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.
Project description:Clostridioides difficile infection (CDI), characterized by colitis and diarrhea, afflicts approximately half a million people in the United States every year, burdening both individuals and the healthcare system. C. difficile 630Δerm is an erythromycin-sensitive variant of the clinical isolate C. difficile 630 and is commonly used in the C. difficile research community due to its genetic tractability. 630Δerm possesses a point mutation in perR, an autoregulated transcriptional repressor that regulates oxidative stress resistance genes. This point mutation results in a constitutively de-repressed PerR operon in 630Δerm. To address the impacts of perR on phenotypes relevant for oxygen tolerance and relevant to a murine model of CDI, we corrected the point mutant to restore PerR function in 630∆erm (herein, 630∆erm perRWT). We demonstrate that there is no difference in growth between 630Δerm and a 630Δerm perRWT under anaerobic conditions or when exposed to concentrations of O2 that mimic those found near the surface of the colonic epithelium. However, 630∆erm perRWT is more sensitive to ambient oxygen than 630∆erm, which coincides with alterations in expression of a variety of perR-dependent and perR-independent genes. Finally, we show that 630∆erm and 630∆erm perRWT do not differ in their ability to infect and cause disease in a well-established murine model of CDI. Together, these data support the hypothesis that the perR mutation in 630∆erm arose as a result of exposure to ambient oxygen and that the perR mutation in 630∆erm is unlikely to impact CDI-relevant phenotypes in laboratory studies
Project description:Clostridium difficile is an anaerobic spore-forming rod-shaped gram-positive bacterium that can infect both humans and animals. Most studies on the pathogenesis of C. difficile have focused on its toxins and their effect on the host cells. Recently, we utilized microarrays to identify conserved and divergent genes associated with virulence in C. difficile isolates from humans and animals. Our data provided the first clue toward a complex mechanism underlying host adaptation and pathogenesis. Microarray technology offers an efficient high-throughput tool to study the transcriptional profiles of pathogens and infected host cells. Transcriptomes of C. difficile after exposure to environmental and antibiotic stresses and those of human epithelial colorectal Caco-2 cells upon TcdA treatment have been analyzed. To our knowledge, there are still no reports on the transcriptomic study of host-pathogen interactions for C. difficile infection (CDI). In vitro analyses of interplay between host and pathogen are essential to unravel the mechanisms of infection and to investigate the host response to infection. We therefore employed microarrays to study both bacterial and human cellular transcriptome kinetics during CDI to Caco-2 cells. Here we present a large-scale analysis of transcriptional profiles to reveal molecular determinants playing a role in C. difficile pathogenesis and the host response. We found that there were 254 and 224 differentially-expressed genes after CDI in C. difficile and Caco-2 cells, respectively. These genes are clustered according to their functional categories and their potential roles in pathogenesis and host response are discussed. Our results will not only increase our understanding on the host-pathogen interaction, but may also provide targets for drug development.
Project description:RNAseq and LC/MS metabolomics analysis of C. difficile strain 630 grown in BHIS media with 50% (vol/vol) faecal water added, compared with control BHIS containing only the additional PBS used for prep of Faecal water. Cells grown in biological triplicates to late log phase (T=6h) prior to harvest. Goal was to determine changes in gene expression caused by exposure to Faecal water, and changes in the metabolite profile of faecal water containing medium when incubated with actively growing C. difficile cells
Project description:The anaerobic and spore-forming bacterium Clostridium difficile has turned into an intensively studied species which can be attributed to increasing numbers of infections and rising costs for the health care system. This dataset represents a benchmark proteome of reference strain C. difficile 630erm and provides mass spectrometry based details on identified proteins which was generally missing from hitherto published datasets. An elaborate annotation and visualization of the 3764 open reading frames will serve as a valuable base for researchers trying to evaluate results of global expression studies. Furthermore, protein expression of late exponentially growing cells in the complex medium BHI and in C. difficile minimal medium was compared. Whereas abundance of proteins of DNA metabolism, protein synthesis and cell envelope showed no variation, enzymes for the biosynthesis of some vitamins and purine as well as proteins involved in butanoate fermentation differed significantly depending on the growth medium.
Project description:Clostridium difficile is an anaerobic spore-forming rod-shaped gram-positive bacterium that can infect both humans and animals. Most studies on the pathogenesis of C. difficile have focused on its toxins and their effect on the host cells. Recently, we utilized microarrays to identify conserved and divergent genes associated with virulence in C. difficile isolates from humans and animals. Our data provided the first clue toward a complex mechanism underlying host adaptation and pathogenesis. Microarray technology offers an efficient high-throughput tool to study the transcriptional profiles of pathogens and infected host cells. Transcriptomes of C. difficile after exposure to environmental and antibiotic stresses and those of human epithelial colorectal Caco-2 cells upon TcdA treatment have been analyzed. To our knowledge, there are still no reports on the transcriptomic study of host-pathogen interactions for C. difficile infection (CDI). In vitro analyses of interplay between host and pathogen are essential to unravel the mechanisms of infection and to investigate the host response to infection. We therefore employed microarrays to study both bacterial and human cellular transcriptome kinetics during CDI to Caco-2 cells. Here we present a large-scale analysis of transcriptional profiles to reveal molecular determinants playing a role in C. difficile pathogenesis and the host response. We found that there were 254 and 224 differentially-expressed genes after CDI in C. difficile and Caco-2 cells, respectively. These genes are clustered according to their functional categories and their potential roles in pathogenesis and host response are discussed. Our results will not only increase our understanding on the host-pathogen interaction, but may also provide targets for drug development. Clostridium difficile: Control vs Infection (time course) mRNA with genomic DNA of tested and reference strains Caco-2 cells: Control vs Infected with Clostridium difficile Time-course experiments of Caco-2 cells infected with C. difficile for 30, 60 and 120 min
Project description:We used microarray to identify the global gene expression profile of recombinant E.coli culture expressing repeating unit of the C. difficile toxin A (rARU) C-terminal region. Controlled condition batch runs were performed with E.coli expressing rARU at both restricted and unrestricted DO conditions for collecting samples of log and late-log phases at unrestricted and restricted DO.