Exploring the genetics underpinning dynamic laryngeal collapse associated with poll flexion in Norwegian-Swedish Coldblooded Trotter racehorses.

BACKGROUND: Dynamic laryngeal collapse (DLC) associated with poll flexion is the most common disorder of the upper respiratory tract (URT) in the Norwegian-Swedish Coldblooded Trotter (NSCT). The disorder, which has also been diagnosed in other breeds of trotters and gaited horses, appears to be related to anatomic phenotypes and only occurs during poll flexion when the horse is exercised 'on the bit'. OBJECTIVES: Identify genomic regions associated with DLC in the NSCT by combining a rigorous phenotyping protocol with genomic data from a high-density equine genotyping array. STUDY DESIGN: Prospective case/control study. METHODS: High-speed treadmill endoscopy was used to phenotype horses (n = 61) for DLC, distinguishing between cases and controls. Genome-wide association (GWA) analysis of DLC status was then performed using a principal component approach (PCA) with haplotype analyses subsequently performed for regions containing single-nucleotide polymorphisms (SNPs) above the suggestive genome-wide significance (GWS) threshold (P<1.0 × 10-5 ). RESULTS: One region containing 10 SNPs (Equus caballus chromosome [ECA] 7: 89,601,935-94,647,192) was above the suggestive GWS threshold. Two inferred haplotypes in this region demonstrated significant differences (P<0.001) between cases and controls, with the most frequent haplotype resulting in a significantly increased risk of DLC. MAIN LIMITATIONS: Small sample size as a result of stringent phenotyping protocols. CONCLUSIONS: The current study highlights a candidate genomic region on ECA7 as potentially important with regard to the manifestation of DLC. Further exploration of this region and the genes included within it will bring veterinarians and researchers closer to fully understanding the biological mechanisms underlying DLC in horses.

Two missense mutations in melanocortin 1 receptor (MC1R) are strongly associated with dark ventral coat color in reindeer (Rangifer tarandus).

The protein-coding region of melanocortin 1 receptor (MC1R) was sequenced to identify potential variation affecting coat color in reindeer (Rangifer tarandus). A T→C sequence variation at nucleotide position 218 (c.218T>C) causing an amino acid (aa) change from methionine to threonine at aa position 73 (p.Met73Thr) was identified. In addition, a T→G sequence variation was found at nucleotide position 839 (c.839T>G), causing phenylalanine to be exchanged by cysteine at aa position 280 (p.Phe280Cys). The two sequence variants (c.218C and c.839G) were found to be closely associated with a darker belly coat compared with animals not having any of these two variants. The aa acid change p.Met73Thr affects the same position as p.Met73Lys previously reported to give constitutive activation of MC1R in black sheep (Ovis aries), whereas p.Phe280Cys is identical to one of two variants previously reported to be associated with dark coat color in Arctic fox (Alopex lagopus), supporting that the two variants found in reindeer are functional. The complete absence of Thr73 and Cys280 among the 51 wild reindeer analyzed provides some evidence that these variants are more common in the domestic herds.

Worldwide frequency distribution of the 'Gait keeper' mutation in the DMRT3 gene.

For centuries, domestic horses have represented an important means of transport and served as working and companion animals. Although their role in transportation is less important today, many horse breeds are still subject to intense selection based on their pattern of locomotion. A striking example of such a selected trait is the ability of a horse to perform additional gaits other than the common walk, trot and gallop. Those could be four-beat ambling gaits, which are particularly smooth and comfortable for the rider, or pace, used mainly in racing. Gaited horse breeds occur around the globe, suggesting that gaitedness is an old trait, selected for in many breeds. A recent study discovered that a nonsense mutation in DMRT3 has a major impact on gaitedness in horses and is present at a high frequency in gaited breeds and in horses bred for harness racing. Here, we report a study of the worldwide distribution of this mutation. We genotyped 4396 horses representing 141 horse breeds for the DMRT3 stop mutation. More than half (2749) of these horses also were genotyped for a SNP situated 32 kb upstream of the DMRT3 nonsense mutation because these two SNPs are in very strong linkage disequilibrium. We show that the DMRT3 mutation is present in 68 of the 141 genotyped horse breeds at a frequency ranging from 1% to 100%. We also show that the mutation is not limited to a geographical area, but is found worldwide. The breeds with a high frequency of the stop mutation (>50%) are either classified as gaited or bred for harness racing.

