北京地区番茄灰霉病菌对嘧霉胺的抗药性检测

为了检测北京地区番茄灰霉病菌对嘧霉胺的抗药性,指导生产上科学用药,从北京12个区、县采集番茄灰霉病样150份,分离纯化获得109个番茄灰霉病菌(Botrytis cinerea)的单孢菌株,采用菌丝生长速率法测定了番茄灰霉病菌对嘧霉胺(pyrimethanil)的抗药性。所测菌株中嘧霉胺抗性菌株出现的比例高达82.57%,且以高抗菌株为主,占78.89%,抗性水平最高的菌株EC50是最敏感菌株的5 096倍;不同区县番茄灰霉病菌对嘧霉胺的抗药性存在差异,门头沟区、密云县、延庆县、怀柔区和平谷区、大兴、通州和顺义8个区县的高抗菌株所占比例达66.67%～100%,海淀、朝阳和房山3个区未检测到高抗菌株。上述结果说明北京地区番茄灰霉病菌对嘧霉胺的抗药性非常普遍,且以高抗菌株为主,生产上应替换新型杀菌剂防治番茄灰霉病。     

辽宁省蔬菜灰霉病菌对嘧霉胺的抗药性

2006年从辽宁省不同地区采集蔬菜灰霉病果或病叶,经分离共获得灰霉病菌株86株。采用菌丝生长速率法测定了其对嘧霉胺的敏感性。试验结果表明,灰霉病菌对嘧霉胺产生中等水平抗药性,抗性频率为17.41%。经紫外、药剂联合诱导获得了高抗菌株,抗性倍数最高达62.78倍。嘧霉胺与多菌灵和腐霉利间不存在交互抗药性,苯胺基嘧啶类杀菌剂之间存在交互抗性。野生抗性菌株具有较好的遗传稳定性,连续转接9次后抗药性无明显下降。不同菌株的菌丝生长速度、鲜重和产孢量存在明显差异,但该差异与灰霉病菌对嘧霉胺的敏感性无相关性。     

山西省蔬菜灰霉病菌对嘧霉胺的抗药性检测

为明确山西省蔬菜灰霉病菌对嘧霉胺的抗药性,2010-2011年用单孢分离法从山西省数十个县、市蔬菜主产区采集、分离到171株灰霉病病原菌(Botrytis cinerea),并采用最低浓度抑制法测定各菌株对嘧霉胺的敏感性。结果表明,山西省灰霉病菌对嘧霉胺的平均抗性频率为50.29%,平均抗性倍数为1 423.4。在山西省不同地区灰霉病菌对嘧霉胺的抗性存在差异,其中嘧霉胺使用较多的朔州地区抗药性频率达到100%。     

武夷菌素对番茄灰霉菌(Botrytis cinerea)的作用方式

 借助光镜和透射电镜观察了武夷菌素对番茄灰霉菌菌丝生长和分生孢子萌发的形态影响,结果表明,武夷菌素处理的菌丝异常生长,分生孢子萌发的芽管膨大缢缩,菌丝体内液泡增多;武夷菌素能引起菌丝细胞膜透性的变化,造成液体培养基电导率增加;同位素标记的¹⁴C-谷氨酸、³H-葡糖胺和³H-腺嘌呤掺入实验表明,武夷菌素能抑制菌丝蛋白质的合成,对细胞壁几丁质和核酸的合成没有影响。     

Inhibition of Methionine Biosynthesis in Botrytis cinerea by the Anilinopyrimidine Fungicide Pyrimethanil

When mycelium of Botrytis cinerea was treated with low concentrations of the anilinopyrimidine fungicide pyrimethanil the total amount of free amino acids increased. Qualitative variations were also induced: alanine, glutamine, lysine, glycine, histidine, asparagine, arginine, threonine and moreover, α-aminobutyrate and β-alanine were accumulated; cyst(e)ine, valine, leucine and citrulline were reduced. When mycelium of B. cinerea was incubated with Na₂[³⁵S]O₄, pyrimethanil at 1·5 μM induced a decrease of [³⁵S]methionine and simultaneously an increase of [³⁵S]cystathionine. These data indicate that the anilinopyrimidine fungicide pyrimethanil inhibits the biosynthesis of methionine and suggest that the primary target could be the cystathionine β-lyase. © 1997 SCI.     

