Activity and Resistance Assessment of a New OSBP Inhibitor, R034-1, in <i>Phytophthora capsici</i> and the Detection of Point Mutations in PcORP1 that Confer Resistance

Abstract

R034-1 is a new member of the piperidinyl thiazole isoxazoline class of fungicides that shows high activity against most plant-pathogenic oomycetes and could effectively inhibit several developmental stages of <i>Phytophthora capsici</i>. Here, the potential resistance risk for R034-1 was evaluated in <i>P. capsici</i>. The baseline sensitivities of 135 isolates to R034-1 showed a unimodal curve, with a mean EC₅₀ value of 0.004 μg/mL. Twelve resistant mutants were generated by fungicide adaptation and displayed lower fitness compared to parental isolates, which suggests that the resistance risk of <i>P. capsici</i> to R034-1 is low. R034-1 and oxathiapiprolin are structurally related, and resistant isolates display cross-resistance to both compounds, suggesting that these fungicides may target the same oxysterol binding protein. Comparison of <i>PcORP1</i> genes in the resistant mutants and their parental isolates revealed (N767S, N767I, and G700V) amino acid substitutions in the R034-1 resistant mutant. Causality was functionally validated using site-directed mutagenesis of the target gene using the CRISPR/Cas9 system.