A Genetic Make Up of Italian Lipizzan Horse Through Uniparental Markers to Preserve Historical Pedigrees

Abstract

Lipizzan is a famous horse breed dating back to 1580 when the original stud of Lipica was established by the Hasburg Archduke Charles II. Currently, the Italian State Stud of Lipizzan Horses (ASCAL) is a conservation nucleus managed through strict mating rules where mitochondrial DNA sequences are used to verify the correct assignment of mares to a historical pedigree maternal lineage. Here, we analyzed the D-loop sequences of Lipizzan horses from the ASCAL in Monterotondo (Rome, Italy) in order to confirm their pedigree assignment to known female founder families. The concurrent investigation of the paternal counterpart based on variation in the male-specific region of the Y chromosome (MSY) showed a prevalence of HT02 (80%), typical of the Neapolitan/Oriental wave. The mtDNA polymorphisms identified shaped nine haplotypes that were unequivocally assigned to each of nine classical mare families of the stud (Africa, Almerina, Argentina, Deflorata, Djebrin, Fistula, Ivanka, Sardinia, Spadiglia), while the Europa and Theodorosta families shared a tenth haplotype. New polymorphisms were identified in a not previously studied region (np 16100-16350). The mtDNA phylogenetic analysis revealed that the Lipizzans of the Monterotondo stud belong to six haplogroups (B, C, G, L, M, Q), out of the 18 recorded for the equine species. This work enabled us to identify and preserve ten haplotypes from the historical maternal lines in a small stud kept in genetic segregation for over 100 years.

Keywords: Lipizzan breed; MSY; haplogroup; haplotype; maternal lines; mtDNA; pedigree.

Figure 1

Polymorphisms among the ten family haplotypes within the mtDNA control region in comparison with the sequence JN398377 for Equus caballus. The dot indicates correspondence with the reference sequence; “nd” indicates that the sequence for that position is not available.

Figure 2

Phylogenetic relationships among the haplotypes/families present in the Monterotondo stud in 2023. Branches corresponding to partitions reproduced in less than 50% of bootstrap replicates are collapsed. Next to the branches are shown the percentages of replicate trees in which the associated taxa clustered together in the 1000 bootstrap test.

Figure 3

Frequency distribution of ten Lipizzan haplotypes within each mtDNA haplogroup.

Figure 4

The median-joining network based on the control-region sequences of all Lipizzan horses from the ASCAL (N = 157, highlighted in yellow) and GenBank (N = 90, black). * indicates a haplotype identical to equine sequence X79547. Each single-nucleotide polymorphism (SNP) indicates a single mutational event; each circle is proportional to the number of samples carrying the same haplotype, and haplogroups are indicated with capital letters.

Figure 5

MSY haplotype frequencies of Lipizzan stallions from ASCAL in Monterotondo (Italy).