Spodoptera frugiperda (J.E. Smith) é uma das pragas mais importantes da cultura do milho no Brasil. Vários casos de resistência de S. frugiperda a inseticidas de grupos químicos distintos já foram documentados. A busca contínua de alternativas para o controle de S. frugiperda tem possibilitado o desenvolvimento de novas tecnologias como plantas de milho geneticamente modificadas, e até mesmo a introdução de inseticidas que aliam segurança ambiental e alta atividade inseticida, como o recente grupo das diamidas. Estudos de caracterização e monitoramento da suscetibilidade de S. frugiperda são fundamentais para a implementação de programas de Manejo da Resistência de Insetos (MRI) no Brasil. Com o intuito de se prolongar a vida útil das diamidas, os objetivos deste trabalho foram: (a) caracterizar a suscetibilidade a diamidas em populações de S. frugiperda nas principais regiões produtoras de milho no Brasil na safra 2011/12; (b) monitorar a suscetibilidade de populações de S. frugiperda a chlorantraniliprole e flubendiamide coletadas na cultura do milho em oito estados do Brasil nas safras 2011/12, 2012/13 e 2013/14; (c) estimar a frequência inicial do alelo de resistência a chlorantraniliprole em populações de S. frugiperda. O método de bioensaio para a caracterização da suscetibilidade de populações de S. frugiperda a chlorantraniliprole e flubendiamide foi o de aplicação superficial do inseticida sobre a dieta. As linhas-básicas de suscetibilidade foram definidas para uma população suscetível de referência e cinco populações de campo de S. frugiperda. Posteriormente, foram definidas concentrações diagnósticas para o monitoramento da suscetibilidade para ambos inseticidas em populações de S. frugiperda. As CLs50 estimadas para as populações de S. frugiperda avaliadas variaram de 1,15 a 2,55 ?g de chlorantraniliprole/mL de água [I.A. (ppm)] (variação de 1,4 vezes), e de 1,75 a 4,17 ?g de flubendiamide/mL de água [I.A. (ppm)] (variação de 2,4 vezes). As concentrações diagnósticas de 10 e 32 ?g de chlorantraniliprole/mL e de 100 e 180?g de flubendiamide/mL foram definidas para o monitoramento da resistência. As populações de S. frugiperda apresentaram alta suscetibilidade nas concentrações diagnósticas de chlorantraniliprole e flubendiamide na safra 2011/12. No entanto, foram observadas variações na sobrevivência de 0 a 12,7% para chlorantraniliprole e de 1 a 6% para flubendiamide de acordo com a safra agrícola e localidade. A frequência do alelo resistente a chlorantraniliprole em S. frugiperda estimado pelo método de \"F2 Screen\" foi baixa para a safra 2011/12 (<0,0033). No entanto, aumento significativo na frequência do alelo resistente foi observado nas safras 2012/13 [0,0134 (0,0082 - 0,0198)] e 2013/14 [0,0176 (0,0084 - 0,0277)] em populações testadas, indicando alto risco de evolução da resistência a chlorantraniliprole no Brasil. / Spodoptera frugiperda (J.E. Smith) is one of the most important maize pests in Brazil. Several cases of S. frugiperda resistance to insecticides of different chemical class have been documented. The continuous search for alternatives to control S. frugiperda has enabled the development of new technologies such as genetically modified corn plants, and even the introduction of insecticides that combine environmental safety and high insecticidal activity, such as the recent class of diamides. Baseline studies and monitoring the susceptibility of S. frugiperda to insecticides are fundamental to the implementation of Insect Resistance Management (IRM) programs in Brazil. In order to prolong the lifetime of diamides, the objectives of this work were: (a) characterize the susceptibility to diamides in populations of S. frugiperda collected from major corn-producing regions in Brazil in 2011/12 season; (b) monitor the susceptibility of S. frugiperda populations to chlorantraniliprole and flubendiamide collected in maize in eight states of Brazil in the 2011/12, 2012/13 and 2013/14 growing seasons; (c) estimate the initial frequency of the allele for resistance to chlorantraniliprole in S. frugiperda populations. The baseline susceptibility of S. frugiperda populations to chlorantraniliprole and flubendiamide was determined using the diet surface treatment bioassays. Baselines were defined for a susceptible population and five field populations of S. frugiperda. Subsequently, diagnostic concentrations for monitoring susceptibility to both insecticides were defined. LCs50 estimated for populations of S. frugiperda ranged from 1.15 to 2.55 mg of chlorantraniliprole / mL of water [IA (ppm)] (1.4-fold variation) and 1.75 to 4.17 mg of flubendiamide / ml water [IA (ppm)] (2.4-fold variation). The diagnostic concentrations of 10 and 32 mg of chlorantraniliprole / mL and 100 and 180?g of flubendiamide / mL were defined for the resistance monitoring. The populations of S. frugiperda showed high susceptibility to chlorantraniliprole and flubendiamide in 2011/12 growing season at diagnostic concentrations. However, variations in survival from 0 to 12.7% for chlorantraniliprole and 1 to 6% for flubendiamide were observed based on growing season and location. The frequency of the resistant allele to chlorantraniliprole in S. frugiperda estimated by using \"F2 Screen\" method was low during 2011/12 growing season (<0.0033). However, significant increase in the frequency of the resistant allele was observed in 2012/13 [0.0134 (0.0082 to 0.0198)] and 2013-14 [0.0176 (0.0084 to 0.0277)] growing seasons, indicating a high risk of resistance evolution to chlorantraniliprole in Brazil.

