Multifactorial inheritance of common white markings in the Arabian horse

The results of a previous study were compatible with the hypothesis that common white facial markings in the Arabian horse have a multifactorial mode of inheritance. I expanded that study to (1) include the legs and therefore obtain insight into the heritability of common white markings in all peripheral regions (face and legs) of the Arabian horse and (2) investigate the influence of sex and the genotypes that produce the bay and chestnut phenotypes on the variation in common white markings. Both studies were based on computerized data obtained from the Arabian Horse Registry of America, Inc. Each leg of a horse was scored from 0 to 5 depending on the amount of whiteness present, and the four leg scores were added to obtain the total leg score for each horse. The facial region was divided into five areas, and each horse was given a score from 0 to 5 according to the number of areas with whiteness. Sire families were analyzed in which each sire family consisted of a sire, his foals, and the dams of those foals. There was a correlation between white facial scores and white leg scores, suggesting that both types of white markings are influenced by the same genetic mechanism. Sire-foal and dam-foal regression analyses were compatible with the hypothesis that common white leg markings also show multifactorial inheritance. Although the results support the model that additively acting genes (polygenes) influence the presence and extent of common white markings, the results also show that males are slightly more marked than are females and that chestnut horses are more heavily marked than are bay horses.(ABSTRACT TRUNCATED AT 250 WORDS)     

Common white facial markings in bay and chestnut Arabian horses and their hybrids

Common white facial and leg markings have a multifactorial mode of inheritance in Equus caballus. Evidence for the complexity of the genetic component is the observation that chestnut (e/e) horses have more extensive white markings than do bay (E/-) horses. Computerized records obtained from the Arabian Horse Registry of America, Inc., were used to determine if heterozygous (E/e) bay horses have more extensive white facial markings than do homozygous (E/E) bay horses. Thirty-five sire families were analyzed. Each sire family consists of a sire, his foals, and the dams of those foals. The facial region was divided into five areas, and each horse was given a score from 0 to 5 according to the number of areas with whiteness. Since dams and foals with E/E genotypes cannot be identified in these sire families, mean facial scores were compared in dams and foals that were E/e and E/-. It was assumed that if a difference exists between E/e and E/E horses, the presence of E/E horses in the E/- group would reduce the mean of the E/- group. The results show that Arabian horses with the genotype E/e have more white markings than do horses with the genotype E/-, leading to the conclusion that horses with the genotypes e/e, E/e, and E/E vary as to the quantitative expression of white facial markings, with heterozygotes having an intermediate expression.     

Does homozygosity contribute to the asymmetry of common white leg markings in the Arabian horse?

Common white and facial markings have a multifactorial mode of inheritance inEquus caballus and result from the absence of melanocytes in the unpigmented areas. Directional asymmetry and fluctuating asymmetry apparently account for the total asymmetry of common white leg markings. Using computerized records obtained from the Arabian Horse Registry of America, Inc., and the International Arabian Horse Association, studies were carried out to determine if homozygosity increases the total asymmetry in common white leg markings by presumably promoting fluctuating asymmetry. The results were as follows: (1) Arabian horses that are symmetrical and asymmetrical for common white leg markings have similar distributions of inbreeding coefficients; (2) Arabian and half-Arabian horses have similar concordance values, in general, for specific white markings in both their forelegs and hind legs. It is concluded that homozygosity does not contribute to the total asymmetry of common white leg markings in the Arabian horse.     

Directional and anteroposterior asymmetry of common white markings in the legs of the Arabian horse: response to selection

Arabian bay horses manifest, on the average, more common white markings in their hind legs than their forelegs (anteroposterior asymmetry) and more common white markings in their left legs than their right legs (directional asymmetry). To determine if genetic variation exists for these types of asymmetry, the phenotypic response was studied in bay foals when their dams and sires were selected for the directions of fore-hind and left-right differences. In the fore-hind studies, the quantitative shifts in the bay foals were in the direction specified by the selection scheme and the observed deviations were all statistically significant. The shifts were also consistently in the direction favored by selection in the left-right studies, but only two of six observed deviations were statistically significant using a one-tailed test of significance. Thus, only marginal statistical evidence is available to support the observed consistent responses to selection in the left-right studies. These differential responses are reflected in the magnitudes of the heritability estimates. Based on the overall results, it is concluded that both types of asymmetry have a genetic basis in the Arabian horse, but much more genetic variation is present for anteroposterior asymmetry than for directional asymmetry. This revised version was published online in July 2006 with corrections to the Cover Date.     

