Дисертації з теми "Lactucine"

Les produits naturels (PNs) ont perdu en popularité depuis l'introduction des petites molécules synthétiques il y a plusieurs années. De nombreuses raisons expliquent ce choix, telles que les difficultés d'accès et d'approvisionnement, la complexité de la chimie des PNs et l'avènement de la chimie combinatoire. Cependant, les PNs offrent de nombreuses propriétés intéressantes par rapport aux molécules synthétiques conventionnelles, ce qui leur confère à la fois des avantages et des inconvénients dans le contexte de la recherche de principes actifs. En général, les PNs se caractérisent par un plus grand nombre de carbones sp3 et de centres stéréogènes, une grande diversité de squelettes et une grande complexité structurelle. La moitié des médicaments approuvés par la FDA depuis 1994 étant des PNs ou des dérivés hémisynthétiques, et étant donnée la récente stagnation de la recherche et du développement de nouveaux médicaments, il devient de plus en plus évident que les produits naturels doivent être utilisés dans le processus de découverte de médicaments en tant que source d'inspiration.Par conséquent, de nombreuses stratégies émergent aujourd'hui pour la construction de chimiothèques inspirées par la nature, selon les stratégies "top-down" et "bottom-up". Dans les approches "bottom-up", la complexité est créée à partir de réactifs simples. En revanche, les approches "top-down" consistent à apporter des modifications structurelles à un produit naturel déjà complexe.Le travail présenté décrit deux approches différentes pour enrichir la chimiothèque de notre unité de recherche avec des composés dérivés de PNs. Une stratégie hémisynthétique "top-down" a été planifiée pour obtenir des dérivés de la lactucine et de la 11β,13-dihydrolactucine, deux lactones sesquiterpéniques extraites des racines de chicorée. 36 dérivés esters ont été synthétisés en trois étapes (synthèse classique), ainsi que deux bibliothèques de dérivés aminés (en utilisant la synthèse parallèle). Tous les composés ont ensuite été testés contre Mycobacterium tuberculosis et certaines molécules ont montré des activités prometteuses (CMI < 1,2 µM).D'autre part, une stratégie "bottom-up" a permis la synthèse de deux analogues de l'antibiotique naturel hygromycine A. Pour cette approche nous sommes partis de réactifs simples disponibles dans le commerce et avons appliqué une stratégie de déaromatisation dans le processus de synthèse.L'ensemble de ces travaux nous a permis d'explorer un espace chimique plus large, d'accroître la diversité structurelle de notre chimiothèque et de découvrir de nouveaux "hits". Nous pourrons également ainsi identifier de nouvelles cibles antibactériennes
Natural products (NPs) have declined in popularity since the introduction of synthetic small molecules several years ago. Many are the reasons behind this choice, such as difficulties in access and supply, complexities of NP chemistry and the advent of combinatorial chemistry. However, NPs offer many interesting properties compared to conventional synthetic molecules, which confer both advantages and challenges for the drug discovery process. Usually, NPs are characterized by a higher number of sp3 carbons and stereogenic centres, large scaffold diversity and structural complexity. With half of the drugs approved by the FDA since 1994 being NPs or hemisynthetic derivatives and the recent stagnation in new drug research and development, it is becoming more and more evident that NPs should be reintroduced in the drug discovery process as a source of inspiration.Therefore, many strategies are now emerging for the construction of nature-inspired chemical libraries, such as “top-down” and “bottom-up” strategies. In “bottom-up” approaches, complexity is created starting from simple building blocks. On the other hand, “top-down” approaches are assumed to make structural modifications to an already complex NP.Our presented work describes two different approaches to enrich the chemical library of our research unit with NP-derived compounds. A “top-down” semisynthetic strategy was planned to obtain derivatives of lactucin and 11β,13-dihydrolactucin, two sesquiterpene lactones extracted from chicory roots. Thirty-six ester derivatives were synthesized in three steps (classical synthesis), together with two amine derivative libraries (using parallel synthesis). All the compounds were then tested against Mycobacterium tuberculosis and some promising hits were found (MICGFP < 1.2 μM). On the other hand, a “bottom-up” strategy allowed the synthesis of two analogues of the known natural antibiotic hygromycin A. This approach started from simple commercially available building blocks and employed a dearomatization strategy in the synthetic process.Together, we explored a broader chemical space, increased the structural diversity of our chemical library and discovered new potential antibacterial hits. Moreover, this work paves the way for the discovery of new antibacterial targets

The effect of temperature and duration of leaf wetness (DLW) on sporulation of Bremia lactucae was determined for lettuce cv. Ithaca. A single spore isolate of B. lactucae was produced and was identified using lettuce differential lines each of which had known gene for resistance. Potted lettuce plants were inoculated with the isolate and incubated at 15 C, at 100% relative humidity for 24 h and then at a relative humidity lower than 70%. Seven days later, the plants were submitted to five different temperatures (5, 10, 15, 20 and 25 C) and six durations of leaf wetness (4, 6, 8, 10, 12 and 14 h). The number of spores produced was determined at the end of each wet period. After 4 h of incubation no spores were observed at any of the temperatures. Highest number of spores was found at 10 and 15 C for more than 10 h of DLW. The rate of sporulation rapidly increased between 8 and 10 h for all the temperatures, including 25 C, where the amount of spores produced was very low. The observed number of spores was transformed into proportion of maximum sporulation (PMS) by providing each data by the maximum number of spores observed for each experiment. The Richards model was used to describe sporulation as a function of leaf wetness duration and the rate and maximum value expressed as a function of temperature. This approach resulted in a three-dimensional equation that explained 87% of the variation in the PMS. Spore viability was also estimated for each temperature and DLW. It was zero after 6 h of incubation and reached almost the maximum after 10 h for all the temperatures. The sporulation model was validated under field conditions and it predicted high, medium, low, or no sporulation in 8 out of 11 times.

3 pp.
This publication provides information on the development and management of Fusarium wilt of lettuce. Topics covered include the characteristics of the plant pathogen, disease development, and disease management considerations.

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Conseguiu-se avançar progênies F3 de alface-crespa resistentes a B. lactucae, até conseguirem estabilidade fenotípica nas gerações F5 e F6, por meio da seleção tanto dentro como entre famílias, com base na coloração verde-clara e alta crespicidade das folhas, tamanho grande, ausência de brotações laterais, tolerância ao pendoamento precoce e queima dos bordos. Diante do exposto, os objetivos deste trabalho consistiram em caracterizar as linhagens em geração F5: L1, L2, L3 e L4, e em geração F6: L5, L6, L7 e L8, em relação à resistência às raças SPBl:03, SPBl:04, SPBl:05, SPBl:06, SPBl:07, SPBl:09, SPBl:10, SPBl:11 e SPBl:12 de B. lactucae identificadas no Estado de São Paulo e à mistura delas, assim como selecionar linhagens superiores por seus caracteres agronômicos, adaptabilidade e estabilidade, em diferentes municípios e épocas de cultivo por meio da metodologia REML/BLUP, para possibilitar a recomendação das referidas linhagens. Para a caracterização da resistência, o delineamento experimental foi o inteiramente casualizado, em esquema fatorial de 10 X 9, determinado pelas nove raças de B. lactucae e a mistura delas, e pelas oito linhagens de alface-crespa e a cultivar Solaris suscetível, em quatro repetições. Cada raça e a mistura delas foram inoculadas isoladamente e avaliaram-se o índice de latência por meio de notas (notas de 0 a 12), dependendo do dia do aparecimento dos primeiros sinais do patógeno, e a porcentagem de plântulas com esporângios em 20 plântulas por parcela. Para avaliar a adaptabilidade e estabilidade das linhagens, o delineamento experimental utilizado foi de blocos ao acaso, em esquema fatorial 10 x 6 x 2, sendo 10 genótipos (oito linhagens, Vanda e Vera), seis áreas de produtores de alface nos municípios de Monte Alto, São Simão, Aramina, Mogi das Cruzes, Biritiba-Mirim e Salesópolis, e duas épocas de cultivo de Outono-Inverno de 2014 e Primavera- Verão de...
It was possible to advance progenies F3 of crisp leaf lettuce resistant to B. lactucae, to achieve phenotypic stability in the generations F5 and F6, by selecting individuals from both, within and between families, based on the bright green color and high roughness of the leaves, large size, absence of side buds and tolerance to bolting and tip burn. Thus, the aims of this work consisted of characterize the lineages in generations F5: L1, L2, L3, L4, and in generations F6: L5, L6, L7, L8, related to their resistance to the races SPBl:03, SPBl:04, SPBl:05, SPBl:06, SPBl:07, SPBl:09, SPBl:10, SPBl:11 and SPBl:12 of B. lactucae identified in São Paulo State and to their mixture, as well as to select lineages superiors for their agronomic characters, adaptability and stability, in different municipalities and cropping seasons using the REML/BLUP methodology, in order to enable the recommendation of such lineages. To characterize the resistance, the experimental model was completely randomized in a factorial design of 10 X 9, determined by the nine races of B. lactucae and their mixture, and the eight lineages of crisp leaf lettuce and the susceptible cultivar Solaris, with four replications. Each race and their mixture were separately inoculated and the index latency was evaluated by means of notes (notes from 0 to 12) depending on the day of appearance of the first signs of pathogen and the percentage of seedlings with sporangia, in 20 seedlings per plot. To assess lineages adaptability and stability, randomized blocks in a factorial design of 10 x 6 x 2 was used as experimental model, being 10 genotypes (eight lineages, Vanda and Vera), six areas of lettuce producers in the municipalities of Monte Alto, São Simão, Aramina, Mogi das Cruzes, Biritiba-Mirim and Salesópolis, and two cropping periods autumn-winter 2014 and spring-summer 2014-2015, with four replications; it was assessed the volume (cm3/plant), the total and commercial ...

