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Infertility is frequently associated with meiotic anomalies which can result in the production of chromosomally abnormal gametes or be concomitant with meiotic arrest. We investigated whether spermatocytes of male interspecific hybrids of the red fox (Vulpes vulpes) and the arctic fox (Alopex lagopus) presented alterations in chromosomal synapses and meiotic checkpoint signalling. Using the immunofluorescence technique with SP1 and SP3 proteins, bivalent structures and their deviations (multivalents, univalents and not fully conjugated bivalents) were analyzed on meiotic preparations. This technique allowed the localization of frequent foci of phosphorylated histones H2AHγ (Ser 139) to the meiotic block in late pachytene. These results indicate a disruption of meiotic division in male fox hybrids, which leads to a high percentage of apoptotic cells in the gonads of these animals and, consequently, sterility. • Synaptonemal complex in spermatocytes of cross-species. • Analysis of asynapsis in spermatocytes of hybrids. • Crosspecies silver fox and blue fox. • arrest of pachyten I by sustained H2AXγ phosphorylation. [ABSTRACT FROM AUTHOR]

Contents The aim of this study was a cytogenetic analysis of stallions semen to find sex chromosome aberrations and to determine if there was an association between stallion's age and aberration frequency for the sex chromosomes. Sperm samples were collected from 22 stallions of various age from 3 to 23 years. Multicolour FISH was performed on each sample, using probes for the sex chromosomes and EGFR gene, localized on 4p12 in domestic horse. A total of 26199 sperm cells were analysed (from 1 070 to 1 532 per animal). Among the analysed cells, there were 50.318% with X chromosome, 48.543% with Y chromosome and 1.139% with aberrant chromosomes. The frequency of aberrations was: sex chromosomes nullisomy (0.466%), XY aneuploidy (0.454%), XX disomy (0.146%), YY disomy (0.041%), diploidy (0.024%) and trisomy XXY (0.008%). Additionally there was a correlation between the age of an animal and the frequency of sex chromosome aberration and a significant positive correlation between age and disomy of XY, XX, YY, trisomy of XXY, autosomal disomy was seen. A Correlation between the age of a stallion and the level of nullisomy was negative. The present study demonstrated that FISH technique is a powerful method to identify sex chromosome aberrations in equine spermatozoa and might be very helpful for a breeder during a selection for the best stallion. [ABSTRACT FROM AUTHOR]

This study describes, for the first time, the relationship between morphology and ploidy in domestic cat embryos. Blastocyst morphology and quality were assessed using time-lapse recordings, while ploidy was analyzed using fluorescence in situ hybridization. Out of 54 blastocysts, clear fluorescence signals for all the molecular probes used were observed in 24 (44.4%) blastocysts, while in another 14 (25.9%) blastocysts, fluorescence signals only allowed for sex assessment. No clear signals were observed in the remaining 16 blastocysts (29.7%). Of the 24 blastocysts with clear signals, normal ploidy was detected in 10 (41.4%), 7 (29.2%) were diagnosed as haploid, and the remaining 7 blastocysts (29.2%) were mosaics. Additionally, results showed the distribution of diploid, haploid, and mosaic blastocysts in relation to the occurrence of morphological disorders and to embryo quality. The presence of abnormal embryo morphology and karyotype disorders may affect further development and the pregnancy rate. Due to the comparable proportion of good and poor quality blastocysts with disturbed ploidy, it is important to implement new methods of embryo assessment, especially when techniques used in humans, such as pronuclear observation, cannot be used. • We analyzed cat embryo morphology, blastocyst quality and ploidy. • Morphologically abnormal embryos may show normal ploidy, haploidy and mosaicism. • Abnormal embryo morphology and karyotype disorders may affect developmental potential of cat embryos. [ABSTRACT FROM AUTHOR]

