The ability in managing reactive oxygen species affects Escherichia coli persistence to ampicillin after nutrient shifts

Bacterial persistence profoundly impacts biofilms, infections, and antibiotic effectiveness. Persister formation can be substantially promoted by nutrient shift, which commonly exists in natural environments. However, mechanisms that promote persister formation remain poorly understood. Here, we investigated the persistence frequency of Escherichia coli after switching from various carbon sources to fatty acid and observed drastically different survival rates. While more than 99.9% of cells died during a 24-hour ampicillin (AMP) treatment after the glycerol to oleic acid (GLY → OA + AMP) shift, a surprising 56% of cells survived the same antibiotic treatment after the glucose to oleic acid (GLU → OOA + AMP) shift. Using a combination of single-cell imaging and time-lapse microscopy, we discovered that the induction of high levels of reactive oxygen species (ROS) by AMP is the primary mechanism of cell killing after switching from gluconeogenic carbons to OA + AMP. Moreover, the timing of the ROS burst is highly correlated (R² = 0.91) with the start of the rapid killing phase in the time-kill curves for all gluconeogenic carbons. However, ROS did not accumulate to lethal levels after the GLU → OA + AMP shift. We also found that the overexpression of the oxidative stress regulator and ROS detoxification enzymes strongly affects the amounts of ROS and the persistence frequency following the nutritional shift. These findings elucidate the different persister frequencies resulting from various nutrient shifts and underscore the pivotal role of ROS. Our study provides insights into bacterial persistence mechanisms, holding promise for targeted therapeutic interventions combating bacterial resistance effectively.