Metodologia e avaliação de resistência de Sphaerotheca fuliginea a fungicidas em cucurbitáceas

Pós-graduação em Agronomia (Proteção de Plantas) - FCA

虎杖对植物病原真菌的杀菌活性及有效成分研究

植物源杀菌剂的开发应用以及从植物中寻找杀菌活性物质作为先导化合物，是目前杀菌研究领域的热点之一。本文以蓼科植物虎杖（Polygonum cuspidatum Sieb.et Zucc.) 为材料，研究了虎杖提取物的杀菌活性和作用机理，确定了虎杖中的有效杀菌活性成分，并以此为先导化合物进行了衍生物合成与结构活性关系的研究。 不同溶剂提取物制备与杀菌活性测定结果表明，乙醇适合作为虎杖植物杀菌剂的提取溶剂，提取率高，对多种植物病原真菌具有广谱的杀菌和抑菌活性，除对黄瓜白粉病（Sphaerotheca fuliginea）表现出很好的防治效果外，对苹果腐烂病菌（Valsa mali）、玉米小斑病菌（Helminthosporium maydis）、葡萄炭疽病菌（Colletotrichum gloeosporioides）、小麦赤霉病菌（Fusarium graminearum）、油菜菌核病菌（Sclerotinia sclerotiorum）、水稻纹枯病菌（Rhizoconia solani）等也具有很好的抑制作用。虎杖回流提取物对黄瓜白粉病的杀菌作用以保护作用为主，兼具一定的治疗作用，并且具有一定的内吸活性，持效期约为4-7 d。温室试验结果表明，虎杖乙醇回流提取物10%可溶性液剂对黄瓜白粉病的EC90值为172.83 mg/L，田间小区试验表明该制剂在800-1600 mg/L的浓度下，对黄瓜白粉病的防效达到76.3-93.4%，具有较好的应用前景。 对苹果腐烂病菌的抑菌作用机理表明，虎杖乙醇提取物对该病原菌有明显的抑制作用，能够抑制蛋白质、葡萄糖等菌体细胞内物质的合成，从而使病菌代谢速度减慢，抑制其生长。虎杖提取物还能够使几丁质酶和β-1,3葡聚糖酶这两种细胞壁相关水解酶的活性升高，降解细胞壁而破坏菌体结构，使菌体自溶。 过测定虎杖乙醇回流提取物对黄瓜体内等一些防御酶和病程相关蛋白活性的影响，表明在40 mg/L和400 mg/L浓度下，虎杖乙醇提取物能够使黄瓜叶片内的过氧化物酶（POD）、多酚氧化酶(PPO)、苯丙氨酸解氨酶(PAL)、几丁质酶等不同程度的升高，从而在一定程度上提高植物对病原真菌的抗病能力。 通过生物活性跟踪测定以及pH梯度提取法确定了虎杖中的主要杀菌活性成分为蒽醌类化合物大黄素（emodin）和大黄素甲醚（physcion），结构通过了HPLC-MS和1H NMR确认，并且通过HPLC确定了虎杖乙醇回流提取物中二者的含量分别为3.28%和1.11%。 以虎杖中的有效成份之一的大黄素为原料，通过羟基的甲基化反应合成了包括已知物大黄素甲醚在内的11个大黄素衍生物，其中5个化合物为首次报道，并进行了初步结构活性关系研究。结果表明通过对大黄素3-OH位置以短直链烷基取代，其衍生物对黄瓜白粉病的活性大大提高，其中以甲基取代的衍生物大黄素甲醚的活性为母体大黄素的16.7倍，而以取代苄基修饰的衍生物的活性没有明显提高。一些目标化合物的活性明显优于三唑酮。研究中还意外发现大黄素的甲基化衍生物三甲氧基大黄素在4000 mg/L时能够明显抑制甜菜夜蛾幼虫的取食与生长发育，而大黄素和大黄素甲醚则无此作用。

