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dc.contributor.authorPerinotto, Wendell Marcelo de Souza
dc.date.accessioned2023-12-21T18:43:59Z-
dc.date.available2023-12-21T18:43:59Z-
dc.date.issued2013-03-05
dc.identifier.citationPERINOTTO, Wendell Marcelo de Souza. Estudo da atividade enzimática e patogênica de isolados brasileiros de Metarhizium anisopliae s.l. visando o uso no controle biológico de Rhipicephalus (Boophilus) microplus. 2013. 67 f. Tese (Doutorado em Ciências Veterinárias) - Instituto de Veterinária, Universidade Federal Rural do Rio de Janeiro, Seropédica, 2013.por
dc.identifier.urihttps://rima.ufrrj.br/jspui/handle/20.500.14407/9770-
dc.description.abstractO fungo Metarhizium anisopliae sensu lato é um dos micro-organismos mais promissores a ser utilizado no controle de carrapato, e por isso, vem sendo estudado em todo o mundo, demonstrando potencial acaricida, principalmente em testes in vitro. Entretanto, tem sido observada variação na virulência entre diferentes isolados deste fungo. Nessa perspectiva, o presente estudo teve como objetivos verificar a ação patogênica em suspensões aquosas e em diferentes formulações oleosas e também analisar a ação enzimática de isolados brasileiros de M. anisopliae s.l. com a finalidade de selecionar isolados mais virulentos para utilização em futuros programas de biocontrole de carrapatos. Além disso, pesquisar a presença de partículas virais de dsRNA e associar as atividades enzimáticas, inclusive da subtilisina Pr 1 secretadas pelos isolados de M. anisopliae s.l. com os percentuais de controle, visando estabeler marcadores moleculares de virulência específicos para o carrapato R. microplus. Para tanto, cinco linhagens de M. anisopliae s.l. isolados de diferentes estados brasileiros foram analisadas. Primeiramente foi realizado um bioensaio para testar a virulência destes isolados sobre fêmeas alimentadas de R. microplus. Concomitantemente foram realizados ensaios enzimáticos para quantificar as atividades de lipases, proteases e quitinases secretadas pelos diferentes isolados de M. anisopliae s.l. cultivados em meio líquido contendo cutícula de carrapato e estearato de colesteril, através da utilização de substratos cromogênicos e fluorescentes. Para a pesquisa de micovírus, foi realizada a extração do ácido nucléico total que foi submetido à digestão por uma endonuclease S1 e uma RNAse A e posteriormente visualizado em gel de agarose. Além desses ensaios, foi realizado um bioensaio utilizando diferentes formulações oleosas, uma a base de óleo vegetal e outra com óleo mineral, ambas a 10%. Neste experimento, fêmeas de R. microplus completamente alimentadas foram imersas por três minutos nos respectivos tratamentos, a eficácia dos tratamentos foi avaliada através de alterações provocadas nos parâmetros reprodutivos destas fêmeas. O estudo demonstrou que há variação na virulência entre os isolados brasileiros de M. anisopliae s.l. para fêmeas alimentadas de R. microplus, além disso, ficou evidenciado que os isolados que apresentaram os maiores percentuais de controle foram também os que tiveram as maiores atividade lipolítica, proteolítica e quitinolítica. Pelo fato da enzima Pr 1 ser sabidamente envolvida no processo de infecção de M. anisopliae s.l., os resultados aqui encontrados demonstraram a possibilidade de usar a atividade desta enzima como um potencial marcador molecular para R. microplus, visto que os isolados que apresentaram maior atividade de Pr 1 foram os mais virulentos para esta espécie de carrapato. Entretanto, não foi evidenciada nenhuma relação entre a virulência com a infecção por partículas de dsRNA, sendo que dos cinco isolados testados, somente um não estava infectado. A partir do bioensaio utilizando formulações oleosas, ficou evidenciada que as formulações contendo óleo mineral foram mais eficientes do que as que continham óleo vegetal, proporciondo os melhores percentuais de controle. Além disso, pode-se concluir que os isolados CG 148 e CG 629 quando em formulação de óleo mineral apresentam alto potencial de controle para fêmeas ingurgitadas de R. micropluspor
dc.description.sponsorshipCoordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPESpor
dc.formatapplication/pdf*
dc.languageporpor
dc.publisherUniversidade Federal Rural do Rio de Janeiropor
dc.rightsAcesso Abertopor
dc.subjectfungos artropodopatogênicospor
dc.subjectvirulênciapor
dc.subjectcontrole biológicopor
dc.subjectarthropod-pathogenic fungieng
dc.subjectvirulenceeng
dc.subjectand biological controlpor
dc.titleEstudo da atividade enzimática e patogênica de isolados brasileiros de Metarhizium anisopliae s.l. visando o uso no controle biológico de Rhipicephalus (Boophilus) micropluspor
