Genet. Sel. Evol.

39 (2007) 609619
c INRA, EDP Sciences, 2007


DOI: 10.1051/gse:2007025

Available online at:

www.gse-journal.org

Original article

A new view on dam lines in Polish Arabian

horses based on mtDNA analysis
Iwona Ga , Anna Wa , Barbara Gb
Renata Pa , Jerzy Sa
a

Department of Genetics and Cytology, University of Gdansk, Kadki 24,

80-822 Gdansk, Poland
b
Polish Academy of Sciences Institute of Genetics and Animal Breeding, Jastrzebiec,
05-552 Wlka Kosowska, Poland

(Received 5 December 2006; accepted 9 March 2007)

Abstract Polish Arabian horses are one of the oldest and the most important Arab populations
in the world. The Polish Arabian Stud Book and the Genealogical Charts by Skorkowski are the
main sources of information on the ancestors of Polish Arabs. Both publications were viewed
as credible sources of information until the 1990s when the data regarding one of the dam lines
was questioned. The aim of the current study was to check the accuracy of the pedigree data
of Polish dam lines using mtDNA analysis. The analyses of a 458 bp mtDNA D-loop fragment
from representatives of 15 Polish Arabian dam lines revealed 14 distinct haplotypes. The results
were inconsistent with pedigree data in the case of two lines. A detailed analysis of the historical
sources was performed to explain these discrepancies. Our study revealed that representatives
of dierent lines shared the same haplotypes. We also noted a genetic identity between some
lines founded by Polish mares of unknown origin and lines established by desert-bred mares.
mitochondrial DNA / horse / pedigree / phylogeny

1. INTRODUCTION
Polish Arabian horses are one of the oldest and most important Arab populations in the world. The first stud described in historical sources was established
in 1778 [16]. Nineteenth century studs were located in the southeast region
of historical Poland, which corresponds now to the Ukrainian territory. The
foundation breeding material was of a dierent origin. The Biaa Cerkiew and
Sawuta stocks are traced back to local mares, and five dam lines originating
from these studs are currently present in Polish breeding (Tab. I).

Corresponding author: [email protected]

Article published by EDP Sciences and available at http://www.gse-journal.org

or http://dx.doi.org/10.1051/gse:2007025

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Table I. Dam lines in Arabian horses in Poland: founders and main branches of the
lines.

* In brackets: percentage of representatives of the line of brood mares registered in 2003;

db: desert-bred mares.
** In brackets: generation number with the founder generation considered as 0;
abbreviations: A: Austro-Hungarian Empire, Radautz stud; PL: Poland; SU-USSR:
Tersk stud.

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611

The second group of founders consisted of horses imported from the Near
East, and included the mares Gazella, Mlecha, and Sahara, the founders of very
valuable dam lines.
World War I and the October Revolution resulted in the total destruction
of most of the Polish studs [7]. Extensive reconstruction of Polish Arabian
breeding was undertaken in the subsequent years, and mares representing the
new dam lines of Rodania, Selma, and Cherifa were imported to Polish studs.
World War II inflicted another ruinous blow to Polish breeding, during which
most of the studs were destroyed, and the horses were either plundered or were
missing [13, 14, 16]. Post-war breeding started with 59 surviving brood mares
and 14 mares from the Babolna stud [7]. The representatives of four new dam
lines were among Babolnas mares (Tab. I), and the Szamrajwka and Rodania
lines were revived by importing representatives from the Tersk stud (former
USSR).
While Polish Arabian breeding started at the end of the 18th century, the history of the Polish studbook dates back to 1926; only a few reports containing
incomplete pedigree data were published beforehand [16]. More or less accurate breeding registers were kept in some studs; unfortunately, they were completely destroyed in 19141918. Eorts to create an ocial studbook were undertaken in the 1920s and the first studbook, edited by Dr. Edward Skorkowski
was published in 1926, which was then revised and published in 1932 as the
Polish Arabian Stud Book (PASB), Vol. I. In 1938, Skorkowski published the
Genealogical Charts of Polish Arabian horses. Due to the scrupulous work performed by the editor in preparing these charts, both publications were considered for years to be credible sources of information on the ancestors of Polish
Arabs. It was not until the 1990s that one of the female lines was called into
doubt. According to Kwiatkowski [11], one of two branches of the Milordka
line, in fact, originates from the mare Malikarda.
Research methods based on mtDNA analysis can permit the verification of
Kwiatkowskis hypothesis. The verification of dam lines, the history of which
was re-created based on incomplete sources, was the second aim of the current
study. The third aim of the work was to evaluate the genetic similarity between
the founders of the oldest Polish lines, the ancestors of which are unknown,
and the founders purchased in the Near East and admitted as purebred Arabian
mares.

