Cas9 modulates Campylobacter jejuni virulence traits inside intestinal epithelial cells.

The CRISPR-associated protein 9 (Cas9) produced by disease-associated strains of Campylobacter jejuni contributes to full virulence, including immune evasion and bacterial survival inside eukaryotic cells. In this work, we explored the role of C. jejuni Cas9 (CjeCas9) in cell envelope integrity, antibiotic resistance, intracellular survival inside Caco-2 intestinal epithelial cells and Toll-like receptor 2 (TLR-2) activation. We show that CjeCas9 modulates the permeability of the C. jejuni cell envelope, sialylated lipooligosaccharide expression and susceptibility to ciprofloxacin, the most commonly prescribed antibiotic to treat C. jejuni infections. Moreover, we reveal that WT production of CjeCas9 increased intracellular survival of C. jejuni inside Caco-2 intestinal epithelial cells by a factor of 550 compared to the respective cas9 gene deletion mutant and that intracellular survival was associated with the activation of TLR-2. In conclusion, we established that CjeCas9 modulates C. jejuni (intracellular) virulence traits, including intracellular survival.