Case Study: Polymelia in a Holstein calf

Polymelia, the manifestation of extra limbs or parts of limbs, occurs when there is a developmental duplication in the novel NHL repeat domain containing protein (NHLRC2) gene in Angus cattle. In Holstein cattle, little is known about the etiology of polymelia. To determine the genetic etiology of polymelia in a Holstein calf, genotyping and sequencing of the Angus developmental duplication locus, karyotyping, and a genome-wide association analysis were performed. The genome-wide association analysis compared the polymelia calf (case) to 2,650 control (normal) Holstein calves using EMMAX-GRM (Efficient Mixed-Model Association eXpedited-Genomic Relationship Matrix) software. In addition, homozygous loci within genes in the case that were never found in a control were also evaluated as putative loci. The polymelia calf had a normal Angus developmental duplication genotype that was confirmed by sequencing. Karyotyping revealed no evidence of chromosomal breakage or gross chromosomal abnormalities in the polymelia calf. The additive genome-wide association analysis identified loci associated with polymelia on BTA13 (P < 1 × 10−281) and BTA10 (P < 2 × 10−110). Seventy-nine candidate genes were identified that contained homozygous genotypes in the case that were never identified in 2,650 controls. Candidate genes include MSH homeobox 2 (MSX2) on BTA20, which has been associated with limb development in mice, and GINS complex subunit 1 (GINS1) on BTA13, which is involved in early embryogenesis in mice. In summary, these results suggest that the etiology of polymelia differs by cattle breed and is the result of genetic variants at more than one locus.