Submitted by (edna.saturno@ufrpe.br) on 2016-06-13T13:01:53Z No. of bitstreams: 1 Carliane Rebeca Coelho da Silva.pdf: 1369576 bytes, checksum: bb63bac1307bc50b7f204a30dfb71ef2 (MD5) / Made available in DSpace on 2016-06-13T13:01:53Z (GMT). No. of bitstreams: 1 Carliane Rebeca Coelho da Silva.pdf: 1369576 bytes, checksum: bb63bac1307bc50b7f204a30dfb71ef2 (MD5) Previous issue date: 2014-03-17 / Various educational institutions and research throughout the world have devoted efforts in the discovery of new insecticidal proteins and their genes for the control of agricultural pests. Studies have shown the potential of some molecules in insect control as the Bacillus thuringiensis Bt toxins, protease inhibitors and alpha-amylase and lectins. Genes coding for these proteins have been stably integrated into the genome of transgenic plants giving them resistance to pests, such as cultures of tobacco, tomato, potato, corn and cotton with Bt resistance to lepidopteran which are widely marketed in various countries. Several studies have confirmed the economy and the ecological benefit of these cultivars in international agribusiness. In cotton field lepidopteran and coleoptera orders are the worldwide major economic problems. Embrapa develops research for the control of cotton pests in order to reduce production costs, of these, 20-30% are for pest control. Despite the important results obtained so far chemical control has still been the best answer although more costly in financial and more aggressive aspect in environmental aspect. The cry1Ia gene derived from Bacillus thuringiensis strain S1451 may be responsible for this type of control. Insecticidal activity of this gene was tested aiming at the analysis of the recombinant protein for the boll weevil (Anthonomus grandis) and Spodoptera frugiperda caterpillar. Indicating that this gene is very promising for use in molecular area aimed at obtaining cultivate transgenic cotton resistant to these two important pests. In previous work this gene was inserted into three commercial cotton cultivars by microinjection technique via ovary drip. At that moment the cultivars received two different buildings, composed of a minimum linear cassette mlc (≈ 3 Kb) and a circular complete construction ccc (≈ 15 Kb), both containing the gene cry1Ia. The integration of the gene and protein expression was analyzed by PCR of genomic DNA, semi-quantitative RT-PCR, Southern blot and ELISA. Over 1,800 transgenes were tested and the full integration of (1 copy) gene was detected by Southern blot assay and found in only a single event called T0 34 which was derived from the cultivar BRS 293 in mlc treatment, suggesting a genotype-dependent trend. The expression of Cry1Ia protein in T0 34, estimated by ELISA was similar to the commercial event Bollgard (Monsanto, USA). The transmission of the transgene to T1 progeny was demonstrated by PCR analysis and no pleiotropic effects were observed in these plants were phenotypically normal, fertile flowers and with abundant production of seeds. In entomological aspect of military caterpillar larvae were used in feeding bioassays with leaves of T0 events resulting from BRS 293. These same plants were also analyzed for expression of Cry1Ia protein via ELISA. A total of 48 plants were only T0 34 selected in both tests, the mortality rate of 89% and a high concentration of the toxic protein in the leaves. Dried flower buds of this plant were provided to larvae of the boll weevil to study immunodetection via optical microscopy, using midgut tissues of the insect. It was found that this protein in flower buds specifically bound to the antibody used to demonstrate that this event is very promising for further studies of improvement of resistance to two important pest of the cotton crop. / Várias instituições de ensino e pesquisa, em todo o mundo têm devotado esforços na descoberta de novas proteínas inseticidas e dos respectivos genes para o controle de pragas agrícolas. Trabalhos têm evidenciado o potencial de algumas moléculas no controle de insetos como as toxinas Bt do Bacillus thuringiensis, inibidores de proteases e de alfa-amilases e lectinas. Genes codificantes para essas proteínas têm sido estavelmente integrados ao genoma de plantas transgênicas conferindo-lhes resistência a pragas, como é o caso das culturas de tabaco, tomate, batata, milho e algodão Bt com resistência a lepidópteros que já são amplamente comercializadas em vários países. Vários estudos comprovam a economia e o benefício ecológico dessas cultivares no agronegócio internacional. Na lavoura algodoeira as pragas das ordens lepidóptera e coleóptera são os principais problemas econômicos em nível mundial. A Embrapa desenvolve pesquisas voltadas para o controle de pragas do algodoeiro com o intuito de reduzir os custos de produção, destes, 20 a 30% são para controle das pragas. Apesar dos importantes resultados obtidos até o momento o controle químico ainda tem sido o de melhor resposta embora mais oneroso no aspecto financeiro e mais agressivo no aspecto ambiental. O gene cry1Ia derivado da estirpe de Bacillus thuringiensis S1451 pode ser responsável por este tipo de controle. A atividade inseticida deste gene foi testada visando a análise da proteína recombinante para o bicudo do algodoeiro (Anthonomus grandis) e para a lagarta Spodoptera frugiperda. Indicando que este gene é bastante promissor para uso na área molecular visando obtenção de cultivar transgênica de algodão resistente a estas duas importantes pragas. Em um trabalho anterior esse gene foi inserido em três cultivares comerciais de algodão pela técnica de microinjeção via ovary drip. Nesse momento as cultivares receberam duas diferentes construções, compostas por um cassete linear mínimo mlc (≈ 3 Kb) e uma construção circular completa ccc (≈ 15 Kb), ambas contendo o gene cry1Ia. A integração do gene e expressão de proteína foram analisadas por PCR de DNA genômico, RT-PCR semi-quantitativo, Southern blot e ELISA. Mais de 1.800 transgenes foram testados e a integração do gene completo (1 cópia) foi detectada pelo ensaio de Southern blot e encontrada em apenas um único evento denominado T0 34 que foi derivado da cultivar BRS 293 no tratamento mlc, sugerindo uma tendência genótipo-dependente. A expressão da proteína Cry1Ia no evento T0 34, estimada por ELISA, foi semelhante a Bollgard comercial (Monsanto, EUA). A transmissão do transgene para progênies T1 foi demonstrada por análise de PCR e nenhum efeito pleiotrópico foi verificado nessas plantas que foram fenotipicamente normais, com flores férteis e produção de sementes abundantes. No aspecto entomológico larvas da lagarta militar foram utilizadas em bioensaios de alimentação com folhas dos eventos T0 resultantes da BRS 293. Essas mesmas plantas também foram analisadas quanto a expressão da proteína Cry1Ia, via ELISA. De um total de 48 plantas, apenas a T0 34 foi selecionada em ambos os ensaios, com taxa de mortalidade de 89% e elevada concentração da proteína toxica nas folhas. Botões florais desidratados dessa planta foram fornecidos a larvas do bicudo do algodoeiro para estudo de imunodetecção via microscopia ótica, utilizando-se tecidos do intestino médio do inseto. Verificou-se que a proteína presente nos botões florais se ligaram especificamente ao anticorpo utilizado demonstrando que esse evento é muito promissor para posteriores estudos de melhoramento visando resistência a duas importantes pragas da lavoura algodoeira.

A soja é uma cultura de grande importância, movimentando aproximadamente 230 bilhões de dólares em todo o mundo, com produção anual estimada de 267,61 milhões de toneladas. Os percevejos sugadores de sementes são considerados uma das pragas de maior importância para a cultura da soja, sendo as espécies Euschistus heros (E.), Piezodorus guildinii (West.) e Nezara viridula (L.) as mais abundantes no Brasil. O ataque por percevejos causa diversos problemas como o atraso da maturação fisiológica, retenção foliar, perdas no rendimento e diminuição da qualidade e potencial germinativo das sementes. Esses insetos são responsáveis ainda pela transmissão de patógenos, e podem causar alterações na composição de óleos, proteínas e ácidos graxos das sementes. A obtenção de cultivares com resistência genética é indispensável, a fim de minimizar a necessidade de utilização de defensivos agrícolas. Dessa forma, este trabalho identificou genes possivelmente associados à resposta de resistência pela comparação da expressão gênica diferencial entre genótipos de soja resistente (IAC-100) e suscetível (CD-215), a partir da metodologia de sequenciamento de RNA (RNA-Seq). As vagens foram submetidas a dois tratamentos por 24 horas: infestação por percevejos Piezodorus guildinii (West.) e não infestação. Foi utilizado o alinhador STAR contra o genoma hardmasked do phytozome e o pacote cufflinks.Com os resultados das análises dos genes diferencialmente expressos e uma posterior filtragem foi possível identificar 128 genes candidatos, dentre estes: genes relacionados com defesa, metabolismo secundário de produção de terpenos e proteínas LRR de sinalização celular. / Soybeans are an important crop with a world-wide annual production of 267.61 tons, worth 230 billion dollars. Stink bugs are a major soybean pest. The most abundant species in Brazil are Euschistus heros (E.), Piezodorus guildinii (West.) and Nezara viridula (L.). Stink-bug attack causes several problems, including delayed physiological maturity, leaf retention, yield losses, decreased seed quality, and decreased germination potential. The bugs are responsible for transmitting pathogens, and they can cause changes in the oil, protein, and fatty acids composition in the seed. Producing genetically resistant cultivars is critical in order to minimize the need for agricultural pesticides. Our work aims to identify genes associated with the stink-bug resistance response in soybean by comparing differential gene expression between resistant (IAC-100) and susceptible (CD-215) genotypes by analyzing RNA sequencing (RNA-Seq) data. Data was from soybean pods that were subjected to two treatments for 24 hours: infestation by the stink bug Piezodorus guildinii (West.), and non-infestation. STAR aligner was used against the hard-masked genome sequence from Phytozome, followed by analysis using the Cufflinks package. Analysis of the results of the differentially expressed genes and a subsequent filtering identified 128 candidate defense-associated genes, among them, defense-related such as LRR-RLKs and production of secondary metabolites like terpenes.

