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A Sample Article Title MINISTÉRIO DA SAÚDE FUNDAÇÃO OSWALDO CRUZ INSTITUTO OSWALDO CRUZ Doutorado no Programa de Pós-Graduação em Biologia Parasitária MORPHOLOGICAL, MOLECULAR AND ECOLOGICAL INTEGRATIVE TAXONOMY OF ACANTHOCEPHALA (ARCHIACANTHOCEPHALA) PARASITE OF BRAZILIAN WILDLIFE MAMMALS ANA PAULA NASCIMENTO GOMES Rio de Janeiro Maio de 2019 INSTITUTO OSWALDO CRUZ Programa de Pós-Graduação em Biologia Parasitária ANA PAULA NASCIMENTO GOMES Morphological, molecular and ecological integrative taxonomy of Acanthocephala (Archiacanthocephala) parasite of Brazilian wildlife mammals Tese apresentada ao Instituto Oswaldo Cruz como parte dos requisitos para obtenção do título de Doutor em Ciências Orientador (es): Prof. Dr. Arnaldo Maldonado Júnior Prof. Dra. Natalie Olifiers RIO DE JANEIRO Maio de 2019 ii INSTITUTO OSWALDO CRUZ Programa de Pós-Graduação em Biologia Parasitária AUTOR: ANA PAULA NASCIMENTO GOMES MORPHOLOGICAL, MOLECULAR AND ECOLOGICAL INTEGRATIVE TAXONOMY OF ACANTHOCEPHALA (ARCHIACANTHOCEPHALA) PARASITE OF BRAZILIAN WILDLIFE MAMMALS ORIENTADOR (ES): Prof. Dr. Arnaldo Maldonado Júnior Prof. Dra. Natalie Olifiers Aprovada em: _____/_____/_____ EXAMINADORES: Prof. Dra. Cláudia Portes dos Santos – Presidente (Instituto Oswaldo Cruz - Fiocruz/RJ) Prof. Dr. Luiz Claudio Muniz Pereira (Instituto Oswaldo Cruz - Fiocruz/RJ) Prof. Dr. Eduardo José Lopes Torres (Universidade do Estado do Rio de Janiero - UERJ) Prof. Dra. Alena Mayo Iñiguez (Instituto Oswaldo Cruz - Fiocruz/RJ) Prof. Dra. Raquel de Oliveira Simões (Universidade Federal do Rio de Janiero – UFRRJ) Rio de Janeiro, 06 de maio de 2019 1 Anexar a cópia da Ata que será entregue pela SEAC já assinada. 2 Dedico a minha família, aos meus pais Maria Natividade e Sérgio Otávio, a minha irmã Cecília e meu namorado Erick que sempre me apoiaram e acreditam em mim. 3 AGRADECIMENTOS Gostaria de agradecer a minha família, ao meu pai Sérgio Otávio V. Gomes, minha mãe Maria Natividade N. Gomes e minha irmã Cecília N. Gomes pelo carinho, amor e paciência neste período que deselvolvi minha tese. Minha família sempre me mostrou o caminho do otimismo, com palavras que me levantaram no momento que me faltava ânimo para continuar a desenvolver este trabalho. Em especial, agradeço a minha irmã que sempre esteve ao meu lado em todos os momentos, de alegria e de choro neste período, inclusive em congressos, compartilhando seus conhecimentos pedagógicos, seu carinho e orgulho de eu me tornar uma pesquisadora. Agradeço também meu irmão de coração Diego de Souza que desde a graduação torceu e me apoiou até chegar neste momento que sempre sonhei. Agradeço a meu namorado Erick Castillo pelo carinho, paciência e por me compreender nos momentos mais difíceis nesta fase. Uma pessoa que tem me ensinado a ter auto-controle, ser menos ansiosa e também me ensina a cada vez mais a acreditar no meu sonho. Agradeço meu orientador Dr. Arnaldo Maldonado por me dar oportunidade de realizar este trabalho, por me ensinar a ser independente e compartilhar seus conhecimentos acadêmicos e de vida pessoal. A minha orientadora Dra. Natalie Olifiers que sempre acreditou no meu potencial. Agradeço pelo carinho, amizade e confiança durante toda minha formação desde a iniciação cientifica até este momento. Agradeço aos meus amigos do LABPRM, principalmente a Taina Monte, ao Thiago Cardoso, Bernardo Teixeira, Natalia Costa, Beatriz Elise, Raquel Simões, Raquel Gonzales, Joyce Souza, Michele Maria, Sócrates Neto, Juliana São Luis, Karina Varela, Renata Souza, Camila Lucio e Rute pelo carinho, pelas conversas, pelo momentos de risadas, pelo apoio na execução do trabalho, pelo apoio emocional e também em compartilhar conhecimentos. As minhas colaboradoras Dra. Rita Bianchi e Clarice Cesário pelo auxílio e amizade. Aos colegas e funcionários do LABPMR pela ajuda e carinho durante estes quatro anos de trabalho. 4 Agradeço ao Dr. Roberto Vilela por ensinar e compartilhar o conhecimento de filogenia molecular, por me ajudar nas análises e pela paciência em discutir meus resultados. Agardeço a Dra. Daniela Lopes e Dr. Marcelo Knoff que me atenderam com profissionalismo no empréstimo dos materiais na Coleção Helmintológica do Instituto Oswaldo Cruz (CHIOC). Agradeço ao Ricardo Baptista por por ter me ajudado na parte de imagens e montagem das pranchas finais. Agradeço a secretária Sra. Rita Gomes por sempre atender com carinho e paciência, ajudando em todo momento de dúvidas para execução do trabalho. À Dra. Helene Santos Barbosa e Sra. Sandra Maria de Oliveira pela disponibilidade e ajuda no protocolo de preparação das amostras para Microscopia Eletrônica de Varredura. À equipe da Plataforma de Microscopia Eletrônica de Varredura Rudolf Barth, em especial ao Sr. Roger que me atendeu com profissionalismo, auxiliando nas analises do material e produção das imagens. À equipe da Plataforma de Sequenciamento de DNA PDTIS/FIOCRUZ pelo atendimento e execução de qualidade para obtenção das sequencias. 