Mining the phage playbook to create a potent, generic phage therapy

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PROJECT SUMMARY Phage therapy, the practice of treating bacterial infections with bacteria-targeting viruses, bacteriophage (or phage), is a promising and urgently needed alternative to antibiotics. A major challenge holding this approach back from widespread adoption is that phage treatments need to be customized for the infecting strains in each patient, a slow and labor-intensive process. This requirement arises from the exquisitely narrow host-range that many phages display, even among closely related bacterial strains. A major factor driving phage host range is the immense collection of bacterial anti-phage immune mechanisms that are unevenly distributed across bacterial strains. However, the underlying molecular arms race between bacteria and phage has given rise to an equally impressive set of corresponding phage counter-defense pathways, and thus collectively phage have already evolved mechanisms by which to overcome most bacterial defenses. Similar to their bacterial counterparts, each phage strain encodes only a miniscule fraction of existing counter-defenses, thus explaining the narrow host-range of individual phages. Developing a phage treatment that could amass these naturally occurring phage solutions into a “super phage cocktail” would enable production of an off-the-shelf phage treatment with a greatly expanded species range and the ability to forestall bacterial resistance. Here, I propose developing a pipeline leveraging existing phage counter-defense mechanisms to create a powerful proof-of-principle phage cocktail for the opportunistic pathogen, Pseudomonas aeruginosa. To realize this vision, I will take an experimental genomic approach to map the immune system of clinically relevant P. aeruginosa isolates and thereby determine which bacterial defenses the phage will encounter during infections. I will then develop a powerful, high throughput screen to identify existing phage counter-defense mechanisms that can overcome these bacterial defenses. Finally, I will create a super phage cocktail encoding an extensive collection of counter-defense gene cassettes with the ability to infect a broad set of P. aeruginosa strains. These studies seek to leverage the existing biology underlying the bacterial-phage molecular arms race to overcome a major hurdle in the development of phage therapy. This work will provide unprecedented insight into the breadth and diversity of both bacterial immunity and phage counter-defenses and uncover a multitude of novel biological mechanisms to be characterized in future studies. The engineered phage cocktail also constitutes an innovative experimental system that can be used to answer fundamental questions about viral population diversity and evolution. This initial study will serve as the blue print for development of phage therapy for other multi-drug resistant opportunistic pathogens.