Susceptibility Screening of Hyphae-Forming Fungi with a New, Easy, and Fast Inoculum Preparation Method
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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 log2 higher MIC values and more than 90 % of the MICs read at 24 and 48 h were within ±2 log2 dilution. MIC end point reading (log2 MIC-RPMI 1640−log2 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.
KeywordsSusceptibility testing New inoculum preparation method MIC distribution Multicentre study Hyphomycetes
Free of charge was provided the antifungal agents anidulafungin, fluconazole, and voriconazole by Pfizer GmbH (Germany), and MSD Sharp & Dohme GmbH (Germany), posaconazole by Essex Pharma GmbH (Germany) and miconazole by Astellas Pharma GmbH (Germany). This work was supported by a grant from Pfizer GmbH (Germany) for purchasing and manufacturing of culture media, ready-to-use microdilution trays with the eight antifungal agents and for transport materials of collected strains. The opinions expressed in this paper are those of the authors and do not necessarily represent those of the pharmaceutical companies.
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