Abstract
Antibiotic susceptibility of bacterial pathogens is typically evaluated using in vitro assays that do not consider the complex host microenvironment. This may help explaining a significant discrepancy between antibiotic efficacy in vitro and in vivo, with some antibiotics being effective in vitro but not in vivo or vice versa. Nevertheless, it is well-known that antibiotic susceptibility of bacteria is driven by environmental factors. Lung epithelial cells enhance the activity of aminoglycoside antibiotics against the opportunistic pathogen Pseudomonas aeruginosa, yet the mechanism behind is unknown. The present study addresses this gap and provides mechanistic understanding on how lung epithelial cells stimulate aminoglycoside activity. To investigate the influence of the local host microenvironment on antibiotic activity, an in vivo-like three-dimensional (3-D) lung epithelial cell model was used. We report that conditioned medium of 3-D lung cells, containing secreted but not cellular components, potentiated the bactericidal activity of aminoglycosides against P. aeruginosa, including resistant clinical isolates, and several other pathogens. In contrast, conditioned medium obtained from the same cell type, but grown as conventional (2-D) monolayers did not influence antibiotic efficacy. We found that 3-D lung cells secreted endogenous metabolites (including succinate and glutamate) that enhanced aminoglycoside activity, and provide evidence that bacterial pyruvate metabolism is linked to the observed potentiation of antimicrobial activity. Biochemical and phenotypic assays indicated that 3-D cell conditioned medium stimulated the proton motive force (PMF), resulting in increased bacterial intracellular pH. The latter stimulated antibiotic uptake, as determined using fluorescently labelled tobramycin in combination with flow cytometry analysis. Our findings reveal a cross-talk between host and bacterial metabolic pathways, that influence downstream activity of antibiotics. Understanding the underlying basis of the discrepancy between the activity of antibiotics in vitro and in vivo may lead to improved diagnostic approaches and pave the way towards novel means to stimulate antibiotic activity.
Highlights
While many biochemical substances that modulate antibiotic activity are known [1,2,3,4,5,6,7,8,9,10], the influence of the local environment at the host-pathogen interface on bacterial responses to antibiotics is still poorly understood [4]
We found that host cell secretions increased antibiotic uptake through stimulation of bacterial metabolism, which in turn resulted in enhanced activity
Our findings highlight that cross-talk between host and bacterial metabolisms contributes to the efficacy of antibiotic treatment
Summary
While many biochemical substances that modulate antibiotic activity are known [1,2,3,4,5,6,7,8,9,10], the influence of the local environment at the host-pathogen interface on bacterial responses to antibiotics is still poorly understood [4]. Whether and how the complex host metabolic environment influences antibiotic activity is not fully understood These insights are important to evaluate the potential role of the host in the clearance of bacterial pathogens during antibiotic treatment of an infection, and may help opening novel avenues to improve the correlation between antibiotic susceptibility profiles in vitro and in vivo. This is relevant for infectious diseases for which antibiotic therapy chosen based on susceptibility assays frequently does not lead to clinical improvement [11,12,13,14], including in respiratory tract infections in people with cystic fibrosis [11]. These findings highlight a significant contribution of the host metabolism to the activity of antibiotic treatment
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