Orientador: Marli Teixeira Almeida Minhoni / Resumo: O tomateiro e o pimentão são importantes culturas no Brasil. Porém, como muitas culturas são atacadas pelos chamados nematóides causadores de galhas, entre estes a espécie Meloidogyne incognita Chitwood. Este nematóide representa grandes perdas em ambas culturas. Estas culturas também são colonizadas por fungos micorrízicos arbusculares, os quais podem trazer vários benefícios às plantas. Dentre esses benefícios temos o auxílio na absorção de fósforo e a redução na severidade de doenças. Este trabalho analisou a interação fungos MA e as raças 2 e 3 de M. incognita em plantas de tomateiro e pimentão. Observou-se que, em tomateiro, os fungos MA, aumentaram os teores de fósforo na parte aérea, sendo que Glomus clarum foi mais eficiente que Gigaspora margarita. Ambas as raças de M. incognita foram bastante agressivas às plantas de tomateiro, porém os fungos MA foram eficientes em auxiliar a planta a superar o dano do nematóide. G. clarum, G. margarita e Glomus etunicatum melhoraram o desenvolvimento de plantas de pimentão, aumentaram os teores de fósforo na parte aérea e auxiliaram as plantas a superar os danos provocados pelo nematóide. / Abstract: Tomato and pepper plants are important cultures in Brazil. But as many cultures are attacked by named root-knot nematodes , among them the Meloidogyne incognita (Kofoid & White) Chitwood races. This nematode represents big losses in both cultures. Those cultures are colonized by VA-mycorrizal fungi and they can help the plants. From among these benefices we have an aid in phosphorus assimilation and a reduction in the hardness of diseases. This work has analyzed the interaction between AM fungi and the races two and three of M. incognita in tomato plants and pepper plants. We observed in tomato plants that AM fungi has increased the phosphorus content in the shoot so Glomus clarum was more efficient than Gigaspora margarita. Both races of M. incognita were too aggressive in tomato plants, but MA fungi were efficient for helping the plants to support the damages of nematodes. G. clarum, G. margarita and Glomus etunicatum got better the development of pepper plants. They increased the phosphorus assimilation in the shoot and helped the plants to support the damage by nematodes. / Doutor

CoordenaÃÃo de AperfeÃoamento de Pessoal de NÃvel Superior / O feijÃo-de-corda [Vigna unguiculata (L.) Walp.] à uma importante leguminosa usada como alimento, sendo cultivada, primariamente, nas savanas secas do Continente Africano, na Ãsia e na AmÃrica do Sul, cobrindo mais de 12 milhÃes de hectares, com produÃÃo anual de cerca de 3 milhÃes de toneladas. O feijÃo-de-corda se constitui numa cultura de extrema importÃncia em zonas semi-Ãridas dos trÃpicos, caracterizadas por baixa precipitaÃÃo pluviomÃtrica, altas temperaturas, solos arenosos e de baixa fertilidade. Dentre os organismos que causam doenÃas nos vegetais, os nematÃides geram prejuÃzos anuais de, aproximadamente, vÃrios bilhÃes de dÃlares na agricultura mundial e as espÃcies Meloidogyne incognita e Meloidogyne javanica sÃo as mais danosas. Uma das principais culturas atacadas pelos nematÃides à o feijÃo-de-corda. Em funÃÃo disso, existe sempre demanda para se buscar melhorar a compreensÃo dessa relaÃÃo entre hospedeiros e patÃgenos objetivando criar novas estratÃgias para minimizar eventuais perdas. O presente trabalho teve como objetivo identificar as proteÃnas que tÃm sua expressÃo diferenciada em raÃzes de feijÃo-de-corda resistente ao Meloidogyne incognita (inoculadas e nÃo inoculadas) usando eletroforese bidimensional (2D) associada à espectrometria de massas de proteÃnas extraÃdas da raiz total e de mitocÃndrias de raiz do cv PitiÃba (CE-31), resistente ao nematÃide, inoculada com este patÃgeno, em comparaÃÃo com plantas nÃo-inoculadas (controle) em conjunto com anÃlise de expressÃo gÃnica semi-quantitativa (PCR semi-quantitativa). Os resultados aqui obtidos demonstraram que houve alteraÃÃo de pelo menos 22 proteÃnas nas amostras de raÃzes inoculadas e mais 22 proteÃnas de mitocÃndrias de raÃzes, quando comparados com seus respectivos controles e que o Ãpice das suas alteraÃÃes ocorriam por volta do 6 dia apÃs a inoculaÃÃo do patÃgeno. As proteÃnas de mitocÃndrias nÃo puderam ser identificadas com confiabilidade. Dentre as proteÃnas de raÃzes totais identificadas podemos destacar as PR-1, 2 e 3, que sÃo reconhecidamente proteÃnas de defesa, ascorbato peroxidase e superÃxido dismutase (enzimas anti-estresse oxidativo) e uma leghemoglobina, que pode ser o primeiro relato dessa proteÃna nesse patossistema. As anÃlises de expressÃo gÃnica concordaram perfeitamente com os achados proteÃmicos. / The cowpea [Vigna unguiculata (L.) Walp.] is an important leguminous used as food, being cultivated, mostly in arid African savannas , Asia and south of America, covering more over 12 millions of hectares, with annual production about 3 millions of tons. The cowpea itâs a culture greatly important in tropics zones characterized by low pluviometric precipitation, high temperature , gravely soils and poor fertility. Among the pathogenic organism that cause plant disease, the nematode, mainly Meloydogine sp. generate annual losses about several billions of dollars all over the world and the Meloidogyne incognita and Meloidogyne javanica species be the most damagers. One of the main cultures attacked by nematodes is the cowpea. Regard it, there