The capsid protein of Helicoverpa armigera stunt virus (HaSV) a T=4 insect virus was expressed in Spodoptera frugiperda 9 cells using a baculovirus vector. When the insect cells were infected at a high MOl the expressed coat protein assembled into virus-like particles (VLPs) that spontaneously underwent maturation and were morphologically indistinguishable from wild-type HaSV. The VLPs were electron dense when viewed under EM and encapsidated their coat protein mRNA. When Sf9 cells were infected at a low multiplicity of infection (MOl) the expressed capsid protein assembled into procapsids that did not spontaneously undergo maturation. These procapsids underwent autoproteolytic maturation cleavage when they were treated with an acidic buffer. The procapsids were used in the encapsidation of a FITC labelled peptide. The peptide encapsidating VLPs showed an increase in their buoyant density that was not collaborated by an increase in the concentration of the FITC labelled peptide detected when these samples were compared to control samples with similar buoyant densities.

Insecticides in the diamide class have a novel mode of action and have become a key component for management of agriculturally important lepidopteran pests since their introduction in 2008. Corn earworm, Helicoverpa zea (Boddie); and the armyworm complex including fall armyworm, Spodoptera frugiperda (J.E. Smith); and Spodoptera exigua (Hübner); are significant pests of agroecosystems in the Mid-southern and Southeastern regions of the United States. They have developed resistance to, and/or inconsistent control has occurred with most chemical classes. The objectives of this study were to establish susceptibility levels of field populations of H. zea, S. frugiperda, and S. exigua collected in the Mid-southern and Southeastern regions of the United States to flubendiamide and chlorantraniliprole. To achieve equivalent levels of mortality for each species, a higher concentration of flubendiamide was required compared to chlorantraniliprole. Furthermore, two experiments were conducted to determine the systemic and residual efficacy of chlorantraniliprole and flubendiamide against H. zea on vegetative and reproductive structures of soybean. Chlorantraniliprole moved systemically and had significantly greater control than flubendiamide in the systemic and residual study out to 31 DAT. Flubendiamide did not move systemically but provided significant residual control out to 31 DAT compared with the untreated control. Neither insecticide was detected in reproductive structures. Finally, to determine the risk of resistance development, a S. exigua colony, originating from a field collection in 2013, was separated into three cohorts that were independently selected with three concentrations (0.016, 0.020, and 0.025 ppm) of flubendiamide incorporated into a meridic diet. These concentrations were chosen from the LC30, LC60 and LC90 of the original colony. Resistance ratios never increased past 2.11old. The highest resistance ratios occurred after 18 generations for the LC30 colony, 19 generations for the LC60 colony, and 13 and 15 generations for the LC90 colony. After reaching their highest point of resistance, the colonies began to decline in egg production and larval survivability and did not recover. After 22 generations the selected colonies were terminated. The results from this portion of the study suggest that the potential for resistance development of beet armyworm to flubendiamide is unclear.

