Orientador: Antonio Carlos Monteiro / Banca: Giane Serafim da Silva / Banca: Érika Barbosa Neves Graminha / Resumo: O presente estudo investigou, em condições de laboratório, a atividade patogênica de diferentes isolados dos fungos Metarhizium anisopliae, Beauveria bassiana e Paecilomyces fumosoroseus, aplicados nas concentrações de 1 x1 06 e 1 x1 08 conídios/mL nos estágios de ovo, larva L2, pupa e adulto de Chrysomya puloria. Grupos de 30 ovos, 20 larvas L2 e 20 pupas foram banhados com as suspensões de conídios e no ensaio com adultos, grupos de 30 moscas, foram pulverizados com as mesmas suspensões. Em todos os ensaios, utilizaram-se quatro repetições por tratamento. Os três fungos e seus respectivos isolados apresentaram baixa atividade patogênica para a fase de ovo. Os isolados de B. bassiana e M. anisopliae, aplicados na concentração de 108 conídios/mL, promoveram 100% de mortalidade de larvas de C. puloria. O tratamento de pupas com o isolado JAB 07 de B. bassiana, na concentração de 108 conídios/mL, reduziu significativamente a emergência de adultos. O efeito dose¬resposta foi melhor evidenciado no ensaio com adultos, pois os isolados de M. anisopliae e B. bassiana dimimuiram a sobrevivência somente quando aplicados na concentração de 108 conídios/mL. Todos os fungos se mostraram patogênicos para C. putoria, havendo diferença entre os isolados quanto a ação patogênica. Os isolados de B. bassiana e M. anisopliae foram mais eficientes, atuando principalmente nas fases de larva e adulto. P. fumosoroseus evidenciou menor atividade patogênica, apenas reduzindo a sobrevivência de adultos emergidos a partir de pupas inoculadas. A eficiência do controle foi influenciada pela concentração de conidios, obtendo-se os melhores resultados com as suspensões mais concentradas. / Abstract: The present study investigated, in laboratory conditions, the pathogenic activity of different isolates of the fungi Metarhizium anisopliae, Beauveria bassiana and Paecilomyces fumosoroseus, applied in the concentrations of 1 x1 06 and 1 x1 08 conidia/ml in the stages of egg, L2 larvae, pupa and adult of Chrysomya putoria. Groups of 30 eggs, 20 L2 larvae e 20 pupae were bathed with the conidia suspensions and in the assay with adult, graups of 30 f1ies were pulverized with the same suspensions. In ali the assays there were used four repiicates per treatment. The three fungus and their respective isolates presented low pathogenic activity for the egg stage. The isolates B. bassiana and M. anisopliae, applied in the concentration of 108 conidia/ml, promoted a 100% of C. puloria larvae morta!ity. The pupae treatment with the isolate JAB 07 of B. bassiana, in the concentration of 108 conidia/ml, decreased significantly adult emergence. The dose-response effect was better evidenced in the assay with adults, because the isolates of M. anisopliae and B. bassiana only reduced the survival when applied in the concentration of 108 conidia/ml. All fungus demonstrated to be pathogenic to C. puloria, although there is a difference between the isolates in their pathogenic activity. The isolates of B. bassiana and M. anisopliae were more efficient, mainly acting in the larva and adult stages. P. fumosoroseus evidenced lower pathogenic activity, decreasing the survival of emerged adult only through inoculated pupae. The control efficiency was influenced by the conidia concentration, obtaining the best results with the most concentrated suspension. / Mestre

Orientador: Paulo Cezar Ceresini / Banca: Margarete Camargo / Banca: Jaqueline Rosemeire Verzignassi / Resumo: Estresse devido a mudanças ambientais pode impactar caracteres quantitativos através de alterações nas variâncias, genética e ambiental. Populações do patógeno da mela da soja, Rhizoctonia solani grupo de anastomose AG-1 IA, são altamente diferenciadas geneticamente ao longo de um gradiente latitudinal nas mais importantes regiões de cultivo de soja do Brasil. Entretanto, os processos evolutivos que guiaram a adaptação regional dessas populações são ainda desconhecidos. Neste trabalho foi testada a hipótese de que o estresse de temperatura elevada pode aumentar a variação genética para caracteres quantitativos, como a resistência a fungicida de amplo espectro, no fungo R. solani AG-1 IA da soja. Isto é o mesmo que testar se um aumento da temperatura, além da temperatura ótima para crescimento, resulta em aumento da herdabilidade de senso amplo para tolerância a um fungicida. Objetivou-se, especificamente, testar o efeito do estresse de temperatura sobre a variação genética para tolerância ao fungicida oxicloreto de cobre e