瑞香狼毒活性物质对草莓灰霉病菌的抑制作用

 为探索瑞香狼毒（Stellera chamaejasme）提取物对草莓灰霉病菌（Botrytis cinerea）的抑菌活性,采用生物活性跟踪法对瑞香狼毒的抑菌活性成分进行初步研究,柱层析法对其甲醇萃取物高活性成分进行分离,经LC-MS和核磁共振仪对最终成分进行结构鉴定,结果表明瑞香狼毒最高活性流份3的主要抑菌活性成分为伞形花内酯,并且随着浓度的提高,该活性成分对草莓灰霉病菌的抑制作用也增强,其抑菌中浓度为1.52 mg·mL^-1。通过扫描电镜观察流份3对草莓灰霉病菌超微结构的影响,结果发现伞形花内酯严重破坏了草莓灰霉病菌孢子和菌丝体的形态结构。     

Synthesis of 2-amino-6-oxocyclohexenylsulfonamides and their activity against Botrytis cinerea

BACKGROUND: With the objective of exploring the fungicidal activity of 2‐oxocyclohexylsulfonamides (2), a series of novel 2‐amino‐6‐oxocyclohexenylsulfonamides (6 to 23) were synthesised, and their fungicidal activities against Botrytis cinerea Pers. were evaluated in vitro and in vivo. RESULTS: The compounds were characterised by IR, ¹H NMR and elemental analysis. Bioassay results of mycelial growth showed that compounds 6 to 23 had a moderate antifungal activity against B. cinerea. N‐(2‐methylphenyl)‐2‐(2‐methylphenylamino)‐4,4‐dimethyl‐6‐oxocyclohexenylsulfonamide (13) and N‐(2‐chlorophenyl)‐2‐(2‐chlorophenylamino)‐6‐oxocyclohexenylsulfonamide (21) showed best antifungal activities, with EC₅₀ values of 8.05 and 10.56 µg mL^?1 respectively. Commercial fungicide procymidone provided an EC₅₀ value of 0.63 µg mL^?1. The conidial germination assay showed that most of compounds 6 to 23 possessed excellent inhibition of spore germination and germ‐tube elongation of conidia of B. cinerea. For in vivo control of B. cinerea colonising cucumber leaves, the compound N‐cyclohexyl‐2‐(cyclohexylamino)‐4,4‐dimethyl‐6‐oxocyclohexenylsulfonamide (19) showed a better control effect than the commercial fungicide procymidone. CONCLUSION: The present work demonstrated that 2‐amino‐6‐oxocyclohexenylsulfonamides can be used as possible new lead compounds for further developing novel fungicides against B. cinerea. Copyright © 2011 Society of Chemical Industry     

拟康宁木霉T-51菌株发酵液对灰葡萄孢的抑菌活性

拟康宁木霉菌株T-51是从土壤中分离得到的一株优秀生防菌株,在前期的研究与应用中表现出对番茄灰霉病的良好防效。本文研究了T-51菌株的PDB发酵液对灰葡萄孢的抑菌活性,结果表明T-51发酵液在0.5%的低浓度下的抑菌率达到52.1%,与高浓度(10%)发酵液的抑菌率(59.7%)没有显著差异。当发酵液浓度下降为0.05%时失去抑菌活性。T-51摇培至第5天时的发酵液的抑菌活性稳定。T-51发酵液中的抑菌物质对高温不敏感,沸水浴处理10 min后抑菌活性没有明显降低。T-51发酵液pH为3.3,呈酸性,当调节pH至7时,发酵液抑菌活性从57.89%下降到46.61%。T-51发酵液能够显著抑制灰葡萄孢侵染番茄叶片后产生的病斑大小,接种后叶片病斑直径比对照减小53.9%。     

新杀菌剂对番茄灰霉病菌的室内毒力及田间防效

室内采用菌丝生长速率法、孢子萌发法和黄瓜子叶法测定了咯菌腈、抑霉唑、福美双、吡唑醚菌酯和嘧霉胺对番茄灰霉病菌( Botrytis cinerea)的抑制作用。结果表明,菌丝生长速率法和孢子萌发法的测定均以咯菌腈毒力最高,EC 50分别为0.005 2 μg/mL和0.087 6 μg/mL;黄瓜子叶法测定以抑霉唑毒力最高,其EC 50为0.675 3 μg/mL。田间药效试验结果表明：50%咯菌腈可湿性粉剂90 g/ hm2对番茄灰霉病的田间防效达到90%以上,显著高于对照药剂50%啶酰菌胺水分散粒剂300 g/hm2和40%嘧霉胺悬浮剂 480 g/hm2的防效。     

