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Journal of Integrative Agriculture 2014年第5期13卷 1137-1145页

作者： CHEN Song-jiang LI Yong-cai BI Yang YIN Yan GE Yong-hong WANG Yi College of Food Science and Engineering Gansu Agricultural University

Effects of different polarity solvents on the ultrastructure and chemical composition of cuticular wax in Pingguoli pear as well as their bioactive role against alternaria alternate were studied and the results showed that the highest wax content was extracted with chloroform, and its wax content was up to 322.2 gg cm2. Long-chain fatty acids predominated in menthol extracts and n-alkanes were predominant in wax extracted with ether, chloroform and n-hexane. Pingguoli pear fruit surface was covered by a smooth and amorphous wax layer with small, scattered crystal. The morphology of recrystallized wax in vitro after removal with different solvents was not similar to that of the intact fruit surface. Removal of cuticular wax with various solvents significantly enhanced A. alternata infection, except for wax removed by methanol. The solvent extracts of methanol and chloroform stimulated the spore germination and mycelium growth ofA. alternata, but the ether and n-hexane extracts showed antifungal activity.

关键词： fruit cuticular membrane alternaria alternata antifungal or stimulus activity

Morphological and Molecular Identification of alternaria alternata Causing Leaf Spot Disease on Huangdi Banana in China

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Plant Diseases and Pests 2023年第3期14卷 5-7页

作者： Xin ZHANG Hong ZHAO Yanfei OUYANG Yixian XIE Jun PENG Fanyun ZENG Yanxiang QI Environment and Plant Protection Institute Chinese Academy of Tropical Agricultural SciencesHaikou 571101China National Key Laboratory of Green Pesticide Key Laboratory of Green Pesticide and Agricultural BioengineeringMinistry of EducationCenter for R&D of Fine Chemicals of Guizhou UniversityGuiyang 550025China College of Plant Science and Technology of Huazhong Agricultural University Wuhan 430070China

[Objectives]The paper was to identify alternaria alternata causing leaf spot disease on Huangdi banana in China.[Methods]Fungal isolates were isolated and identified by morphological features,molecular identification and pathogenicity test.[Results]There were light to dark brown,tiny oval spots on leaves.The causal agent isolated from affected leaves was identified as A.alternata based on the morphological properties,coupled with sequence analyses of the internal transcribed spacer(ITS)region,large subunit ribosomal DNA(LSU rDNA)and the translation elongation factor 1-alpha(TEF-1α)gene.Koch s postulates were fulfilled by successful re-isolation of pathogen from the artificial inoculated leaves.[Conclusions]To our knowledge,this is the first report of leaf spot caused by A.alternata on Huangdi banana in China.The identification of A.alternata as the causal agent of the observed leaf spot disease on Huangdi banana is critical to the prevention and control of this disease in the future.

关键词： alternaria alternata Huangdi banana Leaf spot

ε-聚赖氨酸对梨果实黑斑病菌alternaria alternata的抑制作用及其机理

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食品科学 2021年第24期42卷 213-220页

作者：刘志恬董玉鹏李永才毕阳李宝军景春苑甘肃农业大学食品科学与工程学院甘肃兰州730070

本实验以梨果实黑斑病菌alternaria alternata为研究对象,通过测定菌落直径与病斑直径研究ε-聚赖氨酸(ε-poly-L-lysine,ε-PL)对A.alternata生长的抑制作用和对黑斑病的控制作用,并进一步通过测定细胞膜完整性、线粒体膜电位及其代谢... 详细信息

本实验以梨果实黑斑病菌alternaria alternata为研究对象,通过测定菌落直径与病斑直径研究ε-聚赖氨酸(ε-poly-L-lysine,ε-PL)对A.alternata生长的抑制作用和对黑斑病的控制作用,并进一步通过测定细胞膜完整性、线粒体膜电位及其代谢产物水平探讨其抑菌作用的机理。结果表明:ε-聚赖氨酸能显著抑制A.alternata的菌丝生长(Palternata孢子细胞膜和线粒体的完整性,提高了菌丝体电导率。但同时研究发现,ε-PL促进了A.alternata黑色素及链格孢酚、交链格烯、链格孢酚单甲醚3种毒素的产生,而对腾毒素的产生起抑制作用。综上,ε-PL主要通过延缓A.alternata生长、破坏孢子细胞膜和线粒体完整性而降低A.alternata的致病性。

