Drivers of Clostridioides difficile hypervirulent ribotype 027 spore germination, vegetative cell growth and toxin production in vitro

Abstract Objectives Clostridioides difficile infection (CDI) is a considerable healthcare and economic burden worldwide. Faecal microbial transplant remains the most effective treatment for CDI, but is not at the present time the recommended standard of care. We hereby investigate which factors derived from a healthy gut microbiome might constitute the colonisation resistance barrier (CRB) in the gut, inhibiting CDI. Method CRB drivers pH, short chain fatty acid (SCFA), and oxidation-reduction potential (ORP) were investigated in vitro using C. difficile NAP1/BI/027. Readouts for inhibitory mechanisms included germination, growth, toxin production and virulence gene expression. pH ranges (3 – 7.6), SCFA concentrations (25 – 200mM) and ORP (-300 - +200mV) were manipulated in brain heart infusion broth cultures under anaerobic conditions to assess the inhibitory action of these mechanisms. Results Conclusion This study highlights the critical role that pH has in the CRB, regulating CDI in vitro and that SCFA can regulate C. difficile function independent of pH.