Dissertationen zum Thema „Pesticide resistance“

In recent years mosquito-borne diseases have reemerged, largely because of pesticide resistance. The mosquitoes develop resistance to pesticides because of broad and repeated uses of pesticides. Preventing the development of pesticide resistance requires proper understanding of the environmental factors potentially associated with the development of the resistance. The purpose of this study was to investigate the environmental factors associated with the resistance to pesticides by the Culex mosquitoes. This correlational study included the analysis of archived data samples (N = 29,794) from the Field Cage Tests results conducted between 2013 and 2017 by a large county public health department. The compartmental model was used to help understand and interpret study findings. To examine the associations between the independent variables (i.e., wind speed, temperature, humidity, time, month, and weather conditions) and the dependent variable (i.e., pesticide resistance), chi-square and multiple logistic regression analyses were performed. The results showed that the odds of mosquitoes developing resistance were 2.1 times higher during high temperatures than at low temperatures [x2(1) = 346.5, p = .000]; the resistance was 1.5 times higher during high humidity than in low humidity [x2(1) = 7.23, p = .007]; and the odds of mosquitoes developing resistance to pesticides in August were 3 times higher than when sprayed in June or July [x2 (2) = 702.606, p = .000]. Study findings may be used to help with the development of more effective methods for vector control thereby reducing the numbers of nuisance and disease-carrying mosquitoes along with a possible reduction in the incidence of mosquito-borne diseases and related human morbidity and mortality.

Bibliography: leaves 165-188. This research examines biological and behavioural interactions among the diamond-back moth, Plutella xylostella L. (Lepidoptera: Plutellidae), Cotesia plutellae Kurdjumov (Hymenoptera: Braconidae) and the microbial insecticide, Bacillus thuringiensis Berliner. A deterministic simulation model is developed describing the influence of C. plutellae on the evolution of resistance to Bt in a diamond-back moth poulation.

Bases para a implementação de um programa de manejo da resistência de Blattella germanica (L.) a inseticidas foram coletadas no presente trabalho. Técnicas de bioensaio de aplicação tópica e de contato tarsal foram comparadas inicialmente para a detecção da resistência a deltametrina e clorpirifós. O bioensaio de aplicação tópica possibilitou uma melhor discriminação entre a linhagem suscetível e as populações de campo de B. germanica testadas. Resultados do monitoramento da suscetibilidade de B. germanica a inseticidas evidenciaram a presença de variabilidade genética que confere resistência a deltametrina, clorpirifós e fipronil em populações coletadas em alguns estabelecimentos comerciais dos Estados de São Paulo e Rio de Janeiro. As razões de resistência encontradas para deltametrina e clorpirifós foram de aproximadamente 43 e 6 vezes, respectivamente. A resistência de B. germanica a deltametrina foi instável, ou seja, houve uma redução significativa na freqüência de resistência na ausência de pressão de seleção. Estudos de relações de resistência cruzada revelaram uma baixa intensidade de resistência cruzada (aproximadamente 2 vezes) entre fipronil e os inseticidas deltametrina e clorpirifós. E por último, a mistura de deltametrina e clorpirifós foi avaliada para o controle de linhagens de B. germanica resistentes a deltametrina. A mistura proporcionou um melhor controle da linhagem suscetível do que os produtos utilizados isoladamente. No entanto, a discriminação entre as linhagens suscetível e resistente a deltametrina foi evidente com a mistura nas doses recomendadas comercialmente, o que inviabilizaria o emprego desta estratégia no manejo da resistência de B. germanica a deltametrina.<br>Bases for implementing a resistance management program of Blattella germanica (L.) to insecticides were collected in this study. Initially, we compared the topical and residual tarsal-contact bioassays for detecting resistance to deltamethrin and chlorpyrifos. The topical bioassay gave better discrimination between susceptible and field-collected populations of B. germanica. Results from a survey of the susceptibility of B. germanica to insecticides revealed the presence of genetic variability that confers resistance to deltamethrin, chlorpyrifos and fipronil in field-collected populations of B. germanica from the States of São Paulo and Rio de Janeiro. The resistance ratios to deltamethrin and chlorpyrifos were approximately 43 and 6-fold, respectively. The resistance of B. germanica to deltamethrin was unstable; that is, a significant reduction in the frequency of resistance was detected under absence of selection pressure. Studies on cross-resistance relationships showed a low intensity of cross-resistance (approximately 2-fold) between fipronil and the insecticides deltamethrin and chlorpyrifos. And finally, the viabilility of the use of deltamethrin and chlorpyrifos mixture was evaluated for controlling deltamethrin-resistant strains of B. germanica. The use of mixtures gave better control of the susceptible strain than the use of the products by itself. However, the discrimination between susceptible and resistant strains was clear with the use of mixtures at recommended rates, which impairs the use of this strategy for managing B. germanica resistance to deltamethrin.

O objetivo do presente trabalho foi o de coletar informações básicas para a implementação de um programa de manejo da resistência de Aphis gossypii Glover, 1877 a inseticidas na cultura do algodão. Para tanto, foram caracterizadas as linhas-básicas de suscetibilidade de A. gossypii aos inseticidas carbosulfam, tiametoxam e endosulfam por meio de um bioensaio de contato residual. Posteriormente, foi realizado um monitoramento da suscetibilidade a esses inseticidas em populações de A. gossypii que foram coletadas nas regiões de Acreúna-GO, Ituverava-SP, Rondonópolis-MT e Unai-MG nas safras de algodão de 2001/2002 a 2003/2004. Em cada safra, as coletas de pulgões foram realizadas no início, meio e final do ciclo da cultura. Não foram observadas diferenças significativas na suscetibilidade de populações de A. gossypii para tiametoxam e endosulfam. Por outro lado, houve redução significativa na suscetibilidade ao carbosulfam ao longo do ciclo da cultura. No entanto, foi observado um restabelecimento parcial da suscetibilidade ao carbosulfam no início da safra subseqüente, sugerindo assim a instabilidade da resistência. A razão de resistência de A. gossypii ao carbosulfam foi de 5,7 vezes. Não foi verificada a presença de resistência cruzada entre carbosulfam e os inseticidas tiametoxam e endosulfam. Estudos de dinâmica da resistência de A. gossypii ao carbosulfam realizados em condições de laboratório mostraram que a resistência é instável, ou seja, a freqüência de resistência diminui significativamente na ausência de pressão de seleção. Comparações de parâmetros biológicos entre as linhagens suscetível e resistente ao carbosulfam mostraram que há custo adaptativo associado à resistência. A linhagem resistente apresentou uma menor fecundidade, menor período reprodutivo e menor longevidade do que a linhagem suscetível. Por fim, trabalhos de avaliação da estratégia de mistura de inseticidas evidenciaram que a mistura de carbosulfam com tiametoxam ou endosulfam não é vantajoso no manejo da resistência de A. gossypii ao carbosulfam.<br>The objective of this research was to collect basic information for building an insecticide resistance management program of Aphis gossypii Glover, 1877 in cotton crop. Baseline susceptibility data of A. gossypii to carbosulfan, thiamethoxam and endosulfan were characterized by the use of direct contact bioassay. Then, a susceptibility monitoring to these insecticides was conducted in populations of A. gossypii collected from Acreúna-GO, Ituverava-SP, Rondonópolis-MT and Unai-MG during 2001/2002 to 2003/2004 cotton growing seasons. The aphid collection was performed at the beginning, middle and end of cotton growing season. No significant differences in susceptibility to thiamethoxam and endosulfan were observed among A. gossypii populations. On the other hand, there was a significant decrease in the susceptibility to carbosulfan throughout the cotton growing season. However, a partial reset to the susceptibility to carbosulfan was observed at the beginning of the following cotton season. The resistance ratio of A. gossypii to carbosulfan was 5.7-fold. No cross-resistance between carbosulfan and the insecticides thiametoxam was observed. Studies on dynamics of A. gossypii resistance to carbosulfan conducted under laboratory conditions revealed that the resistance is unstable; that is, the frequency of resistance declines significantly in the absence of selection pressure. Life-history comparisons between carbosulfan-susceptible and resistant strains showed that there is some fitness cost associated to resistance. The resistant strain showed a lower fecundity, lower reproductive period and lower longevity than the susceptible strain. And finally, studies on evaluation of the strategy by mixing insecticides showed that the mixture of carbosulfan with thiamethoxam or endosulfan was not advantageous in the management of carbosulfan resistance in A. gossypii.

The overall purpose of this study was to examine the effects of an insecticide vapor pulse on the dispersal response of the German cockroach, Blattella germanica (L). An apparatus was designed to expose test cockroaches to vapors from commercial pesticide formulations. Insecticide vapor-induced dispersal responses were recorded during a two hour period from different strains that had been allowed to acclimate to a harborage. The dispersal response of large nymphs from a pesticide susceptible laboratory strain (VPI) was compared to two propoxur resistant field strains (Carver, Kenly) after exposure to four propoxur formulations and their blanks. Vapors from the petroleum-based oil and aerosol formulations induced significantly more dispersal than vapors from water-based emulsifiable concentrate and wettable powder formulations. Vapors from formulations containing the toxicant generally induced a significantly faster dispersal response than did their blanks. Exposure of cockroaches to the vapors of diazinon, malathion, and cyfluthrin indicated that the class of pesticide can also influence the dispersal response. Strain differences were found in experiments with the propoxur formulations, their blanks, malathion, and cyfluthrin. Slow dispersal precluded demonstration of significant strain differences in experiments with diazinon. Exposure of mixed age groups of four strains to vapors from 1% propoxur-in-oil, an aerosol and their blanks indicated that dispersal patterns were similar to the single age class experiments. Inter- and intra-strain differences were found. Dispersal of the field strains was distinguished from that of the laboratory strain by more variable responses and differences among age classes. The strongest inter-and intra-strain differences were in response to the solvents rather than the complete formulation. The air concentration of propoxur from a 1% oil formulation was estimated at 146 pg/ml. Condensation of the toxicant onto the apparatus was also observed. Vapor pulse characteristics for the four propoxur formulations indicated that the equilibrium vaporization rate in the test apparatus was approximately 0.5 mg/min for the oil and aerosol formulations and 1.0 mg/min for the WP and EC formulations. Results suggest that effective control strategies must be tailored for each target population.<br>Ph. D.

Includes bibliographical references (leaves 204-230). Investigates the behaviour and ecology of the two major larval parasitoids of the diamondback moth, Plutella xylostella; Cotesia plutellae and Diadegma semiclausum.

O carrapato do boi Rhipicephalus microplus é um ectoparasita de bovinos de grande importância sanitária e econômica, que produz grandes prejuízos na bovinocultura do Brasil. O uso de ivermectina como meio de controle químico, ao longo dos anos, levou à seleção de linhagens de R. microplus resistentes a ivermectina. A consequência desse processo é a perda da eficácia do acaricida. Este trabalho teve por objetivo determinar as bases moleculares da resistência a ivermectina. Para o estudo dos mecanismos envolvidos na resistência a ivermectina de R. microplus foram utilizadas duas cepas de referência: a cepa Mozo, suscetível a ivermectina, e a cepa Juarez, resistente, e duas abordagens experimentais complementares: ensaios enzimáticos com sinergistas e estudos de expressão gênica por meio de sequenciamento massivo em paralelo do RNA ou RNA-Seq. Os resultados deste trabalho permitem afirmar que o mecanismo de desintoxicação metabólica de maior importância é mediado por transportadores ABC, seguido de esterases, glutation-S-transferases e citocromo-oxidases. As diferenças de expressão gênica observadas entre as cepas, e dentro das cepas a diferentes concentrações de ivermectina, confirmam os resultados e permitem concluir que R. microplus apresenta uma resposta poligênica à ivermectina<br>The cattle tick Rhipicephalus microplus is one of the most important ectoparasites with great sanitary and economic importance for cattle rearing in Brazil. Ivermectin is a drug used in the chemical control of R. microplus. The utilization of ivermectin in the last 30 years has led to the selction of resistant populations of R. microplus, and thus, the loss of efficacy in the cattle tick control. To study the molecular basis of ivermectin resistance in R. microplus, two strains of ticks were used: the susceptible strain Mozo, and the resistant strain Juarez. Two complementary approaches were carried out: enzimatic assays using sinergists and gene expression studies using massive parallel sequencing of RNA or RNA-Seq. The results of this work indicate the involvement of various mechanisms of metabolic resistance. Detoxification mechanisms mediated by ABC transporters are probably the most important. Esterases, gluthathion-S-transferases and citochrome-oxidases play a less important role in detoxification. Differences in gene expression were observed between strains and among samples of the same strain exposed to different concentrations of ivermectin, confirming the result with the assays. Hence, the response of R. microplus to ivermectin is, probably, complex and poligenic

Global climate change promotes warming temperatures and altered salinities that pose threats to aquatic ecosystems and species, such as Hyalella azteca. Moreover, these threats to aquatic ecosystems are exacerbated by agricultural, urban, and industrial pesticide runoff. In the state of California in 2012, pyrethroid insecticides were the seventh most applied group by licensed professional applicators for pest control and landscape maintenance. Some species, specifically H. azteca have developed non-target resistance to pyrethroids in California. It is imperative to understand whether the bioenergetic cost of resistance makes H. azteca more susceptible to warming and salinity effects in the presence of contaminants. This research presents an assessment on how multiple stressors can affect the toxicity of permethrin (pyrethroid insecticide) on one Wild-type and two pyrethroid- resistant species of H. azteca, belonging to different clades. A series of 96-h acute toxicity tests exposing animals to a concentration range of permethrin were performed with compounding stress from temperatures (18, 23 and 28 °C) and salinities (0.2, 1.0, and 6.0 practical salinity units [PSU]). Findings indicate resistant H. azteca cultured in pyrethroid-free settings have maintained resistance to permethrin over time, whereas the wild-type population did not develop any resistance over the course of experimentation. For resistant H. azteca, changes in salinity and temperature both increased and decreased survival of H. azteca exposed to permethrin. Between the two resistant clades, not only was survival affected, but the average slope of the dose-response curve was significantly different (p < 0.05); clade D was more susceptible to pyrethroids when coping with warming and higher salinity than clade C., Differential susceptibility potentially indicates that distinct resistance mutations confer a difference in the potency and mode of toxic action. The results provide insight to how changes posed by climate change, coupled with pyrethroid pesticides could be detrimental to this species, and conversely, how in some scenarios, changes to temperature and salinity might actually benefit the survival of H. azteca. These findings further indicate the importance of considering global climate change effects into risk assessments of emerging and legacy use contaminants.

