Cryopreserved sperm does not affect larval ontogeny and quality in Rhamdia quelen.

Abstract

Fish sperm cryopreservation is an important technique for optimizing juvenile production in aquaculture stations and laboratories and contributing to the conservation of endangered species. Despite its benefits, the cryopreservation process can cause cellular damage, affecting spermatozoa quality and offspring viability. This study aimed to evaluate the larval development of jundiá Rhamdia quelen originating from cryopreserved sperm. Larvae were obtained from artificial reproduction using oocyte samples from four females combined with fresh (Control) or cryopreserved/thawed sperm. The semen was diluted in the cryoprotective solution (1:3 ratio) consisting of skimmed milk powder (5%), methanol (10%), and fructose (5%), and was packaged into 0.25 mL straws. The straws were then stored and cooled in liquid nitrogen vapor for 18 h. The straws were individually warmed in a water bath at 25 °C for 10 s to thaw the samples. The experiments were performed in triplicates. Sperm quality, fertilization, hatching, and larval development were evaluated. After larval hatching, six larval collections were performed (5, 10, 15, 20, and 25 days after hatching), and 15 larvae were sampled per collection per treatment. Cryopreservation reduced sperm motility (70.48 ± 7.70 fresh to 41.36 ± 4.80 cryopreserved semen), progressivity (3874 fresh to 2505 cryopreserved semen), and beat cross frequency (55.83 ± 155 fresh to 50.22 ± 190 cryopreserved semen). Increased the percentage of sperm with abnormal morphology and increased most sperm pathologies. Furthermore, the fertilization rate was lower in the cryopreserved group (63.1 ± 18, and 83.72 ± 7.59 for fresh semen), while hatching was not different between groups (65.3 ± 18.05 fresh, 48.89 ± 21.77 cryopreserved semen) Otherwise, the initial larval development morphology showed no difference in the appearance of structures such as the presence of the vitelline structure, pigmentation pattern, development of the anal pore, embryonic membrane, eye, barbells, notochord flexion, and fin rays, for both treatments. There was no significant difference in the frequency of structures between larvae from fresh and cryopreserved/thawed sperm, revealing a similar developmental pattern in both treatments. In conclusion, the cryopreservation protocol affects sperm quality; however, those sperm able to fertilize the oocytes originate normal larvae with regular larval development of R. quelen up to 25 days old.