**1.3 Meningoencephalitis in a polar bear**

A 12-year-old female polar bear (*Ursus maritimus*) developed a sudden onset of neurological symptoms in 2007 (Schrenzel et al., 2008; Donovan et al., 2009). Nonsuppurative pleocytosis of cerebrospinal fluid was observed. The bear was euthanized. Multifocal, random nonsuppurative meningoencephalitis was microscopically found involving most prominently the rostral cerebral cortex, as well as the thalamus, midbrain, and rostral

Virology and Pathology of Encephalitis in Alien Hosts by Neurotropic Equine Herpesvirus 9 131

Fig. 3. Scheme of the EHV-9 genome based on the complete nucleotide sequence (AP010838)

ORF Codons Identity (%) HSV-1 Predicted or confirmed functions 1 202 96 - C-terminal hydrophobic domain 2 204 93 - C-terminal hydrophobic domain

5 470 95 UL54 Post-translational regulator of gene

7 1079 95 UL52 Component of DNA helicase–primase

9 326 94 UL50 Deoxyuridine triphosphatase

6 343 99 UL53 Glycoprotein (gK)

8 245 97 UL51 Tegument protein

10 100 95 UL49A Envelope protein

expression

complex; primase

3 257 93 - 4 200 93 UL55

medulla. Lesions consisted of inflammation, neuronal necrosis, gliosis, and both neuronal and glial basophilic intranuclear inclusion bodies. PCR for the herpesvirus DNA polymerase gene segment was positive on DNA extracted from frozen tissues and from paraffinembedded fixed brain. The nucleotide sequence of the PCR product indicated the presence of EHV-9, which was further confirmed by following PCR for the EHV-9 gB gene segment. Schrenzel et al. (2008) described that EHV-9 had been detected at the same zoological garden in 2 Grevy's zebras (*Equus grevysi*) from the same herd, which had been relocated near the polar bears before the polar bear case. One of the infected Grevy's zebras was 8 days old and had viral interstitial pneumonia; the other was an adult with rhinitis and intranuclear inclusion bodies. Both zebras were immunocompromised as a result of other concurrent conditions.

#### **1.4 Abortion in an onager**

Schrenzel et al. (2008) described that EHV-9 was found by a retrospective analysis of tissues from an aborted Persian onager (*Equus hemionus onager*) fetus from a zoological park in Washington, DC (Montali et al., 1985). The onager fetus was aborted after the dam came in close proximity to a Grevy's zebra. A herpesvirus was isolated from the fetus. The virus was identified as EHV-1 based on DNA fingerprinting and serological analyses (Montali et al., 1985). PCR and DNA sequencing analyses of the DNA polymerase showed that the zebras and the onager had an EHV-9 strain identical to that found in the polar bear (Schrenzel et al., 2008).

#### **1.5 Burchell's zebras from the Serengeti ecosystem**

Zebras have been suspected to be the source of EHV-9 infection. To prove the hypothesis, serological analysis was examined by using 43 sera from Burchell's zebras (*Equus burchelli*) and 21 Thomson's gazelles from the Serengeti cocsystem for neutralizaing antibodies (Borchers et al., 2008). Seven zebra sera were positive for EHV-1 and EHV-9. The trigeminal ganglia of 17 other Burchell's zebras and one Thomson's gazelles were examined by PCR for EHV-9 gB and EHV-1 ICP0 genes. One zebra ganglion was positive for EHV-9 by PCR and confirmed by sequencing. These results suggest that the Burchell's zebras were latently infected by EHV-9.