Maize is an important food crop in most parts of the world including the United States. The plants growing in the field are constantly challenged with various biotic stresses like insect herbivores and fungal pathogens. The physical wounds produced on the growing crops by the insects render the plants more vulnerable to the fungal pathogens. Hence developing both insect and fungal resistant maize varieties is crucial to benefit more from the harvest. Several studies have been in advance in this direction and as a consequence insect, in particular lepidopteran larve resistant maize genotype Mp708 and Aspergillus flavus resistant genotype Mp313E were developed. This study particularly focuses on understanding the functional involvement of the major phytohormones in the signal transduction and expression of the unique defense protein, Maize insect resistance 1-cysteine protease (Mir1-CP) shown to accumulate in response to herbivory by lepidopteran larvae, Spodoptera frugiperda (Fall armyworm, FAW) as a defense mechanism. Using a pharmacological approach involving exogenous hormone and hormone inhibitor treatments and analyzing the expression and accumulation of Mir1-CP protein and mir1 transcript by immunoblot and qRT-PCR analysis respectively, both JA and ET were found to be involved in mediating Mir1-CP accumulation with JA acting upstream of ET. Results also indicate that Mir1-CP accumulation involves both transcriptional and post-transcriptional (or post-translational) regulations. A different part of the study involved in understanding and evaluating the performance of Aspergillus flavus on the resistant and susceptible maize genotypes during infection. As of part of this study I also analyzed and compared the defense response offered by the resistant maize genotype, Mp313E and the susceptible genotype, Va35 by looking at the expression levels of the various defense related genes. The potency of the resistant maize genotype in sustaining the fungal infection in the field was of particular focus. Resistant maize genotype Mp313E was found to potentially oppose A.flavus proliferation and colonization and also delay aflatoxin biosynthesis unlike Va35. The up regulation of the maize defense genes during the early time points of infection, in Mp313E, indicate the potential role of these genes in conferring resistance against fungal pathogens.

Transgenic corn hybrids expressing Bt toxins are widely deployed to control pests such as Helicoverpa zea (Boddie). However, Bt resistance can reduce the efficacy of Bt corn hybrids. Volunteer corn expressing Bt toxins may impact Bt resistance management of pests. Surveys in Mississippi revealed the occurrence of fall season volunteer corn in high densities. Helicoverpa zea larvae were found feeding during both vegetative and reproductive stages. However, Bt parentage and low to moderate water and nitrogen stresses did not significantly influence H. zea larval growth and development on Bt positive volunteer plants. Additionally, pollen mediated Bt gene flow to non-Bt refuge ears from Bt hybrids significantly reduced H. zea larval growth on cross-pollinated refuge ears. The implications of these findings are that volunteer corn in most Mississippi corn production regions will have little impact on H. zea Bt resistance management, but resistance management could be compromised in more southern regions.

Canada thistle is an aggressive perennial weed throughout temperate areas for both the northern and southern hemisphere. As various single tactics have shown limited success in controlling this weed, I believe that a combination of different stress factors is necessary for effective sustainable control of Canada thistle. This study evaluates the competitive abilities of tall fescue grass and crownvetch in conjunction with a thistle feeding insect for Canada thistle control. The goal of the study was to determine what levels of the herbivore and plant competition are needed to suppress Canada thistle. Field studies were conducted to determine the effects of multiple stresses caused by defoliation and plant competition on growth and survival of Canada thistle (<i>Cirsium arvense</i> Scop.). Plant competition was from a cornbination of tall fescue (<i>Festuca arundinaceae</i> Schreb.) and crownvetch (<i>Coronilla varia</i> L). Artificial defoliation was used to determine the influence of level and frequency of defoliation in combination with plant competition on Canada thistle. The effects of various densities of a defoliator, <i>Cassida rubiginosa</i> (Coleop:Chrysonlelidae), in the presence and absence of plant competition were also determined. A three-season study determined the combined effects of <i>C. rubiginosa</i> and plant competitors at various densities on the growth and productivity of Canada thistle plantings of various ages. / Ph. D.

