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Quantitative trait loci (QTL) analysis for Sclerotinia stem rot resistance was carried out in five doubled haploid (DH) populations of Brassica napus. Sclerotinia stem rot is caused by the necrotrophic fungus Sclerotinia sclerotiorum (Lib.) de Bary. Sclerotinia stem rot has worldwide occurrence and causes significant yield losses in many crop species. Several screening methods have been recommended in the literature to evaluate plant resistance to Sclerotinia stem rot. Four controlled environment based screening methods: 1) excised leaf assay, 2) cotyledon assay, 3) mycelial stem inoculation technique and 4) petiole inoculation technique compared for their ability to differentiate between plant susceptibility/resistance, their reliability and suitability for large scale screening using eight B. napus cultivars/lines of varying reaction to S. sclerotiorum. The petiole inoculation technique and the mycelium stem inoculation technique were identified as reliable methods in this study. Previously developed, five B. napus DH populations (H1, H2, H3, DH179 and DH180) segregating for resistance to Sclerotinia stem rot were used in this study. The petiole inoculation technique was used to evaluate resistance to Sclerotinia stem rot. Data on days to wilting was recorded for a two week period. Twelve plants per line were screened in each evaluation and each population was evaluated three times. Two to three day-old mycelial cultures of S. sclerotiorum isolate Canada 77 was used. QTL analyses were carried out using a LOD threshold value of 2.5 on each individual replicate and on the average of all the replicates. In the H1 population, the number of QTL detected ranged from four to six in each analysis. In the H2 population, there were three to six QTL in each analysis. There were two to six QTL in each analysis of the H3 population. In the DH179 population, the number of QTL detected ranged from three to five in each analysis. In DH180 population, the number of QTL identified varied from three to six in each analysis. A number of common QTL were found between the replicates of each population. Five common QTL were identified between these populations. The markers linked to these QTL are now available for marker assisted selection.

Quantitative trait loci (QTL) analysis for Sclerotinia stem rot resistance was carried out in five doubled haploid (DH) populations of Brassica napus. Sclerotinia stem rot is caused by the necrotrophic fungus Sclerotinia sclerotiorum (Lib.) de Bary. Sclerotinia stem rot has worldwide occurrence and causes significant yield losses in many crop species. Several screening methods have been recommended in the literature to evaluate plant resistance to Sclerotinia stem rot. Four controlled environment based screening methods: 1) excised leaf assay, 2) cotyledon assay, 3) mycelial stem inoculation technique and 4) petiole inoculation technique compared for their ability to differentiate between plant susceptibility/resistance, their reliability and suitability for large scale screening using eight B. napus cultivars/lines of varying reaction to S. sclerotiorum. The petiole inoculation technique and the mycelium stem inoculation technique were identified as reliable methods in this study. Previously developed, five B. napus DH populations (H1, H2, H3, DH179 and DH180) segregating for resistance to Sclerotinia stem rot were used in this study. The petiole inoculation technique was used to evaluate resistance to Sclerotinia stem rot. Data on days to wilting was recorded for a two week period. Twelve plants per line were screened in each evaluation and each population was evaluated three times. Two to three day-old mycelial cultures of S. sclerotiorum isolate Canada 77 was used. QTL analyses were carried out using a LOD threshold value of 2.5 on each individual replicate and on the average of all the replicates. In the H1 population, the number of QTL detected ranged from four to six in each analysis. In the H2 population, there were three to six QTL in each analysis. There were two to six QTL in each analysis of the H3 population. In the DH179 population, the number of QTL detected ranged from three to five in each analysis. In DH180 population, the number of QTL identified varied from three to six in each analysis. A number of common QTL were found between the replicates of each population. Five common QTL were identified between these populations. The markers linked to these QTL are now available for marker assisted selection.

