First case of sterility associated with sex chromosomal abnormalities in a jenny

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.     

Y‐chromosomal variation of local goat breeds of Turkey close to the domestication centre

Genetic variations in chromosome Y are enabling researchers to identify paternal lineages, which are informative for introgressions and migrations. In this study, the male‐specific region markers, sex‐determining region‐Y (SRY), amelogenin (AMELY) and zinc finger (ZFY) were analysed in seven Turkish native goat breeds, Angora, Kilis, Hair, Honaml?, Norduz, Gürcü and Abaza. A SNP in the ZFY gene defined a new haplotype Y2C. All domestic haplogroups originate from Capra aegagrus, while the finding of Y1A, Y1B, Y2A and Y2C in 32, 4, 126 and 2 Turkish domestic goats, respectively, appears to indicate a predomestic origin of the major haplotypes. The occurrence of four haplotypes in the Hair goat and, in contrast, a frequency of 96% of Y1A in the Kilis breed illustrate that Y‐chromosomal variants have a more breed‐dependent distribution than mitochondrial or autosomal DNA. This probably reflects male founder effects, but a role in adaptation cannot be excluded.     

Coat colour genes in diversity studies

In this paper we describe the use of polymorphic genes affecting coat colour as a tool in diversity studies of domestic animals. Although phenotypic data has been the main criteria for establishing different breeds, calculation of genetic distances between breeds is normally performed using noncoding microsatellite markers. As anticipated, MC1‐R (melanocyte stimulating hormone receptor) allele frequencies vary greatly between cattle breeds expressing different coat colours. In multicoloured breeds, like Icelandic cattle, a high frequency of the E⁺ allele appears to be essential for colour variation. Whereas black breeds have a high frequency of the dominant acting allele E^D, entirely red breeds have no E^D. Animals being homozygous for the defective allele e occurred frequently in some cattle breeds, indicating that the MC1‐R does not have crucial impact on animal physiology other than coat colour. The E⁺ and e alleles were observed in the closely related river buffalo as well. None of the breeds included in this study express the roan phenotype. Consequently, they were monomorphic at the MGF locus. As for the MC1‐R locus, a correlation to colour pattern was observed for two c‐kit alleles as well, confirming that selection of specific phenotypes strongly affect the allelic variation of underlying loci. Information on genes affecting the phenotype is therefore well suited for describing different breeds of livestock and, consequently, a practical tool in breed conservation.     

Detecting selection signatures on the X chromosome of the Chinese Debao pony

The X chromosome shows a special interaction between demographic factors and genetic variation, and the analysis of X‐linked genomic variation can therefore provide insights into the unique effects of demography and selection on the horse genome that cannot be readily detected by autosomal markers. Debao (DB) ponies have experienced intense selective pressure for the development of their small stature (<106 cm at adult height). To identify selective sweeps on the X chromosome of the DB pony, we performed a genome‐wide scan of three Chinese horse breeds using an Equine SNP70 BeadChip. Using Yili and Mongolian horses (>134 cm at adult height) as reference groups, both F_ST and XP‐EHH revealed that five regions on the X chromosome were under strong selection, resulting in 95 overlapping genes. Seven of these genes, SMS, PHEX, ACSL4, CHRDL1, CACNA1F, DKC1 and CDKL5, are involved in bone development, growth hormone secretion and fat deposition. The region showing the strongest selection pressure was located at the position of 86.6–87.5 Mb. The subsequent genome‐wide association analysis of the adult height of three Chinese horse breeds detected the two most significant SNPs in the same region, and these two SNPs overlapped with the gene CHRDL1. As a member of the bone morphogenetic protein (BMP) superfamily, CHRDL1 antagonizes the function of BMP4 and plays an important role in embryonic bone formation and cartilage generation. Our results provide new insights into the X‐linked selection in Chinese Debao pony.     

