ARTIGOS

Artigos científicos ou traballos académicos sobre resultados aplicados á investigación da criopreservación e conservación da biodiversidade mariña, xa sexa teórica ou aplicada, cun obxectivo divulgativo.

Cryopreservation of Platynereis dumerilii larvae

Aug 2025

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The marine annelid Platynereis dumerilii is a functional molecular model organism for developmental, evolutionary and chronobiological studies. Research on Platynereis is rapidly growing, and with it, the number of genetic variants that laboratories isolate or generate and must subsequently maintain and propagate. Therefore, there is an urgent need to alleviate the burden of live culture maintenance by developing cryopreservation techniques for this species. We report the first cryopreservation protocol for P. dumerilii larvae, which combined with a careful post-thawing culturing regime, allowed us to obtain animals that survived to adulthood and successfully reproduced. Our experiments show highest survival rate in 6-8 day-old larvae. Equilibration with cryoprotecting agents takes 1h in 5% (v/v) Me2SO + 0.1%(v/w) sucrose, followed by transfer to 0.25ml straws. The protocol cools larvae at 2,5C/min from 20C to -35C using a programmable freezer, followed by a rapid transfer to liquid N2. Larvae are thawed in a water bath at 18C. The post-thaw larvae feeding regime consisted of 50% Tetraselmis + 50 % diatom strains mixture (Grammatophora marina and Nitzschia laevis). The maximum survival obtained with this protocol so far produced 34% survival after over 5 months.

Cryoprotection and cryodamage in sea urchin eggs

Jun 2025

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Cryobiology is a well-developed and worldwide-recognized technique for achieving long-term storage of biological material at low temperatures. Cryopreservation consists of freezing, banking and thawing living organisms, cells or tissues in the presence of cryoprotecting agents. When cryopreserving cells, one way to avoid lethal Internal Ice Formation (IIF) is to cool cells slowly enough so that the freezable water in the cell flows out of the cell and freezes externally. Another alternative approach is to avoid intracellular ice formation by vitrifying cellular water which involves converting water into a non-crystalline or amorphous glass. Several species of marine invertebrates can be cryopreserved, most of the developed protocols are either for sperm or for larvae. This is due to the impossibility of cryopreserving oocytes of marine invertebrates with a good post-thaw survival. The oocytes of aquatic organisms have proven to be recalcitrant to cryopreservation and there are several factors that have been quoted as responsible for this: large volumes, low surface/area ratio, low permeability to water and cryoprotecting agents, high sensitivity to toxicity of cryoprotecting agents and chilling sensitivity are some examples. The aim of this work is to use this experimental methodology to try to cryopreserve marine invertebrate´s oocytes which have remained as a challenge up until now. This work aims to answer some important questions regarding damage during traditional slow cooling, about the technical challenges of the application of vitrification to marine invertebrates and the implementation of technologies like vitrification coupled to ultra-rapid laser warming as a solution to slow freezing damage.

Understanding osmoregulation, cryoprotectant toxicity, and cold tolerance in sea urchin eggs: Implications for cryopreservation

Mar 2025

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Successful cryopreservation of marine invertebrates immune cells enables long-term studies of common octopus, Octopus vulgaris Cuvier 1797, hemocyte immune functions

