Clostridioides difficile infection (CDI) is associated with significant patient morbidity and mortality worldwide. Recurrent CDI is extremely common due to the ability of C. difficile to form spores, which involves a specific set of proteins called penicillin-binding proteins (PBPs). Cephamycin antibiotics can inhibit C. difficile sporulation by targeting the sporulation-specific PBP, CdSpoVD [1]. Recently, a cephamycin-resistant accessory spore-PBP, CdSpoCR, has been identified in C. difficile. CdspoCR exists in a gene complex alongside genes encoding a putative regulatory signalling system comprised of an antibiotic sensor CdSpoCR_R and a transcriptional repressor CdSpoCR_I. This work aimed to investigate the currently unknown role of this signal transduction system in CdSpoCR expression in C. difficile.
In this study, CdspoCR_R was disrupted in a CdspoCR-positive C. difficile stain. Sporulation assays were then performed using cephamycins to examine changes in sporulation and antibiotic resistance profiles. Following antibiotic exposure, spore counts were high for the wild-type strain, however, no spores were observed for the CdspoCR_R mutant. This result suggests that this putative sensor protein is important for CdSpoCR expression following cephamycin exposure. Additionally, PBP profile analysis and mass spectrometry demonstrated a loss of CdSpoCR in cephamycin-treated CdspoCR_R mutants. These findings further suggest that CdspoCR_R disruption impairs CdSpoCR production and abolishes sporulation in the presence of cephamycins. This study has confirmed a role for CdSpoCR_R in CdSpoCR-mediated cephamycin resistance in C. difficile. Overall, these findings may facilitate the development of anti-sporulation agents capable of reducing spore-mediated transmission and recurrence of CDI.