Cytogenetic mapping of 11 functional genes to chicken chromosomes by fluorescence in situ hybridization

With chicken bacterial artificial chromosome (BAC) DNA as probes, 11 non‐assigned functional genes were localized to chicken chromosomes 1 or 2 by fluorescence in situ hybridization (FISH). The 11 genes and their chromosomal positions are as follows: ALVEB5, 1p26‐24; ACO2, 1p16‐14; HSP108, 1p14‐13; CD4, 1q11; HSD3B; 1q11; SOD1, 1q14‐21; LAMP1, 1q24‐31; P2Y5, 1q35‐36; EN2, 2p31‐24; NPY, 2p14‐13 and CA2, 2q31‐32. Metaphase chromosome spreads used for hybridization were prepared from embryonic chicken fibroblast cultures. The gene position was identified according to the international standardized G‐banded karyotype of chicken by measuring the relative fractional length from the telomere of the p‐arm to the hybridization signal (FLpter). The 11 genes mapped newly will enrich the cytogenetic map and serve as additional anchor markers for integrating the cytogenetic map with the genetic map of chicken.     

Genetic mapping of dominant white (W), a homozygous lethal condition in the horse (Equus caballus)

Dominant white coat colour (W) is a depigmentation syndrome, known in miscellaneous species. When homozygous in the horse (similar in mice), the mutation responsible for the white phenotype is lethal in a very early stage of gestation. It seems, that the action of the dominant white allele is not always fully penetrant, resulting occasionally in spotted look alike offspring. These horses resemble a coat colour pattern known as sabino spotting. So far, it is not known whether dominant white (W) and sabino spotting (S) share a common genetic background. In this study, a pedigree consisting of 87 horses segregating for dominant white (W) was used to genetically localize the horse (W)‐locus. Microsatellite ASB23 was found linked to (W), which allowed us to map dominant white to a region on horse chromosome 3q22. Tyrosine kinase receptor (KIT) was previously mapped to this same chromosome region (3q21–22). KIT and its ligand (KITLG) are responsible for the normal function of melanogenesis, haematopoiesis and gametogenesis. So far, sequence analysis of different KIT gene fragments did not lead to new polymorphisms, except for a SNP detected in KIT intron 3 (KITSNPIn3). Additional microsatellites from ECA3q (TKY353 and LEX7), together with KITSNPIn3 allowed us to state more precisely the (W)‐mutation. The positional results and comparative functional data strongly suggest that KIT encodes for the horse (W)‐locus.     

Morphological study of tracheal shape in donkeys with and without tracheal obstruction

Reason for performing study: There is limited information on the gross tracheal morphology of donkeys with or without tracheal abnormalities. Objectives: To: 1) examine the morphology of tracheas of donkeys with and without clinical and/or post mortem evidence of tracheal obstruction; 2) record the cross‐sectional dimensions and shapes of tracheal rings at fixed sites; and 3) document prevalence, sites and characteristics of detected tracheal abnormalities. Methods: The tracheas of 75, predominantly aged (median age 30 years, range 7–48 years) donkeys that died or were subjected to euthanasia on humane grounds were examined. Five had severe dyspnoea due to tracheal obstruction (with intercurrent lung disease in 3), while 7 had post mortem evidence of severe tracheal airway obstruction. Every 5th tracheal ring was dissected free and the inner and outer vertical and transverse dimensions and cross sectional areas were measured. Each dissected ring was photographed and the shape of the trachea was classified as normal or, in one of 6 abnormal grades, according to the type and degree of structural abnormality present. Results: The tracheas had a mean of 43 (range 34–50) tracheal rings that tended to be more oval in shape in the distal cervical region. Only 31.2% of rings examined had a circular to oval shape. Dorso‐ventral flattening was present in 0.9% of tracheal rings, dorsal ligament separation in 24.4%, slight cartilage deformity in 26.0%, moderate cartilage deformity in 10.4%, marked cartilage deformity in 1.9% and miscellaneous other abnormalities in 4.9% of tracheal rings. The 12 donkeys with ante or post mortem evidence of tracheal obstruction had significantly increased tracheal abnormality grade in comparison to the remaining donkeys. Conclusions and potential relevance: Structural tracheal abnormalities are present in most old donkeys, but generally do not cause clinical problems in these sedentary animals unless intercurrent pulmonary disease is present.     

