焦脱镁叶绿酸a对黄瓜枯萎病菌的抑制作用

doi:10.16420/j.issn.0513-353x.2022-1113

摘要/Abstract

摘要：

为了探究焦脱镁叶绿酸a（PPa）对植物病原菌的抑制作用，以黄瓜枯萎病病原菌尖孢镰刀菌黄瓜专化型菌（Fusarium oxysporium f. sp. cucumerinum）H₄₆ 5菌和‘长春密刺’黄瓜为材料，测定不同浓度PPa对菌丝生长的影响，以及经不同强度和时长光照的PPa对感枯萎病的黄瓜叶片的病斑面积，丙二醛和游离脯氨酸含量，过氧化物酶、过氧化氢酶和超氧化物歧化酶活性及活性氧的影响。结果表明：PPa对H₄₆ 5菌株生长有明显抑制作用，其抑菌效果与浓度呈正相关，浓度为2 g · L^-1效果最佳。PPa具有光敏性，其经适当强度和时长的光照后喷施感染枯萎病的黄瓜叶片，可显著降低叶片感病程度与MDA含量，提升脯氨酸水平与抗氧化能力，维持活性氧平衡。上述结果表明，PPa具有很好的防治黄瓜枯萎病效果。

关键词: 黄瓜, 枯萎病, 焦脱镁叶绿酸a, 活性氧, 抗病

Abstract:

In order to explore the inhibitory effect of pyropheophorbide-a（PPa）on plant pathogens，and to find a new agent with low toxicity and high efficiency against the cucumber Fusarium wilt（CFW），Fusarium oxysporium f. sp. cucumerinum H₄₆ 5 strain and cucumber cultivar‘Changchun Mici’were used as materials，and the effect of different concentrations of PPa on the mycelial growth of H₄₆ 5 pathogen was deteced. The area of infected cucumber leaves，the malondialdehyde（MDA）content，the free proline content were measured. The activities of peroxidase（POD），catalase（CAT）and superoxide dismutase（SOD）were determined after PPa treatment under different lighting condition. The reactive oxygen species（ROS）including H₂O₂ content and superoxide anion production rate was estimated using 3,3′-diaminobenzidine（DAB）staining and nitro-blue tetrazolium（NBT）staining. The results showed that PPa had a significant inhibitory effect on H₄₆ 5，and its inhibitory effect was positively correlated with its concentration. The optimal inhibitory concentration of PPa was 2 g · L^-1. PPa treatment also significantly reduced the level of CFW disease and MDA content of the plant，and increased the proline content and antioxidant capacity of the plant，and maintained the ROS balance of the plant. Although the inhibitory ability of PPa was limited by the effects of light intensity and light duration，and the results proved that PPa had a good effect on inhibiting the disease of CFW.

Key words: cucumber, Fusarium wilt, pyropheophorbide-a, reactive oxygen species, disease resistance

于静, 冯向君, 金英学, 丁国华. 焦脱镁叶绿酸a对黄瓜枯萎病菌的抑制作用[J]. 园艺学报, 2024, 51(4): 859-874.

YU Jing, FENG Xiangjun, JIN Yingxue, DING Guohua. Inhibitory Effect of Pyropheophorbide-a on Cucumber Fusarium Wilt[J]. Acta Horticulturae Sinica, 2024, 51(4): 859-874.

图/表 9

图1 不同浓度焦脱镁叶绿酸a（PPa）处理对尖孢镰刀菌黄瓜专化型菌株（FOC）菌落生长的影响不同小写字母表示处理间差异显著（P < 0.05）。下同。

Fig. 1 Growth of Fusarium oxysporium f. sp. cucumerinum（FOC）colonies after treatment with different concentrations of（PPa）and inhibition rate of hyphal growth Different lowercase letters indicate a significant difference in the level of 0.05 among different treatments. The same below.

图2 不同浓度、光照强度和光照时长的焦脱镁叶绿酸a（PPa）对感染枯萎病的黄瓜叶片的防治效果

Fig. 2 Effect of pyropheophorbide-a（PPa）at different concentrations，light intensity and light duration on cucumber leaf infected with Fusarium wilt

图3 不同浓度、光照强度和光照时长的焦脱镁叶绿酸a（PPa）处理对感染枯萎病黄瓜叶片丙二醛（MDA）含量的影响

Fig. 3 Effects of different concentration，light intensity and light duration of pyropheophorbide-a（PPa）treatment on the content of malondialdehyde（MDA）in cucumber leaf infected with Fusarium wilt

