Acquired CRISPR spacers and rhamnose-glucose polysaccharide defects confer resistance to phage ɸAPCM01.

is a major cause of dental caries worldwide. Targeted therapeutic strategies to eradicate include oral phage rinses. In this study, we investigated how phage resistance develops in . As a model phage, we used ɸAPCM01, which is known to infect a serotype e strain. We isolated and sequenced the genomes of 15 spontaneous resistant mutants and found that 10 had acquired novel CRISPR spacers targeting the phage, with a total of 18 new spacers identified. Additionally, eight strains contained mutations in rhamnose-glucose polysaccharide (RGP) biosynthetic genes, three of which also acquired spacers. Only the mutants exhibited defects in phage absorption, supporting the role of these cell surface glycans as the phage receptor. Mutations in and the newly identified gene led to severe cell division defects and impaired biofilm formation, the latter of which shared by the mutant. Thus, mutations confer phage resistance but impose severe fitness costs, limiting pathogenic potential. Surprisingly, we found that ɸAPCM01 was capable of binding to and injecting its genome into UA159, a model serotype c strain. However, UA159 was resistant to infection due to an unknown post-entry defense mechanism. Consequently, ɸAPCM01 has the potential to infect both major serotypes associated with dental caries.