A cana-de-açúcar é uma das principais fontes de energia renovável, constituindo a matéria-prima mais importante na busca por energia limpa e sustentável. Os benefícios ambientais provenientes da produção e do uso dos derivados de cana-de-açúcar fomentam o desenvolvimento de métodos e produtos que aumentem, de modo sustentável, a geração de bioenergia. Dentre os diversos fatores limitantes para o aumento da produção de cana-de-açúcar, destaca-se a ocorrência e a severidade de fitopatologias como a escaldadura das folhas. Essa doença, causada pela bactéria Xanthomonas albilineans, causa diminuição da produtividade, necessidade de reforma precoce dos canaviais e queda de qualidade do caldo extraído que determinam prejuízos econômicos significativos para os agricultores. Atualmente, não há alternativas disponíveis para o controle químico ou biológico dessa fitopatologia. Portanto, existe uma necessidade urgente de desenvolvimento de novas moléculas como defensivos agrícolas que sejam eficazes, seletivas, de baixo custo e impacto ao meio ambiente. A X. albilineans produz uma família de antibióticos e fitotoxinas conhecida como albicidinas. As enzimas envolvidas na biossíntese de albicidinas são alvos moleculares extremamente atrativos para o planejamento de novos agroquímicos. Entre as enzimas dessa via, destaca-se a fosfopanteteinil transferase (XaPPT, E.C. 2.7.8.7), uma enzima essencial para o desenvolvimento da X. albilineans. Essa dissertação está dividida em duas partes: i. estudos computacionais e ii. estudos experimentais. Na parte computacional foi desenvolvida uma nova ferramenta, denominada ViTaMIn para auxiliar o desenvolvimento de modelos tridimensionais (3D) de proteínas. A enzima XaPPT foi utilizada como estudo de caso para validar o programa. Os resultados obtidos indicaram que ViTaMIn foi capaz de auxiliar a construção de um modelo 3D robusto da XaPPT. Além disso, foi verificado que ViTaMIn é uma alternativa útil para usuários iniciante e experientes na modelagem molecular. O modelo de XaPPT construído foi utilizado na triagem virtual para a identificação de novos candidatos a inibidores. A estratégia incluiu a aplicação de filtros moleculares e estudos de docagem molecular que resultaram na seleção de 10 candidatos a inibidores da enzima-alvo. Na parte experimental, o cultivo de culturas de X. albilineans foi padronizado e estudos de biologia molecular com a XaPPT conduzidos a partir da extração de DNA genômico da bactéria. Estratégias clássicas e modernas foram empregadas para a clonagem da XaPPT. Os resultados obtidos indicaram a obtenção de proteína expressa na forma solúvel. Os trabalhos integrando estudos computacionais e experimentais apresentados nessa dissertação de mestrado significam importantes contribuições no desenvolvimento de bases científicas sólidas para o desenvolvimento de novos candidatos a agroquímicos para a cultura de cana de açúcar. / Sugarcane is a major source of renewable energy, representing the most important source for clean and sustainable energy. The environmental benefits collected from the production and use of sugarcane derivatives boost the development of new methods and products to improve, in a sustainable way, bioenergy generation. However, the occurrence and severity of plant diseases, such as leaf scald, hinder the productivity of sugarcane crops. Sugarcane leaf scald is a widespread and devastating disease caused by the bacteria Xanthomonas albilineans. The disease has a dramatic impact on crop productivity, including reduced yields and drop in quality of the juice. Currently, there is no chemical or biological treatment for disease control. Therefore, there is an urgent need for new effective and selective pesticides with low cost and environmental impact. The X. albilineans produces a family of antibiotics and phytotoxins known as albicidin. The enzymes involved in the biosynthesis of albicidin are attractive targets for the design of new agrochemicals. Among them, the phosphopantetheinyl transferase (XaPPT, EC 2.7.8.7) enzyme plays an essential role to the development of X. albilineans. This dissertation is divided into two parts: i. computational studies and ii. experimental studies. In the computational part, we developed a new tool, named ViTaMIn to assist the development of three-dimensional models (3D) of proteins. The XaPPT enzyme was employed as a case study to validate the program. The results indicated that ViTaMIn was capable of assisting the construction of a robust 3D model of XaPPT. Moreover, Vitamin is a useful alternative for beginners and advanced modelers. The XaPPT model was used in virtual screening approach to identify new candidate inhibitors. The strategy included the application of molecular filters and molecular docking studies which afforded 10 inhibitor candidates for the target enzyme. In the experimental part, X. albilineans was cultured and standardized. On the basis of that, molecular biology studies were conducted on XaPPT. Classical and modern strategies were employed to cloning XaPPT. The results indicated that protein is expressed soluble. The integration of computational and experimental studies presented in this dissertation are important contributions in the development of strong scientific basis for the design of new agrochemicals for sugarcane cultures.