A crescente utilização do controle químico para o manejo de pragas na cultura dos citros tem afetado significativamente a dinâmica populacional de inimigos naturais como os ácaros predadores que são importantes no controle biológico de pragas como o ácaro-da-leprose Brevipalpus phoenicis (Geijskes). Dentro desse contexto, a integração dos controles químico e biológico, seja pela utilização de produtos seletivos ou linhagens de ácaros predadores resistentes a pesticidas, poderia resultar em uma forma de manejo racional de ácaros fitófagos. Sendo assim, a presente pesquisa foi desenvolvida para avaliar a suscetibilidade de diferentes espécies de ácaros fitoseídeos (Amblyseius chiapensis De Leon, Euseius concordis (Chant) e Iphiseiodes zuluagai Denamark & Muma) a dicofol e deltametrina, em populações coletadas em pomares de citros. Não houve variabilidade interespecífica na suscetibilidade ao dicofol. E também, não foi constatada variabilidade intraespecífica na suscetibilidade das populações de E. concordis testadas. Por outro lado, E. concordis apresentou-se aproximadamente 50 vezes mais tolerante do que I. zuluagai à deltametrina. Foi detectada variabilidade intraespecífica na suscetibilidade à deltametrina nas populações de E. concordis e I. zuluagai, sendo que a razão de resistência estimada foi maior do que 14 vezes para as populações de E. concordis e de 18 vezes para as populações de I. zuluagai testadas. E por último, verificou-se diferenças significativas quanto a respostas de repelência apresentadas pelas populações de E. concordis a resíduos de deltametrina.<br>The increase in the use of chemical control for managing pests of citrus has significantly affected the population dynamics of natural enemies such as predator mites that are important in the biological control of phytophagous mites such as Brevipalpus phoenicis (Geijskes). Within this context, the integration of chemical and biological control through the use of selective pesticides or strains of predator mites resistant to pesticides could be a more rational way to manage phytophagous mites. Thus, the objective of this research was to evaluate the susceptibility to dicofol and deltamethrin in populations of different phytoseiid species (Amblyseius chiapensis DeLeon, Euseius concordis (Chant) e Iphiseiodes zuluagai Denamark & Muma) collected in citrus groves. There was no interespecific variability in response to dicofol. And also, no intraespecific variability in the susceptibility to dicofol in E. concordis populations was detected in this study. On the other hand, E. concordis was 50 times more tolerant than I. zuluagai to deltamethrin. Intraespecific variability in the susceptibility to deltamethrin was detected for both E. concordis and I. zuluagai populations. A resistance ratio of > 14- fold and 18- fold was detected to deltamethrin in E. concordis and I. zuluagai populations, respectively. And finally, there was a significant difference in the repellency of E. concordis populations in deltamethrin residues.

Bibliography: leaves 120-145. Reports Bt crystal protein toxicity to a phthirapteran species. Although Bt strain WB3516 may produce other unidentified toxins effective against B. ovis, the results provide strong evidence that the [delta]-endotoxin crystal proteins of strain WB3516 significantly contribute to the lousicidal toxicity of this strain.

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.<br>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.

O gorgulho do milho, Sitophilus zeamais (Motsch., 1855) é a principal praga no armazenamento de milho. Seu controle é feito com a aplicação de inseticidas curativos e ou preventivos, mas o método apresenta inconvenientes relacionados com fatores toxicológicos e de resistência da praga devido ao pequeno número de inseticidas registrados e estes pertencerem a grupos químicos antigos, como os organofosforados e piretróides. Com o objetivo de testar um grupo químico diferente e mais moderno no controle desta praga e analisar a variação de umidade e atividade de água dos grãos de milho no decorrer do experimento, foi instalado este ensaio. Os grãos de milho foram pulverizados na proporção de 5 litros de calda por tonelada de grão, e depois mantidos em sacos de pano para manter os insetos em contato com os grãos e ao mesmo tempo, possibilitar a troca de ar. O ensaio foi instalado com sete tratamentos no início do experimento e mais quatro depois de dois meses, para testar novas doses e misturas do Etofenprox no controle do gorgulho. Os produtos utilizados para comparação com Etofenprox foram: Sumigran 500 (15 mL/ton. grão), Sumigran Plus (20 mL/ton. grão), K-Obiol (15 mL/ton. grão) e óleo de eucalipto (12,4 Lt/ton. grão), além da mistura do Etofenprox (20mL/ton. grão) com Butóxido de Piperonila (0,5%) e com Sumigran (10 mL/ton. grão). O Etofenprox não apresentou eficiência no controle de S. zeamais, porém sua mistura com Butóxido de Piperonila e Sumigran apresentaram eficiência superior aos tratamentos com Etofenprox, sugerindo um possível efeito sinérgico e uma possível alternativa para o controle da resistência, uma vez que o Etofenprox pertence a um grupo químico diferente dos registrados atualmente, sendo muito menos tóxico. O efeito sinergista do Butóxido de Piperonila já é conhecido e foi testado o possível efeito sinérgico do Etofenprox com o Fenitrotion. O teste foi instalado sobre placas de concreto que tiveram metade de suas superfícies aplicadas com doses de Fenitrotion (0,8 mL/m² ; 0,6 mL/m²; 0,4 mL/m²; 0,2 mL/m²; 0,1 mL/m² e 0,05 mL/m²) e a outra metade com as mesmas doses de Fenitrotion adicionadas ao Etofenprox (1,25 mL/m²). Outra observação foi que o óleo de eucalipto, mesmo não sendo eficiente no controle de S. zeamais, teve eficiência superior à testemunha e inibiu a oviposição. Observou-se que o dano dos insetos alterou a umidade, uma vez que os tratamentos que tinham insetos vivos apresentaram sempre umidade superior em relação aos tratamentos eficientes no controle desta praga. A maior umidade dos grãos nos quatro tratamentos instalados posteriormente fez com que o tempo de controle fosse menor. Esse experimento conclui que: Etofenprox não é eficiente no controle de S. zeamais, nas doses testadas; Etofenprox não apresenta efeito sinérgico com Fenitrotion no controle de S. zeamais; o óleo de eucalipto obteve controle superior à testemunha e inibiu a oviposição; o dano do inseto afeta diretamente a umidade da massa de grãos; a umidade dos grãos reduz o tempo de controle dos produtos utilizados.<br>The maize weevil Sitophilus zeamais (Motsch., 1855) is the main pest found in maize grains storage. The control of this pest is done with the application of curative or preventive insecticides, but these methods have brought unwanted effects concerning toxicological factors in addition to pest resistance factors, once there was only a small number of pesticides registered and they belong to old chemical groups, such as Organophosphorus and Pyrethroids. Due to reasons listed above, the present experiment was carried out with the aim of testing alternative and modern chemical groups that could be able to control the pest and also, of evaluating the moisture content variation and water activity of maize grains during the period of the experiment. The maize grains were sprayed within the proportion of 5 litres of mixture per ton of grains, and after that, they were stored in cloth bad in order to keep the insects in touch with the grains, and at the same time, allowing aeration. Seven treatments were applied at the beginning of the experiment and other four treatments were applied two months after that, with the intention of testing new rates and mixtures of Etofenprox to control the maize weevil. Some insecticides were used to be compared with Etofenprox, as it follows: Sumigran 500 (15 mL/ton of grains), Sumigran Plus (20 mL/ton of grains), K-Obiol (15 mL/ton of grains) and eucalypt oil (12,4 Lt/ton of grains), as well as the mixture of Etofenprox (20 mL/ton of grains) with Piperonyl Butoxide (0,5%) and Sumigran (10 mL/ton of grains). Etofenprox has not shown any efficiency in controlling the S. zeamais, on the other hand, its mixture with Piperonyl Butoxide and Sumigran has shown higher levels of efficiency than the treatments with pure Etofenprox, which suggests a possible synergistic effect and a possible alternative for the control of pest resistance, once Etofenprox belongs to a chemical group which is different from the ones currently registered and is less toxic than they are. The synergistic effect of Piperonyl Butoxide is already known and the possibility of a synergistic effect of Etofenprox with Fenitrotion was also tested. This test was installed on concrete slabs which had rates of Fenitrotion (0,8 mL/m² ; 0,6 mL/m²; 0,4 mL/m²; 0,2 mL/m²; 0,1 mL/m² and 0,05 mL/m²) applied over half of their surfaces while the other half received applications of the same rate of Fenitrotion added to Etofenprox (1,25 mL/m²). Besides that, it was also relevantly observed that although eucalypt oil not being efficient enough to control S. zeamais, it is more efficient than the untreated and also prevented oviposition. It was observed that the attack of insects affected the moisture, once the treatments containing alive insects have always shown moisture rate higher than the efficiently controlling treatments. The higher grain moisture rate of the four last treatments caused the control time to be shorter. After this experiment, it is possible to conclude that: the Etofenprox it´s not efficient in the control of Sitophilus zeamais, on the tested rates; the Etofenprox it´s not present any synergistic effect with Fenitrotion; eucalypt oil was more controlling than the untreated and prevented oviposition; the attack of insects directly affects moisture content of grains; the moisture rate of grains interfered in the control time of the products used.

Apesar da espécie Vigna unguiculata (L.) Walp. ser bastante rústica e estar adaptada às condições adversas de clima e solo brasileiros, seu rendimento é muito baixo. Diversas causas têm sido levantadas para explicar esse comportamento; entre elas destacam-se as doenças fúngicas, sobretudo aquelas cujos patógenos são transmitidos por sementes, em especial a podridão cinzenta do caule, causada por Macrophomina phaseolina (Tassi) Goid. A abordagem analítica desse patossistema revelou alguns problemas emergentes. Entre eles, destacam-se: a) desconhecimento da qualidade sanitária das sementes de caupi, utilizadas para semeadura; b) desunifomidade na metodologia usada para detectar os patógenos presentes nas semente; c) dificuldade na esporulação do patógeno, máxime de alguns isolados reticentes em esporular em meios artificiais de cultivo, cujo comportamento dificulta os trabalhos de seleção de genótipos resistentes; d) carência de medidas de controle do patógeno, que empreguem práticas naturais, como uso de sementes sadias, de indutores de resistência e de cultivares resistentes, de fácil uso e passível de adoção por parte dos produtores. Na estruturação da matriz lógica do presente estudo, referidos problemas foram transformados em objetivos. Os trabalhos foram conduzidos no Departamento de Entomologia, Fitopatologia e Zoologia Agrícola da ESALQ/USP, em Piracicaba-SP. Os resultados indicaram o teste de sanidade de sementes de caupi empregando o método do papel de filtro com restrição hídrica utilizando NaCl a –0,8Mpa, como o mais adequado para detecção dos fungos presentes nas sementes de caupi, especialmente M. phaseolina. A análise sanitária das amostras de sementes originadas de vários estados brasileiros revelou que, em 62% das amostras analisadas, o fungo M. phaseolina estava presente, sendo as amostras originadas do estado da Paraíba, Piauí, Pará e Bahia as que apresentaram maiores níveis de incidência do patógeno. Os melhores níveis de esporulação do patógeno foram conseguidos com a combinação de sobreposição de discos de folhas de trigo ao meio BDA, com temperatura de 25oC. Quanto à identificação de indutores de resistência, capazes de controlar M. phaseolina, os resultados revelaram que o acibenzolar-S-metil (ASM) foi o mais eficiente, quando comparado com quitosana e com um produto silicatado derivado de rocha micronizada (PSiM), apresentando um controle residual por mais de 40 dias após a semeadura. A maior eficiência verificada pelo ASM ocorreu devido a sua capacidade de ativar mecanismos bioquímicos de defesa, configurando-se em efetivo ativador da resistência induzida nas plantas de caupi, por atuar na cinética de importantes enzimas relacionadas à defesa, como a fenilalanina amônia-liase, peroxidase e quitinase. Quanto à reação de cultivares de caupi à doença, foi possível verificar razoável nível de resistência de algumas cultivares, tendo sido consideradas resistentes Mulato, Guariba e Maratauã.<br>Notwithstanding the specie Vigna unguiculata (L.) Walp is sufficiently rustic and adapted to the adverse conditions of the Brazilian soil and climate, its improvement is very low. Many causes have been raised in order to explain such behavior; among them the fungal diseases stand out, over all those whose pathogens are transmitted by the seeds especially the charcoal rot disease caused by Macrophomina phaseolina (Tassi) Goid. The analytical approach of such pathosystem has revealed some emerging problems. Among them, it stands out: a) the ignorance of the sanitary quality of the cowpea seeds used for sowing; b) the non-uniformity in the used methodology in order to detect the pathogens, which are present in the seed; c) the difficult in pathogen sporulation, principally of some isolated reticent in forming spores in cultivation artificial environments whose behavior hampers the selection works of the resistant genotypes; d) lack of pathogen control measures, which utilize natural practices, such as the use of healthy seeds, resistance inducers and resistant cultivars of easy utilization and liable to adoption by the producers. In structuring the logical matrix of this study, such problems were transformed into objectives. The works were conducted at the Entomology, Phytopathology and Agricultural Zoology Departments of ESALQ/USP, in Piracicaba-SP. The results have pointed out the sanity test of the cowpea seeds through the method of filter paper with hydric restriction using NaCI – 0,8Mpa, as the most suitable for detecting the current fungus in cowpea seeds, especially M. phaseolina. The sanitary analysis of the seeds samples originated from several Brazilian states has revealed that in 63% of the analyzed samples, the fungus M. phaseolina was present, and the samples originated from the states of Paraíba, Piauí, Pará and Bahia were those that have presented higher incident levels of pathogen. The best levels of sporulation were obtained with the combination of the superposition of wheat leaves disks in the middle of BDA in 25ºC. As to the identification of the resistance inducers, capable of controlling the M. phaseolina, the results have revealed that the acinbezolar-S-methyl (ASM) was more efficient when compared to chitosan and with a silicate product originated from micronized rock (PsiM), presenting a residual control for more than 40 days after the sowing. The greatest efficiency ascertained by ASM has occurred due to its capacity of activate the defense biochemistries mechanisms, forming itself in an activator effect of the induced resistance in cowpea plants because it acts in the kinetic of important enzymes related to the defense, such as the phenylalanine ammonia-lyase, peroxidase and chitinase. As to the cowpea cultivars reaction to the disease, it was possible to ascertain a reasonable resistance level of some cultivars, and BR 14 Mulato, Guariba e Maratauã were considered as resistant.