Contents Chromosomal abnormalities are one of the main causes of genetic infertility in horses. Currently, their detection rate is rising due to the use of new diagnostic tools employing molecular markers linked to the sex chromosome pair. Despite genetic similarities, there are no previous reports of sterility associated with chromosomal abnormalities in the domestic donkey ( Equus asinus). Hereby, we determined the presence of a chromosomal mosaicism in a female donkey with reproductive problems using molecular methodologies developed for horses. A two-and-a-half-year-old jenny characterized by morphological abnormalities of the reproductive tract was cytogenetically analysed using conventional and fluorescent techniques and a group of microsatellite markers (short tandem repeat, STR). At the same time, five ultrasound measures of the reproductive tract were taken and compared with eight contemporary jennies of the same breed. After slaughter, morphological examinations showed that the case study had a blind vaginal vestibule defining an empty pouch that covered the entrance of the cervical os. Histopathological studies demonstrated that this abnormal structure was compatible with a remnant hymen. Molecular markers, STR and fluorescent in situ hybridization determinations revealed that the animal was a 62, XX/61,X mosaic and, therefore, the first case of chromosomal abnormalities in the sex pair reported in donkeys. [ABSTRACT FROM AUTHOR]

In the current study, transcriptome profiles of mare endometrium, classified into categories I, IIA, and IIB according to Kenney and Doig, were compared using RNA sequencing, analyzed, and functionally annotated using in silico analysis. In the mild stage (IIA) of endometrosis compared to category I endometrium, differentially expressed genes (DEGs) were annotated to inflammation, abnormal metabolism, wound healing, and quantity of connective tissue. In the moderate stage (IIB) of endometrosis compared to category I endometrium, DEGs were annotated to inflammation, fibrosis, cellular homeostasis, mitochondrial dysfunction, and pregnancy disorders. Ingenuity pathway analysis (IPA) identified cytokines such as transforming growth factor (TGF)-β1, interleukin (IL)-4, IL-13, and IL-17 as upstream regulators of DEGs associated with cellular homeostasis, metabolism, and fibrosis signaling pathways. In vitro studies showed the effect of these cytokines on DEGs such as ADAMTS1, -4, -5, -9, and HK2 in endometrial fibroblasts at different stages of endometrosis. The effect of cytokines on ADAMTS members' gene transcription in fibroblasts differs according to the severity of endometrosis. The identified transcriptomic changes associated with endometrosis suggest that inflammation and metabolic changes are features of mild and moderate stages of endometrosis. The changes of ADAMTS-1, -4, -5, -9, in fibrotic endometrium as well as in endometrial fibroblast in response to TGF-β1, IL-4, IL-13, and IL-17 suggest the important role of these factors in the development of endometrosis. [ABSTRACT FROM AUTHOR]

Abstract: Sex chromosome aberrations are known to cause congenital abnormalities and unexplained infertility in horses. Most of these anomalies remain undiagnosed because of the complexity of the horse karyotype and the lack of specialized laboratories that can perform such diagnoses. On the other hand, the utilization of microsatellite markers is a technique widely spread in horse breeding, mostly because of their usage in parentage tests. We studied the usage of a novel combination of diagnostic approaches in the evaluation of a very uncommon case of chromosomal abnormalities in a Spanish purebred colt, primarily detected using a commercial panel of short tandem repeat (STR) makers. Based on these results, we performed a full cytogenetic analysis using conventional and fluorescent in situ hybridization techniques with individual Equus caballus chromosome X and Equus caballus chromosome Y painting probes. We also tested the presence of two genes associated with the sexual development in horses and an extra novel panel of eight microsatellite markers specifically located in the sex chromosome pair. This is the first case report of a leukocyte chimerism between chromosomally normal (64,XY) and abnormal (63,X0) cell lines in horses. Our results indicate that the use of the short tandem repeat markers as a screening technique and as a confirmation utilizing cytogenetic techniques can be used as a very interesting, easy, and nonexpensive diagnostic approach to detect chromosomal abnormalities in the domestic horse. [Copyright &y& Elsevier]

