Study design

This study was designed to search for the metabolites that potentiated the currently available antibiotics to kill multidrug-resistant bacteria. We hypothesized that the reactivation of the metabolic pathways that were suppressed in antibiotic-resistant bacteria could potentiate the killing efficacy of antibiotics. Based on our assumption that metabolic state determines antibiotic resistance and sensitivity, we started our experiments with clinically isolated uropathogenic Escherichia coli. By comparing the metabolomic profiles of antibiotic-sensitive bacteria and antibiotic-resistant bacteria, the most impacted pathway and crucial biomarker were identified. The reactivation of the most impacted pathway by the crucial metabolite potentiated the antibiotic-induced killing efficacy. The potentiation efficacy was tested in the currently antibiotic classes in clinic. In addition to the multidrug-resistant E. coli, this effect is also assessed in other multidrug-resistant bacteria including Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella pneumoniae, Vibrio alginolyticus, V. parahaemolyticus, and Edwardsiella tarda, and other forms of antibiotic tolerance and resistance, the persister and bacterial biofilms. To test the effect in vivo, two mouse infection models were tested. In the urinary infection model, catheters were colonized, and treatment was applied 48 h post-surgery. In the systemic infection model, mice were infected with multidrug resistant bacteria and treatment was applied at 1 h and 12 h post-infection. Then, mouse survival was accounted and bacterial loads were determined in blood, spleen, kidney, and liver. The in vivo effect was further rationalized through phamokinetic analysis of the metabolite and antibiotics. To explore the possible effect of metabolite in slowing new resistance, antibiotic-sensitive and –resistant bacteria were evolved in vitro and in vivo with antibiotic only or plus the metabolite. The resistance was determined by MICs. To reveal the mechanism of the metabolite in potentiating antibiotic efficacy, intracellular concentration of antibiotics, antibiotic hydrolysis and efflux were quantified. Bacterial knock out libraries were used to identify the potential targets of the metabolite.