摘要
【目的】探明花椒精油对番茄链格孢菌(Alternaria alternata)的抑菌效果及作用机制,以期为利用花椒精油作为植物源熏蒸剂进行茄果类蔬菜链格孢腐烂病的生物防治提供理论依据。【方法】以番茄链格孢菌菌株HJF-1为试验对象,设不同浓度(40、80、120、160和200μg/mL)花椒精油及其3种主要成分芳樟醇、柠檬烯、月桂烯熏蒸HJF-1菌丝试验,以无菌水处理作为空白对照,评价不同物质对番茄链格孢菌的抑菌效果。通过显微镜观察、生理生化检测及离体接种等方法,进一步分析花椒精油和芳樟醇对病原菌的菌丝形态、孢子萌发、芽管长度、细胞膜完整性及致病力的影响,阐明花椒精油和芳樟醇熏蒸对番茄链格孢腐烂病菌的抑菌效果及作用机制。【结果】不同浓度的花椒精油及其3种主要成分熏蒸后,HJF-1菌落直径与对照之间存在明显差异,熏蒸抑菌效果表现为花椒精油>芳樟醇>月桂烯>柠檬烯,花椒精油和芳樟醇的抑菌效果相对较强,二者的最小抑菌浓度(MIC)均为160μg/mL。不同浓度花椒精油和芳樟醇熏蒸处理的菌丝生长抑制率存在显著差异(P<0.05,下同),以200μg/mL的浓度熏蒸72 h后,其菌丝生长抑制率分别达99.10%和99.00%。熏蒸浓度与生长抑制率呈剂量效应关系,花椒精油和芳樟醇对菌株HJF-1的半致死浓度(EC_(50))分别为85.98和89.84μg/mL。花椒精油和芳樟醇熏蒸处理不仅能显著降低孢子萌发率,还可显著抑制孢子芽管伸长,以浓度为200μg/mL花椒精油和芳樟醇熏蒸6 h后,菌孢子萌发率最低,分别为1.0%和1.2%,此时孢子芽管长度也最小,分别为9.2和10.8μm。此外,花椒精油和芳樟醇还能引起病原菌畸形和细胞膜氧化破损,导致菌丝丙二醛含量在熏蒸12 h时上升至6.3和4.0 nmol/mL;熏蒸24 h时,胞外可溶性蛋白含量分别上升至11.9和9.1μg/L,胞外可溶性糖含量分别上升至263.8和237.2μg/L,最终在熏蒸72 h时导致胞外相对电导率达最大值,分别为62.5%和50.5%。离体试验结果表明,2×MIC的花椒精油及芳樟醇熏蒸72 h可大幅降低链格孢菌的致病力,对链格孢腐烂病的相对防效分别达83.9%和75.6%。【结论】花椒精油及其主要成分芳樟醇通过抑制链格孢菌菌丝生长和孢子萌发而阻断病原菌侵染及延缓扩展,破坏细胞表面完整性而引起胞内物质渗漏及细胞功能紊乱,最终导致病原菌致病力降低。
【Objective】This research aimed to explore the antifungal effect and mechanism of Chinese prickly ash essential oil against Alternaria alternata in tomato,which would provide theoretical reference for the application of Chinese prickly ash essential oil as a botanical fumigant to control A.alternata rot disease in solanaceous vegetable.【Method】Using tomato A.alternata strain HJF-1 as the the tested material,a fumigation experiment was conducted on the mycelium of HJF-1 with different concentrations(40,80,120,160,200μg/mL)of Chinese prickly ash essential oil and its 3 main components(linalool,limonene and mycrene),with sterile water treatment as the blank control.The antifungal effect of different substances on A.alternata were evaluated.Through microscopy observations,physiological and biochemical analysis and in vitro inoculation methods,further analysis was conducted to study the effect of Chinese prickly ash essential oil and linalool on the mycelial morphology,spore germination,germ tube length,cell membrane integrity and pathogenicity of A.alternata,aiming to clarify the antifungal effect and mechanism of fumigation of Chinese prickly ash essential oil and linalool against A.alternata.【Result】After fumigation with different concentrations of Chinese prickly ash essential oil and its 3 main components,there were great differences in the colony diameter of HJF-1 compared to the control.The inhibition effect showed as Chinese prickly ash essential oil>linalool>myrcene>limonene.The inhibition effects of Chinese prickly ash essential oil and linalool were stronger than those of other components,and the minimum inhibitory concentration(MIC)of the two was both 160μg/mL.There was significant difference in the inhibition rates of mycelial growth fumigated with different concentrations of Chinese prickly ash essential oil