A b s t r a c tA population of the Russian White chickens, bred at the gene pool farm of ARRIFAGB for 25 generations using individual selection, is characterized by resistance to a lowered temperature in the early postnatal period and white colour of the embryonic down. In 2002-2012, breeding was carried out by panmixia, and by now a new population of the Russian White chickens has been formed on the basis of the surviving stock. Comparison of the genetic variability of this population and the archival DNA of representatives of the 2001 population using microarray screening technology will help to assess the population structure and the preservation of the unique characteristics of its genome. The material for the study was DNA extracted from 162 chicken blood samples. Two groups of the Russian White breed were studied, the 2001 population and the current population. Genome-wide analysis using single nucleotide markers (SNP) included screening by means of the Illumina Chicken 60K SNP iSelect BeadChip microarray. Quality control of genotyping, determination of the population genetic structure by multidimensional scaling (MDS), calculation of linkage disequilibrium (LD) and allele frequency in the groups were carried out using PLINK 1.9 software program. The construction of a cluster delimitation model based on SNP genotypes was carried out using the ADMIXTURE program. According to the MDS analysis results, the current population can be divided into four MDS groups, which, when compared to the data of the pedigree, adequately reflect the origin of the studied individuals. The representatives of the ancestral population were genetically similar to the MDS3 group of the current population. Using the F-statistic of the twoway analysis of variance, a significant effect of the group, chromosome, chromosome in the group, and the distance between SNP markers on LD (r 2 ) values was observed. In the 2001 group, the maximum r 2 and the high incidence of LD equal to 1 were observed for all chromosomes, with a distance between SNP markers being 500-1000 Kb. There was also the greatest number of monomorphic alleles in this group. Based on the SNP analysis, we may conclude that the current Russian White chicken population is characterized by the disintegration of long LD regions of the ancestral population. Modelling clusters using the ADMIXTURE program revealed differences between the current population groups determined by MDS analysis. The groups composed of individuals included in MDS1 and MDS2 had a homogeneous structure and differed from each other at K = 4 and K = 5. The MDS4 group formed a genetically heterogeneous cluster different from the MDS1 and MDS2 groups at K of 2-5. The MDS3 group was phylogenetically close to the 2001 population (at K of 2-5). In general, the analysis of the current gene pool population of the Russian White chickens showed its heterogeneity while one of its groups (MDS3) was similar to the ancestral population of 2001, which in turn is characterized by a large number of monomorphic alleles and...

The fertilizing ability of stallion sperm after freezing is lower than in other species. The search for the optimal extender, combination of extenders, and the freezing protocol is relevant. The aim of this study was to compare lactose-chelate-citrate-yolk (LCCY) extender, usually used in Russia, and Steridyl® (Minitube) for freezing sperm of stallions. Steridyl is a concentrated extender medium for freezing ruminant semen. It already contains sterilized egg yolk. Semen was collected from nine stallions, aged from 7 to 12 years old. The total and progressive motility of sperm frozen in Steridyl was significantly higher than in semen frozen in LCCY. The number of spermatozoa with normal morphology in samples frozen in LCCY was 60.4 ± 1.72%, and with Steridyl, 72.4 ± 2.10% (p < 0.01). Semen frozen in Steridyl showed good stimulation of respiration by 2.4-DNP, which indicates that oxidative phosphorylation was retained after freezing–thawing. No differences among the extenders were seen with the DNA integrity of spermatozoa. Six out of ten (60%) mares were pregnant after artificial insemination (AI) by LCCY frozen semen, and 9/12 (75%) by Steridyl frozen semen. No differences among extenders were seen in pregnancy rate. In conclusion, Steridyl was proven to be a good diluent for freezing stallion semen, even though it was developed for ruminants.

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.

