# P24 Antimicrobial susceptibility profiles of hypervirulent strains of Clostridioides difficile of pig and human origin

## TL;DR

This study examines antibiotic resistance in hypervirulent Clostridioides difficile strains from pigs and humans, finding resistance to certain antibiotics but not to vancomycin or metronidazole.

## Contribution

The study provides new insights into antimicrobial resistance patterns in C. difficile strains from both pigs and humans, highlighting the potential role of agricultural antibiotic use.

## Key findings

- All isolates were susceptible to vancomycin and metronidazole but showed resistance to cefoxitin, ceftriaxone, and others.
- ST11 isolates were resistant to ciprofloxacin, with delafloxacin showing lower resistance.
- Whole genome sequencing revealed resistance determinants for quinolones and tetracyclines in most isolates.

## Abstract

Clostridioides difficile places an increasing burden on healthcare because of emerging hypervirulent strains such as RT078/ST11 that may be resistant to commonly prescribed antibiotics e.g. fluoroquinolones. Pigs are often colonized by RT078/ST11 and act as potential reservoir for human C. difficile infection (CDI). They are the highest consumers of veterinary antimicrobials in many countries.1 We investigated the susceptibility of C. difficile to antibiotics that are commonly prescribed and/or used to treat CDI.

Sequenced C. difficile (humans=50, pigs=40), collected as part of a One Health investigation into the epidemiology of pig and human C. difficile, were chosen for phenotypic antimicrobial susceptibility testing (AST). Strains included ST11 (n=70), ST44 (n=9), ST16 (n=6), ST8 (n=5). Susceptibility to a panel of antibiotics were determined using ANAERO3 Sensititre™ plates and Etest®, according to the manufacturer’s instructions. Selected C. difficile ST11 isolates (n=30) were tested for ciprofloxacin and delafloxacin susceptibility using Etest®. Breakpoints were interpreted as per EUCAST, CLSI and Freeman.2C. difficile genomes (n=90) were screened for AMR markers using NCBI AMRFinderPlus.

Phenotypic testing found all isolates were susceptible to vancomycin and metronidazole, and to penicillin, amoxicillin, amoxiclav, piperacillin/tazobactam, piperacillin, imipenem and chloramphenicol. Resistance was found to cefoxitin (100%), ceftriaxone (100%), erythromycin (48.94%), clindamycin (28.72%), moxifloxacin (48.94%), tetracycline (31.91%) and rifampicin (2.13%). All 30 ST11 isolates were resistant to ciprofloxacin, while the mean MIC for delafloxacin was 0.47 mg/L. There was no difference between resistance rate and C. difficile host. Bioinformatic analysis of 90 C. difficile genomes revealed resistance determinants for quinolones (87%) and tetracyclines (69%).

No resistance to vancomycin and metronidazole in these C. difficile isolates is reassuring, although larger datasets for surveillance should be investigated. Finding resistance determinants in the isolates in this study demonstrates that WGS is a valuable tool for detecting AMR. Quinolone and tetracycline resistance may be linked with use of these antibiotics on Irish pig farms (64.1% and 85.1% of farms, respectively).1