ABSTRACT
Fungicide resistance is a constant threat to agricultural production worldwide. Molecular mechanisms of fungicide resistance have been studied extensively in the wheat pathogen Zymoseptoria tritici. However, less is known about the evolutionary processes driving resistance development. In vitro evolutionary studies give the opportunity to investigate this. Here, we examine the adaptation of Z. tritici to fluxapyroxad, a succinate dehydrogenase (Sdh) inhibitor. Replicate populations of Z. tritici derived from the sensitive isolate IPO323 were exposed to increasing concentrations of fluxapyroxad with or without UV mutagenesis. After ten increases in fungicide concentration, sensitivity had decreased dramatically, with replicate populations showing similar phenotypic trajectories. Sequencing the Sdh subunit B, C, and D encoding genes identified seven mutations associated with resistance to fluxapyroxad. Mutation frequency over time was measured with a pyrosequencing assay, revealing sequential lineage replacement in the UV-mutagenized populations but not in the untreated populations. Repeating selection from set time-points with different fungicide concentrations revealed that haplotype replacement of Sdh variants was driven by dose-dependent selection as fungicide concentration changed, and was not mutation-limited. These findings suggest that fungicide field applications may select for highly insensitive Sdh variants with higher resistance factors if the fungicide concentration is increased to achieve a better disease control. However, in the absence or presence of lower fungicide concentrations, the spread of these strains might be restricted if the underlying Sdh mutations carry fitness penalties.