<p>Dhurrin is a cyanogenic glucoside (CG), an important compound that can interplay with primary and secondary metabolism in sorghum. Dhurrin metabolism contributes to insect resistance, growth, nitrogen (N) metabolism, drought tolerance, and safety for animal consumption when used as a forage. Through chemical mutagenesis with ethyl methanesulfonate (EMS), a mutation in the gene encoding CYP79A1 (<i>cyp79a1</i>), the first enzyme in the biosynthetic pathway of dhurrin, was discovered that inhibits the production of dhurrin. The acyanogenic phenotype of this mutant could be a major benefit in reducing the risk of hydrogen cyanide (HCN) toxicity within animals; however, understanding the effects of inhibiting dhurrin biosynthesis is important in understanding metabolic tradeoffs that could occur. This dissertation describes research to assess impacts and tradeoffs of the dhurrin-free trait on susceptibility to Fall Armyworm [<i>Spodoptera frugiperda</i> (J.E. Smith)] (FAW) feeding, seedling growth, effects on post-flowering drought tolerance, cold stress and utilization as a forage. Insect susceptibility and seedling growth were examined using near-isogenic lines (NILs) within the greenhouse utilizing non-destructive phenotyping technologies for green plant area and in the field comparing total leaf area and dry weight. Post-flowering drought stress was induced within a greenhouse, growth chamber, and field environments. The <i>cyp79a1 </i>mutation was tested in NILs, a near-isogenic backcross (NIBC) population, and near-isogenic hybrids (NIH), to understand the impacts of the <i>cyp79a1 </i>mutation<i> </i>on the stay-green trait. Palatability as forage was examined by comparing the feeding preference of ruminant animals with multiple conventional hybrids and an experimental hybrid carrying the <i>cyp79a1</i> mutation. This preference was also examined using a set of NILs varying in the <i>cyp79a1 </i>mutation<i>. </i> Safety was assessed in preference trials by testing for HCN release before grazing. To further our understanding of the benefits of sorghum as a forage, the dhurrin-free experimental hybrid was compared to seven conventional hybrids as a dry product. The dry sorghum product was tested for the release of HCN and dhurrin content. Lastly, the effects of low temperatures and frost were assessed for their effects on the production of dhurrin in cyanogenic and dhurrin-free sorghum genotypes. </p> <p>Overall, the biosynthesis of dhurrin had a significant effect on the deterrence of FAW and on the growth of sorghum seedlings. Dhurrin-free lines were more susceptible to FAW feeding but also exhibited a significantly higher growth rate. Dhurrin-free lines and hybrids only exhibited a slight increase in susceptibility to post-flowering drought stresses with only one dhurrin-free hybrid discovered to senesce faster than its wild-type NIH. Comparisons of the effects of dhurrin biosynthesis on stay-green in a NIBC population in Tx642 (B35), one of the most important sources of the stay-green trait, did not show any variation in chlorophyll concentration (CC) and normalized difference vegetation index (NDVI). Analyses of the impact of dhurrin on palatability as a forage showed that ewes preferred grazing on the dhurrin-free hybrids and NILs, showing that the ewes were able to detect the presence or absence of dhurrin while feeding. Experiments to assess the safety and stability of dhurrin in dried plant material demonstrated that dhurrin content did not change during drying and HCN was released after rehydration. Furthermore, high levels of HCN were immediately released when rumen fluid was added to dried plant materials containing dhurrin; however, no detectable HCN was released from dhurrin-free genotypes. Finally, sorghum plants exposed to freezing temperatures exhibited an increase in dhurrin content in conventional sorghum hybrids while no detectable dhurrin was noted within <i>cyp79a1 </i>mutants. </p> <p>Taken together, these studies demonstrate pleiotropic effects for the <i>cyp79a1 </i>mutation. Dhurrin-free genotypes were more susceptible to insect herbivory and may be slightly more susceptible to post-flowering drought within the hybrids; however, these genotypes exhibited higher seedling growth rates, feeding preference by ewes, no release of HCN in fresh or dry plant material, and frost did not cause an increase in dhurrin content.</p>

Maize (Zea mays) is an important agricultural crop frequently targeted by pests that pose a threat to plant development and survival. To deal with this problem, maize generates a wide variety of responses to attack by pests, from activation of woundresponse pathways to the release of volatile compounds. Several maize lines have been developed that show resistance to one common pest, the larvae of the fall armyworm (Spodoptera frugiperda). Analysis of the volatiles released by the resistant and susceptible lines in the presence and absence of the fall armyworm was conducted using SPME coupled to GC/MS. Caryophyllene, a commonly released plant volatile, was identified in the resistant line. In the susceptible line, caryophyllene was detected in smaller quantities or not at all. The results of a preference study demonstrated that fall armyworm larvae show a statistically significant preference for yellow-green whorl tissue from the susceptible over the resistant line.

