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Copepoda, Calanoida) UNIVERSIDADE DE SÃO PAULO ESCOLA DE ENGENHARIA DE SÃO CARLOS PROGRAMA DE PÓS-GRADUAÇÃO EM CIÊNCIAS DA ENGENHARIA AMBIENTAL DENISE TIEME OKUMURA Estudos ecotoxicológicos com as espécies Argyrodiaptomus furcatus e Notodiaptomus iheringi (Copepoda, Calanoida) Orientadora : Profa. Tit. Odete Rocha São Carlos - SP 2011 DENISE TIEME OKUMURA Estudos ecotoxicológicos com as espécies Argyrodiaptomus furcatus e Notodiaptomus iheringi (Copepoda, Calanoida) Tese apresentada à Escola de Engenharia de São Carlos, da Universidade de São Paulo, como parte dos requisitos para obtenção do título de Doutor em Ciências – Programa em Ciências da Engenharia Ambiental. Orientadora : Profa. Tit. Odete Rocha São Carlos - SP 2011 Dedico este trabalho a meus pais, a minha Família. AGRADECIMENTOS À minha orientadora, Profa. Dra. Odete Rocha, pelo apoio, estímulo e, principalmente, pela orientação com que pude contar e valiosas lições que pude aprender. À FAPESP pelo apoio financeiro, por meio da concessão da bolsa de Doutorado. À doutoranda Erika Cavalcante-Silva e seu orientador Prof. Dr. João Dias de Toledo Arruda Neto, do Programa de Pós-Graduação Interunidades em Biotecnologia da Universidade de São Paulo (USP/Instituto Butantan/IPT), pelo trabalho de cooperação dentro do projeto de pesquisa “Estudo da ação combinada de radiações ionizantes com campos elétricos e magnéticos em cianobactérias - viabilidade celular e atenuação de toxinas” (Processo FAPESP nº 2006/60096-8). À amiga e colega que tornou essa fase mais fácil, muito mais produtiva. Obrigada, Ferrr. Às meninas do laboratório de Ecotoxicologia, Erika, Amanda, Laira e Emanuela, pelos momentos de descontração. À Edna pela boa vontade e carinho com que sempre me ajudou quando solicitada. Ao Zezinho, Luis, Airton e Alcídio pela boa vontade sempre demonstrada nos trabalhos técnicos realizados em vários momentos durante este trabalho. À Roberta, Toshiro, Cintia, Naty, Lidiane, Raphael, Ana Lúcia, Patrícia, Edna, Katy, Renatinha, Andressa, Tiago, Marco e Renata pelas ajudas, bate-papos e amizade. Às minhas amigas Paty Rabs, Mari, Mawusi, Grá, Fer, Rô, Ana, Pat, Carol, Edna. A todos os funcionários e estagiários do Depto. de Ecologia e Biologia Evolutiva/UFSCar e do CRHEA/USP. Ao meu pai, João, porque me ensinou que tudo é possível quando se tem vontade de vencer. À minha mãe, Marina, porque sempre acreditou em mim e me incentivou ao longo da vida. Aos meus queridos irmãos, Rafa e Erika. À minha Família. A todos que direta ou indiretamente contribuíram para a realização deste trabalho. “Mude suas opini ões, sustente seus princ ípios; troque suas folhas, mantenha intacta suas ra ízes” (Victor Hugo) SUMÁRIO RESUMO __________________________________________________________________ i ABSTRACT _______________________________________________________________ ii LISTA DE TABELAS _______________________________________________________ iii LISTA DE FIGURAS _______________________________________________________ xi 1. INTRODUÇÃO __________________________________________________________ 1 2. JUSTIFICATIVA ________________________________________________________ 3 3. REVISÃO BIBLIOGRÁFICA ______________________________________________ 4 3.1. Descrição das espécies ________________________________________________________ 7 3.2. Os calanoides como organismos-teste em estudos ecotoxicológicos __________________ 11 4. OBJETIVOS ___________________________________________________________ 13 4.1. Objetivos Específicos ________________________________________________________ 13 5. HIPÓTESES ___________________________________________________________ 14 6. MATERIAL E MÉTODOS ________________________________________________ 15 6.1. Origem do material biológico _________________________________________________ 16 6.1.1. Descrição dos Corpos de Água de Origem das Espécies ____________________ 16 6.2. Manutenção e cultivo dos Copepoda Calanoida Argyrodiaptomus furcatus e Notodiaptomus iheringi _______________________________________________________________________ 19 6.3. Avaliação dos aspectos biológicos e ecológicos das espécies de Argyrodiaptomus furcatus e Notodiaptomus iheringi __________________________________________________________ 23 6.3.1. Desenvolvimento Embrionário e Pós-Embrionário ________________________ 23 6.3.2. Parâmetros da Dinâmica Populacional _________________________________ 26 6.4. Testes ecotoxicológicos com Argyrodiaptomus furcatus e Notodiaptomus iheringi _______ 28 6.4.1. Substâncias de Referência ___________________________________________ 29 6.4.2. Amostras Ambientais _______________________________________________ 34 6.4.3. Composto Tóxico Orgânico: Cianotoxinas ______________________________ 38 7. RESULTADOS _________________________________________________________ 