Chromosomal abnormalities associated to sex chromosomes are reported as a problem more common than believed to be in horses. Most of them remain undiagnosed due to the complexity of the horse karyotype and the lack of interest of breeders and veterinarians in this type of diagnosis. Approximately 10 years ago, the Spanish Purebred Breeders Association implemented a DNA paternity test to evaluate the pedigree of every newborn foal. All candidates who showed abnormal or uncertain results are routinely submitted to cytogenetical analysis to evaluate the presence of chromosomal abnormalities. We studied the case of a foal showing 3 and even 4 different alleles in several loci in the short tandem repeat (STR) -based DNA parentage test. To confirm these results, a filiation test was repeated using follicular hair DNA showing normal results. A complete set of conventional and molecular cytogenetic analysis was performed to determine their chromosomal complements. C-banding and FISH had shown that the foal presents a sex chimerism 64,XX/64,XY with a cellular percentage of approximately 70/30, diagnosed in blood samples. The use of a diagnostic approach combining routine parentage QF-PCR-based STR screening tested with classical or molecular cytogenetic analysis could be a powerful tool that allows early detection of foals that will have a poor or even no reproductive performance due to chromosomal abnormalities, saving time, efforts and breeders' resources. Copyright © 2013 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Abstract: Nucleolar organizing regions (NORs) containing rDNA gene clusters have been assigned to the equine autosomes ECA1, ECA28, and ECA31. Active NORs (Ag-NORs) are associated with argyrophilic proteins, which allow them to be readily identified using silver staining techniques. Fluorescence in situ hybridization (FISH) for rDNA can also be used to visualize all NOR clusters in the nucleus, regardless of whether they are active or inactive. The present study analyzed the distribution and behavior of equine Ag–NOR and NOR clusters in horse spermatozoa and during male meiosis by FISH and silver staining. The NOR foci were observed to be variable in number, size, and shape, but were usually located centrally and appeared as one or two nucleolus-like structures in the spermatozoa head. Three distinctive FISH signals identified the NOR-bearing chromosome pairs during the synaptic cell stage of meiosis I. At diakinesis/metaphase I, as well as different stages of meiosis II, FISH signals clearly depicted the NOR-bearing sister chromatids. The synaptonemal complexes of primary spermatocytes consistently showed three rDNA foci following FISH, but variably demonstrated two or three Ag–NOR bodies following silver staining. We propose rDNA loss and gain during unequal crossing-over events could be both a direct and indirect cause of variation in equine NOR foci. Additionally, our cytogenetic analysis did not confirm the presence of a fourth pair of NORs-bearing chromosomes in the horse, which is contrary to previously mitotic published data. [Copyright &y& Elsevier]

Summary: Chromosomal abnormalities are a common cause of infertility in horses. However, they are difficult to detect using automated methods. Here, we propose a simple methodology based on single nucleotide polymorphism (SNP)‐array data that allows us to detect the main chromosomal abnormalities in horses in a single procedure. As proof of concept, we were able to detect chromosomal abnormalities in 33 out of 268 individuals, including monosomies, chimerisms, and male and female sex‐reversions, by analyzing the raw signal intensity produced by an SNP array‐based genotyping platform. We also demonstrated that the procedure is not affected by the SNP density of the array employed or by the inbreeding level of the individuals. Finally, the methodology proposed in this study could be performed in an open bioinformatic environment, thus permitting its integration as a flexible screening tool in diagnostic laboratories and genomic breeding programs. [ABSTRACT FROM AUTHOR]

We investigated the activity of chondrogenic markers and variation of methylation patterns in equine cartilaginous cells cultivated in monolayer. The transcriptional and epigenetic effect of the long-term culture of chondrocytes has been evaluated using several passages of chondrocyte cell-lines derived from equine articular cartilage. Using 3 genes as endogenous control we tested the expression of 7 genes important for different stages of chondrocyte differentiation and maturation. CpG islands in RUNX3 locus were inspected for the evaluation of differential methylation state of passaged cell-lines. The general decline of transcript abundance of marker loci was detected in passage 11 which is the sign of dedifferentiation of cultivated chondrocytes in prolonged monolayer culture. Passages 13 and 14 were characterized by the upregulation of a number of genes, possibly due to the heterogeneity of developed cell lines at this stage of the culture. Instead, gradual increase of methylation percent at particular CpG sites of RUNX3 locus was associated with the growing number of passage. This finding led us to the conclusion that epigenetic alterations better describe the stage of cultivated chondrocytes. • Transcriptional decline in prolonged monolayer culture affects markers of early and late chondrocyte differentiation. • Gene upregulation affects certain chondrogenic markers in latter passages of the monolayer culture. • CpG hypermethylation of RUNX3 locus coincides with the growing number of passages of chondrocytes in monolayer. [ABSTRACT FROM AUTHOR]