As principais táticas de controle de Spodoptera frugiperda (Lepidoptera: Noctuidae) têm sido o uso de inseticidas e cultivos Bt no Brasil. Devido à intensa pressão de seleção, a evolução da resistência de S. frugiperda aos principais inseticidas de diferentes grupos químicos e a proteínas Bt já foram documentados no Brasil. Sendo assim, estudos para preservar a vida útil de inseticidas com novos mecanismos de ação como a de chlorfenapyr (desacoplador da fosforilação oxidativa mediante a disrupção do gradiente de prótons) são fundamentais em programas de Manejo da Resistência de Insetos (MRI). Para a implementação de programas proativos de manejo da resistência de S. frugiperda a chlorfenapyr, foram conduzidos estudos para caracterizar e monitorar a suscetibilidade de populações de campo de S. frugiperda a chlorfenapyr, estimar a frequência do alelo da resistência a chlorfenapyr pelo método de F2 screen e avaliar a resistência cruzada entre chlorfenapyr e outros inseticidas e proteínas Bt. O método de bioensaio utilizado foi o de tratamento superficial da dieta. Os dados das linhas-básicas de suscetibilidade das populações de campo a chlorfenapyr demonstraram uma variação da CL50 de 13,87 a 25,07 &#181;g/mL. A concentração diagnóstica de 56 &#181;g/mL foi estimada mediante análise conjunta dos dados de linha-básica de suscetibilidade baseada na CL99. A suscetibilidade foi monitorada a partir de populações de campo de S. frugiperda coletadas nas principais regiões produtoras de milho no Brasil entre 2016 e 2018. A taxa de sobrevivência na concentração diagnóstica variou de 0 a 8,4%. Pelo método de F2 screen a frequência alélica estimada nas safras para a 2ª safra 2016 e a 1ª safra 2017 foi de 0,0008 e a frequência para a entressafra 2016-2017 foi de 0,0012. Não foi verificada resistência cruzada entre chlorfenapyr e os inseticidas: lambda-cyhalothrin, chlorpyrifos, lufenuron, teflubenzuron, spinosad e chlorantraniliprole e as proteínas Bt: Cry1F, Cry1A.105/Cry2Ab2, Cry1A.105/Cry2Ab2/Cry1F e Vip3Aa20 expressas em milho geneticamente modificado. Os resultados demonstraram uma alta suscetibilidade das populações de campo a chlorfenapyr, uma baixa frequência do alelo da resistência e ausência de resistência cruzada aos principais inseticidas e proteínas Bt. Portanto, o inseticida chlorfenapyr pode ser utilizado como uma ferramenta em programas de MRI em S. frugiperda no Brasil. / Chemical control and Bt crops are the major control tactics to control Spodoptera frugiperda (Lepidoptera: Noctuidae) in Brazil. Due to intense selection pressure, the evolution of S. frugiperda resistance to major insecticides from different chemical groups and to Bt proteins has already been documented in Brazil. Therefore, studies to preserve the lifetime of insecticides with new mode of action such as chlorfenapyr (uncouplers of oxidative phosphorylation via disruption of the proton gradient) are important in Insect Resistance Management (IRM) programs. For implementing a proactive resistance management program of S. frugiperda to chlorfenapyr, studies were conducted to characterize and monitor the susceptibility of field populations of S. frugiperda to chlorfenapyr, to estimate the frequency of resistance alleles to chlorfenapyr using F2 screen method and to evaluate the cross-resistance patterns of chlorfenapyr and other insecticides and Bt proteins. Diet overlay bioassays was used to characterize the baseline susceptibility data of field population of S. frugiperda to chlorfenapyr. The LC50 ranged from 13.87 a 25.07 &#181;g/mL. A diagnostic concentration of 56 &#181;g/mL was estimated from the joint analysis of the baseline susceptibility data, based on LC99. Susceptibility was monitored from field populations of S. frugiperda collected from major corn growing regions in Brazil from 2016 to 2018. The survival rate at the diagnostic concentration ranged from 0 to 8.4%. Using the F2 screen method, the estimated allele frequency for the 2nd crop season 2016 and 1st crop season 2017 was 0.0008 and the frequency for the offseason 2016-2017 was 0.0012. No cross-resistance was verified between chlorfenapyr and the insecticides: lambda-cyhalothrin, chlorpyrifos, lufenuron, teflubenzuron, spinosad and chlorantraniliprole and to Bt proteins: Cry1F, Cry1A.105/Cry2Ab2, Cry1A.105/Cry2Ab2/Cry1F and Vip3Aa20 expressed in genetically modified maize. The results demonstrated a high susceptibility of field populations of S. frugiperda to chlorfenapyr, a low resistance allele frequency and lack of cross-resistance to major insecticides and Bt proteins. Therefore, chlorfenapyr can be used as an important tool in IRM programs of S. frugiperda in Brazil.

