDI is the use of bacteriocin therapy. Bacteriocins are ribosomally synthesised protein antibiotics that kill closely related species to the producing micro-organism and have been shown to be important factors in the establishment of individual strains from mixed populations of E. coli, Pseudomonas aeruginosa and Klebsiella pneumoniae. Bacteriocins have recently been shown to be produced by Clostridium spp. with activity against a range of C. difficile isolates. Aim: The aim of this project is to identify novel bacteriocins produced by clostridia and other bacteria from environmental samples, establish their host-range by testing against a range of clinically relevant C. difficile isolates including the hypervirulent PCR-ribotype 027 strains and determine the mechanisms of bacteriocin import and activity against sensitive cells. Strategy: The strategy of this project will involve (i) screening for bacteriocins against strains of C. difficile, which will then be characterised (ii) in terms of activity-spectrum by assessing biological activity against a range of clinically relevant C. difficile ribotypes and (iii) through purification and microscopic analysis. The bacteriocins will be investigated further including (iv) fractionation, SDS-PAGE and Mass spectronomy. (v) The producer will be sequenced (whole genome sequencing), to identify the bacteriocin genes, with a view to potential manipulation to increase host-range and/or potency. Training: This project will be carried out in Nottingham's Clostridia Research Group (CRG), which comprises 80+ graduate and postdoctoral researchers (www.clostron.com/people.php) and is the largest academic group worldwide working on genetic modification of Clostridia. The CRG as recently become part of Nottingham's new synthetic biology research centre (SBRC) and will benefit from state of the art research technologies and equipment. The outlined study will allow for training in an exciting multidisciplinary environment, incorporating anaerobic microbiology, the application of advanced gene tools, in vivo models and links to the BRU providing access to clinical samples. Approaches will include the application of the newest developments in biological engineering and in silico analysis, using bioinformatics.