avaliar a importância relativa da variação genética neutra e da seleção natural sobre a divergência e adaptação regional de populações de R. solani AG-1 IA da soja. Para tanto, avaliou-se o crescimento micelial in vitro de três populações brasileiras de R. solani AG-1 IA da soja sob dois regimes de temperatura, ótimo (25°C) e acima do ótimo (33,5°C), e sob três concentrações de oxicloreto de cobre: nenhum fungicida, 0,42 e 0,84 g.L-1. Foram determinadas os componentes de evolutibilidade: variância genética (IG), ambiental (IE) e a herdabilidade no sentido amplo (h2) para o crescimento micelial nas diferentes condições. Comparou-se, também, a diferenciação fenotípica por caracteres quantitativos (QST) e a diferenciação genética neutra (baseada em dados... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Stress due to environmental changes can impact quantitative traits through changes in variances, genetic and environmental. Populations of soybean foliar blight pathogen, Rhizoctonia solani anastomosis group AG-1 IA, are highly genetically differentiated along a latitudinal gradient in the most important Brazilian cropping areas. However, the evolutionary processes that have guided the regional adaptation of these populations are still unknown. In this study we tested the hypothesis that high temperature stress can increase the genetic variation for quantitative traits such as the tolerance to a broadspectrum fungicide, in the fungus R. solani AG-1 IA from soybean. This is the same as testing whether a temperature increase, beyond the optimum temperature for growth, results in increased broad sense heritability for fungicide tolerance. The specific objective was to test the effects of temperature stress on the genetic variation for tolerance to cupper oxychloride and to assess the relative importance of neutral genetic variation and natural selection on the divergence and local adaptation of R. solani AG-1 IA populations from soybean. We evaluated the in vitro mycelial growth of three Brazilian populations of R. solani AG-1 IA from soybean under two temperature regimes, optimal (25°C) and above optimal (33.5°C), and under three concentrations of cupper oxychloride: no fungicide, 0.42 and 0.84 g.L-1. We determined the components of evolvability: genetic (IG) and environmental (IE) variances and the broad-sense heritability (h2) for mycelial growth under these conditions. We also compared the phenotypic differentiation for quantitative traits (QST) and neutral genetic differentiation (based on microsatellite data) between three pairs of populations (FST). As measures of fungal phenotypic responses... (Complete abstract click electronic access below) / Mestre

A degradação enzimática rápida, eficiente e robusta de polissacarídeos derivados de biomassa lignocelulósica é atualmente um grande desafio na produção de biocombustíveis e considerada uma alternativa viável e promissora para se enfrentar a crise energética mundial e diminuir a dependência das fontes fósseis de energia. O bagaço de cana-de-açúcar no Brasil é a principal matéria lignocelulósica sustentável de grande potencial para a produção do etanol de 2ª geração. O principal requisito para a consolidação dessa abordagem é a disponibilidade de enzimas que hidrolisam a celulose, hemicelulose e outros polissacarídeos em açúcares fermentescíveis e em condições adequadas para a utilização industrial. O presente estudo visou à caracterização molecular, estrutural e funcional da endoglucanase GH12 do fungo Aspergillus terreus (AtGH12) por diferentes técnicas. O gene que codifica para essa enzima foi clonado e expressado no fungo filamentoso A. nidulans linhagem A773. A cepa com maior secreção foi selecionada e a sequência da enzima confirmada por espectrometria de massas MALDI TOF MS. Posteriormente, através de estudos funcionais de parametrização enzimática como pH e temperatura ótimos, estabilidade térmica, efeitos supressores e potencializadores de aditivos, a enzima AtGH12 foi caracterizada bioquímica e fisicamente. A espectrometria de massas do substrato hidrolisado pela catálise enzimática foi tomada como uma forma de investigar o padrão de clivagem da hidrólise e estudo do reconhecimento enzima/substrato para a AtGH12. As caracterizações estruturais das enzimas recombinantes obtidas utilizando as técnicas de espalhamento dinâmico de luz, dicroísmo circular, espalhamento de raios X a baixo ângulo e gel nativo serviram para determinação do enovelamento e estado oligomérico em solução da AtGH12. Com o intuito de fornecer subsídios para o desenvolvimento de coquetéis enzimáticos mais eficazes para hidrólise