关键词： ε-聚赖氨酸梨果实黑斑病 alternaria alternata 细胞膜线粒体真菌毒素

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新疆苹果褐色斑点果腐病病原菌的鉴定

中国果树 2024年第5期 106-110,134页

作者：苏宣乐常晓云张王斌塔里木大学南疆农业有害生物综合治理兵团重点实验室农业农村部阿拉尔作物有害生物科学观测实验站新疆阿拉尔843300

为明确新疆苹果褐色斑点果腐病病原菌的种类,对采自阿拉尔的苹果褐色斑点果腐病病果的病原菌种类进行了鉴定。利用组织分离法对病原菌进行分离纯化,获得菌株PGFL,采用柯赫氏法则进行致病性验证,结合形态学特征观察和ITS、RPB2、EF序列... 详细信息

为明确新疆苹果褐色斑点果腐病病原菌的种类,对采自阿拉尔的苹果褐色斑点果腐病病果的病原菌种类进行了鉴定。利用组织分离法对病原菌进行分离纯化,获得菌株PGFL,采用柯赫氏法则进行致病性验证,结合形态学特征观察和ITS、RPB2、EF序列构建多基因联合系统发育树对病原菌的种类进行确定。菌株PGFL的人工培养菌落形态呈现灰色至暗青褐色,与链格孢属alternaria形态观察比对结果高度相似,且接种离体果实后可发病。分子生物学分析菌株PGFL和菌株alternaria alternata MAFF239887及A.alternata JLUAF6聚为一簇,与alternaria属A.alternata的序列相似度最高,且自展支持率为100%。研究确定引起新疆苹果褐色斑点果腐病病原菌为链格孢A.alternata。

关键词：菌落形态分生孢子 ITS RPB2 EF alternaria alternata

Stem Histopathology of Sesame Seedlings Infected with alternaria alternata

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Microscopy Research 2016年第2期4卷 11-19页

作者： Maryam Ajmal Abida Akram Anum Ara Shaista Akhund Brian Gagosh Nayyar Wajiha Seerat Department of Botany PMAS Arid Agriculture University Rawalpindi Pakistan

In the present study, histopathology of three varieties of sesame TS 3, TS 5 and SG 27 infected with alternaria alternata was carried out to understand the mechanism of fungal infection and penetration in sesame plant as well as to determine the histological manifestation in sesame cells by light microscopy. Fungus was identified in infected tissues as a dark bluish black with toluidine blue O staining. Light microscopic examination of sesame stem showed that the fungus was present in epidermis, hypodermis and cortical parenchyma tissue as the symptoms became visible by naked eye ten days after inoculation (DAI). As the disease progress, the fungus moved from cortical parenchyma to vascular bundle, xylem and phloem. Later on, it completely overlapped the vascular bundle and entered in pith. When necrotic lesion appeared, fungus was present abundantly in epidermis, hypodermis, cortical parenchyma, vascular bundles and in pith. Due to its excessive growth and complete overlapping of cells, disorganization or destruction of cells of sesame took place. It was concluded that the alternaria alternata was not a tissue limited pathogen instead of this it spread in to all tissues of stem from epidermis to pith.

