Evaluation of Short-Time Method with Tetrazolium Salt and Electron Carrier for Measuring Minimal Inhibitory Concentration of Antimicrobial Agents

Background: Minimal inhibitory concentration (MIC) measurements are usually conducted using the agar plate dilution test or the microdilution test. Methods for rapid evaluation of antimicrobial susceptibility using ATP measurements with luciferin-luciferase reagents and a tetrazolium salt have been reported.

Material and methods: We recently measured the minimal inhibitory concentrations (MICs) of various antimicrobial agents against 96 strains of Escherichia coli (1 standard strain and 95 clinically isolated strains) with Cell Counting Kit-8 containing a tetrazolium salt and an electron carrier using the Dry Plate Eiken for MIC measurement and compared the results with that obtained using the standard method.

Results: Agreement exceeded 90% for ampicillin, cefotiam, ceftazidime, flomoxef, aztreonam, imipenem, meropenem, gentamicin, and levofloxacin, and complete agreement exceeded 90% for cefotiam, ceftazidime, flomoxef, aztreonam, imipenem, meropenem, and levofloxacin. Agreement was in the range of 78.1- 86.5% for piperacillin, cefazolin, cefpodoxime, amikacin, and minocycline with complete agreement in the range 55.2-78.1%. Cefaclor had the lowest agreement and complete agreement (42.7% and 38.5, respectively). For the antimicrobial agents with low agreement (piperacillin, cefaclor, cefazolin, amikacin and minocycline), MICs measured by the short-time method were often below onehalf or one-fourth of that measured by the standard method, and regrowth of the bacteria appeared to occur 6 hours after the start of the short-time method.

Conclusion: The results of this study indicate that measuring absorbance with Cell Counting Kit-8 enables counting of viable bacteria. The use of this method for measuring MICs (short-time method) allowed the results of the susceptibility testing to be obtained in 6 hours, and the results correlated well with the results obtained using the standard method. Therefore, this method appears useful for the early detection of drug-resistant bacteria.

Author(s):

Yoshida M, Horino T, Nakazawa Y, Yoshikawa K, Hori S

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