Antimicrobial susceptibility testing of Clostridioides difficile: a dual-site study of three different media and three therapeutic antimicrobials.

Objectives: Increasing resistance to antimicrobials used for the treatment of Clostridioides difficile infections necessitates reproducible antimicrobial susceptibility testing. Current guidelines take a one-size-fits-all approach and/or offer limited guidance. We investigated how the choice of medium affects measured MIC values across two sites.

Methods: We determined MIC values for the antimicrobials fidaxomicin, metronidazole and vancomycin for a representative collection of European C. difficile strains (n=235) using agar dilution on three different media: Brucella Blood Agar (BBA), Fastidious Anaerobe Agar supplemented with horse blood (FAA-HB) and Wilkins-Chalgren (WC) agar. The study was conducted at two sites to compare reproducibility. Useability (ease of preparation of the media as well as read-out of the assay) was assessed through a survey.

Results: We found that all media result in highly consistent aggregated MIC data for all antibiotics, with MIC and MIC within 2-fold of each other across sites. For fidaxomin, MIC values on WC were lower than on the other media (MIC: WC = 0.125-0.25 mg/L; BBA and FAA-HB = 0.5 mg/L). Metronidazole showed the lowest MIC on BBA, and the highest on WC (MIC: WC = 2 mg/L; BBA = 0.5-1 mg/L; FAA-HB: 1-2 mg/L). For vancomycin, MIC values were similar across media (MIC: all media = 1-2 mg/L). Though absolute values for individual isolates differed between sites, identified resistant isolates were similar. Results obtained on FAA-HB were most consistent between sites and results obtained on WC showed the most divergence. FAA-HB was positively evaluated in the usability survey.

Conclusions: This study shows medium-dependent differences in C. difficile MICs for at least two antimicrobials across two sites. We suggest the use of FAA-HB to align with general EUCAST recommendations for susceptibility testing of anaerobic bacteria and deposited reference strains for standard susceptibility testing of C. difficile to increase interlaboratory reproducibility.