Figure 4.

Locking the flagellar switch in the ON orientation exacerbates disease and increases persistence of C. difficile in a mouse model of CDI. Antibiotic-treated male and female C57BL/6 mice were inoculated with 100,000 spores of wildtype R20291 (WT), flg-Δ3 ON, and flg-Δ3 OFF. Mock-inoculated animals were included in each experiment. Data are combined from two independent experiments testing strains in 3 male and 3 female mice, for 12 total mice per strain. (a) CFU enumerated in fecal samples collected every 24 hours post-inoculation (p.i.). Asterisks indicate statistical comparison to WT data at that time point. (b) CFU per gram feces collected on d 6 p.i.; data for d 5–10 shown in Figure S4. (c) Animal weights determined every 24 hours post-inoculation, expressed as a percentage of the mouse’s weight at d 0. Asterisks indicate statistical comparison to mock data at that time point. (d) Animal weights at d 2 p.i.; data for d 1–6 shown in Figure S5. (e) Toxin titers in fecal samples calculated as the reciprocal of the highest dilution to cause ≥80% rounding of Vero cells. No cell rounding occurred when treated with diluted fecal contents from mock-inoculated animals. (a–e) Symbols indicate values from individual animals. (a, b, c) Bars indicate the medians. Statistical significance was determined using the Kruskal–Wallis test and Dunn’s posttest. (c, d) Bars indicate means and standard errors. Statistical significance was determined by one-way ANOVA with Tukey’s posttest. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.