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What's a SNP between friends: The lineage of Clostridioides difficile R20291 can effect research outcomes.

ABSTRACT: Clostridioides difficile R20291 is the most studied PCR-Ribotype 027 isolate. The two predominant lineages of this hypervirulent strain, however, exhibit substantive phenotypic differences and possess genomes that differ by a small number of nucleotide changes. It is important that the source of R20291 is taken into account in research outcomes.

SUBMITTER: Monteford J

PROVIDER: S-EPMC8556159 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Clostridioides difficile R20291

Project description:Clostridium difficile R20291 genome sequencing project

| PRJNA38039 | ENA

Clostridioides difficile R20291

Project description:Clostridioides difficile R20291 Transcriptome or Gene expression

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Clostridioides difficile R20291

Project description:Clostridioides difficile R20291 Transcriptome or Gene expression

| PRJNA416052 | ENA

Clostridioides difficile R20291

Project description:Clostridioides difficile CRISPR-cas9 system

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Clostridioides difficile R20291

Project description:Clostridioides difficile R20291 Genome sequencing and assembly

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Cwl0971, a novel peptidoglycan hydrolase, plays pleiotropic roles in Clostridioides difficile R20291.

Project description:Clostridioides difficile is a Gram-positive, spore-forming, toxin-producing anaerobe that can cause nosocomial antibiotic-associated intestinal disease. Although the production of toxin A (TcdA) and toxin B (TcdB) contribute to the main pathogenesis of C. difficile, the mechanism of TcdA and TcdB release from cell remains unclear. In this study, we identified and characterized a new cell wall hydrolase Cwl0971 (CDR20291_0971) from C. difficile R20291, which is involved in bacterial autolysis. The gene 0971 deletion mutant (R20291Δ0971) generated with CRISPR-AsCpfI exhibited significantly delayed cell autolysis and increased cell viability compared to R20291, and the purified Cwl0971 exhibited hydrolase activity for Bacillus subtilis cell wall. Meanwhile, 0971 gene deletion impaired TcdA and TcdB release due to the decreased cell autolysis in the stationary/late phase of cell growth. Moreover, sporulation of the mutant strain decreased significantly compared to the wild type strain. In vivo, the defect of Cwl0971 decreased fitness over the parent strain in a mouse infection model. Collectively, Cwl0971 is involved in cell wall lysis and cell viability, which affects toxin release, sporulation, germination, and pathogenicity of R20291, indicating that Cwl0971 could be an attractive target for C. difficile infection therapeutics and prophylactics.

| S-EPMC11217927 | biostudies-literature

Hypervirulent R20291 Clostridioides difficile spores show disinfection resilience to sodium hypochlorite despite structural changes.

Project description:BackgroundClostridioides difficile is a spore forming bacterial species and the major causative agent of nosocomial gastrointestinal infections. C. difficile spores are highly resilient to disinfection methods and to prevent infection, common cleaning protocols use sodium hypochlorite solutions to decontaminate hospital surfaces and equipment. However, there is a balance between minimising the use of harmful chemicals to the environment and patients as well as the need to eliminate spores, which can have varying resistance properties between strains. In this work, we employ TEM imaging and Raman spectroscopy to analyse changes in spore physiology in response to sodium hypochlorite. We characterize different C. difficile clinical isolates and assess the chemical's impact on spores' biochemical composition. Changes in the biochemical composition can, in turn, change spores' vibrational spectroscopic fingerprints, which can impact the possibility of detecting spores in a hospital using Raman based methods.ResultsWe found that the isolates show significantly different susceptibility to hypochlorite, with the R20291 strain, in particular, showing less than 1 log reduction in viability for a 0.5% hypochlorite treatment, far below typically reported values for C. difficile. While TEM and Raman spectra analysis of hypochlorite-treated spores revealed that some hypochlorite-exposed spores remained intact and not distinguishable from controls, most spores showed structural changes. These changes were prominent in B. thuringiensis spores than C. difficile spores.ConclusionThis study highlights the ability of certain C. difficile spores to survive practical disinfection exposure and the related changes in spore Raman spectra that can be seen after exposure. These findings are important to consider when designing practical disinfection protocols and vibrational-based detection methods to avoid a false-positive response when screening decontaminated areas.

| S-EPMC9986864 | biostudies-literature

What's a SNP between friends: The influence of single nucleotide polymorphisms on virulence and phenotypes of Clostridium difficile strain 630 and derivatives.

