Development and multicentre validation of an agar-based screening method for echinocandin susceptibility testing of Aspergillus species.

ABSTRACT:

Background

Reference antifungal susceptibility testing of echinocandins against Aspergillus spp. relies on the determination of the minimal effective concentration, which is difficult to perform, time-consuming and subjective. We developed and evaluated in a multicentre study an agar-based screening method for echinocandin susceptibility testing of Aspergillus spp.

Methods

Forty WT isolates [10 Aspergillus fumigatus species complex (SC), 10 Aspergillus flavus SC, 10 Aspergillus terreus SC and 10 Aspergillus niger SC] and 4 non-WT A. fumigatus isolates with or without known fks alterations were used. The optimal test conditions and stability over time were evaluated in preliminary studies monitoring colony growth. Twenty-microlitre aliquots of 1-2 McFarland inocula in 0.1% Tween 20 aqueous solution were added to each well and plates were incubated for 24/48?h at 35?±?2°C. Subsequently, all isolates were tested blindly at three centres using four-well screening plates, containing anidulafungin, caspofungin, micafungin or no antifungal in each of the four wells, respectively.

Results

WT isolates produced fluffy colonies on drug-free agar wells only. The non-WT isolates produced fluffy colonies on echinocandin-containing and control agar wells. Using the echinocandin concentrations of 0.25?mg/L anidulafungin, 1?mg/L caspofungin and 0.125?mg/L micafungin, and the compact (non-fluffy) versus fluffy colony morphology endpoint, all centres successfully discriminated non-WT and WT strains even after 24?h. Among the three echinocandins, anidulafungin produced the clearest endpoints.

Conclusions

The four-well plate agar method is suitable for echinocandin susceptibility screening of Aspergillus spp. and can be used to detect echinocandin non-WT isolates.

SUBMITTER: Meletiadis J 

PROVIDER: S-EPMC6640300 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature