Thesis (doctoral)--Christian-Albrechts-Universität zu Kiel, 2002.

Universiẗat, Diss., 2003--Giessen.

University, Diss., 2005--Köln.

Ingeniero en Biotecnología Molecular / La vid (Vitis vinífera) es uno de los cultivos más importantes dentro de las plantas frutícolas. En los viñedos estas se encuentran sometidas constantemente a patógenos que pueden afectar en la producción y calidad final del fruto destacando el hongo filamentoso Erysiphe necátor agente causal del oídio de la vid. Actualmente la mayor parte de las variedades que se comercializan para la producción de uva de mesa y vino son afectadas por este patógeno. RUN1 y REN1 son dos loci de carácter dominante estudiados en variedades no comerciales de vid, que por si solos confieren resistencia contra esta enfermedad, ambos traducirían para proteínas capaces de reconocer en alguna etapa las alteraciones que produce el patógeno sobre la planta, generando una respuesta de defensa efectiva contra la infección del hongo, por lo que la piramidación de ambos loci en una variedad de interés podría mejorar su defensa, haciéndola más efectiva y duradera en el tiempo. Esto permitiría en los viñedos disminuir la cantidad de químicos necesarios que se utilizan actualmente para combatir este hongo. En este trabajo se identificaron y estudiaron variedades portadoras de RUN1 y REN1 provenientes de iterativos retrocruzamientos entre diversas variedades comerciales y parentales resistentes que portaban uno de estos loci. Para conocer la respuesta que se generó a partir de la piramidación, se evaluó y caracterizó la defensa frente a la infección de oídio bajo condiciones de invernadero. Para identificar las líneas positivas para RUN1 se utilizaron dos parejas de partidores dirigidos al gen MrRUN1 presente dentro del locus, mientras que para identificar aquellas vides portadoras de REN1 se usaron marcadores moleculares tipo microsatélites dirigidas a dos regiones que segregan junto con la resistencia. Se analizaron los resultados, obteniendo plantas portadoras de ningún, uno o ambos loci. Mediante análisis de tinciones histológicas y microcortes 2 transversales de plantas infectadas, se logró observar una fuerte producción de especies reactivas de oxígeno y muerte celular en aquellas plantas portadoras de por lo menos algún loci, teniendo un efecto mayor en aquellas líneas que poseían ambos. Para determinar si la presencia de estos loci provocaba un cambio en la inducción de hormonas relacionadas a procesos de defensa, se midieron las concentraciones de ácido salicílico (SA), ácido jasmónico (JA) y ácido abscísico (ABA) a través de HPLC, observando diferentes cambios según el genotipo infectado. Para saber si existía algún efecto provocado por estos loci en la inducción de genes que participan en el proceso se realizó qPCR de algunos genes relacionados con el proceso infectivo o la defensa, así para todos los genotipos se vio un aumento en la inducción de MLO7 (MLO-like protein 7), gen involucrado en la susceptibilidad de la enfermedad, mientras que para PEN1 (PENETRATION 1), gen involucrado en procesos de resistencia, se observó un aumento mayor en aquellas líneas resistentes. Estos resultados sugieren una respuesta efectiva, robusta y contenida frente a la infección del hongo en aquellas líneas portadoras de ambos loci. / The vid (Vitis vinífera) is one of the most important crops among fruit plants. At the vineyards grapevines are constantly submitted to different pathogenic microorganisms that can affect the production and the final quality of fruit. One of the most important pathogens that affect its growth is the filamentous fungus Erysiphe necator, which causes the disease called powdery mildew, taking the nutrients from green tissue and berries. Currently, most of the marketable varieties for table grapes and wine production are affected by this pathogen. RUN1 and REN1 are two dominant loci studied in non commercial vineyards varieties, which by themselves confer resistance against powdery mildew. It has been suggested that both loci codify for proteins capable of recognizing the alterations produced by the pathogen on the plant in certain infection stage, generating an effective defence response against fungal infection. Therefore, the pyramiding of the two loci in a variety of commercial interest could improve their defense, making it most effective and lasting over time. This would lead to a decrease the amount of chemicals compounds currently used to combat this fungus in vineyards. In this work, RUN1 and REN1 plant carriers were identified and studied from iterative backcrosses between several commercial and resistant varieties that carried one of these loci. In order to analyze the defense response generated from the pyramiding plants, it was characterized against the infection of powdery mildew under greenhouse conditions. To identify the RUN1 positive lines, two pairs of primers directed to the MrRUN1 gene (located within the locus) were used; while to identify those vines carrying the REN1 locus, molecular markers (SSR) were used, identifying two regions that segregate 4 together with the resistance. We obtained plants carrying none, one or both loci. Through histological staining and transverse microcutting of infected plants, it was possible to observe a strong production of reactive oxygen species (ROS) and cell death in the plants carrying at least one locus, finding a greater effect in those lines that carry both loci. To determine if the presence of these loci changes the profile of hormones related to defense processes, the concentrations of salicylic acid (SA), jasmonic acid (JA) and abscisic acid (ABA) were measured through HPLC. We found differences according to the infected genotype. To find out if there was any effect caused by these loci in the induction of genes involved in the process, qPCR of some genes related to infection or defense was performed. We found increased induction of MLO7 (MLO-like protein 7) in all genotypes, involved in the susceptibility of the disease, whereas for PEN1 (PENETRATION 1), a gene involved in resistance processes, a greater induction was observed only in resistant lines. These results suggest an effective, robust and contained response to fungal infection in those lines carrying both loci

