Phage-mediated virulence loss and antimicrobial susceptibility in carbapenem-resistant <i>Klebsiella pneumoniae</i>

Bacteriophages, known for their ability to kill bacteria, are hampered in their effectiveness because bacteria are able to rapidly develop resistance, thereby posing a significant challenge for the efficacy of phage therapy. The impact of evolutionary trajectories on the long-term success of phage therapy remains largely unclear. Herein, we conducted evolutionary experiments, genomic analysis, and CRISPR-mediated gene editing, to illustrate the evolutionary trajectory occurring between phages and their hosts. Our results illustrate the ongoing "arms race" between a lytic phage and its host, a carbapenem-resistant <i>Klebsiella pneumoniae</i> clinical strain Kp2092, suggesting their respective evolutionary adaptations that shape the efficacy of phage therapy. Specifically, Kp2092 rapidly developed resistance to phages through mutations in a key phage receptor (<i>galU</i>) and bacterial membrane defenses such as LPS synthesis, however, this evolution coincides with unexpected benefits. Evolved bacterial clones not only exhibited increased sensitivity to clinically important antibiotics but also displayed a loss of virulence in an <i>in-vivo</i> model. In contrast, phages evolved under the selection pressure against Kp2092 mutants and exhibited enhanced bacterial killing potency, targeting mutations in phage tail proteins gp12 and gp17. These parallel evolutionary trajectories suggest a common genetic mechanism driving adaptation, ultimately favoring the efficacy of phage therapy. Overall, our findings highlight the potential of phages not only as agents for combating bacterial resistance, but also a driver of evolution outcomes that could lead to more favorable clinical outcomes in the treatment of multidrug resistance pathogens.IMPORTANCECarbapenem-resistant <i>Klebsiella pneumoniae</i> represents one of the leading pathogens for infectious diseases. With traditional antibiotics often being ineffective, phage therapy has emerged as a promising alternative. However, phage predation imposes a strong evolutionary pressure on the rapid evolution of bacteria, challenging treatment efficacy. Our findings illustrate how co-evolution enhances phage lytic capabilities through accumulated mutations in the tail proteins gp12 and gp17, while simultaneously reducing bacterial virulence and antibiotic resistance. These insights advance our understanding of phage-host interactions in clinical settings, potentially inspiring new approaches akin to an "arms race" model to combat multidrug-resistant crises effectively.