Brain Infection by Hepatitis E Virus Probably via Damage of the Blood-Brain Barrier Due to Alterations of Tight Junction Proteins

Extrahepatic injury, particularly neurologic dysfunctions such as Guillain-Barré syndrome, neurologic amyotrophy, and encephalitis/meningoencephalitis/myositis were associated with HEV infection, which was supported by both clinical and laboratory studies. Thus, it is crucial to figure out how the virus invades into the central nervous system (CNS). In this study, CNS lesions were determined in rabbits and Mongolian gerbils inoculated with genotype 4 HEV. Junctional proteins were detected in HEV infected primary human brain microvascular cells (HBMVCs). Viral encephalitis associated perivascular cuffs of lymphocytes and microglial nodules were observed in HEV infected rabbits. Both positive- and negative-strand of HEV RNA was detected in brain and spinal cord in rabbits intraperitoneally infected with HEV at 28 dpi (days postinoculation), but not in rabbits gavaged with HEV. HEV ORF2 protein was further examined in both brain and spinal cord sections of infected rabbits, with positive signals located mainly in neural cells and perivascular areas. Ultrastructural study showed thickened and reduplicated basement membranes of capillary endothelium in HEV RNA positive brain tissues. In vitro study showed loss of tight junction proteins including Claudin5, Occludin, and ZO-1 (zonula occludens-1) in HBMVCs inoculated with HEV for 48 h. These findings indicated that disruption of the blood-brain barrier (BBB) might be potential mechanisms of HEV invasion into the CNS. It provides new insights to further study HEV associated neurologic disorders and will be helpful for seeking potential therapeutics for HEV infection in the future

Detection and localization of rabbit hepatitis e virus and antigen in systemic tissues from experimentally intraperitoneally infected rabbits.

Rabbit hepatitis E virus (HEV) is a novel genotype of HEV, and is considered to pose a risk of zoonotic transmission. Research into the systemic distribution of rabbit HEV in rabbits during different periods of infection has rarely been reported. To better understand this virus, we infected rabbits with second-passage rabbit HEV via an intraperitoneal route. After inoculation, the infection showed two types, temporary and constant infection. The detection of HEV RNA in the feces varied with time, and serum antigen correlated with fecal HEV RNA. Viremia only appeared 72 days after inoculation. The rabbits remained antibody negative throughout the experimental period. When HEV was localized, several organs besides the liver were HEV RNA positive. Tissue antigen was observed immunohistochemically in the different cells of various organs, especially in parts of the small intestine and the characteristic rabbit gut-associated lymphoid tissue. These data provide valuable information for future research into the pathogenesis of HEV

Case Report Associated with Aspergillosis and Hepatitis E Virus Coinfection in Himalayan Griffons

This study involved a death which occurred in four Himalayan griffons housed in Beijing zoo, China. Based on pathogen identification and the pathological changes observed, we did characterize the fungi and Hepatitis E virus (HEV) in four dead Himalayan griffons. Pathological changes were severe. Membranous-like material was observed on the surface of the internal organs. Spleen was necrotic. Focal lymphocyte infiltration in the liver and many sunflower-like fungi nodules were evident in the tissues, especially in the kidney. PCR was used to identify the pathogen. Based on the 18SrRNA genomic sequence of known fungi, the results confirmed that all four dead Himalayan griffons were infected with Aspergillus. At the same time the detection of HEV also showed positive results. To the best of our knowledge, this work appears to be the first report of concurrent presence of Aspergillosis and Hepatitis E virus in rare avian species

Detection of Genotype 4 Swine Hepatitis E Virus in Systemic Tissues in Cross-Species Infected Rabbits

