A. Schurink

Genetic variables of various manifestations of osteochondrosis and their correlations between and within joints in Dutch warmblood horses

Osteochondrosis (OC) is an important orthopedic developmental disorder in many horse populations. A review of the literature revealed widely variable heritability estimates for the disorder. We estimated the genetic variables (heritabilities and genetic correlations) of various manifestations of OC. Femoropatellar, tarsocrural, and metacarpophalangeal and metatarsophalangeal joints of 811 randomly selected yearlings from the Royal Warmblood Studbook of The Netherlands, descending from 32 representative stallions, were scored for OC at 28 predilection sites. At each site, OC was scored in 5 categories, distinguishing between flattened bone contours and fragments. At the animal level, the overall heritability of OC was 0.23, the heritability of flattened bone contours was 0.08, and the heritability of fragments was 0.22. At the joint level, heritability was greatest in the tarsocrural joints, intermediate in the metacarpophalangeal and metatarsophalangeal joints, and least in the femoropatellar joints. The heritability estimates for the contralateral joint homologs were very similar. The genetic correlation between the tarsocrural and femoropatellar joint was strong, whereas correlations between the metacarpophalangeal and metatarsophalangeal and other joints were moderate. The genetic correlation between flattened bone contours and fragments at the animal level was 0.80. Scoring OC on a 5-point categorical scale resulted in greater heritability on the observed scale than when analyzing OC as a binary trait. Our results suggest that selection against OC could best be performed by taking into account the OC status of all 4 joints, the femoropatellar, the tarsocrural, and the metacarpophalangeal and metatarsophalangeal joints, and discerning between flattened bone contours and fragments.

Heritability and repeatability of insect bite hypersensitivity in Dutch Shetland breeding mares

Insect bite hypersensitivity (IBH) is a seasonal recurrent allergic reaction of horses to the bites of certain Culicoides spp. and is found throughout the world. The aim of our study was to estimate the heritability and repeatability of IBH in the Dutch Shetland pony population. A total of 7,924 IBH scores on 6,073 mares were collected during foal inspections in 2003, 2005, and 2006. Mares were scored for clinical symptoms of IBH from June until February by 16 inspectors. Of all mares, 74.4% (n = 4,520) had a single observation, 20.7% (n = 1,255) had 2 observations, and 4.9% (n = 298) had 3 observations in different years. The overall mean IBH prevalence was 8.8%. Heritability was 0.08 (SE = 0.02) on the observed binary scale and 0.24 (SE = 0.06) on the underlying continuous scale. Repeatability was 0.30 (SE = 0.02) and indicates that including repeated observations of the clinical symptoms of IBH will improve the accuracy of breeding values for IBH. We conclude that IBH, based on clinical symptoms, is a heritable trait in the Dutch Shetland pony population. Therefore, the IBH prevalence in this population can be decreased by selection.

Genome-wide association study of insect bite hypersensitivity in two horse populations in the Netherlands.

BackgroundInsect bite hypersensitivity is a common allergic disease in horse populations worldwide. Insect bite hypersensitivity is affected by both environmental and genetic factors. However, little is known about genes contributing to the genetic variance associated with insect bite hypersensitivity. Therefore, the aim of our study was to identify and quantify genomic associations with insect bite hypersensitivity in Shetland pony mares and Icelandic horses in the Netherlands.MethodsData on 200 Shetland pony mares and 146 Icelandic horses were collected according to a matched case–control design. Cases and controls were matched on various factors (e.g. region, sire) to minimize effects of population stratification. Breed-specific genome-wide association studies were performed using 70 k single nucleotide polymorphisms genotypes. Bayesian variable selection method Bayes-C with a threshold model implemented in GenSel software was applied. A 1 Mb non-overlapping window approach that accumulated contributions of adjacent single nucleotide polymorphisms was used to identify associated genomic regions.ResultsThe percentage of variance explained by all single nucleotide polymorphisms was 13% in Shetland pony mares and 28% in Icelandic horses. The 20 non-overlapping windows explaining the largest percentages of genetic variance were found on nine chromosomes in Shetland pony mares and on 14 chromosomes in Icelandic horses. Overlap in identified associated genomic regions between breeds would suggest interesting candidate regions to follow-up on. Such regions common to both breeds (within 15 Mb) were found on chromosomes 3, 7, 11, 20 and 23. Positional candidate genes within 2 Mb from the associated windows were identified on chromosome 20 in both breeds. Candidate genes are within the equine lymphocyte antigen class II region, which evokes an immune response by recognizing many foreign molecules.ConclusionsThe genome-wide association study identified several genomic regions associated with insect bite hypersensitivity in Shetland pony mares and Icelandic horses. On chromosome 20, associated genomic regions in both breeds were within 2 Mb from the equine lymphocyte antigen class II region. Increased knowledge on insect bite hypersensitivity associated genes will contribute to our understanding of its biology, enabling more efficient selection, therapy and prevention to decrease insect bite hypersensitivity prevalence.