Thesis (PhD)--Stellenbosch University, 2014. / ENGLISH ABSTRACT: The Russian wheat aphid (Diuraphis noxia, Kurdj., Hemipetra, Aphididae, RWA) is an important pest of wheat, causing large-scale damage and yield losses. Various studies have been done at a transcriptomics level, including complementary DNA-amplified fragment length polymorphisms (cDNA-AFLPs), suppressive subtractive hybridization (SSH) and micro-array, which have identified genes putatively involved in RWA resistance. Even though these candidate genes have been identified, their role in host defence still needs to be verified using a functional genetics approach. In this study virus induced gene silencing (VIGS) using a barley stripe mosaic virus (BSMV) vector, has been utilized to knock-down candidate genes of interest in a wheat cultivar with the Dn1-resistance gene (TugelaDN). In this study it was hypothesized that genes involved in the hypersensitive response (HR) may contribute towards resistance and were thus targeted for silencing. These include glutathione-S-transferase (GST), superoxide dismutase Cu/Zn (SOD) and thylakoid-associated ascorbate peroxidase (tAPX). However, since aphid feeding also results in wounding, the genes were also analyzed under wounding only. Aphid fecundity is considered an indicator of involvement in RWA resistance, as susceptible plants result in higher aphid fertility. Findings in the study suggest that with wounding only, that Dn1 containing plants produce a greater hypersensitive response than susceptible controls. Ascorbate peroxidase was found to be important for wounding-induced resistance in Dn1 wheat plants. Under infestation conditions, silencing of superoxide dismutase Cu/Zn (SOD) and thylakoid-associated ascorbate peroxidase (tAPX) was found not to have an effect on aphid fertility and thus are not directly involved in resistance signaling. Knock-down of a phi-class glutathione-S-transferase F6 (TaGSTF6) transcripts however, had a large effect on aphid nymph numbers and thus may contribute to Dn1-resistance. Putative resistance genes silenced under aphid infestation conditions were a nucleotide binding protein (NBP) and resistance gene analogue 2 (RGA2). Analysis of NBP revealed its identity as a part of the iron homeostasis machinery in the cytosol, responsible for Fe-cluster assembly. Silencing of both NBP and RGA2 resulted in the expression of a susceptible phenotype. T10rga2-1A is an NBS-LRR protein known to be required for rust resistance in concert with resistance gene Lr10. T10rga2-1D silenced treatments resulted in susceptibility and plant death after aphid infestation, suggesting that T10rga2-1D may be a good up-stream candidate in Dn1-resistance. / AFRIKAANSE OPSOMMING: Die Russiese-koringluis (RWA) is ‘n pes wat ‘n belangrike ekonomiese invloed op koring opbrengste het en infestasie kan tot grootskaalse skade en oes verlies lei. Verskeie studies, onder andere komplimentêre DNA amplifiseerde fragment polimorfismes (cDNA-AFLPs), onderdrukkende onderskeidende hibridisaie (SSH) en mikro-reekse wat voorheen op transkriptomiese vlak gedoen is, het moontlike gene wat by RWA weerstand betrokke is, geïdentifiseer. Alhoewel hierdie gene reeds geidentifiseer was, hulle rol is nogtans onbekend. Dié gene moet nog getoets word, duur funksionele genetiese benaderingste maak. In hierdie studie is ‘n gars streep mosaïek virus vektor (BSMV) gebruik om kandidaat-gene van belang in ‘n Dn1-weerstandige geen-bevattende kultivar (TugelaDN) te onderdruk. Ondrukking van gene het deur middel van virus geïnduseerde geen onderdrukking (VIGS) plaasgevind. In hierdie studie is die hipotese gestel dat die gene betrokke by die hipersensitiewe reaksie (HR) ‘n invloed op plantweerstand kan hê en is dus geteiken vir geen-onderdrukking-studies. Hierdie gene het die volgende ingesluit: glutatioon-S-transferase (GST), superoksied dismutase Cu/Zn (SOD) en askorbien peroksidase (APX). Egter, omdat luisinfestasie ook tot verwonding aanleiding gee, is die onderdrukte gene ook onder alleenlik verwondingstoestande getoets. Luis vrugbaarheid is gebruik as indikator van betrokkenheid omdat meer vatbare plante ‘n hoër luis vrugbaarheid tot gevolg het. In die studie is gevind dat onder alleenlik verwondingkondisies, plante wat Dn1 bevat, ‘n groter hipersensitiewe respons vertoon, as vatbare kontroles. Daar is verder gevind dat askorbien peroksidase ‘n belangrike rol tydens verwondings-geïnduseerde weerstand in Dn1-plante speel. Daar is verder bevind dat die onderdrukking van superoksied dismutase Cu/Zn (SOD) en ‘n tilakoïed-geassosïeerde askorbien peroksidase (tAPX). Onder luis-infestasie kondisies, geen effek op luisvrugbaarheid gehad het nie en dus nie direk by die weerstandsrespons betrokke is nie. Die onderdrukking van ‘n phi-klas glutatioon-S-transferase F6 (TaGSTF6) het egter ‘n groot invloed op luis-vrugbaarheid gehad en kan dus ‘n rol in Dn1-weerstand speel. Die moontlike weerstands gene, geïdentifiseer as nukleotied bindings proteïen (NBP) en weestandsgeen anoloog 2 (T10rga2-1D), is getoets onder luis-infestasie kondisies. Die analise van NBP het getoon dat dit ‘n integrale deel van die yster homeostase meganisme in die sitosol, wat vir Fe-kluster samestelling verantwoordelik is, vorm. Onderdrukking van beide die NBP en T10rga2-1D het tot die uitdrukking van ‘n vatbare fenotipe aanleiding gegee. T10rga2-1A is ‘n NBS-LRR proteïen wat bekend is om noodsaaklik te wees tydens roes weerstandigheid in teenwoordigheid van die weerstandsgeen Lr10. T10rga2-1D-onderdrukte behandelings het tot vatbaarheid aangeiding gegee en daartoe gelei dat plante na luis-infestasies doodgaan. Hierdie resultate dui dus ‘n rol vir T10rga2-1D in Dn1-weerstandigheid aan, en suggereer verder dat hierdie geen ‘n goeie stroom-op kandidaat in Dn1-weerstandigheid is.