Orientador: Leila Trevisan Braz
Banca: Rita de Cassia Panizzi
Banca: Renata Castoldi
Banca: Rinaldo Cesar de Paula
Banca: Pablo Forlan Vargas
Resumo: Conseguiu-se avançar progênies F3 de alface-crespa resistentes a B. lactucae, até conseguirem estabilidade fenotípica nas gerações F5 e F6, por meio da seleção tanto dentro como entre famílias, com base na coloração verde-clara e alta crespicidade das folhas, tamanho grande, ausência de brotações laterais, tolerância ao pendoamento precoce e queima dos bordos. Diante do exposto, os objetivos deste trabalho consistiram em caracterizar as linhagens em geração F5: L1, L2, L3 e L4, e em geração F6: L5, L6, L7 e L8, em relação à resistência às raças SPBl:03, SPBl:04, SPBl:05, SPBl:06, SPBl:07, SPBl:09, SPBl:10, SPBl:11 e SPBl:12 de B. lactucae identificadas no Estado de São Paulo e à mistura delas, assim como selecionar linhagens superiores por seus caracteres agronômicos, adaptabilidade e estabilidade, em diferentes municípios e épocas de cultivo por meio da metodologia REML/BLUP, para possibilitar a recomendação das referidas linhagens. Para a caracterização da resistência, o delineamento experimental foi o inteiramente casualizado, em esquema fatorial de 10 X 9, determinado pelas nove raças de B. lactucae e a mistura delas, e pelas oito linhagens de alface-crespa e a cultivar Solaris suscetível, em quatro repetições. Cada raça e a mistura delas foram inoculadas isoladamente e avaliaram-se o índice de latência por meio de notas (notas de 0 a 12), dependendo do dia do aparecimento dos primeiros sinais do patógeno, e a porcentagem de plântulas com esporângios em 20 plântulas por parcela. Para avaliar a adaptabilidade e estabilidade das linhagens, o delineamento experimental utilizado foi de blocos ao acaso, em esquema fatorial 10 x 6 x 2, sendo 10 genótipos (oito linhagens, Vanda e Vera), seis áreas de produtores de alface nos municípios de Monte Alto, São Simão, Aramina, Mogi das Cruzes, Biritiba-Mirim e Salesópolis, e duas épocas de cultivo de Outono-Inverno de 2014 e Primavera- Verão de...
Abstract: It was possible to advance progenies F3 of crisp leaf lettuce resistant to B. lactucae, to achieve phenotypic stability in the generations F5 and F6, by selecting individuals from both, within and between families, based on the bright green color and high roughness of the leaves, large size, absence of side buds and tolerance to bolting and tip burn. Thus, the aims of this work consisted of characterize the lineages in generations F5: L1, L2, L3, L4, and in generations F6: L5, L6, L7, L8, related to their resistance to the races SPBl:03, SPBl:04, SPBl:05, SPBl:06, SPBl:07, SPBl:09, SPBl:10, SPBl:11 and SPBl:12 of B. lactucae identified in São Paulo State and to their mixture, as well as to select lineages superiors for their agronomic characters, adaptability and stability, in different municipalities and cropping seasons using the REML/BLUP methodology, in order to enable the recommendation of such lineages. To characterize the resistance, the experimental model was completely randomized in a factorial design of 10 X 9, determined by the nine races of B. lactucae and their mixture, and the eight lineages of crisp leaf lettuce and the susceptible cultivar Solaris, with four replications. Each race and their mixture were separately inoculated and the index latency was evaluated by means of notes (notes from 0 to 12) depending on the day of appearance of the first signs of pathogen and the percentage of seedlings with sporangia, in 20 seedlings per plot. To assess lineages adaptability and stability, randomized blocks in a factorial design of 10 x 6 x 2 was used as experimental model, being 10 genotypes (eight lineages, Vanda and Vera), six areas of lettuce producers in the municipalities of Monte Alto, São Simão, Aramina, Mogi das Cruzes, Biritiba-Mirim and Salesópolis, and two cropping periods autumn-winter 2014 and spring-summer 2014-2015, with four replications; it was assessed the volume (cm3/plant), the total and commercial ...
Doutor

Dissertação (mestrado)—Universidade de Brasília, Instituto de Ciências Biológicas, Departamento de Fitopatologia, 2008.
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Sintomas e sinais do míldio foram observados e coletados em plantas de alface (Lactuca sativa) no Distrito Federal. Foi observado o estágio assexual do patógeno (Bremia lactucae). Descrições detalhadas, com observações morfológicas e medidas morfométricas de microscopia óptica do estágio assexual foram realizadas. A patogenicidade do míldio proveniente do Núcleo Rural da Vargem Bonita e seu período de latência, em quatro diferentes estádios de desenvolvimento de cultivares comerciais de alface foram estudados por inoculação artificial em ensaios realizados na Estação Biológica da Universidade de Brasília. Quando observado o “ranking” de suscetibilidade das cultivares nos quatro diferentes estádios de desenvolvimento para o período de latência, observou-se diferentes níveis de resistência entre as cultivares. Além disso, observou-se que a resistência parece aumentar com o desenvolvimento fenológico do hospedeiro. O “ranking” relativo das cultivares não foi afetadopelo estádio de desenvolvimento. _________________________________________________________________________________ ABSTRACT
Symptoms and signs of downy mildew were observed and collected in lettuce plants (Lactuca sativa) in Distrito Federal, Brazil. Only the asexual stage of the pathogen (Bremia lactucae) was observed. Detailed descriptions, including fungal morphology and morphometrics were performed with light microscopy. The pathogenicity of downy mildew, collected in Núcleo Rural da Vargem Bonita, as well as it latent period in four phenological stages of lettuce commercial cultivars were studied by artificial inoculation in experiments conducted at Biological Station of University of Brasilia - UnB. Latent period was useful to separate cultivars by resistance level. In addition, results indicated that genetic resistance to downy mildew in lettuce seems to increase with plant development. The relative ranking of susceptibility among cultivars remained constant in all four phenological stages.

Mestrado em Engenharia Agronómica - Instituto Superior de Agronomia
Neste trabalho estabeleceu-se um teste molecular, rápido e fácil, para detectar Sclerotinia sclerotiorum, responsável pela doença da podridão branca da alface, em folhas assintomáticas. Desenharam-se primers específicos (SSM) para ampliar sequências parciais da região ITS de S. sclerotiorum, e utilizaram-se em reacções de PCR com alfaces infectadas. A análise das sequências obtidas indicou homologia significativa com sequências de S. sclerotiorum já conhecidas. Usaram-se os primers SSM com alfaces infectadas com Bremia lactucae e Botrytis cinerea verificando-se que apenas ampliavam fragmentos nas amostras com B. cinerea. A sequenciação destes fragmentos sugeriu a presença de sequências parciais de B. cinerea e também de Alternaria sp., além de S. sclerotiorum. Os fragmentos de S. sclerotiorum e Alternaria sp., com tamanho semelhante, poderão distinguir-se pela digestão dos produtos de PCR com RsaI. Alguns isolados de B. cinerea poderão ser discriminados com HindIII, mas não todos, já que outros apresentam sequências idênticas às identificadas em S. sclerotiorum. Estabeleceu-se uma reacção de PCR multiplex conjugando os primers SSM com os primers Br, específicos para ampliar sequências parciais da região ITS de B. lactucae, para detectar simultaneamente os patogénios referidos. A posterior digestão enzimática com RsaI e HindIII permite discriminar alguns daqueles patogénios
N/A

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Lettuce is the highest consumed vegetable in Brazil. It can however be severely affected by the fungus Sclerotinia sclerotiorum, Sclerotinia minor and Bremia lactucae; diseases considered to be ground and aerial part, respectively. The objective of this experiment was to evaluate the effect of different mulches, fungicides and one foliar fertilizer on the incidence of lettuce drop and severity of downy mildew in crisp type lettuce cultivars. Two experiments were carried out on the Fazenda Escola Capão da Onça, within the Universidade Estadual de Ponta Grossa, during the summer and winter season, with natural conditions of fungus infection. The experimental design was a completty randomized blocks with treatments arranged in split plots. This involved the combination of 3 types of mulches as black polypropylene, oats straw and bare ground. It also included two cultivars of crisp type lettuce Vera and Isabela. And four methods of control were used: 3 fungicides, metalaxyl + chlorothalonil, fenamidone and procimidone, a foliar fertilizer (Hortifós PK), and a control. For each treatment four repetitions were performed. The doses of each product applicateded were: metalaxyl + chlorothalonil 100 g a.i/100 L H2O, fenamidone 15 g a.i/100 L H2O, procimidone 75 g a.i/100 L H2O and foliar fertilizer 135 dosage of 135 g P2O5 + 100 g K2O/100 L H2O in the first application after seedling transplanting. In the subsequent applications were added 67.5 g P2O5 and 67.5 g K2O/100 L H2O and then continued until a maximum of 405 g P2O5 + 405 g K2O/100 L H2O was reached. This occurred in the fifth spraying, and this dosage remained until crop harvest. The first spray was performed at transplanting with subsequent applications occurring at intervals of 7 days. The incidence of lettuce drop was evaluated during the rosette phase and at the time of harvest, in 24 plants per parcel. The severity and the percentage of leaves with downy mildew symptoms and the plant fresh weight and biomass were evaluated in 8 plants per parcel at the time of harvest. Harvest occurred 45 and 57 days after the transplanting, for the summer and winter season experiments, respectively. For both seasons the incidence of lettuce drop was higher in the harvest phase than the phase of rosette. Covering with oat straw and black polypropylene reduced the lettuce drop in the summer season. Black polypropylene also reduced the percentage of leaves with downy mildew symptoms. The fungicide procymidone provided the best control of lettuce drop, while metalaxyl + chlorothalonil and foliar fertilizer showed better control of downy mildew. The Vera cultivar was revealed to be more susceptible to lettuce drop and downy mildew, however, it presented greater plant fresh weight and biomass than Isabela cultivar in the summer season. Keywords: ground covering, methods of control, Lactuca sativa L, Bremia lactucae and Sclerotinia sclerotiorum.
A alface é a hortaliça folhosa mais consumida pelos brasileiros. O seu cultivo é severamente afetado pelos fungos Sclerotinia sclerotiorum, Sclerotinia minor, e Bremia lactucae, doenças consideradas de solo e parte aérea, respectivamente. Com o objetivo de avaliar o efeito de coberturas de solo, diferentes fungicidas e um fertilizante foliar sobre a incidência da podridão de Sclerotinia e sobre a severidade do míldio em cultivares de alface (Lactuca sativa L.) tipo crespa, foram realizado dois experimentos na Fazenda Escola Capão da Onça pertencente a Universidade Estadual de Ponta Grossa no município de Ponta Grossa, PR nas estações de verão e inverno em condições naturais de infecção. O delineamento experimental utilizado foi inteiramente ao acaso, onde os tratamentos foram arranjados em parcela sub-subdivididas, provenientes da combinação de 3 sistemas de cultivo (polipropileno preto, palhada de aveia e solo nú), duas cultivares de alface tipo crespa (Vera e Isabela) e 4 metódos de controle (3 fungicidas – metalaxyl + chlorothalonil, fenamidone e procimidone; um fertilizante foliar - Hortifós PK) além da testemunha, com 4 repetições. As doses de cada produto foram: metalaxyl + chlorothalonil 100 g i.a./100 L H2O, fenamidone 15 g i.a./100 L H2O, procimidone 75 g i.a./100 L H2O e fertilizante foliar na dosagem de 135 g P205 + 135 g K2O/100 L H2O na primeira aplicação, logo após o transplantio das mudas. Nas aplicações seguintes foram adicionados 67,5 g P205 e 67,5 g K2O/100 L H2O até que se atingisse o máximo de 405 g P205 + 405 g K2O/100 L H2O, o que aconteceu na quinta pulverização, permanecendo com essa dosagem até a colheita. As aplicações tiveram início no transplantio das mudas e as subseqüentes em intervalos de 7 dias. Avaliou-se incidência de podridão de Sclerotinia na fase de roseta e no momento da colheita da cultura da alface em 24 plantas/parcela, severidade e porcentagem de folhas com sintomas de míldio, massa fresca e biomassa em 8 plantas/parcela. A colheita foi realizada 45 e 57 dias após o transplantio, para o experimento conduzido na estação de verão e inverno, respectivamente. A incidência de podridão de Sclerotinia foi maior na fase de colheita quando comparada com a fase de roseta para ambas estações. A palha de aveia e polipropileno preto reduziram a podridão de Sclerotinia na estação de verão, sendo que esta última cobertura de solo também proporcionou redução da porcentagem do número de folhas com sintomas de míldio. O fungicida procimidone proporcionou o melhor controle da podridão de Sclerotinia e os produtos metalaxyl + chlorothalonil e o fertilizante foliar foram os que melhor controlaram o míldio. A variedade Vera mostrou-se mais suscetível a podridão de Sclerotinia e ao míldio, entretanto, apresentou maior massa fresca e biomassa que a variadade Isabela na estação de verão.

