Search

In this study the hematological and biochemical parameters in the blood of indigenous Croatian working horse breeds were investigated. The Posavina and Croatian Coldblood horses are adapted to harsh environmental conditions and their blood parameters might differ from other horse breeds. The study was carried out on 100 mares and 12 stallions of ages from 2 to 19 years. Fifteen hematological and 19 biochemical parameters were analysed. Values of 22 parameters showed considerable overlapping with values obtained for other horse breeds, and substantial resemblance is evident with values reported for Pakistani working horses. Several reference values showed statistically significant effect of sex (Hb, ALB, CREA, TBIL and Ca where significantly higher in stallions; EOS, CK, ALP, UREA and Mg were significantly higher in mares). None of the parameters studied showed any differences associated with age. The adaptation of Posavina and Croatian Coldblood horses to the harsh environment of flooded pastures and the way of breeding might be reflected in their specific reference values. The obtained results enabled the establishment of hematological and biochemical reference values in the blood of Posavina and Croatian Coldblood horses that will in turn support clinical diagnosis and further research into horse physiology. U studiji je obavljeno ispitivanje hematoloških i biohemijskih parametara krvi kod radnih hrvatskih autohtonih rasa konja. Posavski i hrvatski hladnokrvnjak su dve rase konja koje su adaptirane na oštre uslove spoljašnje sredine pa se pretpostavlja da njihovi parametri krvi mogu da budu različiti u poređenju sa drugim rasama. Ispitivanje je obavljeno na 100 kobila i 12 pastuva starosti od 2 do 19 godina. Obavljena je analiza u odnosu na pedeset hematoloških i 19 biohemijskih parametara. Vrednosti koje su dobijene u odnosu na 22 parametra pokazale su značajna preklapanja u poređenju sa drugim rasama konja, pri čemu je ustanovljena značajna podudarnost sa vrednostima koje su karakteristične za pakistanskog radnog konja. Nekoliko referentnih vrednosti je pokazalo statistički značajnan utacaj pola (Hb, ALB, CREA, TBIL i Ca vrednosti su bile značajno veće kod pastuva; EOS, CK, ALP, UREA i Mg vrednosti su bile značajno veće kod kobila). Nijedan parametar koji je ispitivan nije pokazao bilo kakvu varijaciju u odnosu na starost životinja. Prilagođenost posavskog i hrvatskog hladnokrvnog konja na oštre i teške uslove spoljašnje sredine koji podrazumevaju ispašu na poplavljenim pašnjacima, kao i način uzgoja, može da ima posledice na specifične referentne vrednosti ispitivanih parametara. Dobijeni podaci omogućavaju defi nisanje hematoloških i biohemijskih referentnih vrednosti krvi posavske i hrvatske rase hladnokrvnih konja, a te vrednosti bi omogućavale kliničku dijagnostiku i buduća ispitivanja fi ziologije konja.

The Posavina and Croatian Coldblood are Croatian autochthonous horse breeds with interwoven breeding histories for which studbooks have only recently been established. The Lipizzan breed has the oldest formalized breeding and no record of recent genetic introgression from other breeds in Croatia. We analyzed the genetic structure, interbreeding, and breed characteristics by genotyping nine dinucleotide microsatel - lite loci for 53 Posavina, 37 Croatian Coldblood, and 33 Lipizzan horses and showed that differing breeding schemes and histories have had a strong and measurable impact on the population genetic structure within and between the three breeds. A Bayesian clustering method demonstrated that two population clusters best explain the genetic structure. Samples from the pre-defined breeds of the Posavina and Croatian Coldblood were assigned to a separate genetic cluster, while Lipizzan specimens were assigned to another distinct genetic group. Twelve samples of the Posavina/Croatian Coldblood cluster (13%) showed admixed ancestry with Lipiz - zan horses. A test for heterozygosity excess, allele frequency distribution mode-shift, and M-ratio test were used to detect genetic evidence of recent population bottlenecks, none of which provided evidence for bot - tlenecks in the Posavina and Croatian Coldblood populations. In contrast, although somewhat ambiguous, evidence suggests a genetic bottleneck in the Lipizzan population in Croatia.

