Porin‐Independent Uptake of Small Molecule Antibiotics Facilitated by <i>Escherichia coli</i> Outer Membrane Vesicles

The development of novel antimicrobial agents that are effective against Gram-negative bacteria is hindered by the dual membrane cell envelope of these bacteria. To reach their intracellular targets, most small-molecule antibiotics must first pass through protein channels called porins; however, a common mechanism of acquired resistance is decreased expression of these outer membrane proteins. Additionally, parameters such as size, shape, and charge regulate passage of antibiotics through porins, further limiting the design space of novel antibiotic molecules. Inspired by the ability of bacterial outer membrane vesicles (OMVs) to deliver cargo to the bacterial cytosol, we hypothesized that encapsulation of small molecule antibiotics within OMVs would improve the activity of the drugs by facilitating uptake. To test this hypothesis, we investigated the ability of imipenem-encapsulated OMVs to inhibit the growth of several Gram-negative bacteria, including multidrug-resistant (MDR) clinical isolates. Our results demonstrated that encapsulation within OMVs significantly lowers the effective concentration of imipenem in several MDR isolates. Using a panel of porin knockout strains, we further demonstrated that this mechanism of antibiotic delivery does not require porin expression. Together, our results demonstrate the potential of OMVs as novel antibiotic delivery vehicles to treat antibiotic-resistant bacterial infections by improving drug uptake.