Y-chromosomal insights into the breeding history and sire line genealogies of two traditional Baroque horse breeds: Lipizzaner and Kladruber.

The paternally inherited, male-specific part of the Y chromosome (MSY) is an ideal marker for studying the origin, genealogies, and historical connections of horse patrilines. Here, we performed fine-scaled MSY haplotype (HT) analysis in two Baroque horse breeds, the Lipizzaner and the Kladruber, both known for their long-standing tradition of sire line breeding and interconnected genealogies. We genotyped 95 MSY markers using KASP technology in 90 stallions representing all patrilines of both breeds. We identified 14 HTs across eight Lipizzaner sire lines and six HTs in eight Kladruber lines. Y-chromosomal analysis confirmed the presumed Arabian and Iberian origins of two Lipizzaner and two Kladruber foundation sires. Interestingly, six Lipizzaner sire lines clustered into the MSY haplogroup (HG) daC_Tb, a HG previously associated with the Turkoman horse, suggesting a Turkish origin of several Lipizzaner foundation sires, contrary to documented records. We also found evidence for an undocumented Arabian founder in the Lipizzaner breed. Furthermore, three private HTs were detected in Kladruber horses, highlighting the uniqueness of their foundation sires. HTs in shared patrilines between Lipizzaner and Kladruber were consistent, while three Lipizzaner sublines showed discrepancies between MSY results and recorded pedigree data. In conclusion, MSY haplotyping validated historical breed documentation, revealed new insights into the origins and distribution of sire lines, and proved effective in resolving parentage issues across generations.