CRISPR spacer profiling and prophage mining reveal diverse bacteriophages associated with Streptococcus Mutans.

BACKGROUND

Streptococcus mutans is a key cariogenic bacterium. Current antimicrobials lack species specificity, while phage-based approaches remain experimental and require more S. mutans phage isolates.

OBJECTIVE

To profile the diversity of S. mutans-associated phages and strain-level heterogeneity in phage exposure using genome-informed CRISPR spacer and prophage analyses.

MATERIALS AND

METHODS

We compiled 944 publicly available S. mutans genomes and dereplicated them into 735 non-redundant strains. CRISPR-Cas systems, spacers, spacer targets, and putative prophages were identified, quality-assessed, and functionally annotated. Phylogenetic relationships of (pro)phages were evaluated using terminase large subunit proteins, and comparative genomics compared spacer-positive and spacer-negative strains.

RESULTS

CRISPR systems were detected in 548/735 strains, yielding 14,263 spacers, 1,864 phage-targeting spacers mapped to 110 viral genomes, including 41 cultured isolates, 51 metagenome-assembled phages, and 18 uncultured viral genomes. The most frequently targeted cultured phage was phiKSM96, whereas metagenome-assembled Caudoviricetes ctNo011 showed broader targeting. Prophage mining identified 186 regions in 130 strains, including 37 of ≥ medium quality and elements related to ctNo011 and phiKSM96. TerL phylogeny showed that most high-quality endogenous prophages clustered with phiKSM96 and ctNo011.

CONCLUSION

These findings reveal a vast, uncultivated phage repertoire targeting S. mutans, providing a critical genomic roadmap to guide the future isolation of novel phages for caries prevention.