大黄素甲基化衍生物的生物活性

在实验室内以天然存在的大黄素(emodin)为原料合成了甲基化衍生物大黄素甲醚(physcion)和三甲氧基大黄素(trimethoxyemodin),并进行了杀菌和杀虫活性研究,发现3个蒽醌类化合物对黄瓜白粉病(Sphaerotheca fuliginea)的活性有明显差异,其EC90值分别为56.291、3.382、102.699mg/L,大黄素甲醚的活性远高于大黄素、三甲氧基大黄素和对照药剂三唑酮,说明大黄素母环上3-OH以甲基取代杀菌活性会显著提高,而3个羟基均被甲基取代活性会降低。在对甜菜夜蛾(Spodoptera exigua)幼虫的活性测定中,发现三甲氧基大黄素对甜菜夜蛾幼虫的生理活动具有明显的抑制作用,在4000mg/L下对2龄和3龄幼虫的体重增长抑制率分别为99.6%和99.2%,而大黄素和大黄素甲醚则对其无明显影响。

Oído das cucurbitáceas.

2007

Efeito de tipos de leite sobre oídio em abóbora plantadas a campo

O trabalho foi realizado a campo para avaliar a eficiência de diversos tipos de leite de vaca sobre o oídio da abóbora cultivar Piramoita, causado por Sphaerotheca fuliginea. Utilizou-se o delineamento experimental de blocos ao acaso com seis tratamentos, cinco repetições e cinco plantas por parcela. Os tratamentos foram: leite cru, leite pasteurizado do tipo C, leite integral do tipo longa vida, leite pasteurizado tipo C + Yakult®, leite integral do tipo longa vida + Yakult® e água como controle. Os melhores resultados no controle da doença foram obtidos com leite de vaca cru, com leite do tipo C e leite do tipo C + Yakult®. Concluiu-se que leite é uma alternativa viável no controle do oídio, mesmo após o início da infecção, no campo. Sua utilização na forma de leite cru mais eficiente e na forma de longa vida o menos eficiente.

Effect of potassium silicate on epidemic components of powdery mildew on melon

The effects of silicon (Si) supplied in the form of potassium silicate (PS) were evaluated on epidemic components of powdery mildew of melon under greenhouse conditions. The PS was applied to the roots or to leaves. In the first experiment, epidemic components were evaluated after inoculation with Podosphaera xanthii. In the second experiment, the disease progress rate was evaluated on plants subjected to natural infection. The area under the disease progress curve was reduced by 65% and 73% in the foliar and root treatments, respectively, compared to control plants, as a consequence of reductions in infection efficiency, colony expansion rate, colony area, conidial production and disease progress rate. However, root application of PS was more effective than foliar application in reducing most of the epidemic components, except for infection efficiency. This can be explained by the high Si concentration in leaf tissues with root application, in contrast to the foliar treatment where Si was only deposited on the external leaf surfaces. The effects of PS reported in this study demonstrated that powdery mildew of melon can be controlled, and that the best results can be achieved when PS is supplied to the roots.

Systematic status of Fejervarya ((Amphibia, Anura, Dicroglossidae) from South and SE Asia with the description of a new species from the Western Ghats of Peninsular India

