Signalment:  

Female, Rocky Mountain elk calf (Cervus elaphus nelsoni).Over a three week period, 11 free-ranging Rocky Mountain elk (mostly calves) were found dead at a Colorado ranch in late January.

Gross Description:  

Postmortem examination was performed on a female elk calf in poor body condition with mild autolysis. The calf was very thin with minimal fat stores throughout and moderate serous atrophy of fat within the bone marrow (emaciation).  Subcutaneous and intramuscular facial edema was prominent in the hind limbs.  The abdomen contained scant serous fluid and large, extensive mats of fibrin along serosal surfaces (fibrinous peritonitis).  Multifocally, the mucosa of the rumen was severely ulcerated, characterized by focal, irregularly round to linear, black, roughened depressions bordered by a hemorrhagic rim.  Ulcerated areas were mural to nearly transmural without perforation.  Rumen pillars were most severely affected, and rumen contents were dry.  The epicardial surface had moderate multifocal petechial hemorrhages.  The ventral margins of the cranioventral lung lobes were dark purple and consolidated.

Histopathologic Description:

Rumen:  Affecting approximately 50% of the section is a wedge-shaped region of the rumen obscured by transmural coagulative necrosis with preservation of tissue architecture but loss of cellular detail (infarction). The center of the lesion has regions of eosinophilic cellular and karyorrhectic debris, fibrin deposition, and multifocal mineralization.  Necrotic regions are ulcerated and coated by myriad mixed bacteria. They are bordered by extensive hemorrhage, edema, fibrin, infiltration by high numbers of neutrophils, lymphocytes, and plasma cells and mild fibroblastic proliferation (fibroplasia).  The walls of vessels throughout the tissue are frequently obscured, either segmentally or diffusely by finely fibrillar, brightly eosinophilic acellular material (fibrinoid necrosis) and moderate to high numbers of neutrophils and lymphocytes admixed with karyorrhectic debris and edema (leukocytoclastic vasculitis).  Affected vessels often contain organizing fibrin aggregates and eosinophilic cellular and karyorrhectic debris (vascular thrombosis).  Endothelial cells lining affected and unaffected vessels (predominately along the margin of the infarct) are hypertrophic and frequently contain basophilic, smudged, 8-10 mm inclusions that fill the nucleus. The adjacent mucosa and submucosa are infiltrated by moderate to high numbers of neutrophils and lymphocytes admixed with edema.  Intact mucosal epithelium also has intracellular edema fluid, occasionally resulting in ballooning epithelial cells.

Microscopic diagnoses of tissues not submitted: Bronchopneumonia, suppurative and necrotizing, subacute, moderate, with mild lymphoid hyperplasia.

2. Lungs:  Vasculitis, mild, subacute, multifocal (no viral inclusions detected).

3. Heart:  Necrotizing myocarditis, multifocal, mild, acute (no viral inclusions detected).

Morphologic Diagnosis:  

Rumen:  Severe necrotizing vasculitis with thrombosis and infarction, subacute to chronic, multifocal, with ulcerative rumenitis and endothelial cell intranuclear viral inclusions consistent with Odocoileus adenovirus.

Lab Results:  

Odocoileus adenovirus PCR of lung:  Positive

Bovine viral diarrhea I&II PCR of lymph node:  Negative

Aerobic culture of lung: Pasteurella multocida, heavy growth

Condition:  

Necrotizing vasculitis/Odocoileus adenovirus

Contributor Comment:  

Adenoviruses are non-enveloped icosahedral viruses that contain double-stranded DNA.  A wide range of veterinary species are affected by this class of virus, often resulting in respiratory, gastrointestinal, or systemic disease.⁴ In most species, clinical disease is mild, unless the animal is young or immunocompromised.⁴  Odocoileus adeno-virus (OdAdV) was first discovered in mule deer in Northern California in 1993 and was determined to be the causative agent of adenovirus hemorrhagic disease (AHD).^11,12  Since that time, the virus has been found to infect multiple species of deer,^11,13 moose,⁸ and more recently, pronghorn and elk (unpublished data).   Transmission has been documented via direct contact of cervids and is thought to occur through infected secretions or feces.¹³ Studies have shown that OdAdV is most closely related to bovine adenovirus,⁵ however, experimental infection studies of domestic cattle with OdAdV have not demonstrated viral replication or disease in this species.¹⁵

As is true with other adenovirus infections, young animals experience a significantly higher morbidity and mortality rate when compared to adults.^1,9,11  Death can occur as rapidly as five days post-infection, but chronic cases can also occur.^12,13  Acute cases usually have no symptoms or may exhibit ptyalism, diarrhea (with or without blood), weakness, lethargy, anorexia, and edema.^1,9,10  Chronic infections are usually associated with ulcers and abscesses of the mouth and throat, emaciation, and/or sepsis .^1,13