is always needs to improve the knowledge about the relationship between host and pathogens aimed develop novels strategies to diminish eventual losses. The present work aimed identify the differential expression of proteins in Meloidogyne incognita resistant cowpea total roots and mitochondria roots cv PitiÃba (CE-31), inoculated and mock inoculated, using 2D electrophoresis assay associated with MS identification and gene expression analyses by semi-quantitative PCR. The results obtained showed at least 22 altered proteins in inoculated total roots and more 22 proteins of mitochondria roots, when compared with their respective controls and the top of alterations occurred next to 6 day after inoculation with pathogen. Mitochondria proteins cannot be identified reliability. Among the total roots proteins we can emphasize PR-1, 2 and 3, recognized as defense proteins, ascorbate peroxidase and superoxide dismutase (anti-oxidative brush enzymes) and a leghemoglobin, which can be the first report about this protein in this pathosystem. The gene expression analyses coincided with proteomics finds.

Submitted by Mario BC (mario@bc.ufrpe.br) on 2016-12-05T12:55:30Z No. of bitstreams: 1 Matheus Silva e Silva.pdf: 927208 bytes, checksum: b577e7fbf3a3419012f5dc12beda0cda (MD5) / Made available in DSpace on 2016-12-05T12:55:30Z (GMT). No. of bitstreams: 1 Matheus Silva e Silva.pdf: 927208 bytes, checksum: b577e7fbf3a3419012f5dc12beda0cda (MD5) Previous issue date: 2015-07-31 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / One of the most economic importance crops in Brazil, the sugarcane (Saccharum spp.) cultivation is severely affected by several biotic and abiotic factors. Within the biotic one the damage caused by Meloidogyne species points out due to lesions in parasite roots resulting in decreases of productivity. The non-availability of root-knot nematode resistant varieties becomes more serious after reduction in nematicides use, increasing efforts in turn resistant varieties a possible alternative. Therefore, the objectives of the present work was screening promising sugarcane genotypes for M. incognita and M. javanica resistance (Study 1) and determine the resistance mechanisms involved through evaluation of penetration, development and reproduction of the nematodes in the selected material (Study 2), under greenhouse. In Study 1, evaluations were carried out 120 days after inoculation with 9000 eggs per plant and based on host development and nematode reproduction. There was significant difference (P≤0.05) within genotypes for all plant development and nematode reproduction variables. Although all genotypes were susceptible (FR≥1.0) to M. incognita and M. javanica, in some of them the reproductive factor (FR) was lower than the control. In Study 2, five genotypes were inoculated with 20000 eggs per plant and evaluations carried out at 5, 10, 15, 20, 40 and 60 days after inoculation. There was difference in nematode penetration among genotypes, and despite juveniles of both nematode species penetrated all genotypes at five days after inoculation, nematode development significantly differed among clones. At 60, but 45, days after inoculation eggs were evident in the genotypes. Reduced penetration and development rates were demonstrate by the genotypes RB041594 e RB071095. / De relevante importância econômica para o Brasil, o cultivo da cana-de-açúcar (Saccharum spp.) sofre grande influência de diversos fatores bióticos e abióticos. Dentre os bióticos, destacam-se os danos causados por espécies de Meloidogyne, devido às lesões nas raízes parasitadas, resultando em redução de produtividade. A não disponibilidade de variedades resistentes aos nematoides das galhas tornou-se mais séria com a redução do uso de nematicidas, intensificando os esforços para tornar a resistência varietal uma alternativa possível. Dessa forma, o objetivo do presente estudo foi selecionar genótipos RB de cana-de-açúcar promissores para o desenvolvimento de variedades resistentes a M. incognita e M. javanica (Estudo 1) e determinar os mecanismos de resistência envolvidos avaliando-se a penetração, desenvolvimento e reprodução das espécies de nematoides no material selecionado (Estudo 2), em condições de casa de vegetação. No estudo 1, as avaliações foram realizadas 120 dias após a inoculação com 9000 ovos por planta e fundamentaram-se no desenvolvimento do hospedeiro e na reprodução dos nematoides. Houve diferença significativa (P≤0,05) entre os genótipos RB para todas as variáveis de desenvolvimento da planta, e para ambas as espécies de Meloidogyne. Todos os genótipos foram suscetíveis (FR≥1,0) à M. incognita e M. javanica, entretanto, em alguns o fator de reprodução foi inferior ao do tratamento controle. No Estudo 2, cinco genótipos foram inoculados com 20.000 ovos por planta e as avaliações realizadas aos 5, 10, 15, 20, 40 e 60 dias após a inoculação. Houve diferença entre os genótipos avaliados com relação à penetração de juvenis de segundo estádio. Cinco dias após a inoculação foi visualizado juvenis no interior das raízes de todos os tratamentos, mas o desenvolvimento dos nematoides diferiu significativamente entre os clones. Aos 45 dias após a inoculação não foi evidenciada a presença de ovos em nenhum dos genótipos testados, o que só ocorreu aos 60 dias após a inoculação. Reduzidas taxas de penetração e desenvolvimento foram demonstradas pelos genótipos RB 041594 e RB071095.

