Biology subjects

Barbut, F.

Publications and source records attributed to Barbut, F..

3 recordsLinked to original sources

bioRxiv2024

In vitro biofilms of Clostridioides difficile undomesticated strains: Morphology and properties according to strain diversity

Clostridioides difficile is a Gram-positive, spore-forming, obligate anaerobe and an entero-pathogen representing a One Health problem. C. difficile infections are difficult to treat and recurrences frequent. C. difficile biofilms may play a role in persistence. We studied the biofilms formed in vitro by a collection of C. difficile undomesticated strains (n=27, 20 toxigenic and 7 non-toxigenic) of 11 PCR-ribotypes (WRT) and isolated from equids. After growth in rich BHIS medium supplemented with glucose for 48h, the biomass of biofilms stained by crystal violet, i. e. adhesive to the surface, was measured, while intact biofilms were observed in situ, by confocal laser scanning microscopy, to determine their total biovolume and 3D morphology. All biofilms were relatively similar, even though adhesive, stained biofilms of non-toxigenic strains of ribotype 009 significantly displayed highest biomass. These results suggested that strains of ribotype 009 were able to form a highly cohesive and polystyrene-adhesive biofilm than any other strains. 8 common, representative ribotypes in equids, including 6 clinically important ribotypes in humans, were then selected and one strain chosen for each of them. Their biofilms and planktonic cultures grown under the same conditions were compared for sporulation, toxin production and susceptibility to vancomycin at a highly inhibitory concentration (8 MIC). In both the biofilm and planktonic culture of all selected strains, sporulation was low and toxin production undetectable. The addition of vancomycin at T0=24h followed by a 24 hours exposure barely reduced the viability of neither planktonic cultures nor biofilms of any strains. On the contrary, for all selected strains, addition of vancomycin at T0=6h followed by a 24 hours exposure significantly and more efficiently reduced the viability of planktonic cultures than that of biofilms, which was not decreased at all in the case of strains of ribotype WRT 005, 009 and 035. The biofilms of these selected strains of eight ribotypes, compared to their planktonic cultures, therefore showed a higher tolerance to vancomycin, suggesting that they could play a role in persistence.

microbiology↗

bioRxiv2024

Clostridioides difficile in Equidae necropsied in Normandie, France from 2019 to 2021: common carriage of toxigenic strains and virulence of emerging ribotype 017

Clostridioides difficile is an anaerobic, spore-forming enteropathogen, which is much more extensively studied in humans than animals, despite growing evidence supporting its importance in One Health. We evaluated C. difficile occurrence, diversity, circulation and virulence in French Equidae (n=100) after their necropsy in northwestern France (Normandy), from 2019 to 2021. We systematically recovered all cecal contents and any watery (possibly diarrheal) intestinal contents. We isolated C. difficile strains, determined their toxin gene profile by PCR and established their PCR-ribotype according to the WEBRIBO database. We also performed free toxin detection. Twenty-seven Equidae were positive for C. difficile and twenty had a toxigenic strain, including one animal also colonized by a non-toxigenic strain. Toxigenic isolates belonged to eight common ribotypes: i) 078 and 126 (tcdA+ tcdB+ cdt+ according to multiplex PCR, i.e. able to produce toxin A, toxin B and binary toxin), ii) 005, 012, 020, AI-53 and FR227 (tcdA+ tcdB+), and iii) 017 (tcdB+). Non-toxigenic isolates were of ribotypes 009, 035 and 439. Ribotypes 017 and 009 were predominant (n=5). In two cases, Equidae of the same premises shared the same ribotype, either 020 or 009. Free toxins were detected in four animals: they displayed signs of diarrhea and a C. difficile of ribotype 126 (n=1) or 017 (n=3) as the only detected enteropathogen, suggesting a C. difficile infection (CDI). Three of them had received antibiotics. Two had died from an entero-toxic infection, for which C. difficile ribotype 017 was the only identified cause. French Equidae were found to display common pathogenic C. difficile, and ribotype 017 was highly virulent. These findings are of concern from a One Health perspective. ImportanceC. difficile, a major enteropathogen widely disseminated in the environment, is a One Health issue. Animals are raising concern as human contamination sources. Equidae are in close contact with humans and also develop post-antibiotic and healthcare-associated CDIs. The systematic survey of Equidae necropsied from 2019 to 2021 in the leading region for horse breeding in France, revealed that 20% harbored pathogenic strains. These belonged to clinically important ribotypes, raising the possibility of cross-species, possibly zoonotic transmission. Free toxins, which are rarely tested in animals, were detected in four animals with signs of diarrhea and a toxigenic C. difficile as the only identified enteropathogen, suggesting CDI. In two of them, C. difficile ribotype 017 was the only identified cause of entero-toxic disease and death. French Equidae could play a role in the dissemination of pathogenic C. difficile and notably ribotype 017. They should be surveilled carefully from a One Health perspective.

microbiology↗

bioRxiv2023

A monoclonal antibody collection for C. difficile typing

Clostridioides difficile is the leading cause of antibiotic-associated diarrhea and pseudomembranous colitis in adults. Various C. difficile strains circulate currently, associated with different outcomes and antibiotic resistance profiles. However, most studies still focus on the reference strain 630 that does not circulate anymore, partly due to the lack of immunological tools to study current clinically important C. difficile PCR ribotypes. Herein, we immunized mice expressing human variable antibody genes with the Low Molecular Weight (LMW) subunit of the surface layer protein SlpA from various C. difficile strains. Monoclonal antibodies purified from hybridomas bound LMW with high-affinity and whole bacteria from current C. difficile ribotypes with different cross-specificities. This first collection of anti-C. difficile mAbs represent valuable tools for basic and clinical research.

immunology↗