Crossbreeding between individuals of different breeds and
introgression, the transfer of genes between breeds and/or populations mediated
primarily by backcrossing, have been characteristic tools used in the
refinement or optimisation of practical horse breeding. In this study
we analysed the genetic contribution of the Arabian horse to the gene pool
of the Lipizzan horse and its association with the overall type via shape
regression analysis in 158 Lipizzan horses from the Austrian federal stud
farm of Piber and the Spanish Riding School. Although crossbreeding
with Arabian horses took place between 1776 and 1945, we found a
significant association between Lipizzan body shape (
Within the refinement processes of horse breeding, throughout the 19th century oriental horses were commonly used for crossbreeding and introgression of positive performance and character traits. Whereas the English Thoroughbred represents the current meliorator breed for warmblood sport horses, Arabian horses played this important role a century before. The Lipizzan horse is known as a classical baroque riding horse, a classification that is supported by the fact that 60 % of founder genes come from baroque Spanish and Italian ancestors born in the 18th century. Nevertheless, Arabian breeding animals were used in the Lipizzan breed between 1776 and 1945, and their founder gene proportion comprised about 23 % in the populations of eight European Lipizzan state stud farms (Zechner et al., 2002). In a genealogical context the introgression of Arabian genes into the Lipizzan gene pool is documented by the classical sire line Siglavy, founded by an original Arabian stallion imported from Syria in 1814, and by several dam lines going back to Arabian founder mares imported between 1830 and 1865. In the 19th century the breeding of Arabian horses was supported by the Austro-Hungarian Empire at the state stud farms Rǎdǎuţi, Bábolna, and Mezőhegyes. The resulting Arabian horse, today called the “Shagya Arabian”, represents a continental Arabian, based on the crossbreeding of original Arabian stallions with Turkish, Kazakh, Moldavian, and Hungarian founder mares that were born before 1800. From 1800 to 1934 original Arabian breeding mares and stallions were constantly imported from Syria in order to consolidate the Arabian population. Due to interactions between the Austro-Hungarian state stud farms and the imperial stud farm Lipica, both breeds, the Lipizzan and the Shagya Arabian, share a common founder pool dating to the time period from 1810 to 1865.
The effects of Arabian genes on morphology and conformation have been the focus of practical breeding for a long time (Bilek, 1914; Schwark, 1984; Schröder et al., 2010). In the case of the Lipizzan horse, the first analysis was published by Bilek in 1914, who studied the morphological differences between Lipizzans with varying Arabian gene proportions based on anatomical body measurements. In a more recent work, Druml and Sölkner (2012) documented differences in the extent of Arabian gene proportions in the gene pools of eight European Lipizzan state stud farms of Austria, Croatia, Hungary, Italy, Romania, Slovakia, and Slovenia using pedigree analysis, where Hungarian and Romanian Lipizzans were characterised by lower genetic contributions of Arabian founder animals. In a morphologic characterisation study by means of anatomical body measurements in the same eight Lipizzan state studs, Zechner et al. (2001) could show significant morphological differences between horses of the single stud farms, which were interpreted as a result of selection in favour for specific types (riding horse or carriage horse).
The aim of our study was to document the Arabian gene pool in the current Lipizzan population of Piber and the Spanish Riding School and to test the association of the body shapes of Lipizzan horses with individual levels of Arabian genetic contributions by means of geometric morphometric (GM) methods. Furthermore, we wanted to test if GM methods based on image analyses are an appropriate tool to differentiate phenotypic traits between animals of varying Arabian genetic background. Further, we used a sample of Shagya Arabian horses from the Slovak national stud farm of Topol'čianky as a reference population for comparing estimated oriental shape transitions in the Lipizzan with the existing differences between Lipizzans and Shagya Arabians.
