Transient host-induced gene silencing targeting fungal sterol biosynthesis genes reduces Fusarium wilt severity in chrysanthemum

Fusarium wilt caused by Fusarium oxysporum is a major soil-borne disease affecting chrysanthemum production. Host-induced gene silencing (HIGS) provides a potential RNA interference-based strategy for controlling fungal pathogens; however, suitable fungal target genes for HIGS-mediated suppression of chrysanthemum Fusarium wilt remain poorly characterized. This study evaluated three fungal sterol-biosynthesis-related genes, CYP51A , CYP51C , and ERG6A , as candidate targets for transient HIGS in chrysanthemum. Synthetic small interfering RNAs targeting CYP51A , CYP51C , and ERG6A inhibited the growth of F. oxysporum in vitro. Artificial miRNA precursor constructs targeting the three genes were subsequently delivered transiently into chrysanthemum using the pCVA/pCVB-based transient-expression system. Across three independent transient-expression and pathogen-inoculation experiments, each comprising 20 plants per treatment, plants subjected to the three transient HIGS treatments exhibited reduced Fusarium wilt symptoms and lower disease severity indices than the WT/pCVA control. At 7 days post inoculation, the corresponding target-specific small RNAs accumulated in inoculated roots, and each construct preferentially reduced the abundance of its respective fungal target transcript. These changes were accompanied by lower root-associated ergosterol accumulation and reduced pathogen accumulation, as supported by qPCR-based fungal biomass analysis, conidial quantification, and microscopic observations. The transient HIGS treatments also showed reduced infection-associated oxidative damage compared with the WT/pCVA control. These findings provide proof-of-concept evidence that transient HIGS targeting fungal sterol-biosynthesis-related genes can reduce F. oxysporum accumulation and Fusarium wilt severity in chrysanthemum. The results support further evaluation of CYP51A , CYP51C , and ERG6A as candidate targets in stable or otherwise deployable RNAi systems.