Blood typing invalidates alleged evidence of fertility of XO mare

A mare with XO gonadal dysgenesis was reported to have produced two foals. Blood samples from the mare, her two foals, their sire and the mare's sire were typed for blood-group and serum protein variants in conjunction with registry requirements. Both foals qualified as offspring of the reported parents. However, the blood-typed mare could be excluded as an offspring of her alleged sire. An alternative hypothesis to explain the blood-type and karyotype findings was that a fertile mare had been substituted for the XO mare, as surrogate mother and blood-type donor. A computer search of 120,000 blood-type records identified only one other horse with the same blood type as the dam of the foals. That horse was a mare of the same breed and owned by the person who had attempted to register foals from the XO mare. These blood-type findings invalidated the allegation of XO fertility and emphasize the need for parentage verification to support reports of unusual reproductive performance.     

Deletion of 2.7 kb near HOXD3 in an Arabian horse with occipitoatlantoaxial malformation

In the horse, the term occipitoatlantoaxial malformation (OAAM) is used to describe a developmental defect in which the first cervical vertebra (atlas) resembles the base of the skull (occiput) and the second cervical vertebra (axis) resembles the atlas. Affected individuals demonstrate an abnormal posture and varying degrees of ataxia. The homeobox (HOX) gene cluster is involved in the development of both the axial and appendicular skeleton. Hoxd3‐null mice demonstrate a strikingly similar phenotype to Arabian foals with OAAM. Whole‐genome sequencing was performed in an OAAM‐affected horse (OAAM1) and seven unaffected Arabian horses. Visual inspection of the raw reads within the region of HOXD3 identified a 2.7‐kb deletion located 4.4 kb downstream of the end of HOXD4 and 8.2 kb upstream of the start of HOXD3. A genotyping assay revealed that both parents of OAAM1 were heterozygous for the deletion. Additional genotyping identified two of 162 heterozygote Arabians, and the deletion was not present in 371 horses of other breeds. Comparative genomics studies have revealed that this region is highly conserved across species and that the entire genomic region between Hoxd4 and Hoxd3 is transcribed in mice. Two additional Arabian foals diagnosed with OAAM (OAAM 2 and 3) were genotyped and did not have the 2.7‐kb deletion. Closer examination of the phenotype in these cases revealed notable variation. OAAM3 also had facial malformations and a patent ductus arteriosus, and the actual malformation at the craniocervical junction differed. Genetic heterogeneity may exist across the HOXD locus in Arabian foals with OAAM.     

A missense mutation in the endothelin-B receptor gene is associated with Lethal White Foal Syndrome: an equine version of Hirschsprung Disease

Lethal White Foal Syndrome is a disease associated with horse breeds that register white coat spotting patterns. Breedings between particular spotted horses, generally described as frame overo, produce some foals that, in contrast to their parents, are all white or nearly all white and die shortly after birth of severe intestinal blockage. These foals have aganglionosis characterized by a lack of submucosal and myenteric ganglia from the distal small intestine to the large intestine, similar to human Hirschsprung Disease. Some sporadic and familial cases of Hirschsprung Disease are due to mutations in the endothelin B receptor gene (EDNRB). In this study, we investigate the role of EDNRB in Lethal White Foal Syndrome. A cDNA for the wild-type horse endothelin-B receptor gene was cloned and sequenced. In three unrelated lethal white foals, the EDNRB gene contained a 2-bp nucleotide change leading to a missense mutation (I118K) in the first transmembrane domain of the receptor, a highly conserved region of this protein among different species. Seven additional unrelated lethal white foal samples were found to be homozygous for this mutation. No other homozygotes were identified in 138 samples analyzed, suggesting that homozygosity was restricted to lethal white foals. All (40/40) horses with the frame overo pattern (a distinct coat color pattern that is a subset of overo horses) that were tested were heterozygous for this allele, defining a heterozygous coat color phenotype for this mutation. Horses with tobiano markings included some carriers, indicating that tobiano is epistatic to frame overo. In addition, horses were identified that were carriers but had no recognized overo coat pattern phenotype, demonstrating the variable penetrance of the mutation. The test for this mutant allele can be utilized in all breeds where heterozygous animals may be unknowingly bred to each other including the Paint Horse, Pinto horse, Quarter Horse, Miniature Horse, and Thoroughbred. Received: 25 November 1997 / Accepted: 3 February 1998     

