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Cryopreservation of canine sperm : preserving for better breeding

(2023)
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Abstract
At the end of the 19th century, the original functional dogs were divided into morphotypes to create the unique phenotypic diversity that modern breeds display. Breed creation, however, has caused the gene flow between distinct dog populations to cease, resulting in a loss of genetic diversity in many dog breeds. Chapter 1 explains how domestication and breed creation took place and details the genetic strategies developed to improve genetic diversity in dogs. Among these, assisted reproduction techniques are particularly helpful in optimizing breeding strategies but are currently limited to artificial insemination and sperm cryopreservation in dogs. Unfortunately, cryopreservation of dog spermatozoa is hampered by individual variability in sperm cryotolerance, confining its application to a selection of dogs. Therefore, this thesis aimed to identify ejaculates of suboptimal quality and optimize sperm cryopreservation in dogs (chapter 2). In chapter 3, we characterized dogs (age, breed, sperm count, and number of sperm collections) presented for sperm cryopreservation and described the use and popularity of frozen sperm by conducting a descriptive analysis of the 3,090 ejaculates frozen over the last nine years at two freezing centers in the Netherlands and Belgium. Our results showed that sperm cryopreservation gained popularity in recent years and that dog breeders used sperm banks for short- and long-term storage of frozen sperm, mainly before international exportation. We also confirmed that dog breed size influenced the total sperm count, and a second semen collection could increase the amount of sperm to cryopreserve. As semen collection in dogs is not always successful, and a complete semen analysis requires experience and specialized equipment, we investigated the value of anti-Müllerian hormone (AMH) as a serum biomarker for gonadal function in chapter 4. Our results showed that serum AMH concentration was negatively correlated with sperm total motility, progressive motility, and normal morphology. Moreover, we defined that a serum AMH concentration below 5.54 µg/L accurately predicted optimal semen quality in 83% of cases. At higher values, however, semen quality could not be reliably predicted. We concluded that assessing serum AMH concentration could serve as a biomarker of spermatogenesis and identify dogs that would benefit from a complete semen analysis to evaluate its quality. Sperm motility analysis has been automated for decades following the development of computer-assisted sperm analysis (CASA) systems. However, these automated systems are expensive and unsuited for field analysis. In chapter 5, we investigated the software iSperm®, designed for the iPad Mini 4, as a potential alternative to CASA systems. Our results showed positive correlations for all kinematic parameters, although the reported values differed between the two devices. However, software improvements are warranted to make it a valid alternative to computerized automated systems. In chapter 6, we explored the potential positive effect of sperm selection by single layer centrifugation (SLC) on sperm freezability. Sperm selection by SLC improved semen quality in dogs with optimal and suboptimal semen before cryopreservation. After thawing, SLC-selected sperm had a better quality compared to unselected sperm. However, the cryosurvival of sperm of suboptimal quality ejaculates remained inferior to that of optimal quality ejaculates. We concluded that isolating a subpopulation of high-quality spermatozoa before cryopreservation was an efficient technique to obtain sperm of better quality after thawing but was not the only treatment that should be undertaken to improve the freezability of lower-quality ejaculates. Finally, the results of this thesis were discussed in chapter 7 in light of current scientific knowledge, and possible lines of future research were suggested.

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MLA
Domain, Guillaume. Cryopreservation of Canine Sperm : Preserving for Better Breeding. Ghent University. Faculty of Veterinary Medicine, 2023.
APA
Domain, G. (2023). Cryopreservation of canine sperm : preserving for better breeding. Ghent University. Faculty of Veterinary Medicine, Merelbeke, Belgium.
Chicago author-date
Domain, Guillaume. 2023. “Cryopreservation of Canine Sperm : Preserving for Better Breeding.” Merelbeke, Belgium: Ghent University. Faculty of Veterinary Medicine.
Chicago author-date (all authors)
Domain, Guillaume. 2023. “Cryopreservation of Canine Sperm : Preserving for Better Breeding.” Merelbeke, Belgium: Ghent University. Faculty of Veterinary Medicine.