湘西黄牛Y-SNPs遗传多样性研究

[目的]通过Y-SNP分子标记方法研究湘西黄牛的遗传多样性、群体遗传结构及父系起源。[方法]采用PCR扩增、测序与生物信息学方法,对24头湘西黄牛的2个Y-SNPs(UTY-19和ZFY-10)标记进行多态性分析。[结果]结果表明,湘西黄牛有Y1和Y3两种单倍型组,频率分别为12.5%和87.5%,表明湘西黄牛可能有普通牛和瘤牛2个父系起源。湘西黄牛的Y-SNP遗传多样度为0.2283±0.0978,表明湘西黄牛具有较低的父系遗传多样性,品种纯度较高。[结论]湘西黄牛的父系起源为瘤牛Y3单倍型组,其Y1单倍型组为国外肉牛杂交所致。     

On ancestors of dog breeds with focus on Weimaraner hunting dogs

Paternally inherited Y chromosomal markers and maternally inherited mitochondrial (mt) DNA sequences were investigated in 27 dog breeds (Canis familiaris), of which the Weimaraner hunting dog was studied in greater detail. Altogether, nine potentially polymorphic markers of the Y chromosome were examined as well as parts of the canine mt genome (1947 base pairs) in 111 male dogs and four wolves for comparison. Twenty Y chromosomal and fifty-nine mitochondrial DNA (mtDNA) haplotypes were identified in the canine breeds and wolves. In 34 Weimaraners, four distinct Y chromosomal haplotypes were observed as well as three mtDNA types thus reflecting at least four male and three female ancestors for the current population in Germany. Tracing patri- and matrilineages, several entries in the Weimaraner stud book cannot be reconciled with the male-only, Y chromosomal neither the female-only, mt inheritance patterns, respectively. The investigated breeds represent 9 of 10 groups defined by the Fédération Cynologique Internationale (FCI). The level of Y chromosomal and especially mtDNA diversity was immense considering the relatively small number of individuals investigated per breed. Unique haplotypes were found only in a few breeds and the wolf. Other haplotypes were shared among several breeds, also across different FCI groups, suggesting that these canine breeds had common male and female ancestors.     

Application of bovine microsatellite markers on Saola (Pseudoryx nghetinhensis)

The aim of the present study was to assess the applicability of bovine microsatellite markers on Saola (Pseudoryx nghetinhensis). A total of 127 microsatellite markers were tested on a male and a young female Saola. An efficient amplification was observed for 123 markers (96.8%), 73 markers (59.3%) were polymorphic. Four loci (BM2304, BMS1928, BMS779 and ILSTS006) on cattle chromosomes 1, 4, 7 and 8, respectively, failed to amplify in Saola. Two cattle Y‐chromosome‐specific microsatellite markers (INRA126 and BM861) were successfully amplified from both sexes in Saola. However, two additional markers (INRA124 and INRA189) on Y‐chromosome failed to amplify in the female animal. These results show that most of the bovine microsatellite markers are applicable in Saola and therefore they can be used to study the phylogenetic relationships and the genetic diversity of the Saola population.     

中国四个地方驴品种mtDNA D-Loop部分序列分析与系统进化研究

对中国4个家驴品种95个个体的mtDNAD—loop部分序列进行测序。用clustalX软件进行同源序列比对。Dnasp4-10软件用于遗传多样性分析，计算单倍型多样度、核苷酸多样度、平均核苷酸差异。MEGA3．1软件采用邻接法构建系统进化树并进行系统发生分析。以欧洲驴线粒体基因组为对照（Genbank登录号为X97337），中国家驴4个品种95个个体385bp序列共检测到核苷酸多态位点33个，其中31个位点为转换，1个位点为颠换，1个位点有插入现象。95个序列由31个单倍型组成，单倍型比例为32．6％。德州驴单倍型比例最高为100％，凉州驴单倍型比例最低为42．42％。31个单倍型与引用Genbank已登录的13条序列构建系统发育树，发现31个单倍型聚为两支，说明中国家驴可能有两个母系起源。以欧洲野驴线粒体D—loop为对照（登录号为AF403063、AF403063、AF403065），揭示本研究涉及的中国家驴品种与非洲野驴的进化亲缘关系较亚洲野驴近，并且从分子水平证明中国家驴可能起源于非洲野驴中的索马里野驴分支。     