8种杀菌剂对番茄灰霉病菌的毒力及田间番茄灰霉病菌对咯菌腈的敏感性

为了解决番茄灰霉病菌的抗性问题,筛选出理想的替代药剂,采用菌丝生长速率法测定了山东寿光和河北徐水两地番茄灰霉病菌对不同类型杀菌剂的敏感性。两地的番茄灰霉病菌对咯菌腈、氟啶胺均高度敏感,对其他不同作用方式的杀菌剂表现出不同的敏感性。用同样方法检测了不同地区的106株番茄灰霉菌对咯菌腈的敏感性,结果显示其EC50值分布在0.003 9～0.044 9μg/mL之间,平均EC50值为(0.014 2±0.008 1)μg/mL。咯菌腈可作为防治番茄灰霉病的候选药剂。     

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%。     

拮抗灰霉菌的海洋细菌甲基营养型芽胞杆菌的筛选、鉴定及其抑菌活性物质的研究

采用Zo Bell’s 2216E培养基从65份渤海海水和底泥样品中分离到海洋细菌464株,通过平板对峙法获得了对灰霉菌具有较强拮抗作用的海洋细菌47株,其中菌株BH21的抑菌作用最强,抑菌带宽度为(11.8±0.3)mm;采用形态学、生理生化和分子生物学相结合的方法,将菌株BH21鉴定为甲基营养型芽胞杆菌Bacillus methylotrophicus,该菌具有较强耐盐、耐甲醇能力;通过排油圈法确定菌株BH21能够产生具有表面活性的代谢产物,随后以脂肽类物质相关基因的特异序列为引物进行菌株BH21抑菌活性成分相关基因的扩增,确定该菌株中含有itu A、itu B、itu C、itu D、fen D、srf AB和ynd J等7种脂肽物质的合成基因;利用盐酸沉淀和甲醇抽提法获得了BH21发酵液中脂肽粗提物,通过牛津杯法确定该脂肽粗提物具有明显抑制灰霉菌菌丝生长的能力,抑制率达到73.9%。     

Preventive and post‐infection control of Botrytis cinerea in tomato plants by hexanoic acid

The antifungal activity of hexanoic acid on the phytopathogen Botrytis cinerea was studied. This chemical inhibited both spore germination and mycelial growth in vitro in a concentration‐ and pH‐dependent manner, and stopped spore germination at a very early stage, preventing germ‐tube development. The minimum fungicidal concentration (MFC) for in vitro spore germination was 16 mm . Hexanoic acid also inhibited in vitro mycelial growth of germinated spores at an MFC of 12 mm . Studies performed to characterize the mechanisms underlying the antimicrobial effect of hexanoic acid showed that it alters fungal membrane permeability. In addition, hexanoic acid treatment increased the levels of spermine, spermidine, putrescine and cadaverine in B. cinerea mycelia. Spray application of hexanoic acid at fungicidal concentrations on 4‐week‐old tomato plants prior to fungal inoculation reduced necrosis diameter by approximately 60%. Application of the same hexanoic acid concentrations on previously infected plants reduced further necrosis expansion by around 30%. The results suggest that this chemical acts as a preventive and curative fungicide. Interestingly, treatments with hexanoic acid at concentrations below the MFC in hydroponic solution prior to fungal inoculation significantly reduced necrosis area. These results suggest an inducer effect of plant responses for hexanoic acid treatments at these concentrations. Hexanoic acid is a good candidate for safe antifungal treatments for the control of B. cinerea, which is responsible for many economic losses on fruits, vegetables and flowers.     

