关键词: 贝莱斯芽孢杆菌, 桃褐腐病, 实生链核盘菌, 生物防治, 作用机制

Abstract:

【Objective】Peach brown rot, caused by Monilinia spp., is a severe fungal disease leading to substantial economic losses to the peach industry. This study aims to evaluate the biocontrol potential of Bacillus velezensis strain C1-17 against peach brown rot and to elucidate its antifungal mechanisms, thereby providing a novel strain resource and theoretical basis for green management of this disease. 【Method】A previously obtained B. velezensis C1-17 strain was used, the in vitro antifungal activity and in vivo biocontrol efficacy of C1-17 against M. fructicola, the dominant pathogen of peach brown rot, were determined by dual-culture plate confrontation assay, fruit immersion with bacterial suspension, and cell-free culture filtrate assay, respectively, to evaluate its biocontrol potential. Morphological and ultrastructural changes in the pathogen hyphae after treatment were investigated by using light microscopy, scanning electron microscopy, and transmission electron microscopy. Plasma membrane integrity and intracellular reactive oxygen species (ROS) levels of the pathogen were detected using propidium iodide (PI) staining and the DCFH-DA fluorescent probe, respectively. The ability of C1-17 to produce extracellular antimicrobial active compounds, including proteases, cellulases, and siderophores, was assessed using specific culture media. Expression changes of defense-related genes (PpPR1, PpNPR1, PpMYC2, PpJAZ5, PpPAL1, PpPOD) in peach fruit following C1-17 treatment and under pathogen infection were analyzed by reverse-transcription quantitative real-time PCR (RT-qPCR). Peach fruits were sprayed with 10% C1-17 bacterial suspension and tebuconazole, respectively, and then inoculated with spore suspensions of DMI-resistant and DMI-sensitive strains. Disease incidence was investigated and control efficacy was calculated to evaluate the control efficacy of C1-17 against different resistant strains. 【Result】The fermented broth of C1-17 showed a control efficacy of 93.25% against peach brown rot at a low concentration of 2% (V/V). Its cell-free culture filtrate at 1% (V/V) inhibited mycelial growth of M. fructicola by 95.93% and reduced the conidial germination rate to 3%. After treatment with 1% (V/V) C1-17 cell-free culture filtrate, pathogen hyphae exhibited swelling, vacuolation, wrinkling and disruption, plasmolysis, and dissolution of organelles such as mitochondria. PI staining and DCFH-DA detection confirmed loss of plasma membrane integrity and massive accumulation of intracellular ROS, suggesting that membrane damage and oxidative burst are important antifungal mechanisms. C1-17 also produced extracellular active substances such as proteases, cellulases, and siderophores. Spraying C1-17 alone did not significantly induce the expression of peach defense-related genes. However, under M. fructicola infection, C1-17 pretreatment specifically up-regulated the expression of salicylic acid pathway-related genes (PpPAL1 and PpPR1) and PpPOD at the late stage of infection (48 h). The 10% C1-17 suspension exhibited good control efficacy (≥94.50%) against both DMI-resistant and DMI-sensitive strains of M. fructicola. Notably, its efficacy against DMI-resistant strains was superior to that of 20 μg·mL^-1 tebuconazole, and no cross-resistance with DMI fungicides was observed. 【Conclusion】B. velezensis C1-17 shows promising biocontrol potential against peach brown rot. By secreting antifungal substances, C1-17 directly disrupts hyphal morphology and plasma membrane integrity of M. fructicola and induces intracellular ROS accumulation. In addition, at the late stage of pathogen infection, it selectively activates the salicylic acid signaling pathway and the phenylpropanoid metabolic pathway, thereby achieving efficient control of peach brown rot.

Key words: Bacillus velezensis, peach brown rot, Monilinia fructicola, biological control, mechanism of action