UNRAVELING THE COMPLEXITY OF CDEC EXPRESSION DURING CLOSTRIDIOIDES DIFFICILE SPORULATION: INSIGHTS INTO THE ROLE OF SIGMA FACTORS AND NON-CODING RNA ON CDEC EXPRESSION

Abstract

Gram-positive, obligatory anaerobic Clostridioides difficile (C. difficile) is a spore- forming, obligate anaerobic bacterium that causes diarrhea and other healthcare-associated illnesses globally. This study aimed to identify the sigma factor boxes in the promoter region of cdeC that affect the early and late stages of C. difficile sporulation. Therefore, we first identified the sigma factors' positions in the promoter region. Then, we constructed transcriptional fusions containing a different deletion of these sigma factors and introduced them into the C. difficile R20291CM210 WT strain. However, no detectable fluorescence was observed due to lack of sporulation in the C. difficile R20291CM210. Both increasing the time of incubation and increasing thiamphenicol concentrations failed to induce sporulation for R20291CM210. After that, we introduced all transcriptional fusions into the C. difficile R20291CM196 WT strain. We found out that only P∆ncRNA-cdeC-mScarlett transcriptional fusion that contains the promoter region with a deletion of the ncRNA shows a modest fluorescence signal compared to other fusions. These results suggest that ncRNA located in the promoter region of cdeC may have an activity for cdeC expression during sporulation.