A utilização de plantas transgênicas expressando proteínas inseticidas de Bacillus thuringiensis (Bt) tem sido a principal estratégia para o controle da lagarta-do-cartucho Spodoptera frugiperda (J.E. Smith) no Brasil. A resistência de S. frugiperda à proteína Vip3Aa20 foi recentemente isolada e caracterizada em condições de laboratório, a partir de uma população proveniente de Correntina-BA. Para subsidiar programas de Manejo da Resistência de Insetos (MRI), foram realizados estudos para entender a distribuição geográfica da resistência de S. frugiperda à proteína Vip3Aa20 no Brasil, mediante uso de métodos fenotípicos (proteína purificada e folhas de milho Bt) e genotípicos (F1 e F2 screen), além da elucidação da presença custo adaptativo associado à resistência a partir de uma linhagem resistente quase-isogênica. Para o monitoramento com proteína purificada, as médias de sobrevivência na dose discriminatória de 3600 ng Vip3Aa20/cm2 para 10 populações/safra de S. frugiperda das principais regiões produtoras de milho do Brasil, tiveram um aumento em número, no decorrer das safras, para as populações que diferiram da sobrevivência da linhagem SUS, por não sobreposição do I.C. (95%). As larvas sobreviventes deste monitoramento morreram quando transferidas para folhas de milho Bt expressando Vip3Aa20. Não houve sobreviventes no monitoramento da resistência utilizando folhas de milho Bt, para todas as populações avaliadas. As médias de frequência alélica da resistência estimadas pelo método F1 screen foram de 0,0069, 0,0051 e 0,0041 para as populações avaliadas na 2ª safra/2016, entressafra/2016 e 1ª safra/2017 respectivamente. Pelo método F2 screen, as médias de frequência alélica foram de 0,0030, 0,0036, 0,0054 e 0,0042 para as populações da 2ª safra/2016, entressafra/2016, 1ª safra/2017 e 2ª safra/2017 respetivamente. Foram selecionadas 3 linhagens resistentes a Vip3Aa20 pelo método F2 screen, a partir de populações provenientes de Peabiru-PR, São João da Boa Vista-SP e Casa Branca-SP. Pelo teste de complementaridade, o mesmo alelo da resistência que havia sido inicialmente isolado da região de Correntina-BA, foi verificado nestas novas linhagens resistentes selecionadas. Não foi verificada a presença de custo adaptativo associado à resistência de S. frugiperda a Vip3Aa20, além de um maior valor adaptativo de indivíduos heterozigotos mediante avaliação de parâmetros de tabela de vida e fertilidade. Os resultados do presente trabalho comprovaram que o alelo da resistência de S. frugiperda para a proteína Vip3Aa20 está amplamente distribuído nas principais regiões produtoras de milho no Brasil. Este fato aliado à ausência de custo adaptativo da resistência reforçam a necessidade de implementação de estratégias eficientes de MRI para retardar a evolução da resistência de S. frugiperda à Vip3Aa20 no Brasil. / The use of transgenic plants expressing Bacillus thuringiensis (Bt) insecticidal proteins has been the main strategy to control the fall armyworm Spodoptera frugiperda (J.E. Smith) in Brazil. The resistance of S. frugiperda to Vip3Aa20 protein was recently isolated and characterized in laboratory conditions from a Correntina-BA population. In order to subsidize Insect Resistance Management (IRM) programs, studies were carried out to understand the geographical distribution of S. frugiperda resistance to Vip3Aa20 protein in Brazil, through the use of phenotypic (purified protein and Bt maize leaves) and genotypic methods (F1 and F2 screen), in addition, the elucidation of fitness-cost presence associated with resistance from a near-isogenic resistant strain. For the purified protein monitoring, the mean survival at the diagnostic