Criopreservação de espermatóforos do camarão marinho Penaeus vannamei em diferentes meios diluidores, adicionado de polissacarídeos sulfatados extraídos de Kelps ascophyllum nodosum, para uso em inseminação assistida

Abstract

Sperm cryopreservation constitutes a strategic biotechnology for genetic improvement programs and the conservation of aquatic resources, including shrimp farming. This thesis aimed to evaluate the effect of adding sulfated polysaccharides (SP) extracted from the brown alga Ascophyllum nodosum, as well as the use of powdered coconut water (ACP®) as a diluent, in cryopreservation protocols for spermatophores of the marine shrimp Penaeus vannamei. Two experiments were conducted: in the first, spermatophores were subjected to different freezing media (ACP®, seawater, or 0.2 M sucrose); in the second, the addition or absence of SP to the 0.2 M sucrose solution was evaluated. Spermatophores were frozen using a dry shipper or a freezing ramp, stored in liquid nitrogen for up to 12 months, and subsequently thawed for evaluation of sperm quality. Plasma membrane integrity, sperm morphology, and DNA fragmentation were analyzed, as well as the structural characterization of SP by Fourier-transform infrared spectroscopy. The results demonstrated that the tested protocols were effective in maintaining cellular integrity after thawing, even after long periods of cryogenic storage. The freezing solution and method significantly influenced sperm quality, with emphasis on the dry shipper, which showed better preservation of plasma membrane integrity, including when associated with the use of ACP®. Supplementation of freezing media with SP from A. nodosum positively contributed to the maintenance of post-thaw sperm quality. Assisted insemination using cryopreserved spermatophores resulted in fertilization. It is concluded that the use of algal-derived sulfated polysaccharides and diluents based on powdered coconut water and seawater represents a promising approach for improving cryopreservation protocols for P. vannamei spermatophores, with potential application in assisted reproduction programs and genetic conservation in shrimp farming.