邻烯丙基苯酚对番茄灰霉病菌全细胞呼吸的影响

 利用Oxygraphy液相氧电极研究了植物源杀菌剂邻烯丙基苯酚对番茄灰霉病菌(Botrytis cinerea)全细胞呼吸的影响。结果显示,邻烯丙基苯酚处理浓度为5、10 mg/L,菌丝细胞的呼吸速率较对照分别增加了19.58%和24.56%;处理浓度为150 mg/L,可显著抑制菌丝细胞的呼吸,抑制率达到77.49%,表明邻烯丙基苯酚具有低浓度刺激而高浓度抑制灰霉病菌细胞呼吸的双重影响。用药剂处理萌芽期、非萌芽期的分生孢子,测定呼吸的结果表明,药剂对孢子的呼吸作用效果与对菌丝的相似,但以萌芽期分生孢子最为敏感。以0.01 mol/L琥珀酸钠作为病菌细胞呼吸底物时,10 mg/L邻烯丙基苯酚对病菌细胞的呼吸速率具有抑制作用,而添加0.1 mol/L葡萄糖和0.01 mol/L丙酮酸钠两种底物时却表现出刺激病菌细胞呼吸的作用,表明邻烯丙基苯酚对细胞呼吸的影响和氧化底物密切相关。反应液中单独添加呼吸电子传递链特异性位点抑制剂鱼藤酮20μmol/L、嘧菌酯10 mg/L和叠氮化钠0.01 mol/L后,病菌菌丝细胞呼吸均受到显著抑制,抑制率分别为48.94%、33.08%和39.18%。随后再添加10 mg/L邻烯丙基苯酚,不改变这种抑制效果。单独添加旁路氧化酶抑制剂水杨肟酸2 mM不影响病菌菌丝的呼吸,随后再添加10 mg/L邻烯丙基苯酚后,两者协同作用则可显著抑制菌丝的呼吸。     

北京地区番茄灰霉病菌的多重抗药性检测

2009年12月-2010年5月,在北京12个郊区县采集番茄病标样150份,分离纯化得到109个灰葡萄孢（Botrytis cinerea）单孢菌株,用最低抑制浓度法（MIC）测定了其对苯并咪唑类（多菌灵）、二甲酰亚胺类（腐霉利）和氨基甲酸酯类（乙霉威）杀菌剂的抗药性。结果表明：番茄灰霉病菌对多菌灵、腐霉利和乙霉威产生抗性菌株的频率分别为96.3%、80.7% 和58.7%;所测菌株对3类杀菌剂的抗性类型有BenRDicSNPCS、BenSDicSNPCR、BenRDicRNPCS和BenRDicRNPCR 4种,所占比例分别是19.3%、3.7%、21.1%和56.0%,表明北京地区番茄灰霉病菌对苯并咪唑类、二甲酰亚胺类和氨基甲酸酯类三类杀菌剂的抗药性严重,在生产中需慎用,应选择一些替代的新型杀菌剂和生物农药。     

番茄灰霉病菌对嘧霉胺的抗药性

2001年从辽宁省不同地区的保护地中采集番茄灰霉病果或病叶，经单孢分离共获得番茄灰霉病菌70株。采用菌丝生长速率法测定了其对嘧霉胺的敏感性。结果表明，番茄灰霉病菌已对嘧霉胺产生中等水平抗药性，抗性频率为20．0％。经室内药剂诱导获得了高抗菌株，抗性倍数最高达35．7倍。嘧霉胺与多菌灵及嘧霉胺与速克灵间不存在交互抗药性。野生抗性菌株具有较好的遗传稳定性，连续转接9次后抗药性无明显下降。不同菌株的菌丝生长速度、鲜重和渗透敏感性存在显著差异，但该差异与灰霉病菌对嘧霉胺的敏感性无相关性。抗性菌株与敏感菌株具有同样的致病能力。     