The fungal pathogen Sclerotinia sclerotiorum (Lib.) de Bary is capable of infecting over 400 plant species including canola (Brassica napus L.). The fungus secretes oxalic acid (OA), which plays an important role in infection and disease progression. An analysis of proteome-level changes associated with infection of susceptible canola leaves by S. sclerotiorum revealed significant changes in the abundance of 32 proteins, including proteins involved in photosynthesis and metabolism, hormone signaling, and antioxidant defense. A similar subset of 37 proteins was affected when leaves were treated with OA alone; this compound also caused a reduction in the activities of a number of antioxidant enzymes, suggesting an OA-mediated suppression of the oxidative burst. To further understand the mechanisms of pathogenesis, the role of Sssp, a predicted secreted protein from S. sclerotiorum, was targeted for analysis. Mutant strains of S. sclerotiorum were generated by disruption of the Sssp gene and characterized for virulence on canola. Based on the extent of symptom development, the virulence of the Sssp-disrupted mutants was significantly reduced relative to the wild-type, indicating that Sssp may play a role in the infection process. Finally, the development of sclerotia, long-term survival structures that serve as a primary source of inoculum for the fungus, was examined. A total of 88 proteins were found to exhibit temporal changes in abundance during sclerotium formation and maturation, including proteins involved in the regulation of melanogenesis. A total of 56 proteins were also identified in the sclerotial exudates, providing a basis for future studies. Collectively, the studies described in this dissertation represent the most comprehensive proteome-level analysis of the canola/S. sclerotiorum interaction and sclerotial development, and could contribute to the development of novel strategies for the management of S. sclerotiorum. / Plant Science

Lettuce (Lactuca sativa L.) seedlings diseases caused by soilborne pathogens are characterised by root rot, stem rot and damping-off of the seedlings that can occur at any time during growth. Fusarium solani, Pythium ultimum and Rhizoctonia solani are known to be the important destructive pathogens of lettuce, causing severe yield losses in South Africa. The aim of this research was to evaluate the effects of three selected fungicides to control these pathogens on lettuce seedlings. In this study the fungicides metalaxyl (Apron®), fludioxonil (Celest®) and mefenoxam (Subdue®) were applied at two concentrations as single and double doses on lettuce seedlings to determine their efficacy to control the pathogens Fusarium solani, Pythium ultimum and Rhizoctonia solani after significant reduction of mycelia growth was observed in vitro. Cultures of P. ultimum (UPGH024), R. solani (UPGH122) and F. solani (UPGH122) were obtained from the culture collection of the Department of Microbiology and Plant Pathology, University of Pretoria and cultivated on PDA for 2 days at 25ºC. Pasteurised soil was artificially inoculated with these pathogens. For the first experiment lettuce seeds were planted in polystyrene seedling trays at a depth of 1.0 cm. There were four replications of 50 seeds per treatment. In Experiment 2 pots (12 cm x 7 cm) were filled with pasteurised growing medium and 3-week old seedlings were transplanted. There were three replications of six pots containing three plants each. Seedling trays and pots were drenched with fungicides and placed in a randomised block design in a controlled environment room at 20- 26°C with a 12h-light/dark regime. The seedling trays and pots were rotated daily in the room. Seedling trays and pots were watered daily to maintain field capacity. The seedlings were able to grow larger in the pots than in seedling trays. It was confirmed that the treatment with fludioxonil (Celest®) at double and single dose inhibited the growth of the three fungi F. solani, P. ultimum and R. solani on lettuce seedlings without causing phytotoxicity. All three fungicides significantly reduced the diseases caused by the three pathogens. These findings are consistent with previous reports that fludioxonil, metalaxyl and mefenoxam can control oomycete fungi. There are few registered fungicides for the control of Fusarium solani, Pythium ultimum and Rhizoctonia solani on lettuce, therefore further work will aim to confirm these results in the field. / Dissertation (MInstAgrar)--University of Pretoria, 2008. / Microbiology and Plant Pathology / unrestricted