Genetic parameters of insect bite hypersensitivity in Dutch Friesian broodmares

Insect bite hypersensitivity (IBH) is a seasonal allergic skin disease in horses caused by bites of certain Culicoides spp. The aim of our study was to investigate the maternal effect on IBH and to estimate the heritability and repeatability of IBH in the Dutch Friesian horse population. Data consisted of 3,453 Dutch Friesian broodmares with 3,763 visual observations on IBH clinical symptoms scored by 12 inspectors during organized foal inspections in 2004 and 2008. Nine percent of the mares (n = 310) were scored in both years. Mares descended from 144 sires and 2,554 dams and 26.2% of the dams (n = 669) had more than 1 offspring in the data set (range: 2 to 6). Insect bite hypersensitivity was analyzed as a binary trait with a threshold animal model with and without a maternal effect, using a Bayesian approach. Observed IBH prevalence in Dutch Friesian broodmare population was 18.2%. Heritability on the liability scale was 0.16 (SD = 0.06); heritability on the observed scale was 0.07; and repeatability was 0.89 (SD = 0.03). Maternal effect was 0.17 (SD = 0.06) and significantly differed from zero, although the animal model without a maternal effect fitted the data better. These results show that genetic and permanent environmental factors affect IBH in Dutch Friesian horses. The dam affected the IBH development of her offspring through an additive genetic influence but also by being part of their rearing environment.

Culicoides obsoletus extract relevant for diagnostics of insect bite hypersensitivity in horses.

Insect bite hypersensitivity (IBH) is an allergic dermatitis in horses caused by the bites of Culicoides species. The aim of the present study was to evaluate the applicability of whole body extracts of C. obsoletus (the main species found feeding on horses in the Netherlands), C. nubeculosus (rarely found in The Netherlands) and C. sonorensis (typical for North America) for diagnosis of IBH in horses in The Netherlands. Blood and serum samples of 10 clinically confirmed IBH affected and 10 healthy control horses were used to evaluate the IgE titers (ELISA) against the Culicoides whole body extracts of the three Culicoides species. Basophil degranulation was assessed by histamine release test (HRT) after stimulation with these extracts at 5, 0.5 and 0.05 μg/ml. IBH affected horses had significantly higher IgE titers against C. obsoletus than against C. nubeculosus and C. sonorensis. Furthermore, C. obsoletus induced significantly higher histamine release in whole blood of IBH affected horses compared to the other extracts at 0.5 μg/ml. Western blot data revealed IgE binding to many proteins in C. obsoletus extract. This interaction was absent or weak in C. nubeculosus and C. sonorensis extracts for IBH affected horses. Results on individual level indicate that the HRT is more sensitive than ELISA in diagnosing IBH. However, ELISA is more practical as a routine test, therefore the ELISA was further evaluated using C. obsoletus extract on 103 IBH affected and 100 healthy horses, which resulted in a test sensitivity and specificity of 93.2% and 90.0%, respectively. The IgE ELISA readings enabled the analysis of the predicted probability of being IBH affected. From an optical density 450nm value of 0.33 onwards, the probability of IBH affected was more than 0.9. The results presented in this paper show that the use of native Culicoides spp. that feed on horse, is important for improved diagnosis and that the described ELISA based on C. obsoletus can be used routinely to diagnose IBH in countries where this species is the main Culicoides feeding on horses.

Genome-wide association study of insect bite hypersensitivity in Dutch Shetland pony mares

Insect bite hypersensitivity (IBH) is the most common allergic disease present in horses worldwide. It has been shown that IBH is under genetic control, but the knowledge of associated genes is limited. We conducted a genome-wide association study to identify and quantify genomic regions contributing to IBH in the Dutch Shetland pony population. A total of 97 cases and 91 controls were selected and matched on withers height, coat colour and pedigree to minimise the population stratification. A blood sample was collected from participating Shetland pony mares, their IBH phenotype was scored and the owner filled in a questionnaire. A total of 40 021 single-nucleotide polymorphisms (SNPs) were fitted in a univariable logistic model fitting an additive effect. Analysis revealed no effects of population stratification. Significant associations with IBH were detected for 24 SNPs on 12 chromosomes [-log(10) (P-value) > 2.5]. Odds ratios of allele substitution effects of the unfavourable allele were between 1.94 and 5.95. The most significant SNP was found on chromosome 27, with an odds ratio of 2.31 and with an allele frequency of the unfavourable allele of 0.72 in cases and 0.53 in controls. Genome-wide association studies on additional horse populations are desired to validate the identified associations, to identify the genes involved in IBH and to develop genomic tools to decrease IBH prevalence.

Risk factors for insect bite hypersensitivity in Friesian horses and Shetland ponies in The Netherlands

Insect bite hypersensitivity (IBH) is an equine skin allergy caused by bites of Culicoides spp. and impacts on the welfare of affected horses. The aim of this study was to identify and quantify risk factors for IBH. Data from 3453 Friesian horse mares and 7074 Shetland pony mares scored for IBH by inspectors during obligatory foal inspections were analysed using breed-specific multivariable logistic regression models. The combined effect of month and year of scoring, Province and inspector were significantly associated with IBH in both breeds. In Shetland pony mares, withers height and coat colour were also significantly associated with IBH, while body condition had a nearly significant effect. The outcomes from this study on risk factors might contribute to the development of more efficient measures to reduce the prevalence of IBH.

