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Updated: May 19, 2026
Broth Microdilution In Vitro Screening: An Easy and Fast Method to Detect New Antifungal Compounds
Published on: February 14, 2018
Susceptibility screening of hyphae-forming fungi with a new, easy, and fast inoculum preparation method
Arno Schmalreck1, Birgit Willinger, Viktor Czaika
1MBS, Munich, Germany.
Abstract:
In vitro susceptibility testing of clinically important fungi becomes more and more essential due to the rising number of fungal infections in patients with impaired immune system. Existing standardized microbroth dilution methods for in vitro testing of molds (CLSI, EUCAST) are not intended for routine testing. These methods are very time-consuming and dependent on sporulating of hyphomycetes. In this multicentre study, a new (independent of sporulation) inoculum preparation method (containing a mixture of vegetative cells, hyphae, and conidia) was evaluated. Minimal inhibitory concentrations (MIC) of amphotericin B, posaconazole, and voriconazole of 180 molds were determined with two different culture media (YST and RPMI 1640) according to the DIN (Deutsches Institut für Normung) microdilution assay. 24 and 48 h MIC of quality control strains, tested per each test run, prepared with the new inoculum method were in the range of DIN. YST and RPMI 1640 media showed similar MIC distributions for all molds tested. MIC readings at 48 versus 24 h yield 1 log(2) higher MIC values and more than 90 % of the MICs read at 24 and 48 h were within ± 2 log(2) dilution. MIC end point reading (log(2 MIC-RPMI 1640)-log(2 MIC-YST)) of both media demonstrated a tendency to slightly lower MICs with RPMI 1640 medium. This study reports the results of a new, time-saving, and easy-to-perform method for inoculum preparation for routine susceptibility testing that can be applied for all types of spore-/non-spore and hyphae-forming fungi.
Insights
A new, sporulation-independent method for preparing fungal cultures simplifies in vitro susceptibility testing. This time-saving approach aids in the routine testing of molds, crucial for immunocompromised patients.
Area of Science:
- Medical Mycology
- Antimicrobial Susceptibility Testing
Background:
- Rising incidence of fungal infections in immunocompromised patients necessitates reliable in vitro susceptibility testing.
- Current standardized methods for mold susceptibility testing are time-consuming and rely on fungal sporulation.
Purpose of the Study:
- To evaluate a novel, sporulation-independent inoculum preparation method for routine in vitro susceptibility testing of molds.
- To compare the performance of this new method with existing standards.
Main Methods:
- A multicenter study evaluated a new inoculum preparation method (vegetative cells, hyphae, conidia) for 180 molds.
- Minimal Inhibitory Concentrations (MICs) for amphotericin B, posaconazole, and voriconazole were determined using the DIN microdilution assay with YST and RPMI 1640 media.
- Quality control strains were tested to validate the new method's consistency.
Main Results:
- The new inoculum preparation method yielded MIC values within DIN ranges for quality control strains at 24 and 48 hours.
- Both YST and RPMI 1640 media showed comparable MIC distributions across all tested molds.
- MIC readings at 48 hours were generally higher than at 24 hours, with over 90% agreement within ±2 log(2) dilutions.
Conclusions:
- The evaluated method offers a time-saving and user-friendly alternative for routine fungal susceptibility testing.
- This approach is applicable to all types of fungi, including spore-forming, non-spore-forming, and hyphae-forming species.
- The new method addresses limitations of current sporulation-dependent techniques.