O gênero Eucalyptus possui diversas espécies que apresentam grande valor comercial, sendo utilizado principalmente para produção de energia (carvão e lenha) e celulose-papel. Na literatura, entretanto, existem poucas informações acerca de nematoses nessa cultura. Além dessa carência e de sua importância econômica, o eucalipto é comumente cultivado em locais de alta incidência de Pratylenchus brachyurus e Meloidogyne incognita, além de ser consorciado com diversas outras culturas intolerantes, como por exemplo a soja. Dessa forma há uma necessidade de conhecer a reação das espécies de eucalipto a estes nematoides. Dito isso, o presente trabalho teve por objetivo testar a reação de algumas espécies de eucalipto a P. brachyurus e M.incognita. Foram realizados 6 experimentos, 5 envolvendo o nematoide das lesões e 1 com o nematoide de galhas. Os tratamentos foram inoculados com uma população inicial de nematoides e, após 90 dias, estimava-se o fator de reprodução (FR) e nematoides por grama de raiz (Nem/g). Adotou-se o critério baseado em Oostenbrink (1966), onde a espécie foi considerada suscetível quando apresenta FR >= 1 e resistente quando FR < 1. Dos eucaliptos testados, 6 apresentaram reação suscetível e 4 reação resistente para P. brachyurus. As espécies com maiores FR estão entre as mais utilizadas no Brasil, exceto E. saligna. Ressalta-se que estas não necessariamente são resistentes, necessitando apenas de um período maior para o estabelecimento do patógeno. Observou-se indícios de que E. saligna e E. camaldulensis sejam intolerantes a P.brachyurus. Para M. incognita raça 3 todas as espécies testadas foram resistentes. Aconselha-se a realização de experimentos com períodos superiores a 100 dias para uma correta classificação da reação de hospedeiro. O eucalipto é uma opção para aproveitamento de áreas infestadas com M. incognita raça 3, porém seu uso mostra-se um risco em áreas infestadas com P. brachyurus. / The Eucalyptus has several species with high commercial value and is used mainly for energy purpose (Charcoal and firewood) and paper. In the literature, however, there is little information about plant parasitic nematodes in this woody specie. In addition to this lack and its economic importance, eucalypts is also commonly cultivated in areas with high incidence of Pratylenchus brachyurus and Meloidogyne incognita, and consorted with several intolerant crops, such as soybean. Thus, there is a need to know the host status of the eucalypts species to these nematodes. With that, this work aimed to test the reaction of some Eucalyptus species to P.brachyurus and M. incognita. For this purpose six experiments were performed, five to test the lesion nematode and one to test the root-knot nematode. The treatments were inoculated and, after 90 days, it was estimated the reproduction factor (FR) and nematodes per gram of root (Nem/g). The present work adopted the criteria based on Oostenbrink (1966), where the species was considered susceptible when presenting FR >= 1 and resistant when FR < 1. Of species tested, six were susceptible hosts and four resistant host to P. brachyurus. The species with highest FR are among the most used eucalypts in Brazil, except E. saligna. It is emphasized that those species with FR < 1 are not necessarily resistant, requiring only a longer period for the pathogen establishment. There was some evidence that E. saligna and E. camaldulensis are intolerant hosts to P. brachyurus. To M. incognita Race 3 all tested species were resistant. It is advisable to perform experiments for periods longer than 100 days to a correct classification of host status. Eucalyptus is an option for utilization of areas infested with M. incognita race 3, but its use appears to be a risk in Pratylenchus brachyurus infested areas.