Genetic analysis of white facial and leg markings in the Swiss Franches-Montagnes Horse Breed

White markings and spotting patterns in animal species are thought to be a result of the domestication process. They often serve for the identification of individuals but sometimes are accompanied by complex pathological syndromes. In the Swiss Franches-Montagnes horse population, white markings increased vastly in size and occurrence during the past 30 years, although the breeding goal demands a horse with as little depigmented areas as possible. In order to improve selection and avoid more excessive depigmentation on the population level, we estimated population parameters and breeding values for white head and anterior and posterior leg markings. Heritabilities and genetic correlations for the traits were high (h(2) > 0.5). A strong positive correlation was found between the chestnut allele at the melanocortin-1-receptor gene locus and the extent of white markings. Segregation analysis revealed that our data fit best to a model including a polygenic effect and a biallelic locus with a dominant-recessive mode of inheritance. The recessive allele was found to be the white trait-increasing allele. Multilocus linkage disequilibrium analysis allowed the mapping of the putative major locus to a chromosomal region on ECA3q harboring the KIT gene.     

Seasonal changes in the white blood cell system, lyzozyme activity and cortisol level in Arabian brood mares and their foals

In 34 pure-breed Arabian horses divided into four groups (Gr. I, ten pregnant mares; Gr. II, seven barren mares; Gr. III, ten foals born in 1981; Gr. IV, seven foals born in 1982) seasonal changes in the white blood cell system, cortisol level and lyzozyme activity were studied. Seasonal periodicity was found in all groups for the number of lymphocytes, segmented neutrophils and eosinophils and cortisol level. Leukocyte periodicity was found in three groups, but not in the barren mares. In lyzozyme activity there was periodicity in three groups but not in the youngest foals. In the stab neutrophils, basophils and monocytes no cycle was observed. The behaviour of the indices studied showed the influence of age of the horses (mature vs young) and the physiological state of the mares (pregnancy or barrenness).     

Characterization of a red blood cell antigen in donkeys and mules associated with neonatal isoerythrolysis

A red cell antigen of donkeys and mules was identified using antibodies in serum from a mare which produced a mule foal affected with neonatal isoerythrolysis (NI). Subsequently antibodies with similar activity were identified in the sera of other mares which had produced mule foals and were produced by immunization of horses with blood from donkeys. The antigen detected by these antibodies does not correspond to any recognized horse red cell alloantigen. This may be a xenoantigen since all donkeys (and mules) tested have shared this antigen and all horses tested have lacked the antigen. The results suggest that all mule pregnancies (donkey sire x horse dam) are incompatible with regard to this factor and a potential for neonatal isoerythrolysis exists in all cases.     

Analyses of TCRB rearrangements substantiate a profound deficit in recombination signal sequence joining in SCID foals: implications for the role of DNA-dependent protein kinase in V(D)J recombination

We reported previously that the genetic SCID disease observed in Arabian foals is explained by a defect in V(D)J recombination that profoundly affects both coding and signal end joining. As in C.B-17 SCID mice, the molecular defect in SCID foals is in the catalytic subunit of the DNA-dependent protein kinase (DNA-PKCS); however, in SCID mice, signal end resolution remains relatively intact. Moreover, recent reports indicate that mice that completely lack DNA-PKCS also generate signal joints at levels that are indistinguishable from those observed in C.B-17 SCID mice, eliminating the possibility that a partially active version of DNA-PKCS facilitates signal end resolution in SCID mice. We have analyzed TCRB rearrangements and find that signal joints are reduced by approximately 4 logs in equine SCID thymocytes as compared with normal horse thymocytes. A potential explanation for the differences between SCID mice and foals is that the mutant DNA-PKCS allele in SCID foals inhibits signal end resolution. We tested this hypothesis using DNA-PKCS expression vectors; in sum, we find no evidence of a dominant-negative effect by the mutant protein. These and other recent data are consistent with an emerging consensus: that in normal cells, DNA-PKCS participates in both coding and signal end resolution, but in the absence of DNA-PKCS an undefined end joining pathway (which is variably expressed in different species and cell types) can facilitate imperfect signal and coding end joining.     