da biomassa lignocelulósica, a atividade da AtGH12 foi avaliada frente ao bagaço de cana-de-açúcar pré-tratados pelos processos hidrotérmicos e organossolve. Posteriormente, o seu grau de sinergismo nesse tipo de substrato foi determinado com o coquetel enzimático comercial Acellerase&reg. / Fast, more efficient and robust enzymatic degradation of lignocellulosic biomassderived polysaccharides is currently a major challenge in the production of biofuels and considered a feasible and promising alternative to confront the global energy crisis and reduce the dependence on fossil energy resources. The sugarcane bagasse in Brazil is the most abundant and sustainable lignocellulosic material for the production of 2nd generation ethanol. The main requirement for the consolidation of this approach is the availability of enzymes that hydrolyze cellulose, hemicelluloses and other polysaccharides into fermentable sugars suitable for industrial use. The present study was aimed at molecular, structural and functional characterization of an endoglucanase from the fungus Aspergillus terreus (AtGH12) using different techniques. The gene encoding this enzyme has been cloned and expressed in the filamentous fungus Aspergillus nidulans strain A773. The strain with increased secretion was selected and the enzyme sequence was confirmed by mass spectroscopy MALDI TOF MS. Later, functional studies such as analysis of optimal pH and temperature, thermal stability, suppression and enhance effects of additives were applied to the AtGH12 characterization. The mass spectrometry of hydrolyzed substrate from the enzyme catalysis was acquired as a way to investigate the cleavage pattern of hydrolysis and the study of the enzyme/substrate interaction. Structural characterization of the recombinant enzymes was obtained using techniques such as dynamic light scattering, circular dichroism as well as small angle X-ray scattering and native gel, aided to determine the folding and oligomeric state of AtGH12 in solution. In order to provide support for the development of more effective enzyme cocktails for hydrolysis of lignocellulosic biomass, the activity of AtGH12 was evaluated using sugarcane bagasse pretreated by hydrothermal and organosolv processes. Subsequently, the degree of synergism in this type of substrate was measured using a commercial enzyme cocktail Acellerase&reg.

A vinhaça e o bagaço de cana são os principais subprodutos oriundos do processamento da cana-de-açúcar nas indústrias sucroalcooleiras, sendo geradas grandes quantidades dos mesmos. O fungo basidiomiceto Pleurotus ostreatus tem a capacidade de degradar materiais lignocelulolíticos e produzir enzimas lignolíticas de interesse para as indústrias. Com o objetivo de avaliar a produção das enzimas lacases e peroxidase, o fungo Pleurotus ostreatus, foi cultivado em meio contendo bagaço pré-tratado e vinhaça, ou em meio contendo apenas vinhaça, em sistema de fermentação semissólido ou submerso; as enzimas extracelulares foram avaliadas após 7, 10 e 12 dias de cultivo. O bagaço peneirado foi considerado pré-tratado fisicamente (T1); para o pré-tratamento T2 o bagaço umedecido foi submetido a autoclave (121°C e 1 atm por 15 min); nos pré-tratamentos químicos, T3 e T4, o bagaço foi tratado com peróxido de hidrogênio e hidróxido de sódio nas seguintes concentrações: 0,75% H2O2 + 0,75% NaOH (T3) e 0,75% H2O2 + 1% NaOH (T4) na proporção 1:10 (p/v) e, em seguida foram submetidos à autoclave (121°C e 1 atm por 15min). A vinhaça utilizada foi proveniente de uma indústria sucroalcooleira (V1) e outra de destilaria (V2); a composição físico-química mostrou que a primeira possuía os índices de matéria orgânica e fósforo mais elevados que na vinhaça V2, enquanto que a relação C:N foi menor na vinhaça V1. Os extratos enzimáticos foram obtidos após filtração do meio submerso; para o meio semissólido foi necessário a adição de tampão citrato (1:5 p/v) antes da filtração. A atividade de lacasse e peroxidase em meio submerso, nos tratamentos com a vinhaça V1, foi superior ao observado em meio semissólido. A produção das enzimas em fermentação submersa, utilizando a vinhaça V1, apresentou valores de atividade de lacase, no tratamento TL1 e TL2, de 784,9 e 707,5 U.L-1, com atividade específica de 3,04 e 2,86 U.mg-1, respectivamente, e a amostra VL1, contendo apenas vinhaça, de 1,91 U.mg-1, no 12º dia de fermentação. Os valores mais altos de atividade de peroxidase foram obtidos nos tratamentos TL1, TL2, VL1, com 133,1; 131,2 e 126,1 U.L-1, respectivamente, após 12 dias de cultivo. A maior atividade específica obtida foi na VL1 (0,86 