短小芽孢杆菌H-46通过诱导系统抗性增强猕猴桃抗病性

Induction of systemic resistance by Bacillus subtilis H-46 enhances disease control in kiwifruit

摘要:

目的 丁香假单胞菌猕猴桃致病变种(Pseudomonas syringae pv. actinidiae, Psa)引起的猕猴桃细菌性溃疡病(kiwifruit bacterial canker, KBC)已成为我国猕猴桃产业可持续发展的首要瓶颈，亟需绿色、无残留的生物防治新策略。方法 以‘红阳’猕猴桃为试验材料，通过叶盘法和枝条接种法系统评价短小芽孢杆菌(Bacillus pumilus) H-46的生防效果。采用分级提取法获得该菌株的3类活性组分(小分子代谢物、蛋白质类和多糖类物质)，并通过抑菌活性和病害防治效果评估确定其主要活性成分。综合运用组织化学染色、抗氧化酶活性测定及防御相关基因RT-qPCR分析等技术解析其活性物质诱导系统抗性(induced systemic resistance, ISR)的作用机理。此外，通过病原菌迁移和定殖实验评估活性成分对病原菌Psa的抑制效果。结果 短小芽孢杆菌H-46对KBC具有良好的防治效果，叶盘防效可达86.54%，其主效活性组分——小分子代谢物(A)通过非抑菌作用对猕猴桃溃疡病的防治效果可达88.16%。作用机理表现为：触发早期H₂O₂与胼胝质沉积，显著提升超氧化物歧化酶(superoxide dismutase, SOD)、过氧化物酶(peroxidase, POD)和过氧化氢酶(catalase, CAT)活性，并激活茉莉酸(jasmonicacid, JA)与乙烯(ethylene, ET)信号通路关键基因(AcMYC2、AcAOC、AcERF2、AcEIN3)的表达诱导植物系统抗性(ISR)，进一步增强猕猴桃抗病性，使Psa在叶脉中的迁移距离缩短37.8%，定殖量降低96.6%。结论 短小芽孢杆菌H-46小分子代谢物(A)通过非抑菌途径激活JA/ET依赖的ISR网络，显著提升猕猴桃对KBC的抗性，为开发基于植物免疫诱抗的绿色农药提供了理论基础和实践依据。

Abstract:

Objective Kiwifruit bacterial canker (KBC), caused by Pseudomonas syringae pv. actinidiae (Psa), has become the primary bottleneck restricting the sustainable development of the kiwifruit industry in China, highlighting an urgent need for eco-friendly and residue-free biocontrol strategies.Methods The kiwifruit variety ‘Hongyang’ was used to systematically evaluate the biocontrol efficacy of Bacillus pumilus H-46 through leaf disc and shoot inoculation assays. The active components were fractionated into three groups (small-molecule metabolites, proteins, and polysaccharides) via sequential extraction, with the major bioactive fraction identified through antimicrobial activity and disease control assessments. An integrated approach combining histochemical staining, antioxidant enzyme activity assays, and RT-qPCR of defense-related genes was employed to elucidate the mechanism of induced systemic resistance (ISR). Furthermore, pathogen migration and colonization assays were conducted to evaluate the inhibitory effects of the active components against Psa.Results B. pumilus H-46 showed excellent control effect against KBC, with the disease control efficacy of 86.54% in leaf disc assays. Its main active components (small-molecule metabolites, fraction A) achieved the control efficacy of 88.16% against KBC through non-antimicrobial mechanisms. The mechanisms included triggering early H₂O₂ accumulation and callose deposition, significantly increasing superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) activities, and activating the expression of key genes (AcMYC2, AcAOC, AcERF2, and AcEIN3) in jasmonicacid (JA) and ethylene (ET) signaling pathways to activate ISR. Consequently, the disease resistance of kiwifruit was enhanced, resulting in a reduction of 37.8% in Psa migration distance in leaf veins and a decrease of 96.6% in colonization ability.Conclusion Our findings demonstrate that the small-molecule metabolites (fraction A) of B. pumilus H-46 activates JA/ET-mediated ISR via non-antimicrobial mechanisms, offering a sustainable solution for the control against KBC and establishing a prototype for next-generation plant immunity activators in crop protection.