Mar 2025

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The common octopus, Octopus vulgaris Cuvier 1797, as all cephalopods, presents highly evolved characteristics compared to other classes of molluscs and the whole invertebrate phyla. However, to date, there is not much information about its immune system, and studying the defense mechanisms is a key step in understanding their response to external aggressions, having the tools to anticipate animal health problems and ensure their welfare. The lack of cell cultures in molluscs is a major problem when carrying out in vitro assays that help to deepen our knowledge of this species’ main immune cells. Cryopreservation becomes an alternative to maintaining viable and functional cells after freezing/thawing processes. Having access to good high-quality cells for long periods allows cover a wider repertoire of studies, time courses, and the avoidance of logistical issues such as loss of viability and/or functionality, time constraints, or sample transport challenges. Additionally, high-quality cell suspensions are essential for successful applications, such as single-cell sequencing, where viability and functionality are the key to optimal identification. The optimal medium, cryoprotective agent, and freezing/thawing protocol for octopus hemocytes have been selected. We show here the first functional results from cryopreserved hemocytes. Cells cryopreserved in MAS medium supplemented with EG maintained viability above 80% after 15 weeks post cryopreservation storage at -80°C, and their functional ability to phagocytize bacteria similar to fresh cells. Moreover, thawed acclimated cells exhibited a gene expression pattern comparable to fresh cells, as opposed to directly thawed cells. The acclimation process after thawing was essential to recover the functional activity of the cells and to return to levels of gene expression involved in oxidative stress similar to fresh cells.The results presented here will facilitate functional studies of octopus immune cells and provide tools for cell preservation in other molluscs species.

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Cryopreservation of Juvenile Mytilus galloprovincialis: Safeguarding Mollusk Biodiversity and Aquaculture

JAN 2025

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Mytilus galloprovincialis Lamarck, 1819, is a globally significant aquaculture species, contributing 25% of fresh seafood landings and leading EU mussel production. In Spain, it accounted for 255,218 tonnes in 2022, around 60% of the global harvest. This study introduces groundbreaking cryopreservation, being the first to successfully cryopreserve the juvenile stage of a marine organism, the largest marine organism ever cryopreserved. By optimizing feeding strategies, equilibration times, and cryoprotectant concentrations, we extended our initial protocol for young larvae to include complex larval stages (72h-26 days postfertilization) and juveniles (40-45 days, up to 1 mm). These protocols mark a significant step in biodiversity conservation, seedstock preservation, genetic line maintenance, and sustainability of overexploited mollusk populations.

Cryopreservation of sea urchin oocytes: Overcoming challenges for future applications

Dec 2024

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Ecotoxicological evaluation of microplastics and associated pollutants in a marine trophic chain

Nov 2024

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The global production of plastics has grown drastically due to their low cost and versatility. In 2022, over 400 million metric tons were produced, with polyethylene (PE) accounting for 26.3%. Microplastics (MPs), which are plastic particles ranging from 1 µm to 5 mm, represent a significant environmental threat, mainly entering the ocean through freshwater sources and inadequate disposal and recycling practices. It is estimated that between 0.13 and 3.8 million metric tons of plastic end up in the world’s seas and oceans each year. MPs accumulate in all parts of the ocean, affecting marine life through physical and chemical interactions and transporting pollutants such as heavy metals and pesticides. The thesis hypothesis suggests that high-density polyethylene (HDPE) MPs can produce toxic effects and adsorb metallic and organic contaminants, modulating their bioavailability and toxicity for aquatic species in the marine trophic chain formed by microalgae, zooplankton, and fish. The general objective is to evaluate the ecotoxicological effects of HDPE MPs alone and in combination with contaminants like mercury (Hg) and the pesticide chlorpyrifos (CPF) in a model marine trophic chain. The toxic impact of oxidized and non-oxidized HDPE MPs on the microalga Rhodomonas lens was examined. The effects of contaminants such as chlorpyrifos (CPF) and mercury (Hg), alone and combined with MPs, on the growth, cell viability, and pigment content of the microalgae were investigated. The results indicated that non-oxidized MPs initially increased cell concentration due to the release of exopolymeric substances by the microalgae, facilitating the aggregation of MPs and microalgae. However, at 96 hours, there were no significant differences in growth rates compared to the controls. Cell viability was significantly reduced due to surface damage caused by physical interaction with MPs. The combination of MPs with contaminants generally showed reduced toxicity compared to contaminants alone, suggesting lower bioavailability of the contaminants when adsorbed on MPs. This highlights the complexity of interactions between MPs and contaminants and their impact on marine microalgae. The thesis also focused on studying the impact of HDPE MPs on the copepods Acartia tonsa, investigating physiological, reproductive, and biochemical responses, as well as the bioaccumulation of Hg and CPF. Oxidized and non-oxidized MPs did not show significant lethal or sublethal effects after 48 hours, but exposure increased the antioxidant system response in the copepods. Bioaccumulation of Hg and CPF was similar between MPs and contaminants dissolved in water, although exposure to MPs loaded with contaminants slightly increased physiological and reproductive toxicity. Furthermore, copepods exposed to MA+Hg/CPF showed lower bioaccumulation and lesser biological effects compared to those exposed to MPs+Hg/CPF, highlighting the importance of particle nature in contaminant toxicity. The effects of trophic transfer of MPs, alone and combined with Hg and CPF, on fish larvae (Scophthalmus maximus) through the ingestion of contaminated copepods were also evaluated. The responses analyzed included bioaccumulation of Hg/CPF, biochemical biomarkers, and physiological responses. Bioaccumulation of Hg was significantly higher through MPs compared to MA, while CPF bioaccumulation showed similar patterns in MPs and MA. The larvae showed alterations in antioxidant enzymes and evident neurotoxicity with non-oxidized MPs; however, no significant effects were observed on growth, condition index, or survival. Exposure to Hg and MPs+Hg produced histological changes in the gills, suggesting that MPs can increase the bioavailability of Hg, causing adverse effects on tissues. The toxic effects were generally lower with MA+Hg/CPF, highlighting the lower risk of natural particles in the transfer of contaminants in the trophic chain.

