Characterization of a Type II-A CRISPR-Cas System in .

and its virulent phages are important members of the human oral microbiota. is also the primary causal agent of dental caries. To survive in this ecological niche, must encode phage defense mechanisms, which include CRISPR-Cas systems. Here, we describe the CRISPR-Cas type II-A system of strain P42S, which was found to display natural adaptation and interference activity in response to phage infection and plasmid transformation. Newly acquired spacers were integrated both at the 5' end of the CRISPR locus and ectopically. In comparisons of the genes of P42S to those of other strains of , , , and appear to be highly conserved within the species. However, more diversity was observed with While the nuclease domains of Cas9 (SmCas9) are conserved, the C terminus of the protein, including the protospacer adjacent motif (PAM) recognition domain, is less conserved. In support of these findings, we experimentally demonstrated that the PAMs associated with SmCas9 of strain P42S are NAA and NGAA. These PAMs are different from those previously reported for the CRISPR-Cas system of the model strain UA159. This study illustrates the diversity of CRISPR-Cas type II-A systems that can be found within the same bacterial species. CRISPR-Cas is one of the mechanisms used by bacteria to defend against viral predation. Increasing our knowledge of the biology and diversity of CRISPR-Cas systems will also improve our understanding of virus-bacterium interactions. As CRISPR-Cas systems acquiring novel immunities under laboratory conditions are rare, strain P42S provides an alternative model to study the adaptation step, which is still the least understood step in CRISPR-Cas biology. Furthermore, the availability of a natural Cas9 protein recognizing an AT-rich PAM opens up new avenues for genome editing purposes.