Isavuconazole nonwildtype Aspergillus fumigatus isolates from a global surveillance study display alterations in multiple genes involved in the ergosterol biosynthesis pathway not previously associated with resistance to other azoles.

Objectives: We evaluated 35 azole nonwildtype Aspergillus fumigatus isolates that were collected during 2017-2018 using whole genome sequencing (WGS) to detect alterations in the genes involved in the ergosterol biosynthesis pathway as well as other genes associated with azole resistance.

Methods: Among 297 A fumigatus isolates collected worldwide, 36 isolates displayed nonwildtype MIC values to isavuconazole, itraconazole, or voriconazole when tested by the CLSI reference broth microdilution method. Isolates were submitted to WGS and results were compared to 2 azolewildtype isolates.

Results: Among the 35 sequenced isolates (1 failed to produce quality sequences), 29 were nonwildtype to isavuconazole, 16 were nonwildtype to itraconazole, and 9 were nonwildtype to voriconazole (CLSI M59Ed2 criteria). A total of 9 isolates carried Cyp51A TR34/L98H alterations (8 from Italy and 1 from Belgium) and had nonwildtype MIC values for ≥2 azoles. A Cyp51B Q42L mutation was detected in 3 isolates, 1 nonwildtype voriconazole and 2 nonwildtype isavuconazole isolates. Three isolates harboured multiple mutations in Cyp51A (F46Y, M172V, E427K ± N248T, and D255E), including 1 isolate with the Cyp51B Q42L mutation. Mutations causing frameshifts, early termination, and duplications were observed among several genes and were more prevalent in isavuconazole nonwildtype isolates (66.7%) than in the isolates that were nonwildtype to 1 or 2 other azoles (22.2%). Nine isolates harboured frameshift mutations in a ERG25 homologue that is usually associated with changes in other genes and should be further evaluated.

Conclusions: Cyp51A L98H/TR34 was the most common alteration observed among the azole nonwildtype A fumigatus isolates from a large surveillance study; however, only isolates that were nonwildtype to isavuconazole had alterations in multiple analysed genes. These isolates deserve further evaluation.