Downy mildew of lettuce caused by Bremia lactucae is a serious disease resulting in yield loss. The population structure of the pathogen in the UK is poorly understood. This PhD project concentrated on developing molecular markers to differentiate the genotypic variation of B. lactucae populations, with the aim of improving methods to investigate lettuce - Bremia interactions. Thirty-seven B. lactucae isolates (including single-spore and new field isolates) were collected and characterized for virulence using the conventional International Bremia Evaluation Board (IBEB) differential set. Microsatellite markers (SSR, ISSR) were investigated for Bremia race specific marker development. Three isolates of B. lactucae were characterized by ISSR (inter simple sequence repeat) primers, although the polymorphic DNA could not be cloned in this project due to the highly variable results of the ISSR process. Some microsatellite repeats were found in B. lactucae isolates sequences that amplified by Plasmopara viticola (grape downy mildew) SSR markers. The development of Simple Sequence Repeat (SSR) markers from Bremia genomic DNA was not successful, which might result from the primers used being unsuitable for Bremia microsatellite enrichment. Bremia specific ITS primers were used for quantitative PCR. RxLR primers obtained from UC Davis (USA) were tested using the collection of B. lactucae isolates. RxLR1 primers distinguished between isolates BL801 and BL806. Eight SNPs were identified in three isolates amplified by RxLR5. No polymorphism was observed on the gel for the remaining RxLR primers on single spore races. Unrefined field isolates showed more polymorphisms on the gel than single spore isolates. The phenotypic differences between these two isolates have been identified by the IBEB differential set. Microscopy and qPCR quantification were used to investigate the compatible and incompatible interactions. The results suggest that BL801 is more virulent than BL806, as more infection structures were observed in IBEB resistant cultivars. Results of qPCR and spore count/unit weight of cotyledons showed that BL801 and BL806 were significantly different. The qPCR quantification results from 4 and 5 dpi were correlated with the spore count/unit weight of cotyledons. Although further work is required to develop race specific markers, the methods used in this project demonstrate the potential use of molecular markers to investigate lettuce - Bremia interactions.

Des nouvelles résistances aux agents pathogènes introgressées dans les plantes cultivées sont fréquemment contournées dans différents pathosystèmes, engendrant des épidémies et des pertes économiques. En conséquence, la compréhension des stratégies évolutives impliquées dans l'adaptation des populations pathogènes est nécessaire pour améliorer la gestion durable des résistances. Bremia lactucae, agent pathogène de la laitue est un organisme diploïde, hétérothallique avec des cycles de reproduction sexuée et asexuée. Cet oomycète est soumis aux fortes pressions de sélection exercées par des gènes de résistance de la plante hôte. Sous cette pression de sélection, les populations de B. lactucae ont montré une rapide adaptation aux résistances hôtes qui se sont donc avérées peu durables. L'étude de la structure génétique d'un agent pathogène peut permettre de comprendre les mécanismes évolutifs impliqués dans le contournement des résistances variétales. Ainsi, une étude de la structure génétique (neutre et potentiellement soumise à sélection) des populations de B. lactucae en France a été conduite afin d'identifier les forces évolutives impliquées dans le contournement des résistances de l'hôte et de déterminer l'influence des pressions de sélection des gènes de résistance de la plante hôte sur cette structure. J'ai validé 12 microsatellites, marqueurs moléculaires neutres, pour analyser la structure génétique de B. lactucae en France. Plus de 800 isolats ont été prélevés dans les plus importants bassins de production de la plante hôte, Lactuca sativa. Ces isolats ont été prélevés sur différentes variétés regroupées selon leur combinaison de gènes de résistance. Par ailleurs, une prospection dans le compartiment sauvage a permis d'échantillonner des isolats de B. lactucae sur la plante hôte adventice L. serriola. Le polymorphisme de plusieurs effecteurs candidats à motif RxLR, a été étudié dans différentes populations de Bremia. La faible diversité génétique et l'excès d'hétérozygotie observés sont en faveur d'une reproduction clonale mais de rares évènements de reproduction sexuée sont également suggérés par les résultats. Par ailleurs, la faible différenciation génétique entre populations suggère des flux de gènes importants à l'échelle des régions. Des flux de gènes ont également été mis en évidence entre le pathosystème du compartiment sauvage et le pathosystème du compartiment cultivé évoquant un possible rôle de réservoir génétique des plantes hôtes adventices. Une structuration des populations en plusieurs lignées clonales résultant de la pression de sélection des gènes de résistance des cultivars est également indiquée par l'analyse des résultats des marqueurs neutres et sélectionnés. La caractérisation de la structure des populations de B. lactucae nous permet de mettre en évidence le fort potentiel évolutif (flux de gènes importants, système de reproduction mixe, sélection de mutants et de migrants par les gènes de résistance) de B. lactucae expliquant la rapide adaptation aux résistances hôtes. Ainsi, nous pouvons suggérer des pistes de gestion des gènes de résistance comme favoriser l'utilisation de résistances quantitatives et l'utilisation de cultures d'association afin d'améliorer la durabilité des résistances.

The aim of this Diploma thesis was to assess the cultivar tolerance of a chosen assortment of butterhead lettuce, leaf lettuce, iceberg lettuce and cabbage to fungal pathogens (Bremia lactucae, Fusarium oxysporum f. sp. conglutinans). The experiment was conducted in the laboratory conditions of the gardening department FAPPZ of the Czech University of Agriculture in growth chambers. The lettuce experiments were done in given conditions with the temperatures of 18 - 20 °C under the regime 12 hrs. of light and 12 hrs. of darkness. The cabbage experiments were conducted in given conditions under the regime 12 hrs. of light and 12 hrs. of darkness with the temperatures of 14 °C during the day and 12 °C in the night. The lettuce plants were planted in dishes on fine sand. All experiments had infected and control variants. After emergence, the experimental plants were inoculated with variants of the selected pathogen. The plants were observed in 2 - 3 day intervals throughout the whole experiment. The experiment was assessed using the modified standard method according to Pawelec et al. (2006). A modified method with a percentage scale was used to assess the lettuce experiments. Cabbage experiments were evaluated with a scale of points ranging from 0 to 9 according to the percentage of infection on the plants. The results of the methodology were determined using the program Statistica 12. In the experiments evaluating the variety tolerance of a chosen assortment leaf lettuce and iceberg lettuce to the pathogen Bremia lactucae Bl:31, a statistically significant difference in the sensitivity to this pathogen was established. The results show that the most resistant varieties are Tarzan, Stamir, Adinal a Verala. The greatest sensitivity to the pathogen Bl:31 has a variety "Dětenická Atrakce" then Traper and Nikolaj. Effect of seed treatment with hot water (HWT) to suppress pathogen B. lactucae was statistically significant in the varieties "Dětenická Atrakce" (10% decrease) and Dubáček (7% decrease) compared with untreated infectious variant with HWT. In the sensitivity evaluation of varieties and lines of white cabbage was a statistically significant difference in infecting of pathogen Fusarium oxysporum f. Sp. conglutinans breed 1. The most significantly susceptible to the pathogen Foc was variety Pourovo late-season (degree of assault after inoculation was 6.52 points).

