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

Abstract

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.

Novel Small Molecule Growth Inhibitor Affecting Bacterial Outer Membrane Reduces Extraintestinal Pathogenic Escherichia coli (ExPEC) Infection in Avian Model

“…The lack of APEC resistance to GI-7 might be due to the membrane-targeting mechanism of GI-7 ( 18 ), although a more thorough search for and characterization of GI-7-resistant mutants is necessary to facilitate industrial application of GI-7 as an anti-APEC therapeutic in the future. Membrane-targeting antibacterial drugs are less likely to acquire resistance and are also effective against antibiotic-resistant strains ( 20 , 40 ). The APEC strain used in our chicken studies is resistant to ampicillin and tetracycline, two of the antibiotics most commonly used in food animals ( 18 ).…”

“…Previously, using microscopy and in vitro studies (crystal violet uptake and loss of 260-/280-nm-absorbing materials) we showed that these GIs induced membrane defects like membrane blebbing, disruption of membrane, and increased membrane permeability ( 18 ), suggesting an effect on the APEC membrane. Antibacterial compounds that target the outer membrane (OM) of Gram-negative bacteria can counter the resistance problem ( 19 , 20 ). The OM acts as a shield for bacteria by providing a permeability barrier, thus precluding the entry of antibiotics in bacteria ( 19 , 21 ).…”

“…Notably, resistance is less likely to occur against OM-targeting antibacterial compounds, as they avoid the most common mechanisms for resistance development in bacteria, which are the entry barriers that prevent access by antibiotics and the efflux pumps that eliminate antibiotics from bacterial cells ( 19 , 21 ). Moreover, OM-acting antibacterial compounds can make resistant bacteria susceptible to antibiotics when used in combination with antibiotics ( 19 , 20 , 30 ). Therefore, antibacterial compounds targeting the OM are promising leads for the development of novel therapeutics to circumvent the resistance problem in ExPECs and other Gram-negative bacteria.…”

The Building Blocks of Antimicrobial Resistance in Pseudomonas aeruginosa: Implications for Current Resistance-Breaking Therapies

“…SPR206 is active against P. aeruginosa with a similar potency to polymyxin B and is currently undergoing phase-one clinical trial (Zhang et al, 2020). OM sensitizers appear to be a promising approach to resistance that can by-pass intrinsic, acquired and spontaneous resistance (Macnair and Brown, 2020). However, further investigation FIGURE 3 | Multi-layered, interacting resistance mechanisms in P. aeruginosa.…”

Rationally designed foldameric adjuvants enhance antibiotic efficacy via promoting membrane hyperpolarization

“…Strikingly, dysregulated membrane potential and perturbation of pH homeostasis also appear to play a critical role in antibiotic induced lethality. [10][11][12] These findings suggest that antibiotic action could be improved by simultaneously targeting bacterial membrane potential during antibiotic treatment. The impact of membrane depolarization on antibiotic efficacy is relatively well-described.…”