The resistance risk of fluopicolide and resistance-associated point mutations in the target protein PlVHA-a in Phytophthora litchii

Litchi, a fruit that is highly sought-after worldwide, faces significant yield challenges due to litchi downy blight, primarily caused by Phytophthora litchii. Fluopicolide has exhibited remarkable efficacy in inhibiting this pathogen and is utilized for the management of litchi downy blight. Although understanding the resistance of P. litchii to fluopicolide is critical, studies on its risk and mechanisms remain limited. In this study, we determined the sensitivity of 125 P. litchii isolates to fluopicolide, revealing an average EC₅₀ value of 0.131 ± 0.037 μg/mL. Through fungicide adaptation, four resistant mutants were obtained with resistance factors exceeding 600, indicating that these strains exhibited high levels of resistance. A compound fitness index analysis demonstrated that the survival fitness of resistant mutants was significantly lower than that of their parental strains. Cross-resistance assays revealed no cross-resistance between fluopicolide and other fungicides with different modes of action. However, positive cross-resistance was observed with fluopimomide. A comprehensive evaluation suggested a moderate risk of P. litchii developing resistance to fluopicolide. PlVHA-a^N771S and PlVHA-a^N846S point mutations in resistant mutants were identified by gene sequencing analyses. These two point mutations were validated as contributors to resistance in P. litchii through genetic transformation and molecular docking.