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
Os objetivos do trabalho foram identificar raças de Bremia lactucae agente causal do míldio no período de 2006 a 2007, na região produtora de alface do Estado de São Paulo; obter linhagens de alface-crespa resistentes. Os experimentos de campo foram realizados no Setor de Olericultura e Plantas Aromático-Medicinais e no Laboratório de Melhoramento Genético de Hortaliças, do Departamento de Produção Vegetal da UNESP-FCAV, Câmpus de Jaboticabal-SP. Foram coletados 36 isolados de B. lactucae em regiões produtoras do Estado de São Paulo, que permitiram identificar três novos comportamentos do fungo, dando origem a três novas raças: SPBl:02 (63/31/19/00), SPBl:03 (63/63/19/00) e SPBl:04 (63/63/03/00). Os genes Dm-14, Dm- 17, Dm-18, Dm-36, Dm-37 e Dm-38 conferem resistência a essas novas raças identificadas. As linhagens JAB 4-2-4, JAB 4-13-7 e JAB 4-13-9 apresentaram os melhores resultados de padrão de plantas e com resistência a B. lactucae. Os genótipos ‘Amanda’, ‘Solaris’, ‘Vanda’, ‘Vera’, JAB 4-13-7 e ‘Malice’ apresentaram melhores desempenhos das características agronômicas no inverno. Para as condições de verão, JAB 4-13-7 e ‘Solaris’ apresentaram melhores desempenhos das características agronômicas. JAB 4-13-7 é o mais indicado para o cultivo de inverno pelo bom desempenho e resistência às raças de B. lactucae. ‘Malice’ apresentou o pior comportamento produtivo, apesar da resistência às quatro raças de B. lactucae avaliadas
The aims of the study were to verify the emergence of races of B. lactucae casual agent of downy mildew in the period from 2006 to 2007 in lettuce-producing region in the State of São Paulo; to obtain lines lettuce with resistance to race SPBI:01; the performance of JAB 4-13-7 before the commercials cultivars as an option to allow for the cultivation of this genotype for both evaluation periods, in the conditions of Jaboticabal and check the resistance of lettuce genotypes to four races of Bremia lactucae. The field experiments were accomplished at the Section of Vegetables Crops and Aromatic-Medicinal Plants and in the Laboratory of Vegetable Breeding of the Department of Vegetable Production from the São Paulo State University at Jaboticabal (UNESP). The monitoring of B. lactucae allowed to identify three new behaviors of the fungus, resulting in three new races SPBI:02 (63/31/19/00), SPBI:03 (63/63/19/00) SPBI:04 (63/63/03/00). The genes Dm-14, Dm-17, Dm-18, Dm-36, Dm-37 and Dm-38, confer resistance to these new identified races. The lines JAB 4-2-4, JAB 4-13-7 and JAB 4-13-9 achieved the best results of standart plants with resistance to B. lactucae. The genotypes ‘Amanda’, ‘Solaris’, ‘Vanda’, ‘Vera’, JAB 4-13-7 and ‘Malice’ showed the best performances of agronomic traits in winter. For summer conditions, JAB 4-13-7 and ‘Solaris’ presented better performances of agronomic traits. The JAB 4-13-7 is the most suitable for the winter cultivation by good performance and resistance for the races of B. lactucae. ‘Malice’ had the worst productive performance, despite the resistance to the four races of B. lactucae evaluated

Orientador: Leila Trevizan Braz
Coorientador: Tagli Dalpian
Coorientador: Margarete Camargo
Banca: Arlete Marchi Tavares de Melo
Banca: Antonio Ismael Inácio Cardoso
Banca: Modesto Barreto
Banca: Érika Auxiliadora Giacheto Scaloppi
Resumo: Os objetivos do trabalho foram identificar raças de Bremia lactucae agente causal do míldio no período de 2006 a 2007, na região produtora de alface do Estado de São Paulo; obter linhagens de alface-crespa resistentes. Os experimentos de campo foram realizados no Setor de Olericultura e Plantas Aromático-Medicinais e no Laboratório de Melhoramento Genético de Hortaliças, do Departamento de Produção Vegetal da UNESP-FCAV, Câmpus de Jaboticabal-SP. Foram coletados 36 isolados de B. lactucae em regiões produtoras do Estado de São Paulo, que permitiram identificar três novos comportamentos do fungo, dando origem a três novas raças: SPBl:02 (63/31/19/00), SPBl:03 (63/63/19/00) e SPBl:04 (63/63/03/00). Os genes Dm-14, Dm- 17, Dm-18, Dm-36, Dm-37 e Dm-38 conferem resistência a essas novas raças identificadas. As linhagens JAB 4-2-4, JAB 4-13-7 e JAB 4-13-9 apresentaram os melhores resultados de padrão de plantas e com resistência a B. lactucae. Os genótipos 'Amanda', 'Solaris', 'Vanda', 'Vera', JAB 4-13-7 e 'Malice' apresentaram melhores desempenhos das características agronômicas no inverno. Para as condições de verão, JAB 4-13-7 e 'Solaris' apresentaram melhores desempenhos das características agronômicas. JAB 4-13-7 é o mais indicado para o cultivo de inverno pelo bom desempenho e resistência às raças de B. lactucae. 'Malice' apresentou o pior comportamento produtivo, apesar da resistência às quatro raças de B. lactucae avaliadas
Abstract: The aims of the study were to verify the emergence of races of B. lactucae casual agent of downy mildew in the period from 2006 to 2007 in lettuce-producing region in the State of São Paulo; to obtain lines lettuce with resistance to race SPBI:01; the performance of JAB 4-13-7 before the commercials cultivars as an option to allow for the cultivation of this genotype for both evaluation periods, in the conditions of Jaboticabal and check the resistance of lettuce genotypes to four races of Bremia lactucae. The field experiments were accomplished at the Section of Vegetables Crops and Aromatic-Medicinal Plants and in the Laboratory of Vegetable Breeding of the Department of Vegetable Production from the São Paulo State University at Jaboticabal (UNESP). The monitoring of B. lactucae allowed to identify three new behaviors of the fungus, resulting in three new races SPBI:02 (63/31/19/00), SPBI:03 (63/63/19/00) SPBI:04 (63/63/03/00). The genes Dm-14, Dm-17, Dm-18, Dm-36, Dm-37 and Dm-38, confer resistance to these new identified races. The lines JAB 4-2-4, JAB 4-13-7 and JAB 4-13-9 achieved the best results of standart plants with resistance to B. lactucae. The genotypes 'Amanda', 'Solaris', 'Vanda', 'Vera', JAB 4-13-7 and 'Malice' showed the best performances of agronomic traits in winter. For summer conditions, JAB 4-13-7 and 'Solaris' presented better performances of agronomic traits. The JAB 4-13-7 is the most suitable for the winter cultivation by good performance and resistance for the races of B. lactucae. 'Malice' had the worst productive performance, despite the resistance to the four races of B. lactucae evaluated
Doutor

Orientador: Leila Trevisan Braz
Resumo: O míldio da alface, causado por Bremia lactucae, é uma das doenças mais importantes dessa cultura, causando inúmeros prejuízos. O surgimento de novas raças torna seu controle desafio constante para os melhoristas de alface, devido à necessidade de sempre buscar novos genes de resistência para o controle da doença. O objetivo deste trabalho foi monitorar as raças de B. lactucae presentes em regiões produtoras de alface do estado de São Paulo, no ano de 2014, e avaliar a sua distribuição no período de 2003 a 2014, a fim de dar suporte ao programa de melhoramento genético de alface da UNESP - FCAV. Amostras de folhas com sintomas de B. lactucae foram coletadas em regiões produtoras do estado de São Paulo, a partir das quais os esporângios foram multiplicados na cultivar Solaris, para então iniciar a fase de diferenciação das raças. A avaliação das diferenciadoras foi realizada quando houve esporulação na cultivar suscetível Green Towers, sendo atribuídos sinais +, (+), - e (-), de acordo com a esporulação observada. Já em relação à distribuição de B. lactucae no período de 2003 a 2014, foi realizado o levantamento da frequências das raças e dos fatores de virulência identificados nesse período. Em 2014, encontraram-se seis raças: SPBl:13, SPBl:14, SPBl:15, SPBl:16, Bl:21 e SPBl:01. No total, foram identificadas 17 raças no estado de São Paulo, entre 2003 e 2014, nos 33 municípios avaliados. A SPBl:01, ocorreu em 35% dos 514 isolados avaliados. Dos 19 fatores de virulência avalia... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: The downy mildew of lettuce caused by Bremia lactucae is one of the most important diseases of this crop, causing major damage during the winter season. The occurrence of new races is a constant challenge for lettuce breeders, due the need of always search for new resistance genes for genetic disease control. The aim of this study was to survey the B. lactucae races, in lettuce producing regions in Sao Paulo State in 2014, and its distribution from 2003 to 2014, to support the lettuce genetic breeding program of the UNESP - FCAV. Leaf samples containing symptoms of B. lactucae were collected from producing regions of Sao Paulo State, from which the sporangia were multiplied in Solaris cultivar, and thereafter it was started the stage of races differentiation. The assessment was performed when the differentiating susceptible cultivar Green Towers showed sporulation, being attributed the signs +, (+) - and (-), according to sporulation observed. To assess the distribution of B. lactucae from 2003 to 2014, it was performed a study of the frequency of the races and virulence factors identified in this period. In 2014, it was found six races - SPBl:13, SPBl:14, SPBl:15, SPBl:16, Bl:21 e SPBl:01. In total, 17 races were identified in Sao Paulo State between 2003 and 2014, in 33 municipalities assessed. The race SPBl:01 occurred in 35% of the 514 isolates tested. From the 19 virulence factors assessed, the ones that did not occur in the population of B. lactucae, in Sao Paulo state,... (Complete abstract click electronic access below)
Mestre

