Effects of Fosfomycin-Resistant Mutants on Bacterial Growth and Efficacy

Introduction: Fosfomycin (FOS) is an antibiotic used in treating urinary tract infections (UTI) and cystitis (bladder infection) in women. The antibiotic, which is bactericidal, works by inhibiting the bacterial cell wall biogenesis by inactivating the enzyme UDP-N-acetylglucosamine-3-enolpyruvyltransferase, also known as MurA. However, several mechanisms of FOS resistance have been reported. The resistance mechanisms involve reduced uptake of FOS due to a defect in one of the two transporters caused by mutations in the structural gene. Objective: This study looked at the differences in virulence and efficacy in the murine UTI model with FOS-resistant mutants. Method: The study started with two FOS-susceptible strains (E. coli 045 and 167). Isolated colonies from each parent strain were inoculated into Tryptic Soy Broth (TSB) then incubated for 5 hours. Next, the broth culture was swabbed onto a TSA plate. A 200 ug FOS disc was placed on the agar surface and incubated. The following day, colonies were selected from inside or around the edge of the zone of inhibition surrounding the FOS disc. These colonies were inoculated into TSB and incubated. This process was repeated for three passages (E. coli 045{A, B, C}and 167{A, B, C}). Next, Mueller-Hinton agar was prepped and used to determine the FOS minimum inhibitory concentration (MIC) for each of the passages of the two strains. Finally, a growth curve in Mueller-Hinton broth was performed to determine the effect the passages had on the growth patterns over time at 0, 1, 2, 4, and 6 hrs. Results: Fosfomycin MICs increased from 8 ug/mL to 512ug/mL from the parent to the resistant generated mutants on the Mueller-Hinton agar. It showed consistency in FOS resistance from the parent to the resistant generated mutants. Growth curves exhibited comparable patterns for parent and resistant mutants. Conclusion: Fosfomycin-resistant mutants were generated both an E. coli 045 and 167 strain following serial passages on plates with a FOS disc. Using the Mueller-Hinton agar method, Fosfomycin MICs increased from parent to the resistant generated mutants (8 ug/mL to 512ug/mL). These results indicate that strains with elevated MICs may still be successfully treatable with Fosfomycin. Future studies will include looking at the same strains in an animal model of UTI and determining if the mutation is stable and can be treated with FOS.