Orientador: Edson Luiz Lopes Baldin / Banca: Ivana Fernandes da Silva / Banca: André Luiz Lourenção / Banca: Pedro Takao Yamamoto / Banca: Bruno Henrique Sardinha de Souza / Resumo: A família Brassicaceae contém aproximadamente 3700 espécies, das quais apenas vinte são consumidas e cultivadas. A couve-de-folha [Brassica oleracea (L.) var. acephala] destaca-se entre as hortaliças como alimento importante para o consumo humano, possuindo níveis significativos de vitaminas A, C, K e ácido fólico, além de ser uma boa fonte de fibra. As brassicas também são conhecidas por terem elevado conteúdo de glucosinolatos, os quais são associados com a colonização de insetos-praga, além de reduzirem os riscos de ocorrecia de vários tipos de câncer em humanos. Um dos desafios para a cultura da couve-de-folha é a alta incidência de pragas em todas as fases da planta, sendo o pulgão Brevicoryne brassicae (L.) (Hemiptera: Aphididade) considerado espécie-chave para várias regiões. Esse pulgão é responsável por danos diretos e indiretos às plantas, prejudicando seu desenvolvimento e reduzindo seu valor comercial. A aplicação de inseticidas sintéticos é a principal tática de controle adotada pelos produtores para o manejo do afídeo, provocando impactos ambientais e colocando em risco a saúde do ser humano. O uso de resistência varietal no controle de pragas pode ser uma ferramenta valiosa para o Manejo Integrado de Pragas (MIP) e pode ser usado junto com outros métodos de controle. Neste estudo foi caracterizada a expressão de antibiose e/ou antixenose em 37 genótipos de couve-de-folha sobre B. brassicae por meio de bioensaios em casa de vegetação. Também foi avaliado o compo... (Resumo completo, clicar acesso eletrônico abaixo) / Doutor

A estratégia de pirâmide de genes tem sido explorada para retardar a evolução da resistência de insetos a plantas geneticamente modificadas que expressam proteínas inseticidas de Bacillus thuringiensis Berliner (Bt). No Brasil, às tecnologias de milho YieldGard VT PRO&trade; (VT PRO) e PowerCore&trade; (PW) que expressam as proteínas Cry1A.105/Cry2Ab2 e Cry1A.105/Cry2Ab2/Cry1F, respectivamente, foram liberadas para uso comercial em 2009. Para subsidiar programas de Manejo da Resistência de Insetos (MRI) foram conduzidos trabalhos para avaliar o risco de evolução da resistência de Spodoptera frugiperda (J. E. Smith) ao milho VT PRO e PW. Inicialmente foram realizados estudos para avaliar a atividade biológica de proteínas Bt expressas em diferentes estruturas da planta de milho VT PRO e PW sobre S. frugiperda e monitorar a suscetibilidade a Cry1A.105 e Cry2Ab2 em populações da praga coletadas em diferentes regiões geográficas do Brasil durante as safras de 2011 a 2014. Houve 100% de mortalidade de neonatas de S. frugiperda quando expostas ao tecido foliar de milho VT PRO e PW. No entanto, em estilo-estigmas e grãos, a mortalidade foi inferior a 50 e 6% respectivamente. Variabilidade geográfica na suscetibilidade de populações S. frugiperda a Cry1A.105 e Cry2Ab2 foi detectada, com reduções significativas na suscetibilidade a essas proteínas para algumas populações de 2011 a 2014. A técnica de \"F2 screen\" foi utilizada para a caracterização da resistência de S. frugiperda ao milho VT PRO e PW a partir de populações coletadas na safra de 2012. Verificou-se uma alta variabilidade na frequência fenotípica de isofamílias resistentes ao milho VT PRO e PW, sendo que as maiores frequências foram observadas em populações coletadas na região Central do Brasil. Com a técnica de \"F2 screen\" foi possível selecionar linhagens resistentes ao milho VT PRO e PW, denominadas de RR-2 e RR-3 respectivamente. Tanto a linhagem RR-2 quanto a RR-3 que foram criadas por 18 gerações consecutivas nos respectivos eventos de milho Bt apresentaram razões de resistência superiores a 3300, 2700 e &asymp; 10 vezes a Cry1A.105, Cry1F e Cry2Ab2, respectivamente. Cruzamentos recíprocos das linhagens RR-2 e RR-3 com uma linhagem suscetível de referência revelaram que o padrão da herança da