Influence of physico-chemical parameters on the settlement of competent larvae of cultured sea urchin, Paracentrotus lividus

Aug 2024

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Achieving efficient metamorphosis and settlement of larvae is a major bottleneck in the culture of sea urchins; therefore, in order for culture operations to be economically viable, it is urgent to optimize this crucial step under captive conditions. This study aimed to investigate how various parameters, including temperature, salinity, light cycle, and inducer type, affect the settlement of competent P. lividus larvae. Three different salinities (35, 30, 25‰), three temperatures (25, 20, 15 °C), four light conditions (16/8, 12/12, 24/0 and 0/24) and five types of biofilms (three of “natural” origin and the other two from monospecific collection cultures) were tested. The results suggest a relationship between low salinity and decreased survival and settlement rates. Conversely, the photoperiod appears not to have significant influence over survival and settlement rates. It appears that specific temperatures did not affect rates of survival and settlement, rather it was the deviation from initial culture temperature that is responsible for these rates. Finally, the multispecific inoculum of benthic diatoms showed a higher survival and settlement rate and lower mortality rate compared to monospecific inoculum.

Cryopreservation Challenges and Cryoprotectant Efficacy in Cyprinodon variegatus Embryos- Cold Biology and Medicine annual meeting 2024

May 2024

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Cold Biology and Medicine Annual Meeting- IPC&C

Comparative Gamete cryopreservation

May 2024

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This is an update of Estefania Paredes, Gamete Preservation, Editor(s): Michael K. Skinner, Encyclopedia of Reproduction (Second Edition), Academic Press, 2018, Pages 769–775, ISBN 9780128151457, https://doi.org/10.1016/B978-0-12-809633-8.20629-8