We carried out a large-scale phylogenetic analysis of fejervaryan (dicroglossid frogs with `Fejervaryan lines' on the ventral side of the body) frogs, distributed in South and SE Asia, using published and newly generated sequences of unidentified individuals from the northern Western Ghats. The results corroborate the presence of a larger fejervaryan clade with a sister relationship to a clade composed of Sphaerotheca. Two sister clades could be discerned within the lager fejervaryan clade. The unidentified individuals formed a monophyletic group and showed a strong support for a sister relationship with Minervarya sahyadris. The species was found to be highly divergent (16S rRNA-4% and tyr-1%) from its sister lineage Minervarya sahyadris, and the clade composed of these two lineages were found to be deeply nested within the larger clade of Fejervarya. Based on this, the genus Minervarya Dubois, Ohler and Biju, 2001 is synonymized under the genus Fejervarya Bolkay, 1915. The unidentified lineage is recognized, based on phylogenetic position, genetic divergence and morphological divergence, as a distinct species and named here as Fejervarya gomantaki sp. nov. The presence of rictal glands was observed to be a synapomorphic character shared by the nested clade members, Fejervarya sahyadris and Fejervarya gomantaki sp. nov. Based on the presence of rictal gland and small size, Minervarya chilapata, a species from a lowland region in the Eastern Himalayas, is synonymized under Fejervarya and evidence for morphological separation from the new species, Fejervarya gomantaki sp. nov. is provided. For the fejervaryan frogs, currently three generic names (Frost, 2015) are available for the two phylogenetic subclades; the genus Fejervarya Bolkay, 1915 for the species of fejervaryan frogs having distribution in the South East Asia; the genus Zakerana Howlader, 2011 for the species of fejervaryan frogs having distribution in the South Asia and the genus Minervarya Dubois, Ohler and Biju, 2001 nested within the `Zakerana clade'. In the phylogenetic analysis Minervarya sahyadris, the new species described herein as Fejervarya gomantaki sp. nov. are nested within the `Zakerana clade', if the `Zakerana clade' for the fejervaryan frogs having distribution in the South Asia is provided a generic status the nomen `Minervarya' should be considered as per the principle of priority of the ICZN Code. Taking into consideration the overlapping distribution ranges of members of the sister clades within the larger fejervaryan clade and the absence of distinct morphological characteristics, we also synonymize the genus Zakerana Howlader, 2011, a name assigned to one of the sister clades with members predominantly distributed in South Asia, under the genus Fejervarya Bolkay, 1915. We discuss the need for additional sampling to identify additional taxa and determine the geographical ranges of the members of the sister clades within Fejervarya to resolve taxonomy within this group.

黄瓜白粉病抗机制的细胞化学及形态学上探讨

黄瓜白粉病[Sphaerotheca fuligenea [Schlecht] Poll]是瓜类尤其是温室瓜类的主要病害之一。本工作用光学显微镜、扫描电镜、透射电镜细胞化学方法，在不同层次上探索抗病和感病寄主，黄瓜，对黄瓜白粉病的不同反应。实验结果如下： 1. 从外观上观察，抗病和感病品种在接种七天后就表现明显的病症差异。在抗病品种上白粉发生较少，感病的则白粉甚多。 2. 从光学显微镜下发现，在抗病品种叶子上，白粉病孢子萌发率虽然比感病的高，但是以后在菌丝生长及产生孢子梗能力上却比感病品种的弱。 3. 从扫描电镜中观察到，接种两天后，抗病品种叶片上，病菌孢子有的菌丝生长不良，至第三天，有的菌丝顶端发生弯曲、变形、甚至降解。而在感病品种上则未见此现象。 4. 从透射电镜的细胞化学定位来看，接种四天后，抗病品种具较高的过氧化物酶活性，酶活性增高的部位是在表皮细胞、叶肉细胞壁、乳突、膜包体(membrane-bound body)和过氧化物体。在感病品种的乳突中，穿透钉壁及外围一圈上缺少过氧化物酶和多酚氧化酶活性，而在抗病品种中则具有酶活性。糖蛋白的相对量在感病品种中较抗病的为多。在感病品种中主要分布在表皮细胞相互连接的壁上和表皮与栅栏细胞连接的壁上以及穿透处附近的表皮细胞外壁。 在电镜下我们观察到还有一些细胞学上差异，即在抗病品种上，有的附着胞不能穿透细胞壁，部分穿透细胞壁后不能穿透乳突，甚至穿透乳突后也不能发育成吸器或发育不良;而在感病品种则一般吸器发育良好。基于以上的实验结果，我们认为过氧化物酶和多酚氧化酶活性可能与壁木质化有关，本质化的细胞壁可作为抵御病原侵入的屏障。糖蛋白则可能与病原和寄主的识别有关。看来黄瓜白粉病抗病机理似是多因素的协调和组合而起作用。

A new species of Tontelea from Amazonian Peru and Ecuador, and notes on the Tontelea attenuata species group (Celastraceae, Hippocrateoideae)

Tontelea fuliginea differs from other species whose stamens alternate with undivided stigma-lobes by its profusely ramified, many flowered, and densely dirty-brown puberulous inflorescences. This paper also provides a synopsis and key of the group of species with alternate stamens and 3-lobed stiomas with undivided lobes (T. attenuata group). Four lectotypes are designated for previously published names: Tontelea longifolia, Tontelea micrantha, Tontelea corrugulata, and Salacia micrantha var. lancifolia.