Heritability estimates of tarsocrural osteochondrosis and palmar/plantar first phalanx osteochondral fragments in Standardbred trotters.

REASONS FOR PERFORMING STUDY: The pathogenesis of osteochondrosis (OC) and palmar/plantar first phalanx osteochondral fragments (POFs) is multifactorial, but specific knowledge of heritability is limited. OBJECTIVES: To improve the precision of heritability estimates and to estimate the genetic correlation between tarsocrural OC and POFs in Standardbred trotters. Further aims were to examine whether the prevalence of OC/POFs was different in the American and French lineages that have contributed to the Norwegian population, and if the prevalence was affected by heterozygosity. STUDY DESIGN: Retrospective cohort study. METHODS: Categorical data on tarsocrural OC and POFs from 2 radiographic studies performed in 1989 and 2007/2008 (n = 1217) were analysed with sire threshold models that included 230 sires. RESULTS: Heritability of OC at the distal intermediate ridge of the tibia and/or the lateral trochlear ridge of the talus was estimated at 0.29 ± 0.15. For OC at the distal intermediate ridge of the tibia only, the estimate was 0.40 ± 0.17. Heritability of POFs in all 4 limbs was estimated at 0.23 ± 0.13; for metatarsophalangeal POFs this was 0.26 ± 0.13 and for medial metatarsophalangeal POFs 0.32 ± 0.14. Estimates of genetic correlation between OC and POFs ranged from 0.68 ± 0.27 to 0.73 ± 0.28 but were not significantly different from a zero-genetic correlation. Effects of lineages or heterozygosity were not observed. CONCLUSIONS AND POTENTIAL RELEVANCE: This study confirmed a moderate to high heritability of tarsocrural OC and POF, providing further evidence of the heritable nature of these diseases. Examination of specific lesions yielded the highest heritability; therefore, breeding programmes and future genome-analysis studies should focus on predilection sites rather than the entire disease complex.

Equine developmental orthopaedic diseases--a genome-wide association study of first phalanx plantar osteochondral fragments in Standardbred trotters.

Palmar/plantar osteochondral fragments (POF) in fetlock joints commonly affect and influence the athletic performance of horses. In this study, we used the Equine SNP50 BeadChip® to perform a genome-wide association study of metatarsophalangeal POF in 176 Norwegian Standardbred trotter yearlings. Putative quantitative trait loci (QTL) for medial and/or lateral POF, and medial POF only were identified on ECA1, 2, 7, 9 and 31, whereas for lateral POF, only on ECA7, 11, 27 and X. The moderate number of QTL evidences a complex inheritance and suggests various genes controlling POF development in medial and lateral locations.

Persistence of vigilance and flight response behaviour in wild reindeer with varying domestic ancestry.

Knowledge about changes in behavioural traits related to wildness and tameness is for most mammals lacking, despite the increased trend of using domestic stock to re-establish wild populations into historical ranges. To test for persistence of behavioural traits of wild reindeer (Rangifer tarandus L.) exposed to hunting, we sampled DNA, vigilance and flight responses in wild reindeer herds with varying domestic ancestry. Analyses of 14 DNA microsatellite loci revealed a dichotomous main genetic structure reflecting their native origin, with the Rondane reindeer genetically different from the others and with least differentiation towards the Hardangervidda reindeer. The genetic clustering of the reindeer in Norefjell-Reinsjøfjell, Ottadalen and Forollhogna, together with domestic reindeer, supports a predominant domestic origin of these herds. Despite extensive hunting in all herds, the behavioural measures indicate increasing vigilance, alert and flight responses with increasing genetic dissimilarity with domestic herds. Vigilance frequency and time spent vigilant were higher in Rondane compared to Hardangervidda, which again were higher than herds with a domestic origin. We conclude that previous domestication has preserved a hard wired behavioural trait in some reindeer herds exhibiting less fright responses towards humans that extensive hunting has, but only slightly, altered. This brings novel and relevant knowledge to discussions about genetic diversity of wildlife in general and wild reindeer herds in Norway in specific.

