Evaluation of the intra and interspecific variability in the genus Perkinsus. Proteomic analysis of the parasite and its interaction with the host

Clam culture industry is rising every year worldwide. The most produced species is the Manila clam Ruditapes philippinarum. China is the main producer country followed by Italy, South Korea, USA and Spain. The introduction of the Manila clam in Europe was between 1970 and 1980. R. philippinarum is well adapted and grows faster than native species R. decussatus, for this, is the most produced species in Europe. The production of venerids in Galicia is a very important socio-economic resource. The production of Manila clam in Galicia rose in the last years and now is the most produced species too. The infection by Perkinsus olseni is one of the most serious diseases affecting clams. The parasite is highly distributed worldwide and affect to a large list of molluscs. Mortality of R. decussatus and R. philippinarum was associated with P. olseni infection in south Europe as well as with R. philippinarum mortality in several Asian countries. Two species of the genus Perkinsus, P. olseni and P. marinus, are included in the list of notifiable diseases of the World Organization for Animal Health, which indicates the interest to stop the expansion of the disease. The knowledge of P. olseni is very scarce in comparison with P. marinus which provokes massive mortalities in oysters Crassostrea virginica in USA. The economic transcendence of the perkinsosis justifies the research for a better knowledge of the disease and minimizes their effects. With this purpose, this study was developed in order to know (1) the variability of P. olseni along the Spanish coast, with the emphasis in virulence variability among populations, and (2) the modulation of the Manila clam protein expression due to P. olseni infection, with emphasis in the search of protein markers of resistance to the disease. The study of the variability was done analyzing the genetic population structure as well as the proteome of P. olseni clons derived from several regions of the Spanish coast. Furthermore, the proteome of P. olseni was compared with the proteome of two species of the genus, P. marinus and P. chesapeaki, in order to amplify the perspective of the variability of P. olseni. The analysis of the modulation of the Manila clam protein expression due to P. olseni infection was focused in haemocytes and haemolymph. The effect of the exposition to P. olseni in clams was measured at short time and a long time with the aim in the modulation of the immune response of the clam to the parasite

Applying genetic technologies to combat infectious diseases in aquaculture

Disease and parasitism cause major welfare, environmental and economic concerns for global aquaculture. In this review, we examine the status and potential of technologies that exploit genetic variation in host resistance to tackle this problem. We argue that there is an urgent need to improve understanding of the genetic mechanisms involved, leading to the development of tools that can be applied to boost host resistance and reduce the disease burden. We draw on two pressing global disease problems as case studies—sea lice infestations in salmonids and white spot syndrome in shrimp. We review how the latest genetic technologies can be capitalised upon to determine the mechanisms underlying inter- and intra-species variation in pathogen/ parasite resistance, and how the derived knowledge could be applied to boost disease resistance using selective breeding, gene editing and/or with targeted feed treatments and vaccines. Gene editing brings novel opportunities, but also implementation and dissemination challenges, and necessitates new protocols to integrate the technology into aquaculture breeding programmes. There is also an ongoing need to minimise risks of disease agents evolving to overcome genetic improvements to host resistance, and insights from epidemiological and evolutionary models of pathogen infestation in wild and cultured host populations are explored. Ethical issues around the different approaches for achieving genetic resistance are discussed. Application of genetic technologies and approaches has potential to improve fundamental knowledge of mechanisms affecting genetic resistance and provide effective pathways for implementation that could lead to more resistant aquaculture stocks, transforming global aquaculture.publishedVersio

Applying genetic technologies to combat infectious diseases in aquaculture

Disease and parasitism cause major welfare, environmental and economic concerns for global aquaculture. In this review, we examine the status and potential of technologies that exploit genetic variation in host resistance to tackle this problem. We argue that there is an urgent need to improve understanding of the genetic mechanisms involved, leading to the development of tools that can be applied to boost host resistance and reduce the disease burden. We draw on two pressing global disease problems as case studies—sea lice infestations in salmonids and white spot syndrome in shrimp. We review how the latest genetic technologies can be capitalised upon to determine the mechanisms underlying inter- and intra-species variation in pathogen/parasite resistance, and how the derived knowledge could be applied to boost disease resistance using selective breeding, gene editing and/or with targeted feed treatments and vaccines. Gene editing brings novel opportunities, but also implementation and dissemination challenges, and necessitates new protocols to integrate the technology into aquaculture breeding programmes. There is also an ongoing need to minimise risks of disease agents evolving to overcome genetic improvements to host resistance, and insights from epidemiological and evolutionary models of pathogen infestation in wild and cultured host populations are explored. Ethical issues around the different approaches for achieving genetic resistance are discussed. Application of genetic technologies and approaches has potential to improve fundamental knowledge of mechanisms affecting genetic resistance and provide effective pathways for implementation that could lead to more resistant aquaculture stocks, transforming global aquaculture