ARMS-MBON: long-term genetic monitoring of marine hard-bottom communities

Oct 2023

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To understand global shifting trends in marine biodiversity it is essential to apply cost-efficient biomonitoring approaches in a coordinated and standardized manner, to generate comparable data across geographical regions and countries. In this context, DNA metabarcoding is particularly useful to document biodiversity over larger spatiotemporal scales and in habitats with limited access, such as marine hard substrata which are difficult to sample for quantitative ecological studies; for this reason, we have established the Autonomous Reef Monitoring Structures Marine Biodiversity Observation Network (ARMS-MBON). ARMS-MBON deploys ARMS in ports, marinas, and nature reserves along the European coastline, including polar regions and the Red Sea. The network maintains 25 observatory sites deploying ARMS on a regular basis. Using metabarcoding data and associated (meta)data (e.g., pictures, voucher specimens), the community composition is analyzed through a dedicated workflow hosted at the Tesseract Virtual Research Environment and built under the Internal Joint and Collaborative Initiative of LifeWatch ERIC. ARMS MBON has now become part of the European Marine Omics Biodiversity Observation Network (EMO BON), a larger European initiative for the observation of genomic biodiversity. The first sampling campaigns (2018 and 2019 deployments) produced 567 physical samples from 56 ARMS across 15 observatories. Preliminary results show the robustness and sensitivity of ARMS in detecting rare, alien, and key species. They also document a significantly increased species richness in marine protected areas, as compared to the other study sites. We suggest that the ARMS-MBON data and protocols should be used in biological monitoring programs and long-term ecological research.

Exploring the feasibility of cryopreserving larvae of the common cockle (Cerastoderma edule) for hatchery production

Aug 2023

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The decline of natural populations of the common cockle (Cerastoderma edule) through the European coast is posing a threat to local small-scale fisheries. These declines are primarily attributed to the prevalence of several pathogens and the disseminated neoplasia in cockle populations. The institution of a biobank of cryopreserved larvae could enhance hatchery production and help the restocking. The present work aimed at the development of a cryopreservation protocol for larvae of the common cockle using the mollusk cryopreservation protocols designed in our laboratory. Toxicity bioassays and short-term cryopreservation experiments were performed for protocol optimization according with cellular tolerance. Once settled, the viability of cryopreserved larvae was studied long term. Toxicity tests evidenced high tolerance of larvae against detrimental effects of Cryoprotecting Agents (CPAs). Cryopreservation of 48 h-old D-larva showed a 100% survival when increasing the equilibrium time from 15 to 60 min and using Propylene-Glycol (PG) + 0.4 M Trehalose (TRE) in Filtered Sea Water (FSW) and 60 min of exposure to CPA solution before slow-cooling. However, when cryopreserving the older larvae, the variation in equilibrium times hardly showed any effect but 10% Ethylene-Glycol (EG) + 0.4 M TRE and 60 min of exposure yielded the best relative survivorship (100%). Cryopreservation caused a significant delay on the growth rate of the latest larval stage. However, cryopreserved larvae survived to day 4-6, while 30 ± 12.17% of control larvae developed into pediveliger stage, of which 50% settled and transformed into juvenile cockles. These results demonstrated the role of the cell-type specificity in cryopreservation and highlight the importance of studying potential long-term effects of this tool to ensure the viability of the protocols.

Marking Methodologies for Sea Urchins: A Review

Apr 2023

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Many authors have tested a large number of tagging methodologies on sea urchins, either with the aim to study the growth of urchins, their movement patterns, the dynamics of their populations, or their ecology and development in the natural environment. Different types of markings, both external and internal, have been used for the individual identification of sea urchins since the year 1935. Since the year 1969, marking techniques based on different chemical substances were also implemented, either by immersion in fluorochrome baths or by injecting these substances directly into the sea urchin. There is a clear need to collect all the existing information to date related to the marking methodologies for sea urchins, to establish the most appropriate marking methodology for each species of sea urchin, depending on the objective and duration of the study to be performed. This report presents for the first time a complete review of tagging methodologies used until the date in different species of sea urchins, from physical methods, such as the use of Passive Integrated Transponder (PIT) Tags, Anchor tags, and so on, to chemical ones, in which different fluorochromes are used that adhere to the calcified structures of the sea urchin and are subsequently detected using a fluorescence microscope. We also analyzed the effectiveness of the methods used in each case, comparing the survival rate, the tag retention rate, or the recapture rate of tagged urchins obtained with each tagging methodology. The evolution of the physical tags for sea urchins should be oriented toward the development of more durable markings over time, which allow marking urchins of smaller diameter and which in turn present a higher retention rate in the long term, both in captivity conditions and in the natural environment. The chemical tags should be aimed at reducing costs through the development of methods, where the mark can be visualized without the need of fluorescence microscopes, the development of methods in which the dyes can be reused, and finally to the development of techniques that allow to visualize the fluorochrome marks externally, without the need to sacrifice the urchin for the mark detection.