Thesis (M.Sc. (Horticulture)) -- University of Limpopo, 2017. / Root-knot (Meloidogyne species) nematodes are economically destructive pathogens of over 3000 species, whereas others have resistance to Meloidogyne species. Wild watermelon (Cucumis africanus) and wild cucumber (Cucumis myriocarpus) are highly resistant to Meloidogyne species, particularly M. incognita and M. javanica. The two Cucumis species are used in inter-generic grafting with watermelon (Citrullus lanatus) as nematode resistant rootstocks. Also, the two Cucumis species are used in traditional medicine and in plant-parasitic nematode management as phytonematicides. The form of nematode resistance, which is essential in plant breeding, is not documented for the two Cucumis species. The objective of this study was to determine the form of nematode resistance in the two Cucumis species to M. incognita and M. javanica under greenhouse conditions. Four parallel experiments were each conducted under greenhouse conditions. Uniform six-week old Cucumis seedlings were transplanted into 250 ml polystyrene cups filled with 200 ml growing medium of steam-pasteurised fine sand. A week after transplanting, Cucumis seedlings were each infested by dispensing approximately 100 M. incognita second-stage juveniles (J2) or M. javanica J2 using a 20 ml plastic syringe by placing into 5-cm-deep furrow around the seedling stem and covered with growing medium. Treatments (periodic harvest intervals) were arranged in a randomised complete block design, replicated five times. Five seedlings from each experiment were harvested every second day, for 30 days, with stained roots being assessed for necrotic spot (suberised cells) number, giant cell number, proliferation of rootlet interference number and root gall number. Periodic harvest intervals were highly significant (P ≤ 0.01) on necrotic spot number, proliferation of rootlet interference number and root gall number in C. africanus-M. incognita relations, but were not significant for giant cell number. Treatments contributed 59, 64 and 50% in total treatment variation (TTV) of necrotic spot number, proliferation of rootlet interference number and root gall number, respectively. Harvest period had highly significant effects on necrotic spot number, giant cell number, proliferation of rootlet interference number and root gall number in C. africanus-M. javanica relations. Treatments contributed 55, 71, 63 and 59% in TTV of necrotic spot number, giant cell number, proliferation of rootlet interference number and root gall number, respectively. Periodic harvest intervals were significant (P ≤ 0.05) on giant cell number and highly significant on root gall number in C. myriocarpus-M. incognita relations. However, there were no significant treatment differences on necrotic spot number and proliferation of rootlet interference number. Treatments contributed 57 and 57% in TTV of root gall number and giant cell number, respectively. Harvest period had highly significant effects on giant cell number, proliferation of rootlet interference number and root gall number, but were not significant on necrotic spot number in C. myriocarpus-M. javanica relations. Treatments accounted for 67, 49 and 53% in TTV of giant cell number, proliferation of rootlet interference number and root gall number, respectively. In conclusion, the mechanism of resistance to M. incognita and M. javanica in both C. africanus and C. myriocarpus was post-infectional nematode resistance, which has attributes for introgression into commercial nematode-susceptible Cucumis cultivars. / Agricultural Research Council (ARC), National Research Foundation of South Africa; and the ARC-Universities Collaboration Centre for Smallholder Farmers

Thesis (M. Agric. (Plant Protection)) -- University of Limpopo, 2019 / Damage and significant losses of potato cultivar due to Meloidogyne incognita has become a serious challenge, after the withdrawal of synthetic chemical nematicides due to their environment-unfriendliness. Various alternatives have been investigated each with a wide range of drawbacks. Most phytonematicides were highly phytotoxic to crops, while their effects on nematode suppression were highly variable. The use of Nemarioc-AG phytonematicide at pre- and post-emergence would help in determining the level that is effective in supressing M. incognita without being phytotoxic. The objective of this study was to determine whether Nemarioc-AG phytonematicide could serve as pre- and post-emergent phytonematicide without inducing phytotoxicity while suppressing population densities of M. incognita. For achieving this objective, treatments, namely, 0, 2, 4, 8 and 16 g of Nemarioc-AG phytonematicide, were arranged in a randomised complete block design (RCBD), with 7 replicates. Potato seed tubers were sown into 20 cm pots, Nemarioc-AG phytonematicide placed above the tubers and covered with soil, after initiation of treatments 5 000 eggs and second stage juveniles (J2) of M. incognita per plant were inoculated. For post-emergent, treatments, replications and design were the same as in pre-emergent. Potato seed tubers were sown and inoculated with 5000 eggs and second-stage