Antibodies targeting phage fiber and nozzle proteins impair Acinetobacter baumannii phage therapy by blocking infection and promoting immune clearance.

Phage therapy represents a promising strategy to tackle the growing threat of antimicrobial resistance. Increasing evidence has demonstrated that phage-specific antibodies may compromise the efficacy of phage therapy. Nevertheless, little is known about how phage protein specificity of antibodies modulates phage therapeutic outcomes. Herein, we utilized AbP20, a podovirus against Acinetobacter baumannii, to explore the effects of antibodies elicited by each structural protein on phage therapy efficacy in a mouse infection model. Intraperitoneal administration of AbP20 once a day for 7 consecutive days induced robust phage-specific antibody responses that impaired phage therapy. Genome-guided antigen screening identified that antibodies elicited by the nozzle and fiber proteins of AbP20, rather than those targeting the portal, capsid, or adaptor proteins, are the dominant drivers of phage therapy failure. Specifically, fiber-specific and nozzle-specific antibodies block bacterial adsorption and genomic injection of AbP20, respectively. Meanwhile, both antibody subsets efficiently induce large phage aggregates and potentiate macrophage phagocytosis via an Fc receptor-independent pathway. An evolved AbP20 variant with improved neutralization escape capacity exhibited comparable antibody-accelerated phagocytosis and host immune clearance, yet partially rescued therapeutic failure caused by neutralizing antibodies. Collectively, this study elucidates that nozzle- and fiber-elicited neutralizing antibodies impair phage therapy via dual synergistic mechanisms, which offers novel insights into the multifaceted modulation of phage-specific antibodies against phage therapeutic efficacy.