Les perturbations environnementales d'origine anthropique favorisent l'établissement de populations invasives. La gestion de ces populations est primordiale pour la santé publique (zoonose, famine), l'environnent (perte de biodiversité), et l'économie (dégâts). L'objectif de cette thèse pluridisciplinaire, menée en conditions naturelles, est d'améliorer les connaissances sur les populations invasives de rongeurs dans des paysages agricoles à grande échelle et d'appréhender les mécanismes d'adaptation qui favorisent une réponse positive aux pressions anthropiques. Les résultats montrent que (1) le rat endémique Rattus tiomanicus, dont la présence est associée à la typologie de l'habitat naturel, et le rat introduit Rattus tanezumi-R3, dont la présence est associée aux activités humaines, constituent les populations invasives des plantations de palmier à huile en Indonésie; (2) leur distribution géographique clinale est probablement contemporaine à l'anthropisation des milieux, et suppose une compétition inter-spécifique; (3) ces grandes populations sont spatialement continues avec un flux génique limité par la distance géographique (caractérisées par un patron d'isolement par la distance) et potentiellement influencé par les transports routiers; (4) R. tanezumi-R3 possède une forte résistance physiologique aux raticides AVK, dont l'origine n'est pas associée à une mutation génétique de la molécule cible mais probablement liée aux enzymes du métabolisme. Ces travaux soulignent des stratégies d'adaptations comportementales et physiologiques des populations invasives de rongeurs en milieux agricoles et procurent des bases pour l'élaboration de stratégies de lutte adaptée

Gray mold of strawberry, caused by Botrytis cinerea, is a very destructive pre- and post-harvest fruit rot. Outside of California, fungicide resistance in B. cinerea has been reported to every site-specific chemical class labeled for use against gray mold. One objective of this study was to characterize the resistance of 888 isolates of B. cinerea from California strawberry fields to ten active ingredients. Isolates were collected from the same planting block in 47 fields during the early-season (0 to 8 fungicide applications) and late-season (16 to 26 fungicide applications) of 2016. Sensitivity of each isolate was determined using the following active ingredients at a discriminatory dosage (μg/ml): boscalid (75), cyprodinil (4), fenhexamid (50), fludioxonil (0.5), fluopyram (10), iprodione (10), isofetamid (5), penthiopyrad (5), pyraclostrobin (10), and thiophanate-methyl (100). Resistance to each active ingredient was observed at varied frequencies (early-season %, late-season %): boscalid (12, 35), cyprodinil (12, 46), fenhexamid (53, 91), fludioxonil (1, 4), fluopyram (2, 7), iprodione (25, 8), isofetamid (0, 1), penthiopyrad (8, 25), pyraclostrobin (77, 98), and thiophanate-methyl (81, 96). Captan, boscalid, cyprodinil, fenhexamid, and fludioxonil were the most commonly used fungicides in surveyed strawberry fields. A selection of 100 isolates was identified to the species level. All isolates were B. cinerea, excluding one isolate of Botrytis mali. A fungicide resistance trial was conducted v to observe resistance responses in populations of B. cinerea. Frequencies of resistance to boscalid and fludioxonil remained unchanged despite consecutive applications of these fungicides. Frequency of resistance to fenhexamid increased when this fungicide was applied and decreased when it was not. This occurred in fungicide treatments including fungicide rotation, tank-mixing with captan, and consecutive applications of fenhexamid. Multi-fungicide resistance was widespread in California strawberries; isolates resistant to fenhexamid, thiophanate-methyl and pyraclostrobin were the most common phenotype. The frequency of resistance increased from the early-season to late-season for multiple active ingredients tested. This within-season change in frequency of resistance was tested and confirmed in a field trial, where common resistance management strategies failed to prevent the buildup of fenhexamid resistance. New and improved methods of resistance management may need to be enacted to ensure the future efficacy of site-specific fungicides.

Pour obtenir des plantes resistantes aux inhibiteurs de l'hydroxyphenylpyruvate dioxygenase (hppd), nous avons souhaite construire une voie metabolique contournant l'hppd en introduisant les genes codant l'hydroxyphenylpyruvate oxydase (hppo) et la 4-hydroxyphenylacetate 1-hydroxylase. Nous decrivons ici l'identification de trois genes bacteriens codant ces deux activites enzymatiques. - le gene hppo, identifie chez arthrobacter globiformis, code l'hppo (e. C. 1. 2. 3. -) qui catalyse l'oxydation du 4-hydroxyphenylpyruvate en 4-hydroxyphenylacetate (4-hpa). C'est la premiere fois que la proteine est purifiee et le gene identifie. - les gene hpah et hpac, identifies chez p. Acidovorans, codent respectivement les proteines hpah et hpac. La combinaison de ces deux proteines produit l'activite enzymatique 4-hydroxyphenylacetate 1-hydroxylase (e. C. 1. 14. 13. 18). L'hpah convertirait le 4-hpa en un metabolite non encore identifie. Ce metabolite serait ensuite convertit en homogentisate par l'hpac. Enfin, nous avons acquis des informations sur le metabolisme de la tyrosine par des cellules vegetales en utilisant la rmn du carbone-13. Nous avons mis en evidence un transporteur de tyrosine localise sur le plasmalemme des cellules d'acer en culture. C'est la premiere fois qu'un transporteur a haute affinite (15 m ; ph 6,2) presentant un ph optimum alcalin (ph 7,0 a 7,8) est caracterise. Nous avons aussi demontre que la tyrosine s'accumule dans la vacuole, apres une phase de latence. Le metabolisme de la tyrosine est etudie dans les cellules d'acer et d'arabidopsis. Le 4-hpa semble identifie comme provenant de la tyrosine seulement dans les cellules d'arabidopsis. Au vue des resultats obtenus, l'analyse par 1 3c-rmn devrait pouvoir etre utilisee pour evaluer les plantes transgeniques obtenues apres introduction des trois genes.

Laboratory, field and semi-field experiments have been carried out on the predatory mites Kampimodromus aberrans, Amblyseius andersoni, Typhlodromus pyri and Phytoseius finitimus in order to optimize biological control strategies against phytophagous mites in apple orchards. The first experiment concerned the augmentative releases of K. aberrans in organic and conventional apple orchards. The most frequent insecticides used in organic orchards were pyrethrins and spinosad, whereas neonicotinoids, OPs and IGRs were used in conventional orchards. K. aberrans releases were successful and predatory mites were significantly higher in released than in control plots. Moreover, populations were high in organic orchards compared to conventional orchards. Amblyseius andersoni was naturally occurring in all orchards. In 2010, no effect of K. aberrans release and orchard management were observed on A. andersoni populations but in 2011 A. andersoni population densities were low in released plots compared to control plots. In organic orchards located at Spresiano releases were made on two cultivars, i.e., Florina and Golden Delicious. In both 2010 and 2011 releases, K. aberrans population densities were higher in release than in control plots of both cultivars. K. aberrans population increased during the three years on Florina but not in Golden Delicious orchard. On the latter A. andersoni population densities were lower in released than in control plots. In conventional orchards K. aberrans did not establish probably because of a series of non-selective insecticide and fungicide treatments. However, the use of spinosad was associated to a decline of predatory mite populations in organic orchards. The second experiment considered interspecific predation among K. aberrans, A. andersoni, T. pyri and P. finitimus. All four predators fed with heterospecific larvae were able to survive some weeks laying a number of eggs. A. andersoni consumed more T. pyri and P. finitimus than K. aberrans larvae and its fecundity was higher on the first two prey species. A diet based on T. pyri larvae was associated to the highest conversion rate of food into egg biomass; longevity was higher for predatory mite females fed with K. aberrans larvae. K. aberrans preyed more P. finitimus than T. pyri and A. andersoni larvae and its fecundity resulted higher on the first species. The conversion of food into egg biomass did not show precise trend while the longevity of K. aberrans resulted higher when fed with A. andersoni. Typhlodromus pyri consumed more P. finitimus and K. aberrans than A. andersoni larvae and laid more eggs on the first two species. Phytoseius finitimus preyed more K. aberrans and T. pyri than A. andersoni larvae but fecundity was not affected by prey; predatory mite females fed with A. andersoni larvae were more efficient in converting food into egg biomass but those fed with K. aberrans larvae lived longer. In terms of predation rate and fecundity, A. andersoni seems to be advantaged over T. pyri, T. pyri over K. aberrans, and A. andersoni, K. aberrans and T. pyri over P. finitimus. The performance of A. andersoni, K. aberrans and T. pyri in terms of predation rate and fecundity proved to be better of P. finitimus larvae than on other prey. The low prey consumption and fecundity of the latter suggest that is disadvantaged in interspecific predation. The comparison between pollen and prey diets confirmed the positive effect of pollen on the fecundity of all four predatory mite species. The third experiment deals with the effects of insecticides, frequently applied in apple orchards, on K. aberrans in field and laboratory conditions. Insecticides including chlorpyrifos, thiacloprid, acetamiprid, lufenuron, indoxacarb, methoxyfenozide and etofenprox were tested. In field studies etofenprox was the most detrimental pesticide to predatory mites. It caused dramatic effects on K. aberrans populations and induced spider mite outbreaks. In other trials tau-fluvalinate and spinosad did similar effects. In laboratory studies we assessed lethal and sub-lethal effects for each pesticide tested. Spinosad, tau-fluvalinate, thiamethoxam, clothianidin and imidacloprid were included. Etofenprox, spinosad with tau-fluvalinate proved to be harmful insecticides to K. aberrans in terms of lethal effects. Neonicotinoids did not affect survival so much but their sub-lethal effects were more prominent in terms of fecundity. Other pesticides such as chlorpyrifos, indoxacarb, lufenuron and methoxyfenozide also significantly reduced fecundity in K. aberrans at a lower level. In the fourth study we explored the role of pollen in alleviating the effects of pesticides on K. aberrans at individual and population levels. In the first part we tested biological and demographic parameters of K. aberrans on pollen and Panonychus ulmi. K. aberrans was reared individually on P. ulmi and on pollen. Both food sources allowed for predatory mites survival, development and reproduction. Pollen was preferred food as compared to P. ulmi. Total fecundity and survival rates of K. aberrans were higher on pollen than on tetranychids. Developmental times of protonymphs, deutonymphs and adults of K. aberrans were longer on P. ulmi than on pollen. Life-table parameters of K. aberrans were positively affected by pollen. The second part deals with the interactions of pesticides, pollen and K. aberrans in semi-field conditions. Experimental factors were: insecticide application and pollen application . Predatory mites were released on 3-4 shoots per plant 7 days before insecticide applications. Typha latifolia pollen was used. Insecticide application, particularly spinosad, determined a reduction in K. aberrans abundance. Pollen applications were associated to an increase in predatory mite population. Significant interaction between pollen applications was observed: pollen application mitigated the effect of chlorpyrifos (because of its compensation of sublethal effects on fecundity) but no effect was observed on spinosad treated plants. In the last work mechanisms of resistance to chlorpyrifos due to mutations in insensitive Acetylcholinesterase (AChE) in K. aberrans was investigated. AChE cDNA was cloned. The cloning strategy relied on the AChE sequence of Tetranychus urticae. To obtain the homologous cDNA sequence in K. aberrans, degenerate primers were designed on conserved functional domains of the annotated AChEs in Metaseiulus occidentalis genome, and combined with 5’-3’ rapid amplification of cDNA ends. When the cloned AChE sequence was compared between K. aberrans sensible and resistant strains, only a Glycine to Serine (G119S) substitution was detected while both AChE sequences had a Phenylalanine residue at 331 position. These results excluded the F331W mutation as responsible for chlorpyrifos target site resistance in K. aberrans but it opened interrogatives about the role of G119S. Indeed the G119S substitution had been already described in mosquitoes Anopheles gambiae and Culex pipiens where it led to AChE1 insensitivity or reduced AChE1 activity. On the contrary, the same aminoacid change in T. urticae AChE sequence was associated with a more moderate decrease in chlorpyrifos response. So a species-specific effect of this substitution cannot be kept out. In any case, site oriented sequencing of AChE cDNA, confirmed the presence of G119S substitution in further three resistant strains of the mite predator as well as the absence of F331W mutation. At the same time, two additional sensible strains, displayed a Glycine at the position 119, instead of Serine, in their AChE sequence. Altogether, these findings might suggest that the G119S polymorphism was involved in the polygenic control of the resistant phenotype. In addition or alternatively, the mutated AChE might be in linkage with one of the genetic determinants which affect the insecticide sensitivity in K. aberrans.<br>Nell’ambito di questo lavoro sono stati condotti esperimenti di campo, semi-campo e laboratorio sugli acari predatori Kampimodromus aberrans, Amblyseius andersoni, Typhlodromus pyri e Phytoseius finitimus allo scopo di perfezionare le strategie di controllo biologico degli acari fitofagi nei meleti. Il primo esperimento ha riguardato l’introduzione di K. aberrans in meleti convenzionali e biologici. I meleti biologici erano trattati soprattutto con piretro e spinosad, mentre i meleti convenzionali erano trattati con neonicotinoidi, esteri fosforici e regolatori della crescita. Le densità dei predatori hanno raggiunto densità superiori nelle tesi oggetto d’introduzione rispetto ai testimoni. Tuttavia, le popolazioni dei predatori hanno raggiunto livelli più elevati nei meleti biologici rispetto ai meleti convenzionali. Amblyseius andersoni risultava naturalmente presente nei meleti. Nel 2010, non sono stati riscontrati effetti delle introduzioni di K. aberrans né della gestione dei meleti sull’abbondanza di A. andersoni. Al contrario, nel 2011 sono state riscontrate densità più contenute di A. andersoni nelle tesi oggetto d’introduzione rispetto ai testimoni. In uno dei siti sperimentali (Spresiano), le popolazioni di K. aberrans sono risultate più elevate, ove introdotte, in due meleti di cultivar Florina e Golden Delicious. Le densità di K. aberrans sono aumentate nel corso del tempo ma in misura più rilevante su Florina, probabilmente per la morfologia fogliare di questa cultivar. Nel meleto di Golden Delicious, le popolazioni di A. andersoni sono risultate più contenute nella tesi oggetto d’introduzione di K. aberrans rispetto ai testimoni; probabilmente, questo risultato è stato influenzato dalla morfologia fogliare di questa cultivar. Nei meleti convenzionali, le introduzioni di K. aberrans sono fallite a causa di trattamenti ripetuti con prodotti fitosanitari non selettivi ma anche gli insetticidi impiegati nei meleti biologici hanno comportato decrementi significativi delle densità dei predatori. La competizione tra K. aberrans, A. andersoni, T. pyri e P. finitimus è stata oggetto di un secondo esperimento. Le femmine dei predatori sono state allevate sulle larve delle altre specie allo scopo di simulare possibili interazioni in condizioni di scarsità di preda. In queste condizioni sperimentali, i predatori hanno esibito buoni tassi di sopravvivenza e di fecondità. A. andersoni ha predato più larve di T. pyri e P. finitimus che di K. aberrans e la sua fecondità è stata superiore sulle prime due prede. Per A. andersoni, una dieta basata sulle larve di T. pyri è stata associata a tassi più elevati di conversione delle prede in biomassa destinata alle uova; la longevità del predatore è risultata più elevata sulle larve di K. aberrans. K. aberrans ha predato più P. finitimus che T. pyri e A. andersoni e la sua fecondità è risultata più elevata sulla prima specie. Il tasso di conversione in uova non ha mostrato tendenze particolari mentre la longevità di K. aberrans è risultata più elevata quando i predatori sono stati alimentati con larve di A. andersoni. Typhlodromus pyri ha consumato più larve di P. finitimus che di A. andersoni e il tasso di ovideposizione è stato conseguente. Phytoseius finitimus ha predato più larve di K. aberrans e T. pyri che di A. andersoni ma la fecondità non ha mostrato la stessa tendenza: i predatori allevati su A. andersoni sono stati più efficienti nella conversione delle prede in uova. Relativamente ai tassi di predazione e di fecondità, A. andersoni sembra avvantaggiato rispetto a T. pyri, T. pyri rispetto a K. aberrans, mentre A. andersoni, K. aberrans e T. pyri sono avvantaggiati rispetto a P. finitimus. Le prestazioni di A. andersoni, K. aberrans e T. pyri (tassi di predazione e fecondità) sono state migliori quando i predatori sono stati allevati con larve di P. finitimus. Le prestazioni di P. finitimus sono apparse le più basse in assoluto e questo suggerisce uno svantaggio competitivo. L’alimentazione pollinica ha garantito prestazioni ottimali in termine di fecondità a sostegno del ruolo del polline nel garantire la persistenza dei predatori generalisti negli ecosistemi agrari. Il terzo esperimento ha riguardato l’effetto degli insetticidi su K. aberrans in condizioni di pieno campo e di laboratorio. E’ stato saggiato l’effetto degli insetticidi chlorpyrifos, thiacloprid, acetamiprid, lufenuron, indoxacarb, methoxyfenozide e etofenprox. Nelle prove di pieno campo, etofenprox è risultato l’insetticida più dannoso nei confronti degli acari predatori ed ha causato pullulazioni di acari fitofagi. In ulteriori prove di pieno campo, spinosad e tau-fluvalinate hanno causato problemi analoghi. Nelle prove di laboratorio sono stati inseriti anche spinosad, tau-fluvalinate, thiamethoxam, clothianidin e imidacloprid. Spinosad, tau-fluvalinate e etofenprox hanno causato effetti drammatici sulla sopravvivenza e sulla fecondità di K. aberrans. I neonicotinoidi non sono apparsi molto tossici ma hanno causato una riduzione della fecondità di K. aberrans. Per altri insetticidi (chlorpyrifos, indoxacarb, lufenuron e methoxyfenozide) sono state osservate riduzioni meno drammatiche della fecondità del predatore. In un altro studio è stato valutato il ruolo del polline nel mitigare gli effetti degli insetticidi su K. aberrans. Nella prima parte di questo studio sono stati calcolati i parametri demografici di K. aberrans allevato su polline o su Panonychus ulmi. Il polline si è dimostrato un alimento ottimale rispetto a P. ulmi in quanto i predatori hanno esibito una fecondità più elevata e tempi di sviluppo più brevi con implicazioni per i parametri demografici. La seconda parte dell’esperimento ha riguardato le interazioni tra pesticidi, polline e K. aberrans in condizioni di semi-campo. Gli acari predatori sono stati introdotti 7 giorni prima dei trattamenti insetticidi e il polline di Typha latifolia è stato impiegato quale alimento. L’applicazione degli insetticidi (soprattutto spinosad) ha ridotto la densità della popolazione di K. aberrans. L’applicazione ripetuta del polline è stata associata a un incremento della popolazione dei predatori. E’ stata riscontrata un’interazione significativa tra l’applicazione di polline e degli insetticidi: l’applicazione di polline ha mitigato l’effetto del chlorpyrifos sulla fecondità di K. aberrans. Tuttavia, questo effetto non è stato osservato sulle piante trattate con spinosad. I meccanismi di resistenza al chlorpyrifos in K. aberrans sono stati oggetto di un ultimo contributo. In particolare, è stata indagata la resistenza associata a mutazioni nell’enzima Acetilcolinesterasi (AChE). Il cDNA dell’enzima è stato clonato mediante una strategia basata sul sequenziamento della AChE di Tetranychus urticae. Al fine di ottenere sequenze omologhe di cDNA in K. aberrans sono stati disegnati primers con domini funzionali delle AChE nel genoma dell’acaro predatore Metaseiulus occidentalis. Combinando le sequenze AChE di ceppi sensibili e resistenti di K. aberrans è stata posta in evidenza una sostituzione di glicina in serina (G119S) mentre entrambe le sequenze AChE avevano un residuo di fenilalanina in posizione 331. Questi risultati hanno escluso la mutazione F331W quale responsabile della resistenza target size al chlorpyriphos ma apre interrogativi sul ruolo della mutazione G119S. Questa sostituzione è già stata descritta in Anopheles gambiae e Culex pipiens ove è coinvolta in una AChE1 insensibile o a ridotta attività. Tale sostituzione è stata associata ad una moderata resistenza al chlorpyrifos in T. urticae. Pertanto, sembra che tale sostituzione abbia un ruolo specie-specifico. Le analisi hanno dimostrato la presenza della sostituzione G119S in tre ceppi resistenti di K. aberrans in cui è assente la mutazione F331W. Allo stesso tempo, due ulteriori ceppi sensibili a chlorpyrifos hanno esibito una glicina in posizione 119, in luogo della serina, nella sequenza AChE. I dati suggeriscono che il polimorfismo G119S sia coinvolto nel controllo poligenico del fenotipo resistente. Le mutazioni della AChE potrebbero essere collegate con uno o più fattori genetici determinanti la resistenza a chlorpyrifos in K. aberrans.

