P ath ology P ages

Oroesophageal Adenovirus-like Infection in a Palm Viper, marchi, with Inclusion Body-like Disease

James T. Raymond1, DVM, MS, DACVP, Michael M. Gamer1, DVM, DACVP, Suzan Murray2, DVM, Robert Nordhausen3, MA

1. Northwest ZooPath, 18210 Waverly Drive, Snohomish, WA 98296, USA Downloaded from http://meridian.allenpress.com/jhms/article-pdf/12/3/30/2207364/1529-9651_12_3_30.pdf by guest on 27 September 2021 2. The Fort Worth Zoo, 1989 Colonial Parkway, Fort Worth, TX 76110, USA

3. California Health and Food Safety Laboratory System, Post Office Box 1770, Davis, CA 95617, USA

A b s t r a c t : An adult, captive-born palm viper, Bothriechis marchi, was found dead in its enclosure. Histologically, there were eosinophilic intracytoplasmic inclusion bodies within epithelial cells in the stomach, intestine, lung, trachea, and esophagus. In addition, there were basophilic intranuclear inclusion bodies in mucosal epithelial cells lining the oral cavity and esophagus with multifocal subacute esophagitis and stomati­ tis. Electron microscopy of the intranuclear inclusion bodies revealed crystalline arrays of hexagonal, 65 - 68 nm, particles morphologically consistent with Adenovirus. This palm viper was diagnosed with inclusion body-like disease, as previously reported in palm vipers, with concurrent esophagitis and stomatitis due to infection with adenovirus-like particles.

K ey W o r d s : palm viper, Bothriechis marchi, adenovirus, inclusion body-like disease.

INTRODUCTION There was transmigration of lymphocytes and plasma cells through the mucosa (Figure 1). The underlying propria-sub- Viral diseases have been commonly reported in collec­ mucosa was infiltrated by lymphocytes and plasma cells tions of , and additional viruses are continuing to be admixed with edema fluid. Several mucosal epithelial cells described. Infections with viruses of the family Adenovirodae contained basophilic intranuclear inclusion bodies (Figure 2). have been previously reported in only a few of snakes The oral mucosa was mildly hyperplastic and had mild multi­ (Heldstab and Bestelli, 1984, Jacobson, et al, 1985, Juhasz and focal degeneration and necrosis of individual epithelial cells. Ahne, 1993, Schumacher, et al, 1994, Wozniak, et al, 2002). Few mucosal epithelial cells contained similar basophilic Recently, an inclusion body-like disease similar to inclusion intranuclear inclusion bodies. Mucosal epithelial cells of the body disease (IBD) of boid snakes was reported in palm vipers, trachea, bronchi, esophagus, stomach, and intestine contained Bothriechis marchi, (Raymond, et al, 2001). This report few eosinophilic intracytoplasmic inclusion bodies. describes a case of oroesophageal adenovirus-like infection in a For transmission electron microscopy, sections of 10% palm viper with concurrent inclusion body-like disease. neutral buffered formalin-fixed esophagus were transferred into modified Karnovsky’s fixative, washed with sodium CASE REPORT cacodylate, and post-fixed in unbuffered 2% osmium tetrox- ide reduced by 2.5% potassium ferrocyanide. Sections of An approximately eight-year-old, captive bred, male palm esophagus were then dehydrated through a graded ethanol viper housed at The Fort Worth Zoo (Fort Worth, TX, USA) series, soaked in propylene oxide, infiltrated with epoxy resin had a history of occasionally holding its mouth open at odd (Spurr’s Low Viscosity Resin, Ted Pella, Redding, CA), sec­ angles. The day prior to death, the was anesthetized to tioned at 70 nm, stained with 6% methanolic uranyl acetate, evaluate the oral cavity, which appeared normal upon visual counterstained with lead citrate, and examined with a trans­ examination. At necropsy, there were no gross lesions. mission electron microscope (EM 10C transmission electron Sections of testis, skeletal muscle, aorta, epididymis, kidney, microscope, Carl Zeiss, Thomwood, NY). Transmission elec­ thyroid gland, esophagus, trachea, liver, oral cavity with teeth, tron microscopy of the intranuclear inclusion bodies in the heart, lung, and stomach were fixed in 10% neutral buffered esophageal mucosa revealed many nonenveloped hexagonal formalin, sectioned at 5.0 pm, and stained with hematoxylin outlined, 65 - 68 nm diameter viral particles with an electron and eosin (HE). dense core (Figure 3). Occasionally, viral particles were Histologically, there was multifocal degeneration and arranged in paracrystalline arrays (Figure 4). necrosis of mucosal epithelial cells lining the esophagus.

30 Journal of Herpetological Medicine and Surgery Volume 12, No. 3,2002 Downloaded from http://meridian.allenpress.com/jhms/article-pdf/12/3/30/2207364/1529-9651_12_3_30.pdf by guest on 27 September 2021

Figure 1. Esophagus of a palm viper, Bothriechis marchi, with suspected adenoviral infection. The mucosa and propria-submu- cosa are infiltrated by macrophages and lymphocytes. HE stain. Bar = 100 pm. Figure 3. Transmission electron microscopy reveals numerous, hexagonal, outlined, suspected adenoviral viral particles dis­ persed throughout the nucleus of esophageal mucosal epithelial cells of a palm viper. Bar = 400 pm.

