Multi-fungicide resistance in both clinical and environmental isolates of the human fungal pathogen <i>Aspergillus fumigatus</i>

Aspergillus fumigatus is a human pathogen and a ubiquitous environmental mould. Consequently, A. fumigatus may be exposed to agricultural fungicides widely used for crop protection. The use of azole fungicides in the environment has been implicated in the emergence of azole antifungal resistance in A. fumigatus. In vitro susceptibility of a collection of both clinical (n = 14) and environmental (n = 6) A. fumigatus isolates was determined against fungicides with different modes of action (demethylation inhibitors, Succinate DeHydrogenase Inhibitor (SDHI), Quinone outside Inhibitors (QoI), Methyl Benzimidazole Carbamate (MBC) and polyene). Sequencing of tubA, CYTB and SDHB, which encode the cellular targets of these fungicides, was performed. Eventually, the impact of resistance on the basal growth kinetics was assessed using a selection of two susceptible and two resistant isolates. Cross-resistance between medical triazole antifungal and agricultural triazole fungicides was confirmed, except for isolates with G54R/E substitutions in the cyp51A protein. No correlation was observed between resistance to triazole antifungals and imidazole fungicides. We identified seven MBC- (five environmental and two clinical isolates), five QoI- (three environmental and two clinical isolates) and one environmental SDHI-resistant isolates. Resistant phenotypes were associated with amino acid substitutions in beta-tubulin (F219Y and E217A), cytB (G143A) and sdhB (H270Y). Multi-fungicide resistance was not systematically associated with reduced growth kinetic. Multi-fungicide resistance, including at least resistance to one non-azole fungicide, was identified in sevenA. fumigatus isolates, of which two were clinical isolates. This last observation supports the hypothesis of the environmental pathway leading to antifungal resistance in A. fumigatus.