Leptinotarsa decemlineata (Say) is the most important defoliating pest of potato Solanum tuberosum L., in North America and Europe.  Management of this pest relies heavily on chemical control and insecticide resistance is a persistent problem.  This phenomenon has increased the need for developing novel insecticides, resistance evaluation, and the development of alternative control strategies regarding this insect pest.  From 2010 to 2013, field and lab experiments were conducted to evaluate the efficacy of a novel insecticide tolfenpyrad on L. decemlineata.  In leaf-dip assays, tolfenpyrad was highly toxic to L. decemlineata with LC50 values of 0.013 and 0.164 g ai/L for larvae and adults, respectively.  Tolfenpyrad was also toxic to eggs with 0% hatching after being dipped in a field rate concentration.  In field efficacy trials, potato plots treated with tolfenpyrad at rates as low as 153 g ai/ha effectively controlled L. decemlineata.  <br />    In 2012, populations of L. decemlineata were collected from the Eastern Shore of VA and subjected to toxicity assays to determine current susceptibility to permethrin and oxamyl.  The toxicity assays indicated an increase in toxicity to permethrin in L. decemlineata larvae (LC50 = 3.931 g ai/L) and an increase in toxicity to oxamyl in adult beetles (LC50 = 9.695 g ai/L) compared with LC50 values previously reported in 1990.  In 2012, populations of L. decemlineata from Cheriton, VA, New Church, VA, Painter, VA, and Plymouth, NC were also evaluated for enzyme activity after exposure to sub-lethal concentrations of permethrin, oxamyl, and tolfenpyrad.  Adult beetles were subjected to enzyme assays to measure the activity of cytochrome P450 mono-oxygenase (P450), glutathione-S-transferase (GST), general esterases, and protein content.  Results from the enzyme assays indicated significantly greater esterase activity in beetles from Painter, VA exposed to permethrin [±-naphthol (F= 11.66, df= 4, 20, P<0.0001) and "-naphthol (F= 11.86, df= 4, 20, P<0.0001)], oxamyl [±- naphthol (F= 10.64, df= 4, 20, P<0.0001) and "-naphthol (F= 6.94, df= 4, 20, P=0.0011)], tolfenpyrad [±- naphthol (F= 407.62, df= 1, 8, P<0.0001) and "- naphthol (F= 28.15, df= 1, 8, P= 0.0007)], and the untreated control [±- naphthol (F= 28.14, df= 3, 16, P<0.0001) and "- naphthol (F= 28.86; df= 3, 16, P<0.0001)] compared to most of the other populations tested.  GST activity was significantly greater in tolfenpyrad exposed beetles compared to the non-treated beetles from Painter VA (F= 17.66, df= 5, 24, P< 0.0001).  <br />    Through laboratory assays and field experiments in potato, the efficacy of a new bio-pesticide derived from the bacterium Chromobacterium subtsugae was evaluated for the control of L. decemlineata.  Results from the laboratory assays showed L. decemlineata feeding was inhibited by the bio-pesticide derived from C. subtsugae.  However, field efficacy trials in 2010, 2011, and 2012, indicated no control of L. decemlineata.<br />    Methyl salicylate is an organic compound produced by potato and other plants in response to insect herbivory.  Abundance of predatory arthropods and L. decemlineata life stages were measured in plots treated with and without 5 g slow-release packets of methyl salicylate (95% methyl salicylate (Predalure")).  Methyl salicylate treatment had no impact on predator recruitment or cumulative mortality of L. decemlineata in potatoes.<br />    This research has provided us with a new tool for L. decemlineata management, as well as more information about resistance trends and alternative control strategies from which we can build on to reduce resistance development in L. decemlineata and ultimately formulate a stronger integrated pest management strategy for this insect pest.<br /><br>Ph. D.

Kiwifruit is a commercial crop threatened by attack of its most dangerous pathogen, Pseudomonas syringae pv. actinidiae (PSA). Currently, the European Economic Community has engaged in a policy of replacing chemical pesticides with natural substances. Following the European trend, this study is dealing with the prevention of disease outbreaks by growing plants able to better resist pathogen infection through the use of chitosan, a biodegradable, well-known elicitor of Systemic Acquired Resistance (SAR). The aim of this work was to improve the quality of Actinidia plants by a sustainable approach and reduce costs for the containment of phytopathological disorders. It was a 3-years project. In the first year, in vitro cultures of Actinidia deliciosa (A. Chev) C.F. Liang & A.R. Ferguson were chosen as model system to investigate the mechanisms of interaction between chitosan (elicitor) and the host plant. In order to understand the response to elicitation in planta, some effects of chitosan on the detoxification of reactive oxygen species (by guaiacol peroxidase [G-POD] and ascorbate peroxidase [APX]) and on the metabolism of phenolic compounds (by phenylalanine ammonia lyase [PAL] and polyphenol oxidase [PPO]) were monitored. Effectively, chitosan has been able to increase activity of the selected SAR biochemical markers in multiplication and rooting stages. In the second year, to study chitosan-host plant interaction in the presence of a pathogen, 2 year old plants belonging to two different species (Actinidia chinensis Planch. and A. deliciosa) were artificially inoculated with PSA in greenhouse experiments. The symptoms and SAR’s onset were investigated using morphological and biochemical analyses. Chitosan had the ability to enhance several activities of enzymes involved in detoxification processes (G-POD and APX) and in increasing plant defence barriers (PAL and PPO). Furthermore, plant’s defence responses were studied at molecular level. After a screening of possible SAR markers, Pathogenesis-Related proteins (PR1 and PR5) were chosen. Chitosan’s elicitation effect was determined by qRT-PCR and compared to the action of the most common SAR elicitors, such as salicylic acid, methyl jasmonate and the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC). As shown by the changes in PR transcription profiles, chitosan elicited a systemic response, with intensity comparable to other well-known signalling molecules. During the third year the validation of previously obtained results was undertaken. The changes in PR transcription profiles were corroborated in 2 year old plants, confirming that chitosan also elicits a systemic response in kiwifruit plants. In addition, a field trial with a natural presence of the pathogen was carried out and the kiwi plant response was determined by molecular techniques. If field trials will confirm the efficacy of chitosan in improving the behaviour of Actinidia plants under pathogen attack, further evidence will be added to support consideration of chitosan as sustainable and eco-friendly alternative for the reduction of pesticide use. In practice, a controlled induction of SAR with chitosan may be useful to alternate and integrate chemical treatments in a modern management of crop protection based on integrated control programs, including the use of environmentally safe products.

Chez les bactéries à Gram-négatif, deux systèmes très bien conservés et essentiels à la survie de la cellule bactérienne ont été identifiés : les systèmes Tol et TonB. Ces deux systèmes utilisent la force proton motrice, issue de la membrane interne et transfert l’énergie associée pour le transport actif de molécules (TonB) ou nécessaire au maintien de l’intégrité membranaire (Tol). Ces 2 systèmes ont été détournés de leurs fonctions initiales et parasités par les colicines, leur conférant un rôle primordial dans le mécanisme d’import de la colicine. Une colicine est une bactériocine (toxine) produite par Escherichia coli pour tuer des souches apparentées. Ce sont des toxines spécifiques et hautement actives. Cependant E.coli a développé des mécanismes de protection afin de résister à l’action cytotoxique des colicines. Ces mécanismes de résistance consistent essentiellement à produire des protéines d’immunité, qui vont pour la plupart se fixer sur le domaine catalytique de la colicine et l’empêcher d’exercer son action létale. La bactérie Yersinia pestis, agent de la peste, possède une colicin-like bactériocine, la pesticine, dont l’activité est de dégrader le peptidoglycane. L’action de la pesticine est inhibée par une protéine d’immunité, Pim, localisée dans le périplasme. Le principal objectif de ce projet est de comprendre les mécanismes d’inhibition de la pesticine par sa protéine d’immunité, grâce à des données biochimiques et structurales, mais aussi d’apporter des solutions pour contourner ce problème de résistance. La structure de la pesticine révèle des homologies structurales avec le T4 lysozyme du bactériophage T4. Pour contourner le problème de la résistance bactérienne liée à la protéine d’immunité, une solution a été de fusionner le domaine de réception/translocation de la pesticine avec le T4 lysozyme. Nous avons ainsi pu créer et résoudre la structure tridimensionnelle d’une protéine chimère fonctionnelle, capable de se fixer sur FyuA (récepteur de la pesticine) et tuer une souche exprimant ce récepteur et dont l’activité létale n’est pas inhibée par Pim<br>In Gram-negative bacteria, two essential systems for cell survival have been characterized: the Tol and TonB system. Both Ton and Tol systems are very well conserved in Gram-negative bacteria and coupled to the proton motive force across the inner membrane, acting as energy transducers for active transport (Ton) or maintenance of outer envelope integrity (Tol). Both systems have been embezzled from their primary function and hijacked by colicins as part of the colicin killing pathway. Colicin is a bacteriocin (toxin) produced by and toxic to some strains of Escherichia coli. Colicins are highly effective toxins. However E.coli could develop protective mechanisms to resist to colicin cytotoxic effect. These mechanisms essentially consist to produce an immunity protein. These proteins bind to colicin catalytic domain and inhibit its lethal activity. Yersinia pestis, plague agent, possesses its own colicin-like bacteriocin, Pesticin, which degrades murein. Pesticin activity is inhibited by an immunity protein, Pim, localized in the periplasm. The main goal of this project is to understand inhibition mechanisms between Pim and Pesticin by biochemical and structural data and to provide solution to overcome the resistance issue, since Pesticin was thought to be used as antimicrobial agent. The Pesticin structure has revealed that Pesticin share structural homologies with the T4 lysozyme from the bacteriophage T4. To overcome the resistance issue due to the immunity protein, one solution has been to fuse the Pesticin binding/translocation domain with the T4 lysozyme. Thus, we could engineered and solved the three-dimensional structure of a chimera protein, able to bind FyuA (Pesticin physiological receptor) and kill a FyuA expressing strain, in which the lethal activity is not affected by Pim

Tobacco SABP2 has been shown to display high affinity for salicylic acid (SA) and methylsalicylate (MeSA) and plays an important role in SAR signal development. Using biochemical approach, SABP2 has been shown to demonstrate strong esterase activity in converting MeSA to SA. Recent study shows that tetra fluoroacetophenone, a synthetic analog of SA, competitively inhibits SABP2 esterase activity as well as suppresses SAR signal development in tobacco mosaic virus (TMV)-infected tobacco plants. Not much has been studied on the effect of pesticides on plant defenses. Because both AChE and SABP2 are esterase-like proteins belonging to α/β hydroxylase superfamily, we hypothesize that pesticides may inhibit the MeSA esterase activity of SABP2 and block SAR development. Biochemical and molecular biology techniques were used to test this hypothesis. SAR in tobacco-TMV plant-pathogen system is measured by significant decrease in TMV-induced lesion sizes in secondarily inoculated distal leaves.