Cette thèse constitue une nouvelle approche pour la prédiction des maladies des plantes dans une plantation par combinaison de fusion de données et modélisation spatio-temporelle. La maladie des plantes est un problème majeur dans le monde de l'agriculture. Par exemple en Malaisie, la maladie de la pourriture de basal de la tige (BSR) causée par le champignon Ganoderma Boninense est la maladie la plus grave pour les plantations de palmiers à huile. Le champignon infecte les palmiers à huile,causant des pertes de rendement et détruisant au final les arbres. Divers facteurs ont été précédemment signalés, qui influencent l'incidence de la BSR, tels que les cultures précédentes, les techniques de replantation, les types de sols et l'âge des arbres. Une gestion efficace et durable des stratégies pour contrôler le BSR se heurte principalement à un manque de compréhension des mécanismes d'établissement de la maladie, de son développement et de sa propagation. La présente recherche est une tentative d'appliquer la technique de fusion de données et la modélisation temporelle en système d'Information géographique (SIG) pour étudier le comportement des maladies des plantes dans un domaine particulier (zone artisanale). Cette recherche portera sur comment les SIG peuvent aider à évaluer la distribution des maladies des plantes dans une plantation de petite échelle. Avec les progrès simultanés dans les systèmes de positionnement global (GPS) et l'utilisation des systèmes d'Information géographique, ces techniques ont fourni de puissants outils d'analyse pour l'agriculture de précision. Les données pour l'analyse proviennent de palmiers à huile des expériences de densité de plantation aux stations de recherche MPOB à Teluk Intan, Perak, Malaisie.Dans le cas de la maladie de la BSR, les résultats de l'émission de modélisation prédictive ont observé une corrélation entre les maladies BSR prédites avec celles visuellement données par le BSR. Il a été constaté que la modélisation prédictive proposée a bien prédit la présence de la maladie de la BSR. Même si au début d'infection des maladies BSR, le modèle n'a pas fixé exactement la distribution de la maladie, la performance du modèle sera améliorée avec la sélection de la source de données. Dans l'ensemble, le modèle a bien prédit la présence de maladies avec une précision allant jusqu'à 98,9 %. / This thesis represents a new approach for predicting plant disease in a plantation through combination of data fusion and spatio-temporal modelling. Plant disease is a major problem in the world of agriculture. Example in Malaysia, basalstem rot disease (BSR) caused by Ganoderma Boinense is the most serious disease for oil palm plantation in Malaysia. The fungus infects oil palm trees, initially causing yield loss and finally killing the trees. Various factors were previously reported to influence incidence of BSR, such as previous crops, techniques for replanting, types of soils and the age of trees. At present effective and sustainable management strategies to control BSR are hampered mainly by a lack of understanding of mechanisms of disease establishment, development and spread. The present research is an attempt to apply data fusion technique and temporal modelling in Geographical Information System (GIS) to investigate the behaviour of plant disease in a specific area (small skill area). This research will focus on how GIS can help to assess the distribution plant disease in a small scale plantation. With concurrent advances in global positioning systems (GPS) and the use of geographical Information Systems(GIS) techniques have provided powerful analysis tools for precision agriculture. Data for analysis were obtained from oil palm planting density experiments at MPOB research stations at Teluk Intan, Perak, Malaysia. In the case of BSR disease, the results of the predictive modelling show a significance correlation between predicted BSR diseases with visually observed BSR data. It found that the proposed predictive modelling has well predicted the presence of BSR disease. Although at the beginning stage of BSR diseases infection, the model has not fitted exactly the distribution of the disease, we believe that with the proper selection of the source of data, the performance of the model will be improved.Overall, the model has well predicted the presence of diseases with accuracy up to 98.9%.

Canola/rapeseed (Brassica napus L.) is a major oilseed crop worldwide. However, its production is largely affected by the fungal disease Sclerotinia stem rot as well as seed glucosinolates. So far the genetic mechanisms controlling these two traits have been poorly understood. In the present study, three bi-parental doubled haploid B. napus populations M730, M692 and ZT were grown in either natural or artificial environments and genotyped using the Brassica 60K Infinium® SNPs and/or sequence related amplified polymorphisms. Three genetic linkage maps covered 2,597.7 cM, 2,474.1 cM and 1,731.6 cM in 19 chromosomes for M730, M692 and ZT, respectively. Plants were inoculated with Sclerotinia sclerotiorum mycelia on stems at the reproductive stage to evaluate their resistivity. Four aliphatic glucosinolates and one indolic glucosinolate were detected in the seeds using high-performance liquid chromatography. 