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2. MATERIAL AND METHODS

The material analysed comprised peripheral blood samples from 44 randomly chosen representatives of 15 dam lines and their most important
branches in Poland (Tab. I). The samples came from the collection for routine
testing of parentage controls carried out at the Institute of Genetics and Animal
Breeding in Jastrzebiec. In 2002, these lines were represented by 98.4% of the
total 722 registered brood mares [14]. In addition to the lines analysed in the
current study, single representatives of dierent dam lines imported in recent
years are also present in Polish studs.
The samples were stored at 20 C. Total DNA was extracted using the
Genomic Mini kit (A&A Biotechnology) according to the manufacturers
guidelines. The mtDNA D-loop hypervariable region was amplified using
the following: forward primer 5 -AACGTTTCCTCCCAAGGACT-3 [4] and
reverse primer IRD 800 5 -ATGGCCCTGAAGAAAGAACC-3 [9]. These
primers amplify a 458 bp fragment between sites 15 400 and 15 857. Amplification of the mitochondrial D-loop region was carried out in a total volume
of 20 L polymerase chain reaction mixture with 1.6 L dNTP (10 mM),
2 L 10 X PCR buer for Taq DNA Polymerase (Invitrogen), 1 L MgCl2
(50 mM), 0.6 L 5 M of each primer, about 50 ng of template and 0.2 L
Taq DNA Polymerase (5U L1 , Invitrogen). PCR conditions were as follows: 5 min at 94 C; 35 cycles of the following: 1 min at 94 C, 45 s at
58.5 C, 1 min at 72 C; final extension at 72 C for 10 min. The amplified
DNA was electrophoresed in 1% agarose gel containing ethidium bromide to
check fragment length. PCR products were sequenced directly in both directions, using the same primers then for amplification. To check the quality of
sequencing, BLAST [1] searches were performed. The sequences were aligned
using BioEdit [8] and the GenBank X79547 reference sequence [21] was included. Finally, the haplotypes of Polish Arabs were compared to those of
Arabian horses in the USA [4] based on the equine mtDNA control region
between 15 427 and 15 740 bp.

3. RESULTS
The analyses of a 458 bp mtDNA D-loop fragment in representatives of
15 Polish Arabian dam lines identified 14 distinct haplotypes. They dier from
the reference sequence (GenBank X79547) by four to ten sites, and altogether
by 32 sites as base substitutions (all transitions; Tab. I). Dierences between
the haplotypes ranged from one to 14 with an average of 9.07 nucleotides.

Table II. Variable site positions in the mtDNA control region for 14 Polish Arabian horse haplotypes defined by sequence analysis.
Dots indicate concordance with the X79547 GenBank sequence; the DNA sequences of all haplotypes are available in GenBank under
Acc. No. EF 437553-EF 437566.

