During infection, Clostridium difficile survives in phage-rich gut communities by relying on efficient anti-invader defence systems including CRISPR-Cas for prokaryotic adaptive immunity. CRISPR arrays are composed of repeat sequences separated by spacers matching with and originating from foreign DNA. They are transcribed into a pre-crRNA, which is cleaved by the Cas proteins to give CRISPR RNAs (crRNAs). In complex with Cas proteins, crRNAs interfere with phage infection and plasmid conjugation by recognizing foreign DNA and targeting it for destruction. crRNAs are among the most abundant ncRNAs detected by RNAseq (Soutourina et al, PLos Genetics, 2013). The originality of C. difficile CRISPR-Cas system is the presence of an unusually large set of CRISPR arrays, the presence of 2 sets of cas genes and the prophage location of several CRISPR arrays. We have recently shown active expression and processing of CRISPR RNAs from multiple CRISPR arrays and provided experimental evidence for CRISPR-Cas system functionality in C. difficile. Through genome sequencing and host-range analysis of several new C. difficile phages and plasmid conjugation experiments we demonstrated the defensive function of CRISPR-Cas system in both reference and epidemic C. difficile strains. Altogether, these data emphasize the original features of active C. difficile CRISPR-Cas system that might be important for C. difficile survival during its infection cycle (Boudry et al, mBio, 2015). This sets the stage for further mechanistic and physiological analyses of CRISPR-Cas-mediated interactions of important human pathogen with its genetic parasites.
We have recently shown the possibility of redirecting this endogenous CRISPR-Cas system to autoimmunity, which allows efficient editing of the C. difficile genome. The new method expands the arsenal of genetic tools available for the study of C. difficile and can serve as a basis for new strategies to fight C. difficile infections (Maikova et al, AEM, 2019; Maikova et al, Frontiers in Microbiology, 2018).