U.mg-1) no 7º dia de cultivo. O pré-tratamento físico do bagaço mostrou melhores condições para a produção das enzimas. Para a produção da lacase e da peroxidase é fundamental a composição da vinhaça. / The vinasse and bagasse are the principal by-products derived from the processing of sugarcane in the sugarcane industry, which generated large amounts of them. The basidiomycete fungus Pleurotus ostreatus, has the ability to degrade lignocellulolytic materials and produce lignolíticas enzymes of interest to industry. In order to evaluate the production of laccase and peroxidase enzymes, fungus P. ostreatus was grown in medium containing pre-treated bagasse and vinasse, or in medium containing only vinasse in semi-solid or submerged fermentation system; extracellular enzymes were evaluated after 7, 10 and 12 days of cultivation. The screened bagasse was considered pretreated physically (T1); for the pretreatment T2 moistened residue was subjected to autoclaving (121°C and 1 atm for 15 min). The chemical pretreatments, T3 and T4, the residue was treated with hydrogen peroxide and sodium hydroxide solution in the following concentrations 0,75% H2O2 + 0,75% NaOH (T3) and 0,75% H2O2 + 1% NaOH (T4) 1:10 (w/v) and then underwent autoclaving (121°C and 1 atm for 15 min). Vinasse used was coming from a sugar and alcohol industry (V1) and a distillery (V2); the physico-chemical composition showed that the former had the rates of organic matter and phosphorus higher than in V2 vinasse, whereas the C: N ratio was lower in V1 vinasse. The enzymatic extracts were obtained after filtration medium the submerged; to semisolid medium was necessary the addition of citrate buffer (1:5 w/v) prior to filtration. The activity of peroxidase and lacasse in submerged medium, in the treatments with the V1 vinasse, was higher than observed in semi-solid medium. The production of enzymes by submerged fermentation using the vinasse V1, presented laccase activity values, in the treatment TL1 and TL2, of 784,9 and 707,5 UI.L-1, with specific activity of 3,04 e 2,86 U.mg-1, on the 12th day of fermentation. Higher values peroxidase activity were obtained in the treatments TL1, TL2, VL1, with 133,1; 131,2 and 126,1 UI.L-1, respectively, after 12 days of culture. The highest specific activity was obtained at VL1 (0,86 U.mg-1) on the 7th day of culture. Physical bagasse pretreatment showed better conditions for the production of enzymes. For the production of the laccase and peroxidase is fundamental composition of vinasse.

Please read the abstract in the dissertation. / Dissertation (MSc)--University of Pretoria, 2013. / gm2014 / Chemistry / unrestricted

Botrytis species affect over 300 host plants in all climate areas of the world, at both pre and post-harvest stages, leading to significant losses in agricultural produce. Therefore, the development of a rapid, sensitive and reliable method to assess the pathogen load of infected crops can help to prescribe an effective curing regime. Growers would then have the ability to predict and manage the full storage potential of their crops and thus provide an effective disease control and reduce post-harvest losses. A highly sensitive electrochemical immunosensor based on a screen-printed gold electrode (SPGE) with onboard carbon counter and silver / silver chloride (Ag/AgCl) pseudo-reference electrode was developed in this work for the detection and quantification of Botrytis species. The sensor utilised a direct sandwich enzyme-linked immunosorbent assay (ELISA) format with a monoclonal antibody against Botrytis immobilised on the gold working electrode. Two immobilisation strategies were investigated for the capture antibody, and these included adsorption and covalent immobilisation after self-assembled monolayer formation with 3-dithiodipropionic acid (DTDPA). A polyclonal antibody conjugated to the electroactive enzyme horseradish peroxidase (HRP) was then applied for signal generation. Electrochemical measurements were conducted using 3,3’, 5,5’-tetramethylbenzidine dihydrochloride / hydrogen peroxide (TMB/H2O2) as the enzyme substrate system at a potential of -200 mV. The developed biosensor was capable of detecting latent Botrytis infections 24 h post inoculation with a linear range from 150 to 0.05 μg fungal mycelium ml-1 and a limit of detection (LOD) as low as 16 ng ml-1 for covalent immobilisation and 58 ng ml-1 for adsorption, respectively. Benchmarked against the commercially available Botrytis ELISA kits, the optimised immuno-electrochemical biosensor showed strong correlation of the quantified samples (R2=0.998) ... [cont.].

