Equid herpesvirus 9 (EHV-9) was isolated from a herd of Thomson's gazelles affected by encephalitis. The natural host of EHV-9 is unknown, but zebras are suspected to be the source of infection in gazelles. To prove this hypothesis, we analyzed 43 sera from Burchell's zebras (Equus burchelli) and 21 Thomson's gazelles (Gazella thomsoni) from the Serengeti ecosystem for neutralizing antibodies. Seven zebra sera were positive for EHV-1, EHV-9 and EHV-1 from Grevy's zebra strains T965 and T616. The trigeminal ganglia of 17 other Burchell's zebras and one Thomson's gazelle were tested by EHV-9 gB and EHV-1 ICP0-specific nested polymerase chain reaction (PCR). PCR sequencing confirmed that one zebra ganglion was positive for EHV-9. These results suggest that the Burchell's zebras were exposed to EHV-9 and latently infected.