The Diagnosis of Herpesvirus Infection in Tortoises by Pathology, Transmission Electron Microscopy and Polymerase Chain Reaction

Between January in 1999 and April in 2005, a total of 56 necropsies of tortoises with clinical signs of oral and nasal discharge, anorexia, lethargy, and palpebral edema and 144 oral swabs were collected. On gross and microscopic examination, 18 of 56 necropsies (32.1%) showed varying degrees of necrotic lesions distributed in the oral cavity and gastrointestinal tract. Three of 18 necropsies (16.6%) with digestive lesions showed marked necrotizing gastroenteritis associated with intralesional eosinophilic intranuclear inclusion bodies (INIB), in which 1 case in the stomach and 2 cases in the small intestine were observed in Geochelone carbonaria, Geochelone sulcata, and Testudo horsfieldi. Electron microscopic examination of INIB in the oral tissue and stomach showed numerous herpesviral particles about 100 nm in diameter in the nuclei and cytoplasm of mucosal epithelium. The viral particles contained envelope, nucleocapsids, and an electron-dense body. Nested polymerase chain reaction analysis using a herpesvirus consensus primer method confirmed the presence of herpesvirus DNA in three oral swabs and stomach. Based on the clinical signs, and the results of pathological examination, electron microscopy and nest polymerase chain reaction, herpesvirus infection was diagnosed. This is the first report to verify herpesvirus infection of tortoises in Taiwan, which induced digestive lesions and caused fatal outcome in some cases.