Target-site mutations in succinate dehydrogenase subunits of <i>Venturia effusa</i> associated with reduced sensitivity to pydiflumetofen during a pecan scab outbreak in Georgia

Venturia effusa, the causal agent of pecan scab, drives the spray program in Georgia's pecan production. Miravis Top (difenoconazole, FRAC 3 + pydiflumetofen, FRAC 7) has been widely adopted for pecan scab management in recent years due to the high efficacy of its SDHI component, pydiflumetofen. In late summer 2025, a severe outbreak occurred in a commercial orchard in South Georgia despite a well-maintained spray program that included Miravis Top. While resistance to demethylation inhibitor (DMI) fungicides and other fungicide classes in V. effusa has been well documented, resistance to pydiflumetofen has not been reported. Symptomatic leaves and nuts were collected from cultivars 'Desirable' and 'Pawnee' located on a commercial farm in Georgia. Thirty-five single-spore isolates were obtained and compared with a historic baseline population (n = 12; 1993-1994) presumed unexposed to SDHIs. In mycelial growth assays, baseline isolates showed a mean EC₅₀ of 0.012 µg/ml pydiflumetofen, whereas values from outbreak isolates ranged from reduced sensitive (0.047-0.091 µg/ml; n = 7) to moderate resistant (0.281-0.895 µg/ml; n = 10) and high resistant (1.17-6.70 µg/ml; n = 18), with resistance factors >500. Sequencing of VesdhB, VesdhC, and VesdhD genes identified six amino acid (aa) substitutions associated with reduced sensitivity: H252R (VeSdhB), K73R; M82I; N84S (VeSdhC), and D150E and D150G (VeSdhD). All moderate to high resistant isolates carried at least one substitution, except for a subset of moderate resistant isolates that exhibited the lowest EC₅₀ values within this group. The most resistant isolate (Ve25-22) carried two mutations, K73R (VeSdhC) and D150G (VeSdhD). Cross-sensitivity assays with fluopyram, isofetamid, and benzovindiflupyr showed incomplete cross-resistance with pydiflumetofen (moderate correlations), but a strong correlation between fluopyram and isofetamid (r = 0.998, P < 0.001). Notably, isolates carrying the M82I substitution in VeSdhC were high resistant to both fluopyram and isofetamid. This study provides the first evidence of reduced sensitivity to pydiflumetofen in V. effusa linked to target-site substitutions in the VeSdhB, VeSdhC, and VeSdhD subunits and highlights the importance of resistance management strategies, including monitoring pathogen populations, rotating fungicide modes of action, using fungicide mixtures, and limiting the number of applications.