juveniles (J2) of M. incognita per plant after 100% emergence. Nemarioc-AG phytonematicide were applied 7 days after inoculation. Trials were conducted in autumn (February-April) 2017 (Experiment 1) and repeated in autumn 2018 (Experiment 2). Plant growth variables and selected nutrient elements were collected and analysed using the Curve Fitting Allelochemical Response Data (CARD) model and lines of best fit, respectively. In pre-emergent application, Experiment 1, MCSP was established at 1.95 g, with the overall xii sensitivity (∑k) being equal to zero. Therefore, in Experiment 1 and Experiment 2, all nutrient elements to increasing concentration of Nemarioc-AG phytonematicide exhibited negative quadratic relations. In both Experiments, nematode variables over increasing concentration of Nemarioc-AG phytonematicide on potato exhibited negative quadratic relations, except in Experiment 1, where J2 in roots exhibited positive quadratic relations, with models ranging between 72 to 99%. In post emergent, Experiment 1, MCSP was established at 1.57 g, with the overall sensitivity (∑k) being equal to 2. In Experiment 1 and Experiment 2, nutrient elements over increasing concentration of Nemarioc-AG phytonematicide exhibited positive and negative quadratic relations, with models ranging from 89 to 97%. In Experiment 1, nematode variables over increasing concentration of Nemarioc-AG phytonematicide exhibited negative quadratic relations, with models ranging between 92 and 98%. Positive and negative relations suggested that the product stimulated and inhibited plant growth or accumulation of selected essential nutrient elements, respectively. Increasing concentration of Nemarioc-AG phytonematicide had stimulated certain plant variables and inhibited population densities of M. incognita in pre- and post emergent application; therefore, this product was suitable for use as pre- and post emergent in management of nematodes on the test crop.

Utilizing resistant cultivars is a main control strategy for root-knot nematodes in flue-cured tobacco (Nicotiana tabacum L.). Most commercial cultivars possess the Rk1 gene, providing resistance to races 1 and 3 of Meloidogyne incognita and race 1 of M. arenaria. This initiated a shift in root-knot populations to other species and races, creating a need for resistance to those populations. Numerous cultivars possess a second resistance gene, Rk2. Greenhouse experiments investigated whether possessing both Rk1 and Rk2 increases resistance to a variant of M. incognita race 3 compared to either gene alone, and if high soil temperatures impact their efficacy. Root galling, numbers of egg masses and eggs, and the reproductive index were compared from roots of Coker 371-Gold (susceptible), NC 95 and SC 72 (Rk1Rk1), T-15-1-1 (Rk2Rk2), and STNCB-2-28 and NOD 8 (Rk1Rk1 and Rk2Rk2). The same data were analyzed from plants in open-top root zone cabinet growth chambers set to 25ºC, 30ºC, and 35ºC to examine if resistance is temperature sensitive. Despite variability, Rk1Rk2 entries conferred greater resistance than entries with Rk1 or Rk2 alone. Entries with Rk1 alone reduced galling and reproduction compared to the susceptible control, whereas T-15-1-1 (Rk2) did not, but often suppressed reproduction. An apparent reduction in nematode reproduction was observed at 25ºC and 30ºC on entries possessing Rk1 and Rk1Rk2 compared to the control and Rk2. However, no apparent differences in reproduction occurred on Rk1 and/or Rk2 entries at 35ºC compared to the control, indicating parasitism increased on resistant entries at higher temperatures. / Master of Science in Life Sciences

Made available in DSpace on 2017-07-10T17:40:55Z (GMT). No. of bitstreams: 1 Edilaine_D_V_GoncalvesTrevisoli.pdf: 2707364 bytes, checksum: 7b7a664f9727d5909c470f377ee155e8 (MD5) Previous issue date: 2016-02-25 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / The induction of resistance in plants to pathogens is an alternative method of disease control, wich involves activation of plant resistance mechanisms such as induction of phytoalexins. The elicitors molecules are able to induce and activate those responses, and therefore, techniques have sought to isolate and characterize fractions with elicitor character. The study aimed to purify, through ion exchange chromatography and gel filtration chromatography, eliciting molecules from pathogenic nematodes, and test them in phaseolin induction in beans hypocotyls beans and gliceolin in soybean cotyledons. The buffer solution Tris HCl 0.05 M (pH 6.8) was used as control and the acibenzolar-S-methyl (50 mg a.i. L-1) and Saccharomyces cerevisiae (20 mg mL-1) were used as induction standard treatments. Ion exchange chromatography (IEC) and gel filtration chromatography (GC) were performed to separate fractions with eliciting power from 500 female nematodes of Meloidogyne incognita and Meloidogyne javanica. For purification of elicitors from Meloidogyne javanica, through IEC, six glycidic fractions and six glycoproteins were obtained. These were purified on GC, obtaining sixty-three fractions. They have been classified according to their nature, as twenty-six glycidic and