关键词： Sesame alternaria alternata Pathogen Fungus Light microscopy

Laboratory and Field Evaluation of Lupo Compound Miroemulsion(LCM) against alternaria alternata in Tobacco

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Plant Diseases and Pests 2014年第2期5卷 18-21页

作者： Lu Jinqing Dai Yi Li Xiaoshuang Liang Huan Cai Junlong Li Qiang Guo Shengnan Medicinal Plant Research and Development Center of Hubei Province Hubei University of Chinese Medicine

[Objective]The paper was to study effect of Lupo compound microemulsion( LCW) on alternaria alternata. [Method]The activity of spore germination and mycelial growth was investigated by filter paper method and mycelial growth rate method,and field test was also conducted to determine the effect of LCW on tobacco plants. [Result]LCW could inhibit spore germination and mycelial growth of A. alternate. The higher concentration led to more severe reduction of sporulation and heavier inhibition against mycelial growth. When LCW was diluted into 200 times,inhibition rate of sporulation reached 91. 23%,and the inhibition rate of mycelial growth reached 93. 50%. Field test also proved that LCW had significant inhibitory effect on tobacco brown spot disease. [Conclusion]The study provided the reference for using botanical pesticide to control A. alternate in fields.

关键词： alternaria alternata Botanical fungicide Lupo compound miroemulsion Disease control

Inhibitory Effects of 12 Chemical Agents against alternaria alternata

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Plant Diseases and Pests 2020年第4期11卷 20-23页

作者： Shen Hong Zhang Jiatao Cui Zhiyan Sun Guangjun Li Bin Zhang Yongfeng Xu Chuantao Ai Yongfeng Song Guanglong Ji Xiuting Wang Xiaoqiang Guizhou Provincial Company of China National Tobacco Corporation Guiyang 550081China Shaanxi Provincial Company of China National Tobacco Corporation Xi'an 710061China Sichuan Provincial Company of China National Tobacco Corporation Chengdu 610017China Institute of Tobacco Research of Chinese Academy of Agricultural Sciences Qingdao 266101China

[Objective]The paper was to screen highly efficient fungicides for the prevention and control of tobacco brown spot(alternaria alternata).[Method]The inhibitory effects of 12 chemical agents against A.alternata were determined indoors by mycelium growth rate method.[Result]Toxicity test showed that 30%kresoxim-methyl·boscalid SC had the best inhibitory effect,with an EC_(50)of 1.05μg/mL,followed by 9%pyraclostrobin CS,with an EC_(50)of 2.09μg/mL.As a broad spectrum protective fungicide,77%copper calcium sulfate WP showed a low sensitivity against A.alternata in the petri dish test since it had a mechanism of action different from that of other chemical agents.[Conclusion]The study lays the foundation for subsequent screening of fungicides for the prevention and control of A.alternata in the field.

关键词： Tobacco brown spot alternaria alternata Fungicide Sensitivity

发育及贮藏期苹果梨果皮蜡质对alternaria alternata侵染的影响

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发育及贮藏期苹果梨果皮蜡质对Alternaria alternata侵染的影响

作者：唐瑛甘肃农业大学

学位级别：硕士

本文以苹果梨作为研究试材,采用冷冻扫描电镜和光学显微镜观察分析发育及贮藏期果实表皮蜡质的结构变化及其病原物侵染过程;并通过体内(in vivo)和体外(in vitro)洋葱膜以及载玻片涂膜技术分析蜡质化学组分对A.alternata侵染结构形成、... 详细信息

