26th Annual Meeting and Symposium of the
Desert Tortoise Council, March 16-18, 2001
Abstracts

Transmission Study with a Tortoise Herpesvirus (THV) In Greek Tortoises

Francesco Origgi, D.V.M.¹, Paul Klein,Ph.D.², Carlos Romero D.V.M.³, and Elliott Jacobson D.V.M.^4
1Departments of Small Animal Clinical Sciences and Pathobiology, College of Veterinary Medicine, University of Florida
²Department of Pathology, College of Medicine, University of Florida
³Department of Pathobiology, College of Veterinary Medicine, University of Florida
⁴Department of Small Animal Clinical Sciences, College of Veterinary Medicine, University of Florida

Two different strains of a pathogenic herpesvirus previously isolated from two captive Herman's tortoises (Testudo hermanni; Marschang et al., 1997) were used in a transmission study in Greek tortoises (Testudo graeca; n=5). Plasma samples were collected before and after a first challenge with 15,000 TCID50 viral suspension (either Intranasally or Intramuscularly). Antibody titers of the tortoises were measured using a new Enzyme-linked Immunosorbent Assay (ELISA; Origgi et al., 2001) and a Serum Neutralization (SN) test (Marschang et al. 1997). Seroconversion was detected 4 to 9 weeks following challenge. Mild clinical signs consistent with those described in the literature for herpesvirus infection of tortoises (Drury et al 1998/1999; Heldstab and Bestetti, 1989; Jacobson et al.,1985; Kabish and Frost 1994; Marschang et al., 1997/1998/1999; Muro et al., 1998) were observed in challenged tortoises. At 11 months following the first challenge, the tortoises in this study were challenged for a second time using a viral dose 10 times higher (150,000 TCID50) than the first challenge dose. Severe clinical signs consisting of stomatis and glossitis were observed in the challenged tortoises. Diphtheritic plaques were observed in areas of the oral mucosa overlying salivary glands. The distribution of the lesions was independent of the route of challenge or strain of virus used. Two of the four challenged animals showed only mild clinical signs. The unchallenged (control) tortoise never developed any clinical signs of herpesvirus infection. At one month following the 2nd challenge all tortoises were euthanized and necropsied. Tissue samples from all major systems were collected and processed for histopathology and DNA/RNA isolation. Using polymerase chain reaction (PCR) analysis all 4 challenged tortoises were positive for presence of herpesvirus nucleotide sequences and negative for the control tortoise. The number of positive tissues varied between two (tortoise #1) and 17 (tortoise #3). PCR evaluation of tissues indicated a specific tropism of the herpesvirus for the central nervous system (CNS), where half of the total number of positive samples was detected. This represents the first transmission study attempted with tortoise herpesvirus in a tortoise and it will serve as a model in investigating the role of herpesvirus in desert tortoises (Gopherus agassizii), a species known to be infected with herpesvirus (Harper et al., 1982, Pettan Brewer et al., 1996)

References

Drury, S. E. N., R.E. Gough, and S. McArthur. 1998. Detection of herpesvirus-like and papillomavirus associated with diseases of tortoises. Vet Rec 143:639

Drury, S. E. N., R.E. Gough, and S. McArthur. 1999. Isolation and identification of herpesvirus and papillomavirus from tortoises in Great Britain. Pages 69-75 in Proceedings of the 6th ARAV Conference, Columbus, Ohio.

Harper P. A. W., D. C. Hammond, and W. P. Heuschele. 1982. A Herpesvirus-like agent associated with a pharyngeal abscess in a desert tortoise. J Wild Dis.18: 491-494

Heldstab, A and G. Bestetti. 1989. Herpesviridae causing glossitis and meningoencephalitis in land tortoises (Testudo hermanni). Herpetopathologia. 1: 5-9

Jacobson, E. R., S. Clubb, and J. G. Gaskin. 1985. Herpesvirus-like infection in Argentine tortoises. J Am Vet Med Assoc 187:1227-1229

Kabish, D. and J. W. Frost. 1994. Isolation of herpesvirus from Testudo hermanni and Agrionemys horsfieldii. Verh Ber Erkrg Zootiere 36: 241- 245

Marschang, R. E., M. Gravendyck, and E. F. Kaleta. 1997. Investigation into virus isolation and the treatment of viral stomatitis in T. hermanni and T. graeca. J Vet Med. Series B. 44: 385-394.

Marschang, R. E., H. Posthaus and M. Gravendyck. 1998. Isolation of Viruses from Land Tortoises in Switzerland. Pages 281-284 in Proceedings of the AAZV and AAWV Joint conference. Omaha, Nebraska.

Marschang, R. E. 1999. Evidence for a new herpesvirus serotype associated with stomatitis in Afghan Tortoises, Testudo horsfieldii. Pages 77-79 in Proceedings of the 6th ARAV Conference, Columbus Ohio.

Muro, J., A. Ramis , J. Pastor , L. Velarde , J. Tarres , S. Lavin. 1998. Chronic rhinitis associated with Herpesviral Infection in captive spur-thighed tortoise from Spain. J Wild Dis. 34:487-495

Origgi F. C., P. A. Klein, K. Mathes, S. Blahak, R. E. Marschang, S. J. Tucker, and E. R. Jacobson. 2001. An Enzyme Linked Immunosorbent Assay (ELISA) for Detecting Herpesvirus Exposure in Mediterranean Tortoises [Spur-Thighed tortoise (Testudo graeca) and Hermann's tortoise (Testudo hermanni)]. J. of Clin Micro (Submitted for publication).

Pettan-Brewer K. C. B., M. E. Drew, E. Ramsey, F. C. Mohr, and L. J. Lowenstine. 1996. Herpesvirus particles associated with oral and respiratory lesions in a California Desert tortoise (Gopherus agassizii). J. Will. Dis. 32: 521-526.

Une Y, K. Uemura, Y. Nakano, J. Kamiie, T. Ishiabashi, and Y. Nomura. 1999. Herpesvirus infection in Tortoises (Malacochersus tornieri and Testudo horsfieldii). Vet Path 36: 624-627.