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Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)
Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
A alface é a hortaliça folhosa de maior consumo no Brasil, no entanto a dificuldade em produzi-la vem aumentando, principalmente pela infestação das áreas de produção por Bremia lactucae, sendo necessário o monitoramento anual para utilização ou desenvolvimento de cultivares resistentes. Diante do exposto, os objetivos do presente trabalho foram: identificação de raças de B. lactucae no Estado de São Paulo, durante os anos de 2008, 2009 e 2010 e obtenção de linhagens de alface crespa resistentes às raças SPBl:01, SPBl:02, SPBl:03, SPBl:04, SPBl:05, SPBl:06 e SPBl:07. Na primeira fase do estudo foram coletadas nos meses de agosto, setembro e outubro de 2008; agosto e setembro de 2009; e junho e julho de 2010 folhas de alface com esporângios de B. lactucae nos principais municípios produtores de alface do Estado de São Paulo. Após a multiplicação dos esporângios na cultivar suscetível Solaris, com posterior inoculação nas diferenciadoras, realizaram-se as avaliações, no mesmo dia do aparecimento da primeira esporulação na cultivar suscetível Cobham Green (DM 0). Na segunda fase do estudo realizou-se o cruzamento entre a cultivar Argeles e a linhagem JAB 4-13-7, bem como autofecundações, até a obtenção da geração F3, quando realizou-se o teste de resistência. Os dados permitiram concluir que: três novos códigos “Sextet” para Bremia lactucae ocorreram no Estado de São Paulo: 63/63/33/00, 63/63/02/00 e 63/31/03/00, denominados raças SPBl:05, SPBl:06 e SPBl:07, respectivamente; recomenda-se a utilização dos genes R-17, R-18 e R-38 como fontes de resistência ao míldio para o desenvolvimento de cultivares no Estado de São Paulo; e 19 progênies resistentes à B. lactucae, raças SPBl:01, SPBl:02, SPBl:03, SPBl:04, SPBl:05, SPBl:06 e SPBl:07, apresentaramse homozigotas, e podem ser avançadas e avaliadas no processo de seleção
Lettuce is the most consumed leafy vegetable in Brazil. Its production, nonetheless, is meeting with increasing difficulties mainly due to the infestation of producing areas with Bremia lactucae. Thus, the annual monitoring of B. lactucae races becomes of essential importance to use or development of resistant cultivars. The objectives of this work were identification of new races of B. lactucae occurring during the years of 2008, 2009 and 2010 in the state of São Paulo, Brazil, and development of resistant lettuce cultivars to races SPBl:01, SPBl:02, SPBl:03, SPBl:04, SPBl:05, SPBl:06 and SPBl:07. In the first phase of experiment in August, September, and October of 2008; August and September of 2009; and June and July of 2010 lettuce leaf samples containing B. lactucae sporangia were collected in some of the lettuce producing areas of the state of São Paulo. After a multiplication procedure in which the susceptible lettuce cultivar ‘Solaris’ was used, the B. lactucae sporangia were inoculated in differentiating lettuce cultivars. Evaluations of the plants response were performed on the same day when plants of the susceptible Cobham Green (DM 0) cultivar showed signs of the first sporulation. In the second phase of experiment made the crossing between Argeles cultivar and JAB 4-13-7 line, and self pollination, to obtain the F3 generation, when made the resistant test. The data have identified: three new codes “Sextet” in the state of São Paulo: 63/63/33/00, 63/63/02/00 e 63/31/03/00. SPBl:05, SPBl:06 and SPBl:07 were the denomination proposed to races, respectively. The genes R-17, R-18, and R-38 are suggested to be used as source of resistance in the lettuce cultivars to state of São Paulo; and 19 resistant progenies to B. lactuae, races SPBl:01, SPBl:02, SPBl:03, SPBl:04, SPBl:05, SPBl:06 and SPBl:07, shown homozygous, and they can be advanced and evaluated in the selection process

The research presented in this dissertation involves the study of two systems in order to analyze and develop resistance in plants to pathogens. The first study considered the interaction between lettuce (Lactuca sativa L.) and Plasmopara lactucae-radicis, a recently described casual agent of downy mildew. This unique fungus is the only known casual agent of downy mildew that is restricted to the roots of its host. Thirty-eight lettuce cultivars were screened for resistance to P. lactucae-radicis. Two-wk-old lettuce plants grown hydroponically were challenged with this fungus and evaluated 2 and 3 wk after inoculation for resistance. Root necrosis and production of sporangia on roots was considered a susceptible reaction. Five cultivars were determined to be resistant to this fungus. Resistant cultivars, however, were colonized by the fungus but did not support the production of sporangia on roots. Data from F2 and F3 progenies demonstrated that resistance was conferred by a single recessive allele (plr). Fungal infection of susceptible plants resulted in significant decreases in fresh root and shoot weights and leaf number compared to decreases associated with infection of resistant plants. RFLP probes have been identified for mapping the plr gene. The second topic of this dissertation dealt with the development of transgenic tobacco plants that express a chimeric gene that consisted of the 35-S promoter and the coat protein gene of Beet Curly Top Virus, a member of the geminivirus group. Expression of viral coat protein genes in plants has resulted in resistance to the virus from which the coat protein gene was obtained. This type of "coat protein-mediated" resistance has not been demonstrated for any geminivirus. Tobacco leaf discs were inoculated with an Agrobacterium tumefaciens line that contained the chimeric gene. Three transgenic lines were determined by Southern and Northern analysis, and ELISA, to express the chimeric gene and produce coat protein. Trials to determine the resistance of these plant lines to BCTV are pending federal approval.

Resumo: A alface é a hortaliça folhosa de maior consumo no Brasil, no entanto a dificuldade em produzi-la vem aumentando, principalmente pela infestação das áreas de produção por Bremia lactucae, sendo necessário o monitoramento anual para utilização ou desenvolvimento de cultivares resistentes. Diante do exposto, os objetivos do presente trabalho foram: identificação de raças de B. lactucae no Estado de São Paulo, durante os anos de 2008, 2009 e 2010 e obtenção de linhagens de alface crespa resistentes às raças SPBl:01, SPBl:02, SPBl:03, SPBl:04, SPBl:05, SPBl:06 e SPBl:07. Na primeira fase do estudo foram coletadas nos meses de agosto, setembro e outubro de 2008; agosto e setembro de 2009; e junho e julho de 2010 folhas de alface com esporângios de B. lactucae nos principais municípios produtores de alface do Estado de São Paulo. Após a multiplicação dos esporângios na cultivar suscetível Solaris, com posterior inoculação nas diferenciadoras, realizaram-se as avaliações, no mesmo dia do aparecimento da primeira esporulação na cultivar suscetível Cobham Green (DM 0). Na segunda fase do estudo realizou-se o cruzamento entre a cultivar Argeles e a linhagem JAB 4-13-7, bem como autofecundações, até a obtenção da geração F3, quando realizou-se o teste de resistência. Os dados permitiram concluir que: três novos códigos "Sextet" para Bremia lactucae ocorreram no Estado de São Paulo: 63/63/33/00, 63/63/02/00 e 63/31/03/00, denominados raças SPBl:05, SPBl:06 e SPBl:07, respectivamente; recomenda-se a utilização dos genes R-17, R-18 e R-38 como fontes de resistência ao míldio para o desenvolvimento de cultivares no Estado de São Paulo; e 19 progênies resistentes à B. lactucae, raças SPBl:01, SPBl:02, SPBl:03, SPBl:04, SPBl:05, SPBl:06 e SPBl:07, apresentaramse homozigotas, e podem ser avançadas e avaliadas no processo de seleção
Abstract: Lettuce is the most consumed leafy vegetable in Brazil. Its production, nonetheless, is meeting with increasing difficulties mainly due to the infestation of producing areas with Bremia lactucae. Thus, the annual monitoring of B. lactucae races becomes of essential importance to use or development of resistant cultivars. The objectives of this work were identification of new races of B. lactucae occurring during the years of 2008, 2009 and 2010 in the state of São Paulo, Brazil, and development of resistant lettuce cultivars to races SPBl:01, SPBl:02, SPBl:03, SPBl:04, SPBl:05, SPBl:06 and SPBl:07. In the first phase of experiment in August, September, and October of 2008; August and September of 2009; and June and July of 2010 lettuce leaf samples containing B. lactucae sporangia were collected in some of the lettuce producing areas of the state of São Paulo. After a multiplication procedure in which the susceptible lettuce cultivar 'Solaris' was used, the B. lactucae sporangia were inoculated in differentiating lettuce cultivars. Evaluations of the plants response were performed on the same day when plants of the susceptible Cobham Green (DM 0) cultivar showed signs of the first sporulation. In the second phase of experiment made the crossing between Argeles cultivar and JAB 4-13-7 line, and self pollination, to obtain the F3 generation, when made the resistant test. The data have identified: three new codes "Sextet" in the state of São Paulo: 63/63/33/00, 63/63/02/00 e 63/31/03/00. SPBl:05, SPBl:06 and SPBl:07 were the denomination proposed to races, respectively. The genes R-17, R-18, and R-38 are suggested to be used as source of resistance in the lettuce cultivars to state of São Paulo; and 19 resistant progenies to B. lactuae, races SPBl:01, SPBl:02, SPBl:03, SPBl:04, SPBl:05, SPBl:06 and SPBl:07, shown homozygous, and they can be advanced and evaluated in the selection process
Orientadora: Leila Trevizan Braz
Coorientador: Tagli Dalpian
Coorientadora: Margarete Camargo
Banca: Pablo Forlan Vargas
Banca: Érika Auxiliadora Giacheto Scalopp
Banca: Rita de Cássia Panizzi
Banca: Joaquim Adelino de Azevedo Filho
Doutor