Metaflumizone é um novo inseticida, pertencente ao novo grupo químico semicarbazone, com excelente atividade no controle de várias espécies da Ordem Lepidoptera. O produto atua no sistema nervoso dos insetos bloqueando os canais de sódio sem a necessidade de ativação metabólica, ou seja, a molécula é tóxica em sua forma original. O mecanismo de ação de metaflumizone é distinto dos inseticidas piretróides que atuam como moduladores de canais de sódio. Para conhecer a atividade de metaflumizone sobre Spodoptera frugiperda (J.E. Smith) e estabelecer estratégias proativas de manejo da resistência para preservar a vida útil desse produto, os principais objetivos da dissertação foram: (a) estabelecer a linha-básica de suscetibilidade a metaflumizone em populações de S. frugiperda coletadas na cultura do milho em diferentes regiões do Brasil; (b) definir concentrações diagnósticas (CL95 e CL99) para o monitoramento da suscetibilidade das populações de S. frugiperda; (c) verificar a possibilidade de resistência cruzada entre metaflumizone e lambda-cyhalothrin; e (d) verificar a eficácia agronômica de metaflumizone no controle de S. frugiperda em condições de campo. O método de bioensaio adotado foi o de tratamento superficial da dieta artificial com inseticidas, utilizando-se lagartas de 3° ínstar. Para o monitoramento da suscetibilidade das populações de S. frugiperda a metaflumizone, foram definidas as concentrações diagnósticas de 32 e 100 µg de metaflumizone/mL de água destilada (ppm). Para as duas concentrações diagnósticas foram encontradas diferenças significativas na suscetibilidade das populações testadas, com sobrevivências de 5,3 a 16,73% na concentração diagnóstica de 32 ppm e de 0,42 a 10,37% na concentração de 100 ppm. Para verificar a presença de resistência cruzada entre metaflumizone e lambda-cyhalothrin, foi realizada a caracterização da curva de concentração-resposta a metaflumizone em uma população de S. frugiperda resistente (R) a esse inseticida e comparada com a resposta obtida com a população suscetível de referência (S), observando-se ausência de resistência cruzada. A eficácia agronômica de metaflumizone em condições de campo foi verificada por meio de 2 ensaios de campo na cultura de milho com infestações de S. frugiperda com alta freqüência de resistência a inseticidas piretróides (40%), realizando-se duas aplicações de inseticidas, espaçadas em sete dias. Os tratamentos foram: 1-testemunha; 2-metaflumizone 200 g I.A./ha, 3-metaflumizone 240 g I.A./ha; 4-indoxacarb 75 g I.A./ha; e 5-lambda-cyhalothrin 7,5 g I.A./ha. Os menores níveis de desfolha, aos 7 dias após a segunda aplicação de inseticida, foram obtidos com os tratamentos de indoxacarb a 75 g I.A./ha e metaflumizone a 240 g I.A./ha, comprovando a eficácia no controle de S. frugiperda. Portanto, metaflumizone pode ser utilizado como uma alternativa para o manejo da resistência de S. frugiperda a inseticidas.<br>Metaflumizone is a new insecticide, belonging to the new chemical group semicarbazone, with excellent activity against many species of Lepidoptera Order. The product acts in the nervous system of insects blocking the sodium channels without metabolic activation, in other words, the molecule is toxic on its original form. The mechanism of action of metaflumizone is distinguished to the pyrethroids that act as sodium channel modulators. To know the activity of metaflumizone against Spodoptera frugiperda (J.E. Smith) and to establish proactive strategies of resistance management to preserve the lifetime of that product, the main objectives of this work were: (a) to establish the baseline susceptibility of metaflumizone in populations of S. frugiperda collected in maize crop in different regions of Brazil; (b) to define diagnostic concentrations (LC95 e LC99) for the susceptibility monitoring of S. frugiperda populations; (c) to verify the possibility of cross-resistance between metaflumizone and lambda-cyhalothrin; and (d) to verify the agronomic efficacy of metaflumizone in the control of S. frugiperda under field conditions. The bioassay method was the surface treatment of an artificial diet with pesticides by using 3rd instar larvae. The diagnostic concentrations of 32 e 100 µg of metaflumizone/mL of distilled water (ppm) were defined for the susceptibility monitoring of S. frugiperda populations to metaflumizone. Significant differences in susceptibility to metaflumizone were found for the two diagnostic concentrations in the populations, with survival percentage from 5.3 to 16.73% in the concentration of 32 ppm and from 0.42 to 10.37% in the concentration of 100 ppm. To verify the cross-resistance between metaflumizone and lambda-cyhalothrin, the characterization of dose-mortality curve to metaflumizone was done in a lambda-cyhalothrin-resistant population of S. frugiperda (R) and the results were compared to the response of the susceptible population (S). The results suggested no existence of cross-resistance. The agronomic efficacy of metaflumizone under field conditions was verified in 2 field trials in maize crop infested with S. frugiperda at high level of pyrethroids resistance frequency (40%), with two applications of insecticides, at seven days interval. The treatments were: 1-untreated; 2-metaflumizone 200 g A.I./ha, 3-metaflumizone 240 g A.I./ha; 4-indoxacarb 75 g A.I./ha; and 5-lambda-cyhalothrin 7,5 g A.I./ha. The lower defoliation levels were achieved by the treatments of indoxacarb at 75 g A.I./ha and metaflumizone at 240 g A.I./ha, at 7 days after the second application of the insecticides, confirming its efficacy against S. frugiperda. Therefore, metaflumizone can be used as an alternative for the resistance management of S. frugiperda to insecticides.

A exploração de ácaros predadores no manejo de ácaros fitófagos em sistemas de produção agrícola pode ser prejudicada pelo uso de agrotóxicos para o controle de outras pragas e doenças. O uso de produtos compatíveis com os ácaros predadores ou de linhagens de ácaros predadores resistentes a agrotóxicos pode favorecer a conservação desses inimigos naturais. Uma outra estratégia a ser explorada pode ser a liberação isolada ou combinada de ácaros predadores com hábitos alimentares e comportamentais distintos. Dentro desse contexto, o presente trabalho foi desenvolvido para coletar subsídios para a integração das estratégias de controle químico e biológico para a conservação e liberação dos ácaros predadores Neoseiulus californicus (McGregor) e Phytoseiulus macropilis (Banks) em programas de manejo do ácaro rajado Tetranychus urticae Koch em sistemas de produção agrícola, tais como cultivos protegidos de plantas ornamentais e hortaliças. Inicialmente a compatibilidade de alguns acaricida-inseticidas, inseticidas e fungicidas com N. californicus e P. macropilis foi avaliada nas concentrações recomendadas para cultivos protegidos de plantas ornamentais e hortaliças. Foram realizados estudos para verificar o efeito desses agrotóxicos sobre a mortalidade de imaturos e adultos e a taxa instantânea de crescimento (ri) de cada espécie. Foram realizadas a caracterização da linhabásica de suscetibilidade dos estágios imaturo e adulto de N. californicus e P. macropilis a deltametrina. Posteriormente, foi efetuado um monitoramento da suscetibilidade a esse inseticida em populações de P. macropilis coletadas em condições de campo. Além do efeito letal dos neonicotinóides acetamiprido, imidacloprido e tiametoxam sobre N. californicus e P. macropilis, estudos envolvendo a resposta funcional desses predadores sobre ovos tratados de T. urticae também foram avaliados. A toxicidade diferencial de abamectina e espiromesifeno foi avaliada para os ácaros predadores e T. urticae. E por fim, a viabilidade das liberações isolada e combinada de N. californicus e P. macropilis foi testada em condições de laboratório e casa de vegetação. Dentre os grupos de agrotóxicos testados, os fungicidas causaram menor impacto sobre os ácaros predadores. A compatibilidade de agrotóxicos testados foi maior com N. californicus do que com P. macropilis. Os estágios imaturo e adulto de N. californicus foram aproximadamente 3.600 e 3.000 vezes mais tolerantes a deltametrina do que os estágios de P. macropilis. No entanto, alta variabilidade na suscetibilidade a esse inseticida foi detectada entre as populações de P. macropilis, com razões de resistência de até 3.500 vezes. Apesar de os neonicotinóides terem apresentado baixa toxicidade e pouco efeito na taxa intrínseca de crescimento de N. californicus e P. macropilis, verificou-se que esses inseticidas podem afetar a resposta funcional desses ácaros predadores sobre ovos de T. urticae contaminados. Abamectina e espiromesifeno foram altamente tóxicos sobre de T. urticae . A toxicidade de abamectina foi também elevada sobre as duas espécies de ácaros predadores; porém, espiromesifeno foi compatível com N. californicus. As liberações combinadas de N. californicus e P. macropilis foram eficientes quando comparadas a liberações isoladas, somente em situações com alta relação predador:presa (igual ou superior a 1:20).<br>The exploitation of predaceous mites to manage phytophagous mites in agricultural production systems can be affected by the use of pesticides for controlling other pests and diseases. The use of pesticides that are compatible with predaceous mites or pesticide-resistant strains can promote the conservation of these natural enemies. Another strategy to be exploited can be the release of single or combined species of predaceous mites with distinct feeding habitat and behavior. Within this context, this research was conducted in order to collect data to integrate the chemical and biological control strategies to preserving and releasing the predaceous mites Neoseiulus californicus (McGregor) and Phytoseiulus macropilis (Banks) for managing twospotted spider mite Tetranychus urticae Koch in agricultural production systems, such as ornamental plants and vegetable crops. Initially, studies were conducted to select pesticides that are compatible with N. californicus and P. macropilis as well as to evaluate the feasibility of combined releases of predaceous mites for managing T. urticae. The compatibility of some acaricide-insecticides, fungicides and insecticides with N. californicus and P. macropilis was evaluated at recommended rates for ornamental and vegetable crops. The effect of these pesticides on mortality of immature and adult stages and on the instantaneous growth rate (ri) of both species was evaluated. The baseline susceptibility to deltamethrin was characterized for immature and adult stages of N. californicus and P. macropilis. Then, monitoring the susceptibility to deltamethrin was conducted in field populations of P. macropilis. Besides the lethal effect of the neonicotinoids acetamiprid, imidacloprid and thiamethoxam on N. californicus and P. macropilis, functional responses of both mites on treated T. urticae eggs were also evaluated. The differential toxicity of abamectin and spiromesifen was evaluated to both predaceous mites and T. urticae. And finally, the feasibility of single or combined releases of N. californicus and P. macropilis was evaluated under laboratory and greenhouse conditions. Among the group of pesticides tested, the fungicides caused the least impact on predaceous mites. Higher compatibility of tested pesticides was observed with N. californicus than with P. macropilis. The immature and adult stages of N. californicus were approximately 3,600 and 3,000-fold more tolerant to deltamethrin than those of P. macropilis. However, high variability in the susceptibility to this insecticide was detected among P. macropilis populations, with resistance ratios of up to 3,500-fold. Although neonicotinoids showed low toxicity and little effect on instantaneous growth rate of N. californicus and P. macropilis, these insecticides can affect the functional response of these predaceous mites on treated T. urticae eggs. Abamectin and spiromesifen were highly toxic to T. urticae. High toxicity of abamectin was also detected to both species of predaceous mites; however, spiromesifen was compatible with N. californicus. Combined releases of N. californicus and P. macropilis were more effective than single species releases, only in situations with high predator:prey ratios (equal or greater than 1:20).

