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Phylum Myxozoa) Infecting the Aquatic Fauna

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Phylum Myxozoa) Infecting the Aquatic Fauna ULTRASTRUCTURAL AND MOLECULAR DESCRIPTION OF SOME MYXOSPOREANS (PHYLUM MYXOZOA) INFECTING THE AQUATIC FAUNA SÓNIA RAQUEL OLIVEIRA ROCHA Dissertation for Master in Marine Sciences – Marine Resources November 2011 i ii SÓNIA RAQUEL OLIVEIRA ROCHA ULTRASTRUCTURAL AND MOLECULAR DESCRIPTION OF SOME MYXOSPOREANS (PHYLUM MYXOZOA) INFECTING THE AQUATIC FAUNA Dissertation for Master’s degree in Marine Sciences – Marine Resources submitted to the Institute of Biomedical Sciences Abel Salazar, University of Porto. Supervisor – Doctor Carlos Azevedo Category – “Professor Catedrático Jubilado” Affiliation – Institute of Biomedical Sciences Abel Salazar, University of Porto. ii Nota prévia Declaro que, como autora desta tese, estive envolvida na realização de todos os procedimentos laboratoriais conduzentes à obtenção dos resultados aqui apresentados pela primeira vez. A minha actividade desenvolveu-se desde a colheita e diagnóstico preliminar de material biológico para amostragem, à execução do processamento protocolar para microscopia óptica, incluindo contraste de interferência diferencial, microscopia eletrónica de transmissão e microscopia eletrónica de varrimento, à realização dos procedimentos laboratoriais necessários para a biologia molecular. O conteúdo desta tese é da minha autoria, embora inclua as recomendações e sugestões positivamente feitas pelo orientador, colaboradores e técnicos. O trabalho realizado e informação obtida resultaram na elaboração de três artigos científicos distintos, aqui apresentados nos capítulos 2, 3 e 4. Rocha S., Casal G., Matos P., Matos E., Dkhil M. and Azevedo C. 2011: Description of Triangulamyxa psittaca sp. nov. (Myxozoa: Myxosporea), a new parasite in the urinary bladder of Colomesus psittacus (Teleostei) from the Amazon River, with emphasis on the ultrastructure of plasmodial stages. Acta Protozool. 50: (In press) Rocha S., Casal G., Al-Quraishy S. and Azevedo C. 2011: Morphological and molecular characterization of Chloromyxum clavatum n. sp. (Myxozoa: Myxosporea), infecting the gall bladder of Raja clavata (Chondrichthyes) from the Portuguese Atlantic coast. J. Parasitol. (Submitted) Rocha S., Casal G. and Azevedo C. 2011: Morphological and ultrastructural re- description of Chloromyxum leydigi (Myxozoa: Myxosporea) form the gall bladder of marine cartilaginous fish Torpedo marmorata (Chondrichthyes: Torpedinidae) from the Portuguese Atlantic coast. (Unsubmitted) iii Preliminary Note I hereby declare that, as the autor of this thesis, I executed the laboratory procedures leading to the acquisition of the results here presented for the first time. My activity included the sampling and preliminary diagnosis of biological material, the execution of specific procedures for light microscopy, including differential interference contrast, transmission electron microscopy and scanning electron microscopy, and the realization of the laboratory procedures necessary for molecular biology. The entire content of this thesis is of my authorship, despite including the positive recommendations and suggestions made by the supervisor, collaborators and technicians. The work developed and the information accquired resulted in the elaboration of three distinct cientific papers here presented in chapters 2, 3, and 4. Rocha S., Casal G., Matos P., Matos E., Dkhil M. and Azevedo C. 2011: Description of Triangulamyxa psittaca sp. nov. (Myxozoa: Myxosporea), a new parasite in the urinary bladder of Colomesus psittacus (Teleostei) from the Amazon River, with emphasis on the ultrastructure of plasmodial stages. Acta Protozool. 50: (In press) Rocha S., Casal G., Al-Quraishy S. and Azevedo C. 2011: Morphological and molecular characterization of Chloromyxum clavatum n. sp. (Myxozoa: Myxosporea), infecting the gall bladder of Raja clavata (Chondrichthyes) from the Portuguese Atlantic coast. J. Parasitol. (Submitted) Rocha S., Casal G. and Azevedo C. 2011: Morphological and ultrastructural re- description of Chloromyxum leydigi (Myxozoa: Myxosporea) form the gall bladder of marine cartilaginous fish Torpedo marmorata (Chondrichthyes: Torpedinidae) from the Portuguese Atlantic coast. (Unsubmitted) iv Agradecimentos A entrega desta tese representa para mim mais uma etapa da minha vida académica e pessoal que termina e que, de forma igual a tudo o que muito se deseja e dificilmente se alcança, tem um sabor simultaneamente doce e amargo. Doce porque sinto-me plenamente satisfeita do meu desempenho nas funções a que me propus e dos objectivos que alcancei, bem como da evolução profissional e pessoal inerente a todo este processo e que hoje me faz seguramente sentir mais completa. Amargo porque apesar de todas as horas que despendi num microscópio