Inclusion Bodies In Hepatocytes Research Articles

Comprehensive diagnosis of abortion in cattle

Abstract. Abortion is one of the most common factors that has an extremely negative eff ect on the reproductive capacity of cattle, resulting in signifi cant economic losses. Abortions can be caused by infectious and non-infectious agents. Diagnosis of abortion is a diffi cult task, since only about 40% of abortions are diagnosed as infectious, and the remaining 60% - as unknown etiology. The goal was to establish the infectious etiology of abortion using complex diagnostics. The research was carried out in the laboratories of "Center of Veterinary Diagnostics" LLC (Kyiv). The selection of samples for research was carried out in the section hall of the LLC "Center of Veterinary Diagnostics" during the autopsy. The material for the study was a fetus and a vaginal smear from an aborted cow. Abortion occurred at 4 months of pregnancy. To establish the etiology, the following research methods were used: patho-anatomical, bacteriological, histological and molecular genetic (PCR-RT). Autolysis, light gray foci ≤ 2 mm in diameter in the lungs, and multiple light gray foci ≤ 1 mm in diameter in the liver were the main pathological fi ndings. The main pathohistological changes detected were focal myocarditis, intranuclear inclusion bodies in hepatocytes, multifocal coagulation necrosis in the lungs and liver, and diff use interstitial nephritis. Bacteriological culture was negative. DNA of infectious bovine rhinotracheitis virus (IBRV) and DNA of bovine herpesvirus type 4 (BHV-4) was isolated from the examined samples of fetal tissues and vaginal smear by PCR-RT. Using a comprehensive study, infectious agents and changes characteristic of the infectious process were identifi ed. On the basis of the obtained results, the etiology of the abortion was established as co-infection of IBRV and BHV-4. Complex diagnostics provides an opportunity to establish a cause-and-eff ect relationship between the identifi ed infectious agent and pathological changes in the fetus and the etiology of abortion. The results of a set of studies provide information on the manifestation of herpesvirus co-infection and complement the existing knowledge that can be used in the diagnosis of abortions in cows. Key words: abortion, cattle, IBRV, BHV-4, coinfection, PCR-RT, pathohistology, diagnosis.

Prevalence and Molecular Diagnosis of Hydropericardium Hepatitis Syndrome in the Poultry Population of Mizoram, India

Background: Hydropericardium hepatitis syndrome (HHS) is an important emerging disease having economic importance and affecting particularly broiler chickens. Although the disease has been reported as a major problem in almost all the regions of India, not much is known from the North Eastern states of India. The present study was aimed to study prevalence, clinico-pathology and molecular diagnosis of HHS in poultry population of Mizoram. Methods: A poultry population of total 2430 from 15 different farms located in different districts of Mizoram were studied for outbreaks of HHS. Clinical manifestation of the disease in affected birds was recorded and pathological studies were carried out in total 190 dead birds. The outbreaks were further confirmed by detection of hexon gene of FAdV in tissue lesions by PCR. Result: Outbreak of HHS was recorded in 5 (33.3%) different broiler farms with sudden onset of mortality (18 to 25.2%) in broiler birds of 4 to 6 weeks of age. Pathological studies revealed characteristic lesions comprising hydro pericardium, hepatitis and basophilic intranuclear inclusion bodies in hepatocytes. Detection of hexon gene of FAdV in tissue lesions by PCR assay has further confirmed the outbreaks. This study reports for the first time, high prevalence of HHS in poultry population of Mizoram.

Molecular characterization and pathogenicity of highly pathogenic fowl adenovirus serotype 4 isolated from laying flock with hydropericardium-hepatitis syndrome