The present study deals with the effect of pesticides on grape mites, with particular emphasis on predatory mites belonging to the family Phytoseiidae. This study comprises experiments carried out in laboratory and in open field. The main results are summarized below. Resistance to acaricides in Tetranychus urticae populations from Italy The two-spotted spider mite Tetranychus urticae was selected as study model. We conducted toxicological tests to assess the acute toxicity of several insecticides and acaricides on different spider mite populations. During these tests, resistance to acaricides in T. urticae was confirmed. This resistance can involve newly introduced formulations. The availability of susceptible and resistant populations of T. urticae allowed us to perform preliminary biochemical assays to assess the activity of detoxifying enzymes related to pesticide resistance. Effects of some fungicides on predatory mites in North-Italian vineyards We evaluated the effects of some fungicides (propineb, metiram, mancozeb, folpet and copper hydroxide) on the predatory mite Kampimodromus aberrans in a vineyard. The predatory mite was known to be poorly susceptible to ethylene-bis-dithiocarbamate (EBDCs) fungicides. In early season it was released in an experimental vineyard inhabited by another predatory mite, i.e. Amblyseius andersoni. Fungicides were applied following a randomized design. Fungicides proved to be not selective towards A. andersoni but this effect was influenced by the incidence of grape downy mildew Plasmopara viticola. In fact, this pathogenic fungus represents an alternative food for A. andersoni. K. aberrans was not influenced by the presence of P. viticola. EBDC fungicides caused significant effects on K. aberrans populations and induced spider mite increases. Effects of a number of pesticides on the predatory mite Kampimodromus aberrans in the laboratory To assess the effect of selected insecticides, we collected and reared some strains of K. aberrans native to different vineyards and apple orchards. We evaluated lethal and sub-lethal effects on predatory mite females. Spinosad and etofenprox were associated with significant effects on mite survival and fecundity. Thiamethoxam caused a significant reduction in fecundity. Resistance to chlorpyriphos in the predatory mite Kampimodromus aberrans Field investigations carried out in the Veneto region showed a potential resistance to chlorpyriphos in K. aberrans populations. Following these observations, we estimated acute toxicity (LC50 and LC90) on K. aberrans populations collected from vineyards (Monteforte d’Alpone and S. Polo di Piave), apple orchards (S. Michele all’Adige) or untreated hackberry trees (Legnaro). The latter exhibited a high susceptibility to chlorpyriphos. On the contrary, LC50 and LC90 in the three populations collected on grapes and apples were relatively high. Resistance factors (RR) were calculated by dividing LD50 of resistant strains by the LD50 of the susceptible strain. RR exceeded 100,000 for all strains being higher for Monteforte strain. These data demonstrated the resistance to organophosphates in K. aberrans. These results have significant implications because lethal concentrations were highest than recommended field doses. Toxicity of thiamethoxam to Tetranychus urticae and Phytoseiulus persimilis (Acari Tetranychidae, Phytoseiidae) through different routes of exposure The spider mite Tetranychus urticae and the predatory mite Phytoseiulus persimilis were exposed to topical (microimmersion bioassay), residual (treating leaves) or ingestion pesticide exposures. In the latter, the predators were fed with contaminated prey confined on treated leaves. We studied the effect of all possible exposure combinations on mites. Topical applications did not affect survival in T. urticae and P. persimilis, but showed significant effects on their fecundity. Toxicity of thiamethoxam to Tetranychus urticae and Kampimodromus aberrans (Acari Tetranychidae, Phytoseiidae) through different routes of exposure Similar experiments were conducted on T. urticae and the predatory mite Kampimodromus aberrans. Thiamethoxam did not affect K. aberrans survival. Regarding fecundity, feeding on contaminated prey was associated to more severe effects than residual and topical exposure. Interactions between K. aberrans – pollen – insecticides: laboratory experiments Field experiments suggested that the effects of pesticides on predatory mites were less pronounced where grass management induced a high pollen flow. We evaluated the influence of pollen availability on survival and fecundity of predatory mites exposed to some insecticides (chlorpyriphos, etofenprox, indoxacarb and spinosad). Survival on chlorpyriphos, etofenprox and spinosad was lower than that observed in the control and indoxacarb. The escape rate was higher on spinosad. Insecticides (exception made for indoxacarb) caused significant effects on the fecundity of K. aberrans. Etofenprox and spinosad showed the lowest levels of fecundity. The amount and the frequency of pollen application improved predatory mite fecundity. We found a significant interaction between insecticides and pollen frequency application. The availability of fresh pollen mitigated the effects of chlorpyriphos on predatory mite fecundity. In conclusion, the availability of alternative foods can mitigate the effects of insecticide application with implications for Integrated Pest Management.<br>L’attività scientifica ha riguardato lo studio degli effetti dei prodotti fitosanitari sulle comunità di acari della vite, con particolare riferimento agli acari appartenenti alla famiglia dei Phytoseiidae. Lo studio si è articolato in una serie di esperimenti effettuati sia in laboratorio sia in pieno campo, di cui vengono di seguito esposti i principali risultati. Resistenza in Tetranychus urticae in Italia L’acaro fitofago T. urticae è stato scelto come primo modello di studio. Su diverse popolazioni dell’acaro tetranichide sono stati svolti dei test per valutare la tossicità acuta di alcuni acaricidi. I test hanno dimostrato che la resistenza agli acaricidi è diffusa e può manifestarsi anche nei confronti di formulati di recente introduzione. E’ stato possibile individuare popolazioni resistenti e sensibili dell’acaro su cui effettuare una serie di saggi biochimici atti a valutare l’attività di diversi enzimi detossificanti potenzialmente correlati con la resistenza ai prodotti fitosanitari. Effetti di prodotti fungicidi sulle comunità di acari in pieno campo In questa seconda ricerca si sono valutati gli effetti di alcuni fungicidi (propineb, metiram, mancozeb, folpet e idrossido di rame) su una popolazione dell’acaro predatore Kampimodromus aberrans introdotta ex-novo in un vigneto in cui era già presente una popolazione autoctona di Amblyseius andersoni. L’acaro è stato inoculato nel vigneto sperimentale all’inizio della stagione vegetativa tramite l’apporto di femminelle prelevate da un vigneto “donatore”. I fungicidi testati non sono apparsi selettivi nei confronti del predatore Amblyseius andersoni, probabilmente a causa dell’alta incidenza della peronospora nel testimone. La peronospora infatti rappresenta un alimento alternativo per tale predatore che ha dimostrato maggiori possibilità di sviluppo su piante non trattate. A differenza di A. andersoni, K. aberrans non è risultato influenzato dalla presenza della peronospora. Alcuni fungicidi (mancozeb e propineb) hanno esercitato effetti negativi significativi sulle popolazioni di K. aberrans a differenza di quanto verificato durante precedenti indagini su popolazioni stabili della stessa specie. Effetto di insetticidi su Kampimodromus aberrans Utilizzando materiale proveniente da diversi vigneti, sono stati allestiti allevamenti di K. aberrans in laboratorio. Su questi ceppi si è valutato l’effetto di alcuni insetticidi scelti sulla base del loro diverso modo di azione. In particolare si sono testati: esteri fosforici (chlorpyriphos), carbammati (indoxacarb), neonicotinoidi (thiamethoxam), chitino-inibitori (flufenoxuron), MAC (methoxyfenozide), fenossiderivati (etofenprox), spinosine (spinosad). Le femmine dell’acaro predatore sono state sottoposte ad applicazioni di insetticidi alla dose di campo per valutare gli effetti su mortalità e fecondità delle femmine e schiusura delle uova deposte. L’impiego di etofenprox e spinosad è stato associato a effetti significativi sulla sopravvivenza e sulla fecondità di K. aberrans. Thiamethoxam ha causato una riduzione significativa della fecondità. Resistenza di Kampimodromus aberrans a chlorpyriphos Osservazioni condotte a Monteforte d’Alpone, S. Polo di Piave e S. Michele all’Adige hanno consentito di appurare un’elevata sopravvivenza nei confronti di chlorpyriphos da parte di K. aberrans. In seguito a tali osservazioni sono stati calcolati i parametri di tossicità acuta in queste popolazioni a confronto con altre non esposte a trattamenti insetticidi. Il calcolo dei parametri di tossicità acuta (DL50, DL90) è stato eseguito su due popolazioni raccolte su vite (Monteforte d’Alpone e S. Polo di Piave), su una popolazione raccolta su melo (S. Michele all’Adige) e su una popolazione prelevata su piante di bagolaro mai trattate con insetticidi. Quest’ultima ha esibito un’elevata sensibilità nei confronti di chlorpyriphos. Al contrario, i valori dei parametri di tossicità acuta nelle tre popolazioni raccolte su vite e melo sono risultati piuttosto elevati. I fattori di resistenza (RR), ottenuti dividendo la DL50 delle popolazioni resistenti per la DL50 della popolazione sensibile sono superiori a 100.000. E’ possibile ipotizzare che i parametri della tossicità acuta si abbassino aumentando l’esposizione a chlorpyriphos (per via topica oltre che per via residuale) ma, in ogni caso, viene dimostrata per la prima volta la resistenza agli esteri fosforici in K. aberrans. I risultati ottenuti sono particolarmente significativi se si considerano le possibili implicazioni pratiche, in quanto le dosi letali superano ampiamente le dosi di pieno campo. Effetti di diverse vie di esposizione a thiamethoxam in T. urticae e P. persimilis Sono stati condotti esperimenti atti a valutare quale possa essere l'effetto degli insetticidi sugli acari in funzione di differenti modalità di esposizione. A tale scopo femmine del fitofago T. urticae e del predatore P. persimilis sono state esposte all’insetticida thiamethoxam per via topica (microimmersione), per via residuale (trattando solo il substrato fogliare) o per ingestione. In quest’ultima modalità, i predatori sono stati alimentati con prede (T. urticae) confinate su foglie di vite contaminate con thiamethoxam tramite applicazione al terreno. Infine, sono stati studiati gli effetti sugli acari delle possibili combinazioni tra i diversi modi di esposizione (topica + ingestione, topica + residuale, ingestione + residuale, topica + ingestione + residuale). La risposta delle diverse specie di acaro è stata valutata calcolando il tasso di sopravvivenza e il tasso di fecondità. In T. urticae e P. persimilis i trattamenti topici non hanno manifestato effetti letali, ma hanno ridotto in modo significativo la fecondità. La combinazione di trattamenti fogliari e radicali ha avuto effetti negativi sia sulla mortalità sia sulla fecondità delle due specie. Effetti di diverse vie di esposizione a thiamethoxam in Tetranychus urticae e Kampimodromus aberrans Nel caso di K. aberrans, thiamethoxam non ha causato effetti negativi significativi a livello di sopravvivenza. Per quanto riguarda la fecondità, l’ingestione di prede contaminate è stata associata a effetti negativi più severi rispetto alle modalità di esposizione residuale e topica. Interazioni tra K. aberrans – polline – insetticidi: prove di laboratorio E’ stata valutata l’influenza dell’apporto pollinico sulla sopravvivenza e sulla fecondità di acari predatori esposti o meno all’effetto di alcuni insetticidi impiegati in viticoltura e in frutticoltura (chlorpyriphos, etofenprox, indoxacarb e spinosad). Nel caso di chlorpyriphos, etofenprox e spinosad la sopravvivenza è stata inferiore rispetto a quella osservata nelle tesi testimone e indoxacarb. Il tasso di fuga è risultato più elevato su spinosad. Gli insetticidi hanno causato una riduzione significativa della fecondità di K. aberrans rispetto al testimone e a indoxacarb. I livelli più bassi di fecondità sono stati osservati nelle tesi etofenprox e spinosad, ma anche chlorpyriphos ha causato riduzioni significative della fecondità. La quantità e la frequenza delle applicazioni polliniche non sono state associate ad effetti significativi sulla sopravvivenza e sul tasso di fuga del predatore. Al contrario, la fecondità è aumentata sia con la dose sia con la frequenza di somministrazione di polline. Infine, è stata riscontrata un’interazione significativa insetticidi*frequenza di applicazione pollinica. A basse frequenze di somministrazione pollinica la fecondità è stata più elevata nel testimone che su chlorpyriphos, mentre a elevate frequenze l’insetticida non ha causato effetti sulla fecondità. Una tendenza simile a questa è stata riscontrata per indoxacarb. In conclusione, la disponibilità di alimenti alternativi può mitigare gli effetti deleteri degli insetticidi con importanti implicazioni di carattere applicativo.

Conselho Nacional de Desenvolvimento CientÃfico e TecnolÃgico<br>O uso de inseticidas tem sido a principal estratÃgia de controle da mosca-branca, Bemisia tabaci (Gennadius). O uso intensivo desses produtos tem levado ao aparecimento de problemas de resistÃncia da mosca-branca aos mesmos em todo o mundo. Devido à carÃncia de estudos nessa Ãrea no Brasil, os objetivos desta pesquisa foram: validar uma tÃcnica de bioensaio para a caracterizaÃÃo de linhas-bÃsicas de suscetibilidade de B. tabaci a inseticidas e verificar a variabilidade genÃtica de populaÃÃes de B. tabaci quanto à suscetibilidade a alguns inseticidas no Brasil. A tÃcnica de bioensaio testada foi uma do tipo contato residual mediante o uso de discos foliares de feijÃo-de-porco como substrato. A influÃncia de plantas de algodÃo e soja sobre a suscetibilidade da criaÃÃo de B. tabaci foi tambÃm avaliada para a definiÃÃo dos procedimentos de bioensaio. Os inseticidas usados na pesquisa foram acetamiprido, imidacloprido, tiametoxam, clorpirifÃs, endosulfan e piridabem. Quatro populaÃÃes de mosca-branca foram testadas em relaÃÃo a uma populaÃÃo suscetÃvel de referÃncia (SusIAC), sendo duas oriundas do Estado de GoiÃs (GO-1 e GO-2) e outras duas do Estado da Bahia (BA-1 e BA-2). Os discos foram tratados por imersÃo em soluÃÃo inseticida e, posteriormente, foram colocados sobre uma camada de soluÃÃo Ãgar-Ãgua no fundo de tubo de vidro. Insetos adultos nÃo separados por sexo e nem por idade foram transferidos para o tubo contendo o disco tratado. As avaliaÃÃes foram realizadas apÃs 24 h da infestaÃÃo de mosca-branca para o endosulfan e 48 h para os demais produtos. As caracterizaÃÃes das linhas-bÃsicas de suscetibilidade de B. tabaci a inseticidas testados foram mais consistentes quando esse inseto foi criado em plantas de algodÃo ao invÃs de soja. Foram detectadas diferenÃas significativas quanto à suscetibilidade de mosca-branca aos inseticidas. A populaÃÃo GO-2 foi significativamente menos suscetÃvel aos inseticidas testados do que a SusIAC, principalmente em relaÃÃo aos neonicotinÃides (acetamiprido, imidacloprido, tiametoxam). A situaÃÃo mais crÃtica de resistÃncia de mosca-branca ocorreu com tiametoxam, seguida pelo imidacloprido.<br>The use of pesticides has been the major strategy to control the whitefly, Bemisia tabaci (Gennadius). The intensive use of these products has resulted in the development of whitefly resistance all around the world. Because of lack of studies in this subject in Brazil, the objectives of this research were: validate a bioassay technique to characterize the baseline susceptibility and evaluate the genetic variability of B. tabaci populations to some pesticides in Brazil. A residual contact bioassay by using foliar discs of Canavalia ensiformis L.. The effect of cotton and soybean plants on susceptibility of the rearing B. tabaci to acetamiprid, imidacloprid, thiamethoxan, chlorpyrifos, endosulfan and pyridaben was evaluated. Four whitefly populations, two from the GoiÃs state (GO-1and GO-2) e two from Bahia state (BA-1 and BA-2), were tested against a susceptible reference one (SusIAC). The foliar discs were treated by immersion on the chemical solutions and; then, they were transferred onto an agar-water solution in the bottom of a glass vial. Adult insect of unknown age and sex were transferred to the vials with treated foliar discs. Evaluations were performed after 24 h for endosulfan and 48 h for the other chemicals. The characterization of the baseline susceptibility of B. tabaci to the tested pesticides was more consistent when whiteflies were reared on cotton than on soybean plants. Significant differences in the susceptibility to pesticides were detected among B. tabaci populations. The population GO-2 was significantly less susceptible to tested pesticides than SusIAC, mainly to neonicotinoids. The most critical whitefly resistance situation was detected to thiamethoxan, followed by imidacloprid.