ou na bancada do laboratório, do quão perdida me senti no meio de mais de uma centena de artigos por vezes conflituosos, do cansaço das horas de olhos colados num computador para escrever dois ou três parágrafos, tenho a certeza que esta época me deixará imensas saudades das experiências que tive e das pessoas que conheci. Como tal, não posso deixar de agradecer a todos os que incondicionavelmente me apoiaram e motivaram. Ao professor Doutor Carlos Azevedo pela orientação, disponibilidade, confiança e optimismo com que me brindou e que estimularam em mim o “bichinho” para a parasitologia. À Doutora Graça Casal, que sendo uma pessoa deveras ocupada, não deixou de me prestar a sua valiosa assistência sempre que necessária. Ao Professor Doutor Alexandre Lobo da Cunha, Director do Departamento de Microscopia, e meu professor de licenciatura, pela sua boa vontade em me acolher no laboratório e simpatia que sempre me dedicou. À Elsa Oliveira e Ângela Alves que não só me prestaram o seu precioso auxílio sempre com boa disposição, como se tornaram a minha maior e melhor companhia durante todo este período. Sem os vossos conselhos e experiência não teria sido possível terminar esta tese como o faço. À Mestre Carla Oliveira pela disponibilidade em me orientar na execução dos procedimentos laboratoriais para a biologia molecular. Às minhas colegas de mestrado Lúcia Barriga Negra, Ângela Ferreira, Cláudia Mendes e Lígia Sousa por todas as dicas que me dispenderam, e por compreenderem todas as vezes em que ao invés de dizer sim tive que dizer não, em que cheguei atrasada, em que tive que sair mais cedo, e nas quais porventura não tive a minha melhor prestação como amiga. À minha irmã Gisela, com quem discuto todos os dias mas a quem amo na certeza de que sempre nos apoiaremos ao longo da nossa vida, tanto nos sucessos como nas dificuldades. Afinal de contas, é esse o papel de uma irmã, uma melhor amiga velada pelos laços fraternais. v Aos meus avós maternos, que estiveram sempre presentes em todas as etapas da minha vida com um sorriso ou uma palavra carinhosa, a quem eu amo profundamente e é para mim uma enorme felicidade dar-lhes mais esta alegria e saber que tem orgulho em mim. Aos meus pais, por lutarem tanto para me providenciarem uma carreira como estudante e profissional, pela liberdade de opções e apoio que sempre me proporcionaram, e pelo amor incondicional que me têm. Bem sei que este é um sonho que realizo conjuntamente com vocês e que constitui igualmente motivo de orgulho próprio. Ao Miguel, por ser o meu melhor amigo, por me dar todo o seu amor e carinho e por me fazer sentir a melhor pessoa do mundo. Não podia ter conhecido melhor pessoa na minha vida, com tanta integridade e que me apoiasse tanto e me aturasse tanto como tu fazes todos os dias. Desculpa as horas infindáveis que passaste a ouvir falar de mixosporídeos, de cápsulas polares, de planches, de escalas, de DNA e do “ultramicrófono”, como dizes. Sei que para ti os meus sucessos são também os teus sucessos. Portanto, aqui vai a nós! vi Preface As part of my master’s degree in Marine Sciences – Marine Resources in the Institute of Biomedical Sciences Abel Salazar of the University of Porto, I developed a work focused in the area of cellular biology, more specifically in parasitology. This type of research has always interested me, since it allows the conjugation of both new and old methodologies and quantitative and qualitative results, thus making the exercise of interpretation much more interesting. With this purpose, I joined the Laboratory of Cellular Biology of the same institute, which develops several projects concerning microparasites infecting the aquatic fauna of different geographical areas, and thus is equipped with all the necessary equipament for both microscopic and molecular studies. Also, the experience demonstrated by the investigators and technicians integrated in those projects reflected a positive receptivity for the development of my work, which culminates with the presentation of this document. The present thesis attempts to provide fundamental information on the class Myxosporea of the phylum Myxozoa. To date, more than 2000 species of myxosporeans have been found infecting several fish species, and more rarely anphibians, birds, mammals, and even humans. Despite most myxosporeans being harmless, some are serious pathogens that provoke devasting damages in both wild and reared populations of fish. Considering environmental sustentability, the socio-economical importance of the aquaculture industry, as well as other activities associated with the aquatic environment, the acquisition of precise knowledge concerning these species is essential. Furthermore, science for the sake of knowledge should never be disregarded, even if a direct advantage is not perceptible. For many years, the microscopic
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