The association analysis between Cytb polymorphism and growth traits in three Chinese donkey breeds

The aim of this study was to investigate the effects of haplotypes in Cytb (Cytochrome b) gene on growth traits in three Chinese donkey breeds, Yunnan donkey (YN), Dezhou donkey (DZ) and Liangzhou donkey (LZ). The samples were genotyped for the Cytb gene by single strand conformation polymorphism (SSCP), and association analysis was conducted with growth traits. The results show that in most cases the effects of the gene on growth traits were not significant and that for the three different donkey breeds, the effects of the gene are different. For YN donkeys, there was a significant effect of the gene on the rump width (P < 0.05) and the effect of genotype AA was larger than that of BB genotype (1.6 cm and 5.4% of the trait mean in YN donkeys, P < 0.05). For DZ donkeys, the effect of gene on body height was significant (P < 0.05). The results also verify that the SSCPs are caused by two cytosine to thymine and one guanine to adenine transitions, corresponding to amino acid substitutions of leucine for proline (L ↔ P) and isoleucine for valine (I ↔ V).     

A Clinical Survey Evaluating the Prevalence of Incisor Disorders in Zamorano-Leonés and Mirandês Donkeys (Equus asinus)

Recent clinical and post-mortem studies documented a high prevalence of dental disorders in donkeys, but less information appears to be available specifically about incisor disorders in donkeys. A study to investigate the prevalence of oral and dental disorders affecting incisor teeth was performed, in two endangered breeds of donkeys: the Mirandês Donkey and the Zamorano-Leonés Donkey, through a prospective cross-sectional study in 800 donkeys, divided in 7 age groups (ranging 0-34 years), in 86 villages inside their geographic area of distribution, thinking on welfare and genetic preservation issues. The 74% of donkeys suffer from incisors disorders, ranging from 56.8% in the youngest group to 90.3% in group 7. Craniofacial abnormalities (49.25%), abnormalities in the occlusal surface (21.63%), fractures (17%), periodontal disease (16.13%) and diastemata (14.38%) were the main disorders recorded. Incisors disorders are significant, presenting at a much higher prevalence when compared to other studies involving the incisor teeth of equids, affecting all ages but particularly in older animals. This study provide essential information in dentistry applied to donkeys but also highlighted the importance of regular dental care in endangered breeds, improving their welfare and preserving a unique genetic heritage.     

Allele frequencies of gene polymorphisms related to economic traits in Bos taurus and Bos indicus cattle breeds

Allele frequencies of 10 representative polymorphisms for beef and milk traits were investigated for a total of 240 animals from Bos taurus and Bos indicus breeds, including two Japanese groups (Japanese Black and Japanese Brown), two East Asian groups (Korean and Mongolian), three European groups (Holstein, Angus and Hereford) and a Bos indicus group in South Asia (Myanmar, Laos and Cambodia). The Japanese Black revealed unique genetic construction in GH, FASN and SREBP‐1 and the other Asian populations show intermediate frequencies between European and Japanese populations. The Bos indicus group showed low favorable allele frequencies in most of the genes. The study showed the variability and distribution of 10 genes affecting economic traits among world representative cattle breeds. The genetic information would contribute to elucidating the genetic background for worldwide cattle breeds and the possibility of improvement using the markers.     

