Engineered Proteins Mimic ssRNA Phage to Disrupt Type IV Pili

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 by identifying the specific phage components necessary for T4P detachment, a key step in viral genome entry. Our data reveals 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 widespread environmental bacteria and members of the ESKAPE group of pathogens, often associated with hospital-acquired infections amongst immunocompromised patients. A major reason for the increase in antibiotic resistance in Acinetobacter species is its use of T4P to evade antibiotic treatment. With antibiotic resistance becoming a global health crisis, targeting these virulence factors represents a promising approach to combat infections. Since T4P are critical for Acinetobacter virulence, understanding mechanisms that can disrupt them is highly important. ssRNA phages provide a natural model for this, as they can target T4P to initiate infection. Recent research has shown that ssRNA phages can detach pili during their infection process. However, the precise mechanism behind this phenomenon remains unknown. Our research addresses part of this knowledge gap by identifying the key components required for this pilus detachment. This work will enhance the current understanding of the virulent T4P in Acinetobacter species, which could ultimately lead to broader medical advancements.