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| 烟草未知叶斑病叶际真菌分离鉴定及其叶际真菌碳代谢特征 |
| Isolation and Identification of Phyllosphere Fungi from Tobacco Leaves with an Unknown Spot Disease and Their Carbon Metabolism Characterization |
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| DOI：doi:10.3969/j.issn.1005-7021.2026.03.004 |
| 中文关键词: 扩增子测序 间座壳属（Diaporthe） 致病力 真菌群落 碳源代谢 |
| 英文关键词: amplicon sequencing Diaporthe pathogenicity fungal community carbon source metabolism |
| 基金项目:国家自然科学基金项目（32160522，32460698）；贵州省“百层次”创新型人才项目（黔科合平台人才-GCC［2022］028-1）；贵州省科技创新人才团队项目（黔科合平台人才-CXTD［2023］021）；贵州省科技创新人才团队项目（黔科合平台人才-CXTD［2023］021)；中国烟草总公司贵州省公司科技项目（2024XM06） |
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| 摘要点击次数: 44 |
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| 中文摘要: |
| 为诊断贵州省遵义市道真县烟田的一未知病害，了解其叶际微生态特征，采用组织分离法对其叶际真菌进行分离纯化及分子生物学鉴定，依据柯赫氏法则验证致病菌，采用扩增子测序技术对其叶际微生物群落组成与多样性进行分析，采用Biolog ECO微平板法研究其叶际微生物的代谢活性。结果表明，使用组织分离法从感病烟叶组织上共分离到10个菌属的24株真菌，致病力验证结果表明间座壳属（Diaporthe）真菌具有致病性，基于ITS、HIS和TEF1-α基因序列构建联合系统发育树并结合形态学鉴定结果，揭示引起该烟草叶斑病的病原为榆树间座壳菌（Diaporthe eres）。扩增子测序结果显示，健康和感病烟叶叶际优势真菌属均为布勒酵母属（Bullera）（相对丰度分别为22.89%、10.33%）和纖柔菌属（Abrothallus）（44.01%、41.21%）。感病烟叶与健康烟叶真菌群落的多样性指数无显著差异，感病烟叶的丰富度指数低于健康烟叶。功能预测结果表明，感病烟叶组织中的植物病原菌类群相对丰度高于健康组织。碳源代谢发现，健康烟叶叶际微生物碳源代谢能力强于感病烟叶，健康烟叶叶际微生物对28种碳源具有较强的代谢能力；感病烟叶叶际微生物对12种碳源有较高的代谢能力。该烟草未知叶斑病由榆树间座壳菌引起，叶际微生物还包含链格孢属、织球壳属、短梗霉属、棒孢属等致病菌，结果揭示了该未知烟草叶斑病的叶际真菌微生态特征。 |
| 英文摘要: |
| To diagnose an unidentified disease affecting tobacco fields at Daozhen County in Zunyi, and to investigate the phyllosphere microecological characteristics associated with the disease, a comprehensive study was conducted. Phyllosphere fungi were isolated and purified using tissue isolation methods, and identified by molecular biology techniques. The pathogenicity of these phyllosphere fungi was assessed based on Koch′s postulates. Amplicon sequencing was employed to analyze the microbial community structure and diversity, while Biolog ECO microplate assays were used to evaluate the metabolic activity of the phyllosphere microorganisms. A total of 24 fungal isolates belonging to 10 genera were obtained from symptomatic tobacco leaf tissues through tissue isolation. Pathogenicity tests demonstrated that fungi of the genus Diaporthe were pathogenic. Phylogenetic analysis based on ITS, HIS and TEF1-α gene sequences combined with morphological identification revealed that the pathogen responsible for the tobacco leaf spot disease was identified as Diaporthe eres. Amplicon sequencing results indicated that the dominant fungal genera in both healthy and diseased tobacco phyllosphere samples were Bullera (22.89% in healthy and 10.33% in diseased samples), Abrothallus (44.01% and 41.21%). There was no significant difference in fungal diversity index between healthy and diseased tobacco leaves; however, the richness index was lower in diseased samples. Functional prediction suggested that the relative abundance of plant pathogenic fungi was higher in diseased tobacco leaf tissues compared to healthy ones. Carbon substrate utilization assays revealed that the metabolic activity of phyllosphere microorganisms from healthy tobacco leaves was stronger than that of those from diseased leaves. Microorganisms from healthy leaves exhibited high metabolic capability toward 28 carbon sources. In contrast, phyllosphere microorganisms from diseased leaves showed relatively higher metabolic activity toward 12 carbon sources. In conclusion, the previously unknown tobacco leaf spot disease was identified and determined to be caused by Diaporthe eres. Additionally, other pathogenic genera such as Alternaria, Abrothallus, Aspergillus,and Colletotrichumwere also present in the disease-associated phyllosphere. These findings provide insight into the fungal microecological characteristics of the phyllosphere associated with the what so called unknown tobacco leaf spot disease. |
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