Hydropericardium-hepatitis syndrome (HHS) is an important emerging disease responsible for huge economic losses to the poultry industry in China. HHS primarily affects 20 to 60-day-old broilers and rarely occurs in laying flock. In this study, the highly pathogenic fowl adenovirus (FAdV) strain, AH-F19, was isolated from the liver samples of 120-day-old laying flock with HHS and its phylogenetic information, genetic mutations, and pathogenicity was evaluated. The phylogenetic analysis revealed that AH-F19 belonged to the FAdV serotype 4 (FAdV-4) cluster, however, 100K differs from the other FAdV-4 strains and is divided into different branches. Amino acid variations in fiber-2 for pathogenic isolates and non-pathogenic isolates indicated that D219, T300, and T380 may not be responsible for virulence. Animal experiments revealed AH-F19 to be a highly pathogenic isolate that can cause 100% mortality in three-week-old specific pathogen-free (SPF) chickens, which exhibited typical hydropericardium and hepatitis. Microscopically, the presence of basophilic intranuclear inclusion bodies in hepatocytes, fractured heart muscle fibers, as well as kidney degeneration and necrosis was observed. Collectively, these findings enriched our understanding of FAdV-4 pathogenicity and provided a reference for further exploration into its pathogenicity.

Molecular Characterization of Fowl Adenovirus D Species in Broiler Chickens with Inclusion Body Hepatitis in Egypt

In the current study, fowl adenovirus (FAdV) was detected and characterized from broiler chicken flocks with inclusion body hepatitis (IBH) in Northern Egypt during the period from March 2017 till March 2018. Grossly, the examined flocks showed hydropericardium and enlarged friable livers with ecchymotic hemorrhages. Histopathologically, liver tissues revealed intranuclear inclusion bodies in hepatocytes and hepatitis. Molecular detection by PCR assay using primer specific for the L1 loop region of hexon gene was conducted, the PCR products were sequenced for typing of FAdV. Seventeen out of 37 flocks tested were positive. Sequence analysis revealed that the 17 FAdVs were identical, suggesting a common ancestor for the FAdVs circulating in northern Egypt during the period of study. Phylogenetically, they clustered with fowl adenoviruses D species (FAdV-2 and FAdV-11). Interestingly, pathogenicity varied from non-pathogenic to more virulent with mortalities up to 20% under field condition. To our knowledge, this is the first report of FAdV-D from broiler chicken in Egypt. Further studies are needed to examine different poultry species and other geographic areas in Egypt. Moreover, studies on the pathogenicity and potential preventive measures against FAdV infection on poultry farms are necessity.

First report of fowl aviadenovirus serotypes FAdV-8b and FAdV-11 associated with inclusion body hepatitis in commercial broiler and broiler-breeder flocks in Turkey.

Inclusion body hepatitis (IBH), hydropericardium syndrome (HS), and gizzard erosion (GE) are all economically important diseases in the poultry industry worldwide and are all caused by fowl aviadenovirus (FAdV). It is important to identify the serotype of the virus to differentiate these diseases. In the present study, a total of six recent FAdV serotypes were isolated and identified in broiler and broiler-breeder flocks in Izmir, Manisa, and Aydın provinces of the Aegean region of Turkey between January and March 2019. The viruses were isolated from livers and pooled organs of chickens using primary chicken embryo kidney cell cultures (CEKC). Virus isolates were identified by PCR amplification of the loop 1 (L1) variable region of the hexon gene followed by Sanger sequencing. Sequence analysis revealed the presence of both FAdV-D (serotype 11) and FAdV-E (serotype 8b). The viruses that were isolated were associated withIBH, which is typically characterized by gross lesions such as enlarged and pale yellow liver with multiple petechial hemorrhages. Histopathological examination also showed necrotizing hepatitis with intranuclear inclusion bodies in hepatocytes. This study is the first report of the isolation and identification of FAdV serotypes associated with IBH in commercial broilers and broiler-breeder flocks in Turkey. The results of sequence analysis showed that FAdV-8b and FAdV-11 were the circulating serotypes that caused recent field outbreaks of IBH in the Aegean region between January and March, 2019.

Multi-tool diagnosis of an outbreak of ranavirosis in amphibian tadpoles in the Canadian boreal forest.

