Joint Pathology Center
Veterinary Pathology Services
Wednesday Slide Conference
25 August 2019
CASE III: Case 1 (JPC 4101226).
Signalment: 8-week-old female Rex cross rabbit, Oryctolagus cuniculus.
History: Found dead in the woods
Gross Pathology: The rabbit was one of several young rabbits housed and hand-reared at a rescue organization. Both this rabbit and its litter mate displayed symptoms of lethargy, anorexia and hypothermia. Within 48 hours of the onset of clinical signs, the rabbit died. The litter mate and several other young rabbits in the organization also died in the days following this death.
Laboratory results: Bacteriology : Yersinia pseudotuberculosis in liver and spleen.
Despite moderate autolysis of the tissues, the liver has multifocal areas of hepatocytes with reasonable preservation. Scattered amongst these areas, and throughout the hepatic lobule, are swollen eosinophilic hepatocytes with varying degrees of pyknosis, karyolysis and karyorrhexis (necrosis) and marked loss of chord architecture. In addition to these findings, there is marked multifocal distention of bile ducts with periductal fibrosis expanding and compressing the surrounding hepatic parenchyma. The biliary epithelium is hyperplastic and forms papillary projections into the lumen. Numerous epithelial cells contain asexual and sexual developmental stages of coccidia and the biliary duct lumina contain large numbers of oocysts. Moderate numbers of lymphocytes and plasma cells and smaller numbers of heterophils are admixed within the periductal fibrous tissue and the connective tissue stroma of the papillary projections.
The kidney diffusely shows recent vascular congestion and tubular and glomerular hemorrhage. Intracapillary fibrin thrombi are present multifocally in glomeruli. Scattered renal tubular cells exhibit hypereosinophilic cytoplasm, pyknotic nuclei, karyorrhexis and karyolysis (necrosis).
Liver: Hepatitis, acute, moderate to severe, random with hepatocellular necrosis
Liver: Cholangitis, proliferative, chronic severe with intraepithelial coccidia
Kidney: Nephritis, haemorrhagic, severe, acute, with multifocal glomerular thrombosis
Contributor Comment: In this case, the rabbit had two concurrent disease processes: hepatic coccidiosis (Eimeria stiedae) and rabbit hemorrhagic disease (RHD) caused by rabbit calicivirus. RHD was the cause of death, with hepatic coccidiosis as an incidental finding.
Rabbit hemorrhagic disease or viral hemorrhagic disease is caused by a calicivirus of the genus Lagovirus, which rapidly infects wild and domestic rabbits (Oryctolagus cuniculi). The virus was first identified in 1984 in China where it killed 140 million rabbits within months.21 It was released in Australia and New Zealand as a method of pest control of wild rabbits and is endemic in the populations in those countries.6,27
The RHD virus is highly infectious, with a greater than 80% mortality rate, and it usually produces death in affected individuals within 48 to 72 hours of infection.1,21 Death is due to acute liver damage and disseminated intravascular coagulation (DIC).24 No clinical signs may be observed if the infection is peracute. Alternatively, they may manifest as anorexia, lethargy, pyrexia, conjunctival congestion and neurological signs such as ataxia, opisthotonos or paralysis in acute infections.13,29 Other signs such as dyspnea, cyanosis, or hemorrhagic epistaxis may also be seen. Subacute infections result in milder clinical signs, with some rabbits surviving. Many rabbits with chronic infections will die within 1 to 3 weeks after a period of jaundice, weight loss and lethargy.11
The primary findings at necropsy include hepatomegaly with an enhanced lobular pattern, splenomegaly, renomegaly, pulmonary hemorrhage and oedema, blood-tinged nasal discharge or bloody foam in the tracheal lumen.1,14 Additionally, hyperemia or sub-serosal hemorrhages may be found on multiple organs such as the intestine, pericardium, kidney and lungs.1
The primary histopathological lesion is an acute necrotizing hepatitis.2 In adult rabbits, the virus has tropism for hepatocytes and can be detected in the liver a few hours post-infection.2 Viral replication occurs primarily in hepatocytes in the centrilobular areas but also in Kupffer cells.2,8-10,22 Additionally, viral antigen has been detected in macrophages of the spleen, alveoli, kidneys and small intestine.22,23 The virus induces apoptosis in these cells, releasing viral progeny to infect other cells.7 In younger rabbits, viral antigen has been detected only in rabbits greater than 4 weeks of age.18,22 Younger rabbits appear to be resistant to the RHD virus. Rabbits less than 3 weeks of age are fully resistant, and this resistance decreases as rabbits increase in age to 8 weeks old, where mortality rates are the same as adult rabbits.1
