Prophage mediated anti-phage defense in Escherichia coli
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brenes-lrbrenes-phd-biology-2026-thesis .pdf
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Author(s)
Brenes, Lucas Rene
Advisor(s)
Laub, Michael T.
Date Issued
February 2026
Publisher
Massachusetts Institute of Technology
Abstract
All cellular life must contend with the threat of viral predators, and bacteria have been locked in an ancient conflict with theirs, known as bacteriophages (phages). Consequently, bacteria have evolved an incredible diversity of immune systems to combat phages, while phages in turn continually evolve counter-defense strategies. In recent years, the number of identified anti-phage defense systems has risen rapidly; however, we are still only beginning to tap into the immense diversity of bacterial immunity. Temperate phages blur the lines of this host-pathogen dynamic because they can undergo an alternate mode of transmission where, instead of producing more virions and lysing the cell, they integrate into the host and exist passively as a prophage. As a prophage, the phage is more closely tied to the success of the host, and as a result, they often encode anti-phage defense systems to protect against subsequent infections. While it has long been appreciated that temperate phages are reservoirs of defense systems, the types of systems they carry and their potential interactions against other temperate phages remains an understudied question. In this work, I isolated 33 temperate phages, named PhiR phages, from a diverse set of 52 E. coli strains by using DNA damage as an induction signal. Lambdoid phages were the most abundant family, and I supplemented the collection with an additional 9 lambdoid phages for a total of 42 temperate phages present as single lysogens across three recipient E. coli strains. For each strain background, every lysogen was tested for defense against every phage present in that background. More than three quarters of the prophages provided defense against at least one other phage, and many defended against multiple phages. Strain background and growth conditions had strong effects on defense outcomes, indicating some systems are context-dependent. Using an iterative deletion-based approach, I identified 4 known and 13 novel anti-phage defense systems. These systems had a range of predicted mechanisms while some only contained domains of unknown function. The prevalence of homologs across the bacterial kingdom varied for each system; however, all were highly enriched in prophage regions. Toxin-antitoxin modules were a prominent fraction of the systems, with 5 different toxin families being ascribed anti-phage defense functions for the first time. Overall, this work contributes to the notion that temperate phages are rich sources of anti-phage defense systems, many of which are yet to be discovered. Additionally, temperate phage – temperate phage interactions are widespread and add a layer of complexity to the ongoing conflict between bacteria and their viral predators.
MIT Department
Massachusetts Institute of Technology. Department of Biology
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