Rapid loss of CRISPR-mediated herd immunity from Pseudomonas aeruginosa populations

CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeats; CRISPR-associated) are prokaryotic adaptive immune systems that insert parasite-derived DNA sequences (spacers) into genomic CRISPR loci to provide sequence-specific immunity [1]. One feature that sets CRISPR-Cas apart from other adaptive immune systems is that immunity is heritable, resulting in trans-generational protection against fixed parasite genotypes [2, 3]. However, both theory and data show that levels of CRISPR-Cas immunity also depend on the diversity of spacers present within a bacterial population, with higher levels of herd immunity when the number of different spacers increases, since this constrains parasites ability to evolve to overcome CRISPR immunity [4-7]. Given its importance for CRISPR herd immunity levels, this raises the question if and how population-level spacer diversity is maintained over time. To examine this, we performed experimental evolution using the opportunistic human pathogen Pseudomonas aeruginosa strain PA14, and its bacteriophage (phage) DMS3vir. Consistent with earlier work [4], we found that upon infection bacterial populations initially generate high population-level spacer diversity, which causes rapid phage extinction. However, levels of spacer diversity rapidly decline after phage extinction as both sensitive bacteria and partially resistant receptor mutants invade the CRISPR population. The key consequence of this process is that immunized bacterial populations that were initially highly effective in driving phage extinct, rapidly lose this ability against the same phage when it is re-introduced into the population. These data can help to explain how bacteria with CRISPR-Cas immune systems and phage can coexist in natural environments.