本文以苹果梨作为研究试材,采用冷冻扫描电镜和光学显微镜观察分析发育及贮藏期果实表皮蜡质的结构变化及其病原物侵染过程;并通过体内(in vivo)和体外(in vitro)洋葱膜以及载玻片涂膜技术分析蜡质化学组分对A.alternata侵染结构形成、角质酶活性的影响,同时分析梨果实表皮及体外涂膜表面的疏水性(接触角)与A.alternata侵染结构形成之间的关系。结果表明:1.经冷冻扫描电镜观察在苹果梨果实发育过程中,花后30d果实表皮的蜡膜已经形成,但膜不完整,花后60d,蜡膜层进一步增厚,局部区域形成微裂纹,在果实发育后期,蜡膜厚度显著增加,皮孔被覆盖,果皮表面形成大量薄片状结晶体。在贮藏期间蜡膜厚度没有明显的变化,在贮藏前期果实表皮蜡质表面覆有片状结晶体,随着贮藏时期的延长,逐步转变为光滑的块状结晶体。2.A.alternata在苹果梨表皮上的侵染过程为孢子萌发形成芽管,芽管伸长、顶端膨大形成附着胞,部分附着胞进一步分化形成侵染菌丝,进而通过皮孔或表皮侵入。体内实验表明,与脱蜡表皮相比,完整果皮能有效的促进A.alternata附着胞和侵染菌丝的形成,不同发育期果实表皮对A.alternata侵染过程及侵染结构的促进作用有差异,其中A.alternata在花后60d果实表皮上2h孢子萌发,9h形成大量附着胞,且形成率高达55%,显著高于其它发育期;体外实验进一步证实了不同发育期果皮蜡质提取物对A.alternata侵染过程以及结构形成均具有促进作用,但由于蜡质提取物在不同膜上重结晶结构的差异,促进作用有所不同,如花后120d蜡质提取物能显著促进侵染菌丝形成,但其形成率在洋葱膜、Formvar膜、硅烷化载玻片上分别为22.3%、11%和4%。不同发育期果实表皮及体外不同膜的疏水性均存在差异,体外蜡质涂膜表面的疏水性高于果实表皮的疏水性,经相关性分析,疏水性与附着胞以及侵染菌丝形成之间是正相关,相同接触角果蜡对A.alternata侵染结构形成的促进作用远高于蜂蜡。体内和体外实验进一步表明发育期果皮蜡质对A.alternata角质酶的基因表达和酶活性也具有诱导作用。同时通过无损接种和化学组分分析表明,A.alternata对苹果梨蜡质也具有一定的降解作用。3.贮藏期完整果皮与脱蜡表皮相比,对A.alternata孢子萌发具有一定的抑制作用,却促进了侵染结构的形成以及芽管长度增加,不同贮藏期果皮蜡质对A.alternata侵染结构的形成具有差异。除了因不同介质导致的差异,体外实验进一步证实了贮藏期果皮蜡质均有效促进了A.alternata附着胞和侵染菌丝的形成,随着贮藏时间延长,果皮蜡质对侵染结构形成促进作用越明显。贮藏期果实表面疏水性均高于发育期,相关性分析同样证实表面疏水性与A.alternata附着胞、侵染菌丝形成呈正相关。贮藏期蜡质提取物对A.alternata角质酶活性具有一定的诱导作用,但对角质酶基因表达影响不大。综上所述,苹果梨发育贮藏过程表皮蜡质化学组分、疏水性以及表皮结构变化对A.alternata侵入过程以及侵染结构的形成均具有一定调控作用。

关键词：苹果梨蜡质 alternaria alternata 侵染结构角质酶

Hog1、Slt2-MAP激酶对梨果蜡质及疏水性诱导alternaria alternata侵染结构分化的调控作用

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Hog1、Slt2-MAP激酶对梨果蜡质及疏水性诱导Alternaria alternata...

作者：王调兰甘肃农业大学

学位级别：硕士

促分裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)级联信号途径对病原真菌生长、形态建成、侵染结构分化、次生代谢、致病性及渗透胁迫响应等多种生理功能具有调节作用,其调控作用因病原物种类、互作系统而异。为进一步阐... 详细信息

