Oral cholestyramine prevents the enrichment of diverse daptomycin-resistance mutations in intestinal Enterococcus faecium populations

<p><em><strong>Background and Objectives</strong></em>: Previously, we showed proof-of-concept in a mouse model that oral administration of cholestyramine prevented enrichment of daptomycin-resistant <em>Enterococcus faecium</em> in the gastrointestinal (GI) tract during daptomycin therapy. Cholestyramine binds daptomycin in the gut, which removes daptomycin selection pressure and so prevents the enrichment of resistant clones. Here, we investigated two open questions related to this approach: 1) can cholestyramine prevent the enrichment of diverse daptomycin mutations emerging <em>de novo</em> in the gut? 2) how does the timing of cholestyramine administration impact its ability to suppress resistance?</p> <p><em><strong>Methodology</strong></em>: Mice with GI <em>E. faecium</em> were treated with daptomycin with or without cholestyramine, and <em>E. faecium</em> was cultured from feces to measure changes in daptomycin susceptibility. A subset of clones was sequenced to investigate the genomic basis of daptomycin resistance.</p> <p><em><strong>Results</strong></em>: Cholestyramine prevented the enrichment of diverse resistance mutations that emerged de novo in daptomycin-treated mice. Whole-genome sequencing revealed that resistance emerged through multiple genetic pathways, with most candidate resistance mutations observed in the clsA gene. Additionally, we observed that cholestyramine was most effective when administration started prior to the first dose of daptomycin. However, beginning cholestyramine after the first daptomycin dose reduced the frequency of resistant <em>E. faecium</em> compared to not using cholestyramine at all.</p> <p><em><strong>Conclusions and Implications</strong></em>: Cholestyramine prevented the enrichment of diverse daptomycin-resistance mutations in intestinal <em>E. faecium</em> populations during daptomycin treatment, and it is a promising tool for managing transmission of daptomycin-resistant <em>E. faecium</em>.</p>