Investigation of mortalities in isolated wild amphibian populations presents diagnostic difficulties that can hinder reaching a definitive diagnosis for the cause of death. Disease can only be diagnosed when pathogen presence (e.g. detection by PCR) is linked to tissue lesions (histopathology) in the host. We report a 2-site outbreak of ranavirosis in wild anuran tadpoles in the boreal forest of Wood Buffalo National Park, Canada, diagnosed by histologic and molecular techniques. Mortalities occurred in wood frog Rana sylvatica tadpoles and boreal chorus frog Pseudacris maculata tadpoles. Lack of mortality in sympatric Canadian toad Bufo (Anaxyrus) hemiophrys tadpoles suggested lower disease susceptibility in this species. In the former 2 species, ranavirosis was diagnosed based on consistent histopathology, immunohistochemistry (IHC), in situ hybridization (ISH), and quantitative PCR results. The most common histopathologic lesion present in wood and boreal chorus frog tadpoles was necrosis of the skin, oral mucosa, renal tubular epithelium, renal hematopoietic tissue, and branchial epithelium. Mild hepatic and pancreatic necrosis and rare intracytoplasmic inclusion bodies in hepatocytes were less common. Skeletal and connective tissues in budding limbs often had multifocal to coalescing necrosis and were intensely positive for ranavirus, with IHC staining even in areas where no obvious necrosis could be observed. Abundant IHC and ISH staining in actively growing tissues support a link between disease emergence and amphibian developmental stage. Our findings provide a definitive diagnosis of ranavirosis in free-living amphibians and highlight the effectiveness of multi-tool approaches to mortality investigation and elucidation of pathogenesis of ranavirosis in wild amphibians.

Pathogenicity and molecular characterization of a fowl adenovirus 4 isolated from chicken associated with IBH and HPS in China

BackgroundSince July in 2015, an emerging infectious disease, Fowl adenovirus (FAdV) species C infection with Hepatitis-Hydropericardium syndrome was prevalent in chicken flocks in China. In our study, one FAdV strain was isolated from commercial broiler chickens and was designated as SDSX1.The phylogenetic information, genetic mutations and pathogenicity of SDSX1 were evaluated.ResultsThe phylogenetic analysis indicated that SDSX1 is a strain of serotype 4, FAdV-C. The amino acid analysis of fiber-2 showed that there were more than 20 mutations compared with the non-virulent FAdV-C strains. The pathogenic evaluation of SDSX1 showed that the mortality of one-day-old chickens inoculated SDSX1 was 100%. The typical histopathological changes of SDSX1 were characterized by the presence of basophilic intranuclear inclusion bodies in hepatocytes. The virus copies in different tissues varied from107 to 1011 per 100 mg tissue and liver had the highest virus genome copies.ConclusionIn conclusion, the isolate SDSX1, identified as FAdV-4, could cause one-day-old chicks’ typical inclusion body hepatitis (IBH) and hepatitis-hydropericardium syndrome (HHS) with 100% mortality. The virus genome loads were the highest in the liver. Molecular analysis indicated that substitutions in fiber-2 proteins may contribute to the pathogenicity of SDSX1.

Pathogenic, Phylogenetic, and Serological Analysis of Group I Fowl Adenovirus Serotype 4 SDSX Isolated From Shandong, China.

Hydropericardium hepatitis syndrome (HHS) caused by group I fowl adenovirus serotype 4 (FAdV-4) is an acute and infectious disease in fowl, particularly in broilers aged 3–5 weeks. In June 2015, a highly pathogenic disease outbroke in 25–40 day-old ducklings in Shandong province, characterized similar symptom to HHS. In order to determine the pathogenic mechanism of FAdV-4 (SDSX strain) in meat ducks. We divided 90 25-day-old Cherry Valley meat ducks into three groups (oral, subcutaneous, and control; 30 ducks in each group) and infected them with the virus. HHS, inclusion body hepatitis, and enlargement and hemorrhage of the spleen, kidney, lung, thymus, and brain were observed in FAdV-4-infected meat ducks. Histopathological changes were mainly characterized by severe fatty degeneration in the liver, basophilic inclusion bodies in hepatocytes, and vacuolation in the bursa. More importantly, viral DNA could be detected by quantitative real-time polymerase chain reaction in several viscera tissues (e.g., heart, liver, spleen) on the third day after infection. Notably, the livers of the two infected groups contained the highest concentration of viral DNA. In addition, immune responses were studied based on titer levels of the virus antibody and the levels of inflammatory cytokines interleukin (IL)-2 and interferon (IFN)-γ, and most levels were significantly upregulated, indicating that the host immune responses were activated early in infection. These findings increase our understanding of the pathogenicity of FAdV-4 (SDSX) in meat ducks and provide the foundation for further in-depth study of the pathogenic mechanism of this virus.