Bayesian Variable Selection to identify QTL affecting a simulated quantitative trait.

BackgroundRecent developments in genetic technology and methodology enable accurate detection of QTL and estimation of breeding values, even in individuals without phenotypes. The QTL-MAS workshop offers the opportunity to test different methods to perform a genome-wide association study on simulated data with a QTL structure that is unknown beforehand. The simulated data contained 3,220 individuals: 20 sires and 200 dams with 3,000 offspring. All individuals were genotyped, though only 2,000 offspring were phenotyped for a quantitative trait. QTL affecting the simulated quantitative trait were identified and breeding values of individuals without phenotypes were estimated using Bayesian Variable Selection, a multi-locus SNP model in association studies.ResultsEstimated heritability of the simulated quantitative trait was 0.30 (SD = 0.02). Mean posterior probability of SNP modelled having a large effect (p^i) was 0.0066 (95%HPDR: 0.0014-0.0132). Mean posterior probability of variance of second distribution was 0.409 (95%HPDR: 0.286-0.589). The genome-wide association analysis resulted in 14 significant and 43 putative SNP, comprising 7 significant QTL on chromosome 1, 2 and 3 and putative QTL on all chromosomes. Assigning single or multiple QTL to significant SNP was not obvious, especially for SNP in the same region that were more or less in LD. Correlation between the simulated and estimated breeding values of 1,000 offspring without phenotypes was 0.91.ConclusionsBayesian Variable Selection using thousands of SNP was successfully applied to genome-wide association analysis of a simulated dataset with unknown QTL structure. Simulated QTL with Mendelian inheritance were accurately identified, while imprinted and epistatic QTL were only putatively detected. The correlation between simulated and estimated breeding values of offspring without phenotypes was high.

Using an Inbred Horse Breed in a High Density Genome-Wide Scan for Genetic Risk Factors of Insect Bite Hypersensitivity (IBH).

While susceptibility to hypersensitive reactions is a common problem amongst humans and animals alike, the population structure of certain animal species and breeds provides a more advantageous route to better understanding the biology underpinning these conditions. The current study uses Exmoor ponies, a highly inbred breed of horse known to frequently suffer from insect bite hypersensitivity, to identify genomic regions associated with a type I and type IV hypersensitive reaction. A total of 110 cases and 170 controls were genotyped on the 670K Axiom Equine Genotyping Array. Quality control resulted in 452,457 SNPs and 268 individuals being tested for association. Genome-wide association analyses were performed using the GenABEL package in R and resulted in the identification of two regions of interest on Chromosome 8. The first region contained the most significant SNP identified, which was located in an intron of the DCC netrin 1 receptor gene. The second region identified contained multiple top SNPs and encompassed the PIGN, KIAA1468, TNFRSF11A, ZCCHC2, and PHLPP1 genes. Although additional studies will be needed to validate the importance of these regions in horses and the relevance of these regions in other species, the knowledge gained from the current study has the potential to be a step forward in unraveling the complex nature of hypersensitive reactions.

A nonsense mutation in B3GALNT2 is concordant with hydrocephalus in Friesian horses.

BackgroundHydrocephalus in Friesian horses is a developmental disorder that often results in stillbirth of affected foals and dystocia in dams. The occurrence is probably related to a founder effect and inbreeding in the population. The aim of our study was to find genomic associations, to investigate the mode of inheritance, to allow a DNA test for hydrocephalus in Friesian horses to be developed. In case of a monogenic inheritance we aimed to identify the causal mutation.ResultsA genome-wide association study of hydrocephalus in 13 cases and 69 controls using 29,720 SNPs indicated the involvement of a region on ECA1 (P <1.68 × 10−6). Next generation DNA sequence analysis of 4 cases and 6 controls of gene exons within the region revealed a mutation in β-1,3-N-acetylgalactosaminyltransferase 2 (B3GALNT2) as the likely cause of hydrocephalus in Friesian horses. The nonsense mutation XM_001491545 c.1423C>T corresponding to XP_001491595 p.Gln475* was identical to a B3GALNT2 mutation identified in a human case of muscular dystrophy-dystroglycanopathy with hydrocephalus. All 16 available cases and none of the controls were homozygous for the mutation, and all 17 obligate carriers (= dams of cases) were heterozygous. A random sample of the Friesian horse population (n = 865) was tested for the mutation in a commercial laboratory. One-hundred and forty-seven horses were carrier and 718 horses were homozygous for the normal allele; the estimated allele frequency in the Friesian horse population is 0.085.ConclusionsHydrocephalus in Friesian horses has an autosomal recessive mode of inheritance. A nonsense mutation XM_001491545 c.1423C>T corresponding to XP_001491595 p.Gln475* in B3GALNT2 (1:75,859,296–75,909,376) is concordant with hydrocephalus in Friesian horses. Application of a DNA test in the breeding programme will reduce the losses caused by hydrocephalus in the Friesian horse population.