Adaptation mechanisms of Clostridioides difficile to auranofin and its impact on human gut microbiota

Auranofin (AF), a former rheumatoid polyarthritis treatment, gained renewed interest for its use as an antimicrobial. AF is an inhibitor of thioredoxin reductase (TrxB), a thiol and protein repair enzyme, with an antibacterial activity against several bacteria including C. difficile, an enteropathogen causing post-antibiotic diarrhea. Several studies demonstrated the effect of AF on C. difficile physiology, but the crucial questions of resistance mechanisms and impact on microbiota remain unaddressed. We explored potential resistance mechanisms by studying the impact of TrxB multiplicity and by generating and characterizing adaptive mutations. We showed that if mutants inactivated for trxB genes have a lower MIC of AF, the number of TrxBs naturally present in clinical strains does not impact the MIC. All stable mutations isolated after AF long-term exposure were in the anti-sigma factor of σ^B and strongly affect physiology. Finally, we showed that AF has less impact on human gut microbiota than vancomycin.