A third allele in the horse albumin system

Starch gel electrophoresis according to Gahne (1966) reveals three new phenotypes designated FI, I and IS in horse serum albumin. Family material comprising 97 sire families with a total number of 2742 foals and their dams and population material comprising 2867 adult horses representative of the Swedish Trotter breed were examined. The results were consistent with a genetic theory of three codominant, autosomal alleles F, I and S controlling the six albumin phenotypes observed. The gene frequencies were estimated to be 0.489, 0.049 and 0.462, respectively, for the F, I and S genes in the Swedish Trotter breed. The risk of making false exclusions in paternity controls in the albumin system, if the I allele is not considered, is discussed.     

Linkage of tobiano coat spotting and albumin markers in a pony family

Genetic segregation patterns among blood type markers and various phenotypically observed traits were studied in a small herd of ponies. The herd consisted of 10 mares without white spotting and a single stallion with the dominant pattern of tobiano spotting. Comparison of segregation patterns at loci for which the stallion was heterozygous showed tight linkage for the Alb-B and tobiano markers. In 17 cases in which the Alb contribution of the sire could be determined, all 10 foals that inherited AlbB from him were tobiano spotted, and all 7 non-spotted foals inherited his AlbA. The use of the symbol To is proposed for dominantly inherited tobiano spotting linked to the albumin.     

A primary production deficit in the thrombocytopenia of equine infectious anemia.

The purpose of this study was to identify the mechanisms responsible for the thrombocytopenia that develops following infection of horses by the lentivirus equine infectious anemia virus (EIAV). Immunocompetent Arabian foals and Arabian foals with severe combined immunodeficiency (SCID), which lack functional B and T lymphocytes, were experimentally infected with EIAV. Levels of viremia and a number of clinical and hematologic parameters were examined prior to and following infection. Thrombocytopenia was not dependent on the immune response: SCID foals were affected as severely as immunocompetent foals. Production of platelets, measured by metabolic incorporation of radioactive label, was significantly reduced. The decrease ranged from 35 to 89% in three SCID and two immunocompetent foals examined. Platelet survival, measured by 51Cr labeling, also declined following infection in both SCID and immunocompetent foals: 51 and 68%, respectively, relative to the preinfection life spans. The difference between immunocompetent and immunodeficient foals was not statistically significant. The number of megakaryocytes (MK) per square millimeter of bone marrow, determined by digitizing morphometry, was not significantly altered in either SCID or immunocompetent thrombocytopenic foals. Numbers of denuded MK nuclei per unit area increased, but the elevation was not statistically significant. No evidence for viral replication in MK was found. Three different parameters of intravascular coagulation (activated prothombin time, fibrin degradation products, and one-step prothombin time) remained normal until after platelet numbers had declined significantly, arguing against an important role for disseminated intravascular coagulation. The findings indicate that EIAV induces thrombocytopenia principally through an indirect, noncytocidal suppressive effect on platelet production, the mechanism of which is unknown. A shortening of platelet life span apparently contributes moderately to the platelet deficit as well. The shortening of platelet life span is multifactorial in origin, including both mechanisms that depend on an active immune response and those that do not.     

Genome-wide linkage and association analysis identifies major gene Loci for guttural pouch tympany in arabian and german warmblood horses

Equine guttural pouch tympany (GPT) is a hereditary condition affecting foals in their first months of life. Complex segregation analyses in Arabian and German warmblood horses showed the involvement of a major gene as very likely. Genome-wide linkage and association analyses including a high density marker set of single nucleotide polymorphisms (SNPs) were performed to map the genomic region harbouring the potential major gene for GPT. A total of 85 Arabian and 373 German warmblood horses were genotyped on the Illumina equine SNP50 beadchip. Non-parametric multipoint linkage analyses showed genome-wide significance on horse chromosomes (ECA) 3 for German warmblood at 16-26 Mb and 34-55 Mb and for Arabian on ECA15 at 64-65 Mb. Genome-wide association analyses confirmed the linked regions for both breeds. In Arabian, genome-wide association was detected at 64 Mb within the region with the highest linkage peak on ECA15. For German warmblood, signals for genome-wide association were close to the peak region of linkage at 52 Mb on ECA3. The odds ratio for the SNP with the highest genome-wide association was 0.12 for the Arabian. In conclusion, the refinement of the regions with the Illumina equine SNP50 beadchip is an important step to unravel the responsible mutations for GPT.     

