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Multicenter evaluation of four methods of yeast inoculum preparation
M A Pfaller1, L Burmeister, M S Bartlett
1Veterans Administration Medical Center, Iowa City, Iowa 52242.
Abstract:
We initiated a comparative study of four methods of yeast inoculum preparation: a spectrophotometric method, the Wickerham card method, a hemacytometer method, and the Prompt inoculation system. The variability in inoculum size obtained when each method was applied to two strains each of Candida albicans, Candida tropicalis, Candida parapsilosis, Torulopsis glabrata, Cryptococcus neoformans, and Saccharomyces cerevisiae was analyzed in a single laboratory. Each method was performed in triplicate on the same day and on three separate days to provide estimates of within-day and between-day variations. Inoculum size was determined by viable colony counts. The greatest range of inoculum sizes was seen with the Wickerham card method. Viable counts ranged from 1.1 X 10(6) to 24.2 X 10(6) CFU/ml among the 12 yeast isolates. The greatest variation was observed with the Prompt system. Within-day coefficients of variation averaged 19% (range, 4 to 45%), and between-day coefficients of variation averaged 22% (range, 3 to 51%). Variation between laboratories was evaluated by comparing inoculum values obtained by each method in three different laboratories for two strains of C. albicans. The spectrophotometric method was the least variable and the Wickerham card and hemacytometer methods were the most variable methods between laboratories. The spectrophotometric method is recommended as the method of choice for preparation of a standardized inoculum suspension for susceptibility testing of yeasts.
Insights
The spectrophotometric method is the most reliable for preparing yeast inoculums for susceptibility testing, showing less variability than Wickerham card, hemacytometer, or Prompt systems. This standardization is crucial for accurate antifungal testing.
Area of Science:
- Clinical Microbiology
- Mycology
- Antimicrobial Susceptibility Testing
Background:
- Accurate yeast inoculum preparation is critical for reliable antifungal susceptibility testing.
- Existing methods for yeast inoculum preparation exhibit variability, potentially impacting test results.
Purpose of the Study:
- To comparatively evaluate four common methods for yeast inoculum preparation: spectrophotometric, Wickerham card, hemacytometer, and Prompt inoculation system.
- To assess the within-day and between-day variability of each method across multiple yeast species.
- To determine inter-laboratory variation for selected methods.
Main Methods:
- Four methods (spectrophotometric, Wickerham card, hemacytometer, Prompt) were used to prepare inoculums for six yeast species (Candida albicans, Candida tropicalis, Candida parapsilosis, Torulopsis glabrata, Cryptococcus neoformans, Saccharomyces cerevisiae).
- Inoculum size was quantified by viable colony counts (CFU/ml).
- Within-day, between-day, and inter-laboratory variations were analyzed.
Main Results:
- The Wickerham card method showed the widest range of inoculum sizes (1.1 x 10^6 to 24.2 x 10^6 CFU/ml).
- The Prompt system exhibited the greatest variation, with average within-day coefficients of variation of 19% and between-day of 22%.
- The spectrophotometric method demonstrated the least variability across laboratories, while Wickerham card and hemacytometer methods were most variable.
Conclusions:
- The spectrophotometric method is recommended for preparing standardized yeast inoculum suspensions for susceptibility testing.
- Standardization of inoculum preparation is essential for consistent and reproducible antifungal susceptibility testing results.
- Further validation of yeast inoculum preparation methods is warranted to ensure clinical accuracy.