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dc.contributor.authorRodrigues, Caio Junior Balduino Coutinho
dc.date.accessioned2023-12-21T18:41:53Z-
dc.date.available2023-12-21T18:41:53Z-
dc.date.issued2019-02-22
dc.identifier.citationRODRIGUES, Caio Junior Balduino Coutinho. Abordagens enzimáticas e proteômicas da resposta de fungos artropodopatogênicos submetidos a estresses e na infecção contra Rhipicephalus microplus. 2019. 96 f. Tese (Doutorado em Ciências Veterinárias) - Instituto de Veterinária, Departamento de Parasitologia Animal, Universidade Federal Rural do Rio de Janeiro, Seropédica, RJ, 2019.por
dc.identifier.urihttps://rima.ufrrj.br/jspui/handle/20.500.14407/9600-
dc.description.abstractNa última década, estudos envolvendo a aplicação de Metarhizium spp. e Beauveria bassiana no controle de Rhipicephalus microplus têm crescido de forma exponencial. O presente estudo avaliou a influência de repiques sucessivos em meio de cultivo sintético (batata-dextrose-ágar - BDA) na atividade enzimática e efeitos sobre virulência para R. microplus em três isolados de Metarhizium. Para isso, amostras foram obtidas do cultivo inicial (Início) e após o décimo (10º ciclo) e vigésimo (20º ciclo) repiques. Apresentando consequências individualizadas entre os isolados, alterações na macromorfologia, diminuição do crescimento radial e produção conidial, além de redução de virulência e atividades de Pr1, Pr2 e lipase foram majoritariamente detectadas após o 20º ciclo. Em outra vertente, foi-se também estudado o efeito de um agente oxidante (menadiona - MEN), potente indutor de espécies reativas de oxigênio, no desenvolvimento, virulência para R. microplus, atividade enzimática e secreção proteica de M. anisopliae sensu stricto (s.s.). Por ser pouco tolerante, para todos os experimentos, uma subdosagem (50% da CL50) de MEN foi inicialmente adicionada aos cultivos. Em meio sólido suplementado com MEN, mudanças relevantes no desenvolvimento das colônias, além de atividades enzimáticas conidiais foram detectadas. Em contrapartida, de forma surpreendente, melhorias significativas no percentual de mortalidade de larvas também ocorreram. Nas análises de meio basal líquido acrescido de cutícula de R. microplus, num total de 654 proteínas identificadas, 31 foram up- e down-reguladas, estando principalmente relacionadas à atividade antioxidante (catalase), patogenicidade (como Pr1B, Pr1D e Pr1K), reparo celular e morfogênese. Por fim, a atividade e secreção de proteínas (secretoma) de Beauveria bassiana s.l. durante a infecção em R. microplus e sua comparação com condições normais de crescimento foram reportadas. A partir de meio basal líquido acrescido de cutícula de R. microplus, e aplicando uma tecnologia multidimensional de identificação de proteínas, de um total de 236 proteínas, 32 proteínas foram identificadas em ambas as condições de estudo e 50 exclusivamente na condição de infecção, sendo estas atribuídas a diferentes aspectos como adesão ao hospedeiro, penetração cuticular, defesa fúngica e estresse. Nesse contexto, um maior número de proteínas detectadas com atividade hidrolítica, classe enzimática de todas as enzimas degradadoras de cutícula, como lipases e proteases, foram identificadas e validadas através de ensaios enzimáticos. Em suma, destaca-se que o repiques sucessivos e o estresse oxidativo, assim como o desconhecimento dos principais metabólitos produzidos durante a infecção por B. bassiana, ainda limitam prospecções futuras para o biocontrole, em especial à aplicação e ao monitoramento de produtos à base de fungos artropodopatogênicos no uso para carrapatos.por
dc.description.sponsorshipCNPq - Conselho Nacional de Desenvolvimento Científico e Tecnológicopor
dc.description.sponsorshipCAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superiorpor
dc.formatapplication/pdf*
dc.languageporpor
