Epidemiology of Azole-Resistance and CYP51A Mutations in Aspergillus Isolates Recovered from Lung Transplant Recipients who Received Voriconazole Prophylaxis

Background: We previously showed that voriconazole (VOR) prophylaxis for 3-6 months after lung transplantation reduced but did not eliminate fungal lung infection. Breakthrough (BT) infections were predominantly due to A. fumigatus (Af). Azole-resistant Af with CYP51A mutations have emerged in Europe, Asia, and the Middle East largely due to agricultural fungicide use.

Methods: We determined itraconazole (ITR), posaconazole (POS), and VOR resistance rates among Aspergillus from bronchoalveolar lavages of lung transplant recipients. Minimum inhibitory concentrations (MICs) were measured by CLSI broth microdilution. Azole susceptibility breakpoints were defined according to EUCAST. CYP51A genes were PCR-amplified and sequences were compared to GenBank references.

Results: 56 isolates were tested (21 Af, 16 A. niger, 11 A. versicolor, 4 A. flavus, 3 A. calidoustus, 1 A. nidulans). 46% of isolates were azole breakthroughs. Median ITR MIC was higher than POS (p=0.01) or VOR (p=0.001). There were direct correlations between ITR, POS, and VOR MICs (R≥0.72). 36% (20), 43% (24) and 4% (7) of isolates were non-susceptible to ITR, POS, and VOR respectively. 5% of isolates were pan-azole resistant (2 A. calidoustus, 1 Af). There were no differences in Aspergillus azole MICs between BT and non-BT isolates. All 3 A. calidoustus were resistant to ITR and POS, and 2/3 were resistant to VOR. Among the 21 Af, 5% (1), 14% (3) and 19% (4) were non-susceptible to ITR, POS, and VOR respectively. 18 Af were available for sequencing, including all resistant isolates. 3 non-resistant Af isolates had a CYP51A 795A>G mutation; none of the resistant isolates had a CYP51A mutation.

Conclusion: Routine VOR prophylaxis post-lung transplantion was associated with very low rates of VOR resistant Aspergillus even among breakthrough isolates. ITR and POS resistance rates were higher than VOR. Azole resistance in Af was not mediated by CYP51A mutations. Discrepancies in azole resistance rates may reflect imprecise ITR and POS breakpoints. The lack of CYP51A mutations is consistent with other North American studies. It is possible that azole resistance in our isolates is mediated by mechanisms such as increased expression of efflux systems which are currently being investigated.