Rational bacteriophage and antibiotic combinations to combat antimicrobial resistance

Abstract

Antimicrobial resistance (AMR) continues to exert a significant toll on global health and progress. Clinical development of new antimicrobial agents has lagged for the last few decades leading to a resurgence of bacteriophage therapy as an alternative therapeutic option against recalcitrant bacterial infections. Chapter two examines the increasingly important role of bacterial porin channel disruptions in antibiotic resistance against β-lactam drugs. In the first study, ceftaroline-avibactam activity was impacted by the porin channel disruptions in isogenic mutant constructs in the presence of selected β-lactamase enzymes while it inhibited 86% of the clinical Klebsiella pneumoniae isolates. The second study determined the potency of imipenem with relebactam, and comparator β-lactams against highly drug resistant clinical Klebsiella pneumoniae isolates with porin defects collected from Italy, Brazil and Australia. Relebactam significantly enhanced the potency of imipenem with 93% of the tested clinical isolates showing susceptibility to the combination. In chapter three, the unexpected finding of bacteriophages protecting bacteria from antibiotic hydrolysis was investigated. Experiments revealed that phage lysis of host bacteria leads to explosive cell lysis and hypervesiculation resulting in the production of outer membrane vesicles containing potent beta-lactamase enzymes that protects the bacteria from antibiotic killing. These data provide evidence that antagonistic phage-antibiotic interactions may contribute to adverse clinical outcomes in bacteriophage therapy. Chapter four explores the development of a scalable bacteriophage purification pipeline using automated FPLC methods. Membrane columns were superior to bead-resin columns with >90% recovery of bacteriophages. EndoTrap HD FPLC column was the most efficient affinity column for the polishing step. This efficient and scalable workflow can be broadly implemented to purify phages for human therapeutic use.