Orientador: Edson Luiz Furtado / Resumo: A seringueira (Hevea brasiliensis) é a terceira cultura florestal mais plantada no Brasil, e o estado de São Paulo é responsável por mais de 60% da produção de látex do país. É uma espécie nativa da região amazônica que possui hábito caducifólio, e durante o período de reenfolha é suscetível ao ataque de fitopatógenos foliares, que prejudicam e até limitam seu estabelecimento em determinadas regiões. Dentre os principais patógenos foliares da seringueira, destacam-se os fungos causadores de oídios, microrganismos biotróficos que infectam tecidos jovens, e podem causar desfolha, principalmente em condições de viveiro e jardim clonal. O agente causal do oídio da seringueira é descrito como Erysiphe quercicola, porém, devido a insuficientes descrições taxonômicas, morfológicas e moleculares desta doença, seu agente causal permanece em discussão, além da fase sexual deste patógeno carecer de qualquer registro neste hospedeiro. Portanto, o objetivo deste trabalho é caracterizar o oídio da seringueira, utilizando caracterizações morfológicas, análises moleculares e testes de patogenicidade. No período de julho a dezembro de 2019, foram coletadas amostras foliares de seringueira em diferentes estágios fenológicos infectadas com oídio na região de Botucatu, São Paulo. Para a caracterização morfológica, foram registradas e mensuradas características das hifas, conídios e conidióforos, que correspondem à fase assexuada do fungo. A caracterização molecular foi realizada através de análi... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Rubber tree (Hevea brasiliensis) is the third most planted forest crop in Brazil, and the state of São Paulo is responsible for more than 60% of the country's latex production. It is a species native to the Amazon region that has a deciduous habit, and during the re-leafing period it is susceptible to the attack of leaf phytopathogens, which harm and even limit its establishment in certain regions. Among the main leaf pathogens of the rubber tree, there are the fungi that cause powdery mildew, biotrophic microorganisms that infect young tissues, and can cause defoliation, especially in nursery and clonal garden conditions. The causal agent of rubber mildew is described as Erysiphe quercicola, however, due to insufficient taxonomic, morphological and molecular descriptions of this disease, its causal agent remains under discussion, in addition to the sexual phase of this pathogen lacking any record in this host. Therefore, the objective of this work is to characterize powdery mildew mildew, using morphological characterizations, molecular analyzes and pathogenicity tests. From July to December 2019, leaf samples of rubber trees were collected at different phenological stages infected with powdery mildew in the Botucatu region, São Paulo. For morphological characterization, characteristics of hyphae, conidia and conidiophores, which correspond to the asexual phase of the fungus, were recorded and measured. Molecular characterization was performed through phylogenetic analyzes o... (Complete abstract click electronic access below) / Mestre