Osteochondrosis and osteochondral fragments in Standardbred trotters: prevalence and relationships.

REASONS FOR PERFORMING STUDY: Developmental orthopaedic diseases (DOD) such as osteochondrosis (OC)/osteochondrosis dissecans (OCD), palmar/plantar osteochondral fragments (POF), ununited palmar/plantar eminences (UPE) and dorsoproximal first phalanx fragments are well recognised in the horse. Aetiopathogeneses are controversial and molecular genetic screening of DNA has recently been employed for their elucidation. Precise phenotypic definition and knowledge of breed-specific prevalence and interrelations are essential for the interpretation of following genomic studies in Standardbred trotters. OBJECTIVES: To assess the prevalence, trend of development and interrelation of DOD in tarsocrural, metacarpophalangeal (MCP) and metatarsophalangeal (MTP) joints in Standardbred trotters. METHODS: The tarsocrural and MCP/MTP joints of 464 Norwegian Standardbred yearlings were radiographed and the prevalence and interrelation of osteochondral lesions calculated. RESULTS: Osteochondral lesions were diagnosed in 50.7% of the horses. The prevalence of tarsocrural OC/OCD at the distal intermediate ridge of the tibia (DIT) and the lateral trochlear ridge of the talus (LTT) was 19.3%. The prevalence of OC/OCD in MCP joints was 3.6%, whereas those of POF and UPE in MCP/MTP joints were 23.1 and 3.9%, respectively. Interrelation was evident for 1) most equivalent lesions in joint homologues, 2) OCD DIT and OCD LTT and 3) POF and UPE. Lesions in hock and fetlock joints were generally not significantly associated. CONCLUSIONS: The prevalence of tarsocrural OC/OCD in Norwegian Standardbreds is apparently increasing, whereas that of other articular DOD appears stable. Association analyses verify bilateralism for most equivalent lesions and suggest aetiological resemblance also between other lesions. The absence of a significant association between tarsocrural OCD and POF implies that the lesions must be considered statistically different disorders. POTENTIAL RELEVANCE: The prevalence results emphasise that DOD should be considered in Standardbred breeding regimens (e.g. by sire selection subsequent to progeny testing). Also, improved phenotypic definitions will help elucidate the true causal genes in following genomic studies.

Genome-wide association analysis of osteochondrosis of the tibiotarsal joint in Norwegian Standardbred trotters.

Osteochondrosis (OC), a disturbance in the process of endochondral ossification, is by far the most important equine developmental orthopaedic disease and is also common in other domestic animals and humans. The purpose of this study was to identify quantitative trait loci (QTL) associated with osteochondrosis dissecans (OCD) at the intermediate ridge of the distal tibia in Norwegian Standardbred (SB) using the Illumina Equine SNP50 BeadChip whole-genome single-nucleotide polymorphism (SNP) assay. Radiographic data and blood samples were obtained from 464 SB yearlings. Based on the radiographic examination, 162 horses were selected for genotyping; 80 of these were cases with an OCD at the intermediate ridge of the distal tibia, and 82 were controls without any developmental lesions in the joints examined. Genotyped horses descended from 22 sires, and the number of horses in each half-sib group ranged from 3 to 14. The population structure necessitated statistical correction for stratification. When conducting a case-control genome-wide association study (GWAS), mixed-model analyses displayed regions on chromosomes (Equus callabus chromosome - ECA) 5, 10, 27 and 28 that showed moderate evidence of association (P ≤ 5 × 10(-5); this P-value is uncorrected i.e. not adjusted for multiple comparisons) with OCD in the tibiotarsal joint. Two SNPs on ECA10 represent the most significant hits (uncorrected P=1.19 × 10(-5) in the mixed-model). In the basic association (chi-square) test, these SNPs achieved statistical significance with the Bonferroni correction (P=0.038) and were close in the permuted logistic regression test (P=0.054). Putative QTL on ECA 5, 10, 27 and 28 represent interesting areas for future research, validation studies and fine mapping of candidate regions. Results presented here represent the first GWAS of OC in horses using the recently released Illumina Equine SNP50 BeadChip.

