Atroshchenko Mikhail Sperm Stallion
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Atroshchenko Mikhail Sperm Stallion
All-Russian Scientific Research Institute for Horse Breeding (ARRIH), Ryazan Oblast, Rybnovskij District 391105, Russia
State Research Center—Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency (SRC-FMBC), 46, Zhivopisnaya Str, Moscow 123098, Russia
Semenov Institute of Chemical Physics, Russian Academy of Sciences, Moscow 119991, Russia
Center for Veterinary Cell Medicine, 8/1 Tvardovsky Str, Moscow 123458, Russia
Moscow Institute of Physics and Technology, Dolgoprudny, Moscow Region 141700, Russia
Institute of Cell Biophysics, Russian Academy of Sciences, Pushchino, Moscow Region 142290, Russia
Author to whom correspondence should be addressed.
Animals 2019 , 9 (12), 1099; https://doi.org/10.3390/ani9121099
Received: 11 October 2019 / Revised: 26 November 2019 / Accepted: 6 December 2019 / Published: 9 December 2019
(This article belongs to the Section Animal Reproduction )
It is very important to evaluate correctly the fertilizing capacity of frozen sperm before carrying out the artificial insemination of mares. It is not always possible to perform complex laboratory tests under the stud farm’s conditions. The purpose of our study was to determine the minimum set of relatively simple laboratory tests sufficient to confirm the feasibility of frozen/thawed sperm for insemination and to predict the pregnancy rate in mares. We found that sperm characteristics such as activity, survival at low temperature, and DNA damage are sufficient to determine the quality of frozen/thawed sperm. These characteristics can be determined by simple tests and assays that can be used for increasing the effectiveness of artificial insemination not only in horse breeding but also in other areas of livestock and even in reproductive medicine.
The main goal of our study was to determine a set of thawed stallion sperm characteristics that have predictive value for the pregnancy rate (PR) of mares after artificial insemination (AI). DNA fragmentation and survival of sperm during hypothermic storage were studied in addition to routinely determined semen characteristics such as concentration, percentage of motile spermatozoa, and morphology. To estimate DNA fragmentation, a modified hallo assay was applied. Sperm survival was determined within hours as the ability of spermatozoa to maintain progressive motility (PM) during the storage of ejaculate diluted with lactose-chelate-citrate-yolk (LCCY) medium at +4 °C. Strong positive correlation between PR and thawed sperm motility ( r = 0.90, p < 0.05) as well as between PR and sperm survival ( r = 084, p < 0.05) was revealed. There was also a strong negative correlation between PR and DNA damages in spermatozoa ( r = −0.94, p < 0.05). We found no dependence of PR on normal morphology spermatozoa percentage in thawed semen. We concluded that the sperm activity, survival, and DNA fragmentation should be considered as the sufficient reproductive characteristics of semen to evaluate the quality of frozen/thawed sperm and prediction of PR.
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Keywords:
horse breeding; sperm; cryopreservation; reproductive characteristics; DNA fragmentation
horse breeding ; sperm ; cryopreservation ; reproductive characteristics ; DNA fragmentation
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Atroshchenko, M.M.; Arkhangelskaya, E.; Isaev, D.A.; Stavitsky, S.B.; Zaitsev, A.M.; Kalaschnikov, V.V.; Leonov, S.; Osipov, A.N. Reproductive Characteristics of Thawed Stallion Sperm. Animals 2019 , 9 , 1099.
https://doi.org/10.3390/ani9121099
Atroshchenko MM, Arkhangelskaya E, Isaev DA, Stavitsky SB, Zaitsev AM, Kalaschnikov VV, Leonov S, Osipov AN. Reproductive Characteristics of Thawed Stallion Sperm. Animals . 2019; 9(12):1099.
https://doi.org/10.3390/ani9121099
Atroshchenko, Mikhail M.; Arkhangelskaya, Ekaterina; Isaev, Dmitry A.; Stavitsky, Sergey B.; Zaitsev, Alexander M.; Kalaschnikov, Valery V.; Leonov, Sergey; Osipov, Andreyan N. 2019. "Reproductive Characteristics of Thawed Stallion Sperm" Animals 9, no. 12: 1099.
