Equine influenza virus (EIV) is a major and highly contagious respiratory virus responsible for outbreaks of equine influenza (EI) worldwide. Nowadays, only EIV of the Clade I and Clade II Florida sub lineage are circulating amongst equids. While the virus continuously evolves and alters its morphology to escape the immune system of the host, it is very important to determine how the structure of the proteins responsible for he induction of a solid immunity change over times. This information is relevant to best evaluate vaccine effectiveness and elaborate recommendations to ensure that contemporary vaccines contain epidemiologically relevant strains of EIV. The objective of this study is to describe the temporal and spatial molecular characteristics of H3N8 EIV isolates collected from horses in the USA from 2006 to 2016 (10 year period).