Resistance of bmr energy sorghum hybrids to sugarcane borer and fall armyworm / Resistência de híbridos de sorgo energia bmr à broca-da-cana-de-açúcar e lagarta-do-cartucho

Abstract The lower lignin content in plants species with energy potential results in easier cellulose breakdown, making glucose available for ethanol generation. However, higher lignin levels can increase resistance to insect attack. The objective of this work was to evaluate the susceptibility of a bmr-6 biomass sorghum (a mutant genotype with a lower concentration of lignin) to important pests of energy sorghum, Diatraea saccharalis and Spodoptera frugiperda. Experiments were performed in the laboratory and greenhouse to evaluate the development of these pests on the biomass sorghum bmr hybrids BR007, BR008, and TX635 and their respective conventional near-isogenic genotypes (without the bmr gene). The lignin content was higher in non-bmr hybrids, but the evaluated insect variables varied between treatments, not being consistent in just one hybrid or because it is bmr or not. The lowest survival of S. frugiperda was observed in the BR008 hybrid, both bmr and non-bmr. The S. frugiperda injury scores on plants in the greenhouse were high (>7) in all treatments. For D. saccharalis, there was no difference in larval survival in the laboratory, but in the greenhouse, the BR007 hybrid, both bmr and non-bmr, provided greater survival. Due the need to diversify the energy matrix and the fact that greater susceptibility of the bmr hybrids to either pests was not found in this study, these results hold promise for cultivation of these biomass sorghum hybrids for the production of biofuels.

Resumo O menor teor de lignina em espécies de plantas com potencial energético resulta na maior facilidade de quebra da celulose, disponibilizando glicose para geração de etanol. Porém, maiores teores de lignina representa um fator de resistência ao ataque de insetos. O objetivo deste trabalho foi avaliar como importantes pragas do sorgo energia, Diatraea saccharalis e Spodoptera frugiperda, se comportam quanto à alimentação e desempenho em sorgo bmr-6, um genótipo mutante com menor concentração de lignina. Foram realizados experimentos em laboratório e casa de vegetação, avaliando o desenvolvimento destas pragas nos híbridos de sorgo biomassa bmr 007, 008, TX635 e seus respectivos genótipos isogênicos convencionais (sem o gene bmr). O teor de lignina foi maior nos híbridos não bmr, mas nos parâmetros avaliados nos insetos, houve variação entre os tratamentos, não sendo consistente em apenas um híbrido e nem por ser ou não bmr. A menor sobrevivência de S. frugiperda foi verificada no híbrido BR008 tanto bmr quanto não bmr. As notas de injúria por S. frugiperda no sorgo em casa de vegetação foram altas (>7) em todos os tratamentos. Para D. saccharalis, não houve diferença significativa para a sobrevivência larval em laboratório, mas em casa de vegetação o híbrido BR007 tanto bmr quanto não bmr proporcionaram maior sobrevivência. Diante da necessidade de diversificar a matriz energética e o fato de que não foi comprovada neste estudo maior suscetibilidade dos híbridos bmr a ambas as pragas, estes resultados são promissores para o cultivo desses híbridos de sorgo biomassa para produção de biocombustíveis.

Efficacy of Citrullus colocynthis seed extract on Earias vittella, Fabricius, (Lepidoptera: Noctuidae): environment sustainable approach / Eficácia do extrato de semente de Citrullus colocynthis em Earias vittella Fabricius (Lepidoptera: Noctuidae): abordagem ambiental sustentável

Abstract Earias vittellaFabricius, 1794 (Noctuidae: Lepidoptera) is deliberated to be one of the most destructive pests of cotton and okra vegetation in the world including Asia. The pest has established resistance to various synthetic insecticides. The use of bio-pesticide is one of the unconventional approaches to develop a vigorous ecosystem without harming non- target pests and beneficial natural insect fauna. In the present study, the toxicity levels of Citrullus colocynthis seed extract have been evaluated against the populations of E. vittellaunder standardized laboratory conditions. The toxic effects of C. colocynthis on development periods, protein contents and esterase activity of the life stages of E. vittella were also evaluated. The toxicity levels of methanol, ethanol, hexane, water and profenofos were evaluated on the 1st instar larvae of E. vittella. LC30 and LC80 concentrations exhibited the effectiveness of methanol-based C. colocynthis seed extract against 1st instar larvae of E. vitella. The enhanced larval and pupal periods were revealed in treated samples during the comparison with untreated samples. The intrinsic rate of increase, net reproductive rate in the LC30 and LC80 concentrations exposed larvae remained less than the control treatment. Fecundity, the esterase activity and protein contents were declined in LC30 and LC80 treated samples as compared to the control. The present findings suggest that C. colosynthis extracts based botanical insecticides are beneficial, ecosystem sustainable and can be integrated with insect management programs from environment safety perspective.