Figure 2. Higher magnification of palm viper with suspected Figure 4. Transmission electron microscopy shows a paracrys­ adenoviral infection. Several mucosal epithelial cells of the esoph­ talline array of nonenveloped, hexagonally outlined viral particles agus contain large basophilic intranuclear inclusion bodies. Arrows with electron dense centers within the nucleus of esophageal indicate two such inclusion bodies. HE stain. Bar = 50 pm. mucosal epitheial cells in a palm viper. Bar = 325 pm.

DISCUSSION nostic testing such as viral culture would be needed for defin­ itive confirmation of adenovirus in this case. This is the first Gastrointestinal diseases due to viral infection have been reported case of stomatitis and esophagitis due to adenovirus­ infrequently reported in snakes, and most of these case like infection in a snake. reports were due to infection with adenoviruses. This palm viper also had inclusion body-like disease, Gastrointestinal infection with adenovirus has been described based upon the characteristic eosinophilic intracytoplasmic in the stomach of a gaboon viper, Bids gabonica, and the duo­ inclusion bodies in epithelial cells of multiple organs. denum of a four-lined rat snake, Elaphe quatuorlineata, Inclusion body-like disease has been previously described in mountain kingsnake, Lampropaltis zonata multicincta palm vipers, which had identical inclusion bodies as this palm (Wozniak, et al, 2002) a boa constrictor, Boa constrictor, and viper (Raymond, et al, 2001). In those previously diagnosed an Aesculapian snake, Elaphe longissima, (Heldstab and cases of inclusion body-like disease, electron microscopy of Bestelli, 1984). Hepatitis due to adenoviral infection has been the inclusions revealed non-membrane bound, electron-dense reported in rosy boas, Lichanura trivirgata, and boa constric­ inclusions of various sizes that were associated with small tors (Jacobson, et al, 1985, Schumacher, et al, 1994, Ramis, electron-dense particles that had morphologic features of et al, 2000). In this palm viper, the morphologic features of retroviruses. Similar electron microscopic findings have been the viral particles noted by electron microscopy (shape, size, described in boid snakes with IBD (Schumacher, et al, 1994). and paracrystalline arrays) and their intranuclear location are This disease affected palm vipers from several different zoo­ most consistent with adenovirus. However, additional diag­ logical parks, including the zoo where this palm viper was

Volume 12, No. 3,2002 Journal of Herpetological Medicine and Surgery 31 housed. Inclusion body-like disease of palm vipers has many Ramis A, Femandez-Bellon N, Majo N, Martinez-Silvestre clinical, histologic, and electron microscopic similarities to A, Latimer K, Campagnoli R. 2000. Adenovirus hepatitis in a IBD of boid snakes. Most palm vipers with clinical inclusion boa constrictor (Boa constrictor). J Vet Diagn Invest, 12:573- body-like disease succumbed to septicemia, and an underlying 576. immunosuppression induced by the proposed retroviral infec­ Raymond J, Gamer M, Nordhausen R, Jacobson E. 2001. A tion is suspected. Inclusion body-like disease in this palm disease resembling inclusion body disease of boid snakes in cap­ viper may have caused immunosuppression, predisposing it to tive palm vipers (Bothriechis marchi). J Vet Diagn Invest, infection by adenovirus. 13:82-86. Schumacher J, Jacobson E, Burns R, Tramontin R. 1994. ACKNOWLEDGEMENTS Adenovirus-like infection in two rosy boas (Lichanura triuirgata). J Zoo Wild Med, 25(3):461-464. We thank R. Brown, HT-ACSP, for preparation of histol­ Schumacher J, Jacobson E, Homer B, Gaskin J. 1994. ogy specimens. Inclusion body disease in boid snakes. J Zoo Wild Med, 25:511- 524. REFERENCES Wozniak EJ, DeNardo DF, Brewer A, Wong V, Tarara RP. 2002. Identification of adenovirus- and dependovirus-like agentsDownloaded from http://meridian.allenpress.com/jhms/article-pdf/12/3/30/2207364/1529-9651_12_3_30.pdf by guest on 27 September 2021 Heldstab A, Bestetti G. 1984. Virus associated gastrointesti­ in an outbreak of fatal gastroenteritis in captive bom California mountain kingsnake, Lampropeltis zonata multicincta. JHMS, nal disease in snakes. J Zoo Anim Med, 15:118-128. 10(2):4-7. Jacobson E, Gaskin J, Gardiner C. 1985. Adenovirus-like infection in a boa constrictor. JAVMA, 187(11): 1226-1227. Juhasz A, Ahne W. 1993. Physiochemical properties and cytopathogenicity of an adenovirus-like agent isolated from a com snake (Elaphe guttata). Arch Virol, 130(3-4): 429-439.

Journal of Herpetological Medicine and Surgery Volume 12, No. 3,2002