Targeting bacterial phosphotransferase system to prevent the dissemination of antibiotic resistance genes

Antimicrobial resistance (AMR) poses a significant challenge to public health and human security, with plasmid-mediated horizontal gene transfer (HGT) being a primary driver for its dissemination. Here, we identify indole analogs containing electron-withdrawing groups as a new category of HGT inhibitors. Using indole-3-acetic acid (IAA) as a representative scaffold, we demonstrate that IAA targets glycolytic enolase to deplete phosphoenolpyruvate (PEP) in donor bacteria; this suppresses phosphotransferase system (PTS) activity, blocks PtsI phosphorylation, reduces cAMP synthesis and prevents catabolite repressor protein (CRP) activation, ultimately limiting intracellular ATP availability. Concurrently, IAA attenuates reactive oxygen species (ROS) generation by inhibiting FADH₂ oxidation and riboflavin biosynthesis. The concerted reduction in ATP and ROS arrests conjugative plasmid transfer. Overall, our work suggests the potential of indole analogs as a new class of conjugative transfer inhibitors and highlights that bacterial phosphotransferase system represents a promising target to prevent the propagation of AMR.