银杏内生真菌拮抗灰霉菌的初步研究

从银杏枝条和叶片中分离到77个真菌菌株,以作物重要病原菌灰葡萄孢(Botrytis cinereaPers.)为指示菌,筛选拮抗菌株。平板对峙测定结果表明:46个菌株对灰霉菌有拮抗作用,占总菌株数的59.7%,其中有7个菌株拮抗性较强,占总菌株数的9.1%。不同内生菌株发酵液离体测定中,2-18和2-9菌株防治离体月季叶片和花瓣灰霉病的效果较稳定,抑制率保持在80.8%~100%。初步鉴定结果表明,2-18和2-9菌株为毛壳菌属真菌(Chaetom iumsp.)。     

Bioluminescence reporter assay system to monitor Arabidopsis MPK3 gene expression in response to infection by Botrytis cinerea

The Arabidopsis MPK3 gene product participates in disease resistance mediated by the MAP kinase cascade. The expression of the MPK3 gene is induced by pathogen inoculation and treatment with chemicals such as salicylic acid (SA) and methyl jasmonate (JA), but the detailed expression pattern of the MPK3 gene has been largely unknown. To investigate MPK3 gene expression in response to disease stress, we fused the MPK3 promoter to the firefly luciferase gene to create a real-time monitoring system for regulated gene expression in planta. The results of an in vivo reporter assay using transgenic Arabidopsis plants harboring MPK3::Fluc showed that the MPK3 promoter activity was induced by treatment with chemicals such as SA and benzo(1,2,3)-thiadiazole-7-carbothioic acid S-methyl ester (BTH), that induce defense gene expression. Inoculation with the fungal pathogen Botrytis cinerea resulted in systemic induction of MPK3::Fluc.     

Activity and metabolism of cyano-oxime derivatives in various strains of Botrytis cinerea

The fungitoxicity of cymoxanil, 4 cyano-oxime analogs (in which the acetylurea group was replaced with various groups: amide, ester, propargyl, and cyanomethyl-amide), and 2 cymoxanil-metabolites was studied against various strains of Botrytis cinerea. The fungicidal effect was measured on germ-tube elongation and mycelial growth. Cymoxanil and the analogs bearing the amide and ester groups showed the best anti-botrytis activity. The strains studied can be classified into 3 groups according to the germ-tube sensitivity to cyano-oximes. These groups fitted well with the 3 phenotypes of sensitivity to cymoxanil previously characterized in our laboratory: CyaS₁ (highly sensitive phenotype); CyaS₂ (moderately sensitive phenotype); and CyaR (tolerant phenotype). The bio-transformations of all the cyano-oximes were monitored in the culture-medium of the different strains using HPLC- and IP-HPTLC-methods. HPLC-studies showed that cymoxanil and the analogs bearing the groups amide and ester were quickly metabolized in the culture-medium of the CyaS₁ strain. Moreover, these studies allowed us to correlate disappearance of these cyano-oximes, to their fungicidal activity towards the CyaS₁ strain. This suggests that cyano-oximes and particularly cymoxanil are probably activated in a fungitoxic compound. IP-HPTLC-studies strengthened the precedent results and allowed us to correlate disappearance of cyano-oximes studied with appearance of two acids metabolites of hydrolysis. These metabolites could be the actual active-principles.     

盾叶薯蓣灰霉病(Botrytis cinerea)的研究

对一种引起盾叶薯蓣叶、茎呈暗绿色至黑褐色水渍状软腐的病害进行了调查 ,并作了鉴定。结果明确其为灰葡萄孢菌 (Botrytis cinerea Pers.ex Fr )引起的一种新病害 ,根据症状和病原鉴定将该病称为盾叶薯蓣灰霉病。该病原菌生长温度 0～30℃ ,最适生长温度 10～ 2 5℃。病害在每年春季、夏初和秋末 ,雨水多的年份发生严重。     

Trichoderma longibrachiatum Rifai 对 Botrytis cinerea 的拮抗机制研究

 从竞争、抗生和重寄生3方面研究Trichoderma longibrachiatum Rifai对Botrytis cinerea的拮抗机制.结果表明:T.longibrachiatum对B.cinerea的拮抗机制主要是竞争作用和抗生作用,没有观察到重寄生现象.     

Biological control of foliar pathogens by means of Trichoderma harzianum and potential modes of action

Biocontrol of foliar diseases is an alternative means of management of foliar pathogens. One of the most studied commercial biocontrol agents is isolate T39 of Trichoderma harzianum which can be regarded as a model to demonstrate biocontrol under commercial conditions and the mechanisms involved. This biocontrol agent (BCA) controls the foliar pathogens, Botrytis cinerea, Pseuperonospora cubensis, Sclerotinia sclerotiorum and Sphaerotheca fusca (syn. S. fuliginea) in cucumber under commercial greenhouse conditions. Control efficacy was similar for three different rates (covering a fourfold range). Involvement of locally and systemically induced resistance has been demonstrated. Cells of the BCA applied to the roots, and dead cells applied to the leaves of cucumber plants induced control of powdery mildew. The BCA suppressed enzymes of B. cinerea, such as pectinases, cutinase, glucanase and chitinase, through the action of protease secreted on plant surfaces. A combination of several modes of action is responsible for biocontrol. However, biocontrol is not achieved by means of antibiotics or by mycoparasitism, in spite of the fact that BCA has the potential to degrade cell-wall polymers, such as chitin.     

