ABSTRACT
BACKGROUND: A spectrophotometric approach to minimum inhibitory concentration (MIC) determination for filamentous fungi may provide an objective and rapid MIC reading, and quantify the hyphal growth of molds. In this study, we evaluated two spectrophotometric broth microdilution methods (SBM) to determine amphotericin B and itraconazole MICs for Aspergillus species isolated from clinical specimens. METHODS: A total of 80 clinical isolates (20 A. fumigatus, 20 A. flavus, 18 A. niger, 20 A. terreus, and 2 A. nidulans) were tested for amphotericin B and itraconazole susceptibility by the broth microdilution method. The MIC endpoint was calculated by the spectrophotometer with microplate reader (SBM-Spec method) or colorimetric XTT (tetrazolium dye) method (SBM-XTT method). The results of the SBM method were compared with those of NCCLS M38-A broth microdilution method. RESULTS: The MICs of amphotericin B by the NCCLS M38-A method ranged from 0.125 to 8 g/mL, and those of itraconazole ranged from 0.25 to 2micrograms/mL. The agreement of SBM-Spec and SBM-XTT methods within one dilution of the NCCLS M38 reference were 98.8% and 96.3% for the ampho-tericin B, and 98.8% and 100% for itraconazole, respectively. The agreements between SBM-Spec and SBM-XTT methods were 97.5% for amphotericin B and 98.8% for itraconazole. CONCLUSIONS: In antifungal susceptibility testing of Aspergillus species, the SBM method includ-ing SBM-Spec and SBM-XTT methods showed high levels of agreements with the NCCLS M38-A method. The SBM methods can be useful in the clinical laboratory.
Subject(s)
Amphotericin B , Aspergillus , Fungi , Itraconazole , Microbial Sensitivity Tests , NigerABSTRACT
BACKGROUND: Minimum inhibitory concentration (MIC) endpoint determination is the major varia-tion source for the fluconazole susceptibility test, especially for Candida albicans. In this study, we evaluated spectrophotometric broth microdilution methods using RPMI 1640 and RPMI supple-mented with 18 g of glucose per liter (RPMI-2% glucose) for determining fluconazole susceptibility of C. albicans. METHODS: A total of 129 clinical isolates of C. albicans were tested by the broth microdilution method using RPMI and RPMI-2% glucose. The MIC endpoint was calculated objectively with the spectrophotometer set at 405 nm. These results were compared to those by the National Commit-tee for Clinical Laboratory Standards (NCCLS) macrodilution method and the agar dilution method. RESULTS: The mean absorbances in the drug-free wells in RPMI and RPMI-2% glucose were 0.208 +/- 0.014 and 0.316 +/- 0.061, respectively, at 24 h and 0.339 +/- 0.094 and 0.530 +/- 0.104, respectively, at 48 h (P < 0.01). The agreement of the microdilution method with the RPMI within two doubling dilutions of the macrodilution reference were 91.5% (118/129) at 24 h and 76.7% (99/129) at 48 h. The percentage of agreement in the microdilution method with the RPMI-2% glucose were significantly higher: 100% (129/129) at 24 h and 99.2% (128/129) at 48 h (P < 0.01). In addition, the MIC endpoints were easier to detect in RPMI-2% glucose, because of the greater difference in absorbance in between grown wells and fluconazole-inhibited wells (P < 0.01). CONCLUSIONS: The spectrophotometric microdilution method with RPMI-2% glucose may have an excellent agreement with the NCCLS broth macrodilution method and may provide more easily determined MIC endpoints for fluconazole susceptibility testing for C. albicans.