Современные репродуктивные технологии позволяют бессрочно сохранять и рационально использовать генофонд исчезающих видов животных, создавать конкурентоспособные селекционные формы. В оленеводстве метод криоконсервации спермы практически не разработан. Суровые условия Арктики усложняют процесс получения спермы северных оленей (Rangifer tarandus)-животных с яркой сезонностью размножения. Гон у них проходит осенью, в течение месяца. Только в этот период можно получить сперму. После гона сперматогенез останавливается. Целью нашей работы была оценка изменения физиологических и морфологических показателей качества спермы северных оленей в процессе криоконсервации. Сперму от северных оленей получали с помощью электроэякулятора или посредством вымывания из придатков семенников. После оценки качества эякулированного и эпидидимального семени (объем, общая и прогрессивная подвижность и концентрация сперматозоидов) сперму разбавляли в среде Steridyl («Minitüb GmbH», Германия) до конечной концентрации 100 млн/мл, фасовали в соломинки по 0,25 мл и охлаждали до 5 С в течение 120 мин. После охлаждения и эквилибрации соломинки выдерживали в парах жидкого азота на поплавке при температуре 110 С в течение 12 мин и затем опускали в жидкий азот. Сперму размораживали при 37 С. Первичная оценка эякулированной спермы показала, что средний объем эякулята составлял 0,5±0,08 мл, концентрация-0,520±0,069 млрд/мл, общая подвижность-64,3±4,07 %, прогрессивная подвижность 47,9±4,24 %. Концентрация эпидидимальных спермиев была в среднем 0,260±0,078 млрд/мл, общая и прогрессивная подвижность-соответственно 43,6±8,49 и 20,8±5,25 %. После криоконсервации общая подвижность снизилась в среднем на 41,9±5,38 %, прогрессивная-на 36,8±5,29 %. Сперматозоиды в 42 % эякулятов теряли более 50 % общей подвижности. В некоторых образцах наблюдалась полная потеря подвижности после замораживания, в некоторых изменения были незначительными. Большая изменчивость по снижению подвижности клеток наблюдалась как в эякулированной, так и в эпидидимальной сперме. Клетки эпидидимальной спермы имели более высокую подвижность после замораживания по сравнению с эякулированной, однако в эпидидимальной сперме наблюдалось больше нарушений в характере движения сперматозоидов. Анализ морфологии сперматозоидов показал, что в сперме северных оленей в сравнении с другими видами животных повышена доля сперматозоидов со сморщенной или отсутствующей акросомой-в среднем 6,9±0,76 %. Значительного увеличения повреждений в области хвоста, шейки сперматозоидов и акросом после замораживания не наблюдалось. Количество клеток с повреждениями хвоста и шейки увеличилось в среднем на 4,2±1,05 %, акросом-на 2,5±0,35 %. Высокая изменчивость наблюдалась по увеличению повреждений плазматических мембран сперматозоидов (в среднем на 10,9±5,02 %). Такая вариабельность обусловлена разной криоустойчивостью спермы оленей и индивидуальной изменчивостью эякулятов. Достоверной разницы по изменениям физиологических и морфологических показателей качества спермы северных оленей после криоконсервации между эя...

Objective:The semen quality of stallions including sperm motility is an important target of selection as it has a high level of individual variability. However, effects of the molecular architecture of the genome on the mechanisms of sperm formation and their preservation after thawing have been poorly investigated. Here, we conducted a genome-wide association study (GWAS) for the sperm motility of cryopreserved semen in stallions of various breeds.Methods: Semen samples were collected from the stallions of 23 horse breeds. The following semen characteristics were examined: progressive motility (PM), progressive motility after freezing (FPM), and the difference between PM and FPM. The respective DNA samples from these stallions were genotyped using Axiom™ Equine Genotyping Array. Results:We performed a GWAS search for single nucleotide polymorphism (SNP) markers and potential genes related to motility properties of frozen-thawed semen in the stallions of various breeds.As a result of the GWAS analysis, two SNP markers, rs1141327473 and rs1149048772, were identified that were associated with preservation of the frozen-thawed stallion sperm motility, the relevant putative candidate genes being NME8, OR2AP1 and OR6C4. Potential implications of effects of these genes on sperm motility are herein discussed. Conclusion:The GWAS results enabled us to localize novel SNPs and candidate genes for sperm motility in stallions. Implications of the study for horse breeding and genetics are a better understanding of genomic regions and candidate genes underlying stallion sperm quality, and improvement in horse reproduction and breeding techniques. The identified markers and genes for sperm cryotolerance and the respective genomic regions are promising candidates for further studying the biological processes in the formation and function of the stallion reproductive system.