LAUSR

Abstract Bacteriophage (phage) therapy in combination with antibiotic treatment serves as a potential strategy to overcome the continued rise in antibiotic resistance across bacterial pathogens. Understanding the impacts of evolutionary and ecological processes to the phage‐antibiotic‐resistance dynamic could advance the development of such combinatorial therapy. We tested whether the acquisition of mutations conferring phage resistance may have antagonistically pleiotropic consequences for antibiotic resistance. First, to determine the robustness of phage resistance across different phage strains, we infected resistant Escherichia coli cultures with phage that were not previously encountered. We found that phage‐resistant E. coli mutants that gained resistance to a single phage strain maintain resistance to other phages with overlapping adsorption methods. Mutations underlying the phage‐resistant phenotype affects lipopolysaccharide (LPS) structure and/or synthesis. Because LPS is implicated in both phage infection and antibiotic response, we then determined whether phage‐resistant trade‐offs exist when challenged with different classes of antibiotics. We found that only 1 out of the 4 phage‐resistant E. coli mutants yielded trade‐offs between phage and antibiotic resistance. Surprisingly, when challenged with novobiocin, we uncovered evidence of synergistic pleiotropy for some mutants allowing for greater antibiotic resistance, even though antibiotic resistance was never selected for. Our results highlight the importance of understanding the role of selective pressures and pleiotropic interactions in the bacterial response to phage‐antibiotic combinatorial therapy.