番茄灰霉病菌拮抗放线菌的筛选、鉴定及其活性评价

从安徽省寿县农田土壤中分离筛选到1株对番茄灰霉病菌Botrytis cinerea有强拮抗作用的放线菌菌株HNU-1。根据菌体形态特征、培养特征、细胞壁组分、生理生化特性和16S rRNA基因序列分析,将该菌株鉴定为吸水链霉菌Streptomyces hygroscopicus。菌株HNU-1发酵滤液可抑制灰霉病菌菌丝生长和分生孢子萌发,且浓度越高,抑制能力越强;当发酵滤液稀释6.67倍时则完全抑制灰霉病菌菌丝生长和分生孢子萌发。盆栽试验结果表明,菌株HNU-1发酵滤液6.67倍稀释液对番茄灰霉病的预防与治疗效果分别为87.8%和77.9%,均显著高于50%多菌灵可湿性粉剂500倍稀释液。     

番茄灰霉生防菌CQ的分子鉴定及其生防效果研究

菌株CQ是一株分离自云南省保山市昌宁县烟草根部的对番茄灰霉病具有较好防治效果的拮抗细菌。通过对该菌株进行形态特征观察、16S rDNA序列分析鉴定,确定菌株CQ为枯草芽孢杆菌Bacillus subtilis。拮抗试验结果表明,菌株CQ能够显著抑制番茄灰霉菌的菌丝生长,使菌丝出现异常分枝和囊泡状畸形。1‰ CQ发酵滤液即可显著抑制番茄灰霉菌孢子的形成。室内生测结果表明,相比对照组,48 h无菌发酵滤液原液在离体叶片上对番茄灰霉病的防效可达100%,而其10、100倍稀释滤液则使发病率分别降低28.5%和21.3%,病情指数分别降低44.01和37.86。     

醚菌酯对番茄灰霉病菌的生物活性及其防病方式研究

离体试验表明,醚菌酯对番茄灰霉病菌菌丝生长、孢子萌发、孢子产量和菌核产量均有一定的抑制作用,其EC50分别为935.670、0.667、0.479和6.975mg/L。其中对菌丝生长的抑制作用较低。但是,用10mg/L和20mg/L的醚菌酯叶片喷雾处理前24h,接种灰霉病菌菌碟,醚菌酯对灰霉病的保护效果分别达到92.3%和100%;处理后24h接种,治疗效果分别达83.3%和100%,说明醚菌酯对番茄灰霉病具有很好的保护和治疗作用。醚菌酯具有优异的内吸活性,能在植物体内向顶传导。     

来源于植物的短肽LD-1对番茄灰霉病的抑制作用及其抗病性研究

短肽LD-1是从牛心朴子草根中新分离的具有抗菌活性的多肽。本文以番茄灰霉病为供试菌株,研究了短肽LD-1对番茄灰霉病的抑制作用及其抗病性。结果表明:短肽LD-1对番茄灰霉病菌菌丝生长及孢子萌发具有较好的抑制作用。离体试验中,短肽LD-1对番茄灰霉病菌菌丝生长抑制作用的EC50在24、48、72h分别为28.3051、30.2134、32.7427mg/L,对孢子萌发的EC50为16.6919mg/L;活体试验中,短肽LD-1对番茄灰霉病具有较好的防治效果,90mg/L短肽LD-1对番茄灰霉病防治效果达到74.24%。短肽LD-1能导致病菌芽管发育畸形,不能正常发育。短肽LD-1对番茄种子的萌发和根系生长具有明显的促进效果,并且经短肽LD-1处理后的番茄须根发达,须根数目明显增多,促进了番茄的生长发育,提高了抗病性。     

Evaluation of anilinopyrimidine and other fungicides for control of grey mould (Botrytis cinerea) in container-grown Calluna vulgaris

BACKGROUND:Grey mould (Botrytis cinerea Pers. ex Fr.) is a damaging disease affecting container-grown Calluna vulgaris (L.) Hull. Trials were conducted to test anilinopyrimidine fungicides (pyrimethanil, cyprodinil and mepanipyrim), azoxystrobin and tolylfluanid for crop safety on cultivars, and to evaluate spray programmes based on alternating these fungicides with different modes of action for disease control. Timing and application at different spray intervals were also investigated. RESULTS:Cyprodinil was slightly phytotoxic on only one of the seven cvs tested. Alternating nine-spray programmes of azoxystrobin with an anilinopyrimidine or tolylfluanid, or an anilinopyrimidine with tolylfluanid, with the first spray applied immediately after potting, were equally effective in reducing disease incidence and severity compared with untreated plants. Spray programmes of five or eight sprays, using an anilinopyrimidine and at least one other fungicide with a different mode of action, applied alternately or in blocks of two sprays of the same fungicide and immediately after potting, were equally effective in reducing disease. Omission of early fungicide sprays until 9 weeks after potting and subsequent alternate application of pyrimethanil and tolylfluanid with a different mode of action at 6 week intervals gave reduced disease control. CONCLUSION:There is potential to use spray programmes based on alternating anilinopyrimidines with fungicides with a different mode of action to provide effective control of B. cinerea on C. vulgaris.     