Evaluation of genetic diversity in Japanese Brown cattle population by pedigree analysis

The Japanese Brown is the second most common domestic beef breed in Japan. However, nowadays this breed is facing reduction in numbers because of pressure from a profitable domestic breed. This breed is uniformly characterized by its brown coat colour, but is comprised of two isolated sub‐breeds, Kumamoto and Kouchi, each possessing a different gene pool. Pedigree analyses were carried out for the two sub‐breeds using the pedigree records of animals born from 1970 to 2000. The effective population size has been found to be consistently reducing during the last three decades in both sub‐breeds. The current effective sizes were estimated to be 25.5 and 6.0 for the Kumamoto and Kouchi sub‐breeds, respectively. The estimate of the effective number of founders (N_ef) in the Kumamoto sub‐breed decreased from 152.1 to 74.4; that of non‐founders (N_enf), from 41.7 to 5.3; and that of founder genome equivalents (N_ge), from 32.7 to 4.9. The corresponding changes in the Kouchi sub‐breed were from 108.2 to 79.4, 16.2 to 4.1, and 14.1 to 3.9. Increasing differences between the two genetic diversity indices in the sub‐breeds indicate that the greater part of the reduction of genetic diversity can be attributed to genetic drift that accumulated in the non‐founder generations. A comparison with published estimates for several cattle breeds suggests the extremely limited genetic diversity of Japanese Brown. In addition to the avoidance of further reduction of genetic diversity, it will be important to counteract the process of breed decline by establishing a production system to efficiently utilize the unique characteristics of this breed and by developing links between the breed and products with market value.     

Combined approaches to identify genomic regions involved in phenotypic differentiation between low divergent breeds: Application in Sardinian sheep populations

Selective breeding has led to modifications in the genome of many livestock breeds. In this study, we identified the genomic regions that may explain some of the phenotypic differences between two closely related breeds from Sardinia. A total of 44 animals, 20 Sardinian Ancestral Black (SAB) and 24 Sardinian White (SW), were genotyped using the Illumina Ovine 50K array. A total of 68, 38 and 15 significant markers were identified using the case–control genome‐wide association study (GWAS), the Bayesian population differentiation analysis (F_ST) and the Rsb metric, respectively. Comparisons among the approaches revealed a total of 22 overlapping markers between GWAS and F_ST and one marker between GWAS and Rsb. Three markers detected by Rsb were also located near (<2 Mb) to highly significant regions identified by GWAS and F_ST analyses. Moreover, one candidate marker identified by GWAS and F_ST approaches was located in a run of homozygosity island that was shared by both breeds. We identified several genes involved in many phenotypic differences (such as stature and growth, reproduction, ear size, coat colour, behaviour) between the two analysed breeds. This study shows that combining several genome‐wide approaches could improve discovery of regions involved in the variability of breeding traits and responsible for the phenotypic diversity even between closely related breeds. Overall, the combination of such genome‐wide methods can be extended to other livestock breeds that share between them a similar genetic background, to understand the process that shapes the patterns of genetic variability between closely related populations.     

Present status of the conservation of livestock genetic resources in Brazil

Brazil has various species of domestic animals, which developed from breeds brought by the Portuguese settlers soon after the discovery of the country. After being submitted to a long process of natural selection, these breeds present characteristics adapted to specific Brazilian environmental conditions, and are known as “criollo”, “local” or naturalized breeds. From the beginning of the 20th century, some exotic breeds, selected in temperate regions, have begun to be imported, and, although more productive, they do not have adaptive traits, such as resistance to disease and parasites found in the naturalized breeds. Even so, they gradually replaced the native breeds, to such an extent that the latter are in danger of extinction. To avoid further loss of this important genetic material, in 1983 the National Research Center for Genetic Resources and Biotechnology (Cenargen) of the Brazilian Agricultural Research Corporation (Embrapa) decided to include conservation of animal genetic resources among its priorities. The conservation activities have been conducted under the Brazilian Genetic Resources Network — Renargen, and is carried out by various research centres of Embrapa, universities and State research institutions, as well as by private farmers, with a single coordinator at national level, Cenargen. The in situ conservation of cattle, horses, buffaloes, donkeys, goats, sheep and pigs is being carried out by Conservation Nuclei, located in the animal's original habitat. Ex situ conservation is centred at the Brazilian Animal Germplasm Bank (AGB), kept at Cenargen. This is responsible for the storage of semen and embryos of various breeds of domestic animals threatened with extinction. Presently the AGB has almost 60,000 doses of semen and more than 250 embryos, as well as over 7000 DNA samples. An important challenge for this program is to increase awareness within the different segments of society in terms of the importance of the conservation of animal genetic resources. The development and evolution of Renargen and its efforts to facilitate the conservation of genetic resources of livestock in Brazil are described in this paper.     