Resumo Earias vittella Fabricius, 1794 (Noctuidae: Lepidoptera) é considerada uma das pragas mais destrutivas de algodão e quiabo no mundo, incluindo a Ásia. Essa praga estabeleceu resistência a vários inseticidas sintéticos. O uso de biopesticidas é uma das abordagens não convencionais para desenvolver um ecossistema saudável sem prejudicar as pragas não alvo e a fauna natural benéfica de insetos. No presente estudo, os níveis de toxicidade do extrato de semente de Citrullus colocynthis foram avaliados nas populações de E. vittella em condições de laboratório padronizadas. Os efeitos tóxicos de C. colocynthis nos períodos de desenvolvimento, conteúdo de proteína e atividade esterase das fases de vida de E. vittella também foram avaliados. Os níveis de toxicidade de metanol, etanol, hexano, água e profenofós foram avaliados em larvas de 1º instar de E. vittella. As concentrações de LC30 e LC80 apresentaram eficácia do extrato de sementes de C. colocynthis à base de metanol contra larvas de 1º instar de E. vittella. Os períodos larval e pupal aumentados foram revelados nas amostras tratadas durante a comparação com as amostras não tratadas. A taxa intrínseca de aumento e a taxa reprodutiva líquida nas concentrações de larvas expostas LC30 e LC80 permaneceram menores do que o tratamento controle. A fecundidade, a atividade da esterase e o conteúdo de proteína diminuíram nas amostras tratadas com LC30 e LC80 em comparação com o controle. As presentes descobertas sugerem que os extratos de C. colocynthis à base de inseticidas botânicos são benéficos, sustentáveis ​​para o ecossistema e podem ser integrados com programas de manejo de insetos do ponto de vista da segurança ambiental.

Nova ocorrência de lepidopterismo por Hylesia nigricans (Berg, 1875) (Lepidoptera: Saturniidae) no Rio Grande do Sul, Brasil / New occurrence of lepidopterism by Hylesia nigricans (Berg, 1875) (Lepidoptera: Saturniidae) in Rio Grande do Sul, Brazil

Adult forms of lepidopterans can cause health problems. This type of incident is known as Lepidopterism, which is caused by contact with urticating setae released by adults, such as those from the Hylesia Hübner (Saturniidae) genus. In this study, the objective is to document a new occurrence of lepidopterism caused by specimens of Hylesia nigricans(Berg, 1875) (Lepidoptera: Saturniidae). The outbreak took place in the summer of 2017 in the municipality of Três Coroas, Rio Grande do Sul, Brazil. During this period, approximately 300 people were affected by dermatitis in the municipality.The outbreak resulted from the release of two types of urticating setae by female moths in the area. We want to emphasize that in case of proliferation of this moth species, it is essential to instruct the population to avoid direct contact with these insects. Monitoring these groups of lepidopterans is fundamental in facilitating decision-making and ensuring public health. (AU)

Formas adultas de lepidópteros podem causar problemas de saúde. Esse tipo de acidente é conhecido como lepidopterismo, causado pelo contato com cerdas urticantes liberadas por adultos, como as do gênero Hylesia Hübner (Saturniidae). Neste estudo, o objetivo é registrar uma nova ocorrência de lepidopterismo causado por espécimes de Hylesia nigricans (Berg, 1875) (Lepidoptera: Saturniidae). O surto ocorreu no verão de 2017, no município de Três Coroas, Rio Grande do Sul, Brasil. No período, aproximadamente 300 pessoas foram acometidas por dermatite no município. O surto foi causado pela liberação de dois tipos de setas urticantes das mariposas fêmeas no ambiente. Ressaltamos que na ocorrência da proliferação dessa espécie de mariposa, é importante instruir a população a evitar o contato direto com esses insetos. O monitoramento desses grupos de lepidópteros é fundamental para facilitar a tomada de decisões e garantir a saúde pública. (AU)

Impact of nitrogen and phosphorus fertilizers on Cry1Ac protein contents in transgenic cotton / Impacto de fertilizantes de nitrogênio e fósforo nos conteúdos de proteína Cry1Ac em algodão transgênico

Abstract Application of different fertilizers to check the efficiency of expression of Bt (Bacillus thuringiensis) gene in one of the leading commercialized crops (cotton) against Lepidopteran species is of great concern. The expression of Cry protein level can be controlled by the improvement of nutrients levels. Therefore, the myth of response of Cry toxin to different combinations of NP fertilizers was explored in three Bt cotton cultivars. Combinations include three levels of nitrogen and three levels of phosphorus fertilizers. Immunostrips and Cry gene(s) specific primer based PCR (Polymerase Chain Reaction) analysis were used for the presence of Bt gene that unveiled the presence of Cry1Ac gene only. Further, the ELISA (enzyme-linked immunosorbent assay) kit was used to quantify the expression of Cry1Ac protein. Under various NP fertilizers rates, the level of toxin protein exhibited highly significant differences. The highest toxin level mean was found to be 2.3740 and 2.1732 µg/g under the treatment of N150P75 kg ha-1 combination while the lowest toxin level mean was found to be 0.9158 and 0.7641 µg/g at the N50P25 kg ha-1 level at 80 and 120 DAS (Days After Sowing), respectively. It was concluded from the research that the usage of NP fertilizers has a positive relation with the expression of Cry1Ac toxin in Bt cotton. We recommend using the N150P50 kg ha-1 level as the most economical and practicable fertilizer instead of the standard dose N100P50 kg ha-1 to get the desired level of Cry1Ac level for long lasting plant resistance (<1.5). The revised dose of fertilizer may help farmers to avoid the cross-resistance development in contradiction of insect pests.

