Use of RNAi-based protection strategy to control stemphylium vesicarium on pear: insight into the molecular mechanisms and silencing effect on target and host

Brown spot disease, caused by the fungal pathogen Stemphylium vesicarium, poses a major threat to European pear production. This study aimed to systematically elucidate the multilayered regulatory effects of exogenous dsRNA on the pear–pathogen interaction, and to explore the host specificity of S. vesicarium and the potential of RNAi–based control strategies. Transcriptomic analysis following treatment with dsRNA-SDHD revealed that, although this treatment did not significantly suppress lesion formation, it pre-induced stress responses in S. vesicarium and persistently inhibited ribosome biogenesis and metabolic activity during the late infection stage, thereby interfering with its normal infection process. On the host side, dsRNA pretreatment activated pear defense responses, including pathogen recognition, hormone signaling, and defense-related gene expression, while enhancing photosynthesis and energy metabolism to establish molecular preparedness for resistance. Moreover, RNAi-related pathways were significantly enriched in pear leaves at 24 hours post-inoculation, highlighting the transcriptional regulation of host–pathogen interactions by exogenous dsRNA. Meanwhile, we systematically analyzed 47 S. vesicarium isolates from pear and other host plants. Pathogenicity assays confirmed host specificity, as only pear-derived isolates caused severe lesions on pear leaves. WGS and phylogenetic analyses further supported this finding, showing that isolates generally clustered according to their host of origin. Orthogroup analysis identified six pear host–specific genes and eleven additional candidate genes potentially involved in host adaptation. These genes may participate in pathogenicity, nutrient acquisition, and metabolic regulation, representing promising molecular targets for brown spot disease management. Based on these results, eight host-specific genes were selected for dsRNA synthesis and evaluated under two application regimes. Among them, dsRNAs targeting HP and FRT reduced lesion development. Collectively, these findings demonstrate that dsRNA can interfere with host–pathogen interactions at multiple stages and provide a conceptual and practical framework for developing environmentally friendly RNAi-based control strategies against pear brown spot disease.