The influence of exposure to various concentrations of five antimicrobial agents on intracellular cytotoxin B production in Clostridioides difficile

Abstract

Background: Clostridioides difficile is an important cause of healthcare-associated diarrhea. Several anti- microbial agents are known to promote C. difficile infection (CDI). The impact of various concentrations of ampicillin (AMP), cefotaxime (CTX), clindamycin (CC), metronidazole (MTZ) and vancomycin (VAN) on intra-cellular cytotoxin B production was investigated in this study.

Methodology: Six clinical strains of C. difficile were grown at minimum inhibitory concentration (MIC) and sub-MIC concentrations of these antibiotics. Inoculum standardization was performed by Miles and Misra method. Intracellular toxin B production was detected using Vero cell cytotoxicity assay in sonicated cultures on days 1, 2, 3, 4, 5 and 7 days of incubation.

Results: There was a heterogeneous relationship between antibiotic exposure and the intra-cellular toxin production by the toxigenic strains. Clinical strains of C. difficile when exposed to MIC and sub-inhibitory concentrations of certain antibiotics produced high cytotoxin levels. All toxigenic isolates produced increased levels of cell-bound cytotoxin after exposure to antibiotics but there was no consistent pattern and the response to different doses varied considerably. Metronidazole was the most potent inducer of cell-bound cytotoxin followed by cefotaxime and clindamycin. Vancomycin induced the least amount of cytotoxin activity.

Conclusion: The effects of sub-inhibitory concentration of antibiotic that predispose to C. difficile infection may partially suppress the normal gut flora, allowing colonization and growth of C. difficile, and may affect the level of toxin produced.