多重环境因子影响下的紫菜黄斑病致病机理研究

Pyropia species were the most important cultivated seaweeds in China, the industries of which were greatly limited by the frequent diseases. Yellow-spot disease was one of the serious diseases occurred during the conchocelis breeding period of Pyropia yezoensis and Pyropia haitanensis. Because yellow-spot disease with strong infectiousness outbreaks at the very beginning of cultivation process and the nosogenesis is complicated, which might make radical damage to Pyropia cultivation. The tri-angle theory of “environment-pathogen-plant” is the principle strategy to study plant pathology. Temperature, pH, salinity, antagonistic bacteria and algae elicitor were considered as the multiple environmental factors in this research. The changes of biomass, metabolits of the pathogenic bacteria (Vibiro 117-T6) under different environmental conditions were studied, and the ability of infect conchocelis were evaluated meanwhile in order to reveal the mechanism of yellow-spot disease pathogenesis in Pyropia species. The physiological and pathological straits of both the free conchocelis and shell-born conchocelis of P. yezoensis and P. haitanensis were detected with the composite actions of environmental factors and the pathogens. Both the metabolomics and transcriptome data of Pyropia conchocelis were applied to reveal the key materials and the related genetic networks involved in the process of yellow-spot disease of Pyropia，which could constructs a metabolic network for the disease. This research systematically elaborate a comprehensive understanding of pathogenesis of the yellow-spot disease in Pyropia and the role of environmental factors in the course of disease, which could direct the prevention and control of the disease.

紫菜是我国重要的经济栽培海藻，病害频发是限制产业发展的重大问题。黄斑病是条斑紫菜和坛紫菜丝状体育苗阶段常见的重大疾病，病因复杂，传染性强，会对紫菜栽培造成根本性损失。基于“环境-病原菌-藻体”三者相互作用的综合考量，本项目通过观察在温度、pH、盐度、拮抗菌、藻类激发子等多重环境因子作用下，病原菌（弧菌117-T6）种群数量和代谢物的变化，探究环境对病原菌感染紫菜丝状体能力的影响，明确黄斑病的致病原因与机理；通过观察紫菜在不同环境条件和病原菌共同作用下的生理与病理反应，结合转录组和代谢组数据，明晰紫菜黄斑病的病理特点，探寻紫菜丝状体在黄斑病发病过程中的标志性代谢物质及其相关代谢网络基因表达的变化规律。本研究旨在系统阐述紫菜丝状体黄斑病的致病机理及环境因子在其中的作用，为该病的防治提供参考。

为系统阐述紫菜丝状体黄斑病的致病机理及环境因子在其中的作用，我们对罹患黄斑病的紫菜丝状体育苗体系的微生物进行了调查。基于16S高通量测序技术分析，明确黄斑病罹病和化学治疗均会破坏紫菜丝状体培养系统的环境及藻际微生物平衡，降低多样性并改变微生物区系组成，增加溶藻细菌并降低有益拮抗菌丰度，从而加大养殖风险。经科赫法则证实地中海弧菌Vibrio mediterranei 117-T6（Vm 117-T6）是紫菜丝状体黄斑病的病原菌，其菌体、胞内和胞外物均可导致紫菜丝状体和叶状体染病。由Vm 117-T6感染导致的紫菜黄斑病病状表现为四个阶段：前期，菌体聚集黏附； I期，细胞器膨大，出现包含体样结构，细胞边缘开始模糊；II期：丝状体内膜系统裂解，胞内物外流，胞内结构疏松；III期：丝状体内容物进一步流失，细胞空胞化，最终细胞壁降解。染病后，藻体短期内调动了抗氧化酶活性以清除氧自由基，提高光合色素以及可溶性蛋白、和游离脯氨酸等渗透调节物的合成，但随着胁迫激烈或时间延长，藻体的生理机能明显下降，藻体防御及修复能力难以维系，快速病烂。Vm 117-T6是一株噬盐菌，低盐会抑制其生长，而高温酸化会提高菌株的毒力。Vm 117-T6的溶藻机制多样且复杂，涉及与细菌黏附、运动、跨膜运输、毒力分泌以及群感效应相关的关键通路，而且吲哚丙烯酸是重要的差异标志物。建立基于“环境-藻体-病原菌”的紫菜黄斑病的综合防控技术体系，控制养殖环境，避免高温和酸化条件出现，利用噬菌体益生菌制剂和琼胶寡糖强化藻体具有良好防护效果。本研究建立了紫菜丝状体黄斑病的原学及病理学研究方法，并为紫菜黄斑病的防治提供了理论参考和实践指导。