West Nile virus (WNV) is an enveloped, positive sense, single-stranded RNA virus in the Flaviviridae family. First identified in Africa in the 1930s, WNV has since gained endemicity on all continents except for Antarctica. Maintained in nature in bird-mosquito-bird transmission cycles, WNV can infect dead-end vertebrate hosts including horses and humans to produce severe, debilitating neuroinvasive disease. Although equine infections are preventable due to the availability of effective commercial vaccines, clinical disease in domestic horses is common worldwide. Occasional epizootics are also seen in atypical vertebrate species, such as farmed alligators. WNV presents diagnostic challenges, due to significant serological cross-reactivity with other flaviviruses and inconsistent availability of viral particles and viral genetic material in clinical samples for antigenic detection. With increasing viral genetic diversity, changing anthropogenic environmental factors favoring viral spread, a lack of preventive options in avian reservoir species, and no commercially available treatments, drugs, or therapeutics, WNV remains one of the most important differentials for viral neurologic disease in the domestic equine and other vertebrate animals.