In vitro fertilization with frozen-thawed blue catfish (Ictalurus furcatus) sperm and implications for gene banking

Abstract

Catfish farming constitutes about 75% of total U.S. finfish aquaculture production, for which the channel catfish female (Ictalurus punctatus) × blue catfish male (I. furcatus) hybrid accounts for nearly 60% of the harvest. Current hatchery technology to produce hybrids is labor intensive and requires the sacrifice of males. This is expensive and time-consuming, as blue catfish males require 4 to 7 years to mature. Sperm, even from mature fish, are often of low quality and quantity and do not necessarily give high fertility and hatch. Therefore, paternal gametes should be used sparingly, using the minimum quantity of sperm for fertilization, while maximizing offspring production. This is particularly important when using cryopreserved cells, as each males' sperm is in limited supply. To optimize the use of available sperm cells, the objectives of this study were to (i) compare sperm swimming kinematics and sperm health metrics before and after cryopreservation; (ii) determine the minimum quantity of frozen-thawed sperm required to maximize hatching success; and (iii) decipher how early offspring development is affected when eggs are sired with fresh and frozen-thawed sperm. Sperm kinematics were evaluated by computer-assisted sperm analysis, while health metrics (i.e., viability, oxidative stress, DNA fragmentation) were assessed using fluorescent imaging and flow cytometry. Over 2 spawning seasons, eggs from 18 females were collected and fertilized using cryopreserved sperm from 36 males at 10 sperm-to-egg ratios (ranging from 1.0 × 104 to 9.0 × 105 sperm per egg). Embryos sired with fresh and frozen-thawed sperm were then incubated under common environmental conditions and growth performance was documented over 40 days post-hatch. Generally, sperm kinematic traits [curvilinear velocity (VCL), motility, progressive VCL, and progressive motility] and health metrics (cell membrane viability, oxidative stress, and DNA fragmentation) declined after cryopreservation. Despite this decrease in sperm quality, hatching success using cryopreserved sperm was high, but dependent on the sperm-to-egg ratio. For instance, at ratios of 1.0 × 104 to 5.0 × 104 sperm per egg, hatch increased from 18.0% ± 8.5 to 43.9% ± 8.2. Adding >5.0 × 104 sperm per egg had no significant effect on hatching success. Furthermore, there were generally no differences in morphology or deformity rates in offspring sired with cryopreserved sperm, compared to a fresh control. These data improve understanding of frozen-thawed sperm quality for blue catfish to improve reproductive sustainability and reduce production costs.