Submitted by Erika Demachki (erikademachki@gmail.com) on 2014-09-04T19:33:18Z No. of bitstreams: 2 Batista, Mariana Pedrosa - Avaliação citogenética e molecular de indivíduos ocupacionalmente expostos aos agrotóxicos - 2014.pdf: 1556018 bytes, checksum: 89fbad76ace350a8cf9c7db5c92f015d (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Made available in DSpace on 2014-09-04T19:33:18Z (GMT). No. of bitstreams: 2 Batista, Mariana Pedrosa - Avaliação citogenética e molecular de indivíduos ocupacionalmente expostos aos agrotóxicos - 2014.pdf: 1556018 bytes, checksum: 89fbad76ace350a8cf9c7db5c92f015d (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) Previous issue date: 2014-02-28 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Rural workers are constantly exposed to pesticides they use on crops, and this exposure may be responsible for genetic damage causing a health risk. A problem with the use of pesticides is the genotoxicity, which can lead to the onset of disease. Little is known about the relationship between genotoxicity and the variation of genetic polymorphisms of xenobiotic metabolism that may modify individual susceptibility to the possible genotoxic effects of pesticides. Therefore, there is a need to study genes as glutathione-S-transferase mu (GSTM1) and theta glutathione S-transferase (GSTT1) encoding detoxification enzymes of genotoxic compounds. Another important assessment in individuals exposed to pesticides is the presence of chromosomal translocation t(14;18) (q31,q21), which can be investigated at the molecular level. Thus, this study evaluated the polymorphisms of GSTM1 and GSTT1 genes in 120 individuals occupationally exposed to pesticides and 115 controls (without exposure to pesticides), by real-time PCR. The frequencies of GSTM1 and GSTT1 genotypes were found in 49 % and 18 %, respectively in the exposed group and 37 % and 18 %, respectively, for the control group. It has been found that there is a greater possibility of poisoning in workers who have null genotypes. There was no statistically significant correlation between the increased risk of intoxication and alcohol consumption, smoking and use of PPE. In addition, 29 workers exposed to pesticides for more than 15 years and with null genotypes in GSTT1 and / or GSTM1, were evaluated for the presence of the t (14;18)(q31, q21), in 100 nuclei, by fluorescent in situ hybridization (FISH) . The translocation was observed in only one individual, which may have been caused by prolonged exposure to pesticides, increasing age or alcohol consumption. Thus, the study of genetic polymorphisms and translocations as biomarkers of susceptibility is of fundamental importance in understanding the processes involved in genotype distribution mutagenesis and carcinogenesis and could help minimize the risks for susceptible individuals who are exposed to pesticides. / Os trabalhadores rurais estão constantemente expostos aos agrotóxicos que utilizam nas lavouras, e esta exposição pode ser responsável por danos genéticos causando um risco para a saúde. Um dos problemas da utilização de agrotóxicos é a genotoxicidade, que pode levar ao aparecimento de doenças. Pouco se sabe sobre a relação entre a genotoxicidade e a variação de polimorfismos genéticos de metabolização de xenobióticos que podem modificar a suscetibilidade individual aos possíveis efeitos genotóxicos dos agrotóxicos. Por isso, há a necessidade do estudo de genes como a glutationa-S-tranferase mu (GSTM1) e glutationa-S–transferase teta (GSTT1) que codificam enzimas de detoxificação de compostos genotóxicos. Outra avaliação importante em indivíduos expostos a agrotóxicos é a pesquisa da translocação cromossômica t(14;18)(q31;q21), a qual pode ser investigada em nível molecular. Nesse sentido, esse estudo avaliou os polimorfismos dos genes GSTT1 e GSTM1, em 120 indivíduos ocupacionalmente expostos aos agrotóxicos e de 115 controles (sem exposição aos agrotóxicos), pela metodologia de PCR em tempo real. As frequências de genótipos GSTM1 e GSTT1 encontradas foram de 49% e 18%, respectivamente, para o grupo exposto e 37% e 18%, respectivamente, para o grupo controle. Verificou-se que não há maior possibilidade de intoxicação em trabalhadores que apresentam genótipos nulos. Não houve correlação estatisticamente significativa entre o aumento do risco de intoxicação e o consumo de álcool, tabaco e uso de EPI. Além disso, 29 trabalhadores, expostos a agrotóxicos por mais de 15 anos e com genótipos nulos, GSTT1 e/ou GSTM1, foram avaliados quanto à presença da translocação t(14;18)(q31; q21), pela análise de 100 células, por Hibridação Fluorescente in situ (FISH). Com essa análise, a translocação foi observada em apenas um indivíduo, a qual pode ter sido causada pela exposição prolongada a agrotóxicos, aumento da idade ou consumo de álcool. Dessa forma, o estudo de polimorfismos genéticos e translocações, como biomarcadores de suscetibilidade é de importância fundamental na compreensão dos processos de distribuição genotípica envolvidos na mutagênese e carcinogênese e poderia ajudar a minimizar os riscos para indivíduos suscetíveis que são expostos a agrotóxicos.

