Phenotypic tolerance: Antibiotic enrichment of noninherited resistance in bacterial populations

When growing bacteria are exposed to bactericidal concentrations of antibiotics, the sensitivity of the bacteria to the antibiotic commonly decreases with time, and substantial fractions of the bacteria survive. Using Escherichia coli CAB1 and antibiotics of five different classes (ampicillin, ciprofloxacin, rifampin, streptomycin, and tetracycline), we examine the details of this phenomenon and, with the aid of mathematical models, develop and explore the properties and predictions of three hypotheses that can account for this phenomenon: (i) antibiotic decay, (ii) inherited resistance, and (iii) phenotypic tolerance. Our experiments cause us to reject the first two hypotheses and provide evidence that this phenomenon can be accounted for by the antibiotic-mediated enrichment of subpopulations physiologically tolerant to but genetically susceptible to these antibiotics, phenotypic tolerance. We demonstrate that tolerant subpopulations generated by exposure to one concentration of an antibiotic are also tolerant to higher concentrations of the same antibiotic and can be tolerant to antibiotics of the other four types. Using a mathematical model, we explore the effects of phenotypic tolerance to the microbiological outcome of antibiotic treatment and demonstrate, a priori, that it can have a profound effect on the rate of clearance of the bacteria and under some conditions can prevent clearance that would be achieved in the absence of tolerance.