Investigating CRISPR spacer targets and their impact on genomic diversification of Streptococcus mutans

Abstract

CRISPR-Cas is a bacterial immune system that restricts the acquisition of mobile DNA elements. These systems provide immunity against foreign DNA by encoding CRISPR spacers that help target DNA if it re-enters the cell. In this way, CRISPR spacers are a type of molecular tape recorder of foreign DNA encountered by the host microorganism. Here, we extracted [~]8,000 CRISPR spacers from a collection of over three hundred Streptococcus mutans genomes. Phage DNA is a major target of S. mutans spacers. Strains have also generated immunity against mobile DNA elements such as plasmids and integrative and conjugative elements. There may also be considerable immunity generated against bacterial DNA, although the relative contribution of self-targeting versus bona fide intra- or inter-species targeting needs to be investigated further. While there was clear evidence that these systems have acquired immunity against foreign DNA, there appeared to be minimal impact on horizontal gene transfer (HGT) constraints on a species-level. There was little or no impact on genome size, GC content and openness of the pangenome when comparing between S. mutans strains with low or high CRISPR spacer loads. In summary, while there is evidence of CRISPR spacer acquisition against self and foreign DNA, CRISPR-Cas does not act as a barrier on the expansion of the S. mutans accessory genome. Impact StatementCRISPR-Cas is a widespread bacterial immune system that has been repurposed as a molecular biology tool. This study investigates the role of these systems in the biology and evolution of the dental caries pathogen Streptococcus mutans. CRISPR spacers, that encode immunity against foreign DNA, were extracted from over three hundred S. mutans isolates. Sequence analysis showed that the CRISPR spacers match against phage, mobile element, and bacterial DNA. This shows that S. mutans is actively acquiring immunity against horizontally acquired DNA. However, additional analysis revealed little to no impact of CRISPR-Cas systems on diversification of the S. mutans genome. This suggests that while these systems are actively acquiring CRISPR spacers to defend against foreign DNA, the overall impact on the S. mutans genome might be small. Data SummarySupporting data provided on the Github platform: https://github.com/theshieldslab/Streptococcus-mutans-CRISPR-Spacers-Analysis The authors confirm all supporting data, code and protocols have been provided within the article or through supplementary data files.