Resumo A aplicação de diferentes fertilizantes para verificar a eficiência da expressão do gene Bt (Bacillus thuringiensis) em uma das principais culturas comercializadas (algodão) contra espécies de lepidópteros é uma grande preocupação. A expressão do nível de proteína Cry pode ser controlada pela melhoria dos níveis de nutrientes. Portanto, o mito da resposta da toxina Cry a diferentes combinações de fertilizantes NP foi explorado em três cultivares de algodão Bt. As combinações incluem três níveis de nitrogênio e três níveis de fertilizantes de fósforo. A análise de PCR (reação em cadeia da polimerase) específica para o gene (s) Immunostrips e Cry (s) foi usada para a presença do gene Bt que revelou a presença do gene Cry1Ac apenas. Além disso, o kit ELISA (ensaio de imunoabsorção enzimática) foi usado para quantificar a expressão da proteína Cry1Ac. Sob várias taxas de fertilizantes NP, o nível de proteína de toxina exibiu diferenças altamente significativas. A média do nível mais alto de toxina foi de 2,3740 e 2,1732 µg / g sob o tratamento da combinação N150P75 kg ha-1, enquanto a média do nível mais baixo de toxina foi de 0,9158 e 0,7641 µg / g no nível de N50P25 kg ha-1 em 80 e 120 DAS (dias após a semeadura), respectivamente. Concluiu-se com a pesquisa que o uso de fertilizantes NP tem relação positiva com a expressão da toxina Cry1Ac no algodão Bt. Recomendamos o uso do nível de N150P50 kg ha-1 como o fertilizante mais econômico e praticável em vez da dose padrão N100P50 kg ha-1 para obter o nível desejado de nível de Cry1Ac para resistência de planta de longa duração (<1,5). A dose revisada de fertilizante pode ajudar os agricultores a evitar o desenvolvimento de resistência cruzada em contradição com as pragas de insetos.

Do fall armyworm's Metaflumizone resistante populations affect the activity of Trichogramma pretiosum? / Populações da lagarta-do-cartucho resistentes à Metaflumizona afetam a atividade de Trichogramma pretiosum?

Abstract The possible interference of resistant pest's populations to insecticides in natural enemies in the action thas not been clarified yet. Thus, this study aimed to evaluate Trichogramma pretiosum Riley (Hymenoptera: Trichogrammatidae) performance on Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae) eggs with resistance frequency to the Metaflumizone over six generations of product exposure. Egg cards (2.0 x 7.0 cm) containing eggs from two populations of S. frugiperda, (resistant to Metaflumizone and the other susceptible), were exposed to T. pretiosum females for 24 hours in free-choice and no-choice testing in three generations (G1, G4, and G6). A completely randomized experimental design was used with 25 replications, each consisting of an egg card (experimental unit) containing 20 eggs. The parameters evaluated were: parasitism (%), emergence (%), sex ratio, number of emerged parasitoids per egg and males/females longevity. ANOVA and Tukey test (P≤ 0.05) were applied on the results. Results showed a reduction in parasitism [41.0% (G1) and 28.4% (G4)], egg emergence (17.5%) and parasitoids/egg [16.2 (G4) and 17.2 (G6)] in eggs originating from the population with resistance frequency. Females emerging from G6 populations eggs without exposure to Metaflumizone had greater longevity (3.5 days more) than the resistant population. The sex ratio and male longevity were not affected. The results indicate a reduction in T. pretiosum activity if S. frugiperda populations have some frequency of resistance to Metaflumizone.

Resumo A possível interferência de populações de pragas resistentes na ação de inimigos naturais ainda não foi esclarecida. Assim, este trabalho teve como objetivo avaliar o desempenho de Trichogramma pretiosum Riley (Hymenoptera: Trichogrammatidae) em ovos de Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae) com frequência de resistência à Metaflumizona ao longo de seis gerações de exposição ao produto. Cartelas (2,0 x 7,0 cm) com ovos de duas populações de S. frugiperda, (resistente à Metaflumizona e outra suscetível), foram expostas às fêmeas de T. pretiosum por 24 horas em condições de livre escolha e sem chance de escolha por três gerações (G1, G4 e G6). O delineamento experimental foi inteiramente casualizado com 25 repetições, sendo cada repetição composta por uma cartela (unidade experimental) contendo 20 ovos. Os parâmetros avaliados foram: parasitismo (%), emergência (%), razão sexual, número de parasitoides emergidos por ovo e longevidade de machos e fêmeas. ANOVA e teste de Tukey (P≤ 0,05) foram aplicados aos dados coletados. Os resultados mostraram redução do parasitismo [41,0% (G1) e 28,4% (G4)], emergência de ovos (17,5%) e parasitoides/ovo [16,2 (G4) e 17,2 (G6)] em ovos oriundos da população com frequência de resistência. As fêmeas emergidas de ovos da população G6 sem exposição à Metaflumizona, tiveram maior longevidade (3,5 dias a mais) do que a população exposta ao inseticida. A razão sexual e a longevidade de machos não foram afetadas. Os resultados indicam uma redução na atividade de T. pretiosum se as populações de S. frugiperda apresentarem alguma frequência de resistência à Metaflumizona.

