Viability and fertilizing capacity of cryopreserved sperm from three North American acipenseriform species: a retrospective study

Abstract

Populations of sturgeon across the globe are threatened due to unregulated harvest and habitat loss, and the status varies among species across North America. Ready access to viable and functional sperm would contribute to recovery programmes for these species. In this study, we examined the motility, viability (cell membrane integrity) of cryopreserved sperm from three North American acipenseriform species and fertilizing capacity. Milt samples were collected from captive shortnose sturgeon (Acipenser brevirostrum), wild paddlefish (Polyodon spathula) and pallid sturgeon (Scaphirhynchus albus) and cryopreserved using combinations of Modified TsvetkovaÕs (MT) extender, Original TsvetkovaÕs extender, and modified HanksÕ balanced salt solution, along with the cryoprotectants methanol (MeOH) or dimethyl sulfoxide (DMSO). A dual-staining technique using the fluorescent stains SYBR-14 and propidium iodide was employed with flow cytometry to determine the percentages of spermatozoa that were viable by virtue of having intact membranes. The percentage of viable spermatozoa ranged from 5% to 12% in shortnose sturgeon, 30-59% in paddlefish, and 44-58% in pallid sturgeon. In the first experiment with shortnose sturgeon sperm, methanol allowed for higher values for dependent variables than did DMSO, and sperm viability generally correlated with post-thaw motility. However, fertilization rate, neurulation, or hatching rates were independent from these factors. In the second experiment with shortnose sturgeon, 5% MeOH combined with MT yielded higher values for all parameters tested than the other combinations: viability was correlated with motility, fertilization rate, and hatching rate. Overall, viability and post-thaw motility was not affected by the use of hyperosmotic extenders (OT) or cryoprotectants (DMSO), but their use decreased fertilization percentages. For paddlefish sperm (experiment 3), MT combined with 10% MeOH was clearly a good choice for cryopreservation; viability and motility results were correlated, but independent of fertilization. For pallid sturgeon sperm (experiment 4), MT with 5-10% MeOH showed significantly higher sperm quality and fertilization parameters. Membrane integrity can be used as a predictor of fertilization by cryopreserved sperm, however additional sperm quality parameters, supplementary to motility and membrane integrity, would be useful in the refining and optimizing cryopreservation protocols with acipenseriform sperm.

Sperm quality assessments for endangered razorback suckers Xyrauchen texanus

“…The lack of accord observed between fertilization rates and standard semen parameters or functional assessments (Linhart et al 2005) may be partially explained by sublethal damage. In a study with sturgeon spermatozoa, various combinations of extenders and cryoprotectants yielding different osmolalities resulted in a variety of fertilization rates, even when post-thaw sperm motility and viability were similar to pre-freeze values (Horváth et al 2008). The diverse biological properties of spermatozoa and dissimilarities among species underscore the use of a combination of sperm quality assessments, particularly in the less well-studied animals.…”

“…For endangered aquatic species and their surrogates (Wayman 2003, Jenkins & Goodbred 2005, Jenkins & Draugelis-Dale 2006, Goodbred et al 2007, Horváth et al 2008, and for fish and shellfish (Segovia et al 2000, Cabrita et al 2001, Paniagua-Chavez et al 2006, Horvá th et al 2008, viability and mitochondrial function assessments have proven useful. Mitochondrial respiration is responsible for ATP synthesis and sperm motility (Johnson et al 1980), whereby mitochondrial membrane electrical potential has been correlated with mammalian spermatozoal normal morphology, concentration, and motility (Ericsson et al 1993, Marchetti et al 2002 and with motilities in Nile tilapia Oreochromis niloticus (Segovia et al 2000) and in eastern oyster Crassostrea virginica (Paniagua-Chavez et al 2006).…”

“…Mitochondrial respiration is responsible for ATP synthesis and sperm motility (Johnson et al 1980), whereby mitochondrial membrane electrical potential has been correlated with mammalian spermatozoal normal morphology, concentration, and motility (Ericsson et al 1993, Marchetti et al 2002 and with motilities in Nile tilapia Oreochromis niloticus (Segovia et al 2000) and in eastern oyster Crassostrea virginica (Paniagua-Chavez et al 2006). Although mitochondrial membrane potential was the most sensitive test in determining human sperm quality (Marchetti et al 2002) and likely provides insight with paddlefish Polyodon spathula (Horváth et al 2008), sperm DNA integrity has been expected to yield superior diagnostic and prognostic information on fertility potential (Agarwal & Said 2003).…”

Amine reactive dyes: An alternative to estimate membrane integrity in fish sperm cells

“…The increasing demand for cryopreserved germplasm goes along with an increasing demand for high quality in post-thaw motility, fertilization rates, and hatching rates (Horváth et al, 2008). Standardized quality control checkpoints along a given process are highly recommended to avoid lack of reproducibility (Baker, 2016).…”

Hatchery-scale trials using cryopreserved spermatozoa of black-lip pearl oyster,Pinctada margaritifera

“…24, 219-223 (2011) obtained using thawed sperm. The integrity of the spermatozoon plasma membrane is required to maintain fertilization capacity (Graham et al 1990;Paniagua-Chávez et al 2006), but this membrane may be damaged by the freezing-thawing process (Grzyb et al 2003;Cabrita et al 2005;Horvat et al 2008). The evaluation of sperm membrane integrity in marine species is based on the use of the fluorescent dyes using microscopy and flow cytometry analysis (Grzyb et al 2003;Adams et al 2003).…”