dc.publisherUniversidade Federal Rural do Rio de Janeiropor
dc.rightsAcesso Abertopor
dc.subjectMetarhizium spp.por
dc.subjectBeauveria bassianapor
dc.subjectsubcultivopor
dc.subjectmenadionapor
dc.subjectMudPpor
dc.subjectsubculturingeng
dc.subjectmenadioneeng
dc.subjectMudPITeng
dc.titleAbordagens enzimáticas e proteômicas da resposta de fungos artropodopatogênicos submetidos a estresses e na infecção contra Rhipicephalus micropluspor
dc.title.alternativeEnzymatic and proteomic approaches to the response of arthropod-pathogenic fungi under stress conditions and the infection against Rhipicephalus micropluseng
dc.typeTesepor
dc.description.abstractOtherStudies using Metarhizium spp. and Beauveria bassiana to control Rhipicephalus microplus have exponentially increased over the last decade. Here, it was reported the subculturing of three Metarhizium isolates in synthetic culture medium (potato-dextrose-agar - PDA) and its effect on enzymatic activity and virulence to R. microplus. For this, samples were collected from the first culture (Initial) and after the tenth (10th cycle) and twentieth (20th cycle) growth. Changes in macromorphology, decrease in radial growth and conidia production, as well as reduction of virulence and Pr1, Pr2 and lipase activities, were detected after the 20th cycle. Next in order, the effects of a potent oxidizing agent (menadione - MEN) were explored in the development, virulence to R. microplus, enzymatic activity and protein secretion of M. anisopliae sensu stricto (s.s.). With low tolerance to MEN, a sub dose (half of LC50) was initially added to the cultures for all experiments. In solid medium supplemented with MEN, significant changes in the macromorphology of colonies and conidial enzymatic activities were detected. In contrast, surprisingly, significant improvements in the percentage of larval mortality also occurred. In basal medium (BM) plus R. microplus cuticle, in 654 identified proteins, 31 were up- and down-regulated, mainly related to antioxidant activity (catalase), pathogenicity (Pr1B, Pr1D and Pr1K), cell repair and morphogenesis. Finally, the activity and secretion of proteins (secretoma) of Beauveria bassiana s.l. during Rhipicephalus microplus infection versus normal growth conditions were described. Applying a multidimensional protein identification technology with samples from BM plus R. microplus cuticle, up to 236 proteins were characterized, being 32 identified in both conditions and 50 exclusively in the infection condition. All proteins were related to different aspects such as host adhesion, cuticular penetration, fungal defense and stress. In this context, a higher number of proteins detected with hydrolysis activity, enzymatic class of all cuticle-degrading enzymes, such as lipases and proteases, were identified and validated through enzymatic assays. In summary, subculturing and oxidative stress, as well as the lack of knowledge about the main metabolites produced during B. bassiana infection, still limit the perspectives of biocontrol, especially the application and the environmental track of products based on arthropodopathogenic fungi used in tick control.eng
dc.contributor.advisor1Bittencourt, Vânia Rita Elias Pinheiro
dc.contributor.advisor1IDCPF: 505.198.676-53por
dc.contributor.advisor-co1Silva, Walter Orlando Beys da
dc.contributor.advisor-co1IDCPF: 982.298.000-06por
dc.contributor.advisor-co2Santi, Lucélia
dc.contributor.advisor-co2IDCPF: 709.760.310-00por
dc.contributor.referee1Bezerra, Simone Quinelato
dc.contributor.referee2Gôlo, Patrícia Silva
dc.contributor.referee3Cardoso, Cristiane Martins
dc.contributor.referee4Fernandes, Éverton Kort Kamp
dc.creator.IDCPF: 124.958.287-36por
dc.creator.Latteshttp://lattes.cnpq.br/9252122827094001por
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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