Horses are one of the longest-living species of farm animals. Advanced age is often associated with a decrease in body condition, dysfunction of immune system, and late-onset disorders. Due to this, the search for new solutions in the prevention and treatment of pathological conditions of the advanced age of horses is desirable. That is why the identification of aging-related changes in the horse genome is interesting in this respect. In the recent years, the research on aging includes studies of age-related epigenetic effects observed on the DNA methylation level. We applied reduced representation bisulfite sequencing (RRBS) to uncover a range of age DMR sites in genomes of blood leukocytes derived from juvenile and aged horses of native Hucul breed. Genes colocated with age-related differentially methylated regions (age DMRs) are the members of pathways involved in cellular signal transduction, immune response, neurogenesis, differentiation, development, and cancer progression. A positive correlation was found between methylation states and gene expression in particular loci from our data set. Some of described age DMR-linked genes were also reported elsewhere. Obtained results contribute to the knowledge about the molecular basis of aging of equine blood cells. [ABSTRACT FROM AUTHOR]

Abstract The impact of exercises on young developing organisms is still of interest to researchers. Similarly like Thoroughbreds, Arabian horses competing at the race track. The high percent of lameness and loss of days in training are often the result of weakness in the condition of the musculoskeletal system. The objective of the presented study was to identify by RNA-Seq method, the possible skeletal system originating transcriptomic profile in peripheral blood of Arabian horses undergoing race training. Obtained results showed that one of the most significantly deregulated pathway involved in bone homeostasis was those involved in osteoclast differentiation. Among the significantly expressed molecules, we recognized twelve genes potentially involved in the metabolism of the skeletal system: BGLAP , CTSK , TYROBP , PDLIM7 , SLC9B2 , TWSG1 , NOTCH2 , IL6ST , VAV3 , NFATc1 , CLEC5A , TXLNG. The panel of identified genes should be evaluated as candidate biomarkers for bone homeostasis indicators of Arabians performing on race tracks to assess bone remodelling states during training for race track competitions. Highlights • Racing training in horses has impact on whole blood transcriptome. • Whole blood transcriptome reflects changes in bone state. • The osteoclast differentiation pathway was significantly deregulated. • We identified genes involved in bone metabolism for further evaluation as biomarkers. [ABSTRACT FROM AUTHOR]

Objective: In this study, for the first time we report the genome-wide DNA methylation profile of skin tumour in horses and describe differentially methylated genomic regions (DMRs) with respect to healthy skin. Materials & Methods: The comparative analysis of DNA methylation patterns detected using Reduced Representation Bisulfite Sequencing (RRBS) technique, allowed identification of 136 regions showing differential methylation between sarcoid and normal skin tissue. Results: Most of the identified DMRs were short fragments, less than 1 kb in size, located in the intergenic regions. Among identified DMRs there were also regions located within genes directly or indirectly related with oncogenesis. We additionally validated 9 CpG sites showing hypomethylation and 9 CpG sites that were hypermethylated in lesional sample, confirming the identified changes in the DNA methylation. Conclusion: Knowledge on the changes taking place in the process of DNA methylation may provide a basis for the development of new alternative diagnostic or therapeutic approaches to equine sarcoids. [ABSTRACT FROM AUTHOR]