图4 不同浓度、光照强度和光照时长的焦脱镁叶绿酸a（PPa）处理对感染枯萎病黄瓜叶片脯氨酸含量的影响

Fig. 4 Effects of different concentration，light intensity and light duration of pyropheophorbide-a（PPa）treatment on the content of proline in cucumber leaf infected with Fusarium wilt

图5 不同浓度、光照强度和光照时长的焦脱镁叶绿酸a（PPa）处理对感染枯萎病黄瓜叶片过氧化物酶（POD）活性的影响

Fig. 5 Effects of different concentration，light intensity and light duration of pyropheophorbide-a（PPa）treatment on peroxidase（POD）activity in cucumber leaf infected with Fusarium wilt

图6 不同浓度、光照强度和光照时长的焦脱镁叶绿酸a（PPa）处理对感染枯萎病黄瓜叶片超氧化物歧化酶（SOD）活性的影响

Fig. 6 Effects of different concentration，light intensity and light duration of pyropheophorbide-a（PPa）treatment on the activity of superoxide dismutase（SOD）in cucumber leaf infected with Fusarium wilt

图7 不同浓度、光照强度和光照时长的焦脱镁叶绿酸a（PPa）处理对感染枯萎病黄瓜叶片过氧化物酶（CAT）活性的影响

Fig. 7 Effects of different concentration，light intensity and light duration of pyropheophorbide-a（PPa）treatment on peroxidase（CAT）activity in cucumber leaf infected with Fusarium wilt

图8 不同浓度、光照强度和光照时长的焦脱镁叶绿酸a（PPa）处理对感染枯萎病黄瓜叶片过氧化氢含量的影响

Fig. 8 Effects of different concentration，light intensity and light duration of pyropheophorbide-a（PPa）treatment on hydrogen peroxide content in cucumber leaf infected with Fusarium wilt

图9 不同浓度、光照强度和光照时长的焦脱镁叶绿酸a（PPa）处理对感染枯萎病黄瓜叶片超氧阴离子的影响

Fig. 9 Effects of different concentration，light intensity and light duration of pyropheophorbide-a（PPa）treatment on superoxide anion in cucumber leaf infected with Fusarium wilt

