Classical antibiotics are losing their efficacy as their application directly selects for resistance. Inducing indirect iron starvation appears to be an alternative and safer strategy as it underlies an extracellular working mechanism. Extracellular mode of action precludesthe gain of resistance via mechanisms such as the prevention of drug entry or the upregulation of efflux pumps, which are very common resistance mechanisms against antibiotics. In this study, we developed a novel approach to induce iron starvation by using siderophores produced by Pseudomonas spp isolated from soil and pond samples. Pyoverdine (PVD) is the main siderophores produced by Pseudomonas. We subjected three human opportunistic pathogens, namely Acinetobacter baumannii, Klebsiella pneumoniae, and Pseudomonas aeruginosa, to three PVDs and investigated their impact on pathogenic growth. The PVDs bind ferric iron and make them inaccessible to the targeted pathogens. We found that PVD treatments are species specific. A. baumannii was inhibited by all three PVDs whereas K. pneumonia was only impacted by one PVD. P. aeruginosa wasinhibited by all three PVDs, but only at low concentrations, while inhibition turned into growth promotion at higher concentration. To test for the rise of resistance evolution against PVDs, we evolved the pathogens for 21 days subjected to PVD treatments in an experimental evolution. The antibiotics ciprofloxacin and meropenem were included as referencesfor fast resistance evolution. We assessed resistance evolution by direct growth comparison of evolved populations and their ancestors. The populations evolved in antibiotic treatments showed an enormous growth increase, whereas the difference between populations evolved in PVD treatments stayed marginal. This result suggests that PVD treatments could be evolutionary more robust than classical antibiotic treatments. Altogether, our work took the first step to use PVDs as an alternative treatment against multi-drug resistant bacteria