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES
Fusarium wilt, caused by Fusarium oxysporum f. sp. lactucae (FOLac), is an important disease of lettuce in the world. This disease was reported in Brazil, recently. From 2008 to 2011 the laboratories of Plant Pathology of Embrapa Hortaliças (Embrapa Vegetable Crops), Sakata Seeds Sudamerica and Instituto Capixaba de Pesquisa, Assistência Técnica e Extensão Rural (INCAPER) collected some FOLac isolates in states from the Southern and the Southeastern regions of Brazil. The isolates were identified as F. oxysporum by the morphological characteristics of conidia and conidiophores. Isolates were inoculated on lettuce plants, cultivars Elisa, Vera, and Red Salad Bowl, conditions in a greenhouse. Isolates were also inoculated on plants of others Asteraceae species (Cichorium endivia, Cichorium intybus, Sonchus oleraceus, Emilia sonchifolia, Bidens pilosa e Tagetes erecta) and others botanical families as (Solanum lycopersicum, Capsicum annuum, Nicotiana tabacum, Gossypium hirsutum, Phaseolus vulgaris e Ocimum basilicum). Lettuce cultivars Elisa and Vera were suscetible to all isolates while the cultivar Red Salad Bowl was resistant. All isolates were reisolated from diseased plants fulfilling the Koch‟s postulates. Others plant species were not susceptible to any isolate, proving that isolates belong to the species F. oxysporum f. sp. lactucae. The isolates were also inoculated in a differential set of cultivars comprising: „Patriot‟ (Susceptible to all races), „Costa Rica No. 4‟ (resistant to race 1) and „Banchu Red Fire‟ (resistant to race 2). Cultivars „Patriot‟ and „Banchu Red Fire‟ were susceptible while „Costa Rica No. 4‟ was resistant, confirming that all the isolates were race 1. Molecular analysis using a primer specific to race 1 isolates was performed using F. oxysporum f. sp. lycopersici (FOL) raça 3 and a non-pathogenic isolate as negative controls. DNAs of FOLac isolates were amplified by PCR and those of the negative controls were not, confirming the specificity of the primers and the presence of only the race 1 of FOLac in Brazil. In addition, it was used the Translation elongation factor 1-α region (tef-1α) for phylogenetic analysis between FOLac isolates races 1 and 2 and isolates of F. oxysporum. Comparison of the sequences obtained with the tef-1α confirmed the polyphyletic origin of the forma specialis lactucae and also showed a greater genetic variability among Brazilian isolates of FOLac race 1compared with isolates of the same race available in GenBank. After the isolates characterization, it was made a screening of 102 accessions for resistance to the isolate Fus-173 and it was selected 47 as highly resistant. After this, the selected genotypes were evaluated for the stability of resistance in three additional assays, using different FOLac race 1 isolates. In all three assays it was used a highly susceptible cultivar (Regina) as susceptible control. In the first assay, carried out in October 2011, it were used the isolates Fus-202 and Fus-205. In the second assay, carried out in November 2011, it were used the isolates Fus-219 and Fus-222. In the third assay, carried out in December 2011, it were used the isolates (Fus-207, Fus-209 e Fus-220). Inoculation was performed on 25 days old seedlings on greenhouse conditions. Seedlings were inoculated by cutting their roots and emerging them in spore suspension of pathogen. Evaluation was carries out 30 days after inoculation, using a grade scale varying from 0 (heath plants) to 4 (dead plants). Data were transformed in Disease Index (DI) submitted to a variance analysis and the media were compared by the Tukey‟s test (5%). Thirty two accessions were identified as having broad spectrum of resistance to different pathogen isolates in the four inoculation seasons.
A murcha de fusário, causada pelo fungo Fusarium oxysporum f. sp. lactucae (FOLac) é uma das doenças mais importantes da alface. Esta doença foi relatada recentemente no Brasil. Nos anos de 2008 a 2011 foram coletados isolados de FOLac nos Laboratórios de Fitopatologia da Embrapa Hortaliças, Sakata Seed Sudamerica Ltda e do Instituto Capixaba de Pesquisa, Assistência Técnica e Extensão Rural (Incaper). Estes eram provenientes de todos os estados das Regiões Sul e Sudeste do Brasil. A identificação foi feita observando-se as características morfológicas de conídios e conidióforos. Os isolados foram inoculados em plantas das cultivares Elisa, Vera e Red Salad Bowl, em condições de casa de vegetação. Os mesmos também foram inoculados em plantas de outras espécies de Asteraceae (Cichorium endivia, Cichorium intybus, Sonchus oleraceus, Emilia sonchifolia, Bidens pilosa e Tagetes erecta) e de outras famílias (Solanum lycopersicum, Capsicum annuum, Nicotiana tabacum, Gossypium hirsutum, Phaseolus vulgaris e Ocimum basilicum). Os isolados foram identificados como Fusarium oxysporum. As cultivares Elisa e Vera foram suscetíveis a todos os isolados e a Red Salad Bowl foi resistente. Efetuou-se o re-isolamento do patógeno, completando-se assim os Postulados de Koch. Nenhuma outra espécie de planta foi suscetível ao patógeno, confirmando a identificação como F. oxysporum f. sp. lactucae. Em seguida, os isolados foram avaliados quanto a sua virulência numa série de cultivares diferenciadoras de raças: Patriot (suscetível), Costa Rica No. 4 (resistente à raça 1) e Banchu Red Fire (resistente à raça 2). As cultivares Patriot e Banchu Red Fire comportaram-se como suscetíveis a todos os isolados, enquanto a cultivar Costa Rica No. 4 comportou-se como resistente. Concluiu-se que os isolados avaliados são da raça 1 de FOLac. Adicionalmente, foi feito um teste com primers específicos para a raça 1 de FOLac, utilizando como controles um isolado de F. oxysporum f. sp. lycopersici (FOL) raça 3 e um isolado não patogênico de F. oxysporum. O fragmento de DNA foi amplificado por PCR para os isolados de FOLac e não foi amplificado para o isolado de FOL e nem para o isolado não patogênico de F. oxysporum. Este resultado confirma a especificidade desse par de primers e a presença apenas da raça 1 de FOLac no Brasil. Além disso, foi utilizada a região do fator de elongação da tradução (tef-1α) para análise filogenética entre os isolados FOLac raças 1 e 2 e isolados de F. oxysporum. Comparação das seqüências obtidas com o tef-1α confirmou a origem polifilética da forma specialis lactucae e também observou-se uma maior variabilidade genética entre os isolados brasileiros de FOLac raça 1 comparados com isolados da mesma raça disponíveis no GenbanK. Posteriormente, 102 acessos (cultivares comerciais ou linhagens) foram avaliados, visando identificar fontes de resistência a FOLac e analisar a estabilidade da resistência de acessos promissores a diferentes isolados do patógeno. Inicialmente foi feita uma seleção preliminar, utilizando um isolado do patógeno (Fus-173). Em seguida, quarenta e sete acessos altamente resistentes mais uma testemunha suscetível (Regina), identificados na seleção preliminar, foram reavaliados para estabilidade da resistência ao FOLac raça 1, utilizando os isolados (Fus-202 e Fus-205) no mês de Outubro de 2011; isolados (Fus-219 e Fus-222) no mês de Novembro de 2011 e isolados (Fus-207, Fus-209 e Fus-220) no mês de Dezembro de 2011. A inoculação foi realizada em condições de casa de vegetação, pelo método de corte das raízes e imersão na suspensão de conídios do patógeno. A avaliação foi realizada após 30 dias, com auxílio de escala de notas variando de 0 (planta sadia) a 4 (planta morta). Os dados obtidos foram transformados em índice de doença (ID) e submetidos a uma análise de variância e comparação de médias pelo teste de Tukey (5%). Foram identificados trinta e dois acessos apresentando amplo espectro de resistência aos diferentes isolados do patógeno nas quatro épocas de inoculação.

Les oomycètes représentent un groupe d’organismes très diversifiés responsables de diverses maladies d’animaux et de plantes. En agriculture le mildiou de la pomme de terre, causé par un oomycète phytopathogène, occasionne à lui seul des pertes économiques mondiales annuelles de 3 à 5 milliards de dollars américains. La lutte contre le mildiou de la pomme de terre et de la laitue, causés respectivement par, Phytophthora infestans et Bremia lactucae, repose en grande partie sur l’utilisation de fongicides. Cependant, l’apparition de nouvelles souches résistantes au métalaxyl a réduit de manière substantielle l’efficacité des fongicides de synthèse. De plus, les fongicides doivent être appliqués au moment opportun pour être efficaces contre le mildiou. L’approche par modélisation constitue donc une option intéressante pour améliorer la lutte contre les mildious en offrant la possibilité de prédire les périodes à risque et d’améliorer la régie d’application des fongicides. Toutefois, l’évaluation de ces modèles a montré qu’ils manquent de précision et que leur efficacité varie d’une année à l’autre. Ce manque de précision peut s’expliquer en partie par l’absence de considération de la présence ou de l’abondance de l’inoculum aérien. P. infestans et B. lactucae produisent un inoculum adapté à la dispersion aérienne. Cependant, il existe un manque de connaissance notable dans la littérature sur l’aérobiologie de ces deux oomycètes phytopathogènes. Un modèle de simulation dynamique qui intègrerait l’information sur l’aérobiologie de l’agent pathogène permettrait de prédire plus efficacement les périodes de risque d’infection par le mildiou. Ainsi, l’objectif général de ce projet de doctorat était d’améliorer la connaissance sur l’aérobiologie de deux oomycètes (P. infestans et B. lactucae) dans l’optique de mieux comprendre et prédire les périodes d’infection du mildiou de la pomme de terre et de la laitue. Dans un premier temps, la distribution spatiotemporelle des spores à l’échelle d’une région de production de monoculture de pommes de terre a été déterminée. Les résultats ont montré que la distribution des spores est hétérogène et l’efficacité des mesures prophylactiques peut être évaluée à l’aide de l’aire sous la courbe de progression de l’inoculum aérien. De plus, l’information dérivée de l’implantation d’un réseau de capteur permet de pondérer le risque d’infection estimé par les modèles prévisionnels. Dans un deuxième temps, la relation entre la concentration aérienne de spores et l’intensité du mildiou a été établie. Cette relation a permis de définir des seuils d’intervention basés sur la concentration aérienne de spores. Dans le pathosystème de la pomme de terre, le seuil d’intervention varie en fonction des lignées clonales de P. infestans alors que dans celui de la laitue, ce seuil ne tient pas compte des lignées clonales de B. lactucae. Un outil moléculaire pour le comptage des spores a été développé pour pallier aux inconvénients liés à la quantification des spores au microscope. Les résultats de cette étude ont montré, entre autres, que sous conditions climatiques au Canada deux heures de mouillure sont suffisantes pour l’établissement d’une infection par B. lactucae, contrairement aux trois à quatre heures de mouillure proposées dans la littérature. Dans un troisième temps, un modèle de simulation de la concentration aérienne des spores a été développé. La relation entre le nombre de spores observées à l’heure et le nombre de spores prédit à l’heure est linéaire. Dans plus de 94% des cas, les coefficients de détermination de la régression entre le nombre de spores observées et celui prédit sont supérieurs à 0,7. Même si ce modèle est une première dans la littérature publiée et représente une avancée significative dans l’aérobiologie des oomycètes, il est à améliorer notamment dans l’optique de pouvoir développer un modèle générique comme outil de recherche pour l’étude de l’aérobiologie des oomycètes phytopathogènes. En effet, l’étape à venir sera d’incorporer ce modèle comme module dans un système de support décisionnel pour prédire efficacement l’apparition des symptômes de mildiou. Les auteurs espèrent que ce modèle constituera la base pour le développement d’un modèle générique pour tous les oomycètes phytopathogènes à dispersion aérienne.

