Outer Membrane Disruption Overcomes Intrinsic, Acquired, and Spontaneous Antibiotic Resistance

Disruption of the outer membrane (OM) barrier allows for the entry of otherwise inactive antimicrobials into Gram-negative pathogens. Numerous efforts to implement this approach have identified a large number of OM perturbants that sensitize Gram-negative bacteria to many clinically available Gram-positive active antibiotics. However, there is a dearth of investigation into the strengths and limitations of this therapeutic strategy, with an overwhelming focus on characterization of individual potentiator molecules. Herein, we look to explore the utility of exploiting OM perturbation to sensitize Gram-negative pathogens to otherwise inactive antimicrobials. We identify the ability of OM disruption to change the rules of Gram-negative entry, overcome preexisting and spontaneous resistance, and impact biofilm formation. Disruption of the OM expands the threshold of hydrophobicity compatible with Gram-negative activity to include hydrophobic molecules. We demonstrate that while resistance to Gram-positive active antibiotics is surprisingly common in Gram-negative pathogens, OM perturbation overcomes many antibiotic inactivation determinants. Further, we find that OM perturbation reduces the rate of spontaneous resistance to rifampicin and impairs biofilm formation. Together, these data suggest that OM disruption overcomes many of the traditional hurdles encountered during antibiotic treatment and is a high priority approach for further development. IMPORTANCE The spread of antibiotic resistance is an urgent threat to global health that necessitates new therapeutics. Treatments for Gram-negative pathogens are particularly challenging to identify due to the robust outer membrane permeability barrier in these organisms. Recent discovery efforts have attempted to overcome this hurdle by disrupting the outer membrane using chemical perturbants and have yielded several new peptides and small molecules that allow the entry of otherwise inactive antimicrobials. However, a comprehensive investigation into the strengths and limitations of outer membrane perturbants as antibiotic partners is currently lacking. Herein, we interrogate the interaction between outer membrane perturbation and several common impediments to effective antibiotic use. Interestingly, we discover that outer membrane disruption is able to overcome intrinsic, spontaneous, and acquired antibiotic resistance in Gram-negative bacteria, meriting increased attention toward this approach.