Project description:Clostridium difficile (C. difficile) strains belonging to PCR ribotype 027, PFGE type NAP1, REA type B1 and toxinotype III, termed NAP1/027, have been implicated in the increased frequency of outbreaks of Clostridium difficile-associated diarrhoea (CDAD) in North America and Europe. The NAP1/027 strains appears to be more virulent with an increased mortality and frequency of relapse. Current European C. difficile microarrays are designed to the first sequenced and annotated C. difficile complete genome - strain 630 (ribotype 12). A high density oligonucleotide microarray was designed to C. difficile 630 (CD630) sequence and extra probes corresponding to two PCR ribotypes O27 strains C. difficile R20291 and QCD-32g58 were also included. Comparative genomic hybridisation was used to identify markers of ribotype 027 strains and markers to identify CD630. Strains hybridised to the array included the most prevalent ribotypes found in the UK and Europe (106 and 001) as well as the emerging hypervirulent ribotype 078.
Project description:Purpose: To investigaet whether an inflammatory immune response at the Ileum can contribute to development and severity of diarrhoea in Merino sheep that are prone and not prone to develop diarrhoea. Methods:Transcriptomes in the Ileum (third part of small intestine) of Merino sheep were analyzed using RNASeq. Functional enrichment of DEGs using GO terms in biological processes and KEGG pathways were studied. PPI (protein-protein interaction) network and sub-network were studied using clusterONE plugin in Cytoscape. Result and conclusion: Although Ileum in not a primary site of helminth (species under study in our experiment), the results indicated that an inflammatory immune response can contribute to development and severity of diarrhoea. The low-diarrhoea animals did not show any signs of immune/inflammatory response. The genes with important roles in the responses to helminth infection could be targeted in breeding programs to prevent diarrhoea.
Project description:Toxin A and B from Clostridium difficile are the primary virulence factors in Clostridium difficile disease. The changes in gene transcription of human colon epithelial cells were investigated in vitro in order to better understand the many effects of both toxins.
Project description:Clostridium difficile is the leading cause of hospital acquired diarrhoea in industrialised countries. Under conditions that are not favourable for growth, the pathogen produces metabolically dormant endospores via asymmetric cell division. These are extremely resistant to both chemical and physical insult and provide the mechanism by which C. difficile can evade the potentially fatal consequences of exposure to heat, oxygen, alcohol, and certain disinfectants. Spores are the primary infective agent and must germinate to allow for vegetative cell growth and toxin production. While spore germination in Bacillus is well understood, little is known about C. difficile germination and outgrowth. Here we use genome-wide transcriptional analysis to elucidate the temporal gene expression patterns in C. difficile 630 endospore germination. We have optimized methods for large scale production and purification of spores. The germination characteristics of purified spores have been characterized and RNA extraction protocols have been optimized. Gene expression was highly dynamic during germination and outgrowth, and was found to involve a large number of genes. Using this genome-wide, microarray approach we have identified 511 genes that are significantly up- or down-regulated during C. difficile germination (p less than or = 0.01). A number of functional groups of genes appeared to be co-regulated. These included transport, protein synthesis and secretion, motility and chemotaxis as well as cell wall biogenesis. These data give insight into how C. difficile re-establishes its metabolism, re-builds the basic structures of the vegetative cell and resumes growth. [Data is also available from http://bugs.sgul.ac.uk/E-BUGS-145]
Project description:Purpose:To explore the underlying biological mechanisms relating to development and control of diarrhoea, we compared sheep that were prone to develop diarrhoea with sheep that were not prone. Methods:Transcriptomes in the tissues of Merino sheep where the parasites were located were analyzed using RNASeq. Functional enrichment of DEGs using GO terms in biological processes and KEGG pathways were studied. PPI (protein-protein interaction) network and sub-network were studied using clusterONE plugin in Cytoscape. Result and conclusion: Our results indicate that suppression of ‘immune response’, ‘antigen processing and presentation’, and a list of biological functional terms related to ‘suppression in immune tolerance and activation of repair processes associated with tissue damage, including ‘extracellular matrix organization’, ‘collagen fibril organization’, ‘tissue morphogenesis’, ‘circulatory system development’, ‘morphogenesis of an epithelium’, and ‘focal adhesion’ were associated with high-diarrhoea phenotype. The genes with important roles in the responses to helminth infection could be targeted in breeding programs to prevent diarrhoea.
Project description:Transcriptional analysis of Clostridium difficile R20291 in biofilm formation, planktonic state and grown on blood agar RNA sequencing was performed on Clostridium difficile R20291 in three different conditions: Biofilm formation, plantonic state and grown on blood agar plates. Each condtion has 3 replicates.
