Assessing Antibiotic Permeability of Gram-Negative Bacteria via Nanofluidics

Abstract

While antibiotic resistance is increasing rapidly, drug discovery has proven to be extremely difficult. Antibiotic resistance transforms some bacterial infections into deadly medical conditions. A significant challenge in antibiotic discovery is designing potent molecules that enter Gram-negative bacteria and also avoid active efflux mechanisms. Critical analysis in rational drug design has been hindered by the lack of effective analytical tools to analyze the bacterial membrane permeability of small molecules. We design, fabricate, and characterize the nanofluidic device that actively loads more than 200 single bacterial cells in a nanochannel array. We demonstrate a gigaohm seal between the nanochannel walls and the loaded bacteria, restricting small molecule transport to only occur through the bacterial cell envelope. Quantitation of clindamycin translocation through wild-type and efflux-deficient (ΔtolC) Escherichia coli strains via nanofluidic-interfaced liquid chromatography mass spectrometry shows higher levels of translocation for wild-type E. coli than for an efflux-deficient strain. We believe that the assessment of compound permeability in Gram-negative bacteria via the nanofluidic analysis platform will be an impactful tool for compound permeation and efflux studies in bacteria to assist rational antibiotic design.