Prokaryotic expression, polyclonal antibody preparation, spatio-temporal expression profile and functional analysis of c-Myc of Helicoverpa armigera (Lepidoptera: Noctuidae) / 生物工程学报

c-Myc protein encoded by c-Myc (cellular-myelocytomatosis viral oncogene) gene regulates the related gene expression through the Wnt/β-catenin signaling pathway, and has received extensive attention in recent years. The purpose of this study was to express Helicoverpa armigera c-Myc gene (Ha-c-Myc) by using prokaryotic expression system, prepare the polyclonal antibody, examine the spatio-temporal expression profile of Ha-c-Myc, and investigate the possible function of Ha-c-Myc in regulating H. armigera sterol carrier protein-2 (SCP-2) gene expression. The Ha-c-Myc gene was amplified by PCR and cloned into a prokaryotic expression plasmid pET-32a(+). The recombinant plasmid pET-32a-Ha-c-Myc was transformed into Escherichia coli BL21. IPTG was used to induce the expression of the recombinant protein. Protein was purified by Ni2+-NTA column and used to immunize New Zealand rabbits for preparing the polyclonal antibody. The Ha-c-Myc expression levels in different developmental stages (egg, larva, prepupa, pupa, and adult) of H. armigera and different tissues (midgut, fat body, head, and epidermis) of the prepupa were determined by real-time quantitative reverse transcription PCR (qRT-PCR). Ha-c-Myc siRNA was synthesized and transfected into H. armigera Ha cells. The relative mRNA levels of Ha-c-Myc and HaSCP-2 in Ha cells were detected by qRT-PCR. Results showed that the pET-32a-Ha-c-Myc recombinant plasmid was constructed. The soluble Ha-c-Myc protein of about 65 kDa was expressed in E. coli. The polyclonal antibody was prepared. Western blotting analysis suggested that the antibody had high specificity. Enzyme linked immunosorbent assay (ELISA) showed that the titer of the antibody was high. Ha-c-Myc gene expressed at all developmental stages, with high levels in the early and late instars of larva, and the prepupal stage. Tissue expression profiles revealed that Ha-c-Myc expressed in various tissues of prepupa, with high expression level in the midgut, but low levels in the epidermis and fat body. RNAi results showed that the knockdown of Ha-c-Myc expression significantly affected transcription of HaSCP-2, leading to a 50% reduction in HaSCP-2 mRNA expression level. In conclusion, the Ha-c-Myc was expressed through a prokaryotic expression system, and the polyclonal anti-Ha-c-Myc antibody was obtained. Ha-c-Myc may promote the expression of HaSCP-2 and play an important role in the lipid metabolism of H. armigera. These results may facilitate further study on the potential role and function mechanism of Ha-c-Myc in H. armigera and provide experimental data for exploring new targets of green pesticides.

Correlation of extracellular enzymes activity of Candida glabrata clinical isolates with in vivo pathogenicity in Galleria mellonella larvae / 中华预防医学杂志

Objective: To explore the relationship between extracellular enzymes activity and virulence of Candida glabrata clinical isolates based on the infection model of Galleria mellonella larvae. Methods: Using experimental research methods, 71 strains of non-repetitive Candida glabrata were collected from Qinghai Provincial People's Hospital from June 2021 to January 2022. Bovine serum protein agar medium, egg yolk agar medium, sheep blood agar medium, Tween-80 agar medium and triglyceride agar medium were used to detect the aspartyl protease activity, phospholipase activity, hemolysis activity, esterase activity and lipase activity of Candida glabrata. Median lethal concentration (LC50) was calculated by using 1.25×108 CFU/ml,2.50×108 CFU/ml,3.75×108 CFU/ml,5.00×108 CFU/ml suspension of Candida glabrata ATCC2001 to infect Galleria mellonella larvae. Histopathological and etiological analysis was performed to determine whether the infection model was successfully established. The clinical isolates of Candida glabrata were configured to infect Galleria mellonella larvae with LC50 concentration to detect the pathogenicity of Galleria mellonella larvae.Spearman test or Pearson test were used to analyze the correlation between the extracellular enzyme activity of Candida glabrata clinical isolates and the pathogenicity of Galleria mellonella larvae. Results: 71 strains of Candida glabrata isolated clinically were detected to have low hemolytic activity after 2 days of culture. Aspartyl protease was detected after 4 days of culture, among which 7 strains (9.86%), 19 strains (26.76%) and 45 strains (63.38%) showed low, medium and high aspartyl protease activity. After 7 days of culture, 71 strains did not detect phospholipase, esterase and lipase activities. Candida glabrata on Galleria mellonella larvae of LC50=2.5×108 CFU/ml Fungal spore were found in the intestinal tissue pathological section of Galleria mellonella larvae in the experimental group, and Candida glabrata was identified by the microbial Mass Spectrometry after culture, while no fungi were found in the pathological section and culture of the control group. Spearman test shows that, there was a linear positive correlation between aspartyl protease activity and the survival rate of Galleria mellonella larvae (r = 0.73, P<0.01), the difference was statistically significant.Pearson test shows that, there was no significant linear relationship between hemolytic activity and survival rate of Galleria mellonella larvae (r = 0.16, P = 0.34), the difference was not statistically significant. Conclusion: The clinical isolates of Candida glabrata in this study had aspartyl protease activity and low hemolytic activity, but no phospholipase, esterase and lipase activity. The activity of aspartyl aspartyl protease of Candida glabrata was positively correlated with the pathogenicity of Galleria mellonella larvae.