thirty-seven glycoprotein with molecular weights ranging from 29.19 to 2989.25 kDa. Regarding the elicitors purification of Meloidogyne incognita through IEC, nine glycidic and five glycoprotein fractions were obtained. From these fractions, a total of fifty-eight fractions was obtained through GC, twenty-five glycidic and thirty-three glycoprotein with molecular weights ranging from 37.42 to 200.32 kDa. From the fractions purified from Meloidogyne javanica eight had inducing potential of phaseolin. For gliceolin fifteen fractions showed inducing effect. Regarding the fractions purified from Meloidogyne incognita, no fraction has inductive potential of phaseolin superior to the standard treatment. However, twenty-two fractions suppressed phytoalexin inducing activity. For gliceolin ten fractions induced the same, whereas, twenty-three fractions suppressed the induction of gliceolin. Chromatography was efficient in the purification of elicitors compounds. Compounds with suppressing characteristics of gliceolin and phaseolin were checked in bioassays. For those fractions obtained through IEC, and then submitted to GC that did not induce phytoalexin, it is suggested that molecules need to act together to have elicitor effect and thus induce defense response in the plant / A indução de resistência em plantas contra patógenos é um método de controle alternativo de doenças, e que envolve a ativação dos mecanismos de resistência da planta, como a indução de fitoalexinas. As moléculas eliciadoras possuem a capacidade de induzir e ativar tais repostas, e assim sendo, técnicas têm buscado isolar e caracterizar frações com caráter eliciador. O trabalho teve por objetivo purificar, por cromatografia de troca iônica cromatografia de filtração em gel, moléculas eliciadoras a partir de nematoides fitopatogênicos, e testá-las na indução de faseolina em hipocótilos de feijoeiro e gliceolina em cotilédones de soja. O tampão Tris HCl 0,05 M (pH 6,8) foi utilizado como tratamento controle e o acibenzolar-S-metil (50 mg i.a. L-1) e o Saccharomyces cerevisiae (20 mg mL-1) foram utilizados como tratamento padrão de indução. Cromatografia de troca iônica (CTI) e cromatografia de filtração em gel (CFG) foram realizadas para separar frações com poder eliciador a partir de quinhentas fêmeas de nematoides de Meloidogyne incognita e Meloidogyne javanica. Para a purificação de eliciadores a patir de Meloidogyne javanica, por CTI, foram obtidos seis frações glicídicas e seis glicoproteicas. Estas, por sua vez, foram purificadas em CFG, sendo obtidos no total sessenta e três frações. As mesmas foram classificadas de acordo com sua natureza, sendo vinte e seis glicídicas e trinta e sete glicoproteicas, com massas moleculares variando de 29,19 a 2.989,25 kDa. Em relação a purificação de eliciadores de Meloidogyne incognita por CTI, foram obtidos nove frações glicídicas e cinco glicoproteicas. A partir destas, foram obtidos por CFG um total de cinquenta e oito frações, sendo vinte e cinco glicídicas e trinta e três glicoproteicas, com massas moleculares variando de 37,42 a 200,32 kDa. Das frações purificadas a partir de Meloidogyne javanica oito apresentaram potencial indutor de faseolina. Para gliceolina quinze frações mostraram efeito indutor. Em relação as frações purificadas a partir de Meloidogyne incognita, nenhuma fração apresentou potencial indutor de faseolina superior ao tratamento padrão. Entretanto, vinte e duas frações suprimiram a atividade de indução de fitoalexina. Para gliceolina dez frações induziram a mesma, enquanto que, vinte e três frações suprimiram a indução da gliceolina. A cromatografia foi eficiente na purificação de compostos eliciadores. Compostos com características supressoras de gliceolina e faseolina foram verificadas nos bioensaios. Para aquelas frações obtidas por CTI e posteriormente submetidas a CFG que não induziram fitoalexina, sugere-se que as moléculas necessitam atuar juntas para haver efeito eliciador e assim induzir a resposta de defesa no vegetal

Meloidogyne incognita est le nématode causant le plus de dégâts en agriculture. Sa particularité est d'être un organisme à reproduction asexuée obligatoire. Une femelle engendre des clones a priori 100% identiques génétiquement. Pourtant, M. incognita est capable de faire preuve d'une grande plasticité phénotypique lui permettant de répondre à de nouveaux environnements. Un exemple est le déterminisme du sexe, un phénotype lié aux conditions environnementales et semblant impliquer des régulations chromatiniennes. Un autre exemple est la capacité à contourner les résistances des plantes (virulence), un caractère héréditaire mais non-Mendelien. Dans le cadre de cette thèse, j'ai cherché à tester l'implication des mécanismes épigénétiques dans la plasticité phénotypique en absence de sexe de M. incognita. A ces fins, j'ai évalué la conservation des mécanismes épigénétiques chez les nématodes à galles. Cette approche a permis de pointer que les mécanismes connus chez C. elegans sont conservés chez les nématodes parasites de plantes. Puis, une méthodologie de ChIP-seq a été mise en place afin de comparer les profils d'accumulation des marques d'histones chez M. incognita