Identification of runs of homozygosity in Western honey bees (Apis mellifera) using whole-genome sequencing data.

Runs of homozygosity (ROH) are continuous homozygous segments that arise through the transmission of haplotypes that are identical by descent. The length and distribution of ROH segments provide insights into the genetic diversity of populations and can be associated with selection signatures. Here, we analyzed reconstructed whole-genome queen genotypes, from a pool-seq data experiment including 265 Western honeybee colonies from Apis mellifera mellifera and Apis mellifera carnica. Integrating individual ROH patterns and admixture levels in a dynamic population network visualization allowed us to ascertain major differences between the two subspecies. Within A. m. mellifera, we identified well-defined substructures according to the genetic origin of the queens. Despite the current applied conservation efforts, we pinpointed 79 admixed queens. Genomic inbreeding (F ROH) strongly varied within and between the identified subpopulations. Conserved A. m. mellifera from Switzerland had the highest mean F ROH (3.39%), while queens originating from a conservation area in France, which were also highly admixed, showed significantly lower F ROH (0.45%). The majority of A. m. carnica queens were also highly admixed, except 12 purebred queens with a mean F ROH of 2.33%. Within the breed-specific ROH islands, we identified 14 coding genes for A. m. mellifera and five for A. m. carnica, respectively. Local adaption of A. m. mellifera could be suggested by the identification of genes involved in the response to ultraviolet light (Crh-BP, Uvop) and body size (Hex70a, Hex70b), while the A. m. carnica specific genes Cpr3 and Cpr4 are most likely associated with the lighter striping pattern, a morphological phenotype expected in this subspecies. We demonstrated that queen genotypes derived from pooled workers are useful tool to unravel the population dynamics in A. mellifera and provide fundamental information to conserve native honey bees.

Determining Objective Parameters to Assess Gait Quality in Franches-Montagnes Horses for Ground Coverage and Over-Tracking - Part 2: At Trot.

In gait quality assessments of horses, stride length (SL) is visually associated with spectacular movements of the front limbs, and described as ground coverage, while the movement of the hind limb under the body is supposedly essential to a longer over-tracking distance (OTD). To identify movement patterns with strong associations to SL and OTD, limb and body kinematics of 24 Franches-Montagnes (FM) stallions were measured with 3D optical motion capture (OMC) on a treadmill during an incremental speed test at trot (3.3-6.5 m/s). These measurements were correlated to the scores of ground coverage and over-tracking from six breeding experts. The amount of explained variance of parameters on SL and OTD were estimated using linear mixed-effect models in two models: a full model with all parameters measurable with OMC, and a reduced model with a subset of parameters measurable with inertial measurement units (IMUs). The front limb stance duration (16%) and OTD (7%) measured with OMC, or the OMC parameters front limb stance duration (24%) and suspension duration (14%) measurable with IMUs explained most variance in SL. However, four of six breeding experts were also significantly correlated (r>|0.41|) to front limb protraction angle. OTD variance was explained with OMC parameters suspension duration (10%) and hind limb contralateral pro-retraction angles (9%) or IMU-measurable parameters suspension duration (20%) and maximal pelvis pitch (5%). Four experts' scores for over-tracking were correlated to suspension duration. These results underscore the need for precise definitions of gait quality traits.

Estimates of Genetic Parameters for Shape Space Data in Franches-Montagnes Horses.

Conformation traits such as joint angles are important selection criteria in equine breeding, but mainly consist of subjective evaluation scores given by breeding judges, showing limited variation. The horse shape space model extracts shape data from 246 landmarks (LM) and objective joint angle measurements from triplets of LM on standardized horse photographs. The heritability was estimated for 10 joint angles (seven were measured twice with different LM placements), and relative warp components of the whole shape, in 608 Franches-Montagnes (FM) horses (480 stallions, 68 mares and 60 geldings born 1940-2018, 3-25 years old). The pedigree data comprised 6986 horses. Genetic variances and covariances were estimated by restricted maximum likelihood model (REML), including the fixed effects birth year, age (linear and quadratic), height at withers (linear and quadratic), as well as postural effects (head, neck, limb position and body alignment), together with a random additive genetic animal component and the residual effect. Estimated heritability varied from 0.08 (stifle joint) to 0.37 (poll). For the shape, the type was most heritable (0.36 to 0.37) and evolved from heavy to light over time. Image-based phenotyping can improve the selection of horses for conformation traits with moderate heritability (e.g., poll, shoulder and fetlock).

