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[Colorimetric broth microdilution for antifungal susceptibility testing]
N Yamane1, M Tosaka, Y Okazawa
1Department of Laboratory Medicine, Kumamoto University Medical School Japan.
Rinsho Byori. the Japanese Journal of Clinical Pathology
|January 1, 1996
Summary
A new colorimetric broth microdilution method using resazurin offers a precise and reproducible way to test antifungal susceptibility. This method provides results comparable to the standard National Committee for Clinical Laboratory Standards macrodilution method.
Area of Science:
- Clinical Microbiology
- Mycology
- Antimicrobial Susceptibility Testing
Background:
- Accurate antifungal susceptibility testing is crucial for effective treatment of fungal infections.
- Established methods like the National Committee for Clinical Laboratory Standards (NCCLS) macrodilution method can be time-consuming and resource-intensive.
- Development of rapid, reliable, and cost-effective alternative methods is needed.
Purpose of the Study:
- To develop and evaluate a colorimetric broth microdilution method for antifungal susceptibility testing.
- To assess the performance of this modified method against the standard NCCLS macrodilution technique.
- To determine the reproducibility and agreement of Minimum Inhibitory Concentrations (MICs) obtained by the new method.
Main Methods:
- A colorimetric modification of the NCCLS broth microdilution method was developed using air-dried trays.
- Serial two-fold dilutions of amphotericin B (AMPH), flucytosine (5-FC), and fluconazole (FCZ) were prepared and reconstituted with yeast inocula.
- An oxidation-reduction indicator (sodium resazurin) was added to RPMI 1640 medium, and trays were incubated at 35°C, read at 24 and 48 hours.
Main Results:
- The resazurin colorimetric method demonstrated excellent reproducibility (99-100% agreement in duplicated dilutions).
- MICs determined by the microdilution method showed good agreement with the NCCLS macrodilution method, particularly at 48 hours (78-97% agreement).
- While 24-hour readings showed slightly lower agreement (61.7%), the 48-hour readings were comparable, with specific high agreements for 5-FC (97%) and AMPH (85%).
Conclusions:
- The resazurin colorimetric broth microdilution method is a practical, precise, and reproducible technique for antifungal susceptibility testing.
- This method offers comparable MIC results to the NCCLS macrodilution reference method.
- The developed method has the potential to be a valuable tool in clinical laboratories for guiding antifungal therapy.