au cours de la réponse aux conditions environnementales. Cette stratégie a permis la mise en évidence 1- de patrons d'histones modifiées marquant le développement du parasite et 2- de régions génomiques comportant plus de 300 gènes dont des candidats facteurs d'avirulence déjà décrits dans la littérature spécifiquement perdue entre M. incognita (a)virulents. Ces travaux de thèse présentent un intérêt fondamental sur la compréhension de l'évolution d'un organisme en absence de reproduction sexuée. / Meloidogyne incognita is the most damaging plant-parasitic nematode in agriculture. M. incognita reproduces in an asexual way by obligatory parthenogenesis. Genetically identical individuals develop from females and form clonal populations. Although these clones share the same genetic heritage, modifications of their phenotype can be observed when they are exposed to unfavorable environments. This phenotypic plasticity is characterized through two phenotypes of interest: sex-differentiation and virulence (i.e. capacity to parasite a resistant crop). Sex-differentiation varies among environmental conditions and was reported to be linked to decondensed chromatin regions. Virulence is an heritable character transmitted in a non-Mendelian way. Our study focuses on identifying the role of epigenome in the generation of phenotypic variability. To this end we detailed the presence of proteins involved in epigenetic regulations in Meloidogyne spp. We also developed a ChIP-seq assay to compare histone modifications between different developmental stages and between virulent and avirulent parasites. Our results allow to detect specific histone patterns associated with M. incognita development. These results lead us to propose a model that could explain sex determination in M. incognita. We also could link virulence acquisition with the loss of some specific genomic regions that contains more than 300 genes including already described potential avirulence factors. This study opens the way for analyzing the role of epigenetic mechanisms at a whole genome scale, and allows to identify novel biological processes involved in phenotypic variation in asexual organisms.

Submitted by Marco Antônio de Ramos Chagas (mchagas@ufv.br) on 2017-04-27T17:02:23Z No. of bitstreams: 1 texto completo.pdf: 404797 bytes, checksum: 73099bf5c75d1919a150a4794868d245 (MD5) / Made available in DSpace on 2017-04-27T17:02:23Z (GMT). No. of bitstreams: 1 texto completo.pdf: 404797 bytes, checksum: 73099bf5c75d1919a150a4794868d245 (MD5) Previous issue date: 2004-02-19 / Conselho Nacional de Desenvolvimento Científico e Tecnológico / A busca de novas alternativas visando o controle de fitonematóides em substituição aos nematicidas convencionais é, hoje, uma preocupação mundial. A utilização de extratos botânicos e a incorporação de plantas ou parte delas têm sido estudadas por pesquisadores em todo o mundo como uma destas alternativas. Diversas espécies do gênero Mucuna têm sido bastante utilizadas no Brasil, seja em programas de adubação verde, seja na redução da população de nematóides em áreas cultivadas. Além disso, algumas substâncias isoladas de mucuna preta (Mucuna pruriens var. utilis) apresentaram atividade nematicida contra M. incognita e Heterodera glycines. Desta forma, objetivou-se neste trabalho verificar a eficácia de extratos aquosos de folhas e de sementes de mucuna preta, aplicados via pulverização foliar, tratamento de sementes ou diretamente ao solo, e também o efeito da incorporação de diferentes quantidades do material vegetal seco da leguminosa sobre a população de M. javanica e M. incognita, em raízes de tomateiro. Na primeira parte do trabalho, os extratos vegetais foram preparados através da mistura das folhas secas ou das sementes de mucuna com água destilada, na proporção 1:10 (m:v), procedendo-se à trituração do material após 24 horas da mistura. Finalmente, os extratos foram filtrados em papel Whatman N 0 1 e utilizados logo em seguida. Nos três ensaios, as avaliações foram realizadas 60 dias após a infestação do solo e transplantio das mudas de tomateiro, quando se mensuraram as seguintes variáveis: altura e peso da parte aérea das plantas e o número de galhas e de ovos presentes no sistema radicular das plantas. No primeiro ensaio, extratos de folhas e de sementes de mucuna, de folhas de manjericão (Ocimum basilicum L.) e água foram pulverizados em plantas de tomate aos 0, 7 e 14 dias após a infestação do solo com ovos de M. incognita e M. javanica. No segundo ensaio, sementes de tomateiro foram tratadas com água ou com os extratos de folhas ou de sementes de mucuna e as mudas originadas destas sementes foram transplantadas em solo infestado com M. incognita e M. javanica. No terceiro ensaio, os extratos de mucuna foram adicionados ao solo aos 0, 7 e 14 dias da infestação deste com ovos das duas espécies de nematóides. A pulverização dos três extratos reduziu o número de galhas de M. incognita em raízes de tomateiro, em relação à testemunha, mas o tratamento de sementes com os extratos não afetou significativamente a altura e massa da parte aérea das plantas, assim como o número de ovos e galhas de M. incognita ou