参考文献 34

[1]	Cai Liang-yuan, Yang Jing, Xiong Yi-ming, Zhang Guo-cai, Lin Lian-nan. 2019. Photosensitive inhibition of sodium pheophorbide a against Pestalotiopsis neglecta under different light conditions. Chinese Journal of Pesticide Science, 21 (3):378-382. (in Chinese)
	蔡良苑, 杨璟, 熊一名, 张国财, 林连男. 2019. 不同光照条件对焦脱镁叶绿酸钠光敏抑菌活性的影响. 农药学学报, 21 (3):378-382.
[2]	ElNagar A, Abdelnaser A, Xuan T D, Gaber M, ElWakeil N, ElSayed Y, Nehela Y. 2022. Metal complexation of bis-chalcone derivatives enhances their efficacy against Fusarium wilt disease,caused by Fusarium equiseti,via induction of antioxidant defense machinery. Plants, 11 (18):36-42.
[3]	Eskandari S, Höfte H, Zhang T. 2020. Foliar manganese spray induces the resistance of cucumber to Colletotrichum lagenarium. Journal of Plant Physiology, 12 (4):246-247.
[4]	Hou Hui, Zhou Zeng-qiang, Wang Li, Wang Sheng-rong. 2016. Control effects of sodium silicate on grape anthracnose. Journal of Plant Protection, 43 (5):836-841. (in Chinese)
	侯珲, 周增强, 王丽, 王生荣. 2016. 硅酸钠对葡萄炭疽病的防治效果. 植物保护学报, 43 (5):836-841.
[5]	Ji Jing-yu. 2021. Study on the antibacterial mechanism of sodium pheophoribide(SPA)on Botrytis cinerea[M. D. Dissertation]. Haerbin:Northeast Forestry University. (in Chinese)
	季竞宇. 2021. 脱镁叶绿酸钠对灰葡萄孢菌抑菌机理的研究[硕士论文]. 哈尔滨: 东北林业大学.
[6]	Jin Peng, Lü Mu-wen, Sun Cui-cui, Zheng Yong-hua, Sun Ming. 2012. Effects of methyl jasmonate in combination with low temperature conditioning on chilling injury and active oxygen metabolism in loquat fruits. Acta Horticulturae Sinica, 39 (3):461-468. (in Chinese)
	金鹏, 吕慕雯, 孙萃萃, 郑永华, 孙明. 2012. MeJA与低温预贮对枇杷冷害和ROS代谢的影响. 园艺学报, 39 (3):461-468.
[7]	Jing Hong-juan, Zhou Guang-zhou, Tan Xiao-rong, Ping Kang-kang, Ren Xue-jian. 2012. Research progress in regulation of reactive oxygen species in plant autophagy. Chinese Bulletin of Botany, 47 (5):534-542. (in Chinese)
	景红娟, 周广舟, 谭晓荣, 平康康, 任雪建. 2012. ROS对植物自噬调控的研究进展. 植物学报, 47 (5):534-542. doi: 10.3724/SP.J.1259.2012.00534
[8]	Li Ge, Meng Xiao-qing, Cai Jing, Dong Ting-ting, Li Zong-yun, Zhu Ming-ku. 2018. Advances in the function of reactive oxygen species in plant responses to abiotic stresses. Plant Physiology Journal, 54 (6):951-959. (in Chinese)
	李格, 孟小庆, 蔡敬, 董婷婷, 李宗芸, 朱明库. 2018. ROS在植物非生物胁迫响应中功能的研究进展. 植物生理学报, 54 (6):951-959.
[9]	Li Xiao-fang, Zhang Zhi-liang. 2016. Experimental guide to plant physiology. 5th ed. Beijing: Higher Education Press. (in Chinese)
	李小方, 张志良. 2016. 植物生理学实验指导. 5版. 北京: 高等教育出版社.
[10]	Li Yue, Chen Zhi-long, Wang Jian-guo. 2004. Photo-activated toxicity of pyropheophorbide-a toward Drosophila melanogaster. Agrochemicals,(2):64-66. (in Chinese)
	李玥, 陈志龙, 王建国. 2004. 焦脱镁叶绿酸a对果蝇的光活化毒性. 农药,(2):64-66.
[11]	Liang Yao. 2021. Study of Trichoderma asperellum against cucumber Fusarium wilt on soil microecosystem[M. D. Dissertation]. Beijing: Chinese Academy of Agricultural Sciences. (in Chinese)
	梁瑶. 2021. 棘孢木霉防治黄瓜枯萎病的土壤微生态研究[硕士论文]. 北京: 中国农业科学院.
[12]	Liu Jia-wang, Fan Ning-bo, Wang Ai-xia, Duan Yu-feng, Wang Ting-xian, Yang Zhan-wei. 2022. Research progress on reactive oxygen species in tobacco. Modern Agricultural Science and Technology,(5):191-194. (in Chinese)
	刘家旺, 范宁波, 王爱霞, 段玉锋, 王廷贤, 杨占伟. 2022. 烟草中ROS的研究进展. 现代农业科技,(5):191-194.
[13]	Liu Dong, Dai Li-ting, Pu Zi-jing, Miao Di, Ma Bo-zhuang, Zhang Yan-ju. 2012. Virulence,vegetative compatibility and ISSR analysis of Fusarium oxysporum f. sp. cucumarinum. Acta Phytopathologica Sinica, 42 (5):456-465. (in Chianese)