Horse breeding in the Republic of Croatia, before its entrance into European Union, is in the transitional period. In the breeding sense, Croatian horse breeding is set according to the model of European countries and is trying to counteract with “modern horse breeding countries”. Croatian Center for Horse Breeding - Đakovo and Lipik State Stud Farms (CCHB), with the Central register of equidae is, as the head national institution, responsible for running and maintaining national central equidae database (49 registers of horses and 27 registers of breeding types). Croatia has registered total of 21.796 equidae, from which 19.306 animals are horses. Registered horses according to groups make the total proportion of 37% warmblood horses, 59% coldblood horses and 4% ponies. Horse breeders are organized in breeders associations, which form federations of associations referring to separate horse breeds. Currently in Croatia there are four federations of breeders associations (with 43 associations and 11 private stud farms). Those verified breeding organizations are conducting independently breeding programs and fully maintain breeding and selection obligations for horse breeds for which they have approval of the Croatian Ministry. Native breeds in Croatia are Croatian Coldblood, Croatian Posavac and Međimurje horse (Murinsulaner), while Lipizzan horse is considered as protected breed. Development of horse breeding is considerably supported with national subsidies. Internationally verified breeds which are bred in Croatia are Lipizzan horse, Pure Arabian horses, Thoroughbred, Haflinger and Gidran horses. The horse breeding in Croatia is developing in four different directions. Ecological breeding implies native horse breeds and is spread on the areas of protected nature resorts in Croatia. Croatians are through their history connected to horse breeding, which is even today irreplaceable in traditional cultural manifestations (Đakovacki vezovi, Vinkovacke jeseni, Sinjska alka, etc.), various horse shows and fairs and also in more “modern” ways through recreation and entertainment. The third direction of development of horse breeding in Croatia is therapeutic riding, which includes 26 associations, over one hundred horses and over one thousand users organized under the Therapeutic Riding Association of Croatia. Sports horse breeding represents fourth direction in Croatian horse breeding. It is formed by four federations: Croatian Trotting Federation (trotting), Croatian Gallop Federation (gallop sport) and Croatian Equestrian Federation (endurance, dressage, military, show jumping, driving). Croatian Equestrian Federation includes 49 equestrian clubs. In the last four years between 455 and 582 competitors and between 495 and 581 horses (for show jumping 45%, dressage 31%, endurance 19%, driving 5% and military) were licensed annually. Croatia achieved noticeable results on the international scene in driving sports and in Paralympics. In all mentioned directions of Croatian horse breeding, the tendency should be directed to achieving higher breeding standards (nutrition, stabling, transport, training) and more rational using of horses. Such breeding and using of horses will lead to top bred horses and successful sport horses. This implies synchronized work through long period of time, respecting institutional and breeding rules.

The genetic variability for a sample of 107 animals from three autochthonous Croatian horse breeds was estimated using 20 microsatellites. The average number of alleles per locus (6.3) and proportion of heterozygosity (0.732) indicated a moderate variability. The expected heterozygosity was similar among all breeds and ranged between 0.724 in the Posavina horse, and 0.737 in the Croatian Coldblood and Murinsulaner horse. The inbreeding coefficient FIS was low and non-significant over the three populations. The genetic differentiation among the three populations was low (FST=0.026), suggesting that only 2.6% of the total genetic variability was due to differences between the breeds, and 97% to individual differences. The results of pairwise genetic differentiation suggest that the Posavina horse and the Croatian Coldblood were the most closely related populations (FST=0.016). These results are confirmed by Nei’s genetic distances with the highest value observed between the Posavina horse and the Murinsulaner (0.082) and the lowest between the Posavina horse and the Croatian Coldblood (0.044). An assignment test correctly assigned 82% of individuals to the correct breed. Strategies for preserving the original native genes in the Croatian native horse breeds should be considered in order to prevent these breeds from becoming extinct and include them in the future breeding programmes.