concentration of 3,600 ng Vip3Aa20/cm2 for 10 populations/season of S. frugiperda from the main maize producing regions of Brazil, had an increase in the number of populations with significant difference by non-overlap in the confidence interval (95 %) in relation with our reference of susceptibility. The surviving larvae of this monitoring died when transferred to leaves of Bt maize expressing Vip3Aa20. There were no survivors in resistance monitoring using Bt maize leaves for all populations evaluated. The mean frequency of resistance alleles estimated by the F1 screen method were 0.0069, 0.0051 and 0.0041 for the populations evaluated in the 2nd/2,016, the offseason/2,016 and the 1st/2,017 crop seasons, respectively. By the F2 screen method, the means of allele frequency were 0.0030, 0.0036, 0.0054 and 0.0042 for the populations of the 2nd/2,016, offseason/2,016, 1st/2,017 and 2nd/2,017 crop seasons, respectively. Three Vip3Aa20 resistant strains were selected by the F2 screen method from populations from Peabiru-PR, São João da Boa Vista-SP and Casa Branca-SP. By the complementarity test, the same resistance allele that had been initially isolated from the Correntina-BA region was verified in these new selected resistant strains. The presence of the adaptive cost associated to the resistance of S. frugiperda to Vip3Aa20 was not verified, besides a higher adaptive value of heterozygous individuals through evaluation of life table parameters and fertility. The results of the present work proved that the resistance allele of S. frugiperda for the Vip3Aa20 protein is widely distributed in the main maize producing regions in Brazil. This fact, coupled with the lack of adaptive cost of resistance, reinforces the need to implement efficient IRM strategies to delay the evolution of S. frugiperda resistance to Vip3Aa20 protein in Brazil.

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.

Visando explorar o potencial bioativo de plantas da família Solanaceae como fonte alternativa para o manejo de Spodoptera frugiperda (Lepidoptera: Noctuidae), a lagarta-do-cartucho-do-milho, realizou-se, primeiramente, uma triagem de extratos etanólicos preparados de diferentes partes de 25 espécies vegetais. Após sete dias de exposição das lagartas em dietas tratadas, constatou-se que os extratos etanólicos de folhas de Acnistus arborscens e de Datura stramonium, na concentração de 4.000 mg kg -1, foram os mais promissores causando inibição do ganho de peso de 97,85 e 95,72%, respectivamente. A seguir, foi realizado um fracionamento biomonitorado, a partir do qual, a fração diclorometânica foi selecionada como a mais ativa para essas duas espécies, causando efeito subletal (inibição do ganho do peso) e letal (mortalidade), porém este inferior ao causado pela formulação comercial Azamax&reg; 1,2 EC (controle positivo). Em sequencia, foi possível estimar as concentrações letais para as frações diclorometânicas durante o período de 168 horas. Em relação ao efeito sobre o desenvolvimento do inseto, os extratos etanólicos foram testados na concentração de 4.000 mg kg -1 e as frações diclorometânicas na concentração letal mediana (4.088 e 3.694 mg kg -1 para D. stramonium e A. arborescens, respectivamente). Na fase larval, os extratos etanólicos de D. stramonium e A. arborescens ocasionaram 77,50 e 62,50% de mortalidade, respectivamente, e, além disso, ambos os tratamentos causaram alongamento dessa fase, e mortalidade, deformidade e redução do peso pupal. A fração diclorometânica de A. arborescens proporcionou 100% de mortalidade das lagartas expostas, enquanto a mortalidade em D. stramonium foi 83,19% e, as lagartas necessitaram de 41,75 dias