Fig. 1: HEPT construction and assessment of payload activities.

a Heterologous effector phage therapeutics (HEPTs) enable pathogen-specific gene delivery and production of antimicrobial effectors. Upon phage-induced host cell lysis, effectors are released alongside progeny virions to exert a secondary antimicrobial activity against defined bacterial targets. HEPTs were designed against the uropathogens E. coli (light green), K. pneumoniae (light purple), and E. faecalis (light blue). b Five phages were employed as HEPT scaffolds to integrate colicins (M, E6, and E7; square star), klebicin M (star), or the E. faecalis-specific cell wall hydrolase EC300 (Pac-Man). The colour of the effector symbol matches the target organism. c Genes encoding for colicin-like bacteriocins or EC300 were codon-optimized for the producing host-organism and integrated within the structural gene cluster of the corresponding phage scaffold alongside a strong ribosomal binding site (RBS) to mediate phage late promoter-driven effector expression. d–f Cross-genus antimicrobial activity of crude WT phage or HEPT lysates (~10⁹−10¹⁰ PFU/mL) was tested against clinical uropathogen isolates using the spot-on-the-lawn method (full lists provided in Supplementary Table 2 and Supplementary Data 1). Clear zone formation at the site of HEPT lysate spot indicates bacterial susceptibility to the corresponding phage-encoded effector. WT phage lysates lacking effectors served as negative controls. cps major capsid protein, gp gene product, hoc highly immunogenic outer capsid protein, 168 phage K1 gene product 168, kb kilobase. Source data are provided as a Source data file.