Validation of high-resolution melting analysis as a diagnostic tool for endothelin receptor B mutation in American Paint horses and allele frequency estimation

Overo lethal white foal syndrome (OLWFS) is a genetic disorder caused by a dinucleotide mutation in the endothelin receptor type B (EDNRB) gene leading to the death of affected foals shortly after birth. The use of rapid and reliable genetic testing is imperative for the early diagnosis of the mutation avoiding, therefore, either additional suffering or the production of affected animals. In the present study, we developed and validated a high-resolution melting (HRM) genotyping assay to detect the OLWFS causative mutation, and we also determined the frequency of heterozygotes among American Paint horses in Brazil. The HRM genotyping assay resulted in a high sensitivity, specificity, and positive and negative predictive values. The overall estimated frequency of heterozygotes was 21.6%; however, this frequency increased to 89.5% when considering only overo horses. The HRM assay optimized here was a reliable and suitable method for the detection of the dinucleotide mutation observed in the EDNRB gene resulting in a fast, accurate, and precise diagnostic tool. The causative gene mutation of OLWFS is present in heterozygosity in the American Paint Horse population in Brazil and is highly frequent among overo horses. •Optimization of a diagnostic tool to detect the causative mutation of Overo Lethal White Foal Syndrome.•Twenty-one percent of all horses evaluated were considered carriers.•High frequency of carriers in overo white spotting phenotypes.