The microbial derived bile acid lithocholate and its epimers inhibitClostridioides difficilegrowth and pathogenicity while sparing members of the gut microbiota

Abstract

AbstractC. difficileinfection (CDI) is associated with antibiotic usage, which disrupts the indigenous gut microbiota and causes the loss of microbial derived secondary bile acids that normally provide protection againstC. difficilecolonization. Previous work has shown that the secondary bile acid lithocholate (LCA) and its epimer isolithocholate (iLCA) have potent inhibitory activity against clinically relevantC. difficilestrains. To further characterize the mechanisms by which LCA and its epimers iLCA and isoallolithocholate (iaLCA) inhibitC. difficile,we tested their minimum inhibitory concentration (MIC) againstC. difficileR20291, and a commensal gut microbiota panel. We also performed a series of experiments to determine the mechanism of action by which LCA and its epimers inhibitC. difficilethrough bacterial killing and effects on toxin expression and activity. Here we show that epimers iLCA and iaLCA strongly inhibitC. difficilegrowthin vitrowhile sparing most commensal Gram-negative gut microbes. We also show that iLCA and iaLCA have bactericidal activity againstC. difficile,and these epimers cause significant bacterial membrane damage at subinhibitory concentrations. Finally, we observe that iLCA and iaLCA decrease the expression of the large cytotoxintcdAwhile LCA significantly reduces toxin activity. Although iLCA and iaLCA are both epimers of LCA, they have distinct mechanisms for inhibitingC. difficile. LCA epimers, iLCA and iaLCA, represent promising compounds that targetC. difficilewith minimal effects on members of the gut microbiota that are important for colonization resistance.ImportanceIn the search for a novel therapeutic that targetsC. difficile, bile acids have become a viable solution. Epimers of bile acids are particularly attractive as they may provide protection againstC. difficilewhile leaving the indigenous gut microbiota largely unaltered. This study shows that iLCA and iaLCA specifically are potent inhibitors ofC. difficile, affecting key virulence factors including growth, toxin expression and activity. As we move toward the use of bile acids as therapeutics, further work will be required to determine how best to deliver these bile acids to a target site within the host intestinal tract.