Our experiments were performed in a glucose free TY broth medium, since glucose is a known repressor of toxin production [17] . Indeed, we observed earlier and higher levels of toxin production in TY broth than in the commonly used Brain-Heat-Infusion broth (BHIS) based media, that does contain low amounts of glucose (0.2%) and to which frequently cysteine is added. However, also in BHIS we did not observe a significant effect of a tcdC deletion on toxin expression (data not shown).