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It is becoming increasingly clear that antibiotics can both positively and negatively impact the infectivity of bacteriophages (phage), but the underlying mechanisms often remain unclear. Here we demonstrate that antibiotics that target the protein translation machinery can fundamentally alter the outcome of bacteria-phage interactions by interfering with the production of phage-encoded counter-defense proteins. Specifically, using Pseudomonas aeruginosa PA14 and phage DMS3vir as a model, we show that bacteria with Clustered Regularly Interspaced Short Palindromic Repeat, CRISPR associated (CRISPR-Cas) immune systems have elevated levels of immunity against phage that encode anti-CRISPR (acr) genes when translation inhibitors are present in the environment. CRISPR-Cas are highly prevalent defense systems that enable bacteria to detect and destroy phage genomes in a sequence-specific manner. In response, many phages encode acr genes that are expressed immediately following the infection to inhibit key steps of the CRISPR-Cas immune response. Our data show that while phage-carrying acr genes can amplify efficiently on bacteria with CRISPR-Cas immune systems in the absence of antibiotics, the presence of antibiotics that act on protein translation prevents phage amplification, while protecting bacteria from lysis.

Articles on CRISPR commonly open with some variant of the phrase "these short palindromic repeats and their associated endonucleases (Cas) are an adaptive immune system that exists to protect bacteria and archaea from viruses and infections with other mobile genetic elements." There is an abundance of genomic data consistent with the hypothesis that CRISPR plays this role in natural populations of bacteria and archaea, and experimental demonstrations with a few species of bacteria and their phage and plasmids show that CRISPR-Cas systems can play this role in vitro. Not at all clear are the ubiquity, magnitude, and nature of the contribution of CRISPR-Cas systems to the ecology and evolution of natural populations of microbes and the strength of selection mediated by different types of phage and plasmids to the evolution and maintenance of CRISPR-Cas systems. In this perspective, with the aid of heuristic mathematical-computer simulation models, we explore the a priori conditions under which exposure to lytic and temperate phage and conjugative plasmids will select for and maintain CRISPR-Cas systems in populations of bacteria and archaea. We review the existing literature addressing these ecological and evolutionary questions and highlight the experimental and other evidence needed to fully understand the conditions responsible for the evolution and maintenance of CRISPR-Cas systems and the contribution of these systems to the ecology and evolution of bacteria, archaea, and the mobile genetic elements that infect them., Competing Interests: The authors declare no competing interest., (Copyright © 2020 the Author(s). Published by PNAS.)

Prokaryotic clustered regularly interspaced short palindromic repeat (CRISPR)/Cas (CRISPR-associated sequences) systems provide adaptive immunity against viruses when a spacer sequence of small CRISPR RNA (crRNA) matches a protospacer sequence in the viral genome. Viruses that escape CRISPR/Cas resistance carry point mutations in protospacers, though not all protospacer mutations lead to escape. Here, we show that in the case of Escherichia coli subtype CRISPR/Cas system, the requirements for crRNA matching are strict only for a seven-nucleotide seed region of a protospacer immediately following the essential protospacer-adjacent motif. Mutations in the seed region abolish CRISPR/Cas mediated immunity by reducing the binding affinity of the crRNA-guided Cascade complex to protospacer DNA. We propose that the crRNA seed sequence plays a role in the initial scanning of invader DNA for a match, before base pairing of the full-length spacer occurs, which may enhance the protospacer locating efficiency of the E. coli Cascade complex. In agreement with this proposal, single or multiple mutations within the protospacer but outside the seed region do not lead to escape. The relaxed specificity of the CRISPR/Cas system limits escape possibilities and allows a single crRNA to effectively target numerous related viruses.