Phage-derived proteins and conjugates induce pilus detachment

ABSTRACT Antimicrobial resistance is a growing global health crisis, with ESKAPE pathogens, such as Acinetobacter species, contributing significantly to hospital-acquired infections. These bacteria employ various virulence factors, including extracellular type IV pili (T4P), which serve as essential appendages for DNA uptake, biofilm formation, and resistance to antibiotics. Single-stranded RNA (ssRNA) bacteriophages (phages) exploit these retractile pili as entry receptors, but the exact mechanism of viral RNA delivery following pilus binding and retraction remains poorly understood. In this study, we investigated the entry mechanism of the ssRNA phage AP205 on its host strain Acinetobacter higginsii . We identified the specific phage components necessary for T4P detachment, a key step in viral genome entry. Our data reveal that the phage’s maturation protein (Mat) is sufficient to induce pilus detachment on its own, and the overall virion structure further enhances the efficiency of this process. These findings provide mechanistic insights into how ssRNA phages exploit bacterial T4P as part of their infection pathway and suggest a conserved strategy for host entry. IMPORTANCE Acinetobacter species are environmental bacteria and members of the ESKAPE pathogens, associated with hospital-acquired infections among immunocompromised patients. Rising antibiotic resistance among Acinetobacter is partly attributed to its use of type IV pili (T4P) to evade treatment. Targeting these virulence factors is a promising strategy to combat these pathogens. Single-stranded RNA bacteriophages naturally interact with T4P, and recent research suggests they can detach pili during infection, but the mechanism remains unclear. Our study identifies components required for this pilus detachment, addressing this knowledge gap. These findings advance understanding of T4P virulence in Acinetobacter species and may guide future approaches to combat antibiotic-resistant infections.