Equine sarcoids are the most commonly detected skin tumours in Equidae. In the present research, a comparative transcriptomic analysis was performed which aimed at looking inside a tumour biology and identification of the expression profile as a potential source of cancer specific genes useful as biomarkers. We have used Horse Gene Expression Microarray data from matched equine sarcoids and tumour-distant skin samples. In total, 901 significantly differentially expressed genes ( DEGs) between lesional and healthy skin samples have been identified (fold change ≥ 2; P < 0.05). The large subset of DEGs, with decreased expression, was associated with a suppression of malignant transformation, whereas several overexpressed genes were involved in the processes associated with growth and progression of a tumour or immune system activity. Our results, as a first to date, showed comprehensive transcriptome analysis of skin tumour in horses and pinpointed significant pathways and genes related with oncogenesis processes. [ABSTRACT FROM AUTHOR]

The aim of this study was to characterize in detail linkage disequilibrium (LD) decay and haplotype block structures in genomes of commercial Limousin (LM, n=201) and Simmental (SM, n=106) cattle and refer this data to the characteristics found in native unselected Polish Red (PR, n=299) cattle. A panel of 40,158 (LM), 40,117 (SM) and 42,118 (PR) high quality SNP (single nucleotide polymorphism) markers from Illumina BovineSNP50 v2 BeadChip were utilized for the LD analysis based on the pairwise r 2 statistic of SNPs at a distance up to 5 Mb. The lower average r 2 of 0.27 was observed in RP cattle at a short distance up to 25 kb when compared to two other studied breeds (0.3 in both LM and SM). The minimum average values of r 2 (0.01) were observed at the distances of 1500–3000 kb in LM and 3000–5000 kb in SM. In PR cattle LD persisted similarly as in SM cattle and the minimum average values of r 2 (0.02) were observed at a distance of 3000–5000 kb. The effects of minor allelic frequency threshold on the extent of LD was also evaluated by applying three different minimum MAF levels (0.05, 0.1 and 0.2). A total of 828 (LM), 667 (SM) and 761 (PR) haplotype block structures spanning 89,781 kb (LM), 72,582 kb (SM) and 70,647 kb (PR) of the genome were detected. In total, 7.92% (LM), 6.43% (SM) and 6.52% (PR) of all SNPs formed blocks with a range of 2–15 SNPs per block in LM and SM breeds and 2–16 SNPs per block in PR cattle. Mean block lengths were slightly lower in RP cattle than in two other breeds and were estimated as 108.4±99 kb, 108.8±97 kb and 92.8±87 for LM, SM and PR respectively. For all studied breeds chromosome 1 showed the longest haplotype block structures in the genome, having 66 blocks spanning 8120 kb in LM, 57 blocks spanning 6754 kb in SM and 59 blocks spanning 5920 kb in PR. The results showed slightly faster LD decay in Polish Red cattle (especially at short distances) than in two other breeds and shorter haplotype block structures which may result from population demographic history, overall genetic diversity as well as extensive breeding applied in this breed. [ABSTRACT FROM AUTHOR]

The study is aimed at identifying selection footprints within the genome of Limousin cattle. With the use of Extended Haplotype Homozygosity test, supplemented with correction for variation in recombination rates across the genome, we created map of selection footprints and detected 173 significant (p < 0.01) core haplotypes being potentially under positive selection. Within these regions, a number of candidate genes associated inter alia with skeletal muscle growth ( GDF15, BMP7, BMP4 and TGFB3) or postmortem proteolysis and meat maturation ( CAPN1 and CAPN5) were annotated. Noticeable clusters of selection footprints were detected on chromosomes 1, 4, 8 and 14, which are known to carry several quantitative trait loci for growth traits and meat quality. The study provides information about the genes and metabolic pathways potentially modified under the influence of directional selection, aimed at improving beef production characteristics in Limousin cattle. [ABSTRACT FROM AUTHOR]

