A rapid microplate method for the proliferation assay of fungi and the antifungal susceptibility testing using the colorimetric microbial viability assay

摘要

A rapid microplate method for the proliferation assay of fungi and the antifungal susceptibility testing using the colorimetric microbial viability assay based on the reduction in a tetrazolium salt 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium, monosodium salt (WST-8) with 2-methyl-1,4-napthoquinone as the electron mediator was developed. The proposed method was useful to measure the proliferation of 18 kinds of moulds and seven kinds of yeasts, including representative pathogens such as Aspergillus spp., Candida spp. and Cryptococcus spp. Linear relationships between the absorbance and viable fungal cell density were obtained for all fungi, suggesting that the absorbance change reflected the fungal proliferation. In addition, the minimum inhibitory concentrations (MICs) against a variety of different pathogenic moulds and yeasts for amphotericin B, itraconazole and 5-flucytosine were determined by susceptibility testing using the proposed method and compared with those obtained using the conventional broth microdilution method. There was an excellent agreement between the results obtained using the WST-8 colorimetric method and those obtained using the conventional Clinical and Laboratory Standard Institute method. The WST-8 colorimetric assay is a useful method for rapid determination of accurate MICs for a variety of different fungi