The World Health Organization classified carbaryl, glyphosate, and paraquat as hazardous to human health. In the Agriculture Health Study in the United States, health problems were associated with the use of these 3 agrochemicals 12 or more times per year. These 3 agrochemicals were commonly used in Grenada. The purpose of this quantitative cross-sectional study was to investigate the relationship between the social and economic characteristics of the farmers who participated in the 2012 agriculture survey and the use of agrochemicals at frequencies that could pose health risks. Five constructs of the social cognitive theory were used as the premise to hypothesize relationships between the variables. Binomial regression analysis was conducted to test the hypotheses of relationships between the characteristics of 8,868 farmers and use of agrochemicals or herbicides -?¥12 timers per year. Statistically significant relationships were found between 16 characteristics of the farmers and use of agrochemicals. Significant relationships were also found between 8 characteristics of the farmers and use of herbicides -?¥12 timers per year. The findings of this study show that several characteristics of the farmers in Grenada were associated with the use of agrochemicals at frequencies that were hazardous to health. By demonstrating the need to implement preventive measures and adopt the precautionary principle in the use of agrochemicals, positive changes can be made in monitoring agriculture practices, health surveillance, and clinical practice.

The aims of this thesis were to evaluate the effects of agriculture on amphibians in terms of (i) population genetic consequences of agriculture-induced spatial changes of the landscape and (ii) local adaptation and tolerance to frequently used agrochemicals. The study was performed using the common frog Rana temporaria as a model. Abundance, occurrence, genetic diversity and gene flow were negatively affected by agriculture in southern Sweden, but unaffected or even positively affected by agriculture in the central and northern regions, respectively. These test parameters correlated positively with landscape diversity both in the south and in the north. Moreover, the size and occurrence of R. temporaria populations decreased towards the north i.e. the margin of the species’ distribution range. In accordance with theoretical expectations, genetic variability decreased and population substructuring increased as a negative function of (effective) population size. Southern Swedish common frogs are naturally exposed to higher levels of nitrates, and thus have a higher tolerance to high nitrate levels than their northern conspecifics. This suggests local adaptation to naturally varying nitrate levels. Consequently, increased anthropogenic supplementation of nitrate could impact more the northern than the southern Swedish common frog populations. Exposure to the pesticides azoxystrobin, cyanazine and permethrin at ecologically relevant concentrations had small or no effects on R. temporaria tadpoles. The populations with lowest microsatellite variation (fragmented populations) in southern Sweden had considerably lower fitness in terms of survival and growth as compared to those with the highest genetic variability (non-fragmented populations). The results indicate that populations with low levels of neutral genetic variability were phenotypically less differentiated than populations with higher levels of variability. One possible explanation for this is that the degree of population differentiation in low variability populations has been constrained due to lack of suitable genetic variation or inefficiency of selection relative to genetic drift.

A range of Proteaceous shrubs and other nursery plants were grown in containers with soilless media and various N levels. Plants demonstrated a range of responsiveness. Supplying soilless media with Osmocote (26% N) and other short term fertilisers proved to be a satisfactory method of studying the comparative nutrition of a wide range of container grown nursery plants using factorial experiments incorporating N, P, K and lime. Nutrient response surfaces were obtained using a central composite incomplete block design. Most Proteaceous shrubs were intolerant of high P levels due to excessive luxury consumption resulting in toxic foliar nutrient levels, especially in the presence of high N. Phosphorus sensitivity in plants appeared to correspond with the soil nutrient levels in their native habitat. This applied to species studied in the 2 main sub-families in the Proteaceae while similar findings were indicated for other Australian genera. A range of optimum N requirements in the Proteaceae was found – lowest needs for Protea which also had the greatest tolerance of very low fertiliser additions, compared to Grevillea robusta with much higher N requirements and strong foliage growth inhibition if nutrient levels are very low. Proteoid root growth on Grevillea rosmarinifolia only occurred at low nutrient levels and was not required for satisfactory foliage growth of container grown plants. Pot plants and seedlings, especially tomato, responded strongly to N and often there were positive NK interactions influencing foliage growth. Lime requirements were studied and in erica increasing lime rates depressed foliage growth. Comparative nutrition studies indicate that general or broad spectrum container media may be unsuitable for some groups of nursery plants and that they could be replaced by potting mixes designed to meet the widely differing needs of the species often grown. The number of specialist mixes would depend on the range of plants and be governed by management considerations.