Effects of habitat loss on three insect assemblages in modified ecosystems of foothills of the Colombian Orinoquia / Efectos de la pérdida de hábitat en tres ensambles de insectos en ecosistemas modificados de Piedemonte en la Orinoquia colombiana

Abstract Introduction: The effects of habitat transformation have been widely studied and the effects are well-known at different levels of biological organization. However, few studies have focused on responses to this process at the level of multiple taxa in diverse taxonomic and functional groups. Objective: Determine the variations in taxonomic and functional diversity of ants, butterflies, and dung beetles, at a spatial and temporal level in a landscape mosaic of the ecoregion of the Colombian foothills. Methods: We assessed amount of natural habitat and landscape composition in four types of vegetation, during the highest and lowest rain periods. We collected butterflies with hand nets and used baited pitfall traps for dung beetles and ants. Results: Habitat loss positively affected ant and butterfly species richness, and negatively affected dung beetles. The abundance of ants and butterflies had a positive effect on the dominance of species in the transformed vegetation, for dung beetles the abundance was negatively affected by the absence of canopy cover. Habitat loss had no negative effect on functional diversity as there is no difference between natural and transformed vegetation. Conclusions: The amount of habitat, habitat connectivity and different types of vegetation cover were important factors in the maintenance of insect diversity in the modified ecosystems of foothills of the Colombian Orinoquia. The lack of a common spatial and temporal pattern shows that studies of multiple insect taxa should be carried out for biodiversity monitoring and conservation processes.

Resumen Introducción: Los efectos de la transformación del hábitat han sido ampliamente estudiados y son bien conocidos los efectos a diferentes niveles de organización biológica. Sin embargo, pocos estudios se han centrado en las respuestas a este proceso a nivel de múltiples taxones en diversos grupos taxonómicos y funcionales. Objetivo: Determinar las variaciones en la diversidad taxonómica y funcional de hormigas, mariposas y escarabajos coprófagos, a nivel espacial y temporal en un mosaico paisajístico de la ecorregión del piedemonte colombiano. Métodos: Evaluamos la cantidad de hábitat natural y la composición del paisaje en cuatro tipos de vegetación, durante los períodos de mayor y menor lluvia. Recolectamos mariposas con redes de mano y usamos trampas de caída con cebo para escarabajos coprófagos y hormigas. Resultados: La pérdida de hábitat afectó positivamente la riqueza de especies de hormigas y mariposas y afectó negativamente a los escarabajos peloteros. La abundancia de hormigas y mariposas tuvo un efecto positivo sobre la dominancia de especies en la vegetación transformada, para los escarabajos coprófagos la abundancia se vio afectada negativamente por la ausencia de cobertura de dosel. La pérdida de hábitat no tuvo un efecto negativo sobre la diversidad funcional ya que no hay diferencia entre la vegetación natural y la transformada. Conclusiones: La cantidad de hábitat, la conectividad del hábitat y los diferentes tipos de cobertura vegetal fueron factores importantes en el mantenimiento de la diversidad de insectos en los ecosistemas modificados del piedemonte de la Orinoquia colombiana. La falta de un patrón espacial y temporal común muestra que se deben realizar estudios de múltiples taxones de insectos para los procesos de monitoreo y conservación de la biodiversidad.

Ethological aspects of Helicoverpa armigera in the reproductive phase of cotton

Pest behavior studies are important to inform the periods when the pest is more exposed to pesticide. This study aimed to evaluate the movement and feeding of Helicoverpa armigera larvae in the first three instars during the reproductive phase of cotton. First, larval behavior was evaluated with a completely randomized design in a 3 × 5 factorial design with eight replications. The treatments were the instars versus behaviors (crawling, resting, waving, eating and dropping-off). In addition, another 3 × 6 factorial design with eight replications was performed to evaluate the plant region most preferred by the caterpillars. The treatments were the instars versus six parts of the plant (abaxial surface, adaxial surface, on floral bud, internal surface of bract, petiole, and terminal growth). Then, the period of the day when caterpillars are more active was also evaluated by a 3 × 2 factorial essay. The treatments were the instars versus periods of the day. Eating and resting are the most frequent behaviors for all instars. The plant regions preferred by the caterpillars for crawling are on the floral bud and the adaxial face of the leaf. There is no preferred period of the day for eating, resting, waving or dropping-off. Caterpillars (until the third instar) prefer to walk in the morning and other behaviors can occur at any period. Therefore, sprays should preferably occur in the morning, and the insecticide drops/spray should reach the flower buds and the adaxial surface of the leaves to contaminate the larvae.

Molecular identification, characterization, and expression analysis of a serine protease inhibitor gene from cotton bollworm, Helicoverpa armigera (Lepidoptera: Noctuidae) / Identificação molecular, caracterização e análise de expressão de um gene inibidor de serina protease da lagarta Helicoverpa armigera (Lepidoptera: noctuidae) na cultura de algodão

Abstract Serine protease inhibitors (serpins), a superfamily of protease inhibitors, are known to be involved in several physiological processes, such as development, metamorphosis, and innate immunity. In our study, a full-length serpin cDNA, designated Haserpin1, was isolated from the cotton bollworm Helicoverpa armigera. The cDNA sequence of Haserpin1 is 1176 nt long, with an open reading frame encoding 391 amino acids; there is one exon and no intron. The predicted molecular weight of Haserpin1 is 43.53 kDa, with an isoelectric point of 4.98. InterProScan was employed for Haserpin1 functional characterization, which revealed that Haserpin1 contains highly conserved signature motifs, including a reactive center loop (RCL) with a hinge region (E341-N350), the serpin signature, (F367-F375) and a predicted P1-P1′ cleavage site (L357-S358), which are useful for identifying serpins. Transcripts of Haserpin1 were constitutively expressed in the fat body, suggesting that it is the major site for serpin synthesis. During the developmental stages, a fluctuation in the expression level of Haserpin1 was observed, with low expression detected at the 5th-instar larval stage. In contrast, relatively high expression was detected at the prepupal stage, suggesting that Haserpin1 might play a critical role at the H. armigera wandering stage. Although the detailed function of this serpin (Haserpin1) needs to be elucidated, our study provides a perspective for the functional investigation of serine protease inhibitor genes.

