Investigating molecular mechanisms of Clostridium difficile colonisation

Lead Research Organisation: University of Warwick
Department Name: School of Life Sciences

Abstract

The gut pathogen Clostridium difficile is a major cause of gastrointestinal infections in humans and animals. It is the leading cause of diarrheoa in piglets in many parts of the world. In recent years it has become evident that animal reservoirs of this pathogen that are important for human infections. Colonisation of the gut is a key determinant of bacterial carriage and disease outcome, although little is known about bacterial or host factors modulating interactions between C. difficile and the gut epithelium. The goal of this work is to identify basic pathways that control host-C. difficile interactions. Human gut epithelial cell layers will be co-cultured with clinically relevant C. difficile strains in novel in vitro systems and genome-wide analyses will be performed to determine the genes important during C. difficile colonisation. The role of selected proteins in modulation of the gut epithelial and microbiota interactions will be studied. Identification and functional analysis of proteins critical to C. difficile colonisation will enhance our understanding of the establishment of pathogen in this gut and lead to effective interventions for CDI.

Publications

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Studentship Projects

Project Reference Relationship Related To Start End Student Name
BB/M01116X/1 01/10/2015 30/09/2023
1782606 Studentship BB/M01116X/1 03/10/2016 31/01/2021 Lucy Rachel Frost
 
Description A vertical diffusion chamber was used to generate a dual-environment in vitro human gut model which facilitated C. difficile infection of human intestinal epithelial cells. A dual RNA-seq approach was used to resolve the transcriptomic profiles of both the host cell and adherent bacteria simultaneously during infection. Samples were taken at 3 h, 6 h, 12 h and 24 h post infection to capture the temporal changes in gene expression over the course of the infection. DNA libraries were generated and sequenced on the Illumina NextSeq 500 system. A bioinformatics pipeline optimised for dual RNA-seq analysis with DESeq2 was used to make comparisons between the gene expression profiles of infected samples and uninfected controls. During infection, we observed differential expression of many bacterial genes and pathways. Interestingly, many cell surface proteins such as Cwp84, Cwp19 and Cwp66 were downregulated, while Cwp10 was upregulated. Several metabolic pathways were also differentially regulated, including butanoate, phenylalanine and tyrosine metabolism. CodY, a global regulator of many processes related to virulence, including toxin production, sporulation and motility, was also downregulated. In the host cells we observed differential expression of genes involved in several pathways including mucus production and wound healing. Muc13 is a cell-surface mucin which has been reported to regulate inflammatory cytokine production in the intestines was significantly upregulated 24 h after C. difficile infection. Interestingly Muc13 was recently reported to be a potential biomarker for Plasmodium infection. Fibrinogen is a glycoprotein complex involved in blood clot formation. Several genes involved in fibrinogen biosynthesis were downregulated at 3 h, 6 h and 12 h post infection.
Exploitation Route Differentially expressed human genes could be biomarker candidates for C. difficile infection which have a potential use in novel diagnostic techniques. Bacterial differentially expressed genes may indicate colonisation factors which can potentially be targeted in new therapies to combat C. difficile infection.
Sectors Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology

 
Description Medical and Life Sciences Research Fund
Amount £6,000 (GBP)
Organisation Medical and Life Sciences Research Fund 
Sector Charity/Non Profit
Country United Kingdom
Start 07/2018