Fowl adenovirus serotype 4-induced apoptosis, autophagy, and a severe inflammatory response in liver

Fowl adenovirus serotype 4 (FAdV-4) is a hepatotrophic virus that causes severe liver diseases. Upon histological examination, the most remarkable findings in the liver are small multifocal areas of necrosis and mononuclear cell infiltration, including basophilic intranuclear inclusion bodies in hepatocytes surrounded by a clear halo or which fill the entire nucleus. Here, we examined the mechanism responsible for FAdV-4-mediated hepatocyte damage in vivo and in vitro. The results showed that FAdV-4 impaired liver integrity and function, which decreased albumin and blood glucose concentrations and increased the plasma activity of aspartate aminotransferase and lactate dehydrogenase, compared with a non-infected control group (P<0.05). FAdV-4 induced hepatocyte apoptosis in a time-dependent manner in vivo and in vitro. Additionally, we found that FAdV-4 also induced the autophagy of hepatocytes, which promoted the conversion of microtubule-associated protein light chain 3 (LC3-I) to LC3-II, which is a hallmarks of autophagy. Furthermore, the mRNA expressions of interleukin (IL)-1β, IL-6, IL-8, and tumor necrosis factor (TNF)-α in vivo and in vitro showed a statistically significant increase (P<0.05) compared to that of the control group. However, the molecular mechanisms underlying the FAdV-4-induced apoptotic and autophagic cell death remain unclear. In summation, our observations suggested that FAdV-4 induced liver injury via apoptosis, autophagy, and a severe inflammatory response.

Phylogenetic Analyses of Fowl Adenoviruses (FAdV) Isolated in China and Pathogenicity of a FAdV-8 Isolate.

Fowl adenoviruses (FAdVs) have a worldwide distribution and are associated with a variety of diseases, causing considerable economic losses to the poultry industry. We characterized 10 FAdVs isolated from China in 2015-2016 and assessed the pathogenicity of a FAdV-8 strain in specific-pathogen-free (SPF) chickens. Phylogenetic analysis of a hexon gene revealed that only 1 of the 10 isolates belonged to FAdV-8, whereas others belonged to FAdV-4, indicating that Chinese FAdVs were mainly FAdV-4 in recent years. The pathogenicity experiment of the FAdV-8 strain CH/SD/2015/09 showed that no clinical signs were observed in infected chickens. Necropsy displayed mild necrotic foci and petechial hemorrhage of livers collected at 5 days postinfection (dpi). Histopathologic examination identified the presence of intranuclear inclusion bodies in hepatocytes. No virus was detected in oral and cloacal swabs at 5 dpi, and only viral DNA could be measured in kidneys collected at the same time. The results revealed that CH/SD/2015/09 had no obvious pathogenicity in 5-wk-old SPF chickens, which could provide a better understanding about the pathogenicity of the FAdV-8 serotype.

First report of a new alphaherpesvirus in a freshwater turtle (Pseudemys concinna concinna) kept in Germany

Herpesviruses represent important pathogenic agents in zoological chelonian collections. Infections in tortoises are actually most commonly associated with necrotizing lesions in the upper digestive tract. Herpesvirus infections in sea turtles are most commonly associated with fibropapillomatosis, although other disease complexes caused by other herpesviruses have been described. Herpesviruses are known to cause latent infections and may be reactivated upon various endogenous or exogenous stimuli resulting in acute and sometimes fatal disease. The present description represents the first report about a new alphaherpesvirus found in a fresh water turtle (Pseudemys concinna concinna). The animal died suddenly without showing clinical signs. Macroscopically, no lesions typically associated with a herpesvirus infection were found. Light microscopic examination showed hepatic lipidosis and countless numbers of intranuclear inclusion bodies in hepatocytes as the only significant light microscopic lesion. Transmission electron microscopy revealed typical herpesvirus particles in the nucleus of hepatocytes. To further substantiate these observations a molecular identification using PCR followed by sequencing of the obtained fragments was performed. Phylogenetic analysis indicated a new alphaherpesvirus called Emydid herpesvirus 1.