Resumo Sabe-se que os inibidores de serina protease (serpinas), uma superfamília de inibidores de protease, estão envolvidos em vários processos fisiológicos, como desenvolvimento, metamorfose e imunidade inata. Neste estudo, um cDNA de serpina de comprimento total, denominado Haserpin1, foi isolado da lagarta Helicoverpa armigera na cultura de algodão. A sequência de ADNc de Haserpin1 tem 1.176 nt de comprimento, com uma grelha de leitura aberta que codifica 391 aminoácidos; existe um éxon, mas nenhum íntron. O peso molecular previsto de Haserpin1 é de 43,53 kDa, com um ponto isoelétrico de 4,98. O InterProScan foi empregado para a caracterização funcional do Haserpin1, que revelou que o Haserpin1 contém motivos de assinatura altamente conservados, incluindo um loop central reativo (RCL) com uma região de dobradiça (E341-N350), a assinatura da serpina (F367-F375) e um local de clivagem previsto de P1-P1' (L357-S358), que são úteis para identificar serpinas. As transcrições de Haserpin1 foram expressas constitutivamente no corpo gordo, sugerindo que é o principal local para a síntese de serpinas. Durante os estágios de desenvolvimento, observou-se uma flutuação no nível de expressão de Haserpin1, com baixa expressão detectada no estágio larval do 5º ínstar. Por outro lado, detectou-se uma expressão relativamente alta no estágio pré-pupal, sugerindo que o Haserpin1 pode desempenhar um papel crítico no estágio errante de H. armigera. Embora a função detalhada dessa serpina (Haserpin1) precise ser elucidada, este estudo fornece uma perspectiva para a investigação funcional dos genes inibidores da serina protease.

Efficiency of biological control for fall armyworm resistant to the protein Cry1F / Eficiência do controle biológico da lagarta-do-cartucho resistente à proteína Cry1F

Abstract Understanding the ecological and toxicological relationship between genetically modified cultivars (GM) and biological control agents is of great importance for discussions related to the compatability of GM cultivars and integrated management strategies for pest resistance. The present study evaluated the search behavior and predatory capacity of Orius insidiosus (Say) (Hemiptera: Anthocoridae) and Doru luteipes (Scudder) (Dermaptera: Forficulidae) on eggs and caterpillars of Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae) resistant or not to the protein Cry1F expressed in Bt corn. To determine the search time, a stopwatch was run until the capture of the first prey, predation capacity was evaluated by counting the prey remaining after 24 hours of infestation. The injuries of S. frugiperda in genetically modified and conventional corn in the presence and absence of predators was also evaluated. The predators were not able to distinguish between resistant and susceptible prey (eggs or caterpillars), given the predatory behaviour observed. There was no difference in searching time or predatory capacity between the predators for eggs and caterpillars of either resistant or susceptible S. frugiperda. In the presence of predators, the injury scores for resistant S. frugiperda on the Bt corn plants were lower. It was concluded that O. insidiosus and D. luteipes did not notice the presence of the protein Cry1F in the prey S. frugiperda, which may facilitate the combined use of GM corn and biological control in integrated management programs and for management of pest resistance.

Resumo O entendimento de relações ecológicas e toxicológicas envolvendo culturas geneticamente modificadas (GM) e agentes de controle biológico é de grande importância para discussões relativas à compatibilidade de culturas GM com estratégias de manejo integrado e manejo de resistência de pragas. Este trabalho avaliou o comportamento de busca e a capacidade predatória de Orius insidiosus (Say) (Hemiptera: Anthocoridae) e Doru luteipes (Scudder) (Dermaptera: Forficulidae) sobre ovos e lagartas de Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae) resistente ou não à proteína Cry1F expressa em milho Bt. Para determinar o tempo de busca foi utilizado um cronômetro que foi disparado até a captura da primeira presa; a capacidade de predação foi avaliada através da contagem das presas remanescentes 24 h após infestação. Também foram avaliadas as injúrias de S. frugiperda em milho transgênico e milho convencional na presença ou ausência dos predadores. Os predadores não foram capazes de distinguir entre presas (ovos ou lagartas) resistentes e suscetíveis, considerando os comportamentos predatórios avaliados. Não houve diferença no tempo de busca e capacidade predatória sobre ovos e lagartas de S. frugiperda resistente ou suscetível entre os predadores. Na presença dos predadores, as notas de injúria de S. frugiperda resistente nas plantas de milho Bt foram menores. Conclui-se que O. insidiosus e D. luteipes não percebem a presença da proteína Cry1F na presa S. frugiperda, o que pode contribuir para o uso integrado de milho GM e controle biológico em programas de manejo integrado e manejo de resistência de pragas.