Cryphonectria cubensis is an ascomycetous fungus that causes a serious canker disease on Eucalyptus trees in many parts of the world. The importance of the disease has led to numerous studies involving the taxonomy, genetic diversity and the control of Cryphonectria canker. However, there remain many questions pertaining to the disease that have not been considered. The objectives of the studies presented in this thesis were, therefore, to investigate the possibility of biological control of Cryphonectria canker, to evaluate the currently used disease screening strategy in South Africa and to establish a transfection system with dsRNA elements in Diaporthe, which is closely related to Cryphonectria. The introductory chapter of this thesis provides a review of the literature pertaining to Cryphonectria cubensis. In addition literature on hypovirulence in fungi is also extensively reviewed, with a special emphasis on the genus Cryphonectria. The aim of study in the second chapter of the thesis was to screen the South African C. cubensis population for the presence of dsRNA viruses. Two viruses were identified and the full sequence of these elements showed a strong homology to the mitochondrial viruses (mitoviruses) within the family Narnaviridae. We, therefore, named the viruses Cryphonectria cubensis mitovirus 1 (CcMV1) and Cryphonectria cubensis mitovirus 2 (CcMV2). The two viral genomes are 2601 nucleotides and 2639 nucleotides in size respectively and encode for a protein that probably functions as an RNA-dependant RNA polymerase (RdRp). Pathogenicity studies indicated that the viruses do not result in a significant reduction in pathogenicity of C. cubensis. In the third chapter, results of a study to consider whether different Eucalyptus clones responded similarly to various South African C. cubensis isolates, are presented. The aim was, therefore, to evaluate the current C. cubensis resistant screening method used on Eucalyptus spp. in South Africa. The statistical analysis of the inoculation data showed a significant isolate x clone interaction. This data also suggest the possibility of vertical resistance, which is different to previous assumptions. Transfection studies (Chapter 4) involving a positive stranded RNA virus, Diaporthe RNA virus (DaRV) from a South African D. perjuncta isolate are presented here. In this study, a virus free D. perjuncta isolate, a virulent C. cubensis isolate and a hypovirulent C. cubensis isolate containing the hypovirus CHV1-EP713 were chosen to be transfected with DaRV. By using electroporation, it was possible to infect a virus free D. perjuncta isolate with the Diaporthe RNA virus, thus extending the transfection range of this virus. The resulting transfection led to altered colony morphology but did not lead to a reduction in pathogenicity. We were also not successful in attempts to transfect isolates of C. cubensis with DaRV, indicating that the virus does not replicate in this host. In a previous study a virulent South African C. cubensis isolate was transfected with the Cryphonectria parasitica hypovirus CHV1-EP713. This resulted in the fungus becoming hypovirulent. Chapter five of this thesis presents the results of a study to evaluate the potential use of this virus in the biological control of Cryphonectria canker in South Africa. A field trial was established and existing cankers were treated with the transfected isolate. The treatment of the cankers did not lead to a significant reduction in canker size, but did alter the morphology of the cankers. The virus was also shown to be transmitted via hyphal anastomosis to the virulent canker causing isolates. In addition the co-inoculation on single trees with both the virulent and virus-containing isolate, resulted in a significant reduction in the size of the lesions. This study also showed that the transfected C. cubensis isolate are characterised by significantly smaller lesions than those associated with the virulent, virus-free isolate. Cryphonectria cubensis and the associated canker disease of Eucalyptus threaten the forestry industry in South Africa. The overall aims of the studies presented in this thesis were to gain a more complete understanding of this fungus and to evaluate potential control strategies. Each of these chapters should contribute towards a better understanding of the viruses associated with C. cubensis and other important aspects of Cryphonectria canker, which will hopefully lead to enhanced control strategies of the disease in South Africa. / Thesis (PhD)--University of Pretoria, 2008. / Microbiology and Plant Pathology / PhD / Unrestricted