Infectious Canine Hepatitis in Free-Ranging Red Foxes (Vulpes vulpes)

INFECTIOUS CANINE HEPATITIS IN FREE-RANGING RED FOXES (VULPES VULPES) A.W. Philbey* and H. Thompsony *Royal (Dick) School of Veterinary Studies, University of Edinburgh, Easter Bush, Edinburgh and yMill House, Camis Eskan, Helensburgh, Scotland, UK Introduction: Infectious canine hepatitis (ICH), caused by canine adenovirus type 1 (CAV-1), has been reported mostly in dogs, farmed foxes and other captive carnivores. Initially called ‘epizootic fox encephalitis’, ICH was first identified in domesticated (‘tamed’) silver foxes, a colour variant of the red fox (Vulpes vulpes). ICH was diagnosed in a free-ranging grey fox (Urocyon cinereoargenteus) in Georgia, USA, in 2005. There is serological evidence of exposure to CAV-1 in free-ranging red foxes in North America, Germany, Australia and the UK, as well as in free-ranging grey foxes in North America. Materials and Methods: Sick free-ranging red foxes were presented to wildlife rehabilitation centres in the UK, where they died naturally. Tissues were submitted for virus isolation and histopathology. Results: ICH was diagnosed in three red foxes found depressed or collapsed in the wild. Grossly, the foxes were jaundiced and had congested livers with accentuation of the lobular pattern. Histologically, there was generalized necrosis and dissociation of hepatocytes, with intranuclear inclusion bodies in hepatocytes, renal glomeruli, proximal convoluted renal tubular epithelial cells and vascular endothelial cells. CAV-1 was isolated from all three foxes. Conclusions: Free-ranging red foxes are considered to be a reservoir of CAV-1 in the UK and are a potential source of infection for domestic dogs. HEALTH STATUS OF BROWN HARES (LEPUS EUROPAEUS) IN EASTERN AUSTRIA A. Posautz, A. K€ ubber-Heiss, F. Knauer, Z.S. Kelemen, F. Suchentrunk and C. Walzer Research Institute of Wildlife Ecology, University of Veterinary Medicine, Vienna, Austria Introduction: The brown hare (Lepus europaeus) population has been declining throughout its range in the past decades. As the brown hare is on the national Red List in Austria, a thorough health screening project was conducted in eastern Austria in the years 2010 and 2011. Materials and Methods: A total of 188 hares underwent thorough pathological screening. Parasitological testing was performed in all hares, bacteriology and virology only on indication. As tularemia is endemic in eastern Austria, all hares were screened for Francisella tularensis. Serological screening for Treponema spp. was undertaken in 119 animals and for European brown hare syndrome (EBHS) in 49. Results: Parasites found were Trichostrongylus spp., Trichuris spp. and Eimeria spp. The infestation rate varied markedly. Three percent of the hares were culture positive for tularemia, 93% were seropositive for EBHS and 58% were seropositive for Treponema spp. Conclusions: The overall health status of the East Austrian brown hare population is good. The occurrence of tularemia is important regarding human health, but does not seem to play a major role in the general health status. The high seroprevalence for Treponema spp. and EBHS shows that the animals have contact with these entities, but seem to cope well. For EBHS this might in part be due to a less virulent strain. Generally, it can be stated that the decline cannot be blamed on diseases circling in this population.

Inclusion Body Hepatitis Caused by Fowl Adenovirus in Broiler Chickens in Japan, 2009–2010

From January 2009 to June 2010, many broiler chicks suddenly died without clinical signs. The mortality rates were from 1.2% to 17.0% in affected flocks. Inclusion body hepatitis (IBH) was detected in 13 prefectures (northern, eastern, western, and southern areas) in Japan. The livers were enlarged and pale. The bursa of Fabricius and thymus had not atrophied. Multifocal necroses of hepatocytes with basophilic intranuclear inclusions were seen in the liver. Eosinophilic intranuclear inclusion bodies in hepatocytes were rare. Focal necrosis of acinar cells with basophilic intranuclear inclusions was found in the pancreas. Basophilic intranuclear inclusion bodies were detected in intact surface epithelial cells of gizzard and epithelial cells of the small intestine. The intranuclear inclusions of liver, pancreas, gizzard, and small intestine were stained positively for immunohistochemistry of fowl adenovirus (FAV) antigen. Ultrastructurally, basophilic intranuclear inclusions consisted of viral particles approximately 70 nm in diameter and arranged in a crystalline array. FAV was isolated from the liver of chickens affected with IBH. The serotype of most isolates was 2. This study suggests that IBH produced by FAV is epidemic in broiler chicks in Japan and that the present cases occurred as the primary disease without the association of infectious bursal disease virus or chicken anemia virus.