Determining Objective Parameters to Assess Gait Quality in Franches-Montagnes Horses for Ground Coverage and Over-Tracking - Part 1: At Walk.

Ground coverage and over-tracking are two gait quality traits describing the forward movement of the front respectively the hind limbs in relation to stride length and over-tracking distance. To investigate the complex interplay of different movement patterns in ground coverage and over-tracking, limb and body kinematics of 24 Franches-Montagnes (FM) stallions were measured with 3D optical motion capture (OMC) on a treadmill during an incremental speed test at the walk (1.4-2.0 m/s). The significance and amount of explained variance of kinematic parameters on stride length and over-tracking distance were estimated using linear mixed-effect models, with speed and horse as random effects. Two separate models were tested: a full model with all parameters measurable by OMC, and a reduced model with a subset of parameters also measurable with inertial measurement units (IMUs). The kinematic parameters were correlated to the subjective scores from six breeding experts to interpret their external validity. The parameter for ground coverage at the walk, explaining most of the variance in stride length, were the maximal forelimb retraction angle (11%) measured with OMC, and the range of pelvis pitch (10%) if measuring with IMUs. The latter was also the most relevant for quantifying over-tracking, explaining 24% to 33% of the variance in the over-tracking distance. The scores from most breeding experts were significantly correlated (r ≥ |0.41|) with the fore- and hind limb protraction angles, which reflect the textual definition of ground coverage and over-tracking. Both gait quality traits can be objectively quantified using either OMC or IMUs.

Social Box: Influence of a New Housing System on the Social Interactions of Stallions When Driven in Pairs.

In order to improve the housing conditions of stallions in individual boxes, we tested a so-called "social box" allowing increased physical contact between neighbouring horses. This study investigated whether housing stallions in social boxes changes the number of social interactions during carriage driving. We hypothesised that the stay in social boxes would decrease the number of unwanted social interactions between stallions when driven in pairs. Eight Franches-Montagnes breeding stallions were observed when driven in pairs with a "neutral" stallion housed in a so-called "conventional box", strongly limiting physical contact. They were driven on a standardised route over the course of four days before, during, and after being housed in social boxes. The type and frequency of behaviours of the pairs and the interventions of the groom and the driver during the test drives were assessed live and using video recordings. Results from linear mixed-effect models show that unwanted social interactions decreased during and after the stallions were housed in the social box (p < 0.001). Stallions' interactions also decreased over the four days (p < 0.01), suggesting a habituation to the test conditions by learning not to interact, or by subtly settling dominance. The social box tended to decrease unwanted social behaviours of stallions driven in pairs and could therefore be used as an environmental enrichment for horses.

Gait quality scoring data of Franches-Montagnes stallions at walk and trot on a treadmill by experts of the breed and their reliability.

This article presents the data obtained from the scoring of 24 stallions of the Franches-Montagnes (FM) horse breed by six experts of this breed. The experts scored six traits at walk and eight at trot from the video recordings of these stallions walking and trotting on a treadmill during an incremental speed test. The scores were given on a scale of one to nine. All experts scored the same videos twice (two scoring tests) with a time interval of two years, and without feedback from the first scoring. Video sequences were presented in a different order between first and second scoring. The inter- and intra-rater reliability of the data was assessed using intraclass correlation coefficients (ICC) to evaluate its quality.

A Novel QTL and a Candidate Gene Are Associated with the Progressive Motility of Franches-Montagnes Stallion Spermatozoa after Thaw.