Project description:Clostridium difficile is a major cause of antibiotic induced diarrhea worldwide, responsible for significant annual mortalities and represents a considerable economic burden on healthcare systems. The two main C. difficile virulence factors are toxins A and B. Isogenic toxin B mutants of 2 independently isolated erythromycin-sensitive derivatives (630E and 630Δerm) of strain 630 were previously shown to exhibit substantively different phenotypes. Compared to 630, strain 630E and its progeny grow slower, achieve lower final cell densities, exhibit a reduced capacity for spore-formation, produce lower levels of toxin and are less virulent in the hamster infection model. By the same measures, strain 630Δerm and its derivatives more closely mirror the behavior of 630. Genome sequencing revealed that 630Δerm had acquired 7 unique Single Nucleotide Polymorphisms (SNPs) compared to 630 and 630E, while 630E had 9 SNPs and a DNA inversion not found in the other 2 strains. The relatively large number of mutations meant that the identification of those responsible for the altered properties of 630E was not possible, despite the restoration of 3 mutations to wildtype by allelic exchange and comparative RNAseq analysis of all 3 strains. The latter analysis revealed large differences in gene expression between the 3 strains, explaining in part why no single SNP could restore the phenotypic differences. Our findings suggest that strain 630Δerm should be favored over 630E as a surrogate for 630 in genetic-based studies. They also underline the importance of effective strain curation and the need to genome re-sequence master seed banks wherever possible.

| S-EPMC5626343 | biostudies-literature

Transcriptomic profile of a Clostridioides difficile rsbW deletion mutant compared to parental R20291 strain

Project description:RNA-seq was conducted to uncover the regulatory roles of RsbW in the lifestyle of Clostridioides difficile strain R20291. RNA were extracted from planktonic cultures, which were grown in BHI-S (Sigma-Aldrich, USA) to early stationary phase (10h). The cells were re-suspended in LETS buffer and lysed using Fast-prep 24 instrument (MP Bioscience) and RNA was extracted using 1 ml Trizol (Ambion, USA). Samples were cleaned and sequenced by Novogene Company Limited, PE150 with approximately 10 million reads per sample. After QC, reads were aligned to R20291 (NC_013316.1) with Bowtie2 and readcounts were generated with Samtools and Bedtools. Differential gene expression analysis was conducted with DESeq2 with a p-value of 0.05, differential gene expression was determined with ± -2 logFC.

2022-08-22 | E-MTAB-12114 | biostudies-arrayexpress

Characterization of Exosporium Layer Variability of <i>Clostridioides difficile</i> Spores in the Epidemically Relevant Strain R20291.

Project description:Clostridioides difficile is a Gram-positive anaerobic intestinal pathogenic bacterium and the causative agent of antibiotic-associated diarrhea. C. difficile spore is a dormant state which acts as a vehicle of transmission and infection. In C. difficile spores, the outermost exosporium layer is the first barrier of interaction with the host and should carry spore ligands involved in spore-host interactions. C. difficile forms two types of spores (i.e., thin and thick exosporium layers). In this communication, we contribute to understand several biological aspects of these two exosporium morphotypes. By transmission electron microscopy, we demonstrate that both exosporium morphotypes appear simultaneously during sporulation and that spore-coat laminations are formed under anaerobic conditions. Nycodenz density-gradient allows enrichment of spores with a thick-exosporium layer morphotype and presence of polar appendage. Using translational fluorescent fusions with exosporium proteins BclA3, CdeA, CdeC, and CdeM as well as with several spore coat proteins, we observed that expression intensity and distribution of SNAP-translational fusions in R20291 strain is highly heterogeneous. Electron micrographs demonstrate that multicopy expression of CdeC, but not CdeM, SNAP translational fusion, increases the abundance of the thick exosporium morphotype. Collectively, these results raise further questions on how these distinctive exosporium morphotypes are made during spore formation.

| S-EPMC7343902 | biostudies-literature

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