Tetrastichus howardi (Hymenoptera: Eulophidae): first report of parasitism in Oxydia vesulia (Lepidoptera: Geometridae) / Tetrastichus howardi (Hymenoptera: Eulophidae): primeiro relato de parasitismo em Oxydia vesulia (Lepidoptera: Geometridae)

The adaptation of native lepidopteran species to eucalyptus plantations reduces the productivity of this crop in Brazil. Oxydia vesulia Cramer (Lepidoptera: Geometridae) is a secondary pest, frequently reported in eucalyptus plantations with population outbreaks and economic damages. Methods of biological control of this pest may include the use of the exotic pupae endoparasitoid Tetrastichus howardi Olliff (Hymenoptera: Eulophidae), reported as efficient to controlling lepidopteran pests. The parasitism of O. vesulia caterpillars and pupae by T. howardi was evaluated under controlled conditions (25 ± 1 ºC, 60 ± 20% humidity and 12:12 h L:D). Each O. vesulia caterpillar or pupae was individually placed in a flat-bottom tube with 10 and 15 females of T. howardi for 48h, respectively. The parasitoids were removed after that period, the caterpillars were fed and the pupae were maintained until emergence of the parasitoid or formation of pupae and emergence of adults of this pest. The fourth-instar caterpillars of O. vesulia, after the parasitism period, were kept in pots with Eucalyptus urophylla leaves, changed daily until the end of the experiment. A total of 40% of the caterpillars died before the pre-pupae stage, 40% reached the pupae stage and died due to inadequate adult formation and 20% generated moths, but none adult parasitoid emerged from the caterpillars. All pupae of O. vesulia were parasitized and showed emergence of parasitoids. The parasitism of O. vesulia caterpillars and pupae by T. howardi shows the potential of this natural enemy for the integrated management of this defoliator pest in eucalyptus plantations.

A adaptação de espécies nativas de lepidópteros a plantios de eucalipto reduz a produtividade dessa cultura no Brasil. Oxydia vesulia Cramer (Lepidoptera: Geometridae) é praga-secundária, frequentemente, relatada em plantios de eucalipto com surtos populacionais e danos econômicos. Métodos de controle biológico, dessa praga, podem incluir o uso do endoparasitoide exótico de pupas Tetrastichus howardi Olliff (Hymenoptera: Eulophidae) relatado como eficiente para o controle de lepidópteros-pragas. O parasitismo de lagartas e pupas de O. vesulia por T. howardi foi avaliado em condições controladas (25 ± 1º C, 60 ± 20% umidade e 12h fotoperíodo). Cada lagarta ou pupa de O. vesulia foi colocada individualmente em tubos de fundo chato com 10 e 15 fêmeas de T. howardi por 48h, respectivamente e, após esse período, os parasitoides foram removidos, as lagartas alimentadas e as pupas mantidas até a emergência do parasitoide ou formação de pupas ou de adultos dessa praga. As lagartas de quarto ínstar de O. vesulia, após o período de parasitismo, foram mantidas em potes com folhas de Eucalyptus urophylla trocadas, diariamente, até o fim do experimento. Um total de 40% das lagartas de O. vesulia morreu antes da fase de pré-pupa, 40% atingiram o estágio de pupa e morreram devido à má formação ou problemas e 20% originaram mariposas, porém nenhum parasitoide adulto emergiu das lagartas. Todas as pupas de O. vesulia foram parasitadas e tiveram emergência de parasitoides. O parasitismo de lagartas e pupas de O. vesulia por T. howardi mostra o potencial desse inimigo natural para o manejo integrado desse desfolhador em cultivos de eucalipto.

Dual effects of entomopathogenic fungi on control of the pest Lobesia botrana and the pathogenic fungus Eutypella microtheca on grapevine

BACKGROUND: Entomopathogenic fungi (EPF) are the natural enemies of insect pests. Nevertheless, research on the use of EPF for simultaneous prevention of pest and disease agents on the same crop is limited. In this study, we explored the potential dual effects of three strains of the EPF Metarhizium anisopliae on the control of detrimental agents of Vitis vinifera L., including different developmental stages (larvae, pupae, and adult) of the insect pest Lobesia botrana and the phytopathogenic fungus Eutypella microtheca. METHODS: Laboratory pathogenicity trials were performed to examine the effects of the three M. anisopliae strains on the mortality rate of L. botrana. In addition, field trials were conducted to assess the biocontrol potential of one selected M. anisopliae strain on the larval stage of L. botrana. Moreover, inhibitory effects of the three EPF strains on E. microtheca growth were examined in vitro. RESULTS: All the M. anisopliae strains were highly effective, killing all stages of L. botrana as well as inhibiting the growth of E. microtheca. The in vitro mortality of larvae treated with the strains was over 75%, whereas that of treated pupae and adults was over 85%. The three EPF strains showed similar efficacy against larvae and adult stages; never-theless, pupal mortality was observed to be strain dependent. Mortality of L. botrana larvae ranged from 64 to 91% at field conditions. Inhibition of E. microtheca growth reached 50% in comparison to the control. CONCLUSIONS: Our study showed that M. anisopliae strains were highly effective in ensuring control of two different detrimental agents of V. vinifera L., providing new evidence to support the dual effects of entomopathogenic fungi.