Submitted by Igor Matos (igoryure.rm@gmail.com) on 2017-02-02T14:06:11Z No. of bitstreams: 1 DISS_2015_Francco Antonio Neri de Souza e Lima.pdf: 3778440 bytes, checksum: 0abb939bae9a4e03d9d98767bf13df6b (MD5) / Approved for entry into archive by Jordan (jordanbiblio@gmail.com) on 2017-02-03T11:50:18Z (GMT) No. of bitstreams: 1 DISS_2015_Francco Antonio Neri de Souza e Lima.pdf: 3778440 bytes, checksum: 0abb939bae9a4e03d9d98767bf13df6b (MD5) / Made available in DSpace on 2017-02-03T11:50:18Z (GMT). No. of bitstreams: 1 DISS_2015_Francco Antonio Neri de Souza e Lima.pdf: 3778440 bytes, checksum: 0abb939bae9a4e03d9d98767bf13df6b (MD5) Previous issue date: 2015-07-22 / A agricultura de monocultivos destinados à exportação demandam grandes porções de terras para sua produção, o que impulsiona os processos de desmatamento, ocupação e exploração de novas áreas. Na região Nordeste de Mato Grosso, a Terra Indígena (TI) Marãiwatsédé, território da etnia Xavante, foi ocupada na década de 1950 por não indígenas. Desde a retirada total dos Xavante de seu território, a TI foi disputada por não indígenas. No ano de 2010, após determinação de posse da TI para os Xavantes, o processo de desintrusão total dos ocupantes não indígenas foi concluído somente em 2013. No entanto, nenhuma avaliação sanitária e ambiental foi realizada pelos órgãos federais para devolução e o retorno dos indígenas à sua terra originária. No período de ausência dos Xavante na TI, a terra foi utilizada para o plantio de monocultivos como arroz, soja, milho e pastagens. Esse modelo de agricultura é químico-dependente de agrotóxicos e fertilizantes em seus processos de produção podendo expor as populações aos seus resíduos químicos diretamente ou indiretamente pelas matrizes ambientais (ar, solo e água) e biológicas (peixes, aves e mamíferos) contaminados. O objetivo desse estudo foi verificar a contaminação hídrica por agrotóxicos na TI Marãiwatsédé e relatar outros impactos negativos do agronegócio na saúde da comunidade. Para isso, foram realizados os estudos de a) quantificação da produção agrícola por uma série histórica e estimativa de consumo de agrotóxicos dentro da TI, b) Verificação da presença de agrotóxicos na água e sedimentos de rios por meio de análises químicas, e c) relato da situação de saúde ambiental, ocasionada pelo processo produtivo agrícola dentro da TI. Verificou-se aumento anual no consumo de agrotóxicos, sendo o glifosato, o metamidofós e o 2,4D, os agrotóxicos mais utilizados. Dos sete pontos amostrados das análises químicas para detecção da presença de agrotóxicos na água superficial e sedimento de rio, foi detectado resíduo do agrotóxico permetrina no ponto P01, na concentração de 0.19µ/l. Essa concentração é menor do que o VMP para esse agrotóxico na legislação de potabilidade da água (Portaria MS 2.914/2011). Foram analisados apenas 12 princípios ativos além de outras limitações do método. No entanto, questionam-se as implicações dessa substância e de outras, prováveis dentro da TI. No ponto amostrado houve relatos de adoecimento após consumo de água deste local. A saúde ambiental na TI estava fragilizada devido à grande degradação do ambiente por queimadas, desmatamento, exposição a montante de fertilizantes químicos e vasilhames de agrotóxicos que estavam espalhados em locais da TI. Além disso há lavouras de monocultivos em atividade no entorno da TI, aumentando o risco de contaminações das nascentes de rios e do ambiente da TI de “fora para dentro” / The monoculture for exportation demands a big portion of land to the production, which impulses the processes of deforestation, occupation and exploration of new areas. In the northeastern part of the Mato Grosso, the indigenous land (IL) Marãiwatsédé, territory of the ethnic Xavante, was occupied in the decade of 1950 by non-indigenous. Since the total removal of the Xavante of their own territory, the area was disputed by the non-indigenous. In the year of 2010, after the determination of possession of the IL to the Xavante, the total process of disintrusion of nonindigenous occupants was concluded in 2013. However, no evaluation sanitary and environmental was made by the federal institutions responsible to the devolution and return of the indigenes to their original land. On the period of absence of the Xavante from the IL, the land was used to the production of crops such as rice, soybeans, corn and pastures. This model of agriculture is chemical-dependent of pesticides and fertilizers in the process of production which may exposure the populations to chemical residues directly or indirectly by environmental matrices (air, soil and water) and biological (fish, birds and mammals) contaminated. The aim of this study is verify the water contamination by pesticides in the IL Marãiwatsédé and report other negatives impacts of the agribusiness in the