Iris yellow spot virus (IYSV), an emerging disease of onion crops, was identified by transmission electron microscopy, enzyme linked immunosorbent assay, and reverse transcription polymerase chain reaction in Costa Rica. Onion plants had straw- colored, elongated lesions and tip dieback. Costa Rican IYSV nucleocapsid partial sequences (15 isolates) grouped with isolates from North and Central America and from New Zealand.