Disease can be systemic or local.¹¹  Typical necropsy findings of systemic infections include severe pulmonary edema expanding the interlobular septa with ecchymotic hemorrhages of the lungs and pulmonary artery, and a hemorrhagic enteropathy characterized by hemorrhage within the lumen of the small and large intestine.^12,13 Additionally, multiple ulcerative lesions of the mouth, nasal cavity, mandible, maxilla, and forestomach can be present in systemic disease.¹¹ Local disease is restricted to the upper alimentary tract (stomatitis, pharyngitis, mandibular/maxillary osteo-myelitis, and/or rumenitis).¹³ In either form, secondary bacterial infections are common, primarily presenting as abscesses of the oral and nasal cavities.^12-15  Secondary bacterial agents include Trueperella pyogenes, Prevotella spp., Fusobacterium necrophorum, Streptococcus spp., Pasteurella multocida, and Pepto-streptococcus spp.^12-15

Histologic findings of OdAdV are predominately associated with vascular changes, including endothelial hypertrophy, fibrinoid vascular necrosis, leukocytoclastic vasculitis, and thrombosis, with resulting mucosal ulcerations, tissue infarctions, and extensive edema.^10,11 All types of vessels can be affected and endothelial cell inclusions can be seen in any tissue, with or without significant vascular changes.^12,13  Animals with infection localized to the upper alimentary tract, especially in more chronic infections, can lack endothelial intranuclear inclusions and viral detection methods may be ineffective.^1,11,13  Diagnosis of adenovirus infection in cervids can be accomplished by histopathology with characteristic endothelial intranuclear inclusions,¹¹ immunohistochemistry of infected tissue,¹² enzyme-linked immuno-sorbent assay (ELISA),⁶ serum neutralization,¹³ virus isolation,¹¹ immuno-fluorescence,¹¹ and PCR (unpublished data).

Differential diagnoses for rumen ulceration include bluetongue (BT) and epizootic hemorrhagic disease of deer (EHD), which are both associated with widespread hemorrhage, edema, necrotizing vasculitis, thrombosis, and ulceration of the alimentary tract.¹⁰  Unlike acute adenovirus infection, inclusions are not detected in tissues with either of these diseases, and additional diagnostic testing may be necessary for definitive diagnosis.¹⁰ In this case, BT and EHD were considered unlikely based on the time of year, since the insect vector (Culicoides) is inactive during the winter in Colorado.  However, in the warmer season and with chronic cases of OdAdV where inclusions may not be present, all three hemorrhagic diseases (BT, EHD, and AHD) should be considered. Additional differentials for rumen ulceration include mycotic rumenitis and rumen acidosis with Fusobacterium necrophorum infection (necrobacillosis).

In this case, the cause of death was likely multifactorial:  malnutrition (emaciation), Pasteurella multocida infection, and OdAdV infection.  Pasteurellosis is a known cause of mortality in elk calves and can result in rapid septicemia and death.³  For this animal, it was unclear if the pulmonary vasculitis and necrotizing myocarditis were related to P. multocida infection and bacterial sepsis, OdAdV, or both. Unfortunately, the definitive role of adenovirus as a primary pathogen in elk and/or a predisposition for other infectious organisms requires further investigation. 

JPC Diagnosis:  

Rumen: Vasculitis, fibrinonecrotic, multifocal to coalescing, subacute, severe with fibrin thrombi, infarction, ulcerative rumenitis, and endothelial intranuclear viral inclusion bodies, Rocky Mountain elk, Cervus elaphus nelsoni.

Conference Comment:  

We thank the contributor for providing an outstanding example and superb overview of Odocoileus adenovirus (OdAdV) causing adenovirus hemorrhagic disease (AHD) in the rumen of a Rocky Mountain elk. Participants readily identified necrotizing arteritis within the rumen wall with throm-bosis and large geographical areas of infarcted tissue.  These infarcted areas correspond to the large ulcerated areas in the rumen seen grossly in the image provided by the contributor. This case nicely demonstrates frequent large basophilic intranuclear inclusion bodies within reactive and hypertrophic endothelial cells, characteristic for endotheliotropic adeno-virus infection.^2,10

Conference participants discussed the top two differentials for necrotizing vasculitis causing severe hemorrhagic disease in elk and white-tailed deer. Bluetongue virus (BTV) and epizootic hemorrhagic disease virus (EHD) of the genus Orbivirus and family Reoviridae, mentioned by the contributor, are the top two differentials that must be considered in cases of widespread hemorrhage, necrotizing vasculitis, edema, thrombosis, and ulceration of the alimentary tract.^{1,2,4,10,11-15} EHD and BTV are both spread by the salivary secretions of the biting Culicoides spp. vector. In North America, EHD is the mist significant viral disease in the highly susceptible white-tailed deer, and devastating epizootics have been reported in the United States. Elk are generally less severely affected by BTV and EHD infection. Both BTV and EHD circulate throughout North America and outbreaks often occur simultaneously.^10,11-15 Interestingly, both viruses can be concurrently isolated from the same individual Culicoides vector. Conference participants also noted that malignant catarrhal fever caused by ovine herpesvirus-2, a highly pathogenic gammaherpesvirus, can also produce intestinal hemorrhage,  pulmonary congestion, edema, and petechiae of the spleen, intestines, heart, and liver with lymphocytic necrotizing vasculitis, and thrombosis in cervids and should also be considered as a potential differential in similar cases.⁷

