Azole-Resistant Aspergillus fumigatus Clinical Isolate Screening in Azole-Containing Agar Plates (EUCAST E.Def 10.1): Low Impact of Plastic Trays Used and Poor Performance in Cryptic Species.

Azole-containing agar is used in routine Aspergillus fumigatus azole resistance screening. We evaluated the impact of the type of plastic used to prepare in-house agar plates on the procedure's performance against A. fumigatus sensu stricto and cryptic species. A. fumigatus sensu stricto ( = 91) and cryptic species ( = 52) were classified as susceptible or resistant (EUCAST E.Def 9.3.2; clinical breakpoints v10). In-house azole-containing agar plates were prepared following EUCAST E.Def 10.1 on three types of multidish plates. We assessed the sensitivity, specificity, and agreement values of the agar plates to screen for azole resistance. Overall, sensitivity and specificity values of the agar screening method were 100% and 93.3%, respectively. The type of tray used did not affect these values. All isolates harboring TR-L98H substitutions were classified as resistant to itraconazole and voriconazole by the agar method; however, false susceptibility (very major error) to posaconazole was not uncommon and happened in isolates with posaconazole MICs of 0.25 mg/liter. Isolates harboring G54R and TR-Y121F-T289A substitutions were correctly classified by the agar method as itraconazole/posaconazole resistant and voriconazole resistant, respectively. False resistance (major error) occurred in isolates showing tiny fungal growth. Finally, agreements between both procedures against cryptic species were much lower. Azole-containing agar plates are a convenient and reliable tool to screen for resistance in A. fumigatus sensu stricto; the type of plastic tray used minimally affects the method. On the contrary, the performance against cryptic species is rather poor.