Fungicidal potential of methoxylated flavones from citrus for in vitro control of Colletotrichum gloeosporioides, causal agent of anthracnose disease in tropical fruits

Four polymethoxylated flavones (3,5,6,7,3',4'-hexamethoxyflavone, 3,5,6,7,8,3',4'-heptamethoxyflavone, 5,6,7,8,4'-pentamethoxyflavone and 5,6,7,8,3',4'-hexamethoxyflavone) were isolated and characterized from cold-pressed orange oil. Their antifungal activities were evaluated against Colletotrichum gloeosporioides (Penz) Penz & Sacc, a major plant pathogen of fruits that causes significant damage to crops in tropical, sub-tropical and temperate regions. Methoxylated flavones were effective in inhibiting mycelial growth of the fungus. As flavone concentration increased, mycelial growth decreased. 5,6,7,8,3',4'-Hexamethoxyflavone completely inhibited the growth of Cgloeosporioides at a concentration of 100 microg ml(-1).     

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     

解淀粉芽胞杆菌MH71抗菌物质理化特性及对番茄灰霉病菌的抑菌活性

解淀粉芽胞杆菌Bacillus amyloliquefaciens MH71是一株高效广谱拮抗活性菌株,明确该菌株发酵液中活性物质的理化特性和抑菌活性,为解淀粉芽胞杆菌生物农药开发应用奠定坚实的基础。本研究在对菌株MH71抑菌活性物质理化特性研究的基础上,以番茄灰霉病菌为供试致病菌,探究了抗菌活性物质对番茄灰霉病菌的抑菌作用;采用酸沉淀和硫酸铵沉淀分别获得脂肽类和蛋白类活性物质,SDS–PAGE电泳和HPLC分别对活性物质进行检测。该菌活性物质具有较强的耐热性和pH适应性,100 ℃处理60 min和121 ℃处理30 min后抑菌活性分别为原有活性的88.98%和79.60%,在pH 3～11时均有抑菌活性,且对紫外线、蛋白酶K、胃蛋白酶和胰蛋白酶不敏感。该抗菌活性物质对番茄灰霉病菌菌丝的毒力测定结果表明,不同浓度发酵液处理的番茄灰霉病菌菌丝呈现出较多局部膨大或溢缩等异常菌丝。其发酵液经酸沉淀法得到抗菌脂肽类活性物质,经硫酸铵沉淀法得到抗菌蛋白,脂肽类的抑菌活性高于抗菌蛋白类物质,SDS–PAGE电泳检测,得到约40 kD的蛋白条带,HPLC检测表明该抗菌物质中有surfactin存在。     

Development of a new dispenser for microbiological control agents and evaluation of dissemination by bumblebees in greenhouse strawberries

BACKGROUND: To date, in modern agriculture, biological control strategies are increasingly becoming the preferred pest management approach. However, the success of microbiological control agents (MCAs) largely depends on efficient dissemination into the crop. The pollinator‐and‐vector technology employs pollinating insects like bees for a better dissemination. In this study, a new dispenser for bumblebee workers of Bombus terrestris L. was developed. Binab‐T‐vector and Prestop‐Mix were used as two typical MCA products for dissemination. RESULTS: In a first series of experiments in the laboratory for optimisation, the newly developed dispenser was a two‐way type dispenser, 20 cm long, with two rectangular compartments and different entrance and exit holes. In addition, the amounts of MCA loaded on the workers were 10 times higher with the new dispenser as compared with the side‐by‐side passageway (SSP) dispenser. Typically, the highest amounts were recovered from the thorax and legs of the workers. In a second series of experiments under greenhouse conditions with the use of queen‐right B. terrestris hives, successful dissemination in strawberry flowers was obtained at different distances from the hive (0–8 m, 8–18 m and 18–21 m), and the workers inoculated the first, second and third flowers that were consecutively visited. In addition, the new dispenser caused no adverse effects on worker foraging intensity, whereas a dramatic reduction was scored with an SSP dispenser. Finally, the data suggested that it is necessary to refill the newly developed dispenser at 3 day intervals. CONCLUSIONS: The results demonstrated that, with the use of the newly developed dispenser, bumblebee workers carried high amounts of MCA, and this resulted in a successful dissemination of MCA into strawberry flowers. Copyright © 2010 Society of Chemical Industry