Outcomes of in vitro fertilization with frozen‐thawed sperm: An analysis of post‐thaw recovery of sperm, embryogenesis, offspring morphology, and skeletogenesis for a cyprinid fish

Abstract

Background Gamete cryopreservation causes cellular damage and death. This study develops cryopreservation techniques for Levantine scraper, and deciphers how early offspring development is affected when eggs are sired with fresh and frozen‐thawed sperm. Results Cryopreserved sperm did not affect embryogenesis at two‐ and four‐cell stages, but impaired embryonic development at eight‐cell stage. Embryonic viability decreased at organogenesis, where only 34‐49% of embryos showed viability with frozen‐thawed sperm. Hatching success and percentage of normal hatched embryos declined when fertilized with frozen‐thawed sperm. Considering only frozen‐thawed cells the dimethyl sulfoxide (DMSO)‐5%, methanol (METH)‐5%, and METH‐10% treatments yielded highest hatch, while METH‐5% and propylene glycol‐5% yielded the most normal hatched embryos. Larval spinal torsion was higher for fresh than frozen‐thawed sperm, where larvae with spinal torsion showed vertebral fusion and shape alterations during exogenous feeding. Both fresh and cryopreserved treatments showed abnormalities in caudal skeleton, while rates of defective yolk‐sacs were higher for cryopreserved sperm, where larvae with defective yolks showed oversized yolk extension. Percentage of larvae with defective heads/eyes were also higher for cryopreserved sperm. Conclusions Results show how frozen‐thawed sperm impairs embryonic/larvae development and identifies frequency and position of abnormalities. Future studies should investigate how sperm DNA damage may have caused these alterations.

Self-reported physical health status of donor sperm-conceived adults

“…71 The long-term health consequences of manual handling and cryopreservation-induced DNA damage on resultant offspring are poorly understood but have been associated with increased incidences of poorer outcomes such as epigenetic disorders, DNA methylation and morbidity in animal studies. [72][73][74] As a method to determine if maternal complications such as preeclampsia had an effect on health outcomes in the cohorts, the data were stratified into those who reported that their mother had maternal complications and those whose mother did not. As was the case with analysis of sex differences, the sample size for those reporting maternal complications in the donor spermconceived cohort of N = 47 reduced the power to detect differences in the donor sperm cohort.…”

Paternal-effect genes revealed through semen cryopreservation inPerca fluviatilis

“…On one hand, sperm motility, fertilization rate and the hatching rates were seen to be high and similar with postthaw semen when compared with use of fresh semen in some Salmoniforms and Esociformes [19][20][21] . On the other hand, studies done so far on few other teleosts (e.g., Levantine scraper, wild brown trout, Atlantic cod) report reduced motility and viability of sperm after freezing-thawing 22 . Also, the post-thaw semen from the same fish when used for in vitro fertilization, there were significant declination in fertilization success when compared with fresh sperm control, followed by abnormalities like cleavage patterns, hatching success, organogenesis etc.…”

Paternal-effect-genes revealed through sperm cryopreservation in Perca fluviatilis

“…On the other hand, studies done so far on few other teleosts (e.g. Levantine scraper, wild brown trout, Atlantic cod) report reduced motility and viability of sperm after freezing-thawing 25 . Also, the post-thaw milt from the same fish when used for in vitro fertilization, there were significant declination in fertilization success when compared with fresh sperm control, followed by abnormalities like cleavage patterns, hatching success, organogenesis etc.…”