Triazole Fungicides Can Induce Cross-Resistance to Medical Triazoles in Aspergillus fumigatus

Abstract

BackgroundAzoles play an important role in the management of Aspergillus diseases. Azole resistance is an emerging global problem in Aspergillus fumigatus, and may develop through patient therapy. In addition, an environmental route of resistance development has been suggested through exposure to 14α-demethylase inhibitors (DMIs). The main resistance mechanism associated with this putative fungicide-driven route is a combination of alterations in the Cyp51A-gene (TR34/L98H). We investigated if TR34/L98H could have developed through exposure to DMIs.Methods and FindingsThirty-one compounds that have been authorized for use as fungicides, herbicides, herbicide safeners and plant growth regulators in the Netherlands between 1970 and 2005, were investigated for cross-resistance to medical triazoles. Furthermore, CYP51-protein homology modeling and molecule alignment studies were performed to identify similarity in molecule structure and docking modes. Five triazole DMIs, propiconazole, bromuconazole, tebuconazole, epoxiconazole and difenoconazole, showed very similar molecule structures to the medical triazoles and adopted similar poses while docking the protein. These DMIs also showed the greatest cross-resistance and, importantly, were authorized for use between 1990 and 1996, directly preceding the recovery of the first clinical TR34/L98H isolate in 1998. Through microsatellite genotyping of TR34/L98H isolates we were able to calculate that the first isolate would have arisen in 1997, confirming the results of the abovementioned experiments. Finally, we performed induction experiments to investigate if TR34/L98H could be induced under laboratory conditions. One isolate evolved from two copies of the tandem repeat to three, indicating that fungicide pressure can indeed result in these genomic changes.ConclusionsOur findings support a fungicide-driven route of TR34/L98H development in A. fumigatus. Similar molecule structure characteristics of five triazole DMIs and the three medical triazoles appear the underlying mechanism of cross resistance development. Our findings have major implications for the assessment of health risks associated with the use of triazole DMIs.

Molecular Epidemiology of Aspergillus fumigatus Isolates Harboring the TR 34 /L98H Azole Resistance Mechanism

“…These observations suggest that azole-resistant Aspergillus is acquired by patients from an environmental source rather than arising through azole therapy. Recently, we provided evidence that exposure of A. fumigatus to 14-␣-demethylase inhibitor (DMI) fungicides might provide a selective pressure leading to the emergence of TR 34 /L98H resistant isolates in the environment (27). On the basis of in vitro crossresistance, molecule alignment studies, and docking simulations, five triazole fungicides that were highly similar to antifungal triazoles used in medicine were identified (27).…”

“…Recently, we provided evidence that exposure of A. fumigatus to 14-␣-demethylase inhibitor (DMI) fungicides might provide a selective pressure leading to the emergence of TR 34 /L98H resistant isolates in the environment (27). On the basis of in vitro crossresistance, molecule alignment studies, and docking simulations, five triazole fungicides that were highly similar to antifungal triazoles used in medicine were identified (27). The TR 34 /L98H resistance mechanism has been shown to be endemic in The Netherlands (36) and is also increasingly being reported in other European countries (11,22,24,30,34).…”

Azole-Resistant Aspergillus fumigatus Isolate with the TR34/L98H Mutation in Both a Fungicide-Sprayed Field and the Lung of a Hematopoietic Stem Cell Transplant Recipient with Invasive Aspergillosis

“…However, in recent years, azole resistance in Aspergillus fumigatus has been increasingly reported in probable association with azole failure (2,3). In the Netherlands, the main mechanism suggested in hematology is the acquisition of resistant isolates from the environment due to the increasingly extensive use of 14-alpha-demethylase inhibitor fungicides in agriculture (4). The presence of the TR34/L98H azole resistance mechanism has been increasingly reported in clinical and environmental A. fumigatus isolates throughout Europe (Netherlands, United Kingdom, Norway, Spain, Denmark, Belgium, France, Germany) and Asia (India, China, Iran) (see reference 5 and references therein).…”

Mechanisms of Antifungal Drug Resistance

“…To date, all known mutations confer resistance to itraconazole, whereas resistance to voriconazole and/or posaconazole depend on the specific modification (Verweij et al , 2009Howard and Arendrup 2011;van der Linden et al 2011;Camps et al 2012b). Resistance has also been identified in azole-naïve patients, which is environmentally derived and appears to be driven by the agricultural use of azoles (Snelders et al 2012;Bowyer and Denning 2013;Verweij et al 2013). These cases involved a Cyp51A substitution at position 98 (from leucine to histidine), and a 34-base tandem repeat (TR) in the cyp51A promoter, which leads to overexpression (Snelders et al 2008).…”