Experimental Infection of Ground Squirrels (Spermophilus tridecemlineatus) with Monkeypox Virus

A proposed new small-animal (rodent) model for studying the pathogenesis and treatment of severe orthopoxvirus infections is described. Thirteen-lined ground squirrels (Spermophilus tridecemlineatus) were infected intraperitoneally and intranasally with monkeypox virus (MPXV). A fulminant illness developed in all animals, and they died 6-9 days after infection. Virus was cultured from the blood and oropharynx several days before death; at necropsy, all of the organs tested contained relatively high titers of MPXV. The major pathologic findings were in the liver, which showed centrilobular necrosis, steatosis, and basophilic inclusion bodies in hepatocytes. Splenic necrosis was also observed, as well as interstitial inflammation in the lungs. The pathologic features of MPXV in ground squirrels are similar to that described with MPXV in macaques and severe variola (smallpox) virus infection in humans.

Cattle mortality in the Thane district, India: a study of cause/effect relationships.

An unexpected mortality of more than 300 cattle was investigated near a metal recovery factory located in a rural area of the Thane district of India. The factory was engaged in reclaiming lead, aluminum, tin, and zinc from discarded lead storage batteries and soft drink cans. The environmental samples (soil, leaves, grass, slag, water, and sediment), human blood and hair and animal samples (blood, urine, peritoneal fluid, liver, kidney, cow dung, ribs, and femur), collected for analysis revealed toxic levels of lead, cadmium, and chromium. Clinical examination of factory workers and school children revealed cough, fever, gastric problems, abdominal pain, skin lesions (scabies), and blue line on gums. Histopathological examination of animal tissues revealed chronic pathology with lead inclusion bodies in hepatocytes and renal tubules. Based on environmental, clinical, analytical, and histopathological observations, the mortality has been attributed to toxic levels of metals in the body and the malnourished status of the animals.

Pathological studies in broiler chicks fed aflatoxin or ochratoxin and inoculated with inclusion body hepatitis virus singly and in concurrence.

Day-old broiler chicks found negative for maternal antibodies against inclusion body hepatitis (IBH) virus by agar gel precipitation test and viral antigen in cloacal swabs by dot enzyme immunoassay were divided into 6 groups of 20 chicks each. Group A was fed aflatoxin B1 at 1.25 ppm from 3 to 38 days of age; group O was fed ochratoxin A at 0.5 ppm from 3 to 38 days of age; group V was inoculated with 1 ml of IBH virus of titre log10 6.5 EID50 per 0.2 ml. Groups AV and OV were given aflatoxin B1 and ochratoxin A, respectively, and also infected with the virus. Group C served as control. There was mild enlargement and paleness of the liver up to 18 days post inoculation in group V; there were no lesions in group A; and there was gradual enlargement of the kidneys from 10 days post feeding of mycotoxin onwards in group O. In the combined groups AV and OV the gross lesions were slightly more severe. In group V, varying degrees of degenerative histopathological changes, congestion and haemorrhages were seen particularly in the liver, followed by the kidneys, bursa, spleen, myocardium and lungs, along with intranuclear inclusion bodies in the hepatocytes, mostly in the early stages of infection. Similar microscopic changes, but without inclusion bodies, were seen in groups A and O and the changes were pronounced in the later stages. In group O, the kidney lesions were more pronounced than the liver lesions. In the concurrently infected groups, AV and OV, the changes were similar but slightly more marked than in the corresponding individual groups. Inclusion bodies in hepatocytes were more frequent, more prominent and appeared earlier in the concurrent groups.

