Azole-Resistant Among Danish Cystic Fibrosis Patients: Increasing Prevalence and Dominance of TR/L98H

Azole-resistant (azole-R) is an increasing challenge worldwide. Patients with cystic fibrosis (CF) are at risk of colonization and disease due to a favorable lung environment for microorganisms. We performed a nationwide study in 2018 of azole-non-susceptible in CF patients and compared with data from two prior studies. All airway samples with mold isolates from patients monitored at the two CF centers in Denmark (RH, Jan-Sept and AUH, Jan-Jun) were included. Classical species identification (morphology and thermo-tolerance) was performed and MALDI-TOF/β-tubulin sequencing was performed if needed. Susceptibility was determined using EUCAST E.Def 10.1, and E.Def 9.3.2. sequencing and STR genotyping were performed for azole-non-susceptible isolates and relevant sequential isolates. In total, 340 mold isolates from 159 CF patients were obtained. The most frequent species were (266/340, 78.2%) and (26/340, 7.6%). Azole-R was cultured from 7.3% (10/137) of patients, including 9.5% (9/95) of patients at RH and 2.4% at AUH (1/42), respectively. In a 10-year perspective, azole-non-susceptibility increased numerically among patients at RH (10.5% in 2018 vs 4.5% in 2007-2009). Cyp51A resistance mechanisms were found in nine azole-R from eight CF patients. Five were of environmental origin (TR/L98H), three were human medicine-driven (two M220K and one M220R), and one was novel (TR /L98H) and found in a patient who also harbored a TR/L98H isolate. STR genotyping identified 27 unique genotypes among 45 isolates and ≥2 genotypes in 8 of 12 patients. This included one patient carrying two unique TR/L98H isolates, a rare phenomenon. Genotyping of sequential TR /L98H and TR/L98H isolates from the same patient showed only minor differences in 1/9 markers. Finally, azole-R was found in three patients including two with Cyp51A alterations (M217I and G51A, respectively). Azole-R is increasing among CF patients in Denmark with the environmentally associated resistance TR/L98H mechanism being dominant. Mixed infections (wildtype/non-wildtype and several non-wildtypes) and a case of potential additional tandem repeat acquisition were found. However, similar genotypes were identified from another patient (and outside this study), potentially suggesting a predominant TR/L98H clone in DK. These findings suggest an increasing prevalence and complexity of azole resistance in .