Museum specimens reveal changes in the population structure of northern Fennoscandian domestic reindeer in the past one hundred years.

Traditional reindeer herding of northern Fennoscandia has been based on seasonal movements independent of national borders. At the beginning of the 19th century, these yearly movements of reindeer were excessive, but during that century the borders between the Fennoscandian countries were closed. By analysing a 190-base pair fragment of the mitochondrial DNA control region in 79 museum samples, we show that the reindeer of northern Fennoscandia were one homogenous population shortly after the national borders were closed. However, anthropogenic activity has effectively ended genetic exchange within northern Fennoscandia and has made the reindeer population within this region heterogeneous. Genetic input of eastern origin is also suggested within the extant Russian reindeer of the Kola Peninsula.

Genome sequence, comparative analysis, and population genetics of the domestic horse.

We report a high-quality draft sequence of the genome of the horse (Equus caballus). The genome is relatively repetitive but has little segmental duplication. Chromosomes appear to have undergone few historical rearrangements: 53% of equine chromosomes show conserved synteny to a single human chromosome. Equine chromosome 11 is shown to have an evolutionary new centromere devoid of centromeric satellite DNA, suggesting that centromeric function may arise before satellite repeat accumulation. Linkage disequilibrium, showing the influences of early domestication of large herds of female horses, is intermediate in length between dog and human, and there is long-range haplotype sharing among breeds.

International Equine Gene Mapping Workshop Report: a comprehensive linkage map constructed with data from new markers and by merging four mapping resources.

A comprehensive male linkage map was generated by adding 359 new, informative microsatellites to the International Equine Gene Map half-sibling reference families and by combining genotype data from three independent mapping resources: a full sibling family created at the Animal Health Trust in Newmarket, United Kingdom, eight half-sibling families from Sweden and two half-sibling families from the University of California, Davis. Because the combined data were derived primarily from half-sibling families, only autosomal markers were analyzed. The map was constructed from a total of 766 markers distributed on the 31 equine chromosomes. It has a higher marker density than that of previously reported maps, with 626 markers linearly ordered and 140 other markers assigned to a chromosomal region. Fifty-nine markers (7%) failed to meet the criteria for statistical evidence of linkage and remain unassigned. The map spans 3,740 cM with an average distance of 6.3 cM between markers. Fifty-five percent of the intervals are < or = 5 cM and only 3% > or = 20 cM. The present map demonstrates the cohesiveness of the different data sets and provides a single resource for genome scan analyses and integration with the radiation hybrid map.

Absorption, retention and tissue distribution of radiocaesium in reindeer: effects of diet and radiocaesium source.

Radiocaesium absorption and retention in reindeer (Rangifer tarandus) calves was compared in groups fed diets containing different proportions of lichen and concentrates, and different chemical forms of radiocaesium ((134)CsCl in solution or fallout from the Chernobyl accident). Daily intakes of fallout radiocaesium were 15-23 kBq, while daily intakes of (134)CsCl ranged from 70 kBq to 1,160 kBq. The half-life for radiocaesium in red blood cells (RBC) in animals fed with a pure lichen diet exceeded that in animals fed with a combined diet of lichen and concentrates by 40% (17.8+/-0.7 days vs. 12.7+/-0.4 days). Corresponding differences in the half-lives for urinary and faecal excretion were about 60% and 40%, respectively. Transfer coefficients (F(f)) to reindeer meat were estimated to be 0.25+/-0.01 days kg(-1) for fallout radiocaesium and 1.04+/-0.03 days kg(-1) for (134)CsCl, reflecting differences in both radiocaesium bioavailability and retention. The bioavailability of the Chernobyl radiocaesium in lichen in 1988 was estimated at ca. 35% compared to (134)CsCl.

Genetic relationship between Mongolian and Norwegian horses?

