Background: The use of simple and effective model organisms like Caenorhabditis elegans and Galleria mellonella in studying host-pathogen interactions and drug discovery is increasing due to their shared immunogenic pattern with humans. The homology of virulence effects between pathogens on G. mellonella, C. elegans, and humans makes them ideal for biological research.

Objective: This study aims to investigate the impact of phages and phage cocktails on clinical pathogens in C. elegans and G. mellonella. We developed a liquid-based assay, in which the efficiency of phage treatment was evaluated using a scoring system based on microscopy and counting of the nematodes, allowing increasing statistical significance compared to other assays such as larvae or mice.

Methods and result: The nematodes were infected with clinical pathogens in liquid-based assays in 96-well titer plates, followed by phage treatment. The results showed that the survival rates of both organisms significantly increased following phage treatment, with phage cocktails producing the best results. In G. mellonella, E. coli and E. cloacae infected larvae required three phage doses at six-hour intervals to achieve a 100% survival rate, while a single phage dose treatment was effective for K. pneumoniae. For mixed bacterial infections, at least four doses of phage cocktail were necessary to recover the larvae. In C. elegans, phage treatment boosted the survival rates from <20% to >90%, including for poly-microbial infections.

Conclusion: Overall, the phages used in this study were highly efficient, and their efficacy was further improved by phage cocktails. Therefore, G. mellonella and C. elegans are excellent model organisms for studying host-pathogen interactions and drug discovery. Our work demonstrates the potential to use Caenorhabditis elegans to test the virulence of strains of Klebsiella pneumoniae, Enterobacter cloacae, and EHEC/EPEC as well as the efficacy of bacteriophages to treat or prevent infections, allowing a more reliable evaluation for the clinical therapeutic potential of lytic phages.