	刘东, 代丽婷, 蒲子婧, 苗笛, 马柏壮, 张艳菊. 2012. 黄瓜枯萎病菌毒力、营养体亲和性及ISSR分析. 植物病理学报, 42 (5):456-465.
[14]	Liu H, Zhang H, Yang F, Chai S, Wang L, de Dios V R, Tian W R, Yao Y N. 2022. Ethylene activates poplar defense against Dothiorella gregaria Sacc by regulating reactive oxygen species accumulation. Physiologia Plantarum, 174 (3):e13726-e13738.
[15]	Meng Zhen, Zhang Weiping, Wang Ying, Li Long, Ji Xiaoxue, Dong Bei, Qiao Kang. 2022. Establishment and application of real-time PCR for quantitative detection of Fusarium oxysporum f. sp. lycopersici. Acta Horticulturae Sinica, 49 (11):2479-2488. (in Chianese) doi: 10.16420/j.issn.0513-353x.2021-0606
	孟臻, 张伟萍, 王莹, 李龙, 姬小雪, 董贝, 乔康. 2022. 番茄枯萎病菌RT-PCR检测技术的建立与应用. 园艺学报, 49 (11):2479-2488. doi: 10.16420/j.issn.0513-353x.2021-0606
[16]	Sayed M, Mirza H, Borhannuddin B, Khursheda P, Masayuki F. 2019. Exogenous tebuconazole and trifloxystrobin regulates reactive oxygen species metabolism toward mitigating salt-induced damages in cucumber seedling. Plants, 8 (10):45-54.
[17]	Sun Y K, Liu Z Y, Lan G P, Jiao C J, Sun Y. 2019. Effect of exogenous melatonin on resistance of cucumber to downy mildew. Scientia Horticulturae, 3 (22):231-241.
[18]	Tao Yong. 2017. The effect of methyl pyropheophorbide a mediated photodynamic therapy on human cell line MG63[M. D. Dissertation]. Chongqing: Chongqing Medical University. (in Chinese)
	陶勇. 2017. MG63 焦脱镁叶绿酸-a甲酯介导的光动力对人骨肉瘤MG63细胞凋亡影响的研究[硕士论文]. 重庆: 重庆医科大学.
[19]	Wang Bin, Zhang Teng-xiao, Liu Chao-qun, Zu Yu-yang, Li Yan-fang, Meng Xiang-cai. 2022. Research progress in the effects of abiotic stress on reactive oxygen species metabolism in medicinal plants. Research and Practice on Chinese Medicines, 36 (3):94-98. (in Chinese)
	王斌, 张腾霄, 刘超群, 祖余洋, 李艳芳, 孟祥才. 2022. 非生物胁迫对药用植物ROS代谢影响的研究进展. 现代中药研究与实践, 36 (3):94-98.
[20]	Wang D L, Liu B, Ma Z Q, Feng J T, He Y. 2020. Reticine A,a new potent natural elicitor:isolation from the fruit peel of Citrus reticulate and induction of systemic resistance against tobacco mosaic virus and other plant fungal diseases. Pest Management Science,(3):77-84.
[21]	Wang Ruo-meng, Dong Kuan-hu, Li Yu-ying, Li Chen, Yang Jing-fang. 2014. Effects of applying exogenous plant hormone on praline metabolism of Swainsonia salsual seedling under NaCl stress. Acta Prataculturae Sinica, 23 (2):189-195. (in Chinese) doi: 10.11686/cyxb20140223
	王若梦, 董宽虎, 李钰莹, 李晨, 杨静芳. 2014. 外源植物激素对NaCl胁迫下苦马豆苗期脯氨酸代谢的影响. 草业学报, 23 (2):189-195. doi: 10.11686/cyxb20140223
[22]	Wang Shu-ren, Zhang Bo-wen, Yang Jing, Ji Jing-yu, Zhang Guo-cai, Chen Guang-sheng. 2020. Inhibitory effect of sodium pheophorbide a on carrot gray mold. Chinese Journal of Pesticide Science, 22 (6):967-972. (in Chinese)
	王姝人, 张博文, 杨璟, 季竞宇, 张国财, 陈广胜. 2020. 脱镁叶绿酸钠对胡萝卜灰霉病菌的抑制作用. 农药学学报, 22 (6):967-972.
[23]	Wang Y J, Ye Z C, Li J H, Zhang Y G, Cheng Y X. 2021. Effects of dielectric barrier discharge cold plasma on the activity,structure and conformation of horseradish peroxidase(HRP)and on the activity of litchi peroxidase(POD). LWT-Food Science and Technology, 141:111-128.
[24]	Wu Han-xiang, Chen Zhi-bin, Xiao Chun-xia, Lai Duo, Yao Guang-kai, Xu Han-hong. 2021. Systemicity of fludioxonil derivatives with a carboxylic acid function and their control effect against Fusarium wilt of banana. Journal of Plant Protection, 48 (4):789-797. (in Chinese)
