Effect of Pefloxacin on <i>Clostridioides difficile</i> R20291 Persister Cells Formation

<i>Clostridioides difficile</i> is a Gram-positive bacterium recognized for its ability to produce toxins and form spores. It is mainly accountable for the majority of instances of antibiotic-related diarrhea. <b>Background.</b> Bacterial persister represent a minor fraction of the population that shows temporary tolerance to bactericidal agents, and they pose considerable medical issues because of their link to the rise of antibiotic resistance and challenging chronic or recurrent infections. Our previous research has shown a persister-like phenotype associated with treatments that include pefloxacin. Nonetheless, the mechanism is still mostly unclear, mainly because of the difficulty in isolating this small group of cells. <b>Objectives.</b> To enhance the understanding of <i>C. difficile</i> persister cells, we made an enrichment and characterization of these cells from bacterial cultures during the exponential phase under pefloxacin treatment and lysis treatment. <b>Results.</b> We demonstrate the appearance of cells with lower metabolism and DNA damage. Furthermore, we noted the participation of toxin–antitoxin systems and Clp proteases in the generation of persister cells. <b>Conclusions.</b> This work demonstrates the formation of <i>C. difficile</i> persister cells triggered by a lethal concentration of pefloxacin.