Resistance evolution against phage combinations depends on the timing and order of exposure

Globally rising rates of antibiotic resistance have renewed interest in phage therapy where combinations of phages have been successfully used to treat multidrug resistant infections. To optimise phage therapy, we first need to understand how bacteria evolve resistance against combinations of multiple phages. Here we use simple laboratory experiments and genome sequencing to show that the timing and order of phage exposure determine the strength, cost and mutational basis of resistance evolution in the opportunistic pathogen Pseudomonas aeruginosa. These findings suggest that phage combinations can be optimised to limit the emergence and persistence of resistance, thereby promoting the long-term usefulness of phage therapy.