resistência é autossômica recessiva. A recessividade genética da resistência também foi confirmada pela mortalidade completa de indivíduos heterozigotos (descendentes provenientes de cruzamentos entre as linhagens RR-2 ou RR-3 com a linhagem suscetível) em tecidos de milho VT PRO e PW, demonstrando que esses eventos atendem ao conceito de alta dose para o MRI. Em retrocruzamentos da progênie F1 dos cruzamentos recíprocos com as linhagens resistentes confirmou-se a hipótese de que a resistência é poligênica. A presença de custo adaptativo associado à resistência foi verificada para as linhagens RR-2 e RR-3, porém ausente para os indivíduos heterozigotos, baseado nos parâmetros biológicos avaliados. Neste estudo fornecemos a primeira evidência do potencial de evolução da resistência de S. frugiperda a eventos de milho Bt piramidados e informações para o refinamento das estratégias de MRI para preservar a vida útil das tecnologias de milho Bt para o controle de S. frugiperda no Brasil. / The strategy of pyramid of genes has been exploited to delay the evolution of insect resistance to genetically modified crops expressing insecticidal proteins from from Bacillus thuringiensis Berliner (Bt). In Brazil, YieldGard VT Pro&trade; (VT PRO) and PowerCore&trade; (PW) corn technologies expressing Cry1A.105/Cry2Ab2 and Cry1A.105/Cry2Ab2/Cry1F proteins respectively were released for commercial use in 2009. Resistance risk assessment were conducted to support an Insect Resistance Management (IRM) program of Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae) to VT PRO and PW corn. Initially, studies were conducted to evaluate the biological activity of Bt proteins expressed in different plant structures of VT PRO and PW corn on S. frugiperda and to monitor the susceptibility to Cry1A.105 and Cry2Ab2 in pest populations collected from different geographical regions in Brazil from 2011 to 2014 growing seasons. The mortality of neonate larvae of S. frugiperda was 100% when fed on leaf tissue of VT PRO and PW corn. However, the larval mortality when fed on silks and grains was less than 50 and 6% respectively. A geographical variation in the susceptibility of S. frugiperda to Cry1A.105 and Cry2Ab2 proteins was detected among populations, with significant reduction in susceptibility to these proteins in some populations from 2011 to 2014. The F2 screen technique was used to characterize the resistance of S. frugiperda to VT PRO and PW corn from populations sampled in 2012 growing season. High variability in the frequency of resistant phenotypic isofamilies to VT PRO and PW corn was obtained with higher frequencies in S. frugiperda populations from Midwestern region of Brazil. Resistant populations to VT PRO and PW corn were selected by using F2 screen which were designated as RR-2 and RR-3 strains respectively. Both RR-2 and RR-3 strains reared on respective Bt maize events for 18 consecutive generations showed resistance ratios greater than 3,300; 2,700 and &asymp; 10-fold to Cry1A.105, Cry1F and Cry2Ab2 respectively. Reciprocal crosses of RR-2 and RR-3 strains with a susceptible reference strain revealed that the inheritance of resistance is autosomal recessive. The genetic recessiveness of the resistance was also confirmed by the complete mortality of heterozygous individuals (offspring from the crosses between RR-2 or RR-3 strains with susceptible strain) on VT PRO and PW corn leaf tissues, indicating that these events meet the concept of high-dose for IRM strategies. Backcrosses of F1 progenies with both resistant strains revealed that resistance is polygenic. Fitness costs associated with resistance were found in RR-2 e RR-3 strains but not in heterozygous individuals, based on life history traits. In this study, we reported the first evidence of the potential of S. frugiperda to evolve resistance to pyramided Bt corn events, as well as provide valuable information to support the current IRM strategies to preserve the useful life of Bt corn technologies for S. frugiperda control in Brazil.