The conference moderator mentioned that orbivirus infection will not produce the characteristic intranuclear viral inclusion bodies of adenovirus. To further differentiate these two diseases, orbiviral disease outbreaks are usually associated with acute onset of fulminant disease with high morbidity and mortality and viral tropism for the microvasculature endothelial cells. OdAdV typically results in a more chronic onset with targeting of the larger vessels and resulting in larger areas of coagulative necrosis.^{1,2,4,5,10-15} Additionally, petechial and/or ecchymotic hemorrhages present at the base of the pulmonary artery has been reported to be highly characteristic and potentially pathognomonic for orbivirus infection in susceptible species. As mentioned by the contributor, diagnosis of adenovirus infection is done by a combination of testing modalities, including visualization of the characteristic endothelial intranuclear inclusions with histopathology, immunohistochemistry (IHC), virus isolation, and polymerase chain reaction (PCR).¹¹

References:

1.      Boyce WM, Woods LW, Keel MK, MacLachlan NJ, Porter CO, Lehmkuhl HD. An Epizootic of Adenovirus-Induced Hemorrhagic Disease in Captive Black-Tailed Deer (Odocoileus hemionus). J Zoo Wildl Med. 2000; 31:370-373.

2.      Fox KA, Atwater L, Hoon-Hanks L, Miller M. A mortality event in elk (Cervus elaphus nelsoni) calves associated with malnutrition, pasteurellosis, and deer adenovirus in Colorado, USA. J Wildl Dis. 2017; 53(3): DOI: 10.7589/2016-07-167. [Epub ahead of print].

3.      Franson, JC, Smith BL. Septicemic pasteurellosis in elk (Cervus elaphus) on the United States National Elk Refuge, Wyoming. J Wildl Dis. 1988;24:715–717.

4.      Gelberg HB. Alimentary system and the peritoneum, omentum, mesentery, and peritoneal cavity. In: McGavin JF, Zachary MD, eds. Pathologic Basis of Veterinary Disease. 6th ed. St. Louis, MO: Elsevier Mosby; 2017:325-375.

5.      Lapointe, JM, Hedges JF, Woods LW, Reubel GH, MacLachlan NJ. The adenovirus that causes hemorrhagic disease of black-tailed deer is closely related to bovine adenovirus-3. Arch Virol. 1999;144:393-396.

6.      Lapointe, JM, Woods LW, Lehmkuhl HD, et al. Serologic detection of adenoviral hemorrhagic disease in black-tailed deer in California. J Wildl Dis. 2000;36:374-377

7.      Palmer MV, Thacker TC, et al. Active and latent ovine herpesvirus-2 (OvHV-2) infection in a herd of captive white-tailed deer (Odocoileus virginianus). J Comp Path. 2013; 149:162-166.

8.      Shilton CM, Smith DA, Woods LW, Crawshaw GJ, Lehmkuhl HD. Adenoviral infection in captive moose (Alces alces) in Canada. J Zoo Wildl Med. 2002;33:73-79.

9.      Sorden SD, Woods LW, Lehmkuhl HD. Fatal pulmonary edema in white-tailed deer (Odocoileus virginianus) associated with adenovirus infection. J Vet Diagn Invest. 2000;12:378–380.

10.  Uzal FA, Plattner BL, Hostetter JM. In: Maxie MG, ed. Jubb, Kennedy, and Palmer’s Pathology of Domestic Animals. Vol 2. 6th ed.  Philadelphia, PA: Elsevier; 2016:131-140.

11.  Woods, LW, Swift PK, Barr BC, et al. Systemic adenovirus infection associated with high mortality in mule deer (Odocoileus hemionus) in California. Vet Pathol. 1996;33:125-132.

12.  Woods, LW, Hanley RS, Chiu PH et al. Experimental adenovirus hemorrhagic disease in yearling black-tailed deer. J Wildl Dis. 1997;33:801-811.

13.  Woods, LW, Hanley RS, Chiu PH, et al. Lesions and transmission of experimental adenovirus hemorrhagic disease in black-tailed deer fawns. Vet Pathol. 1999;36:100-110.

14.  Woods LW, Lehmkuhl HD, Swift PK, et al. Experimental adenovirus hemorrhagic disease in White-tailed deer fawns. J Wildl Dis. 2001;37:153–158.

15.  Woods LW, Lehmkuhl HD, Hobbs LA, Parker JC, Manzer M. Evaluation of the pathogenic potential of cervid adenovirus in calves. J Vet Diagn Invest. 2008;20:33–37.

Click the slide to view.

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2-1. Rumen, elk calf:

2-2. Esophogeoruminal junction:, elk calf:

2-3. Esophogeoruminal junction:, elk calf:

2-4. Esophogeoruminal junction:, elk calf:

2-5. Esophogeoruminal junction:, elk calf:

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