Modulation of stress factors for cryopreservation of Paracentrotus lividus (Lamarck 1816) larvae

Feb 2023

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In the present work we modulated two stress factors salinity and temperature, whose ranges have been previously determined by bioassays using six pre-treatments (18 ºC-29.5‰; 18 ºC-35‰; 18 ºC-39‰ and 20 ºC-29.5‰; 20 ºC-35‰; 20 ºC-39‰), in order to obtain a successful cryopreservation protocol for pluteus larvae of the sea urchin P. lividus (Lamarck 1816). Toxicity tests were performed with different cryoprotectants in a range of 0.5-3 M. Best results pointed out to METH and Me2SO as those more suitable for cryopreservation. First an exploratory cryopreservation experiment with Me2SO supplemented with 0.04 M trehalose (TRE) was tested following the protocol for cryopreservation of embryos (8-hour blastula) of Bellas and Paredes, 2011, which did not give satisfactory results. A cryopreservation experiment was performed with both cryoprotectants supplemented with 0.04 M trehalose on 4-arm pluteus larvae (48h-old) developed in these pre-treatment conditions, followed by a simpler and shorter protocol with a cooling rate of 1 °C/min to -35 °C, achieving for the first time the successful cryopreservation of P. lividus larvae. When larvae were incubated in low salinity or low temperature pre-treatments, they showed delayed larval development and abnormalities. In contrast, pretreatments with high temperature and salinity showed good results. Dimethyl sulfoxide with trehalose proved to be the only effective cryoprotectant for successful cryopreservation of P. lividus larvae. The success of dimethyl sulfoxide is consistent with that described for other cases in previous literature, where dimethyl sulfoxide, although not the least toxic compound, gave the best cryopreservation result.

The Importance of Developing and Standardizing Gamete, Embryo and Larvae Handling in Aquatic Animals

Jan 2023

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Artificial reproduction in aquatic animals usually involves the collection and handling of gametes both from males and females in a way that secures their quality and optimizes the fertilization event […]

Evaluation of physico-chemical factor interactions in the cryopreservation of Paracentrotus lividus pluteus larvae

Dec 2022

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Probl Cryobiol Cryomed 2022; 32(4):295

Modulation of stress factors for cryopreservation of P. lividus larvae

Dec 2022

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Handling, Reproducing and Cryopreserving Five European Sea Urchins (Echinodermata, Klein, 1778) for Biodiversity Conservation Purposes

Nov 2022

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Simple Summary This is a basic study about the handling, reproduction and cryopreservation of cells from five different sea urchin species present in the Northeastern Atlantic Ocean: Sphaerechinus granularis, Psammechinus miliaris, Echinus esculentus, Paracentrotus lividus and Echinocardium cordatum. Abstract In this work, five local sea urchin species found in European waters were studied. Four were regular species: Sphaerechinus granularis, Psammechinus miliaris, Echinus esculentus (Linnaeus, 1758) and the edible sea urchin Paracentrotus lividus; and one was an irregular species, Echinocardium cordatum. These five species of sea urchins have been studied regarding their fertility, toxicity of cryoprotecting agents, cryopreservation of different cell types and chilling injury. The baseline fertility is similar in P. lividus, P. miliaris and S. granularis. Nonetheless, the sperm:egg ratio, contact time and development of the fertilization envelope would need to be studied further on a case-by-case basis. Sperm can be maintained inactively in the gonad (4 °C), and oocytes also maintain quality in sea water (4 °C), even after 72 h. Sperm was cryopreserved for four species with some post-thaw intra specific variability, and embryo cryopreservation was only possible for S. granularis. Overall, this study provided a wider vision of the biology and reproduction of these species that will help us develop tools for their biodiversity conservation through cryopreservation.

