Fig. 4

DOC represses both sporulation and toxin production of C. difficile. a CFU of viable cells (black) and heat-resistant spores (gray) in the absence or presence of 240 µM CHO or 240 µM DOC. The sporulation assays performed with unwashed biofilms. b Relative expression levels of tcdA and tcdB measured by qRT-PCR in cells grown in the absence or presence of 240 µM DOC or 240 µM CHO. Relative expression levels (∆∆Ct method) are the ratio of mRNA level in the presence of bile salts to the mRNA level in the absence of bile salts. Reactions of qRT-PCR were normalized using DNA polIII (CD1305), rpoA (CD0098), pgi (CD3285), and tpi (CD3172). c ELISA-based quantification of TcdA production by cells grown in the absence or presence of 240 µM DOC or 240 µM CHO. Concentrations were standardized to the amount of protein as measured by the Bradford method. d ELISA-based quantification of TcdA release into the supernatant by cells grown in the absence or presence of 240 µM DOC or 240 µM CHO. Asterisks indicate statistical significance determined with a two-way ANOVA followed by a Fisher LSD test (***p ≤ 0.001, ****p ≤ 0.0001 vs BHIS). Each bar represents the mean of at least 6 biological replicates performed on different days and error bars represent the standard error of the mean