Spotted wolffish (Anarhichas minor) sperm cryopreservation in 5-mL cryovials

Abstract

In spotted wolffish Anarhichas minor aquaculture, cryopreservation is used to secure sperm availability throughout the entire spawning season. Under current protocols, sperm is cryopreserved in 0.5-mL straws. This implies thawing a considerable number of straws for insemination with cryopreserved sperm. In this work, we scale up the spotted wolffish sperm cryopreservation procedure through the development of a protocol for sperm cryopreservation in 5-mL cryovials. Different freezing (distances from the liquid nitrogen surface) and thawing rates were tested. The best results were obtained with cryovials frozen at a distance of 1.5 cm from the liquid nitrogen surface and thawed either at 15 or 10°C for 4 and 6 min, respectively. Under these conditions, similar percentage of motile cells, sperm velocity and percentage of viable cells were obtained in comparison with the sperm cryopreserved in the traditional 0.5-mL straws. This protocol will facilitate the process of insemination with cryopreserved sperm in the spotted wolffish hatcheries.

The status of spotted wolffish, Anarhichas minor: A commercially ready species for U.S. marine aquaculture?

“…Unpredictable sperm quality and quantity require maintenance of large numbers of males, which results in poor utilization of hatchery facilities. Beirao et al (2020) observed a desynchronization between male and female maturation, with males maturing earlier than females. The level of plasma androgens (11-ketotestosterone; 11-Kt) that direct testicular development is considered low in spotted wolffish, compared to other fish species, possibly as a result of physiological stress associated with a suboptimal rearing environment usually mediated by the hormone cortisol.…”

Spotted Wolffish Broodstock Management and Egg Production: Retrospective, Current Status, and Research Priorities

“…This method provides an initial assessment of the motility and is often used in fertilization trials [110]. Nonetheless, the use of computer-assisted sperm analysis systems (CASA) provides a more objective evaluation and has also been employed in spotted wolffish sperm quality studies [25,102,106,111]. The CASA systems, besides the percentage motility, measure different sperm swimming velocity parameters, such as curvilinear velocity (VCL) or straight-line velocity (VSL), parameters associated with the trajectory such as linearity (LIN), and other parameters such as wobble of the sperm head; for a detailed description, see [112].…”

“…It is often difficult to find spermiating males and with enough sperm volume at the end of the spawning season [1], and this represents a challenge to fertilize the last egg batches. Consequently, the development of sperm storage techniques has been extensively researched [25,102,105,106,109,111]. Cryobanking of the sperm of genotyped broodstock populations is justified in order to control undesirable inbreeding levels.…”

“…Cryobanking of the sperm of genotyped broodstock populations is justified in order to control undesirable inbreeding levels. Different studies have optimized different aspects of the cryopreservation process such as cryoprotectant and concentration [102,109], freezing and thawing rate [102,109], or cryopreservation volume [109,111]. Compared with other marine fish, spotted wolffish sperm is quite resilient to cryopreservation.…”

“…This suggests that the current procedure necessitates a substantial number of small straws which proved to be complicated and time-consuming. Thus, large cryovials (5-10 mL) are the option preferred by the industry [111].…”