Human populations of Central Asian origin have contributed genetic material to northern European populations. It is likely that migrating humans carried livestock to ensure food and ease transportation. Thus, eastern genes could also have dispersed to northern European livestock populations. Using microsatellite data, we here report that the essentially different genetic distances DA and (deltamu)2 and their corresponding phylogenetic trees show close associations between the Mongolian native horse and northern European horse breeds. The genetic distances between the northern European breeds and Standardbred/Thoroughbred, representing a southern-derived source of horses, were notably larger. We suggest that contribution of genetic material from eastern horses to northern European populations is likely to have occurred.

Evaluation of factors affecting individual assignment precision using microsatellite data from horse breeds and simulated breed crosses.

Assignment tests have been utilized to investigate population classification, measure genetic diversity and to solve forensic questions. Using microsatellite data from 26 loci genotyped in eight horse breeds we examined how population differentiation, number of scored loci, number of scored animals per breed and loci variability affected individual assignment precision applying log likelihood methods. We found that both genetic differentiation and number of scored loci were highly important for recognizing the breed of origin. When comparing two and two breeds, a proportion of 95% of the most differentiated breeds (0.200 < or = FST < or = 0.259) could be identified scoring only three loci, while the corresponding number was six for the least differentiated breeds (0.080 < or = FST < or = 0.139). An identical proportion of simulated breed crosses, differentiated from their parental breeds by FST estimates in the range 0.050-0.069, was identified when scoring 12 loci. This level of source identification was not obtained for the less differentiated breed crosses. The current data further suggested that population sample size and locus variability were not critical for the assignment precision as long as moderately large sample sizes (> or = 20 animals per population) and fairly variable loci were used.

Breed demarcation and potential for breed allocation of horses assessed by microsatellite markers.

Population demarcation of eight horse breeds was investigated using genotype information of 306 horses from 26 microsatellite loci. The breeds include the indigenous Norwegian breeds Fjord Horse, Nordland/Lyngen Horse, Døle Horse and Coldblooded Trotter together with Icelandic Horse, Shetland Pony, Standardbred and Thoroughbred. Both phylogenetic analysis and a maximum likelihood method were applied to examine the potential for breed allocation of individual animals. The phylogenetic analysis utilizing simple allele sharing statistics revealed clear demarcation among the breeds; 95% of the individuals clustered together with animals of the same breed in the phylogenetic tree. Even breeds with a short history of divergence like Døle Horse and Coldblooded Trotter formed distinct clusters. Implementing the maximum likelihood method allocated 96% of the individuals to their source population, applying an assignment stringency of a log of the odds ratio larger than 2. Lower allocation stringency assigned nearly all the horses. Only three individuals were wrongly allocated a breed by both methods. In conclusion, the study demonstrates clear distinction among horse breeds, and by combining the two assignment methods breed allocation could be determined for more than 99% of the individuals.

Genetic relationships in the peregrine falcon (Falco peregrinus) analysed by microsatellite DNA markers.

Microsatellite DNA markers were developed from a peregrine falcon (Falco peregrinus) and genetic relationships among peregrine falcons in southern Norway were analysed using the markers. The genomic DNA library was screened for the presence of dinucleotide microsatellite repeats. Twelve loci revealed polymorphism through the initial analysis of 24 unrelated peregrine falcons, and Mendelian inheritance was confirmed in two peregrine falcon families bred in captivity. The estimated mean probability of identical genotypes in two unrelated individuals was 3 x 10-8, and the combined exclusion probability for parentage testing was 0.99 and 0.94 for one or both parents unknown, respectively. The markers were used to investigate the parentage of peregrine broods from the same nest site from different breeding seasons, and subsequently the nest-site fidelity of the breeding peregrines. High nest-site fidelity was found by studying pairwise comparisons of relatedness (rxy) estimates among chicks at six nest sites from three different breeding seasons. Cross-species amplifications showed that most loci also appeared to amplify polymorphic products in the gyrfalcon (F. rusticolus), merlin (F. columbarius), hobby (F. subbuteo) and kestrel (F. tinnunculus), demonstrating that the loci will provide powerful genetic markers in these falcons too.