Vancouver
1.
Domain G. Cryopreservation of canine sperm : preserving for better breeding. [Merelbeke, Belgium]: Ghent University. Faculty of Veterinary Medicine; 2023.
IEEE
[1]
G. Domain, “Cryopreservation of canine sperm : preserving for better breeding,” Ghent University. Faculty of Veterinary Medicine, Merelbeke, Belgium, 2023.
@phdthesis{01HFKCR4HBQ55EQVBVYKRAFQ31,
  abstract     = {{At the end of the 19th century, the original functional dogs were divided into morphotypes to create the unique phenotypic diversity that modern breeds display. Breed creation, however, has caused the gene flow between distinct dog populations to cease, resulting in a loss of genetic diversity in many dog breeds. Chapter 1 explains how domestication and breed creation took place and details the genetic strategies developed to improve genetic diversity in dogs. Among these, assisted reproduction techniques are particularly helpful in optimizing breeding strategies but are currently limited to artificial insemination and sperm cryopreservation in dogs. Unfortunately, cryopreservation of dog spermatozoa is hampered by individual variability in sperm cryotolerance, confining its application to a selection of dogs. Therefore, this thesis aimed to identify ejaculates of suboptimal quality and optimize sperm cryopreservation in dogs (chapter 2).
In chapter 3, we characterized dogs (age, breed, sperm count, and number of sperm collections) presented for sperm cryopreservation and described the use and popularity of frozen sperm by conducting a descriptive analysis of the 3,090 ejaculates frozen over the last nine years at two freezing centers in the Netherlands and Belgium. Our results showed that sperm cryopreservation gained popularity in recent years and that dog breeders used sperm banks for short- and long-term storage of frozen sperm, mainly before international exportation. We also confirmed that dog breed size influenced the total sperm count, and a second semen collection could increase the amount of sperm to cryopreserve. 
As semen collection in dogs is not always successful, and a complete semen analysis requires experience and specialized equipment, we investigated the value of anti-Müllerian hormone (AMH) as a serum biomarker for gonadal function in chapter 4. Our results showed that serum AMH concentration was negatively correlated with sperm total motility, progressive motility, and normal morphology. Moreover, we defined that a serum AMH concentration below 5.54 µg/L accurately predicted optimal semen quality in 83% of cases. At higher values, however, semen quality could not be reliably predicted. We concluded that assessing serum AMH concentration could serve as a biomarker of spermatogenesis and identify dogs that would benefit from a complete semen analysis to evaluate its quality.
Sperm motility analysis has been automated for decades following the development of computer-assisted sperm analysis (CASA) systems. However, these automated systems are expensive and unsuited for field analysis. In chapter 5, we investigated the software iSperm®, designed for the iPad Mini 4, as a potential alternative to CASA systems. Our results showed positive correlations for all kinematic parameters, although the reported values differed between the two devices. However, software improvements are warranted to make it a valid alternative to computerized automated systems.
In chapter 6, we explored the potential positive effect of sperm selection by single layer centrifugation (SLC) on sperm freezability. Sperm selection by SLC improved semen quality in dogs with optimal and suboptimal semen before cryopreservation. After thawing, SLC-selected sperm had a better quality compared to unselected sperm. However, the cryosurvival of sperm of suboptimal quality ejaculates remained inferior to that of optimal quality ejaculates. We concluded that isolating a subpopulation of high-quality spermatozoa before cryopreservation was an efficient technique to obtain sperm of better quality after thawing but was not the only treatment that should be undertaken to improve the freezability of lower-quality ejaculates.
Finally, the results of this thesis were discussed in chapter 7 in light of current scientific knowledge, and possible lines of future research were suggested.}},
  author       = {{Domain, Guillaume}},
  language     = {{eng}},
  pages        = {{VIII, 191}},
  publisher    = {{Ghent University. Faculty of Veterinary Medicine}},
  school       = {{Ghent University}},
  title        = {{Cryopreservation of canine sperm : preserving for better breeding}},
  year         = {{2023}},
}