In order to derive the individual Arabian gene proportions of 158 Lipizzan
horses (93 stallions from the Spanish Riding School, 65 mares from the
federal stud farm Piber) at a mean age of 10.9 years (minimum age
For the same set of animals, which represents the actual state-owned
Austrian Lipizzan breeding population, we collected standardized digital
photographs following the protocol of Druml et al. (2015, 2016) (see
Fig. S1 in the Supplement). The photos were taken by one of the authors and the imaging process
was repeated several times where a minimum of two line-ups per horse were
performed per session. For the selection of the pictures used, an optimal
fit criterion regarding the stance of the horse (open posture) was applied
by visual assessment (Fig. S1). The Arabian reference population was imaged
by the same person in one session following the above-mentioned procedure in
the Slovak national stud farm of Topol'čianky, where 32 Shagya Arabians
(5 stallions, 27 mares) from 11 sires (mean number of 2.9 offspring per
sire) at a mean age of 6.1 years (minimum age
In order to retrieve informative shape data from the standardised images, we applied a shape model that includes the coordinates of parts of the outline and 31 somatometric and anatomical landmarks. The 215 outline coordinates were defined as sliding semi-landmarks, which allow the single points to slide along the outline curve during the rotation process of the generalised Procrustes analysis (GPA). Hence the outline coordinates result in homologous-like points along a curve (Gunz and Mitteröcker, 2013) and can be compared directly to each other (Fig. 1).
Shape model for phenotyping the horses. On the Lipizzan stallion
Before proceeding to statistical analyses of shape data and/or variables the
individual coordinates need to be normalised, as images also contain
information concerning different body size, orientation, and location of the
animal. Such uninformative effects can be eliminated or corrected by means
of a generalised Procrustes superimposition, which scales, rotates, and
centres every single specimen onto the mean configuration of the sample.
This statistical procedure eliminates non-shape-associated variation by
superimposing landmark configurations using least-squares estimates for
translation and rotation parameters. First, the centroid of each
configuration is translated to the origin, and configurations are scaled to
a common unit size. The centroid is defined as the mean of all
At the end of this procedure, the original coordinate data are replaced by
substitute Cartesian coordinates (shape coordinates; Bookstein, 1991), as
they vary around their own sample mean, and are corrected for effects of
scale (body size), for effects of orientation, and for effects of location of
the original specimens. The resulting shape coordinates ((
To evaluate the association of Arabian genetic contribution with the body shape
of 158 Lipizzan horses, we applied so-called shape regressions, in which shape
coordinates were regressed onto the individual Arabian gene proportions
(
Descriptive statistics were calculated using the procedures proc freq and
proc means of the SAS software package, version 9.1 (SAS Institute, 2009).
For the analysis of differences in shape between 158 Lipizzan horses and 32
Shagya Arabians, we applied a generalised linear model (glm) as follows:
The most important founder animals of the Lipizzan population from the federal stud farm of Piber and the Spanish Riding School defining 50 % of the gene pool. Among these 15 founders, four Arabian stallions (marked by bold letters) together contribute 16.6 % of genes to the total founder gene pool.
The pedigree used for this analysis was maximally 33 generations long. In the 12th generation 90 % of ancestors were known and in the 17th generation the percentage of known ancestors dropped beyond 50 %. In total the Lipizzan gene pool of this sample was defined by 434 founder animals, whereas the 15 most influential founder animals, including four Arabian stallions, accounted for 50 % of the pool (Fig. 2).
The proportion of genetic contribution of Arabian founders in the Lipizzan population of the Austrian federal stud farm of Piber.
The mean genetic contribution of Arabian founders to the gene pool of the
Lipizzan population of the federal stud farm of Piber and the Spanish Riding School
achieved, in total, 26 %, and it was contributed by 72 founder animals (22 mares
contributing 1.4 % and 50 stallions contributing 24.6 %). Of
the Arabian genes, 20.1 % were due to imported original Arabians from Syria between
the years 1800 and 1865. Most of these 22 original Arabians were the founder animals for the Shagya Arabian breed in the state stud
farms of the Austro-Hungarian monarchy at the same
time. Furthermore, 32 Shagya Arabians
contributed 1.4 % of genes within the time period from 1817 to 1934. The
remaining 4.5 % of the Arabian gene pool was contributed by so-called Arabian founders
(15 horses), Anglo-Arabian founders (one horse), purebred Arabians (three purebred
Arabian stallions), one Tunisian stallion, and three Turco-Arabian stallions
(see Table 1; Table S1 in the Supplement). On an individual basis, the Arabian genetic
contributions varied from 21.0 to 29.0 % (SD
Proportion of shape variance explained by PC (Var. explained);
coefficient of determination of the linear model (
n.s.: not significant.