4-hydroxy-3-indolylmethyl predominated over aliphatic glucosinolates in canola, but inversely constituted a small portion of total glucosinolate content in semi-winter rapeseed. In rapeseed, 2-hydroxy-3-butenyl predominated in 4C aliphatic glucosinolates, which in turn predominated in total aliphatic glucosinolates, which likewise predominated in total glucosinolate content. QTLs regulating major glucosinolates were located on chromosome A9 for high glucosinolate content populations M730 and ZT, and on chromosome C7 for low glucosinolate content population M692. Major QTLs for Sclerotinia stem rot resistance were located on chromosomes A7 and C6 in M730, on chromosomes A3 and A7 in ZT, while no major QTLs were found in M692. Additive genetic effect was the major factor explaining phenotypic variations of the two traits. No direct genetic relationship was observed between Sclerotinia stem rot resistance in adult plants and seed glucosinolates in B. napus. The findings in the studies could be used to formulate breeding and research strategies in B. napus and the major QTLs controlling the two traits and their closely linked SNP markers could be validated over wide germplasm and used in marker assisted selection. / October 2016

Submitted by (lucia.rodrigues@ufrpe.br) on 2017-03-23T14:09:27Z No. of bitstreams: 1 Romulo Diniz Cavalcante.pdf: 683087 bytes, checksum: 8124aad4adf327b8c664300dfebcb472 (MD5) / Made available in DSpace on 2017-03-23T14:09:27Z (GMT). No. of bitstreams: 1 Romulo Diniz Cavalcante.pdf: 683087 bytes, checksum: 8124aad4adf327b8c664300dfebcb472 (MD5) Previous issue date: 2013-02-25 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / Stem rot caused by Lasiodiplodia theobromae is an important postharvest disease of papaya in Brazil. The disease symptoms start on the stem region and then progressing around the fruit. The tissues remain with soaked aspect and with progression of symptoms, lesions become dark brown and depressed. The infected area becomes gradually devoid of parenchymal tissue and fruits lose their consistency and strength. It is recommended that the disease control starts in the field and continue until the commercialization phase since the symptoms are difficult to be perceived until an advanced stage of maturation. The use of Fungicides is one of the main disease management measures. However, there are no data available on the sensitivity of L. theobromae to thiophanate methyl, the most common fungicide used in papaya orchards in Northeastern Brazil. Thus, the EC50 of 109 isolates of the fungus, representing five populations of the pathogen was estimated in vitro for the fungicide. Of the 109 isolates, 20,2% were non-sensitive (NS) to the fungicide with EC50 values greater than 300 mg ml-1, for the remaining 79.8% sensitive (S) isolates the average EC50 was 1.87 μg ml−1. The EC50 values for the NS isolates were significantly (P≤0.05) higher than those for the sensitive isolates. Seven components of fitness were measured for the 10 isolates with lower and high values of EC50. When the fitness components were evaluated, the non-sensitive (NS) isolates showed sporulation capacity significantly lower than the sensitive isolates, indicating a fitness costs. / A podridão peduncular causada por Lasiodiplodia theobromae é uma doença em pós-colheita importante na cultura do mamoeiro no Brasil. Os sintomas da doença iniciam na região do pedúnculo e então progride para todo o fruto. Os tecidos permanecem com aspecto encharcado e com a progressão dos sintomas, as lesões se tornam marrom escura e deprimida. A área infectada torna-se gradualmente desprovida de tecido parenquimatoso e os frutos perdem sua consistência e vigor. É recomendado que o controle da doença seja iniciado no campo e continue até a fase de comercialização, uma vez que os sintomas são difíceis de serem percebidos até um estádio avançado de maturação. O uso de fungicidas é uma das principais medidas de manejo da doença. Todavia, dados sobre a sensibilidade de L. theobromae a tiofanato metílico, fungicida comumente empregado em pomares de mamoeiros no Nordeste brasileiro são inexistentes. Sendo assim, a CE50 de 109 isolados do fungo, representando cinco populações do patógeno foi estimada in vitro para o fungicida. Dos 109 isolados, 20,2% foram não sensíveis (NS) ao fungicida com valores de CE50 superiores a 300 μg ml-1, para 79,8% dos isolados remanescente denominados de sensíveis (S), a média de CE50 foi 1,87 μg ml-1. Os valores da CE50 dos isolados NS foram significativamente (P≤0.05) superiores quando comparados com os isolados S. Sete componentes de adaptabilidade foram mensurados para os 10 isolados com menores e maiores valores de CE50. Quando foram avaliados os componentes de adaptabilidade, os isolados não sensíveis apresentaram capacidade de esporulação significativamente inferior aos isolados sensíveis, indicando um custo de adaptabilidade.