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A murcha de fusário, causada pelo fungo de solo Fusarium oxysporum f.sp. lactucae é uma das doenças mais severas que ataca m a cultura da alface em todo o mundo. Em 2011, foram encontrados em uma plantação de alface n os município s de Capão Bonito e Campin as n o estado de São Paulo, sintomas característicos do ataque do patógeno. Plantas sintomáticas foram coletadas para identificação do patógeno. O s exame s em microscóp io óptico revel aram a presença de macroconídios falcados de coloração hialina, microconídi os hialinos unicelulares e testes de patogenicidade revelaram a presença de Fusarium oxysporum f.sp. lactucae . Efetuou - se o re - isolamento do patógeno, completa ndo - se assim o Postulado de Koch . No total, for am encontrados 4 isolados patogê nicos atacando pla ntas de alface. A partir do s isolados identificados , fo ram caracterizada s as variações patogênicas através de diferenciadoras de raça. Para esta finalidade foram utilizadas Patriot ( suscetível às raças 1, 2 e 3 ), Costa Rica ( resistente à raça 1 ; suscetível às raças 2 e 3 ), Summer Green ( suscetível às raças 1 e 3 ; resistente à raça 2 ) e Banchu Red Fire ( suscetível às raças 1 e 3, resistente à raça 2 ) . Os resultados obtidos identificaram a ocorrência da raça 3 de Folac. Este é o primeiro relato de ocorrência desta raça no Brasil . A partir do isolado selecionado como sendo o mais agressivo , e caracterizado como pertencente a raça 3 , r ealizou - se a triagem de genótipos de alface, afim de identificar fontes de resistência para uso em programas de melhoramento . No total de 63 genótipos testados apenas Onondaga, Itha ca, Sudenvil e JP - 11 apresentara m - se como resistente a raça 3 de Fusarium oxysporum f.sp. lactucae
The fusarium wilt, caused b y the soil fungus Fusarium oxysporum f.sp. soil lactucae is one of the most severe diseases that attack lettuce crop worldwide. In 2011, were found in a pla nting lettuce in Capão Bonito and Campinas in São Paulo, symptoms characteristic of this pathogen. S ymptomatic plants were collected for pathogen identification. The optical microscope examinations revealed the presence of hyaline staining falcate macroconidia, microconidia unicellular hyaline and pathogenicity tests revealed the presence of Fusarium oxy sporum f.sp. lactucae. We performed the re - isolation of the pathogen, there by supplementing the Koch's postulate. For this purpose were used: Patriot(susceptible to races 1, 2 and 3), Costa Rica (susceptible to races 2 and 3, resistant to race 1); Summer Green (susceptible to races 1 and 3; resistant to race 2) and Banchu Red Fire (susceptible to races 1 and 3, resistant to race 2). The results show the occurrence of race 3 of Folac. This is the first report of the case in Brazil. From the isolated select ed as being the 3 most aggressive, and characterized as belonging to race 3 was held screening of lettuce genotypes in order to identify sources of resistance for use in breeding programs. From of 63 genotypes tested only Onondaga, Ithaca, Sudenvil and JP - 11 were identified as resistant to race 3 of Fusarium oxysporum f.sp. lactucae

Orientador: Marcelo Agenor Pavan
Coorientador: Denise Nozaki
Banca: Rumy Goto
Banca: Romulo Fujito Kobori
Resumo: A murcha de fusário, causada pelo fungo de solo Fusarium oxysporum f.sp. lactucae é uma das doenças mais severas que ataca m a cultura da alface em todo o mundo. Em 2011, foram encontrados em uma plantação de alface n os município s de Capão Bonito e Campin as n o estado de São Paulo, sintomas característicos do ataque do patógeno. Plantas sintomáticas foram coletadas para identificação do patógeno. O s exame s em microscóp io óptico revel aram a presença de macroconídios falcados de coloração hialina, microconídi os hialinos unicelulares e testes de patogenicidade revelaram a presença de Fusarium oxysporum f.sp. lactucae . Efetuou - se o re - isolamento do patógeno, completa ndo - se assim o Postulado de Koch . No total, for am encontrados 4 isolados patogê nicos atacando pla ntas de alface. A partir do s isolados identificados , fo ram caracterizada s as variações patogênicas através de diferenciadoras de raça. Para esta finalidade foram utilizadas Patriot ( suscetível às raças 1, 2 e 3 ), Costa Rica ( resistente à raça 1 ; suscetível às raças 2 e 3 ), Summer Green ( suscetível às raças 1 e 3 ; resistente à raça 2 ) e Banchu Red Fire ( suscetível às raças 1 e 3, resistente à raça 2 ) . Os resultados obtidos identificaram a ocorrência da raça 3 de Folac. Este é o primeiro relato de ocorrência desta raça no Brasil . A partir do isolado selecionado como sendo o mais agressivo , e caracterizado como pertencente a raça 3 , r ealizou - se a triagem de genótipos de alface, afim de identificar fontes de resistência para uso em programas de melhoramento . No total de 63 genótipos testados apenas Onondaga, Itha ca, Sudenvil e JP - 11 apresentara m - se como resistente a raça 3 de Fusarium oxysporum f.sp. lactucae
Abstract: The fusarium wilt, caused b y the soil fungus Fusarium oxysporum f.sp. soil lactucae is one of the most severe diseases that attack lettuce crop worldwide. In 2011, were found in a pla nting lettuce in Capão Bonito and Campinas in São Paulo, symptoms characteristic of this pathogen. S ymptomatic plants were collected for pathogen identification. The optical microscope examinations revealed the presence of hyaline staining falcate macroconidia, microconidia unicellular hyaline and pathogenicity tests revealed the presence of Fusarium oxy sporum f.sp. lactucae. We performed the re - isolation of the pathogen, there by supplementing the Koch's postulate. For this purpose were used: Patriot(susceptible to races 1, 2 and 3), Costa Rica (susceptible to races 2 and 3, resistant to race 1); Summer Green (susceptible to races 1 and 3; resistant to race 2) and Banchu Red Fire (susceptible to races 1 and 3, resistant to race 2). The results show the occurrence of race 3 of Folac. This is the first report of the case in Brazil. From the isolated select ed as being the 3 most aggressive, and characterized as belonging to race 3 was held screening of lettuce genotypes in order to identify sources of resistance for use in breeding programs. From of 63 genotypes tested only Onondaga, Ithaca, Sudenvil and JP - 11 were identified as resistant to race 3 of Fusarium oxysporum f.sp. lactucae
Mestre