As formigas cortadeiras (Hymenoptera-Formicidae) estão distribuídas desde o sul dos Estados Unidos até a Argentina. São herbívoros comuns de florestas que coletam material vegetal para cultivar um fungo mutualista específico. São conhecidas pelo seu papel ecológico na aeração do solo, na infiltração da água e na ciclagem de nutrientes. Atividades humanas, como o desmatamento e a agricultura, afetam o ambiente, alterando também o comportamento das saúvas, que acabam atacando os espécimes cultivados. Devido aos prejuízos causados à agricultura, novos inseticidas específicos são necessários. Muitos trabalhos têm sido desenvolvidos usando metabólitos secundários para essa finalidade. Este trabalho tem como objetivo avaliar o efeito da cafeína na mortalidade de Atta sexdens rubropilosa (Forel, 1908) e no crescimento in vitro de seu fungo mutualístico Leucoagaricus gongylophorus (Möller) Singer (Leucocoprineae: Agaricaceae), obtidos de sauveiros mantidos em laboratório. Foram utilizadas quatro concentrações de cafeína, 0,01%, 0,05%, 0,10% e 0,50%. Mortalidade das formigas foi avaliada pelo ensaio de ingestão, acrescentando a cafeína a dietas artificiais sólidas. A cafeína foi incorporada ao meio de cultura para medir a sua influência no crescimento in vitro. Independente das concentrações de cafeína, esse metabólito parece atuar como repelente para a saúvas. A respeito do fungo, quanto maior a concentração de cafeína, menor o crescimento in vitro. Inibição do crescimento foi observada em 0,10% e 0,50% e morte do fungo foi observada em algumas amostras Em conclusão, os resultados sugerem que a cafeína pode ser usada como fungicida, sendo adicionada a iscas que serão coletadas pelas formigas e carregada aos ninhos, causando a redução do fungo e/ou a morte e consequentemente, a morte das formigas. / The leaf-cutting ants (Hymenoptera-Formicidae) are found from south of United States to Argentina. They are common florest herbivorous which collect plant material to feed a specific mutualist fungus. These insects are known by their ecological role at soil aeration, water permeation and nutrient cycling. Human activities, like deforestation and agriculture, affect the environment, affecting the behavior of leaf-cutting ants, which start to attack the crops. Due to crops lost, new specific pesticides are needed. Several researches have been developed using secondary metabolites for this purpose. The present work aimed to evaluate the effect of caffeine at Atta sexdens rubropilosa (Forel, 1908) mortality, and at in vitro growth of the mutualist fungus Leucoagaricus gongylophorus (Möller) Singer (Leucocoprineae: Agaricaceae), collected from laboratory nests. Four caffeine concentrations were tested: 0.01%, 0.05%, 0.10% and 0.50%. Ant\'s mortality was evaluated by ingestion assay, adding caffeine to artificial diets. Caffeine was added to culture medium, to measure its influence on in vitro fungus growth. Despite caffeine concentrations, this compound seems to act as repellent to ants. Concerning to the fungus, the higher the caffeine concentration, the lower the in vitro fungus growth. Growth inhibition was observed at both 0.10% and 0.50% concentrations and death of fungus was observed in some samples. In conclusion, the results suggest that caffeine could be used as fungicide, being added to baits which could be collected by ants and carried to the nests, causing fungus reduction and/or death and consequently, the death of the nests.