The GPA of coordinate data from 158 Lipizzans and 32 Shagya Arabians
resulted in 189 shape variables (PCs), from which 17 PCs explained 95 % of
total shape variation (Table 2). According to the glm analysis, 12 of these
17 PCs remained uninformative (
The largest shape differences were found for PC2 (explaining 17.5 % of
shape variation) and PC3 (explaining 9.5 % of shape variation) for the
breed effect (
For the shape variables PC1 (explaining 36.0 % of shape variation), PC5 (explaining 5.3 % of shape variation), and PC6 (explaining 4.3 % of shape variation) no significant results could be derived by the linear model as these three variables mostly accounted for individual posing differences (PC1 for neck posture; PC5 and PC6 for leg and stance posture).
Scatter plot of shape variables and plot of PC2 and PC3 containing
158 Lipizzan and 32 Shagya Arabian horses
Significant (
The scatter plot for PC3 and PC4 in Fig. 3 shows that younger
Shagya Arabians (
Mean shapes for 32 Shagya Arabians and 158 Lipizzan horses. The
differences in landmark configurations between both mean shapes are shown in
panel
The mean shapes for Lipizzans and Arabians are shown in Fig. 4 and illustrate the typical conformation of the two breeds. The mean Lipizzan configuration is characterised by a heavier head with a tendency toward a ram-shaped profile; a muscled, well curved neck; and a curved top line. The withers are less expressed and the hindquarters are well muscled. The mean Shagya Arabian configuration shows a clear concave head profile with pronounced cheekbones. The withers are long and well expressed; the back is straight with a high-positioned tail. The detailed shape differences are illustrated in Fig. 4c, where the arrows indicate the landmark deviations of the mean Arabian shape from the mean Lipizzan shape. The main deviations were found in the conformation of neck and head, followed by the withers and the loans. Whereas the slope of shoulder, hip bone, and neck (neck pose/ankle) was similar in both mean configurations, we can observe that in the Arabian shape the extremities were longer (Fig. 4c).
Estimated Lipizzan shapes along the regression curve for the
Although the variation in the Arabian gene proportions from 21 to 29 %
can be considered low, a significant association between the Arabian
genetic contribution (
The mean Arabian gene proportions in the stud farm population of Piber did not substantially change throughout the last 20 years, as Zechner et al. (2002) calculated a nearly identical contribution of 26.2 % at the population level. The major part of Arabian genes (21.2 %) in the current Austrian Lipizzan population is derived from original Arabian founders imported from Syria from 1800 to 1865. The gene pool calculated in this study supports the previous findings of Druml and Sölkner (2012), in which the highest levels of Arabian genes were documented in the stud farms Lipica (28.1 %), Piber (26.2 %), and Monterotondo (26.0 %). The breeding objectives in these three stud farms are defined by the classical dressage riding horse, whereas the Hungarian and Romanian Lipizzan populations represent multipurpose carriage-driving horses and are characterised by lower Arabian gene proportions varying between 17.4 % (Beclean and Fǎgǎraş) and 22.7 % (Szilvásvárad). The current mean Lipizzan shape of Piber as described using the Procrustes analysis, represents the classical Iberian or baroque riding horse type and is significantly different from the Shagya Arabian population of the stud farm Topol'čianky.
Although the Lipizzan population mean represents the Iberian or baroque type
and the crossbreeding events with Arabian horses took place between 1776 and
1945, we were able to detect an association of Arabian gene proportions
(
Previous analyses of shape variation and conformation judging preferences in the Lipizzan population of Piber (Druml et al., 2015, 2016) demonstrated that the classical Iberian or baroque Lipizzan type represents the ideal breeding goal. The advantage of this image-based morphometric approach over conventional linear measuring methods is that the retrieved model horses can be easily understood and can be used for the adjustment of conformation scoring protocols and/or breeding goals in the context of breeding programs.
From the results of this study we can assume that an underlying conserved oriental genotype is responsible for the shape differences shown between Iberian and oriental Lipizzan horse types. The presented phenotyping and analysing method can be used for identifying major effects of genetic introgression and it can be used to predict transitions of conformation with increasing and decreasing genetic contributions of foreign breeds.
Data are available upon reasonable request.
TD and GGS designed the experiments and TD, MD, and MH carried them out. MH and TD analysed the data and GGS and TD prepared the paper.
The authors declare that they have no conflict of interest.
The authors wish to thank the Austrian Research Promotion Agency (FFG) and Xenogenetik for financial support and the federal stud farm of Piber and the Spanish Riding School in Vienna and the National stud farm Topol'ćianky for assistance and cooperation. Edited by: Manfred Mielenz Reviewed by: four anonymous referees