community's health. Thus it were conducted the studies of a) quantification of the agriculture production by a historical series and the pesticide consumption estimate within IL, b) Verify the presence of pesticides in the water and sediments of rives through chemical analyses, and c) reports of the environmental health situation, due to the process of agricultural production inside of the IL. It was found the increase of the annual consumption of pesticides, being the glyphosate, the methamidophos and the 2,4-D, being glyphosate, the methamidophos and the 2,4-D, the most commonly used pesticides. From the seven collected points of the chemical analysis to the detection of the pesticides presence in the superficial water and sediment of rives it was detected the presence of the pesticide permethrin in the point P01, in the concentration of 0.19µ/l. This concentration is smaller than the LMR to this pesticide in the water potability legislation (Portaria MS 2.914/2011). It was analyzed only 12 substances besides of other limitations of the method. Though, it is questioned the effects of this substances inside of the IL. In this point there were reports of sickness after drinking water from this location. The environmental health inside of the IL was fragile because to the large environmental degradation by wildfires, deforestation, exposure to a pile of chemical fertilizers and bottles of pesticides that are spread in different places of the IL. Beside that there is crops around the area of the IL, increasing the risk of contamination of the rivers sources and the environmental of the IL from "outside to inside"

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Agrochemicals are intensely used on rice (Orysa sativa L.) paddy culture, and depending on their environmental persistence and toxicity they may contaminate water bodies and affect non-target organisms. Both herbicides and insecticides are recommended for use in rice culture in southern Brazil. This study aimed was divided into three different experiments, with the following aims: (i) to determine the persistence of the herbicides imazethapyr + imazapic (Only®), bispyribac-sodium, penoxsulan, clomazone, and quinclorac and of the insecticides carbofuran and fipronil in irrigation water; (ii) to investigate the effect of these agrochemicals on physical, chemical and biological water quality; and, (iii) to investigate the effects of these agrochemicals on the native zooplanktonic biota of rice paddy fields cultivated under the direct seeding system. The field experiment was carried out in Santa Maria (RS), at the Federal University of Santa Maria (UFSM), in crop years 2007/08 and 2008/09; in 9.7 x 7.6 m= 73.7 m2 plots separated by soil levees. The experimental design used was randomized blocks with three replications. The treatments applied were imazethapyr (75 g a.i. ha-1) + imazapic (25 g a.i. ha-1) (Only®), bispyribacsodium (50 g a.i. ha-1), penoxsulan (48 a.i. g ha-1), clomazone (600 g a.i. ha-1), quinclorac (375 g a.i. ha-1), carbofuran (400 g a.i. ha-1) and fipronil (37.5 g a.i. ha-1) as well as a control block. In Experiment I, the agrochemicals persistence decreased in the following order: quinclorac > bispyribac-sodium = penoxsulan > carbofuran > imazapic > imazethapyr = fipronil > clomazone, detected for 84, 60, 60, 42, 39, 28, 28 e 25 days, respectively, (means of crop years 2007/08 and 2008/09). Quinclorac shows the greatest dissipation time (DT50) with 15.4 days, whilst clomazone exhibited the smallest DT50 value, 1.9 days. On Experiment II, results show that the isolated application of the herbicides and insecticides tested did not result in noticeable water quality alteration when compared to the control treatment. In general, physical, chemical and biological parameters remained below the values recommended by the environmental agencies used as reference for this study. In Experiment III, results indicated that Rotifers had the highest abundance in the zooplankton community, with Polyarthra as the most abundant genus; it was also concluded that the agrochemicals had little effect over the Rotifera and Copepoda groups (adult and nauplii). / A lavoura de arroz (Orysa sativa L.) irrigado utiliza agroquímicos intensamente, que, dependendo da sua persistência no ambiente e toxicidade, podem contaminar corpos d água e afetar organismos não-alvo. Os herbicidas e os inseticidas são agrotóxicos recomendados para o arroz irrigado no sul do Brasil. Este estudo foi desenvolvido em três experimentos, os quais tiveram por objetivos: (i) determinar a persistência dos herbicidas imazetapir + imazapique (Only®), bispiribaque-sódico, penoxsulam, clomazona, quincloraque, e dos inseticidas carbofurano e fipronil na água de irrigação em áreas cultivadas com arroz irrigado; (ii) investigar o efeito destes agrotóxicos na qualidade