Osmotic, temperature, and cytotoxic damage in sea urchin oocytes

Jul 2022

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Sea urchin (Paracentrotus lividus) oocytes are an important species for aquaculture and as a model species for multiple scientific fields. Despite their importance, methods of cryopreserved biobanking of oocytes are currently not possible. Optimized cryoprotectant loading may enable vitrification methods of cryopreservation and thus long term storage of oocytes. Determining an optimized protocol requires membrane characteristics and models of damage associated with the vitrification loading protocol, namely osmotic, temperature, and cytotoxic damage. We present and experimentally evaluated state of the art models alongside our novel models. We experimentally verify the damage models throughout time at difference treatment intensities. Osmotic damage experiments consisted of hypertonic solutions composed of seawater supplemented with NaCl or sucrose and hypotonic solutions composed of seawater diluted with deionized water. Treatment times ranged from 2 to 30 minutes. To test temperature damage (in particular chill injury), oocytes were exposed to 1.7 °C, 10 °C, and 20 °C (control) for exposure times ranging from 2 to 90 minutes. Cytotoxicity was investigated by exposing oocytes to solutions of Me2SO for exposure times ranging from 2 to 30 minutes. We identify appropriate models and use these to search for an optimal loading protocol, namely the time dependent osmotic damage model (for osmotic damage), the temperature dependent model (for temperature damage), and the external molality Arrhenius power model (for cytotoxicity). We combined these models to estimate total damage during a cryopreservation loading protocol and performed a exhaustive grid search for optimal loading for a given goal intracellular cryoprotectant concentration. Given our fitted models, we find sea urchin oocytes can only be loaded to 0.13 Me2SO v/v with a 50% survival, For reference, levels for vitrification are approximately 0.45 v/v. Our synthesis of damages is the first of its kind, and enables a fundamentally novel approach to modelling survival for cells in general.

Can Sea Urchin Eggs Shelf-Life Be Extended by Cold Storage?

Mar 2022

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Probl Cryobiol Cryomed 2022; 32(1): 068–071

OCIMER Dietas

Dec 2021

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Video protocols for a sea urchin feed preparation

FV6.2 Informe técnico sobre mejores prácticas en los biobancos de invertebrados marinos

Dec 2021

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Guía sobre conceptos básicos de biobancarización especialmente dirigida a la preservación de la biodiversidad marina en biobancos de pequeño y medio tamaño, tanto a nivel institucional como pertenecientes a laboratorios individuales o empresas. El objetivo es dar a conocer procesos críticos para la conservación de muestras y proporcionar un conocimiento básico sobre ellos. Igualmente los autores queremos destacar la necesidad de organizar un biobanco para garantizar la calidad de las muestras almacenadas y su buena conservación. Así como, proporcionar una ventana a todos los recursos actuales disponibles para los gestores e investigadores en caso de que se estén acercando a la temática de la criopreservación y biobancarización por primera vez.

Effect of Low Temperature Storage In Sea Urchin Eggs Viability

Dec 2021

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Mollusk cryopreservation and variability along development and species

Dec 2021

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The main objective of this doctoral thesis consists of the development of cryopreservation protocols for larval stages of marine mollusks, by carrying out a study of involved steps. This will optimize the method to obtain maximal survival rates and provide larvae with unaltered fitness by cryopreservation procedures. Therefore, developed cryopreservation protocols will be suitable for their implementation into research field, aquaculture sector and species conservation. Moreover, a comparative study of data collected from the work with related species could provide general trends about the response or behavior to cryopreservation, hence some standards could be formulated for the development of further cryopreservation protocols for larval stages of related species. The application of cryopreservation techniques on areas of knowledge and aquaculture industry can offer several benefits, as well as the avoidance of severe problems. For example: • In research field, cryopreservation would enhance international interactions between institutions or researchers; it would provide the transport of unaltered biological material until handling. It would also allow greater flexibility in obtaining competent larvae on demand, without the need of sampling, thus avoiding the need to wait for administrative permits and the economic and personal effort they entail.