The use of frozen-thawed semen is an important reproduction tool to preserve the biodiversity of small, native horse breeds such as the Franches-Montagnes (FM). However, not all stallions produce cryotolerant semen with a progressive motility after thaw ≥ 35%. To improve our understanding of the genetic background of male fertility traits in both fresh and frozen-thawed semen, we performed genome-wide association studies (GWAS) on gel-free volume, sperm cell concentration, total sperm count, and progressive motility in fresh and frozen-thawed semen from 109 FM stallions using 335,494 genome-wide single nucleotide polymorphisms (SNPs). We identified one significant (p < 1.69 × 10-7) quantitative trait locus (QTL) on ECA6 within the SCN8A gene for progressive motility after thaw, which was previously associated with progressive motility in boars. Homozygous stallions showed a substantial drop in progressive motility after thaw. This QTL could be used to identify cryointolerant stallions, avoiding the costly cryopreservation process. Further studies are needed to confirm whether this QTL is also present in other horse breeds.

Using Different Combinations of Body-Mounted IMU Sensors to Estimate Speed of Horses-A Machine Learning Approach.

Speed is an essential parameter in biomechanical analysis and general locomotion research. It is possible to estimate the speed using global positioning systems (GPS) or inertial measurement units (IMUs). However, GPS requires a consistent signal connection to satellites, and errors accumulate during IMU signals integration. In an attempt to overcome these issues, we have investigated the possibility of estimating the horse speed by developing machine learning (ML) models using the signals from seven body-mounted IMUs. Since motion patterns extracted from IMU signals are different between breeds and gaits, we trained the models based on data from 40 Icelandic and Franches-Montagnes horses during walk, trot, tölt, pace, and canter. In addition, we studied the estimation accuracy between IMU locations on the body (sacrum, withers, head, and limbs). The models were evaluated per gait and were compared between ML algorithms and IMU location. The model yielded the highest estimation accuracy of speed (RMSE = 0.25 m/s) within equine and most of human speed estimation literature. In conclusion, highly accurate horse speed estimation models, independent of IMU(s) location on-body and gait, were developed using ML.

Reliable and clinically applicable gait event classification using upper body motion in walking and trotting horses.

Objectively assessing horse movement symmetry as an adjunctive to the routine lameness evaluation is on the rise with several commercially available systems on the market. Prerequisites for quantifying such symmetries include knowledge of the gait and gait events, such as hoof to ground contact patterns over consecutive strides. Extracting this information in a robust and reliable way is essential to accurately calculate many kinematic variables commonly used in the field. In this study, optical motion capture was used to measure 222 horses of various breeds, performing a total of 82 664 steps in walk and trot under different conditions, including soft, hard and treadmill surfaces as well as moving on a straight line and in circles. Features were extracted from the pelvis and withers vertical movement and from pelvic rotations. The features were then used in a quadratic discriminant analysis to classify gait and to detect if the left/right hind limb was in contact with the ground on a step by step basis. The predictive model achieved 99.98% accuracy on the test data of 120 horses and 21 845 steps, all measured under clinical conditions. One of the benefits of the proposed method is that it does not require the use of limb kinematics making it especially suited for clinical applications where ease of use and minimal error intervention are a priority. Future research could investigate the extension of this functionality to classify other gaits and validating the use of the algorithm for inertial measurement units.

Should We Agree to Disagree? An Evaluation of the Inter-Rater Reliability of Gait Quality Traits in Franches-Montagnes Stallions.

Gait quality, that is, the way horses move according to functional and aesthetic principles, englobes many traits that are scored by experts during breeding competitions. The experts can score a trait on a subjective valuating (SV) scale or on a linear profiling (LP) scale representing the biological extremes of the population. However, the reliability of the appraisal of gait quality traits has not been extensively evaluated. In this study, seven breed experts appraised the walk and trot quality of 24 Franches-Montagnes stallions presented in hand on a sand track. Inter-rater reliabilities of six traits (five SV and one LP) at the walk and eight traits (five SV and three LP) at the trot were estimated with intraclass correlation coefficients (ICCs). The inter-rater reliabilities were poor (ICC < 0.50). The scale anchoring varied between experts, and the variance of scores was low. There were no systematic differences in inter-rater reliability between LP and SV traits. Future studies should determine whether the inter-rater reliabilities may be increased by a more precise definition of the scores within each trait to improve the absolute agreement between experts, by a more uniform scale anchoring between experts, and by decreasing the number of scale items. However, considering the inherent limitations of the human eye in observing high-speed movement, the use of a field-applicable kinematic measurement system may support breeding experts in the appraisal of gait quality traits in the future.