física, química e biológica da água; e, (iii) investigar os efeitos destes agrotóxicos na densidade da biota zooplanctônica natural no arroz cultivado no sistema de plantio direto. O experimento foi realizado a campo, na UFSM, em Santa Maria (RS), nas safras de 2007/08 e 2008/09; e conduzido em parcelas de 9,7 x 7,6 m= 73,7 m2 isoladas por taipas. O experimento foi conduzido com delineamento de blocos ao acaso com três repetições. Cada parcela recebeu de forma aleatória um tratamento, onde foram aplicadas a mistura dos herbicidas imazetapir (75 g i.a. ha-1) com imazapique (25 g i.a. ha-1), (Only®) bispiribaque-sódico (50 g i.a. ha-1), penoxsulam (48 i.a. g ha-1), clomazona (600 g i.a. ha-1), quincloraque (375 g i.a. ha-1), e os inseticidas carbofurano (400 g i.a. ha-1) e fipronil (37,5 g i.a. ha-1), além do tratamento controle. No Experimento I, os resultados mostraram que na média das duas safras (2007/08 e 2008/09) a persistência dos agrotóxicos decresceu na seguinte ordem: quincloraque > bispiribaque sódico = penoxsulam > carbofurano > imazapique > imazetapir = fipronil > clomazona com persistência de 84, 60, 60, 42, 39, 28, 28 e 25 dias, respectivamente. Quincloraque registrou maior tempo de dissipação (DT50) com 15,4 dias, enquanto clomazona apresentou o menor valor de DT50 na água com 1,9 dias. No Experimento II, os resultados mostraram que a aplicação isolada dos herbicidas e os inseticidas testados não acarretam grande alteração na qualidade da água quando comparada à qualidade da água de irrigação do tratamento controle (padrão). Em geral, os parâmetros físicos e químicos da água estiveram abaixo dos utilizados pelas agências ambientais e usados como referência neste estudo. No Experimento III, os resultados indicaram que Rotífera foi o grupo que apresentou maior abundância na comunidade zooplanctônica, destacando-se o gênero Polyarthra como o mais abundante; e também que agrotóxicos provocaram poucas alterações na abundância de organismos dos grupos Rotifera e Copepoda (adulto e nauplio).

A cana-de-açúcar é uma das principais fontes de energia renovável, constituindo a matéria-prima mais importante na busca por energia limpa e sustentável. Os benefícios ambientais provenientes da produção e do uso dos derivados de cana-de-açúcar fomentam o desenvolvimento de métodos e produtos que aumentem, de modo sustentável, a geração de bioenergia. Dentre os diversos fatores limitantes para o aumento da produção de cana-de-açúcar, destaca-se a ocorrência e a severidade de fitopatologias como a escaldadura das folhas. Essa doença, causada pela bactéria Xanthomonas albilineans, causa diminuição da produtividade, necessidade de reforma precoce dos canaviais e queda de qualidade do caldo extraído que determinam prejuízos econômicos significativos para os agricultores. Atualmente, não há alternativas disponíveis para o controle químico ou biológico dessa fitopatologia. Portanto, existe uma necessidade urgente de desenvolvimento de novas moléculas como defensivos agrícolas que sejam eficazes, seletivas, de baixo custo e impacto ao meio ambiente. A X. albilineans produz uma família de antibióticos e fitotoxinas conhecida como albicidinas. As enzimas envolvidas na biossíntese de albicidinas são alvos moleculares extremamente atrativos para o planejamento de novos agroquímicos. Entre as enzimas dessa via, destaca-se a fosfopanteteinil transferase (XaPPT, E.C. 2.7.8.7), uma enzima essencial para o desenvolvimento da X. albilineans. Essa dissertação está dividida em duas partes: i. estudos computacionais e ii. estudos experimentais. Na parte computacional foi desenvolvida uma nova ferramenta, denominada ViTaMIn para auxiliar o desenvolvimento de modelos tridimensionais (3D) de proteínas. A enzima XaPPT foi utilizada como estudo de caso para validar o programa. Os resultados obtidos indicaram que ViTaMIn foi capaz de auxiliar a construção de um modelo 3D robusto da XaPPT. Além disso, foi verificado que ViTaMIn é uma alternativa útil para usuários iniciante e experientes na modelagem molecular. O modelo de XaPPT construído foi utilizado na triagem virtual para a identificação de novos candidatos a inibidores. A estratégia incluiu a aplicação de filtros moleculares e estudos de docagem molecular que resultaram na seleção de 10 candidatos a inibidores da enzima-alvo. Na parte experimental, o cultivo de culturas de X. albilineans foi padronizado e estudos de biologia molecular com a XaPPT conduzidos a partir da extração de DNA genômico da bactéria. Estratégias clássicas e modernas foram empregadas para a clonagem da XaPPT. Os resultados obtidos indicaram a obtenção de proteína expressa na forma solúvel. Os trabalhos integrando estudos computacionais e experimentais apresentados nessa dissertação de mestrado significam importantes contribuições no desenvolvimento de bases científicas sólidas para o desenvolvimento