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IDENTIFICACIÓN MORFOLÓGICA Y MOLECULAR DE Cochliomyia Spp

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IDENTIFICACIÓN MORFOLÓGICA Y MOLECULAR DE Cochliomyia Spp UNIVERSIDAD CENTRAL DEL ECUADOR FACULTAD DE MEDICINA VETERINARIA Y ZOOTECNIA CARRERA DE MEDICINA VETERINARIA Y ZOOTECNIA “IDENTIFICACIÓN MORFOLÓGICA Y MOLECULAR DE Cochliomyia spp. DE LA COLECCIÓN CIENTÍFICA DEL CENTRO INTERNACIONAL DE ZOONOSIS” Trabajo de Grado presentado como requisito para optar por el Título de Médico Veterinario Zootecnista AUTOR: Jenny Soraya Carrillo Toro TUTOR: Dr. Richar Iván Rodríguez Hidalgo Ph.D. Quito, Mayo, 2015 ii DEDICATORIA A mis padres y abuelos por su esfuerzo, sacrificio e incondicional apoyo, han sido un ejemplo de superación y lucha en mi vida; además con su templanza han sabido mostrarme el camino para alcanzar una meta; también a mi hermano Fernando y a mi compañero Roberto quienes a pesar de las circunstancias adversas han estado ahí durante mi carrera profesional. Jenny iii AGRADECIMIENTOS Quiero expresar mi más sincero agradecimiento Al Dr. Richar Rodríguez por brindarme la oportunidad de formar parte de su Proyecto y, a la vez, ser tutor de esta investigación. A todos mis profesores que, de una u otra forma, me brindaron sus conocimientos y experiencias para mi formación a lo largo de esta carrera. Agradezco al personal científico del Centro Internacional de Zoonosis, especialmente al Ing. Gustavo Echeverría, Dr. Juan Carlos Navarro y a la MSc. Sandra Enríquez, cuyos conocimientos me han enriquecido y han permitido concluir este proyecto. A mis padres, hermano y a mi compañero Diego Cushicóndor, quienes contribuyeron, directa e indirectamente, para la materialización y finalización de este proyecto de tesis. iv AUTORIZACIÓN DE LA AUTORÍA INTELECTUAL v INFORME DEL TUTOR vi APROBACIÓN DEL TRABAJO/TRIBUNAL “IDENTIFICACIÓN MORFOLÓGICA Y MOLECULAR DE Cochliomyia spp. DE LA COLECCIÓN CIENTÍFICA DEL CENTRO INTERNACIONAL DE ZOONOSIS” vii ÍNDICE GENERAL pp. DEDICATORIA .......................................................................................................... ii AGRADECIMIENTOS ............................................................................................... iii AUTORIZACIÓN DE LA AUTORÍA INTELECTUAL .................................................. iv INFORME DEL TUTOR ............................................................................................ v APROBACIÓN DEL TRABAJO/TRIBUNAL .............................................................. vi “IDENTIFICACIÓN MORFOLÓGICA Y MOLECULAR DE Cochliomyia spp. DE LA COLECCIÓN CIENTÍFICA DEL CENTRO INTERNACIONAL DE ZOONOSIS” ........ vi ÍNDICE GENERAL .................................................................................................. vii LISTA DE FIGURAS ................................................................................................. ix LISTA DE TABLAS ................................................................................................... x RESUMEN ................................................................................................................ xi ABSTRACT ............................................................................................................. xii CAPÍTULO I .............................................................................................................. 1 INTRODUCCIÓN ...................................................................................................... 1 CAPÍTULO II ............................................................................................................. 4 REVISIÓN DE LITERATURA.................................................................................... 4 Antecedentes ........................................................................................................ 4 Generalidades ....................................................................................................... 5 Cochliomyia hominivorax ...................................................................................... 6 Ciclo de vida de C. hominivorax- GBG .................................................................. 7 Distribución Geográfica ......................................................................................... 8 Cochliomyia macellaria ......................................................................................... 9 Ciclo Biológico de Cochliomyia macellaria ............................................................ 9 Distribución Geográfica ......................................................................................... 9 Morfología diferencial entre C. hominivorax y C. macellaria ................................ 10 Epidemiología ..................................................................................................... 14 Impacto Económico ............................................................................................. 17 Erradicación del GBG – Cochliomyia hominivorax .............................................. 18 Biología Molecular en la Identificación de Insectos ............................................. 19 ADN Mitocondrial ................................................................................................ 20 Estudios Moleculares realizados en Cochliomyia ................................................ 21 Filogenia ............................................................................................................. 21 viii CAPÍTULO III .......................................................................................................... 23 MATERIALES Y MÉTODOS ................................................................................... 23 Fase de Laboratorio: Unidad de Entomología Aplicada .......................................... 24 Identificación y cultivo de larvas .......................................................................... 24 Identificación Morfológica de Adultos de Cochliomyia spp .................................. 25 Fase de laboratorio: Biología Molecular .................................................................. 25 Extracción y cuantificación de ADN ..................................................................... 25 Reacción en Cadena de la Polimerasa (PCR) .................................................... 27 Análisis de Datos ................................................................................................ 30 CAPÍTULO IV ......................................................................................................... 31 RESULTADOS Y DISCUSIÓN ............................................................................... 31 Identificación Morfológica .................................................................................... 31 Identificación Molecular ....................................................................................... 40 Análisis Filogenéticos .......................................................................................... 41 Citocromo Oxidasa subunidad I (COI) ............................................................. 41 ARN ribosomal (12S) ....................................................................................... 44 Genes combinados COI y 12S......................................................................... 46 CAPÍTULO V .......................................................................................................... 50 CONCLUSIONES Y RECOMENDACIONES .......................................................... 50 Conclusiones ...................................................................................................... 50 Recomendaciones .............................................................................................. 51 BIBLIOGRAFÍA ....................................................................................................... 52 Anexos ................................................................................................................... 65 ix LISTA DE FIGURAS pp. Figura 1. Ciclo de vida de C. hominivorax ............................................................... 8 Figura 2. Características del estadío larvario 3 de C. hominivorax. ....................... 11 Figura 3. Características del estadío larvario 3 de C. macellaria ........................... 11 Figura 4. Características morfológicas diferenciales entre adultos de C. hominivorax y C. macellaria. ....................................................................................................... 12 Figura 5. Mapa de distribución de los países libres del GBG ................................. 14 Figura 6. Mapa de distribución de miasis por C. hominivorax ................................ 15 Figura 7. Genoma mitocondrial de Insectos ........................................................... 20 Figura 8. Adultos de C. hominivorax (izq) y C. macellaria (dcha). .......................... 34 Figura 9. Larvas y Adultos de C. hominivorax y C. macellaria ................................ 39 Figura 10. Árbol de consenso de mayoría producido por análisis de parsimonia del gen COI. ................................................................................................................. 43 Figura 11. Árbol de consenso de mayoría producido por análisis de parsimonia del gen 12S. ................................................................................................................. 45 Figura 12. Árbol de consenso de mayoría producido por análisis de parsimonia del gen COI y 12S. ....................................................................................................... 47 x LISTA DE TABLAS Tabla 1. Características
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