Combination of glucose and oxygen gradients dictates metabolic regime and antibiotic tolerance of biofilms
摘要
Biofilms are structured bacterial communities that underlie many persistent infections, exhibiting strong survival under antibiotics that cannot be explained by genetic resistance alone. Here we uncover a biofilm-specific mechanism of antibiotic tolerance driven by spatial gradients. Using an agarose-based Escherichia coli biofilm system, we found a non-monotonic relationship between glucose availability and biofilm growth: high glucose paradoxically reduced final biomass. This arose from the overlap of the glucose penetration zone and the anaerobic core of the biofilm at high glucose, triggering mixed-acid fermentation. We identified acetate, a fermentation by-product secreted by the biofilm, as a potent inducer of tolerance to gentamicin and ciprofloxacin. This effect was biofilm-specific: acetate enhanced antibiotic killing instead of tolerance in planktonic cultures. In biofilms, acetate entered the TCA cycle via the glyoxylate shunt, driving respiration that depleted oxygen and maintained a persistent anaerobic core, thereby blocking antibiotic efficacy. Supplementing the alternative electron acceptor fumarate fully reversed biofilm tolerance. Our work demonstrates that community-level metabolic organization can invert the relationship between metabolic activity and antibiotic susceptibility, and reveals that interventions effective against planktonic bacteria may be counterproductive in biofilms. These findings underscore the necessity of considering spatial physiology in the development of anti-biofilm therapies.
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