para atingir a fase pupa, que apresentou baixo peso (158,60 mg) e altas deformidades (40%) em comparação ao controle (255,90 mg e 8,26%, respectivamente). Além disso, as frações diclorometânicas ocasionaram atividade fagodeterrente nas CL25, CL50 e CL90. Novamente, utilizando-se a fração diclorometânica, foi realizado novo fracionamento, e após sete dias de exposição das lagartas neonatas, todas as subfrações ocasionaram efeitos subletais. Com isso, tornaram-se necessárias novas investigações químicas a fim de se compreender o (s) composto (os) responsável (is) pela bioatividade, porém a complexidade de algumas subfrações não permitiu uma caracterização química. Ainda assim, foi constatada nas subfrações acetato de etila e metanólica de D. stramonium a presença de compostos pertencentes à classe dos vitanolidos. Dessa forma, os derivados de Solanaceae apresentam potencial bioativo passíveis de serem estudados mais profundamente visando ao emprego dos mesmos no manejo de S. frugiperda. / Aiming to explore the bioactive potential of Solanaceae plants as alternative resources for the management of Spodoptera frugiperda (Lepitoptera: Noctuidae), the fall armyworm, it was performed, firstly, a screening of ethanolic extracts prepared from different parts of 25 plant species. After seven days of exposure of caterpillars to treated diets, it was verified that ethanolic extracts of Acnistus arborscens and Datura stramonium leaves in the concentration of 4,000 mg kg-1 were the most promising, causing an inhibition of 97.85% and 95.72%, respectively, in weight gain. After this, a bioguided fractionation was performed, from which the dichloromethane fraction was selected as the most active in both species, causing sublethal (inhibition of weight gain) and lethal (mortality) effects, although both were less effective than the Azamax&reg; 1,2 EC commercial formulation (positive control). Following the previous step, it was possible to estimate the lethal concentrations for dichloromethane fraction during the period of 168 hours. In relation to the effect on the insect development, the ethanolic extracts were tested in the concentration of 4,000 mg kg-1 and the dichloromethane fractions in their median lethal concentrations (4,088 and 3,694 mg kg-1 for D. stramonium and A. arborescens, respectively). In the larval phase, ethanolic extracts of D. stramonium and A. arborescens lead to 77.50% and 62.50% of mortality, respectively, and both treatments lead to the elongation of this phase, mortality, deformity and pupal weight reduction. The dichloromethane fraction of A. arborescens lead to 100% of mortality of the exposed caterpillars, while mortality of D. stramonium was 83.19%, and the caterpillars needed 41.75 days to reach the pupal stage, which presented low weight (158.60 mg) and were highly deformed (40%) in comparison with the negative control (255.90 mg and 8.26%, respectively). Furthermore, the dichloromethane lead to antifeedant activity in CL25, CL50 and CL90. Once again, using the dichloromethane fraction, another fractionation was performed, and after seven days of exposure of neonates caterpillars, all subfractions lead to sublethal effects. Therefore, new chemical evaluations are necessary to understand the compounds responsible for bioactivity, however, the complexity of some subfractions did not permit chemical characterization. Nevertheless, it was verified in the ethyl acetate and methanolic subfractions of D. stramonium the presence of withanolides compounds. Therefore, derivatives of Solanaceae present bioactive potential that may be liable for further evaluation aiming their employment in the management of S. frugiperda.

