Cryopreservation of yellowtail kingfish (Seriola lalandi) sperm: Effects on metabolic enzymes and sperm quality

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Villalobos, Elias Figueroa ; Wilson, Rodolfo ; Pereira, Wellison Amorim ; Merino, Osvaldo ; Perez-Atehortua, Maritza ; Niedmann, Paola ; Avila, Sebastian ; Alvarado, Claudia ; Risopatron, Jennie ; Oliveira, Ricardo Pinheiro S. ; Farias, Jorge G. ; Isler, Ivan Valdebenito ; Villasante, Alejandro ; Romero, Jaime

• Document type: Journal article
• Source: Aquaculture; v. 596, p. 9-pg., 2025-02-15.

Abstract

This study aimed to determine the effects of cryopreservation on sperm functionality and assess post-thaw sperm quality in yellowtail kingfish ( Seriola lalandi). Milt designated for cryopreservation was frozen in a solution containing Cortland (R) medium +1.3 M Me2SO + 0.2 M glucose +2 % BSA in a 1:3 ratio (milt:cryoprotectant). Fresh milt was used as the control. The samples were frozen in 0.5 mL straws using a programmable freezing system. After thawing, the samples were evaluated for motility using Computer-Assisted Sperm Analysis (CASA) with a phase-contrast optical microscope. Additionally, flow cytometry and a multimodal reader were employed to assess various sperm parameters, including plasma membrane integrity (SYBR-14/PI), mitochondrial membrane potential (Delta Psi MMit: JC-1), DNA fragmentation (TUNEL), ATP content (CellTiter-Glo (R)), intracellular calcium levels (Fluo-4 AM), lipid peroxidation (LPO), intracellular superoxide (DHE/SYTOX) and glutathione (GSH/ GSSG) production, activities of glutathione peroxidase (GPx), catalase (CAT) and 8-oxoguanine glycosylase (OGG1), and detection of 8-hydroxydeoxyguanosine (8-OHdG). In cryopreserved milt, significant differences in motility (64.14 +/- 2.2 %), plasma membrane integrity (61.5 +/- 2.7 %), mitochondrial membrane potential (55.4 +/- 2.4 %), DNA fragmentation (6.7 +/- 1.1 %), ATP content (7.5 +/- 1.5 nmol/109 spermatozoa), and intracellular calcium levels (59.8 +/- 2.5 nM/109 spermatozoa) were observed compared to fresh milt. Differences were also noted in LPO, O 2i- , GPx, GSH/GSSG, CAT, 8-OHdG and OGG1 between the cryopreserved and control groups. Additionally, correlations between sperm quality parameters, oxidative stress markers, and enzyme activity in cryopreserved spermatozoa were determined. These results suggest that cryopreservation induces significant alterations derived from oxidative damage in the functionality of yellowtail kingfish spermatozoa, and thus further research is necessary to explore the need for using antioxidants in the cryopreservation process of sperm from this species. (AU)

• FAPESP's process: 18/25511-1 - Bioprospecting of bacteriocin-producing bacteria: from culture optimization to the application in animal production systems
• Grantee: Ricardo Pinheiro de Souza Oliveira
• Support Opportunities: Research Projects - Thematic Grants
• FAPESP's process: 24/00151-3 - Utilization of agar extraction by-product from Gelidium spp. as an alternative ingredient for aquafeeds: Towards the circular economy and the sustainability
• Grantee: Wellison Amorim Pereira
• Support Opportunities: Scholarships abroad - Research Internship - Post-doctor