Programas de melhoramento genético são fundamentais para suprir à crescente demanda por maiores rendimentos. Contudo, existem alguns fatores restritivos à produtividade, como a incidência de insetos-praga e doenças. Dentre os insetos que atacam a cultura da soja, os percevejos são considerados pragas-chave, pois se alimentam diretamente da parte reprodutiva das plantas, danificando vagens e sementes. A seleção para resistência a percevejos pode levar a redução da produtividade, na ausência dessas pragas. Dessa forma o objetivo geral desse trabalho foi avaliar linhagens elite de soja em duas safras (2013/2014 e 2014/2015), sendo a primeira com controle dos percevejos e a outra sob ataque natural desses insetos. Na safra de 2013/2014 foram avaliadas 295 linhagens de soja, previamente selecionadas para resistência ao complexo de percevejos, e nove testemunhas (BMX Ativa, NA5909 RR, FTS Arapoty, FTS Campo Mourão, NA4823RR, IAC-100, V Max, CD-215 e BMX Potência) em dois ambientes. Os caracteres avaliados foram: número de dias para a maturidade (NDM); período de granação (PEG); valor agronômico (VA); acamamento (AC); produtividade de grãos (PG); peso de cem sementes (PCS). Foram calculados os componentes de variância e valor genotípicos pelo método REML/BLUP, herdabilidade e correlação genotípica. A seleção dos genótipos foi baseada no caráter PG, sendo considerado para seleção valores acima de 3000 kg.ha-1. A herdabilidade entre os caracteres avaliados variou de 0,084 (PEG) a 0,854 (PCS). Sete correlações apresentaram significância (P < 0,05) pelo Teste t, sendo que para PG apenas as correlações com VA e AC foram significativas. A seleção baseada na PG foi satisfatória para gerar ganho para todas as características avaliadas, sendo que alguns genótipos tiveram valor genotípico favorável ou contrário ao ganho de seleção estimado. Na safra de 2014/2015, as mesmas linhagens e testemunhas foram avaliadas sob infestação natural de percevejos. Os caracteres avaliados foram número de dias para a maturidade (NDM); altura da planta na maturidade (APM); acamamento (AC); valor agronômico (VA); produtividade de grãos (PG); período de granação (PEG); peso de cem sementes (PCS); retenção foliar (RF); e peso de sementes boas (PSB). As análises foram semelhantes à da safra anterior, com exceção que seleção foi baseada na característica PSB, visando a obtenção de genótipos superiores, considerando resistência ao complexo de percevejos e alto rendimento. A população de percevejos foi avaliada pelo método pano de batida, sendo que houve aumento dos mesmo durante o período de granação, chegando a um nível superior ao patamar para danos econômicos. A variância genotípica teve maior magnitude que a variância residual para VA, PSB e PCS, indicando alta variabilidade genética para essas características. As herdabilidades observadas foram de intermediária a alta, variando de 0,553 (APM) a 0,838 (PSC). A maioria das correlações foram significativas pelo teste de t (P < 0,05). A maior correlação positiva encontrada foi entre PSB e PG (0,765), enquanto que a negativa foi entre VA e RF (- 0,835). PSB teve correlações significativas com todas as outras características e foi eficiente para selecionar genótipos com alto rendimento, boas características agronômicas e resistentes ao complexo de percevejos. / Breeding programs are essential to meet the growing demand for high yield crops. However, there are some factors that can preclude production increase such as pests and diseases incidence. Among these pests, stinkbugs are considered a major issue and may hamper soybean farming due its feeding behavior, spoiling the reproductive parts of the plants (e.g. pods and seeds). Selection for resistance to stink bugs may reduce productivity when these insects are absent. Thus the aim of this study was to evaluate soybean lines in two seasons (2013/2014 and 2014/2015), the first controlling the stinkbug\'s population and the other under natural attack of these insects. In the 2013/2014 season 295 soybean lines, previously selected for resistance to stinkbugs complex and nine commercial cultivars (BMX Ativa, NA5909 RR, FTS Arapoty, FTS Campo Mourão, NA4823RR, IAC-100, V Max, CD-215 e BMX Potência) were evaluated in both environments. The evaluated traits were: number of days to maturity (NDM); graining period (PEG); agronomic value (VA); lodging (AC); grain yield (PG); one hundred seeds weight (PCS). The variance components and genotypic value were estimated by REML / BLUP method, heritability and genetic correlations were calculated. The selection of genotypes was based on PG, being considered for selection values above 3000 kg ha-1. The heritability of the traits ranged from 0.084 (PEG) to 0.854 (PCS). Seven correlations were significant (P <0.05) by t test, and for PG only correlations with VA and AC were significant. Selection based on PG was satisfactory to generate gains for all traits evaluated, and some genotypes had genotypic values favorable or contrary to gain selection. In the 2014/2015 season, the same lines and commercial cultivars were evaluated under natural infestation of stinkbugs. The characters evaluated were number of days to maturity (NDM); plant height at maturity (APM); lodging (AC); agronomic value (VA); grain yield (PG); graining period (PEG); one hundred seeds weight (PCS); leaf retention (RF); and good seed weight (PSB). The analyzes were similar to the previous season, except that selection was based on the PSB in order to obtain superior genotypes, considering resistance to complex bugs and high yield. The population of stinkbugs was controlled by beating cloth method, and the population increased during the graining period, reaching a level higher than the level for economic losses. The genotypic variance was higher than the residual variance for VA, PSB and PCS, suggesting high genetic variability for these traits. The heritability observed were intermediate to high, ranging from 0.553 (APM) to 0.838 (PSC). Most of the correlations were significant by t test (P <0.05). The highest positive correlation was found between PSB and PG (0.765), while the negative was between VA and RF (-0.835). PSB had significant correlations with all other features and was efficient to select genotypes with high yield, good agronomic characteristics and better resistance to the stinkbugs incidence.