	吴瀚翔, 陈志彬, 肖春霞, 赖多, 姚光凯, 徐汉虹. 2021. 咯菌腈羧酸衍生物内吸传导性及对香蕉枯萎病的防效测定. 植物保护学报, 48 (4):789-797.
[25]	Xia Ming, Li Jing-wei, Luo Zhang-rui, Zu Gui-dong, Wang Ya, Zhang Wan-ping. 2022. Research on mechanism of exogenous melatonin effects on radish growth and resistence to Alternaria brassicae. Acta Horticulturae Sinica, 49 (3):548-560. (in Chinese) doi: 10.16420/j.issn.0513-353x.2021-0423
	夏铭, 李经纬, 罗章瑞, 祖贵东, 王娅, 张万萍. 2022. 外源褪黑素影响萝卜生长及对链格孢菌抗性的机理研究. 园艺学报, 49 (3):548-560. doi: 10.16420/j.issn.0513-353x.2021-0423
[26]	Xiang Miao-lian, Zhao Xian-yang, Chen Ming, Fu Yong-qi, Zeng Xiao-chun. 2017. Involvement of induced resistance by methyl jasmonate to bacterial wilt and reactive oxygen species metabolism in pepper ceedlings. Acta Horticulturae Sinica, 44 (10):1985-1992. (in Chinese)
	向妙莲, 赵显阳, 陈明, 付永琦, 曾晓春. 2017. 茉莉酸甲酯诱导辣椒抗青枯病与活性氧代谢的关系. 园艺学报, 44 (10):1985-1992. doi: 10.16420/j.issn.0513-353x.2017-0004
[27]	Xu J, Xian Q Q, Wang K, Dong J P, Zhang C Y, Du S L, Xu X W, Chen X H. 2022. Screening and identification of candidate Fusarium wilt-resistance genes from pumpkin. Horticultural Plant Journal, 8 (5):583-592.
[28]	Xu Lei, Tang Lei, Shao Wei-lan, Zhang Feng, Chen Yun, Wu Kang. 2005. The inhibition of pheophorbide a on Staphylococcus aureus and Escherichiacoli. Journal of Food Science and Biotechnology,(4):66-68. (in Chinese)
	徐磊, 唐蕾, 邵蔚蓝, 张峰, 陈蕴, 吴亢. 2005. 脱镁叶绿酸a对金黄色葡萄球菌、大肠杆菌的抑制作用. 食品与生物技术学报,(4):66-68.
[29]	Yang Jing. 2021. Study on the photoactivated abti fungal activity of sodium pheophoribide a against Pestalotiopsis neglecta and the possible mechanism involved[Ph. D. Dissertation]. Harbin:Northeast Forestry University. (in Chinese)
	杨璟. 2021. 脱镁叶绿酸钠对忽视拟盘多毛孢的光活化抑制作用机理研究[博士论文]. 哈尔滨: 东北林业大学.
[30]	You Yu-ming, Tang Jie, Zhang Mei-xia, Wang Da-ping, Lu Jia-hong. 2021. 2,4-Epibrassinolide alleviates chilling injury in baby ginger rhizome by regulating active oxygen and phenolic metabolism. Food Science, 42 (3):273-280. (in Chinese)
	游玉明, 汤洁, 张美霞, 王大平, 陆佳红. 2021. 外源2,4-表油菜素内酯调控仔姜ROS及酚类代谢减轻冷害. 食品科学, 42 (3):273-280.
[31]	Zhang Bao-cheng, Liu Yun-fang, Li Ying-lu, Chen Wen-fang, Li De-hui, Qian Zheng-min. 2021. Preliminary study on the effect of manganese stress on the growth and physiological and biochemical characteristics of Conyza canadensis. Journal of Northeast Agricultural Sciences, 46 (4):110-112,127. (in Chinese)
	张宝成, 刘云芳, 李应禄, 陈文芳, 李德辉, 钱正敏. 2021. 锰胁迫对小飞蓬生长与生理生化特征影响初探. 东北农业科学, 46 (4):110-112,127.
[32]	Zhang Lei, Wu Jingchun, Zhang Haiying, Gong Guoyi, Liu Tangjing, Zhang Jie, Xu Yong. 2023. Advances in functional genes for important traits in watermelon. Acta Horticulturae Sinica, 50 (12):2748-2764. (in Chinese) doi: 10.16420/j.issn.0513-353x.2022-1218
	张磊, 武竞春, 张海英, 宫国义, 柳唐镜, 张洁, 许勇. 2023. 西瓜重要性状功能基因研究进展. 园艺学报, 50 (12):2748-2764. doi: 10.16420/j.issn.0513-353x.2022-1218
[33]	Zhou Deng-bo, Jing Tao, Zhang Xi-yan, Qi Deng-feng, Chen Yu-feng, Wang Fei. 2016. Screening and antibacterial activity of antagonistic bacteria against banana Fusarium wilt disease. Journal of Plant Protection, 43 (6):913-921. (in Chinese)
	周登博, 井涛, 张锡炎, 起登凤, 陈宇丰, 王飞. 2016. 香蕉枯萎病拮抗菌筛选及其抑菌活性. 植物保护学报, 43 (6):913-921.
[34]	Zhou Hai-lin. 2020. The application of pyropheophorbide a(PPa)nanomaterials in cancer theranostics[M. D. Dissertation]. Suzhou: Suzhou University. (in Chinese)
	周海琳. 2020. 焦脱镁叶绿酸a纳米材料在癌症诊疗中的应用[硕士论文]. 苏州: 苏州大学.