de novos candidatos a agroquímicos para a cultura de cana de açúcar. / Sugarcane is a major source of renewable energy, representing the most important source for clean and sustainable energy. The environmental benefits collected from the production and use of sugarcane derivatives boost the development of new methods and products to improve, in a sustainable way, bioenergy generation. However, the occurrence and severity of plant diseases, such as leaf scald, hinder the productivity of sugarcane crops. Sugarcane leaf scald is a widespread and devastating disease caused by the bacteria Xanthomonas albilineans. The disease has a dramatic impact on crop productivity, including reduced yields and drop in quality of the juice. Currently, there is no chemical or biological treatment for disease control. Therefore, there is an urgent need for new effective and selective pesticides with low cost and environmental impact. The X. albilineans produces a family of antibiotics and phytotoxins known as albicidin. The enzymes involved in the biosynthesis of albicidin are attractive targets for the design of new agrochemicals. Among them, the phosphopantetheinyl transferase (XaPPT, EC 2.7.8.7) enzyme plays an essential role to the development of X. albilineans. This dissertation is divided into two parts: i. computational studies and ii. experimental studies. In the computational part, we developed a new tool, named ViTaMIn to assist the development of three-dimensional models (3D) of proteins. The XaPPT enzyme was employed as a case study to validate the program. The results indicated that ViTaMIn was capable of assisting the construction of a robust 3D model of XaPPT. Moreover, Vitamin is a useful alternative for beginners and advanced modelers. The XaPPT model was used in virtual screening approach to identify new candidate inhibitors. The strategy included the application of molecular filters and molecular docking studies which afforded 10 inhibitor candidates for the target enzyme. In the experimental part, X. albilineans was cultured and standardized. On the basis of that, molecular biology studies were conducted on XaPPT. Classical and modern strategies were employed to cloning XaPPT. The results indicated that protein is expressed soluble. The integration of computational and experimental studies presented in this dissertation are important contributions in the development of strong scientific basis for the design of new agrochemicals for sugarcane cultures.

Submitted by Adriana Martinez (amartinez@unoeste.br) on 2017-08-03T20:13:27Z No. of bitstreams: 1 Jamile Tomiazzi.pdf: 1140169 bytes, checksum: 85ab1353f7aab9dbdd62efcda0bc0e0c (MD5) / Made available in DSpace on 2017-08-03T20:13:27Z (GMT). No. of bitstreams: 1 Jamile Tomiazzi.pdf: 1140169 bytes, checksum: 85ab1353f7aab9dbdd62efcda0bc0e0c (MD5) Previous issue date: 2017-03-27 / The growing increase in the use of agrochemicals by small and large rural producers has generated environmental impacts and in the health of the exposed population. Another relevant public health problem, whose adverse effects have been widely documented, is smoking. Studies have indicated that isolate exposure to these xenobiotics can lead to ototoxicity and genotoxicity. Thus, the objective of this study was to evaluate the possible auditory and cytogenetic alterations in rural workers exposed to smoking and agrochemicals in isolation or in combination and to identify possible classification patterns of exposure groups. Were evaluated 127 workers of both sexes, with ages ranging from 18 to 39 years, divided into four groups: Control Group - CG; Group exposed to smoking - GT; Group exposed to agrochemicals - GA and Group exposed to the association between smoking and agrochemicals - GTA. Initially, a questionnaire was used to measure the exposure to the study compounds and general and auditory health. Auditory examinations (meatoscopy, conventional and high-frequency audiometry, logoaudiometry and imitanciometry) and cytogenetic evaluation (from cells of the bucal mucosa, stained by Giemsa method) were performed. The data were evaluated by the following pattern recognition algorithms Artificial Neural Network (ANN), Bayes Classifier (BAY) and Support Vector Machine (SVM). The results of the audiological evaluation demonstrated a lowering of high frequency thresholds, a higher incidence of descending type, type A tympanometry and absence of reflex of the stapedic muscle in the three exposed groups (GT, GA and GTA). In addition, these groups showed an increase in the total number of nuclear alterations and in the number of micronuclei, binucleate cells, karyotype, karynx, pycnotic cells and nuclear bud. The computational analysis did not recognize the GTA group as a real value, as with GT and GA in relation to GC, in which the data were distributed with standard