Este estudo foi realizado com o objetivo de identificar genótipos de feijão Phaseolus vulgaris resistentes aos carunchos Acanthoscelides obtectus e Zabrotes subfasciatus, bem como avaliar o efeito associado desses genótipos resistentes com inseticidas de origem vegetal. Para tanto, foram utilizados acessos de P. vulgaris do Banco de Germoplasma do Instituto Agronômico de Campinas e inseticidas comerciais de origem botânica. No screening inicial, amostras dos genótipos foram infestadas com cada uma das espécies de bruquídeos separadamente, avaliando-se o número de insetos emergidos aos 50 dias após a infestação. Dos 49 genótipos testados contra A. obtecus, não houve emergência naqueles com números de acesso 525, 584 e 615, podendo ser considerados os mais resistentes. Em relação a Z. subfasciatus, os genótipos com números de acesso 2, 35, 251, 570, 583, 584, 610, 621, 634, 816, 818 e 819 se mostraram mais resistentes entre os 185 avaliados. Destes, os genótipos portadores de arcelina 583, 584, 816, 818 e 819, além de 570 e 610, foram selecionados como os mais promissores para os testes subseqüentes, juntamente com a variedade Bolinha, que foi utilizada como controle de suscetibilidade. Não foi observada correlação entre as características morfoagronômicas dos genótipos de P. vulgaris e a sua resistência às espécies de bruquídeos avaliadas, indicando que a resistência a estas pragas não está associada às características da flor, vagem, semente e fenologia dos genótipos. A massa de mil sementes, que é indicativa da origem dos genótipos, foi um dos descritores analisados, mostrando também que a resistência de P. vulgaris a A. obtectus e a Z. subfasciatus não está relacionada à origem dos genótipos. Em testes de livre escolha e de confinamento, avaliou-se o efeito dos genótipos selecionados no screening, juntamente com a variedade Bolinha, sobre o comportamento e biologia de Z. subfasciatus. Verificou-se que a avaliação da preferência de Z. subfasciatus por genótipos de P. vulgaris em teste de livre escolha pode ser feita com 1 dia após a infestação e que \'Bolinha\', apesar de ser suscetível a Z. subfasciatus e favorecer o seu desenvolvimento, apresenta antixenose para oviposição em relação à praga. Nos testes realizados, os genótipos contendo arcelina tenderam a ser mais resistentes que os demais sem essa proteína, sendo que os seus efeitos sobre Z. subfasciatus incluíram o aumento da mortalidade no período de desenvolvimento, alongamento desse período e redução do peso de adultos emergidos, mantendo-se, de certa forma, estáveis ao longo de duas gerações da praga. A resistência conferida pela arcelina revelou ser do tipo antibiose, tendo como causas a impropriedade nutricional e a ação no metabolismo do inseto. Com relação aos inseticidas botânicos, foram testados 3 produtos comerciais, sendo 2 à base de azadiractina e um à base de rotenona. Entre estes, o produto que mais afetou o desenvolvimento de Z. subfasciatus foi NeemPro®, derivado de nim (Azadirachta indica), o qual apresentou efeito ovicida e prolongou a duração do período de desenvolvimento de Z. subfasciatus. Frente a isto, avaliou-se o efeito associado de NeemPro® com o genótipo resistente portador de arcelina 818 sobre alguns parâmetros biológicos de Z. subfasciatus. Verificou-se que os efeitos mais severos sobre Z. subfasciatus foram causados pelo genótipo resistente, independentemente do inseticida à base de nim e que o uso associado de ambos não provoca efeito aditivo ou sinérgico, não sendo recomendado para o manejo de Z. subfasciatus.<br>This research was carried out to identify Phaseolus vulgaris genotypes resistant to the bean weevils Acanthoscelides obtectus and Zabrotes subfasciatus, as well as to evaluate the effect of these genotypes in association with botanical insecticides. To reach this objective, P. vulgaris accessions from the Germplasm Bank of Instituto Agronômico de Campinas and commercial insecticides from botanical origin were tested. In the initial screening, samples of bean genotypes were infested with the weevil species separately and the number of adults emerged at the 50th day after infestation was evaluated. There was no A. obtectus emergence in genotypes 525, 584 and 615, among the 49 ones screened against this pest. In relation to Z. subfasciatus, genotypes with accession numbers 2, 35, 251, 570, 583, 584, 610, 621, 634, 816, 818 and 819 showed themselves resistant among 185 screened ones. The arcelin-containing genotypes 583, 584, 816, 818 and 819, plus 570 and 610 (both lacking this protein), were selected as the most promising for additional evaluations. Bolinha variety was also used as the susceptible standard. No correlation between morpho-agronomical characteristics of the P. vulgaris genotypes and their resistance to the weevils was observed, indicating that resistance to these two pests is not associated to genotypes flower, pod and seed characters or plant phenology. The mass of 1000 seeds, which indicates the origin of genotypes, was one of the used descriptors, showing that P. vulgaris resistance to A. obtectus and Z. subfasciatus is not related to genotypes origin too. In free- and no-choice tests, it was evaluated the effect of the screened genotypes on Z. subfasciatus behavior and biology, compared to \'Bolinha\'. It was verified that in free-choice tests, the evaluation of Z. subfasciatus preference for P. vulgaris genotypes can be done 1 day after infestation. Despite being susceptible to Z. subfasciatus and supporting its development, \'Bolinha\' holds antixenosis for oviposition in relation to the pest. In the bioassays carried out, genotypes containing arcelin tended to be more resistant than those lacking this protein and their effects on Z. subfasciatus include increasing of the mortality in the developmental period, enlargement of this period and reduction in adult weight, also being stable during two pest generations. The resistance provided by arcelin revealed itself to be antibiosis, by acting as antinutrients and also as antimetabolics. In relation to botanical insecticides, 3 commercial products, 2 of them based on azadirachtin and 1 based on rotenone, were evaluated. The insecticide NeemPro®, extracted from neem (Azadirachta indica) was the only one significantly causing ovicidal effect and enlarging Z. subfasciatus developmental period. Based on these results, the associated effect of NeemPro® and the resistant arcelin-containing P. vulgaris genotype 818 on some Z. subfasciatus biological parameters was evaluated. It was verified that the most severe effects on Z. subfasciatus were caused by the resistant bean genotype, independently of the neem based insecticide. The associated use of these two control methods no results in additive or synergistc effect and is not recommended for the management of Z. subfasciatus.

La mise au point d'un système pour la sélection in vitro de deux solanacées diploïdes et autogames résistantes a l'atrazine, au chlortoluron et au terbutryne, est décrite. Les effets de ces produits ont été détermines sur des cultures d'organes (explants différencies) et sur des cals (tissus indifférencies). Aucune sélection n'est possible a partir de disques foliaires, entre-nœuds, anthères et cals non chlorophylliens. On procede ensuite au repiquage des tissus sur un milieu organogène. On réalise le micro greffage des apex des tiges ainsi formées sur de jeunes plantes issues de germination. On met en évidence que le caractère de résistance est transmis a certains des embryons produits par les plantes greffées

Une strategie de mutagenese-selection de cultures de protoplastes de tabac a permis d'obtenir des cals resistant a des pesticides de type n-alkyl-morpholine ou triazole, inhibiteurs de la biosynthese des sterols. Les differentes classes de cals resistants, discrimines en fonction de leur composition sterolique, suggerent l'existence de plusieurs mecanismes de resistance a ces pesticides, mettant en jeu soit une modification de la voie des sterols, soit des modifications dans d'autres voies metaboliques. En particulier, un mutant surproducteur de sterols a ete caracterise. L'analyse genetique montre que cette surproduction segrege avec la resistance, ce qui suggere la coincidence des deux phenotypes. Le mutant parvient ainsi a contrer l'effet des inhibiteurs phytotoxiques en surproduisant les sterols puis en esterifiant avec des acides gras les intermediaires biosynthetiques accumules. Ce mecanisme de regulation permet de maintenir une fraction de sterols libres, destines a etre incorpores dans les membranes, approchant en quantite et en qualite celle du genotype sauvage cultive en l'absence d'inhibiteur

Resumo: Os fungicidas agrícolas consistem num componente importante da produção agrícola, exercendo papel primordial principalmente no momento em que os demais métodos de controle mostram-se ineficientes. Sua intensa utilização vem provocando um sério problema, que é a resistência dos fungos a alguns princípios ativos utilizados. Quando se utilizam os fungicidas em um programa de manejo de doenças, é muito importante o conhecimento do mecanismo de ação das moléculas utilizadas, visando evitar erros que podem comprometer sua vida útil e, em alguns casos, gerar situações de difícil resolução como está. Porém, nem sempre as informações necessárias sobre os fungicidas utilizados na agricultura estão disponíveis, necessitando-se recorrer a inúmeras obras e trabalhos científicos para melhor compreender seu funcionamento. Este trabalho teve como objetivo a elaboração de uma revisão que abordasse todos os grupos de fungicidas utilizados no mercado nacional e internacional, disponibilizando informações importantes àqueles que fazem uso desta modalidade de controle, quer para recomendação direta no controle de doenças, quer para pesquisa e desenvolvimento que esclareçam sua performance regional, ou ainda na consulta acadêmica. Utilizou-se como diretriz para formatação deste trabalho as normas estabelecidas pelo FRAC (Fungicide Resitance Action Committee), que são internacionalmente conhecidas e respeitadas pelos usuarios de fungicidas. Juntamente com o mecanismo de ação, procurou-se também esclarecer alguns pontos importantes no metabolismo dos fungos, principalmente aqueles caracterizados como alvo dos fungicidas.<br>Abstract: The fungicides are efficient tools for agriculture, exerting primordial role especially when different methods of control are inefficient. However, the intensive use may result in a serious problem, the fungi resistance against the fungicide. When the fungicides are used in a program of disease management, knowing the mechanism of action of the molecules used is of the most importance, in order to prevent errous which can compromises its lifetime and, in some cases, to produce situations of difficult resolution. However, its not always that the information about the fungicides used in agriculture are readily available, and in that case, there's the need to consult several books and scientific pappers. The target of this work is to review and organize the information about all groups of fungicides used in the market, providing important information to those who are going to use this tool, for direct recommendation in the control of diseases, or for research and development which will help its regional performance, or for academic purposes. This work was based on the norms used by FRAC (Fungicide Resistance Action Committee), which are internationally known and respected by the users of fungicides. Additionally to the mechanism of action, it was also presented some important points in the fungi metabolism, mainly related to fungicide targets.<br>Orientador: Nilton Luiz de Souza<br>Coorientador: Wilson Story Venâncio<br>Banca: Edson Luiz Furtado<br>Banca: Dauri José Tessmann<br>Mestre

Made available in DSpace on 2014-06-11T19:28:37Z (GMT). No. of bitstreams: 0 Previous issue date: 2006-01-13Bitstream added on 2014-06-13T20:37:49Z : No. of bitstreams: 1 rodrigues_mat_me_botfca.pdf: 1701093 bytes, checksum: a0f9979d96649f11849b3f734c7eb846 (MD5)<br>Os fungicidas agrícolas consistem num componente importante da produção agrícola, exercendo papel primordial principalmente no momento em que os demais métodos de controle mostram-se ineficientes. Sua intensa utilização vem provocando um sério problema, que é a resistência dos fungos a alguns princípios ativos utilizados. Quando se utilizam os fungicidas em um programa de manejo de doenças, é muito importante o conhecimento do mecanismo de ação das moléculas utilizadas, visando evitar erros que podem comprometer sua vida útil e, em alguns casos, gerar situações de difícil resolução como está. Porém, nem sempre as informações necessárias sobre os fungicidas utilizados na agricultura estão disponíveis, necessitando-se recorrer a inúmeras obras e trabalhos científicos para melhor compreender seu funcionamento. Este trabalho teve como objetivo a elaboração de uma revisão que abordasse todos os grupos de fungicidas utilizados no mercado nacional e internacional, disponibilizando informações importantes àqueles que fazem uso desta modalidade de controle, quer para recomendação direta no controle de doenças, quer para pesquisa e desenvolvimento que esclareçam sua performance regional, ou ainda na consulta acadêmica. Utilizou-se como diretriz para formatação deste trabalho as normas estabelecidas pelo FRAC (Fungicide Resitance Action Committee), que são internacionalmente conhecidas e respeitadas pelos usuarios de fungicidas. Juntamente com o mecanismo de ação, procurou-se também esclarecer alguns pontos importantes no metabolismo dos fungos, principalmente aqueles caracterizados como alvo dos fungicidas.<br>The fungicides are efficient tools for agriculture, exerting primordial role especially when different methods of control are inefficient. However, the intensive use may result in a serious problem, the fungi resistance against the fungicide. When the fungicides are used in a program of disease management, knowing the mechanism of action of the molecules used is of the most importance, in order to prevent errous which can compromises its lifetime and, in some cases, to produce situations of difficult resolution. However, its not always that the information about the fungicides used in agriculture are readily available, and in that case, there's the need to consult several books and scientific pappers. The target of this work is to review and organize the information about all groups of fungicides used in the market, providing important information to those who are going to use this tool, for direct recommendation in the control of diseases, or for research and development which will help its regional performance, or for academic purposes. This work was based on the norms used by FRAC (Fungicide Resistance Action Committee), which are internationally known and respected by the users of fungicides. Additionally to the mechanism of action, it was also presented some important points in the fungi metabolism, mainly related to fungicide targets.

Dans un contexte de lutte intégrée contre les moustiques, l'Entente Interdépartementale pour la Démoustication (E.I.D. Ain, Isère, Rhône, Savoie Rhône-Alpes s'est tournée vers une lutte totalement biologique (Bti) pour lutter contre les moustiques. Mon sujet de thèse s'inscrit dans la suite d'une collaboration scientifique constante depuis 40 ans entre l'E.I.D. et le laboratoire de recherche dans lequel j'ai effectué ma thèse. Cet organisme de gestion utilise le Bti depuis 20 ans. Et bien qu'à ce jour, aucune population de moustique ne soit apparue résistante au Bti, ce gestionnaire s'interroge sur la possibilité d'apparition de populations résistantes aux traitements insecticides. Des travaux antérieurs ont laissé supposer qu'il existait une différence de sensibilité des larves de moustiques aux insecticides en fonction de leur gîte d'origine, les larves originaires de gîtes herbacées étant moins tolérantes que celles provenant des gîtes arborescents. Il nous a semblé nécessaire de comprendre et ainsi de s'intéresser aux différents mécanismes de résistance des larves de moustiques pour permettre, demain, une lutte plus efficace contre cet insecte. Et nous nous intéresserons à la résistance à divers xénobiotiques alimentaires : du téméphos (insecticide organophosphoré) au Bacillus thuringiensis var. israelensis (Bti - bactério-insecticide) en passant par de la litière naturelle issue de la décomposition de feuilles dans les gîtes à moustiques se révélant toxique pour les larves. L'intérêt de cette thèse est double. D'un point de vue fondamental, la connaissance et la compréhension de la résistance (des enzymes impliquées aux facteurs environnementaux en passant par les gènes mis en jeu) stimulent mes recherches. Et d'un point de vue appliqué, il est nécessaire de mettre au point, enfin, un système de lutte efficace non polluant, qui passe par la compréhension globale des résistances. La démarche expérimentale utilisée dans ce travail est d'identifier les dysfonctionnements environnementaux sur le terrain, les analyser au laboratoire sur des espèces modèles (ici Aedes aegypti) les mécanismes à l'origine de ces perturbations, puis revenir sur le terrain pour confronter les résultats de laboratoire avec ceux obtenus in natura (ici Ochlerotatus cataphylla, Aedes rusticus). Ainsi cette étude va porter à la fois sur des espèces de terrain (Aedes rusticus, Ochlerotatus cataphylla, Culex pipiens ...) que sur des espèces de laboratoires (Aedes aegypti, Aedes albopictus ...). Pour comprendre les mécanismes de résistance mis en jeu par ce nuisant, nous avons travaillés à plusieurs niveaux d'études avec des études écotoxicologiques réalisées grâce à des études de terrain en collaboration avec l'E.I.D. (Entente Interdépartementale pour la Démoustication), des études biochimiques nous permettant de caractériser les enzymes de résistance mises en jeu, et des études génétiques et moléculaires pour approfondir ces mécanismes, en espérant trouver les gènes impliqués.

