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Giardia Duodenalis and Blastocystis Sp UNIVERSIDAD COMPLUTENSE DE MADRID FACULTAD DE FARMACIA TESIS DOCTORAL Epidemiología molecular y factores de riesgo de protistas enteroparásitos asociados a diarrea en poblaciones pediátricas sintomáticas y asintomáticas en España y Mozambique MEMORIA PARA OPTAR AL GRADO DE DOCTOR PRESENTADA POR Aly Salimo Omar Muadica Directores David Antonio Carmena Jiménez Isabel de Fuentes Corripio Madrid © Aly Salimo Omar Muadica, 2020 UNIVERSIDAD COMPLUTENSE DE MADRID FACULTAD DE FARMACIA DEPARTAMENTO DE MICROBIOLOGÍA Y PARASITOLOGÍA TESIS DOCTORAL Epidemiología molecular y factores de riesgo de protistas enteroparásitos asociados a diarrea en poblaciones pediátricas sintomáticas y asintomáticas en España y Mozambique MEMORIA PARA OPTAR AL GRADO DE DOCTOR PRESENTADA POR: Aly Salimo Omar Muadica Madrid, 2020 D. DAVID ANTONIO CARMENA JIMÉNEZ, Investigador Distinguido del Laboratorio de Referencia e Investigación en Parasitología, Centro Nacional de Microbiología, Instituto de Salud Carlos III. DÑA. ISABEL FUENTES CORRIPIO, Responsable de la Unidad de Toxoplasmosis y Protozoos Intestinales del Laboratorio de Referencia e Investigación en Parasitología, Centro Nacional de Microbiología, Instituto de Salud Carlos III. CERTIFICAN: Que la Tesis Doctoral titulada “EPIDEMIOLOGÍA MOLECULAR Y FACTORES DE RIESGO DE PROTISTAS ENTEROPARÁSITOS ASOCIADOS A DIARREA EN POBLACIONES PEDIÁTRICAS SINTOMÁTICAS Y ASINTOMÁTICAS EN ESPAÑA Y MOZAMBIQUE” presentada por el graduado en Biología D. ALY SALIMO MUADICA ha sido realizada en el Laboratorio de Referencia e Investigación en Parasitología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Majadahonda, bajo su dirección y cumple las condiciones exigidas para optar al grado de Doctor en Microbiología y Parasitología por la Universidad Complutense de Madrid. Majadahonda, 30 de junio de 2020 V.º B.º Director V.º B.º Directora D. David Antonio Carmena Jiménez Dña. Isabel de Fuentes Corripio Notas 1. Esta tesis doctoral ha dado lugar a la publicación, o envío para publicación, de los siguientes artículos revisados por pares: a) Reh L*, Muadica AS*, Köster PC, Balasegaram S, Verlander NQ, Chércoles ER, Carmena D. Substantial prevalence of enteroparasites Cryptosporidium spp., Giardia duodenalis and Blastocystis sp. in asymptomatic schoolchildren in Madrid, Spain, November 2017 to June 2018. Euro Surveill. 2019;24(43). b) Muadica AS, Köster PC, Dashti A, Bailo B, Hernández-de-Mingo M, Reh L, Balasegaram S, Verlander NQ, Ruiz Chércoles E, Carmena D. Molecular diversity of Giardia duodenalis, Cryptosporidium spp. and Blastocystis sp. in asymptomatic school children in Leganés, Madrid (Spain). Microorganisms. 2020;8(4). c) Muadica AS*, Balasegaram S*, Beebeejaun K, Köster PC, Bailo B, Hernández-de-Mingo M, Dashti A, Dacal E, Saugar JM, Fuentes I, Carmena D. Risk associations for intestinal parasites in symptomatic and asymptomatic schoolchildren in central Mozambique. Clin Microbiol Infect. 2020; doi: 10.1016/j.cmi.2020.05.031. d) Muadica AS*, Messa Jr AE*, Dashti A, Santin M, Manjate F, Chirinda P, Garrine M, Vubil D, Acácio S, KösterPC, Bailo B, Nhampossa T, Calero-Bernal R, Mwenda J, Mandomando I, Carmena D. First identification of genotypes of Enterocytozoon bieneusi (Microsporidia) among symptomatic and asymptomatic children in Mozambique. PLoS Negl Trop Dis. 2020;14:e0008419. e) Muadica AS, Köster PC, Dashti A, Bailo B, Hernández de Mingo M, Balasegaram S, Carmena D. Molecular diversity of Giardia duodenalis, Cryptosporidium spp. and Blastocystis sp. in symptomatic and asymptomatic school children in Zambézia province (Mozambique). En revisión. Los autores identificados con un asterisco contribuyeron en igual medida al trabajo citado. 2. Siguiendo las recomendaciones de la Real Academia Española, esta tesis doctoral ha sido escrita utilizando el masculino genérico como mecanismo inclusivo. 3. Siguiendo las recomendaciones del Centro Europeo para la Prevención y el Control de las Enfermedades para la presentación de datos epidemiológicos, en esta tesis doctoral se ha priorizado el redondeo de valores numéricos con decimales a cifras significativas, con excepción de algunos cálculos estadísticos. "Lo importante en la ciencia no es tanto obtener nuevos datos, sino descubrir nuevas formas de pensar sobre ellos". William L. Bragg Agradecimientos La elaboración de este trabajo no hubiera sido posible sin la colaboración, el aliento y el compromiso de varias personas. Por lo tanto, expreso toda mi gratitud y aprecio a todos aquellos que, directa o indirectamente, contribuyeron a hacer realidad esta tarea. Debo agradecer de manera especial, sincera y profunda, a mi director Dr. David Antonio Carmena Jiménez. ¡Muchas gracias! Me diste una oportunidad cuando más la necesitaba, creíste