Occurrence of triazole-resistant Aspergillus fumigatus with TR34/L98H mutations in outdoor and hospital environment in Kuwait.

Invasive aspergillosis due to triazole-resistant Aspergillus fumigatus, a common airborne fungal pathogen, has emerged in some European and Asian countries, likely due to exposure to azole fungicides in the environment. This study determined the occurrence of triazole-resistant A. fumigatus in outdoor and hospital environment in Kuwait, a semi-arid desert country in Arabian Peninsula. Outdoor/indoor air, water and cotton swab samples were cultured on malt extract agar and A. fumigatus colonies were identified by phenotypic and molecular methods. Drug susceptibility of A. fumigatus isolates to itraconazole, posaconazole and voriconazole was carried out by an Etest and a broth microdilution method. Resistance mechanisms involving cyp51A mutations were probed by mixed-format real-time (MF-rt)-PCR assays. Triazole-resistant isolates were typed by nine-locus microsatellite analysis. A multiplex allele-specific (MAS)-PCR assay was developed for detection of L98H mutation in cyp51A. Of 115 A. fumigatus isolates obtained from 362 environmental samples from across Kuwait, 8 isolates were resistant to itraconazole, posaconazole and voriconazole. All itraconazole-resistant isolates contained a 34-bp tandem repeat (TR34) in the promoter region and a L98H mutation at codon 98 (TR34/L98H) in cyp51A. These mutations were absent in all itraconazole-susceptible isolates. MAS-PCR accurately detected L98H mutation in all triazole-resistant isolates. Three microsatellite patterns were observed among resistant isolates with one pattern clustering with Indian clinical and environmental isolates. Triazole-resistant A. fumigatus with TR34/L98H mutations in cyp51A is prevalent in Kuwait. Although triazole-resistant A. fumigatus has not yet been isolated from clinical specimens, its presence in the environment suggests that the possibility of susceptible individuals getting infected with such strains exists and may pose therapeutic challenges in its management.