The Escherichia coli TolC efflux pump protein is immunogenic and elicits protective antibodies.

Abstract

Antimicrobial resistance is an increasing worldwide public health burden that threatens to make existent antimicrobials obsolete. An important mechanism of antimicrobial resistance is the overexpression of efflux pumps, which reduce the intracellular concentration of antimicrobials. TolC is the outer membrane protein of an efflux pump that has gained attention as a therapeutic target. Little is known about the immune response against TolC. Here, we evaluated the immune response against TolC from Escherichia coli. TolC in silico epitope prediction showed several residues that could bind to human antibodies, and we showed that human plasma presented higher titers of anti-TolC IgG and IgA, than IgM. E. coli recombinant TolC protein stimulated macrophages in vitro to produce nitric oxide, as well as interleukin-6 and tumor necrosis factor α, assessed by Griess assay and enzyme-linked immunosorbent assay, respectively. Immunization of mice with TolC intraperitoneally and an in vitro restimulation led to increased T cell proliferation and interferon γ production, evaluated by flow cytometry and enzyme-linked immunosorbent assay, respectively. TolC mouse immunization stimulated anti-TolC IgM and IgG production, with higher levels of IgG1 and IgG2, among the IgG subclasses. Anti-TolC murine antibodies could bind to live E. coli and increase bacterial uptake and elimination by macrophages in vitro. Intraperitoneal or intranasal, but not oral, immunizations with inactivated E. coli also led to anti-TolC antibody production. Finally, TolC immunization increased mouse survival rates to antimicrobial-sensitive or resistant E. coli infection. Our results showed that TolC is immunogenic, leading to the production of protective antibodies against E. coli, reinforcing its value as a therapeutic target.