Cryopreservation of Marine Invertebrates: From Sperm to Complex Larval Stages

Jan 2021

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Marine invertebrates represent the vast majority of marine biodiversity; they are extremely diverse playing a key role in marine ecosystems, thus playing an important role at the socioeconomic level. Some invertebrates such as sea urchins, ascidians, and horse-shoe crabs are very well-known model organisms for research and biocompound discovery. In this chapter we revisit the importance of cryopreservation for the conservation and rational use in research, fisheries management, or aquaculture and provide comprehensive protocols for the cryopreservation of sperm, embryos, and larvae.

Compliance with the Access and Benefit Sharing (ABS) and the regulations derived from the Nagoya Protocol and implications for Biobanking and biodiversity

Jul 2020

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Video Poster: https://www.youtube.com/watch?v=kRoTf1W7Ln0&t=4s

POTENTIAL EFFECTS OF CRYOPRESERVATION ON THE DEVELOPMENT OF POST-THAWED MYTILUS GALLOPROVINCIALIS LARVAL STAGES

Jul 2020

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Selected 72 hold D larva and 10% Ethylene-Glycol + 0.4 M Trehalose in Sea Water. Cryopreservation protocol: equilibration of 60 min. Samples loaded into 0.25 mL straws. Using a programmable freezer: 2 min at 4°C,-1°C/min to-12°C, 2 min at-12°C (seeding),-1°C/min to-35°C, plunging into LN, thawing by immersion into a water bath at 35°C. Larval rearing. Post-thawed larvae cultured into 150 L tanks (Fig. 1A) during 22 days post-fertilization. Provided aeration and feeding. Sampling periodically for survival counts and shell measurements. Settlement. Cultured into settlement drums (Fig. 1B) for 33 days. At the end, mussel juveniles collected for total counts. Culture in rafts. Spat cultured for 27 days to allow the settlement on ropes (Fig. 1C) and transported to rafts to study its development (Fig. 1D). Objective: Study potential long-term effects of cryopreservation and test the capacity of cryopreservation to produce competent larvae. IN ORDER TO: application on improvement of aquaculture production; diminish the reliance of seasonal supply of mussel spat from natural environment; avoid hazards of natural populations; capacity to implement other techniques on aquaculture (selective breeding). Introduction. The shellfish aquaculture is reliable to catches of wild seed for hatchery spat production. Several factors affect the natural shellfish recruitment: environmental variations, the climate change, presence of pathogens, competitors or, predators maritime transport, the pollution , limitations imposed by the seasonal spawning. Results Acknowledgements.

Non-lethal sex identification of sea urchins: method and advantages

Nov 2019

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A simple method for sex determination can reduce the number of sea urchins used in the lab and help to optimize experimental design.

LONG-TERM EFFECTS OF MOLLUSK CRYOPRESERVATION A COMPARATIVE STUDY OF LARVAL SURVIVAL

Jul 2019

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Cryopreservation of marine invertebrate early-life stagesapplications in marine quality assessment and aquaculture

Jul 2014

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El objetivo de esta tesis doctoral es desarrollar protocolos de criopreservación para estados tempranos de desarrollo para varias especies de invertebrados marinos: erizo de mar Paracentrotus lividus, ostra Crassostrea gigas y dos especies de mejillón, Mytilus galoprovinciallis y Perna canaliculus. Este trabajo de divide en cinco capítulos, una introducción general, tres capítulos de experimentación como compilaciones de siete artículos, cada uno de ellos con sus introducciones, material y métodos, resultados y conclusiones. Al final de cada capítulo se resumen las conclusiones específicas del capítulo. El capítulo final es una discusión general de los resultados de la tesis