dc.title.alternativeStudy of the enzymatic activity and pathogenic of Metarhizium anisopliae s.l. Brazilian isolates targeting the biological control of Rhipicephalus (Boophilus) micropluseng
dc.typeTesepor
dc.description.abstractOtherThe fungus Metarhizium anisopliae sensu lato (s.l.) is one of the most promising micro-organisms to be used in the biological control of ticks. This fungus has been studied worldwide showing accurate acaricidal potential, especially in in vitro tests. However, a large variation on the virulence of different fungal isolates has been observed. From this perspective, the present study aimed to verify the pathogenic action of different fungal formulations (oil-based and aqueous) on ticks and also analyze the enzymatic action of M. anisopliae s.l. isolates from different Brazilian geographical regions in order to select the most virulent isolates for further use in programs of biological control of ticks. The presence of dsRNA viral particles (mycovirus) associated to the fungal enzymatic activities was also studied. Among the studied enzymes was the subtilisin Pr 1; whose enzymatic activity was associated with the percentage of tick control caused by the fungi aiming establish virulence markers specific for Rhipicephalus microplus ticks. Five M. anisopliae s.l. isolates from different Brazilian states were analyzed. First, a bioassay was performed on R. microplus engorged females to test the virulence of these fungal isolates. Concomitant assays were performed to quantify the enzymatic activity of lipases, proteases and chitinases secreted by the same M. anisopliae s.l. isolates. Fungi were grown in liquid medium containing ticks’ cuticle and cholesteryl stearate; fluorescent and chromogenic substrates were used to quantify the enzymatic activity. Mycovirus survey was carried out following steps of total nucleic acid extraction before an endonuclease S1 and RNAse A digestion and consecutive visualization on agarose gel. Furthermore, a bioassay was conducted using different oil formulations (10 % vegetable oil and 10% mineral oil). In this experiment, R. microplus engorged females were immersed in the respective formulations for three minutes. The efficacy of the treatments was assessed through analysis in the females’ reproductive parameters. The present study showed that there is variation in the virulence among M. anisopliae s.l. isolates against R. microplus, moreover, it was evident that the isolates that yielded the highest tick control percentages were also those which had the highest lipolytic, proteolytic and chitinolytic activities. Pr1 is known to be involved in the infection process of M. anisopliae s.l.; accordingly the present results demonstrate that this enzyme can be used as a molecular marker for R. microplus, since the isolates that showed higher Pr 1 activity were also the most virulent for this tick species. However, there was no evident analogy between the infection with dsRNA virulent particles and the enzymatic activity levels of the five isolates tested, considering that only one was not infected. Regarding the bioassay with oil-based formulations, it was clear that formulation containing mineral oil were more effective than those containing vegetable oil which provided low percentage of tick control. Furthermore, it can be concluded that the isolates GC 148 and GC 629 have a high potential to control R. microplus engorged females when formulated in mineral oileng
dc.contributor.advisor1Bittencourt, Vânia Rita Elias Pinheiro
dc.contributor.advisor1ID505.198.676-53por
dc.contributor.advisor1Latteshttp://lattes.cnpq.br/3888832724995864por
dc.contributor.advisor-co1Salles, Cristiane Martins Cardoso de
dc.contributor.advisor-co1ID035.399.287-90por
dc.contributor.advisor-co1Latteshttp://lattes.cnpq.br/3610279707231709por
dc.contributor.advisor-co2Silva, Walter Orlando Beys da
dc.contributor.advisor-co2Latteshttp://lattes.cnpq.br/1353100437268465por
dc.contributor.referee1Costa, Gisela Lara da
dc.contributor.referee2Reck Júnior, José
dc.contributor.referee3Famadas, Kátia Maria
dc.contributor.referee4Coumendouros, Katherina
dc.creator.ID306.937.188-46por
dc.creator.Latteshttp://lattes.cnpq.br/1812730341180904por
dc.publisher.countryBrasilpor
dc.publisher.departmentInstituto de Veterináriapor
dc.publisher.initialsUFRRJpor
dc.publisher.programPrograma de Pós-Graduação em Ciências Veterináriaspor
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dc.subject.cnpqMedicina Veterináriapor
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