In ihrer Evolution mussten Pflanzen Strategien entwickeln um sich sowohl gegen Pathogene aus der Luft als auch solche im Boden zu verteidigen. Diese Resistenzmechanismen der Pflanzen zu verstehen ist von höchster Wichtigkeit für die moderne Gesellschaft. Die Weltbevölkerung wächst schnell, was zu der Notwendigkeit führt, die landwirtschaftlichen Flächen möglichst optimal zu nutzen. Ohne die Weiterentwicklung der landwirtschaftlichen Methoden wird eine ausreichende Versorgung mit Grundnahrungsmitteln nicht möglich sein. Obwohl nicht viele Daten zu diesem Thema vorliegen, ist es sehr wahrscheinlich, dass ein hoher Prozentsatz der jährlichen Ernteverluste auf Pflanzenkrankheiten zurückzuführen ist (Orke et al. 1994, Pinstrup-Andersen; 2001). Der Ernteverlust ist nicht ausschließlich auf den Tod der infizierten Pflanze zurückzuführen, sondern vielmehr auf die sogenannten Resistenzkosten Walters und Heil; 2007). Um sich gegen das Pathogen zu schützen müssen Ressourcen genutzt werden, die sonst für die korrekte Entwicklung der Pflanze, sowie der Samen und Früchte verwendet würden. Die pflanzliche Cuticula, welche die Blattoberfläche bedeckt, ist die erste Verteidigungslinie gegen pathogene Microorganismen, die durch die Luft verbreitet werden. Um diese Barriere zu umgehen nutzen Bakterien und einige Pilze die Stomata als Eingang in den Apoplasten der Blätter. Dies kann durch die Pflanze allerdings verhindert werden, indem diese Poren geschlossen werden. Diese Schließzellantwort wurde zunächst als Teil der Immunantwort auf Bakterien angesehen (Melotto et al. 2006). Nichtsdestotrotz konnte beobachtet werden, dass die Stomata auch während der Infektion des Mehltaupilzes schließen, obwohl dieser nicht durch die Stomata in das Blatt eindringen. Daher haben wir Einzelzellstudien an intakten Gerstenpflanzen vorgenommen um zu klären, wie die Signale erkannt und weitergeleitet werden, die schließlich zum pathogen-induzierten Stomaschluss führen (Koers et al. 2011). Zusammengefasst kann gesagt werden, dass der Stomaschluss ein wichtiger Bestandteil der pflanzlichen Immunantwort ist. Innerhalb dieser Antwort der Stomata auf durch Wind übertragene Pathogene, spielt die Aktivierung der S-Typ Anionenkanäle eine entscheidende Rolle. Es konnte dabei gezeigt werden, dass die Immunantwort die Licht-induzierte Inhibierung dieser Anionenkanäle außer Kraft setzt. S-Typ Anionenkanäle sind aber nicht allein in der Pathogenabwehr von Bedeutung, sondern auch in der Reaktion der Pflanzen auf Trockenstress. Es ist jedoch nicht bekannt, in wie weit sich die beiden Signalwege überschneiden. Zusammen mit den neuen mutierten Gerstenlinien, werden die in dieser Arbeit beschriebenen Techniken zur Messung von Einzelzellen tiefere Einsichten in das Zusammenspiel zwischen Trockenstress und Pathogenabwehr in Pflanzen ermöglichen. Die daraus resultierenden Ergebnisse können zur Optimierung von Getreide für die moderne Landwirtschaft genutzt werden. Dies wird einer der wichtigsten Ansätze sein, um die Menschheit auch in Zukunft mit ausreichend Nahrung versorgen zu können. / During evolution, plants had to evolve potent strategies to defend themselves against airborne pathogens, as well as against those encountered in the soil. Understanding the mechanisms that provide plant immunity is crucial for modern society. The world population is growing at rapid pace, leading to the necessity of using agricultural areas as productive as possible. Without improvement of agricultural practice, a sufficient supply with staple foods will not be possible. It is very likely that an important percentage of crop loss is due to plant diseases, even though precise data on this issue are lacking, (Orke et al. 1994, Pinstrup-Andersen; 2001). Crop loss is not exclusively caused by the death of infected plants, but more often by so called costs of resistance (Walters and Heil; 2007). To gain protection against an attacking pathogen, resources have to be consumed, which otherwise would be used for proper plant, crop and fruit development. Plant cuticles, that cover the leaf surface, are the first line of defence to airborne pathogenic microorganisms. To bypass this barrier, bacteria and some fungi use stomata as an entry site into the apoplastic space of leaves. The entry of pathogens through stomata can be prevented by plants upon closure of these pores. This guard cell response was proposed to contribute to plant innate immunity against bacteria (Melotto et al. 2006). However, stomata were found to close during the infection of powdery mildew fungi, which do not use open stomata to enter the leaf. We therefore pursued single cell studies within intact barley plants to elucidate the signal perception and transduction mechanisms that evoke stomatal closure during a pathogen attack (Koers et al. 2011). All results taken together, stomatal closure is an integral part of plant innate immunity. Within the stomatal response to airborne pathogens, the activation of S-type anion channels is essential. It is shown, that the immunity responses of guard cells bypass light induced inhibition of anion channels. S-type anion channels are not only crucial for responses to pathogens, but they are also involved in the response of guard cells towards drought. However, it is unknown to which extent both signals share mutual components. Together with the, now available, mutant lines of barley, the single cell techniques described in this thesis can provide a further insight into the interplay of drought and pathogen responses in plants. The results are likely to be used for optimizing crops for the future agriculture, which is a pivotal step in providing enough food for mankind in the near future.