Resíduos da indústria pesqueira na indução de supressividade a Cylindrocladium spathiphylli e Ralstonia solanacearum e de biofertilizantes no controle de oídios

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Primera cita de Podosphaera fusca, responsable del oídio de la caléndula en Mendoza, Argentina

La caléndula (Calendula officinalis) es una planta herbácea utilizada por sus hermosas flores amarillas o naranjas en borduras, jardinería y como flor de corte. Está ampliamente difundida en el mundo, no sólo por la belleza de sus flores o por su uso en la cosmética sino por ser resistente, de fácil cultivo y poco exigente en condiciones ambientales. En Mendoza es muy empleada como ornamental y todos los años es afectada por una enfermedad denominada oídio. En Argentina se mencionan como agentes etiológicos del oídio diversas especies del género Erysiphe y Oidium erysiphoides, pero no se había descripto la forma teleomórfica sino hasta 1991 en Corrientes y Córdoba, cuando fue citada como Sphaerotheca fusca. En 2004, en Mendoza, se produjo un intenso ataque de oídio sobre esta especie floral y al estudiar la pulverulencia típica de la enfermedad se observaron cuerpos negros, lo que motivó el estudio etiológico de la enfermedad en dicha provincia. Se muestrearon ejemplares de caléndula afectados procedentes de distintos orígenes. En laboratorio se realizaron observaciones y estudios morfométricos que permitieron reconocer Podosphaera fusca (=Sphaerotheca fusca) como agente responsable del oídio de la caléndula en Mendoza.

Filogenia molecular do gênero Morganella Zeller (Fungi, Basidiomycota) utilizando marcadores ITS

With the development and improvement of techniques for molecular studies and their subsequent application to the systematic, significant changes occurred in the classification of gasteroid fungi. The genus Morganella belongs to the family Lycoperdaceae, and is characterized mainly by lignicolous habit and presence of paracapilicium. Recent data demonstrate the discovery of new species for the group and the existence of a wide variety of species occurring in tropical ecosystems. However, the phylogenetic relationships of the genus, as well as the taxonomic classification, still require revisions to be better understood, the literature studies that address this issue are still very scarce. Thus, the objective of this study was to conduct studies of molecular phylogeny with species of the genus Morganella, to enhance understanding of the phylogeny of the group by including tropical species data. For this, the specimens used both for DNA extractions as for morphological review were obtained from Brazilian and foreign herbaria. For morphological analysis were observed characters relevant to the group's taxonomy. For phylogenetic analysis the Maximum Parsimony and Bayesian Analyzes were used, using the internal transcribed spacer (ITS) of nuclear ribosomal DNA. In phylogenetic analyzes, representatives from Morganella form a monophyletic clade with good support value and based on these results the genus should not be included as subgenus of Lycoperdon. The analysis indicated that M. pyriformis was not grouped with other representatives of Morganella, and therefore should not be included in the group as representative of Apioperdon subgenus because it is a Lycoperdon representative. Moreover, M. fuliginea, M. nuda, M. albostipitata, M. velutina, M. subincarnata are grouped with high support values within the genus Morganella. Morganella arenicola based on morphological and molecular studies does not aggregate in Morganella. Morganella nuda was grouped with M. fuliginea giving indications that can be treated as an intraspecific variation. The results of the analyzes favor to a better understanding of the species of Morganella. However, additional studies using a greater number of species, as well as other molecular markers are needed for a better understanding of the phylogenetic of Morganella.