1. Insights in the molecular mechanisms of an azole stress adapted laboratory-generated Aspergillus fumigatus strain
- Author
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Patricia Flamant, Isabelle Mouyna, Samantha Gozel, Alix T. Coste, Dominique Sanglard, Frédéric Lamoth, Josie E. Parker, Daniel Bachmann, Marion Aruanno, Lausanne University Hospital, Université de Lausanne = University of Lausanne (UNIL), Aspergillus, Institut Pasteur [Paris] (IP), Swansea University, and This work was supported by the Swiss National Science Foundation (SNSF Ambizione-Score, grant number: PZ00P3 161140) and by the Santos-Suarez foundation.
- Subjects
Azoles ,Antifungal Agents ,ATP-binding cassette transporter ,Aspergillus fumigatus ,chemistry.chemical_compound ,MESH: HMGB1 Protein ,Cytochrome P-450 Enzyme System ,MESH: Reverse Transcriptase Polymerase Chain Reaction ,MESH: Azoles ,MESH: Ku Autoantigen ,HMGB1 Protein ,[SDV.MP.MYC]Life Sciences [q-bio]/Microbiology and Parasitology/Mycology ,chemistry.chemical_classification ,0303 health sciences ,Ergosterol ,biology ,MESH: Real-Time Polymerase Chain Reaction ,Reverse Transcriptase Polymerase Chain Reaction ,MESH: Voriconazole ,General Medicine ,isavuconazole milbemycine ,Sterols ,Infectious Diseases ,MESH: CCAAT-Binding Factor ,MESH: Cytochrome P-450 Enzyme System ,MESH: ATP-Binding Cassette Transporters ,MESH: Fungal Proteins ,Efflux ,MESH: Aspergillus fumigatus ,efflux pumps ,transporters ,Real-Time Polymerase Chain Reaction ,Gas Chromatography-Mass Spectrometry ,MESH: Drug Resistance, Fungal ,Microbiology ,resistance ,Fungal Proteins ,03 medical and health sciences ,Drug Resistance, Fungal ,Ku Autoantigen ,Transcription factor ,030304 developmental biology ,030306 microbiology ,MESH: Transcriptome ,Cell Membrane ,Transporter ,MESH: Antifungal Agents ,biology.organism_classification ,posaconazole ,Major facilitator superfamily ,MESH: Gas Chromatography-Mass Spectrometry ,CCAAT-Binding Factor ,chemistry ,MESH: Sterols ,Azole ,ATP-Binding Cassette Transporters ,Voriconazole ,erogsterol ,Transcriptome ,MESH: Cell Membrane - Abstract
Aspergillus fumigatus is the main cause of invasive aspergillosis, for which azole drugs are the first-line therapy. Emergence of pan-azole resistance among A. fumigatus is concerning and has been mainly attributed to mutations in the target gene (cyp51A). However, azole resistance may also result from other mutations (hmg1, hapE) or other adaptive mechanisms. We performed microevolution experiment exposing an A. fumigatus azole-susceptible strain (Ku80) to sub-minimal inhibitory concentration of voriconazole to analyze emergence of azole resistance. We obtained a strain with pan-azole resistance (Ku80R), which was partially reversible after drug relief, and without mutations in cyp51A, hmg1, and hapE. Transcriptomic analyses revealed overexpression of the transcription factor asg1, several ATP-binding cassette (ABC) and major facilitator superfamily transporters and genes of the ergosterol biosynthesis pathway in Ku80R. Sterol analysis showed a significant decrease of the ergosterol mass under voriconazole exposure in Ku80, but not in Ku80R. However, the proportion of the sterol compounds was similar between both strains. To further assess the role of transporters, we used the ABC transporter inhibitor milbemycine oxime (MLB). MLB inhibited transporter activity in both Ku80 and Ku80R and demonstrated some potentiating effect on azole activity. Criteria for synergism were reached for MLB and posaconazole against Ku80. Finally, deletion of asg1 revealed some role of this transcription factor in controlling drug transporter expression, but had no impact on azole susceptibility. This work provides further insight in mechanisms of azole stress adaptation and suggests that drug transporters inhibition may represent a novel therapeutic target. Lay Summary A pan-azole-resistant strain was generated in vitro, in which drug transporter overexpression was a major trait. Analyses suggested a role of the transporter inhibitor milbemycin oxime in inhibiting drug transporters and potentiating azole activity.
- Published
- 2021