Role of strawberry volatile organic compounds in the development of Botrytis cinerea infection

Botrytis cinerea, the main pathogen of strawberry, has the ability to remain quiescent in unripe tissue and develop disease symptoms in ripe fruit. As strawberry ripening is characterized by an increase of aroma compounds, the role of volatile emission in the development of infection was investigated. Thirty‐five strawberry volatile organic compounds (VOCs) were tested on B. cinerea in vitro and volatile emission was analysed in strawberry harvested at four ripening stages by headspace solid‐phase microextraction/gas chromatography–mass spectrometry and proton transfer reaction–time of flight–mass spectrometry. The coupling of such data sets made it possible to conclude that key strawberry aroma compounds stimulate B. cinerea conidial germination and some typical wound‐volatiles stimulate pathogen conidial germination or mycelial growth. This study is the first report of fungal stimulation by some VOCs naturally occurring in strawberry: the esters ethyl butanoate, cis‐3‐hexenyl acetate, trans‐2‐hexenyl acetate, methyl butanoate and hexyl butanoate, the furanones furaneol and mesifurane, and the alcohol trans‐2‐hexenol. The results of this work provide advances in understanding the functional role of fruit VOCs and suggest, for the first time, that fruit VOCs may influence the development of B. cinerea from the latent phase and that they could favour the invasive growth of B. cinerea after wounding. In particular, ethyl butanoate and furaneol could signal strawberry ripening, and the green leaf volatiles trans‐2‐hexenol, trans‐2‐hexenyl acetate and cis‐3‐hexenyl acetate could signal the presence of damaged tissues that are easier sites for penetration by B. cinerea.     

解淀粉芽胞杆菌MH71的生防活性及脂肽类抗生素基因检测

从漠河多年永冻土层采集的冻土样品中分离的拮抗菌解淀粉芽胞杆菌MH71,对多种植物病原真菌和细菌具有较强的拮抗作用。本文对该菌株的生防活性相关特性及其对番茄灰霉病的离体防效进行了研究,同时,应用特异性引物对MH71中脂肽类抗生素合成有关基因进行检测。结果表明,该拮抗菌产生嗜铁素、蛋白酶、纤维素酶和吲哚-3-乙酸(IAA),不产生磷酸酯酶和几丁质酶;番茄离体叶片第5天调查结果表明该菌发酵液对番茄灰霉病的防效为92.8%,而在番茄果实上第7天和第11天调查的结果表明,与清水对照对比,发酵原液、发酵液5倍稀释液与50%啶酰菌胺1 000倍液化学农药对番茄灰霉病均有良好的防治效果,防效均为100%;从该菌的基因组DNA中扩增得到了fenB、mycB、ituA、sfp等脂肽类抗生素合成酶基因。此研究结果为该拮抗菌生防潜力的深入评价及活性物质的分离、鉴定和番茄灰霉病的生物防治提供了重要的参考资料。     

草莓花瓣脱落对果实灰霉病的影响

【目的】明确衰老花瓣在草莓果实灰霉病中的作用以及草莓花瓣脱落早晚与果实灰霉病发病的关系,为果实灰霉病的防控和抗（耐）灰霉病品种选育提供理论依据。【方法】试验1：以草莓品种‘甜查理’为材料,进行田间试验,摘除至少2 000个果实表面的衰老花瓣,以不摘除花瓣的处理为对照。保持塑料棚内高湿度8 d后,调查不同处理草莓果实灰霉病发病率。试验2：分别在2013、2014和2015年3月草莓盛花期,对11个草莓品种的田间花瓣脱落动态进行调查,每个品种选择20朵刚现蕾的花,记录现蕾后第7、14、21和28天果实上残留的花瓣数量,计算花瓣脱落率;于果实灰霉病发病高峰期,调查不同草莓品种果实灰霉病的发病率,分析现蕾后第7天花瓣脱落率与果实灰霉病发病率的相关性。试验3：对33个草莓品种现蕾后不同天数的花瓣脱落率与果实上残留花瓣数量进行调查,并分析两者的相关性。【结果】试验开始前棚内的草莓果实灰霉病发病率为4.2%;利用自然雨雪天气保持塑料棚内高湿度8 d后,摘除果实上残留衰败花瓣的处理,果实灰霉病的平均发病率为7.3%,对照小区果实灰霉病的平均发病率为25.3%,大部分果实均从与花瓣接触的地方开始感染。在不同试验年份,草莓品种之间的花瓣脱落趋势始终一致。所有草莓品种在花朵现蕾后,随时间延长,花瓣脱落率不断提高,但是不同草莓品种的花瓣脱落速度却有显著差异。不同年份草莓品种‘晶瑶’在现蕾后7 d的花瓣脱落率分别为99.0%、79.8%和94.4%,显著高于‘晶玉’（61.0%、24.0%和62.5%）和‘甜查理’（26.0%、3.8%和31.9%）。2013年调查的5个草莓品种的果实灰霉病发病率与花朵现蕾后第7天花瓣脱落率显著负相关 (P＜0.05);2014年调查的6个草莓品种,2015年调查的11个草莓品种的果实灰霉病发病率与花朵现蕾后第7天花瓣脱落率都显著负相关 (P ＜ 0.01)。对33个草莓品种的调查发现,不同草莓品种现蕾后第7天花瓣脱落率与果实上残留花瓣数量显著负相关(P ＜0.01)。【结论】衰老花瓣是灰葡萄孢（Botrytis cinerea）侵染果实的主要途径,摘除草莓果实上衰老花瓣可显著降低果实灰霉病的发病率。现蕾后第7天的草莓花瓣脱落率与果实灰霉病的发生具有显著负相关性,利用这种避病性,在选育抗灰霉病草莓品种时可优先考虑花瓣脱落早的材料。