Polyphosphate and pyoverdine synthesis modulates Pseudomonas aeruginosa PAO1 virulence in a zebrafish model

Abstract Understanding how bacterial metabolism shapes virulence requires infection models that are both biologically informative and experimentally accessible. Here, we used zebrafish (Danio rerio) larvae infection by static immersion as a simple and robust vertebrate model to study Pseudomonas aeruginosa host-pathogen interactions and virulence regulation. Using this approach, we show that disruption of polyphosphate (polyP) synthesis by loss of ppk1 gene (τ1ppk1) leads to marked attenuation of virulence, whereas interrupting ppk2 gene (τ1ppk2) results in a hypervirulent phenotype. Phenotypic assays and quantitative proteomics revealed that loss of polyP selectively impairs pyoverdine production, with minimal effects on other canonical virulence factors. Consistently, a pyoverdine-deficient mutant (τ1pvdF) exhibited reduced virulence in zebrafish, validating the capacity of the zebrafish model to functionally resolve key virulence factors. Together, our results highlight polyphosphate metabolism as a central regulator of P. aeruginosa virulence and position zebrafish immersion assays as an efficient and ethically aligned alternative to mammalian models for studying bacterial pathogenesis. Competing Interest Statement The authors have declared no competing interest.