Lufenuron é um novo inseticida do grupo dos inibidores da biossíntese de quitina utilizado para o controle de Spodoptera frugiperda J. E. Smith (Lepidoptera: Noctuidae) na cultura do milho. O uso deste inseticida para o controle de S. frugiperda tem aumentado muito nos últimos anos. Este incremento pode ser atribuído à sua alta eficiência e devido ao desenvolvimento da resistência desta praga aos produtos tradicionalmente recomendados para o seu controle (por exemplo, fosforados e piretróides). Neste trabalho, a linha básica de suscetibilidade de S. frugiperda foi obtida com o intuito de implementar um programa proativo de manejo da resistência. Para a caracterização da suscetibilidade de populações de S. frugiperda ao lufenuron, utilizou-se o bioensaio que consistiu no tratamento superficial de dieta artificial. Lagartas de 2° e 3° instar foram expostas à dieta tratada por um período de até 5 dias (120 h). A partir deste estudo concluiu-se que a utilização de lagartas de 3° instar expostas por um período de 4 dias (96 h) foi o procedimento de bioensaio mais adequado. Baseado na curva de concentração-resposta da população suscetível de referência, as concentrações de 3,2 e 10,0 mg de lufenuron / mL de água destilada foram escolhidas para o monitoramento da suscetibilidade. Resultados do monitoramento da suscetibilidade de populações de S. frugiperda provenientes dos principais Estados produtores de milho do Brasil (Rio Grande do Sul, Paraná, São Paulo, Goiás e Minas Gerais) mostraram diferenças significativas quanto à suscetibilidade ao lufenuron. Esses resultados sugerem a necessidade da implementação de estratégias de manejo da resistência para preservar a vida útil do lufenuron no controle de S. frugiperda no Brasil. / Lufenuron is a newly benzoylphenyl urea chitin synthesis inhibitor insecticide. The use of this compound to control Spodoptera frugiperda J. E. Smith (Lepidoptera: Noctuidae) has increased significantly in the past years. This increase can be attributed to its high activity against S. frugiperda and to the reduction of efficacy due to resistance to some traditional insecticides (such as phosphates, carbamates and pyrethroids) recommended for controlling this pest. In this study, a baseline susceptibility data of S. frugiperda populations to lufenuron were obtained for implementing a proactive resistance management program. An artificial diet treatment surface bioassay was used to characterize the response to lufenuron in a susceptible strain. Second and third instar larvae were exposed to the treated diet up to 5 days (120 h). A period of 4 days (96 h) and third instar larvae were defined as standard procedures for bioassays. After the characterization of the susceptible population, diagnostic concentrations of 3.2 and 10.0 mg of lufenuron / mL of distilled water were chosen for monitoring the susceptibility. A survey of susceptibility to lufenuron was conducted in field populations of S. frugiperda collected from major corn-growing regio ns located in the States of Rio Grande do Sul, Paraná, São Paulo, Goiás and Minas Gerais. Significant differences in susceptibility to lufenuron were detected among S. frugiperda populations. These results suggest the need to implement resistance management strategies to preserve the lifetime of lufenuron for controlling S. frugiperda in Brazil.