Tese (doutorado)—Universidade de Brasília, Instituto de Ciências Biológicas, Departamento de Fitopatologia, Programa de Pós-Graduação em Fitopatologia, 2016.
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A alface (Lactuca sativa L.) é uma das hortaliças mais importantes no Brasil e no mundo. A murcha de fusário (causada por distintas raças do fungo Fusarium oxysporum f. sp. lactucae – FOLAC) é uma das principais doenças de da alface em regiões tropicais e subtropicais. Devido à dificuldade de implementação de estratégias eficazes de controle químico e cultural, o uso de cultivares com resistência genética é o método mais prático de manejo da doença. Fontes estáveis de resistência à raça FOLAC 1 foram encontradas, mas a base genética dessa característica ainda se encontra mal caracterizada. A integração da seleção assistida por marcadores (SAM) em programas de melhoramento convencional irá acelerar o desenvolvimento e lançamento de cultivares de alface com resistência genética a esse patógeno. O presente estudo teve como objetivos: elucidar os fatores genéticos associados à resistência a murcha de fusário (Capítulo 2) e idenficar marcadores moleculares potencialmente ligados aos fatores de resistência a isolados de FOLAC raça 1 identificados na cultivar ‘Vanda’ (Capítulo 3). O estudo da herança genética da resistência a FOLAC raça 1 foi conduzido com populações segregantes (F2 e F3) obtidas do cruzamento entre um parental suscetível ‘Gizele’ e ‘Vanda’. Populações foram inoculadas com uma suspensão de esporos do patógeno (3 x 106 microconídios/mL) por meio de corte e imersão de raízes e avaliadas quanto à resistência por meio de escala de notas. O DNA genômico das plantas resistentes e suscetíveis foi extraído e usado como molde em diferentes sistemas de marcadores (RAPD, SCAR, DR analogs, SRR e CAPS), visando identificar polimorfismos ligados a essa característica. Com relação à resistência a FOLAC raça 1, os estudos indicaram um controle genético relativamente simples na cultivar ‘Vanda’, com os resultados de segregação indicando um locus monogênico com uma provável combinação de efeitos de dosagem e penetrância incompleta. No entanto, os diferentes sistemas de marcadores moleculares utilizados nessa primeira etapa do trabalho não permitiram encontrar polimorfismos com estreita ligação com o(s) fator(es) de resistência. Desta forma, uma nova etapa do trabalho foi conduzida visando identificar por meio do método genotyping-by-sequencing (GBS) marcadores moleculares ligados a resistência (Capítulo 4). O GBS foi explorado na identificação de SNPs (single nucleotide polymorphisms) empregando DNA extraído dos dois parentais e de um conjunto de 82 indivíduos da população F2 derivados do cruzamento entre ‘Vanda’ x ‘Gizele’. Cada indivíduo foi genotipado utilizando o Illumina Hiseq 3000, que produziu 4,5 milhões de reads por amostra. Um total de 10.017 SNPs foi identificado entre os parentais. Estes SNPs foram avaliados para ligação com o fenótipo de resistência nos 82 indivíduos F2. Os dados genotípicos foram condensados em 1.484 scaffolds ou supercontigs. Um subconjunto de 417 scaffolds contendo polimorfismos foi selecionado para a construção de um mapa de ligação após filtragem baseada em missing data (< 20%), teste de qui-quadrado (3:1 p>0,05) e número de SNPs por scaffold. O mapa foi composto por 17 grupos de ligação, com um comprimento total de 1.132,984 cM. A análise de QTL (quantitative trait loci) para resistência a FOLAC raça 1 foi realizada no mapa de ligação com os conjuntos de dados de severidade da doença obtido nas populações F2 e F3. Dois QTLs de efeito maior foram identificados no cromossomo 9, explicando 30 a 40% da variação fenotípica observada na população F3 e F2, respectivamente. Um QTL de efeito menor foi detectado no cromossomo 4, explicando 0,06% da variação fenotípica na população F3. Desta forma, o presente estudo estabelece a porção mediana do cromossomo 9 como sendo a localização física do principal locus de resistência para FOLAC raça 1 presente na cultivar ‘Vanda’. Portanto, os marcadores moleculares localizados na proximidade desta região genômica são candidatos para o desenvolvimento de ferramentas de SAM em programas de melhoramento genético visando incorporar essa característica em linhagens elite de alface.
Lettuce (Lactuca sativa L.) is one of the most important leafy vegetable crops in Brazil and worldwide. Fusarium wilt (caused by distinct races of the fungus Fusarium oxysporum f. sp. lactucae – FOLAC) is one of the main soil-borne diseases of lettuce in tropical and subtropical regions. Due to the complexity of implementing effective cultural and chemical control, the most practical disease management strategy has been the use of cultivars with genetic resistance. Stable sources of resistance to FOLAC race 1 have been found, but the genetic basis of this trait is yet poorly characterized. The integration of marker assisted selection (MAS) in conventional breeding programs would be an important contribution to accelerate the development and release of lettuce cultivars with genetic resistance to this pathogen. In this context, the present study aimed to elucidate the inheritance of resistance to FOLAC race 1 detected in the cultivar Vanda (Chapter II) and to search for molecular markers linked to FOLAC race 1 resistance factor(s) (Chapter III). Inheritance studies were carried out using segregating populations (F2 and F3 families) derived from the cross between a susceptible cultivar (Gizele) and a FOLAC race 1 resistant cultivar (Vanda). The parental lines and the segregating populations were inoculated via root dipping technique with a conidial suspension adjusted to 3 x 106 microconidia/mL. Reaction to one FOLAC race 1 isolate was evaluated using a disease rating scale ranging from 1 (= no symptoms) to 5 (= dead plant). The studies indicated a simple genetic control of FOLAC race resistance in cultivar Vanda, with segregation results indicating a single gene locus with a likely combination of dosage effects and incomplete penetrance. Genomic DNA of resistant and susceptible plants was extracted and used as a template in PCR assays with different marker systems (RAPD, SCAR, DR Analogs, SSRs and CAPS) aiming to identify polymorphisms linked to the resistant reaction. However, none of the evaluated molecular techniques were able to identify markers in close linkage with the resistance factor(s). Therefore, a new phase of the present study was conducted with the objective of searching for markers linked to resistance through the employment of the genotyping-by-sequencing (GBS) strategy (Chapter IV). The GBS strategy was employed using DNA extracted from the parental lines and 82 phenotyped F2 individuals derived from the same cross between Gizele x Vanda. Each individual was genotyped using the Illumina Hiseq 3000. A total of 4.5 million reads was obtained per sample with 10,017 SNPs being identified among the parental lines as well as among the 82 F2 individuals. Genotypic data were condensed into 1484 scaffolds. Four hundred seventeen (417) scaffolds containing polymorphisms were selected for the construction of the linkage map after filtering based on missing data (< 20%), chi-square test (3: 1 p> 0.05) and number of SNPs per scaffold. The final map consisted of 17 linking groups with a total length of 1,132.984 cM. The QTL analysis for resistance to FOLAC race 1 was performed on the linkage map based upon the disease severity datasets of F2 population and F3 families. Two major effect QTLs were identified on chromosome 9, explaining 30 to 40% of the phenotypic variation observed in the population F3 and F2, respectively. One QTL of minor effect was also identified on chromosome 4, explaining 0.06% of the phenotypic variation in the F3 population. This study establishes the central region of chromosome 9 as the physical location of a major resistance locus to FOLAC race 1 found in Vanda. Molecular markers in close proximity to this genomic region are candidates for the development of MAS tools for breeding programs aiming to incorporate this trait into elite lettuce lines.

The hawkweed subgenus Hieracium s. str. is notoriously known for its extreme morphological variability and variation in ploidy levels that is associated with differences in modes of reproduction. Extensive past hybridization is supposed for the subgenus, but recent hybridization was evidenced only in few cases. The subgenus attracts the attention of botanists already for more than a century. Therefore the species diversity is largely examined and the taxonomy of the subgenus is well elaborated, although several contradictory taxonomic concepts exist. However the relationships among the species are unknown and haven't been studied yet. The investigation of these relationships from a phylogenetic perspective using molecular approaches was the main aim of the presented thesis. Basic species (both diploid and polyploid), representing morphologically unique taxa, that are supposed to be the basic evolutionary units of the subgenus were studied. The sequences of two intergenic spacers of the cpDNA (trnT-trnL and trnV-ndhC) and the external transcribed spacer of the nuclear ribosomal DNA (nrDNA ETS) were analyzed. Moreover, three new low-copy nuclear markers with higher variability than nrDNA and cpDNA markers were developed and their suitability for phylogenetic studies in Hieracium s. str. was...

碩士
國立中興大學
植物病理學系
93
In Taiwan, lettuce Fusarium wilt caused by Fusarium oxysporum f. sp. lactucae is a serious soil-borne disease in monoculture fields during summer season. The disease plants of lettuce appeared yellowing, vascular discoloration of taproots and stems, stunning and wilt symptoms. The disease is one of the limiting factors of yield in commercial lettuce cultivation. Three lettuce cultivars, Costa Rica No.4, Benchu Red Fire and Patriot were used as differential cultivars for identification of the physiological races of F. oxysporum f. sp. lactucae. The results indicated that 12 isolates of the pathogen obtained from Taiwan were identified as race1, howeven the pathogenical characteristics of three isolates LFO 32-14, LFO 106-1 and LFO 106-3 were different from ones of three races of Japanese and USA isolates. The results suggest that a new race of F. oxysporum f. sp. lactucae may has occurred in Taiwan. Susceptibility of thirty lettuce cultivars to isolates LFO 11-13 and LFO 32-14 was apparently varied degree. Among the cultivars tested, asparagus lettuce and crisp head lettuce were the most resistant to Fusarium wilt. However round-leaf lettuce, pointed leaf lettuce and indian lettuce were very susceptible. In this study, amendment of soil with crab and shrimp shell powder, soybean meal and CH100-S was able to enhance lettuce growth and reduce disease severity of lettuce Fusarium wilt. CH100-S, a new formulation of Plant health guard CH100 paste, was obtained from precipitate substance of a mixture solution for producing CH100 that includes major ingredients of S-H mixture, extracts of tobacco and cabbage, and is recommended for control of a wide range of plant pests. Application of CH100-S at 1000-fold dilution was significantly effective in reducing disease severity of lettuce Fusarium wilt in greenhouse test. Lettuce seeds were sown in infested soil which had been treated with CH100-S for six days. The disease severity of lettuce Fusarium wilt was markedly reduced 27 – 50 %. CH100-S did also promote growth of lettuce plants and stimulate proliferation of beneficial microorganisms including fungi, bacteria and actinomycetes. Beneficial microorganisms were isolated from the CH100-S-treated rhizosphere soils, and showed either antagonistic ability to the pathogen or promoting plant growth. These bacterial isolates including B122, CHB139, CHB143, CHB180, CHB184, CHB186 and CHB190 were significantly effective in reducing disease severity and increasing root weight of lettuce. Especially, amendment of LFO 11-13-infested soil with CH100-S mixed with CHB184 was more effective than CH100-S alone in reducing disease severity.

碩士
國立中興大學
植物病理學系所
98
Fusarium wilt of lettuce (Lactuca sativa L.), caused by the vascular wilt pathogen Fusarium oxysporum (Schlechtend.: Fr.) f. sp. lactucae (Fola) was found in Yunlin lettuce field in Taiwan since 1996, become one of the major factors restricting the stable production of lettuce on summer in Taiwan. Identification of pathogenic races of Fola is fundamentally necessary for lettuce growers to choose appropriate cultivars for production and for breeders to develop Fola-resistant cultivars. In our study, Fola isolates in Taiwan and reference isolates from Japan were subjected to inoculation test for race identification. The results showed that isolates SB1-1 (race 1 reference isolate), Fo-2, Fo-18, LFO11-13, LFO32-14, LFO106-1, and LFO106-3 were identified as race 1; 744085 (race 3 reference isolate), Fo-10, and Fo-40 isolates, with their races unknown, were identified as race 3 in this study. Furthermore, primer sets of FLA0101 F/R and Hani3’/Hanilatt3rev, and FLA0201 F/R were unable to amplified the DNA bands by polymerase chain reaction (PCR) specific to race 1 and race 2, respectively, in the DNA samples of Fo-10 and Fo-40. The random amplified polymorphic DNA (RAPD) patterns also showed differences between Fo-10 and Fo-40, and the other Fola isolates tested. Combining the results of inoculation tests and PCR assay, our results suggested that a new race of Fola may have occurred in Taiwan. In addition, we use the published primer sets (FLA0001 F/R, FLA0101 F/R, and Hani3’/Hanilatt3rev) to test the specificity and sensitivity for Fola isolates. The sensitivity can be increased by 10 to 100 folds by modified PCR conditions. Besides, RAPD analysis of the representative isolates of each race could be a useful mean for rapid and unambiguous race identification of Fola. The RAPD patterns shown in this study could differentiate race 1, race 2, and race 3 of Fola isolates using different random primers (OP-2, OP-3, OP-5, OP-6, and OP-7) that amplified DNA bands specific to race 1, race 2, and race 3, individually. These bands could be organized as a schematism to differentiate races of Fola isolates in Taiwan. In this study, we can differentiate different races of Fola by using RAPD and PCR technique with the combinations of different random primers and primer sets, to reduce the time and efforts for Fola identification and detection.