• Differences in level of LD were found between breeds representing different horse types. • Breeds representing the same horse type showed similar trends of historical N e. • Longer haplotype block structure was found in Arabian breed than in native horse breeds. • Population history and breeding schemes may influence patterns of LD. This study was conducted to investigate linkage disequilibrium (LD), historical effective population size (N e) and haplotype block structures in genomes of highly selected Polish Arabian (AR; n=124) horses and conserved horse breeds derived from Polish native breeds, in particular, Malopolski (MLP; n=56), Sokolski (SOK; n=107), Sztumski (SZTUM; n=69), Hucul (HC; n=116) and Polish Konik (KN; n=99) horses, based on single nucleotide polymorphisms (SNPs) from the Neogen Equine Community BeadChip assay (Illumina, San Diego, CA). LD analysis was performed based on the pairwise r2 statistic of SNPs at a distance up to 5Mb. At a short distance, up to 100kb, the lower average r2 was observed in SOK (0.09) and SZTUM (0.10) breeds, representing draft horse type, and the highest average r2 was observed in AR (0.19) and MLP (0.17) breeds, representing light horse type. At a distance of 4-5Mb, the average values of r2 were 0.03 for HC and KN, 0.02 for AR and MLP and 0.01 for SOK and SZTUM. About 100 generations ago, effective population size (N e) ranged from 518 (SOK) to 310 (AR) and five generations ago from 85 (SOK) to 40 (HC). A total of 6,162 (AR), 5,792 (MLP), 5,744 (HC), 5,429 (KN), 5,542 (SOK) and 4,848 (SZTUM) haplotype block structures, spanning 656.2Mb (AR), 439.4Mb (MLP), 381.2Mb (HC), 344.3 (KN), 206.7 (SOK) and 171.0Mb (SZTUM) of the genome were detected. Mean block lengths were estimated as 106.5±138kb, 75.9±120kb, 66.4±109kb, 63.4±109kb, 37.3±80kb and 35.3±81kb for AR, MLP, HC, KN, SOK and SZTUM, respectively. A visible similarity in the decay of LD across the genome, as well as in trends of historical N e, were found for breeds representing the same horse type. Differences in the span of haplotype block structures were found between AR and Polish native horse breeds, which may reflect different demographic population histories and differences in intensity of selection. [ABSTRACT FROM AUTHOR]

Background: Apoptosis plays an important role in the regulation of healthy tissue growth and development as well as in controlling the maintenance of homeostasis in exercising muscles. During an intensive physical effort, the regulation of cell death by apoptosis results in the replacement of unaccustomed muscle cells by new cells that are better suited to exercise. The aim of this study was to determine the expression of two genes (SH3FR1 and SH3RF2) that control apoptosis in muscle tissues during training periods characterized by different intensities. The gene expression levels were estimated using real-time PCR method in skeletal muscle biopsies collected from 15 Arabian horses (untrained, after an intense gallop phase, and at the end of the racing season). An association study was performed on 250 Arabian horses to assess the effect of the SH3RF2:c.796 T > C (p.Ser266Pro) variant on race performance traits in flat gallop-racing. Results: A gene expression analysis confirmed a significant decrease (p < 0.01) in the anti-apoptotic SH3RF2 (POSHER) gene during training periods that differed in intensity. The highest SH3RF2 expression level was detected in the muscles of untrained horses, whereas the lowest expression was identified at the end of the racing season in horses that were fully adapted to the exercise. A non-significant decrease in SH3RF1 gene expression following the training periods was observed. Moreover, a serine substitution by proline at amino acid position 266 (CC genotype) was negatively associated with the probability of winning races, the number of races in which a horse occurred and the financial value of the prizes. Horses with the TT genotype achieved the highest financial benefits, both for total winnings and for winnings per race in which the horses participated. Conclusions: The present study showed the supposed regulation mechanism of exercise-induced apoptosis in horses at the molecular level. The identified SH3RF2: c.796 T > C missense variant was associated with selected racing performance traits, which is important information during the evaluation of horses’ exercise predisposition. The association results and frequencies of the CT and TT genotypes suggest the possibility of using SH3RF2 variant in selection to improve the racing performance of Arabian horses. [ABSTRACT FROM AUTHOR]

The article presents a study that investigates the implications of melanocortin 1 receptor (MC1R) on the color coat variation of nutrias in Krakow, Poland. Researchers collected blood from the animals, on the farms controlled by the National Animal Husbandry Center, which have nutria color coat variants and examined their blood with the use of DNA purification kit. Moreover, they discovered the presence of dark color patterns on coat colors.