The Kushum is a relatively new breed of horses in Kazakhstan that was established in the middle of the 20th century through a cross between mares of Kazakhstan local horses and stallions of Thoroughbred, Trotter, and Russian Don breeds to supply military horses. To reveal the genetic characteristics of this breed, we investigated haplotypes of mitochondrial DNA (mtDNA) and single-nucleotide polymorphisms of the Y chromosome, as well as genotypes of five functional genes associated with coat color, body composition, and locomotion traits. We detected 10 mtDNA haplotypes that fell into 8 of the 17 major haplogroups of horse mtDNA, indicating a unique haplotype composition with high genetic diversity. We also found two Y-chromosomal haplotypes in Kushum horses, which likely originated from Trotter and/or Don breeds. The findings regarding the mtDNA and Y-chromosomal haplotypes are concordant with the documented maternal and paternal origins of the Kushum horses. The allele frequencies of ASIP, MC1R, and MATP associated with coat color were consistent with the coat color variations of Kushum horses. The allele frequencies of MSTN associated with endurance performance and those of DMRT3 associated with gait suggested that the observed allele frequencies of these genes were the result of selective breeding for these traits. As a result of this study, we were able to obtain useful information for a better understanding of the origin and breeding history of the Kushum horse breed using molecular markers.

Horses in Japan have historically been important domestic animals for agricultural power and transportation and many native horses have been raised in Japan. They also played an important role in military affairs. However, from the Meiji Era, many Japanese native horses were crossed with modern Western breeds to increase their body size and physical performance in order to meet military demands. Furthermore, Japanese native horse populations markedly decreased as their roles were replaced by artificial machines, including automobiles. As a result, currently, only eight local populations of Japanese native horses, namely, Hokkaido, Kiso, Noma, Taishu, Misaki, Tokara, Miyako, and Yonaguni horses, remain for mainly conservation purposes in several locations in Japan (Hayashida, 1958; Ichikawa, 1984; Nozawa, 1992). However, since these horse populations may retain the genetic properties of ancestral Japanese horses that are considered to originate from con