A Comprehensive Analysis of the Agreement and Performance of Variant Annotation Programs in Equine Genomes

Background/Objectives: Advances in whole-genome sequencing (WGS) technology have led to the widespread adoption of WGS for investigating genetic diseases and traits in domestic animals. This has created a need for improved methods for prioritizing candidate causal variants. One way variants are prioritized is using variant annotators that predict variant effects based on their proximity to genomic features and effect on amino acid sequence. However, validation of variant annotators for domestic animal genomes is lacking. Methods: In this study, we calculated the agreement of three popular variant annotators, Ensembl Variant Effect Predictor (Ensembl-VEP), SnpEff, and ANNOVAR, across >58 million variants identified in 1065 horse genomes. Results: Comparisons showed that agreement across all three variant annotators was >90% when terminology was standardized. Terminology standardization was the most important factor affecting agreement, as agreement dropped to 0-67% when terminology was not standardized across variant annotators. Genomic context was also a major factor, as exonic, and specifically loss-of-function, variants showed lower agreement rates than intergenic variants. In addition to annotation agreement, differences in computational resource requirements were identified. ANNOVAR required ~28× more memory and ~1.5× more time than the next best tool. Conclusions: These results demonstrate that tool selection for annotating variants should not be based on a single metric; rather, a study's needs and available computational resources should be considered when selecting the appropriate variant annotators(s) along with the standardization of terminology across annotators. These findings are a resource for guiding decisions on the use of variant annotators in domestic animals and suggest areas for improvement in the standardization of variant prioritization.