Study of Expression of Indigenous Bt cry2AX1 Gene in T3 Progeny of Cotton and its Efficacy Against Helicoverpa armigera (Hubner)

Abstract Development of transgenic Bt crops with stable and high level of Bt protein expression over generations under different environmental conditions is critical for successful deployment at field level. In the present study, progenies of transgenic cotton Coker310 event, CH12 expressing novel cry2AX1 gene were evaluated in T3 generation for stable integration, expression and resistance against cotton bollworm, Helicoverpa armigera. The cry2AX1 gene showed stable inheritance and integration in the T3 progeny plants as revealed by PCR and Southern blot hybridization. The expression of Cry2AX1 protein on 90 days after sowing (DAS) was in the range of 1.055 to 1.5 µg/g of fresh leaf tissue except one plant which showed 0.806 µg/g of fresh leaf tissue and after 30 days (i.e., on 120 DAS) three plants recorded in between 0.69 to 0.82 µg/g and other plants are in range of 0.918 to 1.058 µg/g of fresh leaf tissue. Detached leaf bit bioassay in T3 progeny on 110 DAS recorded mortality of 73.33 to 93.33 per cent against H. armigera and severe growth retardation in surviving larvae. These results indicate that the expression of chimeric cry2AX1 is stable and exhibits insecticidal activity against H. armigera in T3 progeny of CH12 event of transgenic cotton.

Owlet moths (Lepidoptera: Noctuoidea) associated with Bt and non- Bt soybean in the brazilian savanna / Noctuóides (Lepidoptera: Noctuoidea) associados a soja Bt e não-Bt no Cerrado brasileiro

Abstract The use of GMO expressing Bt toxin in soybean production has increased significantly in the last years in Brazil in order to manage the damage caused by lepidopteran pests. In this study, we compared the richness and abundance of owlet moths (Noctuoidea) associated with Bt and non-Bt soybean. We determined the temporal variations as a function of phenology, and correlated the population variations of the most common species with meteorological variables. The research was conducted at the experimental area of Embrapa Cerrados. The collection method used was differentiated being suppressive and absolute. A total of 13 species were collected, of which eight occurred on Bt soybeans. The most representative taxa were Chrysodeixis includens (72.87%), Anticarsia gemmatalis (18.17%) and Spodoptera spp (5.22%). The number of larvae belonging to species targeted by the Bt technology was 10 times lower on Bt than on non-Bt soybeans. Utetheisa ornatrix and Elaphria deltoides were recorded on soybean for the first time, observing larvae of both species in non-Bt soybean and those of U. ornatrix also in Bt soybean. Only A. gemmatalis larvae correlated (p <0.05) negatively with precipitation. This study provided field information on the abundance and species richness of owlet moths on non-Bt soybeans, associated with the effects of Bt soybean. When considering the different levels of infestation between cultivars as a criterion, larvae monitoring is of substantial importance in order to develop the lost control program.

Resumo O uso de OGM que expressam toxina Bt na produção de soja tem aumentado significativamente nos últimos anos no Brasil e são utilizados para conter os danos causados ​​pelos lepidópteros pragas. Neste estudo comparamos a riqueza e a abundância de Noctuoides (Noctuoidea) associados à soja Bt e não-Bt. Determinamos as variações temporais em função da fenologia e correlacionamos às variações populacionais das espécies mais comuns com variáveis ​​meteorológicas. A pesquisa foi conduzida na área experimental da Embrapa Cerrados. O método de coleta utilizado foi diferenciado sendo supressivo e absoluto. Um total de 13 espécies foram coletadas, das quais oito ocorreram em soja Bt. Os taxa mais representativos foram Chrysodeixis includens, Anticarsia gemmatalis e Spodoptera spp. O número de larvas pertencentes às espécies alvo da tecnologia Bt foram 10 vezes menores na soja Bt do que em soja não-Bt . Utetheisa ornatrix e Elaphria deltoides foram registradas na soja pela primeira vez, observando-se larvas de ambas espécies na soja não-Bt e as de U. ornatrix também na soja Bt. Somente as larvas de A. gemmatalis se correlacionaram (p <0,05) negativamente com a precipitação. Este estudo forneceu informações em campo sobre a abundância e riqueza de espécies na soja não- Bt, associada aos efeitos da soja Bt. A importância do monitoramento das lagartas é substancial, a fim de tomar a melhor decisão de controle, considerando-se os diferentes níveis de infestação entre cultivares como critério.

Megaselia Rondani (Diptera: Phoridae) larvae as a Sphingidae (Lepidoptera) parasitoid / Larvas de Megaselia Rondani (Diptera: Phoridae) como parasitoide de Sphingidae (Lepidoptera)

Detritivoria and parasitism are dietary habits already registered for the phorids of the genus Megaselia Rondani. The results of this study confirm the parasitic habit of Megaselia larvae. This is the first Pachylia ficus (Linnaeus, 1758) (Lepidoptera: Sphingidae) adult record as a host of Megaselia scalaris (Loew, 1866) and Megaselia sp. (Diptera: Phoridae).(AU)

Detritivoria e parasitismo são hábitos alimentares já registrados para os forídeos do gênero Megaselia Rondani. Os resultados do presente estudo confirmam o hábito parasitário das larvas de Megaselia. Este é o primeiro registro de adulto de Pachylia ficus (Linnaeus, 1758) (Lepidoptera: Sphingidae) como hospedeiro de Megaselia scalaris (Loew, 1866) e Megaselia sp. (Diptera: Phoridae).(AU)