Genome-Wide Association Studies Based on Equine Joint Angle Measurements Reveal New QTL Affecting the Conformation of Horses.

The evaluation of conformation traits is an important part of selection for breeding stallions and mares. Some of these judged conformation traits involve joint angles that are associated with performance, health, and longevity. To improve our understanding of the genetic background of joint angles in horses, we have objectively measured the angles of the poll, elbow, carpal, fetlock (front and hind), hip, stifle, and hock joints based on one photograph of each of the 300 Franches-Montagnes (FM) and 224 Lipizzan (LIP) horses. After quality control, genome-wide association studies (GWASs) for these traits were performed on 495 horses, using 374,070 genome-wide single nucleotide polymorphisms (SNPs) in a mixed-effect model. We identified two significant quantitative trait loci (QTL) for the poll angle on ECA28 (p = 1.36 × 10-7), 50 kb downstream of the ALX1 gene, involved in cranial morphology, and for the elbow joint on ECA29 (p = 1.69 × 10-7), 49 kb downstream of the RSU1 gene, and 75 kb upstream of the PTER gene. Both genes are associated with bone mineral density in humans. Furthermore, we identified other suggestive QTL associated with the stifle joint on ECA8 (p = 3.10 × 10-7); the poll on ECA1 (p = 6.83 × 10-7); the fetlock joint of the hind limb on ECA27 (p = 5.42 × 10-7); and the carpal joint angle on ECA3 (p = 6.24 × 10-7), ECA4 (p = 6.07 × 10-7), and ECA7 (p = 8.83 × 10-7). The application of angular measurements in genetic studies may increase our understanding of the underlying genetic effects of important traits in equine breeding.

Repeatability, reproducibility and consistency of horse shape data and its association with linearly described conformation traits in Franches-Montagnes stallions.

Linear description (LD) of conformation traits was introduced in horse breeding to minimise subjectivity in scoring. However, recent studies have shown that LD traits show essentially the same problems as traditionally scored traits, such as data converging around the mean value with very small standard deviations. To improve the assessment of conformation traits of horses, we investigated the application of the recently described horse shape space model based upon 403 digitised photographs of 243 Franches-Montagnes (FM) stallions and extracted joint angles based on specific landmark triplets. Repeatability, reproducibility and consistency of the resulting shape data and joint angles were assessed with Procrustes ANOVA (Rep) and intra-class correlation coefficients (ICC). Furthermore, we developed a subjective score to classify the posture of the horses on each photograph. We derived relative warp scores (PCs) based upon the digitised photos conducting a principal component analysis (PCA). The PCs of the shapes and joint angles were compared to the posture scores and to the linear description data using linear mixed effect models including significant posture scores as random factors. The digitisation process was highly repeatable and reproducible for the shape (Rep = 0.72-0.99, ICC = 0.99). The consistency of the shape was limited by the age and posture (p < 0.05). The angle measurements were highly repeatable within one digitiser. Between digitisers, we found a higher variability of ICC values (ICC = 0.054-0.92), indicating digitising error in specific landmarks (e.g. shoulder point). The posture scores were highly repeatable (Fleiss' kappa = 0.713-0.857). We identified significant associations (p(X2) < 0.05) with traits describing the withers height, shoulder length and incline, overall leg conformation, walk and trot step length. The horse shape data and angles provide additional information to explore the morphology of horses and therefore can be applied to improve the knowledge of the genetic architecture of LD traits.