灰霉菌(Botrytis cinerea)采后致病性研究

 灰霉菌(Botrytis cinerea)是引起葡萄采后病害的主要病原真菌之一。葡萄灰霉菌可在田间潜伏侵染,采后由健康果实携带进入销售市场,该菌的显著致病症状为果实软腐和脱落。灰霉菌与葡萄的其它采后致病菌,如链格孢菌(Alternaria alternata)、镰刀菌(Fusarium sp.)、芽枝霉(Cladosporium sp.)、青霉菌(Penicillium sp.)、黑曲霉(Aspergillus nigar)和粉红单端孢菌(Trichothecium roseum)相比,不仅表现出明显的潜伏侵染优势,而且具有较强的低温(4℃)条件下的致病优势。4℃低温下灰霉菌在寄主葡萄体外和体内分泌多聚半乳糖醛酸酶(PG)活力均显著高于以上各菌,而在25℃下无显著差异,这一结果与该2种温度下灰霉菌接种果实后的症状表现一致。     

Inhibitory effect of bionic fungicide 2‐allylphenol on Botrytis cinerea (Pers. ex Fr.) in vitro

BACKGROUND: 2‐Allylphenol is a registered fungicide in China to control fungal diseases on tomato, strawberry and apple. It is synthetic and structurally resembles the active ingredient ginkgol isolated from Ginkgo biloba L. bark. 2‐Allylphenol has been used in China for 10 years. However, its biochemical mode of action remains unclear. An in vitro study was conducted on the biochemical mechanism of 2‐allyphenol inhibiting Botrytis cinerea (Pers. ex Fr.). RESULTS: The inhibition was approximately 3 times stronger when the fungus was grown on non‐fermentable source, glycerol, than that on a fermentable carbon source, glucose. Inhibition of B. cinerea and Magnaporthe oryzae (Hebert) Barr mycelial growth was markedly potentiated in the presence of salicylhydroxamic acid (SHAM), an inhibitor of mitochondrial alternative oxidase. Furthermore, at 3 h after treatment with 2‐allylphenol, oxygen consumption had recovered, but respiration was resistant to potassium cyanide and sensitive to SHAM, indicating that 2‐allylphenol had the ability to induce cyanide‐resistant respiration. The mycelium inhibited in the presence of 2‐allylphenol grew vigorously after being transferred to a fungicide‐free medium, indicating that 2‐allylphenol is a fungistatic compound. Adenine nucleotide assay showed that 2‐allylphenol depleted ATP content and decreased the energy charge values, which confirmed that 2‐allylphenol is involved in the impairment of the ATP energy generation system. CONCLUSION: These results suggested that 2‐allylphenol induces cyanide‐resistant respiration and causes ATP decrease, and inhibits respiration by an unidentified mechanism. Copyright © 2009 Society of Chemical Industry     

酵母挥发性物质环辛四烯抑制灰霉菌的研究

为了探明生防酵母菌菌株C410产生的挥发性物质对灰霉菌的抑制作用机理,本文以挥发性物质的主要成分之一环辛四烯为研究对象,测定其对灰霉菌的糖、蛋白、DNA、RNA、细胞的形态及细胞膜通透性的影响。结果表明,在16.5μL/L环辛四烯的作用下,灰霉菌生长速率显著降低,总糖含量基本不变;还原糖含量显著降低;蛋白含量变化不显著;RNA的完整性显著降低。在49.5μL/L环辛四烯的作用下,灰霉菌的生长几乎完全停止,但细胞通透性不受影响,不导致细胞死亡。环辛四烯孵育下灰霉菌DNA分子没有被打断。由此推测还原糖和RNA代谢过程是环辛四烯作用于灰霉菌的重要靶标,抑制其生长及侵染。     

药用植物提取物对草莓灰霉病菌抑菌活性研究

用丙酮为溶剂提取7种药用植物材料粉末,得到提取物母液。采用生长速率法和叶片离体接菌法测试提取物对草莓灰霉病菌的室内抑菌作用。在供试的7种植物材料中,五味子对菌丝生长的抑制率最高,达到95.65%,其次为薏苡,抑菌率为91.30%,决明、酸浆的抑菌率均达到了85%以上。结合叶片离体接菌法的试验结果,认为五味子和酸浆可以作为重点进行深入研究。     

