SynopsisBackgroundAntimicrobial resistance (AMR) plays an important role in the pathogenesis and spread of Clostridioides difficile infection (CDI). Many antimicrobials, such as fluoroquinolones, have been associated with outbreaks of CDI globally.ObjectivesThis study aimed to characterise AMR among clinical C. difficile strains in Thailand, a country where the use of antimicrobials remains inadequately regulated.MethodsStool samples were screened for tcdB and positives were cultured. C. difficile isolates were characterised by toxin profiling and PCR ribotyping. Antimicrobial susceptibility testing was performed using an agar incorporation method, and whole-genome sequencing and AMR genotyping performed on a subset of strains.ResultsA total of 321 C. difficile strains were isolated from 326 stool samples. The most common toxigenic ribotype (RT) was RT 017 (18%), followed by RTs 014 (12%) and 020 (7%). There was a high resistance prevalence (≥ 10%) to clindamycin, erythromycin, moxifloxacin and rifaximin, and resistance prevalence was greatest among RT 017 strains. AMR genotyping revealed a significant correlation between resistance genotype and phenotype for moxifloxacin and rifampicin. The presence of erm-class genes was associated with high-level clindamycin and erythromycin resistance. Point substitutions on the penicillin-binding proteins (PBP1 and PBP3) were not sufficient to confer meropenem resistance, however, a Y721S substitution in PBP3 was associated with a slight increase in meropenem MIC. No resistance to metronidazole, vancomycin or fidaxomicin was observed.ConclusionThere was a large proportion of C. difficile RT 017 in Thailand and significant AMR among these strains. The concordance between AMR phenotype and genotype was significant.