促分裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)级联信号途径对病原真菌生长、形态建成、侵染结构分化、次生代谢、致病性及渗透胁迫响应等多种生理功能具有调节作用,其调控作用因病原物种类、互作系统而异。为进一步阐明Hog1与Slt2-MAP激酶是否参与及如何调控alternaria alternata响应梨果表皮疏水性和蜡质信号从而启动侵染的过程,本试验在药理学试验的基础上,对梨果致病真菌A.alternata(JT-03)MAPK基因Hog1、Slt2进行鉴定、克隆和生物信息学分析,并构建缺失突变株和回复菌株,从分子水平上系统研究了Hog1和Slt2的生物学功能及对A.alternata响应梨果皮蜡质物化信号进而启动侵染的调控机制,结果如下:1.药理学结果表明MAPK特异性的抑制剂SB203580可显著地抑制梨果蜡质和疏水性诱导的A.alternata孢子萌发及附着胞的形成,同时也减弱了A.alternata对早酥梨的致病能力。2.对Hog1、Slt2-MAP激酶进行基因克隆后分别得到大小为1615bp和1747bp的克隆产物AaHog1和AaSlt2。同源序列比对得知AaHog1和AaSlt2基因编码产物均分别含有高渗反应所需的独特磷酸化基序“TGY”和自我磷酸化位点“TEY”。二级结构的主要元件是α-螺旋和无规卷曲,三级结构具有稳定的空间结构,均属于亲水性稳定蛋白。系统进化树表明AaHog1与AaSlt2基因分别与alternaria longipes的Hog和alternaria arborescens的Slt基因的同源性高达99%和91%。3.利用同源重组法构建ΔAaHog1,ΔAaSlt2,ΔAaHog1-C和ΔAaSlt2-C突变株。与野生型相比,ΔAaHog1和ΔAaSlt2的孢子形态和大小均发生了变化,菌落生长缓慢,产孢量分别显著降低了12.53%和68.13%(palternata孢子的形态、侵染结构、生长、细胞壁完整性和致病性等均具有调控作用。其中AaSlt2在参与A.alternata对梨果蜡质与疏水性的识别和响应过程中发挥更重要的作用。

关键词： alternaria alternata Hog1-MAPK Slt2-MAPK 基因敲除功能验证

两种致病真菌(Botryosphaeria dothidea和alternaria alternata)T-DNA突变体库的建立与分析

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两种致病真菌(Botryosphaeria dothidea和Alternaria alternata)T...

作者：顾雪迎新疆农业大学

学位级别：硕士

由葡萄球座菌(B.dothidea)及链格孢(A.alternata)引发的病害在世界范围内流行,也是影响我国水果生产的两大主要病害,给我国水果生产造成巨大的经济损失。目前,对于该两种病原菌的研究仍处于初级阶段。虽对AM毒素(AM-toxin)合成基因的研... 详细信息

由葡萄球座菌(B.dothidea)及链格孢(A.alternata)引发的病害在世界范围内流行,也是影响我国水果生产的两大主要病害,给我国水果生产造成巨大的经济损失。目前,对于该两种病原菌的研究仍处于初级阶段。虽对AM毒素(AM-toxin)合成基因的研究取得一些进展,但对其他相关基因及功能的研究仍较为缺乏。农杆菌介导的T-DNA随机插入突变法是目前应用于植物和真菌基因功能研究中常用且有效的方法。为能够尽早的揭示B.dothidea及A.alternata与寄主之间的互作关系,开展致病相关基因的研究,本研究旨在建立农杆菌介导的T-DNA转化体系,初步构建起两种致病菌的突变体库,并对部分突变体菌株进行分析,以获得一些性状差异显著的菌株。主要结果如下:1.成功构建了农杆菌介导的B.dothidea原生质体遗传转化体系,转化效率为220株/106个原生质体;筛选获得901株突变体,初步建立了B.dothidea突变体库,PCR验证eGFP基因及抗潮霉素B基因均成功整合到B.dothidea基因组中。2.对120株突变体菌株表型分析发现,43.3%的突变体菌株色素沉积加深,14.2%的菌株色数沉淀减弱;对其生长速率测量发现,59.3%的突变体菌丝生长速率加快,仅有6.2%的菌丝生长速率有较为明显的减慢。3.对500个突变体菌株致病性检测,筛选获得16株致病性明显减弱的菌株,占供试突变体菌株的3.2%。4.构建了链格孢苹果致病型农杆菌介导的孢子转化体系,优化了转化程序,以培养36-48 h所产孢子4℃处理6 h用于转化,转化效率为200株/106个分生孢子。5.筛选获得880个突变体,初步构建起链格孢苹果致病型XP-1菌株突变体库,通过PCR检测及Southern blot杂交分析,抗潮霉素B基因成功整合到病原菌的染色体中,且多数为单拷贝插入,单拷贝率达63.6%。6.对100个突变体菌株进行致病性检测,筛选获得7个致病性显著下降的突变体。

关键词： Botryosphaeria dothidea alternaria alternata 农杆菌介导突变体库