M. javanica. Apenas o número de ovos de M. javanica foi reduzido pela adição dos extratos de sementes de mucuna ao solo. A parte aérea seca e picada de mucuna preta ou de tomateiro (0, 2, 4, 6 e 8 g por vaso) foi adicionada a vasos de argila de 0,6 litro de capacidade, infestados com 2000 ovos de M. incognita ou M. javanica, visando estudar o efeito da matéria orgânica de uma planta antagonista e de uma boa hospedeira no controle desses nematóides. Sessenta dias após a infestação do solo, foi realizada a avaliação da altura e peso da parte aérea das plantas, além do número de galhas e de ovos presentes no sistema radicular destas plantas. A incorporação ao solo da parte aérea de mucuna preta ou tomate, independente da quantidade utilizada, não afetou a altura e o peso da parte aérea das plantas inoculadas com M. incognita e M. javanica. A adição de matéria orgânica de mucuna preta ao solo, a partir de 2 g/vaso, reduziu o número de galhas das duas espécies de nematóides, sendo que nas parcelas infestadas com M. incognita, as duas maiores doses promoveram maior efeito. A incorporação do material vegetal seco de mucuna, nas doses de 6 e 8 g/vaso, influenciou negativamente a reprodução de ambas as espécies de nematóides. Não foi observado nenhum efeito significativo na redução do número de galhas e de ovos de M. incognita e M. javanica nas parcelas nas quais foram adicionados os materiais vegetais de tomateiro. Esse trabalho confirma que a utilização de extratos e/ou matéria orgânica de mucuna preta e outras plantas pode ser muito útil no manejo de nematóides. / The search for new alternatives to control nematodes, to replace the use of conventional nematicides is, today, a world concern. Several natural products obtained from different plant species with nematicidal or nematostatic properties have shown to be promising to be used in field conditions. Consequently, the use of botanical extracts and soil amendment with plant parts have deserved special attention by researchers, all over the world. Several species of Mucuna have been used in Brazil, not only as green manure but also to reduce nematode populations in cultivated areas. Some substances isolated from velvetbean (Mucuna pruriens var. utilis) have shown activity against M. incognita, Heterodera glycines and other nematodes. Thus, this study was conducted to verify the effectiveness of aqueous extracts of leaves and seeds of velvetbean; applied as foliar spray, seed treatment or directly in the soil, as well as the effect of soil amendment using that plant species on the control of M. javanica and M. incognita. First, the botanical extracts were prepared through the mixture of the dry leaves or seeds with distilled water (1:10 w/v). The material was ground 24 hours later, filtered in Whatman paper and used right away. In all the three experiments, the evaluations were accomplished 60 days after soil infestation and the following parameters were evaluated: height and weight of the tops and the numbers of galls and eggs in the root systems. In the first assay, the extracts of leaves and seeds of velvetbean and of sweet basil leaves (Ocimum basilicum L.), used as control, were sprayed on tomato plants at 0, 7 and 14 days after soil infestation with M. incognita and M. javanica eggs. In the second assay, tomato seeds were treated with velvetbean extracts (from seeds and leaves) and the seedlings resulting from them were transplanted in soil infested with M. incognita and M. javanica. In the third assay, velvetbean extracts were poured in the soil 0, 7 and 14 days after infestation with eggs of both nematodes species. The foliar application of the three extracts reduced the number of galls of M. incognita in the root system of tomatoes as compared to the control, but the seed treatment with the extracts did not affect any parameter. Only the number of eggs of M. javanica was reduced by the addition of velvetbean seed extract to the soil. In the studies involving soil amendment, dry aboveground plant parts of velvetbean (0, 2, 4, 6 and 8 g) were added to clay pots (0,6 liter capacity) infested with 2000 eggs of M. incognita or M. javanica. A treatment using dry tomato tops was included in the experiment, in the same rates, to compare the effect of either type of organic amendment. Sixty days after soil infestation, the experiment was evaluated. The incorporation of velvetbean or tomato tops in the soil, independently of the amount used, did not affect the height and weight of the plants inoculated with M. incognita or M. javanica. Gall numbers were reduced for both nematode species, starting from 2 g/pot, and in the plots infested with M. incognita, the two larger rates of amendment promoted the best effect. The rates of 6 and 8 g/pot influenced the reproduction of both nematodes species negatively. No significant effect was observed in the reduction of the numbers of galls and eggs of M. incognita or M. javanica in the plots where tomato tops were used as soil amendment. This work corroborates others that show that the use of extracts and/or organic material from velvetbean and other plants can be very useful in nematode management. / Dissertação importada do Alexandria