Abstract
Fusarium oxysporum is a widespread soil-borne fungal pathogen that can infect various plants, causing wilt and root rot diseases. The root rot disease of Atractylodes macrocephala caused by F. oxysporum is among the most serious diseases associated with continuous cropping, significantly hindering its sustainable development. In this study, we aimed to investigate the effect of exogenous application of double-stranded RNA (dsRNA) on silencing the F. oxysporum Tup1 gene to reduce its virulence and to evaluate its potential application in controlling root rot disease in A. macrocephala. The Tup1 gene was amplified from the F. oxysporum genome, and different lengths of Tup1-dsRNA were designed and synthesized. The uptake of dsRNA by the fungus was verified using Tup1-dsRNA labeled with fluorescein, and in vitro dsRNA treatment experiments were conducted to assess its impact on the growth and virulence of F. oxysporum. Additionally, Tup1-dsRNA was applied to the roots of A. macrocephala to evaluate its effectiveness in controlling root rot disease. The experimental results showed that F. oxysporum could effectively uptake exogenously applied Tup1-dsRNA, significantly reducing Tup1 gene expression. All lengths of Tup1-dsRNA inhibited fungal growth and caused morphological changes in the fungal hyphae. Further plant experiments and Reverse Transcription Quantitative Polymerase Chain Reaction (RT-qPCR) analysis indicated that Tup1-dsRNA treatment significantly reduced the incidence of root rot disease in A. macrocephala, which was supported by the reduction in peroxidase (POD) and catalase (CAT) enzyme activities, malondialdehyde (MDA) content, and proline (Pro) levels in treated root tissues. This study demonstrated that exogenous dsRNA could reduce the virulence of F. oxysporum by silencing the Tup1 gene and effectively mitigate the root rot disease it causes in A. macrocephala. The successful application of Tup1-dsRNA provided strong evidence for the potential of RNA interference (RNAi) technology in plant disease control. Future research could further optimize the design and application of dsRNA to enhance its practical value in agriculture.