Cryptic reservoirs of micro-eukaryotic parasites in ecologically relevant intertidal invertebrates from temperate coastal ecosystems unveiled by a combined histopathological, ultrastructural, and molecular approach

271 p.La mayoría de los eucariotas son organismos unicelulares (protistas), muchos de ellos pertenecientes a linajes que divergieron temprano en la historia evolutiva de este Dominio de organismos nucleados. Microscópicos, enormemente diversos y fenotípicamente convergentes, su clasificación cladística ha sido históricamente compleja, dejando atrás un extenso registro de taxones y de términos parafiléticos y polifiléticos. Teniendo que investigar atributos estructurales, celulares, biológicos y ecológicos en un mundo de rápidas interacciones y difícilmente accesible a simple vista, la protistología es particularmente dependiente de la sistemática. Ésta permite inferir rasgos de especies crípticas a partir de especies evolutivamente relacionadas.Las moléculas de ADN (y ARN), representan un "registro" preciso de estos eventos de diversificación, que preceden incluso a los más antiguos registros fósiles. En los últimos años, la maduración de los métodos de filogenia molecular, catalizados por una mayor accesibilidad a la secuenciación de próxima generación (NGS), está permitiendo resolver preguntas e hipótesis sobre la evolución y la especiación de estos organismos micro-eucariotas que no se habían podido responder mediante otros métodos. Por una parte, arboles filogenéticos construidos mediante concatenaciones de cientos, incluso miles de genes, están permitiendo rastrear la historia evolutiva de los linajes protistas hasta el último ancestro común de todos los eucariotas (LECA). Concomitantemente, análisis moleculares basados en genes e incluso fragmentos cortos (especialmente 18S rRNA), recuperados principalmente de matrices ambientales u orgánicas (eDNA o ADN ambiental), están revelando una ¿caja de Pandora¿ de diversidad micro-eucariota. Ésta diversidad ¿oculta¿ está transformando nuestra percepción de los protistas en la cadena trófica y la estructura ecológica. En el medio marino, sus papeles como autótrofos, heterótrofos (predadores, saprófitos, parásitos) o mixótrofos crece en importancia día a día. El aumento simultáneo de diversidad e importancia ha sido particularmente pronunciado entre los linajes de parásitos protistas, que adaptados a la vida dentro de un huésped son más inaccesibles y morfológicamente indistinguibles que sus homólogos de vida libre. Muy competitivo como estilo de vida, el parasitismo ha evolucionado de forma independiente varias veces en prácticamente todos los grupos eucariotas, en algunos incluso cientos de veces. De hecho, es posible que el efecto parapátrico que implica una existencia endosimbiótica, haya exacerbado la especiación entre los parásitos, que representan la que posiblemente sea la más común estrategia de consumo entre los organismos vivos. Es más, el número de especies crípticas que están, a día de hoy, siendo descubiertas en la mayoría de los linajes de parásitos protistas sigue aumentando abruptamente o apenas comienza a mostrar una desaceleración. Cabe destacar, que el descubrimiento de esta diversidad oculta, incluidas las especies crípticas, va más allá de la escalada en el número de especies; afecta los estudios sobre biología celular, ciclos biológicos, y ecología. Inexorablemente, esta fuerza mostrada por los métodos de análisis y secuenciación del ADN está abriendo una brecha entre la diversidad genética existente y nuestra comprensión de la morfología, patología, transmisión y posibles hospedadores de los parásitos protistas que la constituyen. Este desequilibrio es particularmente evidente entre los parásitos que infectan linajes de invertebrados, los cuales, salvo algunos taxones con interés comercial, permanecen en gran parte sin analizar, a pesar de constituir un grupo mucho más diverso que los vertebrados. Por una parte, es lógico que los parásitos protistas causantes de infecciones en especies marinas de interés comercial (peces, bivalvos, crustáceos¿) hayan sido priorizadas, pero hay que tener en cuenta que muchos de estos micro-eucariotas tienen ciclos de vida complejos, en los que pequeños invertebrados actúan muchas veces como vectores o reservorios. Descubrir y contextualizar estas asociaciones puede ser determinante a la hora de comprender cuándo y dónde puede variar la presión y capacidad infectiva de algunas de estas infecciones en la comunidad o huéspedes específicos. Al mismo tiempo que su diversidad e importancia aumenta, la inclusión progresiva de parásitos en modelos ecológicos está experimentando variaciones de gran alcance en la dinámica poblacional de las especies animales, vegetales o fúngicas en los ecosistemas. En consecuencia, las asociaciones entre parásitos y hospedadores se investigan cada vez más como una parte importante de la estructura de la comunidad, y no exclusivamente como una "molestia" para el ser humano y sus intereses. Por desgracia la inclusión de parásitos en modelos ecológicos está siendo lastrada por un profundo desconocimiento de estas interacciones. A diferencia de los organismos multicelulares, que han podido ser observados por científicos y aficionados durante siglos, la distribución espaciotemporal de la mayoría de los organismos unicelulares sigue siendo un profundo misterio. No obstante, dadas sus importantes funciones como vectores, huéspedes intermediarios y reservorios, una comprensión mucho más profunda del patobioma (patógenos asociados a un hospedador) y su variabilidad espacio-temporal es de suma importancia para un mayor poder de predicción de los factores de presión causantes de epidemias o zoonosis en el huésped, la población y el medioambiente.En este contexto, la hipótesis de este estudio plantea que especies de invertebrados comunes en la zona inter-mareal de ecosistemas costeros en climas templados son reservorios crípticos de un número significativo de parásitos micro-eucariotas (protistas) de interés para el medio y los recursos marinos. Eldescubrimiento progresivo de estas asociaciones ocultas de parásitos-huéspedes (mediante exámenes combinando técnicas histopatológicas, ultraestructurales y moleculares) permite una mejor comprensión de la morfología, patología, biología celular y ciclo de vida de dichos patógenos, lo que a su vez consiente un seguimiento más estrecho de los factores y presiones que promueven epidemias y zoonosis en una escala espacio-temporal.PIE:Plentziako Itsas Estazio

