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However, a major problem with phage therapy is the overly high specificity of phages for their hosts, which is currently addressed by a personalized approach involving screening a bank of wild-type phages against each clinical isolate. To shorten this process, we propose that a focused library of synthetic phages could be rapidly selected for a member binding to a given clinical isolate. We created libraries of recombinant M13 phages expressing receptor-binding proteins based on the collective metagenome of inovirus phages, a diverse group whose members appear to infect nearly all bacterial phyla. Using two rounds of a pull-down selection, phage variants were identified against several Gram-negative pathogens, including a variant (M13PAB) that bound to several Pseudomonas aeruginosa strains, including clinical isolates. To confer bactericidal activity to the nonlytic phage, a last-line but nephrotoxic lipopeptide, colistin, was cross-linked to the M13PAB virions. The colistin-M13PAB phage conjugate lowered the minimal inhibitory concentration\u00a0of colistin by 1\u20132 orders of magnitude for multiple strains of P. aeruginosa and showed a lack of hemolytic or cytotoxic activity in vitro, suggesting high potency combined with low toxicity. Thus, a metagenome-inspired library displayed on the M13 phage scaffold, when subjected to a short selection for binding to a bacterial clinical isolate, could yield a phage variant that targets the specified strain. This approach may improve the speed, consistency, and cost-effectiveness of personalized phage therapy.<\/jats:p>","DOI":"10.1093\/nar\/gkaf984","type":"journal-article","created":{"date-parts":[[2025,9,16]],"date-time":"2025-09-16T11:41:23Z","timestamp":1758022883000},"source":"Crossref","is-referenced-by-count":2,"title":["Metagenome-inspired libraries to engineer phage M13 for targeted killing of Gram-negative bacterial species"],"prefix":"10.1093","volume":"53","author":[{"given":"Yanxi","family":"Yang","sequence":"first","affiliation":[{"name":"University of California Department of Chemical and Biomolecular Engineering, , Los Angeles , CA 90095 ,","place":["United States"]},{"name":"University of California Department of Chemistry and Biochemistry, , Los Angeles , CA 90095 ,","place":["United States"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dayeon","family":"Kang","sequence":"additional","affiliation":[{"name":"University of California Department of 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