Exploring the in-situ evolution of Nitrofurantoin resistance in clinically derived Uropathogenic Escherichia coli isolates

BACKGROUND Nitrofurantoin has been re-introduced as a first-choice antibiotic to treat uncomplicated acute urinary tract infections in England and Wales. Its mode of action involves initial reduction by nitroreductases, to generate electrophilic intermediates that inhibit protein and nucleic acid synthesis. Highly effective against common uropathogens such as Escherichia coli , its use is accompanied by a low incidence (<10%) of antimicrobial resistance. Resistance to Nitrofurantoin is predominantly via the acquisition of loss-of-function, step-wise mutations in the nitroreductase genes nfsA and nfsB . OBJECTIVE To explore the in situ evolution of Nit R , longitudinal uropathogenic E. coli isolates recovered from two rUTI patients. RESULTS Growth rate analysis identified a 2-10% slower doubling time for Nitrofurantoin resistant strains, but statistically, these data suggested there was no fitness advantage of evolved strains over their sensitive predecessor (ANOVA P-value = 0.13). Genetic manipulation of E. coli to mimic nitrofurantoin resistance evolution, again confirmed no fitness advantages (ANOVA P-value = 0.22). Rather, further analysis argued that a first-step mutant gained a selective advantage, at sub-MIC (4-8 mg/L) nitrofurantoin concentrations. CONCLUSION Correlation of these findings to Nitrofurantoin pharmacokinetic data suggests that the low incidence of E. coli Nit R , within the community, is driven by urine-based nitrofurantoin concentrations that selectively inhibit the growth of E. coli strains carrying the key first-step loss-of-function mutation.