The impact of ocean acidification, increased seawater temperature and a bacterial challenge on the immune response and physiology of the blue mussel, Mytilus edulis.

Anthropogenic activities are fundamentally altering the chemistry of the world’s oceans. Many of these modifications could have a significant impact on the health of marine organisms. Yet, despite being proposed as one of the most significant threats that marine ecosystems face, to date very little is known about the impact of anthropogenic climate change, and ocean acidification in particular, on host defence. The aims of this thesis are to investigate the impact of environmental stressors on the invertebrate immune response, providing empirical data on how anthropogenically induced stressors will impact the invertebrate immune system and how this will impact organism condition and subsequent physiological trade-offs. Exposure to reduced seawater pH and increased temperature significantly reduced the immune response in the blue mussel, Mytilus edulis. This reduction in immune response could indicate stress-induced immune dysfunction. However, the immune system protects an organism from infectious disease, ensuring survival, and should therefore be evaluated functionally rather than immunologically. By subsequently exposing mussels to a bacterial challenge this study demonstrated that an earlier study which measured a reduction in host defence represented a trade-off of immune system maintenance costs, with mussels maintaining a capacity to up-regulate immune defence when required. However, whilst this immune plasticity ensures mussels are able to survive a pathogen exposure, such a strategy appears to be physiologically costly. This cost is seen as a reduction in reproductive investment, an altered energy metabolism and an altered fatty acid composition in organisms exposed to low pH. Therefore the overarching picture that emerges is, without measuring physiological processes functionally, and in neglecting any physiological trade-offs, it is possible that many studies may misinterpret the complex physiological responses of marine organisms to ocean acidification.Natural Environment Research Counci

CMFRI Annual Report 2011-2012

CMFRI Annual Report 2011-201

Analisi e gestione informatica di sequenze trascritte in organismi non-modello

2011/2012Il tema principale di questo lavoro di tesi è la discussione dei metodi che, mediante l’utilizzo di strumenti creati ad-hoc e di software di terze parti, hanno permesso analizzare sequenze trascritte di 5 organismi non-modello: Mytilus galloprovincialis, Ruditapes philippinarum, Latimeria menadoensis, Astacus leptodactylus e Procambarus clarkii.XXV Ciclo198