Phage-Mediated Amikacin Dose Sparing Against Pseudomonas aeruginosa and Klebsiella aerogenes: In Vitro Synergy and In Vivo Validation

Background/Objectives: Phage–antibiotic synergy (PAS) offers a potential strategy to enhance antibacterial activity. This study evaluated phage–amikacin interactions against Pseudomonas aeruginosa and Klebsiella aerogenes using in vitro assays and a Caenorhabditis elegans infection model. Methods: Two lytic bacteriophages ɸPh_Pae_AS2 and ɸPh_Eae_AS3 were characterized for morphology, host range, lytic activity, and environmental stability. Phage–amikacin interactions were assessed using checkerboard and time-kill assays, bacterial viable counts, phage-titre measurements, and biofilm assays. In vivo efficacy was evaluated using C. elegans. Results: Both phages exhibited lytic activity and environmental stability. Checkerboard assays demonstrated synergistic interaction yielding fractional inhibitory concentration indices (FICIs) of 0.072 and 0.041 against P. aeruginosa and K. aerogenes, respectively. Under the tested conditions, phage addition reduced the amikacin concentration required to achieve ≥90% growth inhibition by approximately 16-fold and 32-fold, respectively. Antibiofilm efficacy was species- and maturity-dependent. Combination treatment enhanced biofilm inhibition in both species and disruption of mature K. aerogenes biofilms but provided no clear additional benefit over monotherapy against mature P. aeruginosa biofilms. In the C. elegans model, combination treatment improved survival, although the magnitude of improvement over monotherapy varied between the two systems. Conclusions: Phage–amikacin combinations demonstrated substantial in vitro synergy against P. aeruginosa and K. aerogenes, markedly reducing the amikacin concentrations required for growth inhibition under the tested conditions. These findings highlight the potential of phage–antibiotic combinations as an antibiotic-sparing strategy, although the in vitro reductions do not establish clinical dose-sparing benefits. Further validation using diverse clinical isolates and mammalian infection models is warranted to determine their translational potential.