Elucidating the mechanism by which synthetic helper peptides sensitize Pseudomonas aeruginosa to multiple antibiotics

The emergence and rapid spread of multi-drug resistant (MDR) bacteria pose a serious threat to global healthcare. There is an urgent need for new antibacterial substances or new treatment strategies to deal with the infections by MDR bacterial pathogens, especially the Gram-negative pathogens. In this study, we show that a number of synthetic cationic peptides display strong synergistic antimicrobial effects with multiple antibiotics against the Gram-negative pathogen Pseudomonas aeruginosa. We found that an all-D amino acid containing peptide called D-11 increases membrane permeability by attaching to LPS and membrane phospholipids, thereby facilitating the uptake of antibiotics. Subsequently, the peptide can dissipate the proton motive force (PMF) (reduce ATP production and inhibit the activity of efflux pumps), impairs the respiration chain, promote the production of reactive oxygen species (ROS) in bacterial cells and induce intracellular antibiotics accumulation, ultimately resulting in cell death. By using a P. aeruginosa abscess infection model, we demonstrate enhanced therapeutic efficacies of the combination of D-11 with various antibiotics. In addition, we found that the combination of D-11 and azithromycin enhanced the inhibition of biofilm formation and elimination of established biofilms. Our study provides a realistic treatment option for combining close-to-nature synthetic peptide adjuvants with existing antibiotics to combat infections caused by P. aeruginosa. Author summary Antimicrobial resistance is a global public health problem that is limiting our antibiotic capacity to control bacterial infections. Despite the enormous efforts in drug development, very few clinically relevant drugs have been approved in the last decades. This is especially critical for Gram-negative bacteria, which remain the most problematic group, because, among other reasons, the outer membrane acts as a permeability barrier preventing antibiotics to reach their targets. Here, we describe the action mechanism through which a human close-to-nature synthetic helper peptide is able to sensitize Pseudomonas aeruginosa to a broad range of antibiotics for which it is normally resistant. We demonstrate that this peptide is active at different levels (membrane, proton motive force, respiration, ATP synthesis, ROS accumulation and efflux pump activity), facilitating antibiotic accumulation inside the bacteria and promoting cell death. Moreover, we have determined the efficacy of the synergistic combinations in complex relevant environments, such as Pseudomonas biofilms, human blood and in vivo in a mice infection model. This work demonstrates the effectiveness of synergistic combinations for fighting an important lung pathogen, providing a new therapeutical option for the treatment of P. aeruginosa infection and extending the use of the current antibiotics.

Automated summary

This study demonstrates that synthetic cationic peptides, particularly D-11, can enhance the therapeutic efficacy of multiple antibiotics against Gram-negative pathogen Pseudomonas aeruginosa by increasing membrane permeability, disrupting proton motive force, promoting ROS production, inducing intracellular antibiotic accumulation, among others. The findings provide a realistic treatment option for combating P. aeruginosa infections by combining synthetic peptide adjuvants with existing antibiotics.