5 “There is a driving force more powerful than steam, electricity and nuclear power: the will” Albert Einstein “Há uma força motriz mais poderosa que o vapor, a eletricidade e a energia atômica: a vontade” Albert Einstein 6 INSTITUTO OSWALDO CRUZ TAXONOMIA INTEGRATIVA, MORFOLÓGICA, MOLECULAR E ECOLÓGICA DE ACANTHOCEPHALA (ARCHIACANTHOCEPHALA) PARASITOS DE MAMÍFEROS SILVETRES BRASILEIROS RESUMO TESE DE DOUTORADO EM BIOLOGIA PARASITÁRIA Ana Paula Nascimento Gomes O filo Acanthocephala é caracterizado por não possuir trato digestório e por apresentar na região anterior uma probóscide munida de ganchos que retrai-se para dentro de um receptáculo. Este grupo é dividido em quatro classes Archiacanthocephala, Palaeacanthocephala, Eoacanthocephala e Polyacanthocephala baseado em características morfológicas, biológicas e ecológicas. Dentre os filos dos helmintos estudados em mamíferos brasileiros, o filo Acanthocephala se destaca por apresentar lacunas no que se refere às informações taxonômicas, filogenéticas e ecológicas. O objetivo geral deste trabalho foi realizar a taxonomia integrativa dos acantocéfalos recuperados em mamíferos das famílias Procyonidae, Myrmecophagidae e Cricetidae de diferentes regiões geográficas do Brasil, armazenados e disponibilizados pela coleção do Laboratório de Biologia e Parasitologia de mamíferos Silvestres Reservatórios (LABPMR) utilizando características morfologicas, moleculares e ecológicas. Os acantocéfalos recuperados foram identificados através da microscopia de luz (ML) e por microscopia eletrônica de varredura (MEV). Foi também realizada a análise filogenética molecular dos acantocéfalos com os marcadores moleculares do gene ribossomal da subunidade maior (28S rRNA) e do gene mitocondrial citocromo oxidase da subunidade 1 (MT-CO1). Além disto, foi determinada a prevalência e abundância dos ovos de Acanthocephala através da análise coproparasitológica de fezes de quati Nasua nasua e de cachorro-do-mato Cerdocyon thous, avaliando a influencia dos fatores bióticos e abióticos na infecção. Os espécimes de acantocéfalos foram descritos e identificados em duas novas espécies Pachysentis n. sp. (Archiacanthocephala: Oligacanthorhynchidae) parasitando Nasua nasua (quati) proveniente do Mato Grosso do Sul do bioma Pantanal e Moniliformis n. sp. (Archiacanthocephala: Moniliformidae) em Necromys lasiurus (ratinho-do-cerrado) da região de Uberlândia, Minas Gerais do bioma Cerrado; e redescrita a espécie Gigantorhynchus echinodiscus (Archiacanthocephala: Gigantorhynchidae) em Myrmecophaga tridactyla (Tamanduá-bandeira) da Estação Ecológica Santa Bárbara, São Paulo, bioma cerrado. As análises filogenéticas moleculares sugeriram que a espécie G. echinosdichus está relacionada com Mediorhynchus sp. formando um grupo monofilético, assim como Moniliformis n. sp. está relacionado com as espécies do gênero Moniliformis também formando grupo monofilético. A análise ecológica foi realizada com 118 amostras fecais de 55 espécimes de cachorro-do-mato e 72 amostras fecais de 61 espécimes de quatis sugerindo a influência da sazonalidade na abundância dos acantocéfalos para ambos os hospedeiros e que os atributos relacionados ao hospedeiro como sexo e idade também constituíram fatores importantes associados à prevalência e às cargas parasitárias. O presente trabalho acrescentou informações morfológicas, moleculares e ecológicas, enfatizando a importância de adotar abordagem da taxonomia integrativa nos estudos com Acanthocephala. 7 INSTITUTO OSWALDO CRUZ MORPHOLOGICAL, MOLECULAR AND ECOLOGICAL INTEGRATIVE TAXONOMY OF ACANTHOCEPHALA (ARCHIACANTHOCEPHALA) PARASITE OF BRAZILIAN WILDLIFE MAMMALS ABSTRACT PHD THESIS IN PARASITE BIOLOGY Ana Paula Nascimento Gomes The phylum acanthocephala is characterized by the presence of a proboscis armed with hooks, which retracts into receptacle, and lack of alimentary tract. This group is divided in four classes Archiacanthocephala, Palaeacanthocephala, Eoacanthocephala and Polyacanthocephala based on morphological, biological and ecological characteristics. Among the helminths studied in Brazilian mammals, the phylum Acanthocephala have a lack of taxonomic, phylogenetic and ecological information. The aim of the present work was to perform the integrative taxonomy of acanthocephalans recovered in mammals of the family Procyonidae, Myrmecophagidae and Cricetidae from different geographic regions, store and made available by the Laboratory of Biology and Parasitology of Wild Reservoirs Mammal (LABPMR) using morphological, molecular and ecological characteristics. The recovered acanthocephalans were identified by
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