Genetic variation in three Croatian coldblood horse populations was analysed using a sequence analysis of the proximal part (nt 15 498-15 821) of the D-loop region of mtDNA. Twenty unrelated horses were chosen from the Posavina horse and the Croatian Coldblood breeds and fifteen horses from the Murinsulaner horse population. Sequencing of the proximal part of the mtDNA D-loop region revealed 26 polymorphic sites representing thirty haplotypes which were clustered into eight haplogroups. A wide variety of mitochondrial haplotypes of the analysed horse breeds were clustered into eight different haplogroups. Two haplogroups (D and F) were specific to Posavina horse, five haplogroups were shared among the Croatian Coldblood and Murinsulaner horse, the fact that can be explained by selection strategy at the beginning of the 20 th century and possible gene flow between the two populations. These results indicate the presence of many ancient maternal lineages with high diversity in mtDNA. The genetic information based on mtDNA typing has a great importance for the future breed conservation strategy, especially for the critically endangered breed such as Murinsulaner horse.

The global biodiversity crisis extends to autochthonous local breeds of livestock. There is an increasing danger that these rare breeds become extinct and with them their locally adapted gene pool. Modern molecular tools such as parentage testing using microsatellite genotyping are powerful in guiding management and conservation. We tested nine microsatellite markers in three Croatian horse breeds and obtained high exclusion probabilities (EPs) for the most common test scenario ‘one parent and offspring known and the other parent tested’ (99.9% in Posavina and Croatian Coldblood and 99.3% in Lipizzaner), despite that Lipizzaner has an overall lower genetic variability at microsatellite loci. To become a useful tool in breed management in countries with developing economies, genetic screening systems must be designed to be statistically powerful yet economically viable. Therefore, a suite of six markers that can be run in two multiplex systems and which still gives high exclusion probabilities (99.5% in Posavina and Croatian Coldblood and 98% in Lipizzaner) was chosen.

Domestic equidae display polymorphism within ELA-DRA locus which is not characteristic for other species. We characterised sequence polymorphism present at ELADRA locus exon 2 and estimated allele frequencies in two autochthonous breeds, Posavina and Croatian Coldblood. In 88 horses, four different alleles were found, one of them not reported before in horses. The new allele shows non-synonymous mutation at position 65 (T→A) causing amino acid change (Phe→Tyr) in antigen binding site and synonymous mutation at position 105 (C→T). Our findings emphasize the importance of DRA polymorphism among equids and some specific DRA frequency pattern potentially specific in draught horses.

Prosječna veličina progenih grupa umatičenih kobila prema pastusima očevima iznosila je 6,27 s rasponom varijacije od 1 do 34 kobile kćerke. Kao očevi umatičenih kobila pojavljuju se 352 pastuha, od kojih je 165 umatičenih (6 %), a 197 pastuha očeva poznato je samo po imenu (7 %). Bez poznatog porijekla po očevoj strani umatičeno je 2277 kobila rodonačelnica (87 %). Istraživanjem zastupljenosti boje dlake u pasmini hrvatski hladnokrvnjak utvrđeno je pojavljivanje 21 boje, a očekivano dorata boja prevladavala je u svim analizama., The average size of progenies groups of registered mares according to fathering sire stallion was 6,27 with a span of variation from 1 to 34 daughter mares. A number of 352 sire stallions appear as fathers of registered mares, from which 165 are registered (6%) and 197 fathering sire stallions are known only by their name (7%). There are 2277 founding mares (87%) registered without any known origin according to the sire line. After researching the hair color representation with the Croatian Coldblood it has been found that 21 colors appear, and expectedly the bay color is predominant in all analysis.

Hrvatski hladnokrvnjak je izvorna pasmina konja, a uzgojno područje je gotovo cjelokupan prostor Republike Hrvatske. U cilju dobivanja više podataka o uzgoju kako bi se uzgojni program u cilju genetskog unaprijeđenja i očuvanja kao izvorne pasmine što uspješnije provodio, u 2006. godini započeta je sistematizacija uzgoja hrvatskog hladnokrvnjaka. U formiranje rodova u obzir su uzeta isključivo umatičena (žigovana) ženska grla. Rezultat sistematizacije je formiranje 2639 rodova kobila s velikom varijacijom u broju kobila prema rodu (1 do 35). Brojno najveći rod kobila s 35 umatičenih ženskih grla je H297, dok je čak 1640 rodova brojilo samo 1 kobilu., The Croatian Coldblood is an authentic horse breed, and the breeding area is almost the entire area of the Republic of Croatia. In order to obtain more information about breeding to implement the breeding program as successfully as possible, to genetically improve and preserve the authentic horse breed, in the year 2006. the systematization of breeding of the Croatian Coldblood was initialized. While forming lines only registered (branded) mares were included. The result of the systematization is the formation of 2639 dam lines with great variation in the number of mares by line (1 to 35). Numerously the largest dam line with 35 registered mares is H297, while even as much as 1640 lines numbered only one mare.

