The arms race between bacteria and phages led to the development of sophisticated antiphage defense systems, including CRISPR-Cas and restriction-modification systems. Evidence suggests that known and unknown defense systems are located in "defense islands" in microbial genomes. Here, we comprehensively characterized the bacterial defensive arsenal by examining gene families that are clustered next to known defense genes in prokaryotic genomes. Candidate defense systems were systematically engineered and validated in model bacteria for their antiphage activities. We report nine previously unknown antiphage systems and one antiplasmid system that are widespread in microbes and strongly protect against foreign invaders. These include systems that adopted components of the bacterial flagella and condensin complexes. Our data also suggest a common, ancient ancestry of innate immunity components shared between animals, plants, and bacteria.

Temperate viruses can become dormant in their host cells, a process called lysogeny. In every infection, such viruses need to decide between the lytic and the lysogenic cycles, i.e., whether to replicate and lyse their host or to lysogenize and keep the host viable. Here we show that viruses (phages) of the spBeta group use a small-molecule communication system to coordinate lysis-lysogeny decisions. During infection of its Bacillus host cell, the phage produces a 6aa communication peptide that is released to the medium. In subsequent infections, progeny phages measure the concentration of this peptide and lysogenize if the concentration is sufficiently high. We found that different phages encode different versions of the communication peptide, demonstrating a phage-specific peptide communication code for lysogeny decisions. We termed this communication system the “arbitrium” system, and further show that it is encoded by 3 phage genes: aimP, producing the peptide, aimR, the intracellular peptide receptor, and aimX, a negative regulator of lysogeny. The arbitrium system enables an offspring phage to communicate with its predecessors, i.e., to estimate the amount of recent prior infections and hence decide whether to employ the lytic or lysogenic cycle.

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In the process of CRISPR adaptation, short pieces of DNA ("spacers") are acquired from foreign elements and integrated into the CRISPR array. It so far remained a mystery how spacers are preferentially acquired from the foreign DNA while the self chromosome is avoided. Here we show that spacer acquisition is replication-dependent, and that DNA breaks formed at stalled replication forks promote spacer acquisition. Chromosomal hotspots of spacer acquisition were confined by Chi sites, which are sequence octamers highly enriched on the bacterial chromosome, suggesting that these sites limit spacer acquisition from self DNA. We further show that the avoidance of "self" is mediated by the RecBCD dsDNA break repair complex. Our results suggest that in E. coli, acquisition of new spacers depends on RecBCD-mediated processing of dsDNA breaks occurring primarily at replication forks, and that the preference for foreign DNA is achieved through the higher density of Chi sites on the self chromosome, in combination with the higher number of forks on the foreign DNA. This model explains the strong preference to acquire spacers from both high copy plasmids and phages.