Memoria para optar al Título Profesional De Ingeniero Agrónomo / Esta investigación surge de la necesidad de encontrar nuevas alternativas para el control del oídio de la vid (Erysiphe necator Schw.) que puedan incorporarse en el concepto de manejo integrado de enfermedades en vides, utilizando productos de origen biológico, como el uso de bacterias antagónicas al hongo y extractos de plantas que tengan efecto inhibitorio de la enfermedad. El trabajo se realizó en un viñedo experimental de 5 años en los cultivares Thompson Seedless y Crimson Seedless ubicado en la Facultad de Ciencias Agronómicas de la Universidad de Chile.

Thesis (doctoral)--Christian-Albrechts-Universität zu Kiel, 1999.

Made available in DSpace on 2014-06-11T19:26:08Z (GMT). No. of bitstreams: 0 Previous issue date: 2008-07-25Bitstream added on 2014-06-13T20:54:10Z : No. of bitstreams: 1 demore_ps_me_jabo.pdf: 1002271 bytes, checksum: a27ac944079c717a8ba5366fe7e35456 (MD5) / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O oídio em soja, trata-se de uma doença praticamente presente em todos os paises produtores. Os marcadores moleculares microssatélites ou SSR (Simple Sequence Repeat) têm sido amplamente utilizados no processo de seleção assistida de genótipos de soja. Assim, o objetivo deste estudo foi obter marcadores microssatélites próximos ao gene de resistência ao oídio em soja. O estudo foi realizado em duas populações F2, oriundas de cruzamentos entre parentais contrastantes quanto à resistência ao oídio. Para o estudo, foram selecionados marcadores microssatélites a uma distância de até 42 cM ao redor do gene Rmd (resistência ao oídio). Utilizou-se o método de BSA (Bulked Segregant Analysis) na avaliação dos marcadores, para a comparação com a análise fenotípica das populações. Na análise foram utilizados dez iniciadores SSRs para as duas populações, sendo identificados quatro marcadores polimórficos para o cruzamento 1 (MGBR95-20937 x IAC-Foscarin 31) e três para o cruzamento 2 (MGBR 46/Conquista x EMBRAPA 48). Pela análise de Qui-quadrado da avaliação fenotípica, confirmou-se à segregação esperada (3:1) de um gene dominante condicionando a resistência. Os marcadores polimórficos também segregaram conforme o esperado (1:2:1) já que possuem natureza codominante. Para as populações 1 e 2, os melhores resultados foram obtidos com os marcadores Sat_366 e Sat_393, respectivamente, localizando-se a 9,41 e 12,45 cM de distância do gene, sendo considerados promissores na seleção assistida para resistência ao oidio em soja. / Powdery mildew in soybeans, it is a disease present in virtually all producing countries. The molecular markers microsatellites or SSR (Simple Sequence Repeat) have been widely used in the assisted selection of soybean genotypes. The objective of this study was to obtain microsatellites markers near the gene for resistance to powdery mildew in soybeans. The study was conducted in two populations F2, from crosses between contrasting parents about the resistance to powdery mildew. For the study, were selected microsatellites markers at a distance of 42 cM around the gene Rmd (resistance to powdery mildew). It was used the method of BSA (Bulked Segregant Analysis) in the evaluation of markers, for comparison with the phenotypic analysis of populations. In the analysis were used in ten initiators SSRs for the two populations, and identified four polymorphic markers for the crossing 1 (MGBR95-20937 x-IAC Foscarin 31) and three for the crossing 2 (EMBRAPA MGBR 46/Conquista x 48). For the analysis of chi-square of the phenotypic evaluation, it is confirmed the segregation expected (3:1) of a dominant gene conditioning the resistance. The polymorphic markers also segregation as expected (1:2:1) that have already codominante nature. For the populations 1 and 2, the best results were obtained with the markers Sat_366 and Sat_393, respectively, finding itself to 9.41 and 12.45 cM distance of the gene and are considered promising in assisted selection for resistance to soybean in powdery mildew.

Annotation: Powdery mildew Erysiphe alphitoides is one of the most important pathogenic fungus infecting pedunculate oak Quercus robur in Europe. As the identification of this species with morphologic markers is unreliable, the molecular markers development is necessary. In this work, we developed primers for PCR amplification of cytochrome b gene fragment. The results show high level of intraspecific variability of this mitochondrial gene and revealed the indistinct taxonomy relations within E. alphitoides s. lat group.