安徽省番茄灰霉菌抗药性测定和治理

测定了安徽省番茄灰霉病菌对多菌灵、腐霉利、乙霉威的抗性。结果表明,田间有B_SD_SN_S、B_SD_SN_LR、B_SD_SN_HR、B_HRD_SN_S、B_MRD_LRN_S、B_HRD_LRN_S、B_HRD_LRN_LR7种抗性类型,对多菌灵、腐霉利、乙霉威具有抗性的菌株分别占试验总菌株数的28.3%、20%和73.3%。     

Effect of the methoxyiminoacetamide fungicide,SSF129, on respiratory activity in Botrytis cinerea

(E)-2-Methoxyimino-N-methyl-2-[2-(2,5-dimethylphenoxymethyl)phenyl]acetamide (SSF129) has been developed as a broad-spectrum systemic fungicide for control of cereal and fruit diseases. This compound inhibited NADH-oxidation by submitochondrial particles from mycelial cells of Botrytis cinerea, with an EC₅₀ value of 14.5 nM , due to blockage of electron transport through the cytochrome bc₁ complex in the mitochondrial respiratory chain. However, SSF129 did not suppress, but rather increased, oxygen consumption by mycelial cells of the fungus. This was because mycelial cells contain an alternative oxidase protein and the cells have the ability to rapidly switch electron flux from the main cytochrome pathway to the alternative pathway on blockage of the former by SSF129. The alternative pathway of the mycelia seems not to be operative when the cytochrome pathway is functional. Naturally occurring flavonoids inhibited the alternative oxidase of the mycelial cells in a dose-dependent manner, with EC₅₀ values of 68.4 µM for flavone and 63.7 µM for flavanone. These observations suggested that plant components play an important role in control of gray mould by SSF129. © 1999 Society of Chemical Industry     

苯霜灵与茶皂素对黄瓜灰霉病菌的联合作用研究

在室内用生长速率法测定了苯霜灵（ｂｅｎａｌａｘｙｌ）、茶皂素（Ｔｅａｓａｐｏｎｉｎ）及二者的不同比例的混剂对黄瓜灰霉病菌的抑制作用。生测结果表明,苯霜灵与茶皂素以１：４混用对抑制黄瓜灰霉病菌菌丝的生长有一定的增效作用,ＥＣ５０＝１１０．６３μｇ／ｍＬ,共毒系数为１６３．８４;二种药剂及其１：４比例的混剂对灰霉菌孢子的萌发没有显著抑制作用,但能迟孢子的萌发;实验还发现二种药剂及其混剂仅起抑菌作用,而无杀菌作用。用２５μｇ／ｍＬ浓度的苯霜灵处理菌丝悬浮液,发现处理后的菌丝短而分枝少,菌丝形成隔膜较早,老化局,细胞壁变薄,透明度增加。     

喜树碱对番茄灰霉病菌的病理反应

以番茄灰霉病菌为供试菌种,采用电导率法、吸光度法和电镜分析法对经过喜树碱（CPT）处理的番茄灰霉病菌进行病理反应研究。试验结果表明经200 mg/L CPT溶液处理后,番茄灰霉病菌菌丝细胞膜的通透性增加了30.7%,细胞外蛋白质浓度提高了13.5%。番茄灰霉病菌经CPT溶液处理后,还原糖和可溶性蛋白的含量随着处理时间的增长先升高后降低最后趋于平缓,在0～1 h之间增长最快。CPT处理后的番茄灰霉病菌的菌丝畸形,出现膨大或缢缩,菌丝生长点缢缩,细胞中的空腔增多,线粒体增多且线粒体嵴变得模糊,细胞核有降解的现象。     

Effects of carbendazim on conidial germination and mitosis in germlings of Fusarium graminearum and Botrytis cinerea

The effects of carbendazim on carbendazim-sensitive (wild-type) and -resistant strains of Fusarium graminearum and Botrytis cinerea were compared. When treated with carbendazim, conidia and germlings of wild-type F. graminearum and B. cinerea germinated and grew to produce stunted, distorted, and abnormally branched germlings; in addition, distinct nuclei were replaced by irregularly distributed chromosome masses, and normal nuclear division was not observed. The CMI (chromosome mitosis index) values of wild-type strains rapidly increased within 60 min of treatment with carbendazim and then rapidly declined. Whereas the wild-type strains of the two fungi responded similarly, the carbendazim-resistant strains of F. graminearum and B. cinerea responded differently to carbendazim. The resistant strain of F. graminearum showed normal mitosis but produced significantly more branches and higher CMI values when treated with carbendazim than when untreated. In contrast, the carbendazim-resistant strain of B. cinerea exhibited normal morphology and mitosis after carbendazim treatment. The results suggest that the mechanism for carbendazim resistance in F. graminearum differs from that in B. cinerea and other fungi.     