La résistance des moustiques aux insecticides pyréthrinoïdes (PYRs) menace les programmes de lutte anti-vectorielle à l'échelle mondiale. Chez le moustique Aedes aegypti, vecteur de la dengue et du Chikungunya, les principaux mécanismes causant cette résistance ont été identifiés. La résistance métabolique joue alors un rôle important et consiste en une biodégradation accrue de l'insecticide par des enzymes de détoxication. Néanmoins, les bases moléculaires de ce mécanisme restent méconnues. La plupart des gènes impliqués dans la résistance métabolique aux PYRs ont été identifiés par des approches transcriptomiques, mais les modifications génomiques à l'origine de leur sur-expression dans les populations résistantes ainsi que les modifications structurales des enzymes en lien avec la résistance restent méconnues. Cette thèse vise alors à utiliser les nouvelles approches de séquençage à haut débit (NGS) pour caractériser les mécanismes moléculaires de la résistance aux PYRs chez le moustique Ae. Aegypti. La première partie de la thèse présente une étude pilote RNA-seq menée sur des populations de laboratoire sélectionnées avec des insecticides. Cette étude a pour objectif d'évaluer les avantages des NGS pour l'étude des mécanismes de résistance chez les moustiques. Le rôle des enzymes de détoxication dans la résistance a ainsi été clairement confirmé. Plusieurs gènes codant pour ces enzymes apparaissent sur-exprimés dans les populations résistantes et un important regroupement de P450 montre une forte empreinte de sélection en lien avec la résistance aux PYRs. La seconde partie de la thèse présente une étude sur des populations naturelles échantillonnées sur divers continents. Cette étude combine les technologies d'enrichissement génomique et de DNA-seq afin d'étudier les variations génomiques liées à la résistance au PYR Deltaméthrine. La comparaison de la couverture de séquençage entre populations résistantes et sensibles a permis d'identifier des variations de nombre de copies (CNVs) de certains gènes de détoxication associées à la résistance à la Deltaméthrine. Des mutations non-synonymes fortement liées au phénotype de résistance ont également été mises en évidence. La comparaison de ces marqueurs de la résistance entre les différentes populations a révélé que les gènes/mutations associés à la résistance à la Deltaméthrine sont peu conservés entre continents, probablement à cause des différences de fond génétique des populations, de leur histoire démographique et des pressions de sélections. La troisième partie de la thèse décrit une étude par RNA-seq portant sur les mêmes populations naturelles, visant à croiser des données de transcriptomique (expression des gènes et polymorphisme des transcrits) avec les données génomiques générées par l'étude précédente. Plusieurs enzymes de détoxications ont été retrouvées sur-exprimées chez les populations résistantes en lien avec les CNVs précédemment identifiées. Des centaines de variations de polymorphisme ont été identifiées par DNA-seq dans les zones cis-promotrices des différents gènes étudiés. Parmi ces variations, plusieurs apparaissent associées à la sur-régulation d'enzymes de détoxication. Enfin, la comparaison des données de polymorphismes obtenues par DNA-seq et RNA-seq a permis d'étudier les phénomènes d'expression d'allèles spécifiques en lien avec la résistance. Cette étude confirme l'intérêt de croiser des données de transcriptomique et de génomique pour caractériser les bases moléculaires de la résistance aux insecticides. D'un point de vue général, cette thèse permet de mieux appréhender les mécanismes de résistance du moustique Ae. aegypti aux PYRs mais aussi d'identifier de nouveaux marqueurs de la résistance potentiellement utilisables pour développer de nouveaux outils moléculaires diagnostiques de la résistance sur le terrain. Ce travail met également en avant les apports des NGS pour l'étude fine des bases moléculaires de l'adaptation d'organismes modèles<br>Mosquito control programs worldwide are increasingly threatened by resistance to pyrethroid insecticides (PYRs). In the dengue and chikungunya vector Aedes aegypti, the key resistance mechanisms include modifications in the protein targeted by insecticides (target-site mutations) and metabolic resistance, consisting in an increased insecticide biodegradation by so called detoxification enzymes. However, as opposed to target-site mutations, the molecular basis of metabolic resistance remains poorly understood. Most metabolic resistance genes have been detected by transcriptomic approaches based on their over-expressed in resistant populations, but genomic changes leading to these expression changes as well as structural changes in enzymes potentially involved in resistance remain unknown. In this context, this thesis aims at using next-generation sequencing approaches for characterizing PYR resistance mechanisms in the mosquito Ae. aegypti.The first chapter of this thesis describes a pilot study on laboratory insecticide-selected populations of Ae. aegypti. This study aims at investigating the benefits of next-generation sequencing for studying resistance mechanisms in mosquitoes. This study confirmed that detoxification enzymes play a key role in resistance, with several of them being over-expressed in resistant populations and a large cluster of cytochrome P450 genes showing a selection imprint associated with resistance to PYRs.The second chapter of this thesis describes a study conducted on natural mosquito populations from various continents. Combining genomic target enrichment (targeting about 800 genes potentially involved in resistance) and DNA-seq allowed unravelling genomic changes associated with resistance to the PYR deltamethrin. Comparing normalized sequencing coverage between resistant and susceptible populations identified significant copy number variations (CNVs) in several detoxification genes strongly associated to deltamethrin resistance. Non-synonymous mutations affecting detoxification enzymes associated to the resistance phenotype were also detected. Comparing resistance markers between populations from various continents revealed that genes/mutations associated with deltamethrin resistance are poorly conserved across continents, probably due to differences in the genetic background of populations but also differences in terms of demographic history and selection pressures.The third chapter describes an RNA-seq study performed on the same natural mosquito populations in order to cross-link transcriptomic data (gene expression and transcript polymorphism) with genomic data obtained from the previous study. Multiple detoxification enzymes were found over-transcribed in resistant populations linked with previously identified CNVs. Hundreds polymorphism variations were identified by targeted DNA-seq in cis-promoter regions of detoxification genes. Among them, several were associated with the upper-regulation of detoxification enzymes in resistant populations. Finally, cross-comparing polymorphism data obtain from DNA-seq and RNA-seq allowed investigating allele specific expression (ASE) events related to PYR resistance. Overall, this study confirmed the benefits of combining transcriptomic and genomic NGS approaches for studying the molecular basis of insecticide resistance.As a whole, this thesis not only contributed to better understand PYR resistance mechanisms in the dengue vector Ae. aegypti but also identified novel genomic markers of resistance opening the way for developing new molecular diagnostic to early detect and monitor resistance mechanisms in the field. This work also highlights the benefits of using NGS technologies for unravelling the molecular bases of adaptation in model organisms

Um dos principais riscos associados às culturas inseticidas que expressam toxinas da bactéria Bacillus thuringiensis (Bt) é a evolução de resistência em pragas alvo, processo governado por fatores genéticos e bioecológicos que se inter-relacionam. Um dos parâmetros chave que influenciam a evolução de resistência é a freqüência inicial do alelo de resistência na população da praga alvo (FreqInicial). Devido à complexidade desse processo, experimentos para estudar sua evolução em larga escala são praticamente impossíveis. Modelos matemáticos de simulação têm sido utilizados para estimar a freqüência do alelo de resistência (FreqR) à toxina Bt ao longo das gerações da praga. O risco de resistência foi estimado utilizado o modelo de Caprio, incorporando incerteza ao parâmetro FreqInicial. Essa abordagem, denominada análise de incertezas, possibilita estimar o risco ao longo das gerações da praga, quantificado pela probabilidade de FreqR exceder um valor crítico. Os principais objetivos deste trabalho foram: (i) discutir o uso de análise de incertezas no contexto da estimação do risco de resistência e (ii) avaliar o efeito de diferentes distribuições da freqüência inicial do alelo de resistência (FreqInicial) sobre as estimativas de risco. Foi desenvolvido um software (RRiskBt), em linguagem Visual BASIC que permite quantificar o risco de resistência utilizando análise de incertezas. Os resultados da análise de incertezas indicaram que as estimativas de risco são afetadas pela distribuição de FreqInicial de modo diferenciado ao longo das gerações. As estimativas do risco de resistência, considerando a distribuição Normal para FreqInicial, são similares àquelas considerando a distribuição Triangular quando as referidas distribuições têm a mesma variância. O uso da distribuição Uniforme, ao invés da Normal ou Triangular em função da falta de informação sobre FreqInicial, leva à superestimação das estimativas de risco de resistência nas gerações iniciais e subestimação nas gerações subseqüentes à geração na qual a freqüência crítica é atingida. A análise de sensibilidade de FreqR a variações nos parâmetros FreqInicial ou DFRes possibilita estimar a geração a partir da qual a estimativa de FreqR independe da variação do parâmetro em questão dentro de um intervalo de valores possíveis preestabelecido. A utilização da análise de incertezas possibilita estimar a geração da praga alvo a partir da qual o risco de resistência é superior a 0,99, independentemente da distribuição utilizada para caracterizar a incerteza associada a FreqInicial.<br>One of the main risks associated to transgenic crops expressing Bacillus thuringiensis (Bt) toxins is the pest resistance evolution, process driven by genetic and ecological inter related factors. A key parameter that influences the rate of resistance evolution is the initial frequency of the resistance allele in the pest population (FreqInicial). Due to complexity of that process, large scale field experiments to investigate resistance evolution are practically impossible. Mathematical simulation models have been utilized to estimate the R allele frequency (FreqR) along pest generations. The resistance risk was estimated using the deterministic Caprio’s model, incorporating uncertainty to FreqInicial. That approach, called uncertainty analysis, allows to estimate resistance risk along generations. The risk is quantified by the probability of FreqR exceeding a critical value. The main objectives of this work were: (i) to discuss the use of uncertainty analysis in the context of risk resistance estimation and (ii) to evaluate the effect of different FreqInicial input probability distributions on the risk estimates. A software (RRiskBt) was developed in Visual BASIC language to quantify risk of pest resistance evolution to Bt toxins using uncertainty analysis. The results of uncertainty analysis showed that the influence of FreqInicial input distributions on the risk estimates changes along pest generations. The risk estimates considering input Normal distribution for FreqInicial are similar to those ones obtained considering Triangular distribution if their variances are equal. The use of Uniform distribution instead the Normal or Triangular due to lack of information about FreqInicial, leads to super estimation of risk estimates for the initial generations and sub estimation for the generations after the generation for which the critical frequency is achieved. The sensitivity analysis of FreqR to FreqInicial or DFRes allows to estimate the generation after which the FreqR estimates become independent of changes in the particular parameter. The uncertainty analysis allows to estimate the pest generation after which the resistance risk is higher than 0.99, independently of the FreqInicial input distribution.

Laboratory, semi-field and field experiments were conducted with the aim of identifying biological and integrated control strategies to be applied against Frankliniella occidentalis. We studied the spatial and temporal distribution of the main thrips pests in greenhouses and potential antagonists in some greenhouses characterized by a different level of lateral openings and a rose garden in the open. The second experiment was planned to show that opening greenhouse structures are not automatically related to an increase in thrips problems on ornamental crops. The third experimental study has been undertaken with the aim of evaluating the impact of some biological control agents (BCAs) on Frankliniella occidentalis population. In the fourth experiment, the effect of a fungal strain of Beauveria bassiana on F. occidentalis reached the highest levels when residual and topical exposures were combined<br>Sono stati condotti esperimenti di laboratorio, semi-campo e campo (serra) volti a individuare strategie di lotta biologica ed integrata da applicare contro Frankliniella occidentalis. E’ stata studiata la distribuzione spazio-temporale dei principali tripidi fitofagi delle serre e dei potenziali antagonisti in alcune serre caratterizzate da un diverso livello di aperture laterali e in un roseto a cielo aperto. La seconda ricerca sperimentale è stata intrapresa con lo scopo di dimostrare che il grado di apertura delle serre verso l'ambiente esterno non implica un maggiore incremento delle problematiche correlate ai tripidi su colture ornamentali. Il terzo studio è stato condotto con l'obbiettivo di valutare l'impatto di alcuni agenti di controllo biologico su F. occidentalis. Nel quarto esperimento, l'effetto di un ceppo fungino di Beauveria bassiana su F. occidentalis ha raggiunto i livelli più elevati quando l’esposizione topica è stata combinata con quella residuale

Genetics, gene flow, and distribution of pesticide resistance traits were examined for organophosphate (OP) resistance in three beneficial phytoseiid mites. Levels and genetics of OP resistance in Amblyseius andersoni were examined first. Laboratory strains from Italy and Oregon, USA, were compared in susceptibility to insecticides used in western Oregon fruit crops. The Italian strain was 80-100 times more resistant to the OPs azinphosmethyl, diazinon, malathion, and phosalone, as well as carbaryl, a carbamate. Significant differences were not found between strains for endosulfan or fenvalerate. Using backcross analysis, response of F1 hybrids to azinphosmethyl indicated OP resistance was semidominant. Through novel statistical analysis, backcross of F1 to parent strains revealed resistance was polygenic, with at least two loci. Reciprocal crosses demonstrated the presence of maternal effects, with increased variation associated with progeny of Oregon females. In the next set of experiments, electrophoresis of allozymes was used to estimate gene flow for Typhlodromus pyri. Ten populations from two apple growing valleys of Oregon were compared. Subpopulations were collected from in and around commercial apple orchards. Four loci unaffected by pesticide use were examined. FST was calculated at 0.115, and Nm as 2.08. No allelic patterns could be discerned for populations among or within valleys; however, more variation was present for mite populations within valleys than between them. Some inbreeding was found within populations. While from dispersal studies one would conclude T. pyri is nondispersive, allozymic analysis indicates there is moderate gene flow. Factors affecting OP resistance distribution in T. pyri and Metaseiulus occidentalis were examined. A diagnostic concentration of azinphosmethyl was used to determine OP resistance frequencies for populations of each species, collected in and near commercial apple orchards in two valleys. OP resistance in T. pyri populations was localized: mites from 10 m or more outside orchards were OP susceptible, while those within orchards were resistant. This indicated limited gene flow. All M. occidentalis populations were resistant, indicating a regional resistance pattern and high gene flow. Factors which were not significant in the distribution of OP resistance were: valley, degree of orchard isolation, host plant, and seasonality.<br>Graduation date: 1994

Bibliography: leaves 165-188.<br>ix, 188 leaves, [2] leaves of plates : ill. (some col.) ; 30 cm.<br>This research examines biological and behavioural interactions among the diamond-back moth, Plutella xylostella L. (Lepidoptera: Plutellidae), Cotesia plutellae Kurdjumov (Hymenoptera: Braconidae) and the microbial insecticide, Bacillus thuringiensis Berliner. A deterministic simulation model is developed describing the influence of C. plutellae on the evolution of resistance to Bt in a diamond-back moth poulation.<br>Thesis (Ph.D.)--University of Adelaide, Dept. of Crop Protection, 1998