<div><p>Increasing evidence demonstrates that hepatitis E virus (HEV) can be transmitted across species. According to previous reports, swine HEV has two genotypes, genotype 3 and 4, and both can infect humans by the fecal-oral route. Thus, it is crucial for the control of HEV zoonotic transmission to evaluate the dynamics of viral shedding and distribution in different tissues during cross-species infection by HEV. In this study, rabbits were infected with genotype 4 swine HEV by the intraperitoneal route. The results showed that HEV RNA not only shed in the feces but also in the saliva of some rabbits during infection with swine HEV. Viremia appeared late after infection, and anti-HEV IgG was not obvious until the appearance of high viremia levels. After the rabbits were euthanized, a histopathological examination showed that the livers developed overt hepatitis accompanied by an elevation of alanine aminotransferase (ALT) and aspartate transaminase (AST). Furthermore, HEV RNA was detected in various tissues, especially in the salivary glands and tonsils. Subsequently, negative-stranded HEV RNA was practiced in tissues with positive HEV RNA, which demonstrated that HEV replicated in the tissues. Next, we harvested additional tissues from the liver, salivary gland, tonsil, spleen, thymus gland, lymph node and intestine, which are known as replication sites of swine HEV. Additionally, we also observed the HEV antigen distributed in the organs above through immunohistochemical staining. These results demonstrate that rabbits could be used as an animal model for researching cross-species infection of genotype 4 HEV. It is also noteworthy that HEV can shed in the saliva and presents the risk of droplet transmission. These new data provide valuable information for understanding cross-species infection by HEV.</p></div

Changes in serum HEV antigen and anti-HEV IgG based on ELISA at various days post-inoculation.

<p>A. Serum HEV antigen was first detected in the T5 rabbit at 35 dpi and was elevated until 49 dpi. Serum HEV antigen was also detected in the T6 and T8 rabbits at 49 dpi. B. Anti-HEV IgG appeared in the T5 rabbit at 42 dpi and was elevated until 49 dpi. Other rabbits had no anti-HEV IgG until the end of the experiment. All rabbits in the control group were designated the “control”.</p

Histopathological analysis of livers and kidneys.

<p>A. There were no gross histopathological lesions in the liver sections from the control group. B. A large number of lymphocytes infiltrated the liver, and the bile ducts proliferated in the portal tracts of the liver in the experimental group. C. No gross lesions were observed in the kidney sections from the control group. D. A large number of lymphocytes infiltrated into the mesenchyme of the renal cortex. H&E staining.</p

HEV ORF2 antigen localization in various tissues from the experimental group detected using immunohistochemical staining.

<p>A. Liver. The positive signal for HEV antigen was detected in hepatocytes and the epithelium of bile ducts. B. Kidney. The positive signal for HEV antigen was detected in the epithelium of the renal tubules. C. Salivary gland. The positive signal for HEV antigen was detected in the duct epithelium of salivary gland. D. Tonsil. The positive signal for HEV antigen was detected in the crypt epithelium and in macrophages in the tonsil. E. Spleen. The positive signal for HEV antigen was detected in macrophages or dendritic cells in the spleen. F. Thymus gland. The positive signal for HEV antigen was detected in thymic epithelial cells in the medulla of the thymus gland. G. Heart. The positive signal for HEV antigen was detected in the myocardium. H. Lung. The positive signal for HEV antigen was detected in the epithelium of the alveoli and bronchioles. I. Pancreas. The positive signal for HEV antigen was detected in the duct epithelium of the pancreas. J. Small intestine. The positive signal for HEV antigen was detected in the mucosal epithelium in the small intestine. K. SR. The positive signal for HEV antigen was detected in the follicle-associated epithelium. L. Appendix. The positive signal for HEV antigen was detected in macrophages or dendritic cells of lymphoid follicles. The positive signal is yellow.</p

HEV ORF2 antigen localization in various tissues from the control group detected using immunohistochemical staining.

<p>No positive signal (yellow) was detected in these tissues of the control group. A-L: liver, kidney, salivary gland, tonsil, spleen, thymus gland, heart, lung, pancreas, small intestine, SR or cecum.</p

Distribution of HEV RNA and negative-stranded RNA in various tissues of rabbits infected with swine HEV at the indicated days post-inoculation.

<p>Distribution of HEV RNA and negative-stranded RNA in various tissues of rabbits infected with swine HEV at the indicated days post-inoculation.</p