and linalool(P<0.05,the same below)with mycelial growth inhibition rates of 99.10%and 99.00%treated with 200μg/mL of Chinese prickly ash essential oil and linalool respectively.There was a dose-effect relationship between fumigation concentration and growth inhibition rate of HJF-1,and the half lethal concentrations(EC_(50))of Chinese prickly ash essential oil and linalool were 85.98 and 89.84μg/mL respectively.Fumigation with Chinese prickly ash essential oil and linalool not only significantly reduced spore germination rate,but also inhibited spore tube elongation.By fumigation with 200μg/mL Chinese prickly ash essential oil and linalool for 6 h,spore germination rates were the lowest(1.0%and 1.2%respectively).At the same time,the lengthes of spore tubes were the minimum(9.2 and 10.8μm respectively).In addition,Chinese prickly ash essential oil and linalool could also cause pathogen deformity,cell membrane oxidative damage,and increased mycelium malondialdehyde content to 6.3 and 4.0 nmol/mL after panthogen fumigatied for 12 h compared with before fumigation.By fumigation with the two essential oil for 24 h,the inextracellular soluble proteins contents showed an increase and reached 11.9 and 9.1μg/L respectively,and a increase in extracellular soluble sugars contents to 263.8 and 237.2μg/L respectively.At 72 h of fumigation,extracellular relative conductivity reached the maximun,compared with before fumigation which were 62.5%and 50.5%.In vitro experiment indicated that fumigation with Chinese prickly ash essential oil and linalool(2×MIC)for 72 h greatly reduced the pathogenicity of A.alternata with relative control effects of 83.9%and 75.6%to Alternaria rot respectively.Chinese prickly ash essential oil and its main components linalool can block the infection and delay the spread of A.alternata by inhibiting mycelium growth and spore germination,causing intracellular leakage and function disorders through damage of cell surface integrity,and ultimately resulting in reduced pathogenicity of A.alternata.
作者
熊诗洁
朱学栋
袁继荣
万发兴
李京花
贾切
XIONG Shi-jie;ZHU Xue-dong;YUAN Ji-rong;WANG Fa-xing;LI Jing-hua;JIA Qie(College of Horticulture and Gardening,Yangtze University/Spice Crops Research Institute,Jingzhou,Hubei 434025,China;Southeast Chongqing Academy of Agricultural Sciences,Chongqing 408000,China)
出处
《南方农业学报》
CAS
CSCD
北大核心
2024年第8期2473-2484,共12页
Journal of Southern Agriculture
基金
湖北省重点研发计划项目(2022BBA0061)
湖北省自然科学基金项目(2023AFB1001)
重庆市自然科学基金面上项目(CSTB2022NSCQ-MSX1558)
宜昌市夷陵区科技计划专项(2023D-101)。
关键词
花椒精油
番茄
链格孢菌
抑菌机理
相对防效
Chinese prickly ash essential oil
tomato
Alternaria alternata
antifungal mechanism
relative control effect
作者简介
第一作者:熊诗洁(1998-),https://orcid.org/0009-0005-9087-0986,研究方向为植物活性成分抑菌保鲜效果及机理,E-mail:xsj19980418@163.com;通讯作者:贾切(1980-),https://orcid.org/0000-0001-8071-6137,博士,副教授,主要从事香辛植物活性成分采后贮藏生物学研究工作,E-mail:jiaqie020@163.com。
