One step forward to improve the latest method of antibacterial susceptibility testing of vitro-cultured bacteria: an implication for antibacterial efficacy of Enrofloxacine on <Emphasis Type="Italic">Aeromonas hydrophila</Emphasis>

Recently a new method of antibacterial susceptibility testing has been developed, which based on the reduction of the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) by bacteria. This study aimed to improve the latest developed method in terms of the period of time, concentration of bacterium, accuracy of assay and minimizing the toxic effect of the MTT solution. The commonly used broth microdilution method was compared in this study, as well. The minimum inhibition concentration (MIC) of the enrofloxacine against Aeromonas hydrophila as an opportunistic and ubiquitous bacterium was tested by using the improved MTT method. Using both methods showed that enrofloxacine exerts an excellent antibacterial effect against A. hidrophyla with MIC value of 62.5 ng/ml. The simplicity of the improved MTT method was shown with performing all steps of the assay in one plate, using rather low concentration of the bacterium (500 CFU/well) and shortening of the incubation time to 9 h. Moreover, 5–30 min incubation of bacteria with MTT solution excludes any toxic effect of tetrazolium salt against bacteria. Thus, our results suggest that enrofloxacine might be considered as a potent antibacterial agent against A. hydrophila induced contaminations. Moreover, MTT method with current improved characteristics such as short incubation time, low concentration of bacteria, and high sensitivity could be more practical alternative for the broth microdilution method in antibacterial susceptibility testing protocol.