Genotyping of Probiotic Bifidobacterium Longum Strains Using CRISPR Loci

Bifidobacterium longum strains are amongst the most widely used probiotics for human consumption due to their ability to modulate the host immune response, and withstand passage through the human gastrointestinal tract. CRISPR-Cas systems occur widely in bacteria and provide adaptive immunity via acquisition of DNA sequences from viruses and plasmids that constitute unique genetic records of immunization. We performed bioinformatics analyses of 66 B. longum genomes that identified 29 CRISPR loci in 25 strains. Each CRISPR locus was characterized and used for phylogenetic analyses. We identify various Type I and Type II CRISPR-Cas systems that are widespread in this species, notably I-C, I-E and II-C. Noteworthy, Type I-C systems showed extended CRISPR arrays, with extensive spacer diversity. The spacer sequences enable high-resolution genotyping of B. longum strains and revealed a common origin for some strains reflecting divergent evolution under putative selective pressure from invasive DNA. Analysis of CRISPR spacer origin also revealed their implication in phages and prophages targeting and novel PAM sequences. Altogether, results suggest that these immune systems are likely active in this species, and provide a basis for high-resolution genotyping. Furthermore, this study opens new avenues for the repurposing of the endogenous Cas machinery for genome editing applications to enhance their probiotic effect.