The cowpea, Vigna unguiculata (L.) Walp., is known as feijão-de-corda and feijão-verde , among others, is one of the main cultures exploited by small producers of the Northeast region of Brazil. Among the phytopathogens that affects its productivity, Sclerotium rolfsii Sacc. is noteworthy, causing the stem rot in several cultures around the world. The objective of this work was to evaluate the alternative control of S. rolfsii Sacc. in saplings of V. unguiculata (L.) Walp. trough the biocontrol of antagonists, organic residues incorporation to the soil, utilization of essential oils, plant extracts and mineral nutrition. The work was developed in the Laboratory de Phytopathology and in the vegetation house of CECA/UFAL. The pathogen was obtained trough the isolations of cowpea with symptoms of the disease and, afterwards, it was cultivated in sterilized rice. For in vitro control, the antagonists isolates were matched with the pathogen in PDA medium, for the purpose of evaluating the reduction of growth and the hyperparasitism. For the biofumigations of the soil, the organic materials poultry litter, mussel, sugar cane bagasse, bean residue, cassava scuff were dehydrated in stove at 55ºC for 96h, grinded in the concentrations of 10% and 20% (v/v) and incorporated to the substrate, infested for 20 days and compared to the group treated with methyl thiophanate and to the control. After thirty days, the seedlings were evaluated about the incidence and the suppression of the disease. In the in vivo control the seeds were microbiolized with antagonists (C110, C21, ENF24, R14 and Trichoderma harzianum), the fungicide and saline solution for the control. The substrate was infested with the pathogen, two days after the sow, and after 30 days were evaluated. For the natural substances, 21 days old seedlings were pulverized with cassava flour wastewater extract (40%), eucalyptus oil (1%), peppermint (1%), Ecolife® (2%), methyl thiophanate (0,7 g/L) and water for the control and after two days, the substrate was infested with the pathogen. Six days after the inoculation, a new pulverization was done. The mineral fertilization was done in the sow trough Sarruge solution and doses of calcium silicate and sodium, 50, 100, 500 and 1000mg/L-1 and water for the control. The substrate was infested two days after that and a second fertilization was done 10 days after the sow. After 30 days the evaluations took place. The antagonists R14, C16, ENF 24 and T. harzianum inhibited the pathogen with RC from 42 up to 57%. Trichoderma has the hyperparasitic capacity. The incorporation of organic material was not efficient in the control of the disease. The in vivo antagonists reduced the incidence of the disease, but it was effective in the suppression. The oils and plant extracts were not efficient in reduced the incidence. The mineral fertilizations was not able to suppress the disease. / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / O feijão-caupi, Vigna unguiculata (L.) Walp., é conhecido como feijão-de-corda e feijão-verde sendo uma das principais culturas exploradas pelos pequenos produtores no Nordeste brasileiro. Dentre os fitopatógenos que afetam sua produtividade, destaca-se Sclerotium rolfsii Sacc. que causa a podridão de colo em diversos cultivos do mundo. O objetivo do trabalho foi avaliar o controle alternativo de S. rolfsii Sacc. em mudas de V. unguiculata (L.) Walp. através do uso de antagonistas, incorporação de resíduos orgânicos ao solo, utilização de óleos essenciais, extratos vegetais e ecolife® e nutrição mineral. O trabalho foi desenvolvido no Laboratório de Fitopatologia e em casa de vegetação do CECA/UFAL. O patógeno foi obtido pelo isolamento de folhas de feijão-caupi com sintomas da doença e depois cultivados em arroz esterilizado. Para controle in vitro , os isolados de antagonistas foram pareados com o patógeno em meio de BDA, para avaliar a redução de crescimento micelial (RC) e o hiperparasitismo. Para a biofumigação do solo, as matérias orgânicas cama de frango, marisco, bagaço de cana, resíduo de feijão, raspa de mandioca foram desidratadas em estufa 55ºC por 96h, moídas e incorporadas ao substrato infestado, em concentrações de 10% e 20% (v/v) por 20 dias e comparadas ao tiofanato metílico e a testemunha. Após 30 dias, as plantas foram avaliadas quanto à incidência da doença e desenvolvimento da planta. No controle in vivo as sementes foram microbiolizadas com os antagonistas (C110, C21, ENF24, R14 e Trichoderma harzianum), o fungicida e água salina para testemunha. O substrato foi infestado com o patógeno, dois dias após o semeio foram avaliadas com 30 dias. Para as substâncias naturais, plantas de feijão-caupi, com 21 dias de idade foram pulverizadas com extrato de manipueira (40%), óleo de eucalipto (1%), hortelã pimenta (1%), Ecolife® (2%), tiofanato metílico (0,7 g/L) e água para a testemunha. Após dois dias, o substrato foi infestado com o patógeno. Ao completar seis dias da infestação, uma nova pulverização foi realizada. A adubação mineral foi realizada no semeio através de solução de Sarruge e doses de silicato de cálcio e sódio, 50, 100, 500 e 1000mg/L-1 e água para testemunha. O substrato foi infestado dois dias depois e uma segunda adubação foi feita 10 dias após o semeio. Os antagonistas R14, C16, ENF 24 e T. harzianum inibiram o patógeno com RC de 42 a 57%. Trichoderma teve capacidade hiperparasitária. A incorporação da matéria orgânica não foi eficiente no controle da doença. Os antagonistas in vivo reduziram a incidência da doença e contribuiu para o desenvolvimento das plantas. Os óleos e extratos vegetais não foram eficientes em reduzir a incidência da doença. A adubação mineral não foi capaz de suprimir a doença.