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antibiotics, aminoglycosides, silver, reactive oxygen species, redox stress, membranes, iron


The rapid dissemination of antibiotic resistance combined with the decline in the discovery of novel antibiotics represents a major challenge for infectious disease control that can only be mitigated by investments in novel treatment strategies. Alternative antimicrobials, including silver, have regained interest due to their diverse mechanisms of inhibiting microbial growth. One such example is AGXX, a broad-spec­trum antimicrobial that produces highly cytotoxic reactive oxygen species (ROS) to inflict extensive macromolecular damage. Due to the connections identified between ROS production and antibiotic lethality, we hypothesized that AGXX could potentially increase the activity of conventional antibiotics. Using the gram-negative pathogen Pseudomonas aeruginosa, we screened possible synergistic effects of AGXX on several antibiotic classes. We found that the combination of AGXX and aminoglycosides tested at sublethal concentrations led to a rapid exponential decrease in bacterial survival and restored the sensitivity of a kanamycin-resistant strain. ROS production contributes significantly to the bactericidal effects of AGXX/aminoglycoside treatments, which is dependent on oxygen availability and can be reduced by the addition of ROS scaveng­ers. Additionally, P. aeruginosa strains deficient in ROS detoxifying/repair genes were more susceptible to AGXX/aminoglycoside treatment. We further demonstrate that this synergistic interaction was associated with a significant increase in outer and inner membrane permeability, resulting in increased antibiotic influx. Our study also revealed that AGXX/aminoglycoside-mediated killing requires an active proton motive force across the bacterial membrane. Overall, our findings provide an understanding of cellular targets that could be inhibited to increase the activity of conventional antimicrobials

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This work was supported by the NIAID grants R15AI164585 and R03AI174033-01A1 as well as the Illinois State University Pre-Tenure Faculty Initiative Grant (to J.-U.D.). G.Y.D. was supported by Weigel and Mockford-Thompson fellowships by the Phi-Sigma Biological Sciences Honors Society and the SIGMA Xi and BIRDFeeder grants. G.M.A. and C.D.O. were supported by the Illinois State University Undergraduate Research Support Program.


First published in mSphere, 2023,

This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license.



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msphere.00190-23-s0002.xlsx (10 kB)
Table 1

msphere.00190-23-s0003.xlsx (10 kB)
Table 2

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