Multiple routes to fungicide resistance: Interaction of Cyp51 gene sequences, copy number and expression

Authors: ARNOLD, CORINNE - John Innes Center; MEYERS, EMILY - North Carolina State University; Whetten, Rebecca; CHARTRAIN, LAETITIA - John Innes Center; CHEEMA, JITENDER - John Innes Center; BROWN, JAMES - John Innes Center; Cowger, Christina

Submitted to: Molecular Plant Pathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/1/2024
Publication Date: 9/20/2024
Citation: Arnold, C., Meyers, E., Whetten, R.B., Chartrain, L., Cheema, J., Brown, J.K., Cowger, C. 2024. Multiple routes to fungicide resistance: Interaction of Cyp51 gene sequences, copy number and expression. Molecular Plant Pathology. https://doi.org/10.1111/mpp.13498.
DOI: https://doi.org/10.1111/mpp.13498

Interpretive Summary: Powdery mildew is an important disease of wheat and other small-grain cereals in many parts of the world. Samples of the powdery mildew fungus from wheat in the USA and UK were compared for sensitivity to an important class of fungicides used to control the disease, azoles. Resistance to two common triazoles, tebuconazole and prothioconazole, was found to be much higher in the UK than the USA. Four different pathways to resistance were identified: (1) a mutation in the fungal Cyp51 gene, to which fungicide molecules are intended to bind; (2) a mixture of mutated and wild-type (original) Cyp51 genes; (3) increased copy number of Cyp51; and (4) increased expression of Cyp51 by fungal cells. Different mechanisms may interact to increase resistance while maintaining fitness and survival of the fungus. When we assess resistance to azoles in mildew populations, we need to take into account the full range of mechanisms that could be providing an advantage to the fungus.

Technical Abstract: Blumeria graminis f.sp. tritici (wheat powdery mildew) isolates from the USA and UK were compared for sensitivity to the lanosterol demethylation inhibitor (DMI) azole fungicides tebuconazole and prothioconazole and assayed for Cyp51 sequences, copy number and expression. Median effective doses (ED50) of tebuconazole spanned a wider range than prothioconazole and US isolates had much greater mean sensitivity than UK isolates. Four genetic models involving multiple resistance mechanisms were identified. (1) The Y136F substitution was associated with resistance in both countries, combined with S509T in the most resistant UK isolates. No other CYP51 mutation was detected. In the US, F136 was most frequent where azoles have been most heavily used on wheat. (2) Some US isolates were heteroallelic for Y136 and F136 but this was not associated with higher resistance. In the UK, however, the most tebuconazole-resistant isolates were heteroallelic with wild-type Y136+S509 and mutant F136+T509 variants. (3) Increased copy number and expression of Cyp51 were closely correlated with each other and with resistance to both azoles in both homoallelic and heteroallelic isolates from both countries. (4) Some US isolates with wild-type CYP51 had elevated Cyp51 copy number and moderate prothioconazole resistance. Different mechanisms may interact to increase resistance while maintaining fitness; in particular, heteroallelism involving wild-type and mutant alleles may provide a selective advantage to powdery mildews if the wild-type protein catalyses lanosterol demethylation when azoles are absent but sequesters azole molecules in fungicide-treated leaves. Molecular field diagnostics should survey the full range of genetic mechanisms involved in azole resistance.