Orientador: Diego Cunha Zied / Resumo: O fungo Lecanicillium fungicola é um importante patógeno no cultivo de Agaricus bisporus, apresentando diversos sintomas e perdas de produção. O objetivo do trabalho foi avaliar interações in vivo e in vitro dos fungos Lecanicillium fungicola e Agaricus bisporus. Os experimentos in vitro foram feitos utilizando discos de micélio do patógeno e composto colonizado pelo A. bisporus. O composto colonizado também foi utilizado para a frutificação de cogumelos tanto em tamanho menor em placas de petri como em maior quantidade. Foram aplicados quatro suspensões de esporos L. fungicola nos cogumelos em placas de petri, de acordo com o seu tratamento. Não ocorreu a paralisação do crescimento micelial in vitro no momento em que o patógeno e o hospedeiro se encontram. Após 36 horas a inoculação do patógeno foi possível a visualização de manchas no píleo e após 60 horas a visualização de hifas e esporos. A linhagem coloração creme apresentou maior massa e diâmetro do píleo e menor porcentagem de rompimento do véu. A linhagem de A. bisporus de coloração branca obteve maior produtividade do que a linhagem de coloração creme. O isolado LF 19/03 apresentou maior agressividade em ambas as linhagens de A. bisporus. Os diversos sintomas encontrados no decorrer da pesquisa possibilitaram a confecção de uma escala diagramática para auxiliar o produtor na comercialização dos cogumelos doentes. / Abstract: The fungus Lecanicillium fungicola is an important pathogen in the cultivation of Agaricus bisporus, presenting several symptoms and production losses. The objective of the work was to evaluate interactions in vivo and in vitro of the fungi Lecanicillium fungicola and Agaricus bisporus. The in vitro experiments were done using mycelium discs of the pathogen and compound colonized by A. bisporus. The colonized compost was also used for the fruiting of mushrooms both in smaller size in petri dishes and in greater quantity. Four suspensions of L. fungicola spores were applied to the boxes according to their treatment. Mycelial growth did not stop in vitro at the time the pathogen and host meet. After 36 hours the inoculation of the pathogen made it possible to see spots on the cap and after 60 hours, to view hyphae and spores. The cream-colored lineage showed greater mass and diameter of the cap and lesser percentage of rupture of the veil. The white colored A. bisporus strain obtained higher productivity than the cream colored strain. The isolate LF 19/03 showed greater aggressiveness in both strains of A. bisporus. The various symptoms found in the course of the research enabled the production of a diagrammatic scale to assist the producer in the sale of sick mushrooms. / Mestre

<p>Per-and  polyfluoroalkyl  substances  (PFAS)  are  a  large  group  of  emerging  contaminants  that include astrong carbon-flourine bond that makes the compounds resistant to physical, chemical and  biological  degradation.  They  are  found  in  drinking  water  supplies,  daily  human  products, manufacturing  facilities,  and  in  areas  where  aqueous  film-forming  foam  (AFFF)was  used  to extinguish fires. Toxicity levels of these chemicals can vary depending on the characteristics of the specific chemical; longer carbon chain has shown to be more bioaccumulative and toxic than shorter chain length PFAS. Many studies have recognized perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA) to be a substantial concern due to their known toxicity to wildlife. For example, studies show strong evidence that PFOA and PFOS suppress the antibody response from animals. Due to adverse health effects and public concern, the U.S stopped perfluorooctanoic acid (PFOA) manufacturing and switched to the production of an alternative fluorinated compound known  as  hexafluoropropylene  oxide  (HFPO)  dimer  acid  or  GenX,  which  is  thought  to  beless bioaccumulative and therefore, potentially less toxic. These anthropogenic pollutants are one of many stressors acting on aquatic organisms like anurans. Natural stressors such as the devastating fungal  pathogen Batrachocytrium  dendrobatidis(Bd)  is  another  stressor  impacting  amphibian populations.  Despite the co-occurrence of these stressors, no studies have examined interactive effects of the fungal pathogen Bd and PFAS, or whether PFAS effects carry over into the terrestrial environment aftera larvae  exposure. This study  tested the growth and developmental effects of PFOS, PFOA, and GenX, on gray treefrog (Hyla versicolor) tadpoles, followed by a Bd challenge in metamorphs. Our results demonstrate that a PFAS larval exposure interacted witha terrestrial Bdchallenge to influence growth and development. Bdexposed animals were significantly shorter (smaller snout vent length)  and had a significant increase in body condition and mass. This is the first study to report effects on amphibian terrestrial life stages after larval exposure to PFAS and to report an increased sensitivity to Bd. The environmentally relevant concentrations tested in this study  (<10  parts  per  billion)  lend  ecological  significance  to  these  results  however,  additional studies are needed to understand the mechanisms behind these effects.</p>