Optimization of management procedures for sea urchin (paracentrotus lividus) artificial reproduction

• Autor(a) principal: Rodrigues, Daniel Alexandre Cardoso
• Data de Publicação: 2021
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10400.1/17707
• topic: Sémen; Reprodução in vitro; Criopreservação; Seleção de progenitores; Paracentrotus lividus; Domínio/Área Científica::Ciências Naturais::Outras Ciências Naturais

Aquaculture industry has a huge contribution for the world food production, and is extremely necessary to develop and optimize the production techniques, especially not to depend totally on wild stocks. In the case of the sea urchin Paracentrotus lividus, the demand increase of their roe is provoking a drastically decrease of their wild stocks, particularity in the recent years. This demand for sea urchin roe is arousing interest for the industry for a new profitable product and for the recovering and preserve wild populations. In this sense, new research needs to guarantee optimization of the techniques used to reduce the effort and the production cost. The selection of breeders is a very important aspect for sea urchin industry, since the success of the reproduction and quality of their product are dependent of the breeders. In addition to the high cost of acquiring wild specimens, it is not viable in the long term. Therefore, is important to study the possibility of use breeders from captive generations. The optimization of in vitro reproduction techniques is crucial for an efficiency production. The time that gametes need to be in contact for the best fertilization rate and the ratio of spermatozoa per oocyte are two of the principal aspects to be considered in an optimization of an in vitro reproduction protocol. Through preservation and cryopreservation technology is possible to promote alternatives techniques to contribute for the resource management efficiency of P. lividus. The objectives of this work are to study the hypothesis of using male wild sea urchins from different locations and captive generations specimens as breeders, optimize in vitro protocols using different contact times and different spermatozoa to oocyte ratio, and compare cryopreservation and preservation techniques for the storage of the sperm. To study the possibility of use captive generations as breeders were evaluated the sperm quality of each case, for this evaluation were analyzed the viability and motility, being used two approaches for the last one (mean values and subpopulation). The optimization of fertilization outcomes was evaluated counting the number of fertilized eggs using a counting chamber (Sedgwick) in optical microscope. Moreover, the comparation of the preservation and cryopreservation techniques was analyzed the motility by mean values, viability and DNA fragmentation (comet assay). The motility analysis were assessed by CASA system, viability through staining techniques and counting at a fluorescent microscope, DNA fragmentation was quantified the DNA tail. Significant differences were observed (p<0.05) in breeders selection, revealing the domestication seems to have a positive effect in P. lividus sperm quality. The in vitro spermatozoa to oocyte ration trial reveled significant differences between all ratios, where the 20 000 spermatozoa to oocyte ratio had the higher value. Contact times chosen differed negatively from the control. The preservation technique had higher viability in the first 6 days comparing to cryopreservation, although had no damage in DNA were detected in both. The present study revealed that domestication seems to be a good step to optimize the breeders sperm quality. With the higher spermatozoa to oocyte ratio used is possible to reach almost the total egg fertilization percentage. The preservation at 4ºC is a good technique to preserve the sperm quality until 6 days, cryopreservation seems to be better from there if the protocol is optimized.