Visando detectar alternativas de manejo para a lagarta-do-cartucho-do-milho, Spodoptera frugiperda (Lepidoptera: Noctuidae), realizou-se, primeiramente, uma triagem em extratos etanólicos preparados das estruturas (folhas, ramos e sementes) de diferentes espécies de Annonaceae (Annona cacans, A. montana, A. mucosa, A. reticulata, A. sylvatica e Duguetia lanceolata). Com base nessa etapa inicial, constatou-se que o extrato etanólico das sementes de A. mucosa (ESAM) foi o mais promissor, causando significativa toxicidade aguda (CL50 e CL90 de 842,97 e 1.882,00 mg kg-1, respectivamente) e pronunciada inibição do desenvolvimento larval (toxicidade crônica), após sete dias de exposição a dietas tratadas. Na CL50, o ESAM reduziu as viabilidades larval e pupal e o peso de pupas com 24 horas, e aumentou a duração da fase larval. No entanto, o ESAM não ocasionou efeito fagodeterrente para larvas de quarto ínstar de S. frugiperda, embora tenha reduzido o consumo ao longo do tempo. Feito isso, a eficácia de ESAM foi comparada com inseticidas comerciais de origem natural (Azamax® 1,2 EC) e sintética (Premio® SC). Nessa comparação, o ESAM (na CL90) apresentou eficácia similar (equitóxico) aos dois produtos comerciais (utilizados na dose registrada para o controle do inseto-praga). A seguir, foi realizado o fracionamento biomonitorado através de diferentes técnicas cromatográficas que conduziu ao isolamento da acetogenina roliniastatina-1, identificada (com base em técnicas espectroscópicas) como componente majoritário da fração mais ativa do extrato etanólico de A. mucosa. Roliniastatina-1 foi então novamente ensaiada frente a larvas de S. frugiperda e ocasionou significativos efeitos agudos (mortalidade larval) e crônicos (redução do desevolvimento larval). Por fim, investigou-se a bioatividade de uma formulação à base de acetogeninas (Anosom® 1 EC) recentemente registrada, tanto isoladamente quanto em mistura com uma formulação à base de limonoides (Azamax® 1,2 EC). Anosom® 1 EC, testado na CL90 estimada (2.959,00 mg kg-1) e Azamax® 1,2 EC, testado na concentração registrada para o controle do inseto-praga (4.000,00 mg kg-1), causaram significativa mortalidade larval, sem ocorrer diferença entre os tratamentos tanto quando testados isoladamente como em mistura binária. Anosom® 1 EC (na CL50) provocou ainda aumento significativo das mortalidades larval e pupal e da duração da fase larval, bem como redução do peso pupal do inseto. Dessa forma, derivados de Annonaceae podem constituir um componente útil para o manejo integrado de S. frugiperda em condições de campo. / A screening with ethanolic extracts prepared from structures (leaves, branches and seeds) from different species of Annonaceae (Annona cacans, A. montana, A. mucosa, A. reticulate, A. sylvatica and Duguetia lanceolata) aiming to detect management alternatives for Spodoptera frugiperda (Lepidoptera: Noctuidae) was done. Based on this initial phase, it was verified that the ethanolic extract from seeds of A. mucosa (AMSE) was the most promising one, causing significant acute toxicity (LC50 and LC90 of 842.97 and 1,822.00 mg kg-1, respectively) and pronounced inhibition of larval development (chronic toxicity) after a 7-day exposition to treated diets. At LC50, AMSE decreased larval and pupal viabilities and also the pupal weight at 24 hours, and increased the larval phase duration. However, AMSE didn\'t cause antifeedant effect for 4-instar S. frugiperda larvae, though it decreased consumption throughout the time. After that, AMSE efficacy was compared to commercial pesticides from natural (Azamax® 1.2 EC) and synthetic (Premio® SC) origin. In this confrontation, AMSE (at LC90) showed efficacy similar to that of commercial products (used at registered dose for S. frugiperda control). Then, the bioguided fractionation using different chromatographic techniques which led to isolation of acetogenin rolliniastatin-1 identified as the majoritary compound of the most active ethanolic extract from A. mucosa. After that, rolliniastatin-1 was again tested against S. frugiperda larvae and caused significant acute (larval mortality) and chronic (decrease of larval development) effects. Finally, the bioactivity of a newly-registered acetogenin-based (Anosom® 1 EC) formulation was investigated, both separately and mixed with a limonoid-based formulation (Azamax® 1.2 EC). Anosom® 1 EC, tested at estimated LC90 (2,959.00 mg kg-1) and Azamax® 1.2 EC, tested at registered dose for S. frugiperda (4,000.00 mg kg-1) led to significant larval mortality, without happening any differences between treatments either tested separately or in a binary mixture. Anosom® 1 EC (at LC50) also led to significant increase of larval and pupal mortality and larval phase, as well as insect pupal weight decrease. Thus, Annonaceae derivates can be a useful component for S. frugiperda integrated management under field conditions.