Hrvatski hladnokrvnjak brojčano je najbrojnija izvorna pasmina konja Republike Hrvatske, ali i općenito najbrojnija pasmina konja u Hrvatskoj. Kroz stoljetnu tradiciju uzgoja, hrvatski hladnokrvnjak je postao sastavni dio povijesti, kulturnih i tradicijskih običaja. Danas, kada je izgubio nekadašnju ulogu izvora radne snage u poljoprivredi, postao je pasmina s novim vrijednostima. Pasminu karakterizira specifičan put formiranja u vrlo ekstenzivnoj tehnologiji uz veliki utjecaj stranih hladnokrvnih pasmina te velika sposobnost adaptacije na nepovoljne klimatske prilike i uvjete držanja. Njegova važna uloga danas kao pasmine konja koja se uzgaja u visokom stupnju ekstenzivne (pašnjačke) tehnologije, je u očuvanju biološke raznolikosti zaštićenih krajolika, a kroz to ima veliki potencijal u ekološkoj proizvodnji hrane sukladno trendovima u prehrani ljudi. Kao sredstvo očuvanja i razvoja ruralnih krajeva kroz profitabilnu ekološku proizvodnju, hrvatski hladnokrvnjak ujedno si samostalno osigurava olakotnije očuvanje kao izvorne pasmine konja., The Croatian Coldblood is the most numerous authentic horse breed in the Republic of Croatia, but also the most numerous horse breed in Croatia in general. Through a hundred year old breeding tradition the Croatian Coldblood is an essential part of history, cultural and traditional customs and heritage. Today, when is has lost its old purpose as a source of labor in agriculture, it has become a breed with new values. The breed is characterized by a specific way of formation in very extensive technology with high implication of foreign coldblood breeds, and a great ability to adapt to unfavorable climatic and keeping conditions. His important role today as a horse breed which is grown in a high degree of extensive (pastoral) technology, is in the preservation of biological diversity of protected landscapes, and through this it has great potential in ecologic food production consistent with human feeding trends. As a means of preservation and development of rural regions through profitable eco – production, the Croatian Coldblood is thereby autonomously ensuring itself easier preservation as an authentic horse breed.

All over Europe, the number of draught horses has decreased drastically during the last 50 years. As a prerequisite for efficient management decisions, we analysed the conservation status in Austrian (Noriker Carinthia – NC, Noriker Salzburg – NS), Croatian (Croatian Coldblood – C, Posavina horse – P) and German (Altmaerkisch Coldblood – A, Black Forest horse – BF, Mecklenburg Coldblood – M, Rhenish German Draught horse – R, Saxon Thuringa Coldblood – ST, Schleswig Draught horse – Sch, South German Coldblood – SG) draught horses (434) using multilocus genotypic information from 30 (effectively 27) microsatellite loci. Populations located in areas with less intensive agricultural production (C, NC, NS, P and SG) had greater diversity within the population and estimated effective population size than A, BF, Sch, M, R and ST populations. The PCA plots revealed that populations form five separate groups. The 'Noriker' group (NC, NS and SG) and the 'Rhenish' group (A, M, R and ST) were the most distinctive (pairwise FST values ranged from 0.078 to 0.094). The 'Croatian' group (C and P) was in the centre, while the BF and Sch populations formed two out-groups. A posterior Bayesian analysis detected further differentiation, mainly caused by political and geographical factors. Thus, it was possible to separate the South German Coldblood from the Austrian Noriker population where no subpopulation structure was detected. The admixture analysis revealed imprecise classification between C and P populations. A small but notable separation of R from A, M and ST populations was detected, while Sch and BF populations remained as out-groups. The information obtained should aid in making efficient conservation decisions.