en mí y la confianza depositada me ha permitido completar con éxito esta tesis doctoral. Me siento muy agradecido por todo lo que me enseñaste, por la paciencia que mostraste y por nunca renunciar a mí. El trabajo realizado con rigor y tesón bajo tu orientación ha sido la clave de los buenos resultados que hemos obtenido juntos. Además de enseñarme mucho, te convertiste en un amigo muy importante en el que me inspiro. Quiero agradecer la oportunidad que me brindaste de formar parte de tu equipo de trabajo, que seguro nos permitirá afrontar nuevos y excitantes retos profesionales en el futuro. Es un privilegio trabajar bajo tu liderazgo. Agradezco también a mi directora Dra. Isabel Fuentes Corripio, para quien no hay gracias suficientes. Las notas dominantes de su orientación fueron la libertad de acción que me permitió, la utilidad de sus recomendaciones y la cordialidad con la que siempre me recibió, todas ellas decisivas para que este trabajo contribuyese a mi desarrollo científico y personal. Como maestra, ella fue la máxima exponente, me abrió horizontes y me enseñó principalmente a pensar. Fue y es fundamental en la transmisión de experiencias, en la creación y consolidación del conocimiento y en mis pequeños éxitos. Como amiga es lo que todos queremos, ella siempre está a nuestro lado sin tener que pedir nada. Un agradecimiento muy especial y profundo a mi tutora académica, la Dra. Carmen Cuéllar del Hoyo, por el interés, simpatía y presteza con la que atendió todos mis requerimientos, consultas y dudas relacionadas con la tramitación de esta tesis doctoral. Quiero agradecerle su empatía, el tiempo que me dedicó cuando necesité que me escuchara y los sabios consejos prestados. El trabajo es arduo pero su constancia y dedicación para conmigo han sido claves para lograr el éxito que he obtenido. ¡Gracias por ser una gran persona! Agradezco especialmente a Pamela C. Köster, Begoña Bailo, Alejandro Dashti, Marta Hernández de Mingo y María Aguilera, del Laboratorio de Referencia e Investigación en Parasitología (CNM-ISCIII). Quiero que sepan que es un honor trabajar con un grupo tan extraordinario de compañeros tan atentos y talentosos. Gracias por todos los logros que hemos alcanzado y por todas las dificultades que hemos superado juntos. Todos ustedes son los mejores. Mi más sincero agradecimiento a la Dra. Esther Ruiz Chércoles, del Centro de Salud Maria Jesús Hereza (Leganés); a la Dra. Sooria Balasegaram, el Dr. Kazim Beebeejaun y el Dr. Neville Q. Verlander, de la Agencia de Salud Pública de Inglaterra (Reino Unido); a la Dra. Lucia Reh, del Centro Europeo para la Prevención y el Control de las Enfermedades (Suecia); al Dr. Rafael Calero Bernal, de la Facultad de Veterinaria (UCM) y a la Dra. Mónica Santín, del Departamento de Agricultura de Estados Unidos, quienes me brindaron su valiosa ayuda y colaboración en los muestreos de campo en Leganés, el diseño de los cuestionarios epidemiológico, el análisis de datos estadísticos y la revisión del trabajo final. A Growen Will, Ana “Vicky” Ibarra, Ramón Tanoira “Moncho”, Carla Franch, Elena Dacal, Aida de Lucio, Andrés Darío Tolosa y Néstor Alberto; gracias por vuestra espontaneidad y alegría a la hora de intercambiar información y material, en una clara demostración de amistad y solidaridad. También dejo una palabra de agradecimiento a los investigadores y técnicos del Laboratorio de Referencia e Investigación en Parasitología (CNM-ISCIII) y de los laboratorios clínicos de los centros médicos colaboradores de Zambézia (Mozambique) por la amabilidad y colaboración prestada siempre que la solicité. A la Dra. Maria Molina y la Dra. Maria Valderrama, coordinadoras del máster de Microbiología y Parasitología (UCM), un cariñoso agradecimiento. Agradezco a todos los niños y sus representantes legales su participación voluntaria en este trabajo de tesis doctoral, y a las autoridades del gobierno local (Direcciones de Administración de Salud y de Educación del distrito en Zambézia) y a los líderes comunitarios por posibilitar las distintas etapas del trabajo de campo realizado en Mozambique. Agradezco a la Universidad Pedagógica de Quelimane (actual, Universidad Licungo), por la concesión de una beca durante mis estudios doctorales, y al Centro Nacional de Microbiología en Majadahonda por abrirme las puertas de esta prestigiosa institución. Finalmente, agradezco al Instituto de Salud Carlos III por la financiación del proyecto de investigación Exp. PI16CIII/00024 (Investigador Principal: David Antonio Carmena Jiménez) que ha permitido el trabajo experimental y de campo necesario para realizar esta tesis doctoral. A mi papá (en su memoria) A mi familia ¡Estímulos que me llevaron a buscar una nueva vida cada día, mis agradecimientos por aceptar que me alejase de su compañía por
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