Multiple Single-Point Mutations in Stemphylium vesicarium Are Associated with the Rapid Development of Resistance to...

Succinate dehydrogenase inhibitor (SDHI) fungicides are used to manage Stemphylium leaf blight (SLB) of onion caused by the fungus Stemphylium vesicarium. The SDHIs commonly used for SLB management in New York (NY) onion production are boscalid (first registered in 2005), fluxapyroxad (2015), fluopyram (2016), and pydiflumetofen (2019). However, reduced field performance of these products across multiple onion-producing regions within NY has been encountered. We quantified the in vitro sensitivity of S. vesicarium isolates collected from five onion-producing regions throughout NY in 2016, 2018, and 2020. To evaluate whether variations in in vitro sensitivity phenotypes were associated with target-site mutations, sequencing of the sdhB, sdhC, and sdhD genes associated with fungicide response was conducted. We identified a shift in sensitivity over a short period, that is, although more than 90% of isolates sampled in 2016 were sensitive to fluopyram and fluxapyroxad (EC₅₀ < 1 mg/liter), more than 50% of isolates sampled in 2018 exhibited reduced sensitivity (EC₅₀ > 1 mg/liter). This change in fungicide sensitivity was observed in three of the four main onion-producing regions of NY and emphasizes the need for improved disease management practices to preserve their efficacy. Primers were developed to sequence the full sdh genes of 176 isolates from all regions and years sampled. Thirteen single-nucleotide polymorphisms were identified in the sdhB, sdhC, and sdhD genes, and 11 were found to predict nonsynonymous amino acid (aa) substitutions. The isolates with genotypes P230H (SDHB) and G79R, H134N/R, and C135R (SDHC) were associated with the reduced sensitivity of S. vesicarium to fluopyram and fluxapyroxad. These putative aa substitutions were not associated with effects on mycelial growth at one temperature. Spatiotemporal analyses revealed a clear shift in population structure from wild-type populations in 2016 to diverse genotypes with multiple substitutions across onion-producing regions by 2020. The rapid, diverse, and widespread distribution of genotypes with putative aa substitutions suggests an ongoing adaptation and the presence of strong selective forces in S. vesicarium in NY.