Effects of fludioxonil and pyrimethanil, two fungicides used against Botrytis cinerea, on carbohydrate physiology in Vitis vinifera L

In Vitis vinifera L, photosynthesis and photosynthate partitioning are affected in the presence of fludioxonil and pyrimethanil, two fungicides commonly used in vineyards against Botrytis cinerea Pers. However, the effects were found to be different according to the model studied: plantlets (cv Chardonnay) grown in vitro, fruiting cuttings (cv Chardonnay) and plants grown in vineyards (cvs Chardonnay, Pinot noir and Pinot Meunier). In the plantlets grown in vitro, both fungicides decreased gas exchanges, photosynthetic pigment and starch concentrations in the leaves, whereas soluble carbohydrates transiently accumulated, suggesting that plantlets mobilised starch in response to photosynthesis inhibition caused by fungicides. In the fruiting cuttings, the fungicides did not affect photosynthesis, although fludioxonil caused starch decrease in parallel with sucrose accumulation, suggesting that the fungicide effects were of lower intensity than in vitro. Conversely, in vineyard, the two fungicides stimulated photosynthesis and increased pigment concentrations in the three vine cultivars tested. In the meantime, glucose, fructose and starch levels of the leaves declined after fungicide exposure, whereas sucrose accumulated, indicating that sucrose synthesis increased in the leaves following the fungicide treatment. Among the three varieties, Chardonnay was the most sensitive to the fungicides as revealed by the intensity of the responses and the longer period for recovery. In vineyard, the results suggested that the two fungicides, in addition to inhibiting B cinerea development, had a beneficial effect on vine physiology through the stimulation of leaf carbon nutrition, which may further enable the plant to rapidly make use of its defence reactions.     

硼抑制灰霉病菌孢子萌发机制的初步研究

 本文研究了硼对灰霉病菌孢子萌发的影响并对其抑制机理作了进一步的分析。用0.1%硼处理灰霉病菌孢子后,显著地抑制了孢子萌发和芽管伸长。与不处理的对照相比,0.1%硼使灰霉病菌孢子细胞内超氧化物歧化酶(SOD)、过氧化氢酶(CAT)和过氧化物酶(POD)的活性逐渐下降,还原型谷胱甘肽(GSH)和抗坏血酸(ASA)含量逐渐降低,丙二醛(MDA)、过氧化氢(H₂O₂)含量和超氧阴离子(O₂^·)产生速率上升。此外,DCHF-DA荧光染色结果也进一步证明硼处理造成了灰霉病菌孢子细胞内活性氧的积累。本试验结果初步表明,0.1%硼处理使细胞内活性氧清除系统破坏,膜脂过氧化作用加强,是抑制灰霉病菌孢子萌发和芽管伸长的重要原因。     

致病杆菌CB43菌株对灰霉病菌的抑制作用

测定了致病杆菌CB43菌株代谢物对灰葡萄病菌菌丝生长和孢子萌发的影响。结果表明:CB43代谢物对灰葡萄孢菌丝生长有较强的抑制作用,500、250、125ml/L代谢物能完全抑制菌丝的生长,并具有杀死作用。62.5ml/L的发酵液处理灰葡萄病菌菌丝72h的抑制率仍达83.35%。20ml/L的发酵液处理灰葡萄孢菌丝能引起菌丝畸形,异常分支、粗短。CB43代谢物能杀死灰葡萄孢分生孢子或抑制其萌发,500~62.5ml/L的发酵液处理16h后,孢子萌发抑制率达88.1%~97.91%。温室试验表明,CB43代谢物对黄瓜灰霉病的控制效果达84.09%,化学农药40%施佳乐的效果为72.94%。