The mechanism of resistance is unclear. It is thought to relate to viral attachment to the carbohydrate group of host-cell histo-blood group antigens (HBGA).24,25 RHD attaches to HBGA H type 2, A type 2 and B type 2 oligosaccharides which are found on the surface of epithelial cells of the upper respiratory and digestive tracts. The expression of HBGA H type 2 seems to be mostly lacking in the upper respiratory and gastrointestinal tracts of resistant rabbits.17 However, other factors must be involved in viral attachment to cells as hepatocytes, which are the main cell involved in viral replication, do not contain these HBGA receptors.25 Additionally, in young rabbits, viral replication occurs only in a small fraction of hepatocytes, indicating other factors are involved in the resistance to this virus.1
In this rabbit, hepatic coccidiosis, was an incidental finding that likely contributed to its poor condition. Eimeria stiedae is a common cause of morbidity and mortality in rabbits. Ten other species of Eimeria spp. infect the domestic rabbits but specifically infect the gastrointestinal tract.15,17 Clinical signs for hepatic coccidiosis include a thin body condition, diarrhoea, a pot-bellied appearance and, in severe cases, icterus.21
Young, weanling rabbits are most often infected when ingestion of the sporulated oocyst from the environment occurs. The oocysts are shed after a prepatent period of 15 to 18 days, and once in the environment, are extremely resistant to disinfectants.21
Sporulated oocysts are ingested, where sporozoites are released to invade the duodenal mucosa and lamina propria.18 It is possible that sporozoites are then transported to the liver via either lymphatic or hematogenous spread. Organisms have been found in macrophages in the lymphatics and in regional lymph nodes within 12 hours of exposure, in bone marrow within 24 hours and in the liver within 48 hours.18,21 Once in the liver, sporozoites invade the epithelial cells of bile ducts to become trophozoites. Trophozoites undergo the asexual division of schizogony and merogony over several generations to eventually form the macro- and microgametocytes involved in sexual division. Fertilization of a macrogametocyte by a microgametocyte to form an oocyst which is then shed in the feces.18,21
Institute of Veterinary Animal and Biomedical Sciences, Massey University
Private Bag 11222, Palmerston North
New Zealand 4442
JPC Diagnosis: Liver: Hepatitis, necrotizing (its really apoptosis), diffuse, severe.
2. Liver, bile ducts: Epithelial hyperplasia, diffuse, mild to moderate, with intraluminal apicomplexan oocysts.
JPC Comment: The
contributor has done an excellent review of a virus of global import in this
species, as well as well-known common parasite of young rabbits.
The Czech v351 strain of rabbit lagovirus has been used in Australia and New Zealand (following illegal release on the South island) to control pest rabbits for a number of years.14 However, even before this virus was released into a naive population, wild rabbits demonstrated cross-reacting antibodies suggesting other similar viruses had previously circulated within this population. The first non-pathogenic rabbit calicivirus was identified in Italy, following seroconversion of animals in a rabbitry with no history of clinical disease. Shortly thereafter, a non-pathogenic strain of rabbit calicivirus with 88% was identified in Australia, and New Zealand. One common factor was that these viruses appeared to prevail in cool high-rainfall areas.14 Additional "benign" viruses have also been identified in European hares in Australia, with evidence of previous recombination, confirming previous hypotheses of the origin of genetic diversity within this genus of virus.12 In 2014, a recombinant strain of RHDV was identified in Australia which contained capsid and non-structural genes of non-pathogenic RHDV variants.12
Other interesting changes have been noted in RHDV since its release thirty years ago in Australia. In contrast to myxoma virus, which was also released into the wild to control pest rabbits and rapidly attenuated in virulence over time as local rabbits developed genetic resistance, pathogenic genotypes of Australian RHDV appear to have increased in virulence.7 A comparison study of deaths in a closed population experiencing outbreaks back to the original release of RHDV noted that in outbreaks from 2007-2009 (as compared to the 1990s), more recent outbreaks demonstrated elevation in case fatality rates, disease duration (time to death), as well as the amount of virus produced in infected animals.7
The first lagovirus other than RHVD infecting rabbits in the United States was identified by Bergin et al in 2009. This virus, referred to as Michigan rabbit calicivirus occurred in a closed rabbitry with an approximately 32% case mortality and clinical and necropsy findings of epistaxis, vulvar hemorrhage, diarrhea, and ocular discharge. This virus averaged 79% homology with the RNA genome of RHDV virus. The rabbitry was ultimately depopulated.5
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