https://doi.org/10.3390/ani9121099
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Reproductive Characteristics of Thawed Stallion Sperm
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Mikhail M. Atroshchenko , PhD, Senior Researcher of the All-Russian Research Institute for Horse Breeding (391105, Russia, Ryazan region, Rybnovskii district, settlement Divovo); e-mail: atromiks-77@mail.ru Elizaveta E. Bragina , Dr.Sc., Senior Researcher of the A.N. Belozersky Research Institute of Physico-Chemical Biology of the Lomonosov Moscow State University (119992, Russia, Moscow, Leninskie gory, 1/40); e-mail bragor@mail.ru Alexander M. Zaitsev , PhD, Director of the All-Russian Research Institute for Horse Breeding (391105, Russia, Ryazan region, Rybnovskii district, settlement Divovo); e-mail: amzaitceff@mail.ru Valeriy V. Kalashnikov , Dr.Sc., Academician of RAS, Research Adviser of the All-Russian Research Institute for Horse Breeding (391105, Russia, Ryazan region, Rybnovskii district, settlement Divovo); e-mail: vniik08@mail.ru Valentina A. Naumenkova , PhD, Senior Researcher of the All-Russian Research Institute for Horse Breeding (391105, Russia, Ryazan region, Rybnovskii district, settlement Divovo); e-mail: naumenkova.00@mail.ru Anna M. Kudlaeva , Assistant of the Ryazan State Medical University (390026, Russia, Ryazan, Vysokovol'tnaya street, 9); e-mail: anyakudlaeva@mail.ru Elena V. Nikitkina , PhD, Leading Researcher of the All-Russian Research Institute of Farm Animal Genetics and Breeding Branch of the L.K. Ernst Federal Science Center for Animal Husbandry (196601, Russia, St. Petersburg, Pushkin, Moscow Shosse, 55a); e-mail: nikitkinae@mail.ru
Atroshchenko M.M., Bragina E.E., Zaitsev A.M., Kalashnikov V.V., Naumenkova V.A., Kudlaeva A.M., Nikitkina E.V. 2019. Conservation of genetic resources in horse breeding and major structural damages of sperm during semen cryopreservation in stallions. Nature Conservation Research 4(Suppl.2): 78–82. https://dx.doi.org/10.24189/ncr.2019.024
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 .
acrosomes, electron microscopy, equids, Equus ferus caballus , mitochondria, progressive motility, ultrastructure
Received: 14.02.2019. Revised: 11.04.2019. Accepted: 15.04.2019.
Atroshchenko M.M., Bragina E.E. 2011. Change in the ultrastructure of stallion spermatozoa under the effect of cryopreservation. Russian Agricultural Sciences 3(2): 175–178. DOI: 10.3103/S1068367411020029 Atroshchenko M.M., Kalaschnikov V.V., Bragina Ye.Ye., Zaitsev A.M. 2017. Comparative study of the structural integrity of spermatozoa in epididymal, ejaculated and cryopreserved semen of stallions. Sel'skokhozyaistvennaya biologiya 52(2): 274–281. DOI: 10.15389/agrobiology.2017.2.274eng Bagirov V.A., Kalaschnikov V.V., Zaitsev A.M., Atroshchenko M.M., Miroshnikov S.A., Zavialov O.A., Frolov A.N. 2017. Reproductive function in purebred arabian stallions as related to the levels of chemical elements in mane hair samples. Sel'skokhozyaistvennaya biologiya 52(6): 1184–1193. DOI: 10.15389/agrobiology.2017.6.1184eng Bragina Ye.Ye., Bocharova Ye.N. 2014. Quantitative electron microscopic examination of sperm for male infertility diagnosis. Andrology and Genital Surgery 15(1): 41–50. DOI: 10.17650/2070–9781–2014–1–41–50 [In Russian] Lebedeva L.F., Atroshchenko M.M., Burmistrova S.A. 2015. Main factors affecting mare insemination with cryopreserved domestic and foreign sperm. Sel'skokhozyaistvennaya biologiya 50(2): 476–485. DOI: 10.15389/agrobiology.2015.4.476eng Naumenkova V.A., Atroshchenko M.M., Lebedeva L.F., Khalilov R.A., Ryabova T.N. 2013. Studying of duration of periods of storage of stallions' sperm in a condition of anabiosis. Konevodstvo i konnyi sport 5: 15–17. [In Russian] Pesch S., Bostedt H., Failing K., Bergmannc M. 2006. Advanced fertility diagnosis in stallion semen using transmission electron microscopy. Animal Reproduction Science 91(3–4): 285–298. DOI: 10.1016/j.anireprosci.2005.04.004 Zamboni L. 1987. The ultrastructural pathology of the spermatozoon as a cause of infertility: the role of electron microscopy in the evaluation of semen quality. Fertility and Sterility 48(5): 711–734. DOI:10.1016/S0015-0282(16)59520-X
CONSERVATION OF GENETIC RESOURCES IN HORSE BREEDING AND MAJOR STRUCTURAL DAMAGES OF SPERM DURING SEMEN CRYOPRESERVATION IN STALLIONS
https://dx.doi.org/10.24189/ncr.2019.024
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