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Eight haplotypes were each represented in a single dam line. All Babolnas
dam lines, Szamrajwka, Selma, and two of four of Sawutas lines
(Szweykowska and Wooszka) were concerned. Two haplotypes were each
shared by representatives of two lines. The G haplotype was found in the
Ukrainka and Mlecha lines, and the K haplotype was registered in the Rodania and Cherifa lines. More than one haplotype was found in two lines. The
D and E haplotypes, diering from each other by 13 nucleotides, were found
in the Milordka line. The D haplotype was identified in the branch established
by Zaira, a daughter of Milordka. The E haplotype was found in the branch
comprised of approximately 28% of the representatives of the line [14] and
founded by Milordkas second daughter, Delfina, whose origin has been questioned by Kwiatkowski [11].
Two haplotypes were also found in the Gazella line. The F haplotype was
identified in the branch continued by Gazella II (Tab. I). The second haplotype,
which is identical to the H haplotype of the Sahara line, was found in the second branch continued by Fryga. This branch is represented by approximately
35% of the mares from the line.
Figure 1 shows the relationships between the haplotypes of Polish and
American lines [4]. The analysis yielded four groups of lines with dierences not greater than four nucleotides. The largest group includes 16 lines,
of which are those founded by Polish mares of unknown origin: Wooszka,
Szweykowska, and Milordkas daughter Zaira. The group of the highest similarity proved to be that of Adjuze, Mlecha, Ukrainka, Szamrajwka and six
foreign lines, with dierences from one to three nucleotides.
Haplotypes identical to those found in American Arabian horses analysed
by Bowling et al. were present in eight lines (Fig. 1). Particular attention was
given to the genetic identity between the line founded by Szamrajwka, a mare
of unknown origin born about 1810, and the line of Balkis, a desert-bred (d.b.)
mare imported to France in 1880. The genetic identity between the largest
Polish line of Gazella d.b. and the line of Murana I d.b., imported in 1816 to
the Weil stud (Wrtenberg, Germany) is also worth noting.

4. DISCUSSION
The current results were inconsistent with pedigree data recorded in the Genealogical Charts by Skorkowski [18] in two cases. A detailed analysis of historical sources was performed to explain these discrepancies.
In the case of the Milordka line, the results indicate a possible error in the
record of the line, which was suggested by Kwiatkowski [11]. Haplotype D

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Figure 1. Relationship among Arabian dam lines based on sequence data of the
15 42715 740 bp mtDNA region. The number of nucleotide substitutions is given;
respective values in brackets correspond to the 15 40015 857 bp fragment.

was found in the branch established by Milordkas daughter Zaira 1826

(Tab. I). This haplotype was identical (within the region analysed by Bowling
et al. [4]) to the A25 haplotype found in representatives of the Milordka line
in the USA. A second haplotype (E) was found in the horses from the branch
that originated from Milordkas second daughter, Delfina 1825. The dierence

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between the haplotypes is significant and refers to 13 sites, which excludes

the common origin of both branches. Considering the accuracy in stud registration and the clear system of foal marking at the Sawuta stud [2], errors
regarding the identity of some mares from the line are rather improbable.
Errors in reading the hand-written data of the studbook, however, cannot be
excluded. In some of the documents from the Sawuta stud, copies of which
are in Kwiatkowskis possession, the mare identified by Skorkowski [18] as
Milordka daughter Delfina, is registered as Delfinka 1825, the daughter of
Malikarda 1810. The last record is described in the documents with the annotation z dawnego. This term can be translated as from the old (stud) and
can be interpreted to be synonymous with the designation of Malikarda as a
founder mare (Kwiatkowski, personal communication). The current findings
provide strong evidence for the validity of the hypothesis that there are two
independent lines in the present Milordka line.
Two haplotypes were also found in the Gazella line. The F haplotype was
found in the branch continued by Gazella II born in 1914 in the Jezupol stud
and her four daughters. The haplotype is identical, within the region compared,
to the A19 haplotype found in the American representatives of the line [4]. The
second haplotype (H) was identified in the branch continued by Fryga born
in 1914 in the Pekinie stud and her two daughters. This haplotype is identical to the haplotype found in the Sahara line in both Polish and American
(A04 haplotype) horses. The presence of the H haplotype in the Fryga branch
undermines the ocial pedigree data. The analysis of historical sources indicates some possible reasons of the error. Firstly, a proper stud register was not
maintained in the Jarczowce stud [16], which might have led to the erroneous
reconstruction of the Fryga pedigree. The faulty identification of one of the
mares ancestors is equally probable; after the death of the first owner in 1884,
the horses from the Jarczowce stud were divided between his heirs into five
studs, including Pekinie and Jezupol [2]. However, an error in the identification of Fryga during World War I appears to be most probable. The Russian
army confiscated almost all the horses from the Pekinie stud in 1914, and only
a few foals were left in the stud, together with Fryga [6]. Without stud data it
is impossible to determine precisely the identity of the mare. Nevertheless, on
the basis of information from three sources [3, 5, 12], it can be speculated that
Fryga was actually a descendant of Saharas great-granddaughter Moodycia,
whose daughter and two granddaughters were in the stud in 1906.
The pedigree errors found in the Milordka and Gazella lines concern 11% of
the brood mares registered in 2002 in PASB. This number is comparable to the
results of the Lipizzan horses analysis, where discrepancies between pedigree