Coagulation profiles of healthy Andalusian donkeys are different than those of healthy horses

Background: Coagulation disorders are frequently diagnosed, especially in hospitalized equidae, and result in increased morbidity and mortality. However, hemostatic reference intervals have not been established for donkeys yet. Objectives: To determine whether the most common coagulation parameters used in equine practice are different between healthy donkeys and horses. Animals: Thirty‐eight healthy donkeys and 29 healthy horses. Methods: Blood samples were collected to assess both coagulation and fibrinolytic systems by determination of platelet count, fibrinogen concentration, clotting times (prothrombin time [PT] and activated partial thromboplastin time [aPTT]), fibrin degradation products (FDP) and D‐Dimer concentrations. Results: PT and aPTT in donkeys were significantly (P < .05) shorter than those of horses. In contrast, FDP and D‐Dimer concentrations were significantly (P < .05) higher in donkeys than in horses. Conclusions and Clinical Importance: The coagulation parameters most commonly determined in equine practice are different in donkeys compared with horses. Thus, the use of normal reference ranges reported previously for healthy horses in donkeys might lead to a misdiagnosis of coagulopathy in healthy donkeys, and unnecessary treatments in sick donkeys. This is the first report of normal coagulation profile results in donkeys, and further studies are warranted to elucidate the physiological mechanisms of the differences observed between donkeys and horses.     

中国家驴生长激素基因内含子2的遗传多样性分析

为了从分子水平揭示家驴的遗传多样性,本研究以中国7个家驴品种(临县驴、关中驴、新疆驴、广灵驴、淮北驴、德州驴、晋南驴)174个个体为试验动物,利用PCR-SSCP技术研究生长激素(GH)基因第2内含子的遗传多样性并进行序列分析.结果显示:第2内含子表现多态性,174个个体检测到3个单倍型,单倍型比例为1.7%.单倍型多样度以临县驴、淮北驴较高,分别为0.678和0.542.广灵驴与关中驴的单倍型多样度(Haplotype diversity,H)比较接近,分别为0.409和0.462.其次为德州驴和晋南驴,分别为0.355和0.304,新疆驴最低为0.077.对该片段的纯合型分别测序发现,B单倍型在第735位碱基G→C.A单倍型在869位碱基G→T.上述结果首次证实驴GH基因内含子2存在多态性.     

The application of genetic markers for cell chimerism diagnosis in lambs

A total of 181 Romanov lambs, including 84 pairs of twins, three litters of triplets and one litter of quadruplets, were investigated. Erythrocyte antigens belonging to six blood group systems were determined with 16 reagents. Six microsatellite loci BMS360, INRA123, McM42, CSSM66, ETH225 and TGLA53 were used to genotype the lambs with automated DNA sizing technology. For cytogenetic diagnosis of leucocyte chimerism, the fluorescence in situ hybridization (FISH) was applied using bovine X and Y chromosome painting probes. The blood‐group typing of lambs from twin and multiple pregnancies allowed us to detect nine cases of erythrocyte chimerism in the investigated population. The analysis of microsatellite DNA sequences showed the presence of cell chimerism in 13 animals. The hybridization of a bovine X and Y chromosome‐specific probe resulted in two yellow brightly luminescent signals in 54,XX cells, and one yellow and one blue signal in 54,XY cells helped us diagnose chromosomal chimerism in seven animals.