42 7.1. Aspectos biológicos e ecológicos das espécies de Argyrodiaptomus furcatus e Notodiaptomus iheringi _______________________________________________________________________ 42 7.1.1. Descrição Morfológica _____________________________________________ 42 7.1.2. Desenvolvimento Embrionário e Pós-Embrionário _______________________ 43 7.1.3. Duração do Tempo de Desenvolvimento Embrionário e Pós-Embrionário _____ 50 7.1.4. Taxas de Crescimento Populacional ___________________________________ 63 7.2. As espécies de Copepoda como organismos-teste: A sensibilidade das espécies Argyrodiaptomus furcatus e Notodiaptomus iheringi a compostos de referência ____________ 68 7.2.1. Sais ____________________________________________________________ 69 7.2.3. Surfactantes _____________________________________________________ 107 7.2.4. Amostras Ambientais _____________________________________________ 124 7.2.5. Composto Tóxico Orgânico: Cianotoxinas _____________________________ 131 8. DISCUSSÃO __________________________________________________________ 133 8.1. Experimentos de ciclo de vida e crescimento populacional obtidos em laboratório ____ 133 8.2. Ensaios ecotoxicológicos com as espécies Argyrodiaptomus furcatus e Notodiaptomus iheringi ______________________________________________________________________ 141 9. CONCLUSÕES ________________________________________________________ 154 10. BIBLIOGRAFIA ______________________________________________________ 157 APÊNDICE A ___________________________________________________________ 172 APÊNDICE B - Sais: Argyrodiaptomus furcatus _______________________________ 175 APÊNDICE C - Sais:Notodiaptomus iheringi __________________________________ 186 APÊNDICE D - Metais:Argyrodiaptomus furcatus _____________________________ 207 APÊNDICE E - Metais: Notodiaptomus iheringi _______________________________ 223 APÊNDICE F - Surfactantes: Argyrodiaptomus furcatus ________________________ 254 APÊNDICE G - Surfactantes: Notodiaptomus iheringi __________________________ 265 APÊNDICE H - Composto Tóxico Orgânico: Argyrodiaptomus furcatus ____________ 286 APÊNDICE I - Composto Tóxico Orgânico: Notodiaptomus iheringi ______________ 292 APÊNDICE J - Proposta de protocolo para realização de testes agudos com Copepoda Calanoida de água doce como organismos-teste ________________________________ 298 i RESUMO OKUMURA, D.T. Estudos ecotoxicológicos com as espécies Argyrodiaptomus furcatus e Notodiaptomus iheringi (Copepoda, Calanoida) . 2011. 312 f. Tese (Doutorado) – Escola de Engenharia de São Carlos, Universidade de São Paulo, São Carlos, 2011. Os Copepoda constituem a maior classe de pequenos crustáceos, com mais de 8.500 espécies descritas. Dentre as cinco ordens de Copepoda de vida livre, as espécies pertencentes à ordem Calanoida apresentam um forte endemismo, com uma restrita distribuição geográfica. Além disso, os Copepoda Calanoida são particularmente sensíveis a contaminantes contidos na coluna de água devido ao seu hábito planctônico. No presente trabalho foram realizados estudos ecotoxicológicos com duas espécies de Copepoda Calanoida, particularmente com as espécies Argyrodiaptomus furcatus Sars, 1901 e Notodiaptomus iheringi Wright, 1935. Estas espécies foram comparadas com relação à sensibilidade e desempenho como organismos-teste na avaliação da qualidade da água em ecossistemas de água doce. Foram testadas e adaptadas metodologias de cultivo em laboratório, e foram estudados os principais aspectos da biologia destes em condições ambientais controladas. Além disso, foram analisados aspectos referentes à dinâmica populacional. A sensibilidade das duas espécies de Calanoida às substâncias de referência sódio, potássio e zinco na forma de cloreto (NaCl, KCl e ZnCl 2, respectivamente), ao dicromato de potássio (K 2Cr 2O7) e ao sulfato de alumínio (Al 2(SO 4)3), aos surfactantes dodecilsulfato de sódio (C 12 H25 NaO 4S) e ácido dodecil benzenosulfonato de sódio (CH 3(CH 2)11 C6H4SO 3Na), foram avaliadas. Além disso, avaliou-se a toxicidade aguda de diferentes amostras ambientais e das toxinas da cianobactéria Microcystis aeruginosa após tratamento por irradiação. Os resultados dos testes ecotoxicológicos foram expressos em concentração efetiva (CE(I)50). Para o cálculo estatístico da CE50 foi utilizado o programa Trimmed Spearman-Karber. Os resultados relativos ao ciclo de vida mostraram que N. iheringi tem um menor tempo de desenvolvimento do que A. furcatus , com tempos de duração do desenvolvimento do ovo determinados pelos métodos indireto e direto. Verificou-se que a espécie de N. iheringi , de menor tamanho, apresentou tempo de desenvolvimento pós-embrionário menor do que a espécie de maior tamanho, A. furcatus . As taxas intrínsecas de crescimento populacional foram mais elevadas para a
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