Antimicrobial Susceptibility Testing of Clostridioides Difficile: a Dual-Site Study of Three Different Media and Three Therapeutic Antimicrobials

Jane Freeman,Ingrid M.J.G. Sanders,Céline Harmanus, Emma V. Clark, Andrea M. Berry,Wiep Klaas Smits

Clinical microbiology and infection the official publication of the European Society of Clinical Microbiology and Infectious Diseases（2025）

Leeds Teaching Hospitals National Health Service (NHS) Trust & University of Leeds | Leiden University Medical Center

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Abstract

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 MIC50 and MIC90 within 2-fold of each other across sites. For fidaxomin, MIC values on WC were lower than on the other media (MIC90: 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 (MIC90: WC = 2 mg/L; BBA = 0.5-1 mg/L; FAA-HB: 1-2 mg/L). For vancomycin, MIC values were similar across media (MIC90: 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.

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Key words

Agar dilution,Antimicrobial susceptibility test,Clostridioides difficile,fidaxomicin,metronidazole,vancomycin

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【要点】：研究考察了不同培养基对艰难梭菌对抗微生物药物最小抑菌浓度（MIC）的影响，提出使用Fastidious Anaerobe Agar补充马血（FAA-HB）培养基以提高实验室间测试的重复性。

【方法】：通过在两个不同位点使用三种培养基（Brucella Blood Agar，Fastidious Anaerobe Agar补充马血，Wilkins-Chalgren agar）对235株欧洲艰难梭菌代表菌株进行琼脂稀释法测定三种治疗性抗生素（fidaxomicin、metronidazole、vancomycin）的MIC值。

【实验】：研究在两个位点进行以比较重复性，并通过调查评估了培养基的易用性；结果发现各培养基得到的总体MIC数据高度一致，但个别抗生素在不同培养基上的MIC值存在差异，FAA-HB培养基在两个位点间的结果最一致，且易用性评价积极。

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