and correctly classified. Therefore, it was concluded that exposure to agrochemicals and cigarettes, in isolation or in combination, has been shown to be potentially ototoxic and genotoxic. However, the concomitant use of xenobiotics did not lead to additive or potentiating effect. / O crescente aumento do uso de agroquímicos por pequenos e grandes produtores rurais tem gerado impactos ambientais e à saúde da população exposta. Outro problema relevante de saúde pública, cujos efeitos adversos têm sido amplamente documentados, é o tabagismo. Estudos têm indicado que a exposição isolada a estes xenobióticos pode levar a ototoxicidade e genotoxicidade. Assim, o objetivo deste estudo foi avaliar as possíveis alterações auditivas e citogenéticas de trabalhadores rurais expostos ao tabagismo e aos agroquímicos de maneira isolada ou em combinação e identificar possíveis padrões de classificação dos grupos de exposição. Foram avaliados 127 trabalhadores de ambos os sexos, com faixa etária de 18 a 39 anos, divididos em quatro grupos: Grupo Controle – GC; Grupo exposto ao tabagismo – GT; Grupo exposto a agroquímicos – GA e Grupo exposto à associação entre tabagismo e agroquímicos - GTA. Inicialmente foi aplicado um questionário, para coleta de dados sobre a exposição aos compostos de estudo e saúde geral e auditiva. Em seguida, foram realizados exames auditivos (meatoscopia, audiometria convencional e de alta frequência, logoaudiometria e imitânciometria) e avaliação citogenética (a partir de células da mucosa bucal, coradas pelo método Giemsa). Os dados foram avaliados pelos seguintes algoritmos de reconhecimento de padrões: Artificial Neural Network (ANN), Bayes Classifier (BAY) e Support Vector Machine (SVM). Os resultados da avaliação audiológica demonstraram rebaixamento de limiares auditivos em alta frequência, maior incidência de curva do tipo descendente, timpanometria tipo A e ausência de reflexo do músculo estapédico nos três grupos expostos (GT, GA e GTA). Além disso, nestes grupos foi observado aumento do número total de alterações nucleares e no número de micronúcleos, células binucleadas, cariólise, cariorréxis, células picnóticas e broto nuclear. A análise computacional não reconheceu o grupo GTA como um valor real, como ocorreu com GT e GA em relação ao GC, onde os dados foram distribuídos com padrão e classificadas corretamente. Diante disso, concluiu-se que a exposição a agroquímicos e cigarro, de maneira isolada ou em combinação, demonstrou ser potencialmente ototóxica e genotóxica. No entanto, o uso concomitante dos xenobióticos não levou a efeito aditivo ou de potencialização.

<p>PPO inhibitors are a valuable group of herbicides that provide soil-residual and foliar control of glyphosate resistant <em>Amaranthus</em> species. The ΔG210 mutation in the <em>PPX2</em> gene confers PPO-inhibitor resistance and has been present in the Midwest for more than a decade. Until recently, PPO-inhibitor resistance in waterhemp was attributable to just the ΔG210 mutation in the <em>PPX2</em> gene, but recently, several new PPO-resistant biotypes have been discovered in waterhemp and Palmer amaranth. A possible explanation is a change in PPO-inhibitor use patterns and commonly used active ingredients.</p> <p>A direct comparison of the ΔG210 mutation and a new mutation, R128G, was conducted in the greenhouse. Results indicate that the R128G mutation in waterhemp is not substantially better than the ΔG210 mutation with respect to conferring resistance to PPO inhibitors applied preemergence. Furthermore, there is no evidence that the utility of PPO inhibitors applied preemergence will diminish any further as a result of the R128G mutation increasing in frequency. A set of field trials was conducted to investigate how a new PPO inhibitor, trifludimoxazin, will select for resistant biotypes in the field. Overall, a greater number of resistant plants survived the foliar herbicide applications than emerged through soil applications. Trifludimoxazin did not increase the frequency of PPO-resistant individuals when applied to soil, but when applied to foliage, increased the frequency of PPO-resistant individuals by 2.5 to 2.6-fold, similar to other PPO inhibitors applied to foliage. In other experiments, investigations of waterhemp populations with unique resistance phenotypes were conducted. In populations that were more resistant than others, yet had the same ΔG210 mutation, there was no evidence of herbicide metabolism contributing to the greater resistance phenotype. In resistant populations that had no target site mutations, target site expression experiments and lipid peroxidation experiments were inconclusive. However, there was some evidence of increased target-site expression or increased antioxidant capacity as being causal mechanisms, although no mechanisms have been fully ruled out.</p>