Engineered superinfective Pf phage prevents dissemination of Pseudomonas aeruginosa in a mouse burn model

Abstract

Pf is a filamentous bacteriophage integrated in the chromosome of most clinical isolates of Pseudomonas aeruginosa. Under stress conditions, mutations occurring in the Pf genome result in the emergence of super-infective variants of Pf (SI-Pf) that are capable of circumventing phage immunity; therefore SI-Pf can even infect Pf-lysogenized P. aeruginosa. Herein, we identified specific mutations located between the repressor and the excisionase genes that result in the emergence of SI-Pf. Based on these findings, we genetically engineered a SI-Pf (eSI-Pf) and tested it as a phage therapy tool for the treatment of life-threatening P. aeruginosa infection of burns caused by strain PAO1. eSI-Pf was able to infect PAO1 biofilms formed in vitro on polystyrene and inhibited their formation when at high concentration. eSI-Pf also infected PAO1 present in burned skin wounds on mice but was not capable of maintaining a sustained reduction in bacterial burden beyond 24 hours. Importantly, and despite not lowering CFU/g of burn skin tissue, eSI-Pf treatment completely abolished the capability of P. aeruginosa to disseminate from the burn site to internal organs. Over the course of 10 days, this resulted in bacterial clearance and survival of all treated mice. We determined that eSI-Pf induced a small colony variant of P. aeruginosa that was unable to disseminate systemically in our burned mouse model during acute infection. Our results suggest that eSI-Pf has potential as a phage therapy against highly recalcitrant antimicrobial resistant P. aeruginosa infections of burn wounds. IMPORTANCEPseudomonas aeruginosa is a major cause of burn related infections. It is also the most likely bacterial infection to advance to sepsis and result in burn-linked death. Frequently, P. aeruginosa strains isolated from burn patients display a multidrug resistant phenotype necessitating the development of new therapeutic strategies and prophylactic treatments. In this context, phage therapy using lytic phages has demonstrated exciting potential in the control P. aeruginosa infection. However, lytic phages have a set of drawbacks during phage therapy including the induction of bacterial resistance and limited bacteria-phage interactions in vivo. Here we propose an alternative approach to interfere with P. aeruginosa pathogenesis in a burn infection model, i.e., using an engineered super-infective filamentous phage. Our study demonstrates that treatment with the engineered Pf phage can prevent sepsis and death in a burn mouse model.