| Summary: | In recent decades, the incidence of <i>Clostridioides difficile</i> infection (CDI) has remained high in both community and health-care settings. With the increasing rate of treatment failures and its ability to form spores, an alternative treatment for CDI has become a global priority. We used the microdilution assay to determine minimal inhibitory concentrations (MICs) of vancomycin and teicoplanin against 30 distinct <i>C. difficile</i> strains isolated from various host origins. We also examined the effect of drugs on spore germination and outgrowth by following the development of OD<sub>600</sub>. Finally, we confirmed the spore germination and cell stages by microscopy. We showed that teicoplanin exhibited lower MICs compared to vancomycin in all tested isolates. MICs of teicoplanin ranged from 0.03–0.25 µg/mL, while vancomycin ranged from 0.5–4 µg/mL. Exposure of <i>C. difficile</i> spores to broth supplemented with various concentrations of antimicrobial agents did not affect the initiation of germination, but the outgrowth to vegetative cells was inhibited by all test compounds. This finding was concordant with aberrant vegetative cells after antibiotic treatment observed by light microscopy. This work highlights the efficiency of teicoplanin for treatment of <i>C. difficile</i> through prevention of vegetative cell outgrowth.
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