Assisted reproductive technologies have been used in zoos and breeding centres to preserve rare and endangered species of wild animals over the past three decades. The most common way to preserve the genetic material of farm and wildlife animals is to create cryobanks of semen producers and embryos. Studies of domestic species under controlled conditions provide an excellent opportunity to develop effective semen handling techniques for application to wild species of the genus Equus. The All-Russian Research Institute for Horse Breeding (ARRIH) began studies on the cryopreservation of genetic resources in the early 1950s. In 1954 the ARRIH got the world's first foal from the artificial insemination by cryopreserved sperm. The collection of genetic resources of ARRIH has been successfully working since 1972 and contains nowadays cryopreserved sperm from 56 stallions of 16 breeds of domestic and foreign selections. Freezing and subsequent thawing of stallion sperm leads to a decrease in progressive sperm motility and an increase in the number of sperm with structural pathologies. Electron microscopy is one of the most accurate methods used to assess the structural integrity of sperm. We conducted an electron microscopic study of native and cryopreserved sperm of 35 stallions of riding and trotting breeds, aged 4 to 22 years (mean age 11.3 ± 0.9 years). We found that organoids with a denser structure are more resistant to freezing and subsequent thawing of sperm. The use of stallion sperm cryopreservation according to the standard technology of ARRIH leads to a decrease in the number of sperms with intact heads by an average of 19.7%, and the share of sperm with intact heads in cryopreserved sperm is 49.1 ± 2.7% (p < 0.001). The most negatively cryopreservation of semen affects an acrosome of sperms. The main pathologies of acrosomes in sperm cryopreservation are acrosome hypoplasia and its degradation (secondary absence of acrosome). The number of spermatozoa with acrosome hypoplasia and the absence of internal content after cryopreservation increases by 20.9%, and the share of such sperms is 31.7% in thawed sperm (p < 0.001). The number of sperm with acrosome degradation during sperm freezing and thawing increases by 10.4%, and is 18.6 ± 2.4% in average. The share of spermatozoa with normal mitochondrial structure in native sperm is 89.3 ± 2.0% in average, in cryopreserved sperm-decreases by 6.5%. The share of spermatozoa with normal axoneme after cryopreservation decreases slowly, averaging by 4.4% and is 81.4 ± 2.0%. Some of the available assisted reproduction technologies will need to be further optimised in domestic horses before they can be applied to the endangered Przewalski's horse Equus ferus przewalskii.

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