Inheritance of guttural pouch tympany in the arabian horse

The objective of the present study was to analyze the mode of inheritance of guttural pouch tympany (GPT) using pedigrees of Arabian horses. Complex segregation analyses were employed to test for the significance of nongenetic transmission and for monogenic, polygenic, and mixed monogenic-polygenic modes of inheritance. Horses affected by GPT comprised 27 Arabian purebred foals. Of these 27 animals, 22 were patients at the Clinic for Horses, School of Veterinary Medicine Hannover, Hannover, Germany, between 1994 and 2001 and 5 Arabian foals were from stud farms. Information on the pedigrees of these patients allowed us to classify the affected foals into four families with a total of 276 animals. The regressive logistic model analysis took into account the nonrandomness of the pedigrees through multiple single ascertainment correction. The complex segregation analysis showed that, among all other models employed, a polygenic and a mixed monogenic-polygenic model best explained the segregation of Arabian foals with GPT. Models including only nongenetic distributions and monogenic inheritance could be significantly rejected. This is the first report in which a genetic component could be shown to be responsible for GPT in horses.     

Chinese Mongolian horses may retain early domestic male genetic lineages yet to be discovered

The Mongolian horse represents one of the most ancient extant horse populations. In this study we determined the male‐specific region of the Y chromosome (MSY) haplotype distribution in 60 Chinese Mongolian horses representing five distinct populations. Cosmopolitan male lineages were predominant in horses from one improved (Sanhe), one Chinese Mongolian subtype (Baicha Iron Hoof) and one indigenous (Abaga Black) population. In contrast, autochthonous Y chromosome diversity was evident among the two landrace populations (Wushen and Wuzhumuqin), as the majority of their MSY haplotypes were situated at root nodes in a network. Our results also suggest gene flow between Chinese Mongolian and Arabian horses, as an appreciable number of Wuzhumuqin horses carried haplotypes that are typically observed in Arabian horses. Although most horses carried modern haplotypes as a direct result of recent breed improvement, authentic Chinese Mongolian horses retain an ancient signature of paternal lineages that has not previously been described in extant horse populations. Therefore, further characterization of MSY variation in these populations will be important for the discovery of lost diversity in modern domestic horses and also for understanding the evolutionary history of equine paternal lineages.     

Evidence for eumelanin and pheomelanin producing genotypes in the Arabian horse

The ultrastructural imaging of melanocytes coupled with analyses to detect sulfur-containing melanosomes by energy-dispersive X-ray spectroscopy were used to test the hypothesis that the yellowish-red and black pigments found in Arabian horses result from pheomelanogenesis and eumelanogenesis, respectively. These procedures detected pheomelanosomes in follicles at the base of hairs in chestnut horses and eumelanosomes in follicles at the base of hairs in black horses. By analyzing tissue obtained by skin biopsy, these procedures also demonstrated that skin melanocytes in a chestnut horse produce eumelanosomes, and follicular melanocytes in the same horse produce pheomelanosomes. It was also shown that the type of follicular melanosome present in light bay horses is correlated with the color of the hair. The results of this study give experimental evidence for the Odriozola-Adalsteinsson hypothesis that the e allele is responsible for the chestnut phenotype; they also give fine structure and chemical confirmation of the action of the A and E loci in the Arabian horse as currently proposed for the mouse and other mammals.     

A comparative study of oculomotor,trochlear and abducens nerves in Arabian foals

We investigated the microscopic structure of transverse sections of the oculomotor, trochlear and abducens nerves of Arabian foals using stereological methods. Bilateral nerve pairs from 2-month-old female Arabian foals were analyzed. The tissues were embedded in plastic blocks, then 1 µm thick sections were cut and stained with osmium tetroxide and methylene blue-azure II. Stereology was performed using light microscopy. Morphometry showed that the right and left pairs of nerves were similar. The transverse sectional areas of the oculomotor, trochlear and abducens nerves were 1.93 ± 0.19 mm², 0.32 ± 0.06 mm² and 0.70 ± 0.08 mm², respectively. The oculomotor nerve exhibited a significantly greater number of myelinated axons (16755 ± 1279) and trochlear (2656 ± 494) and the abducens nerves (4468 ± 447). The ratio of the axon diameter to myelinated nerve fiber diameter was 0.58, 0.55 and 0.55 for the oculomotor, trochlear and abducens nerves, respectively. Of the three nerves studied, the abducens nerve exhibited the greatest nerve fiber area, myelin area, nerve and axon diameters, and myelin thickness. The ratio of small myelinated nerve fibers was greatest in the oculomotor nerve.