Duck Viral Enteritis in Domestic Muscovy Ducks in Pennsylvania

Duck viral enteritis (DVE) outbreaks occurred at two different locations in Pennsylvania in 1991 and 1992. In the first outbreak, four ducks died out of a group of 30 domestic ducks; in the second outbreak, 65 ducks died out of a group of 114 domestic ducks, and 15 domestic geese died as well. A variety of species of ducks were present on both premises, but only muscovy ducks (Cairina moschata) died from the disease. On necropsy, gross lesions included hepatomegaly with petechial hemorrhages, petechial hemorrhages in the abdominal fat, petechial hemorrhages on the epicardial surface of the heart, and multifocal to coalescing areas of fibrinonecrotic material over the mucosal surface of the trachea, esophagus, intestine, and cloaca. Histologically, the liver had random multifocal areas of necrosis and eosinophilic intranuclear inclusion bodies in hepatocytes. DVE virus was isolated and identified using muscovy duck embryo fibroblast inoculation and virus neutralization.

Pathogenicity studies with a strain of fowl adenovirus serotype 8 (VRI-33) in chickens.

Experiments were undertaken to study the pathogenesis of VRI-33, a strain of fowl adenovirus serotype 8 isolated from the liver of a broiler chicken with inclusion body hepatitis. A 30% death rate resulted from oral infection of one-day-old specific pathogen free chickens with 10(6) plaque forming units of VRI-33. Chickens 10, 14, 21 and 28 days of age did not die following infection via natural routes but there were some motalities following infection via parenteral routes. Immunodepression by neonatal cyclophosphamide treatment, followed by infection with VRI-33 via non-parenteral routes, caused varying degrees of hepatitis with basophilic intranuclear inclusion bodies in hepatocytes. The mortality rate of cyclophosphamide-treated, VRI-33 infected chickens, was not significantly altered by post-infection temperature stress. Infection with infectious bursal disease virus, followed by infection with VRI-33 via natural routes at 14 days of age, was not associated with mortalities.

Ultrastructure of intracytoplasmic inclusion bodies in hepatocytes of griseofulvin-treated mice. Fibrils with myelin figure and rod-like fibrils.

Intracytoplasmic eosinophilic inclusion bodies light-microscopically identical to human alcoholic hyalin were produced in hepatocytes of griseofulvin-treated mice. Electron-microscopic investigation on these inclusion bodies revealed two morphologically different fibrils. The most frequently observed fibrils in liver tissues appeared straight or slightly curved in longitudinal sections and granular in cross sections. High-power electron micrographs on these fibrils revealed that each fibril consisted of parallel-running fine filaments of about 1.25 nm thick which were separated by spaces of about 1.25 nm. The ultrastructural appearance of these fibrils resembled that of myelin sheath in longitudinal sections. They usually measured from 13 to 32 nm wide and up to 0.9 micron long. Fibrils with larger diameter of up to 62 nm were infrequently encountered. Rod-like fibrils ultrastructurally similar to those seen in human alcoholic hyalin were also found in isolated materials. All these fibrils were isolated in the same fraction by a slight modification of a discontinuous sucrose gradient ultracentrifugation and a two phase polymer centrifugation. The present observation suggests that intracytoplasmic inclusion bodies produced in this animal model are composed of morphologically different fibrils. These different fibrils could share the same origin.

Intracytoplasmic Inclusion Bodies in Hepatocytes During Prolonged Extracorporeal Circulation

Hyaline inclusion bodies appeared in hepatocytes of acutely congested livers produced by prolonged extracorporeal circulation. The inclusions were positive by PAS, PTAH and Ziehl-Neelsen staining and were colored light green to green by Masson's trichrome staining. In methylene blue-stained sections, they were divided into two groups, pale to light blue (Type I) and dark blue (Type II). By electron microscopy, small vacuolar structures which contained small amounts of flocculent material appeared near the vascular pole of hepatocytes at early stages of the experiment. With lapse of time, they became larger and had compact amorphous material. These inclusions corresponded to the Type I inclusions seen by light microscopy. By electron microscopic cytochemistry, these inclusions were usually positive to the acid phosphatase reaction and negative to the DAB reaction. With an increase in serum free hemoglobin, electron-dense inclusions corresponding to the Type II inclusions appeared in the hepatocytes. They were strongly positive to the DAB reaction. Large ones were usually positive to the acid phosphatase reaction in the peripheral narrow rim. Large invaginations of cytoplasmic membranes and large hydropic vacuoles were observed at the late stage of the experiment. They contained frequently fibrin strands or a few erythrocytes. Autophagic vacuoles and myelin figures were also increased in the hepatocytes at the late stage of the bypass circulation.