A targeted cell lysis mechanism facilitates toxin release in Clostridioides difficile

Clostridioides difficile infection is associated with the production of two large toxins, TcdA and TcdB. The toxins are encoded in a pathogenicity locus along with a phage-like holin protein, TcdE, the remnants of a phage endolysin, TcdL, and a positive transcriptional regulator, TcdR. Previous studies investigating the mechanism of toxin secretion from the bacterium have yielded disparate results as to whether the bacteria undergo lysis and whether TcdE is required. Here, we demonstrate that TcdE and TcdA are only expressed in a small subset of cells in culture. The cells with high TcdE and TcdA expression show disrupted membranes, consistent with a form of bacterial lysis. The artificial overexpression of TcdE and TcdA through TcdR induction promotes tcdE -dependent bacterial lysis and cell death, even in 630Δerm, a strain previously thought to have tcdE -independent toxin secretion. We propose a general mechanism where a minority subpopulation undergoes TcdE-mediated lysis to release toxins, reconciling studies with differences stemming from strain variability and population-level assays.