In this study we aimed to determine the development of resistance against subinhibitory concentrations of ciprofloxacin and levofloxacin of the Escherichia coli strains. Fifty E.coli strains that are susceptible to ciprofloxacin and levofloxacin have been included in this study. Minimal Inhibitory Concentration (MIC)s were detected by macro dilution method. In order to investigate the resistance development, the E.coli strains were left to incubate at 37°C for one night in subinhibitory concentrations (1/2 x MIC), then incubated in antibiotic free Mueller-Hinton broth for one night. In this way, after eight sequential passages, second MICs were detected. The third MICs were determined by repeating the same process based on the newly determined MIC values. Accordingly, as of the first MICs, after 16 successive passages an increase of MIC values against subinhibitory concentrations of ciprofloxacin was observed in all E.coli strains. This was the same for levofloxacin except for three strains; however, there were striking increases such as 64 or 128 times in the remaining strains. The use of antimicrobials for long treatment durations, taking care to monitor dose regulations and proper usage is recommended.