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data and mtDNA haplotypes were found in 18 of the 56 lines analysed, which
also covered about 11% of the population [10].
The occurrence of the same haplotype in two lines is another issue worth
consideration. The K haplotype was found in the Rodania and Cherifa lines.
There is no reason to question the pedigree data of both lines. The occurrence
of this haplotype in dierent lines was also confirmed by a study of American
Arabs [4]. Haplotype A1, the 397 nucleotide-long fragment of the K haplotype, was noted in the Rodania, Wadduda, Urfah, and Noura lines. The same
haplotype occurring in five lines may be recognised as evidence of their common origin. The existence of similar haplotypes in dierent dam lines has also
been demonstrated in Lipizzan [10] and Thoroughbred [9] horses.
A common haplotype (G) was also found in the Mlecha and Ukrainka lines.
The accuracy of the pedigree data of the Mlecha line is beyond all question.
The main branches of the line were established by two of Mlechas daughters
(Tab. I). The G haplotype is identical to the A13 haplotype (within the region
analysed) found in the American descendants of Mlecha [4]. The Ukrainka
line was bred in Sawuta and then in the Gumniska (from the fifth generation)
studs until 1944. There were no representatives of the Mlecha line in these
studs [14, 15], which precludes potential errors in horse identification during
stud activity. There is also no convincing evidence to suggest that the only continuator of the line (Forta 1943) was misidentified during wartime ([13, 14, 20],
Pankiewicz, personal communication). Furthermore, it should be pointed out
that identical haplotypes (within the region compared) were also found in
American representatives of the Dahma line [4].
The presence of identical haplotypes in lines established by mares recognised as desert-bred Arabs and in those of unknown origin may be admitted as
evidence of the Arabian origin of the latter. Dierent opinions on the ancestry
of foundation stocks in the oldest Polish studs are presented in the historical sources. According to Skorkowski [17], the stocks were comprised of local breeding Tarpan-type mares (so-called Polish mares) that were repeatedly
mated by oriental stallions. Bojanowski and Tarnowski [2,19], in turn, reported
that the foundation stock of the Biaa Cerkiew stud in 1778 was composed of
Polish, Persian, and Cherkesian mares. The results of the current study shed
new light on the origin of the ancestors of the oldest Polish Arabian dam lines.
This issue will be considered fully in the authors next study.
It should be emphasised that any size dierence in the mtDNA region analysed may influence the accuracy of the evaluation of genetic similarity. As
shown in Figure 1, there are dierences in the number of nucleotides that differentiate particular lines depending on the length of the mtDNA control region

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analysed. For example, the Szweykowska and Wooszka haplotypes dier by

three (397 bp region) or seven sites (458 bp region), while the Szweykowska
and Scherife haplotypes vary by three or eight sites, respectively.
The current analysis of Polish Arabian dam lines, similarly to those by
Hill et al. [9] and Kavar et al. [10], demonstrates the usefulness of mtDNA
analysis in the verification of historical pedigree records. As Hill et al. justly
point out, the findings of such analyses may be of great importance for horse
breeders since their decisions often depend on pedigree information.

ACKNOWLEDGEMENTS
This paper is dedicated to memory of Dr. Edward Skorkowski (18991985),
the preeminent authority on Polish Arabian horse pedigrees, the first Secretary
of the Arab Horse Breeding Society of Poland, the first editor of Polish Arabian
Stud Book, and the compiler of Genealogical Charts of Polish Arabian horses.

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