SPF rabbits infected with rabbit hepatitis E virus isolate experimentally showing the chronicity of hepatitis

Abstract

This study focused on investigating the pathogenesis seen in specific-pathogen-free (SPF) rabbits following infection with a homologous rabbit HEV isolate (CHN-BJ-rb14) and comparing it to that seen following infection with a heterologous swine genotype 4 HEV isolate (CHN-XJ-SW13). Three of the four animals inoculated with the homologous rabbit HEV became infected, exhibiting an intermittent viremia, obvious fluctuations of liver function biomarkers alanine aminotransferase (ALT) and aspartate aminotransferase (AST), and persistent fecal virus shedding throughout the nine month study. In addition, liver histopathology showed both chronic inflammation and some degree of fibrosis. Both positive and negative-stranded HEV RNA and HEV antigen expression were detected in liver, brain, stomach, duodenum and kidney from the necropsied rabbits. Inflammation of extrahepatic tissue (duodenum and kidney) was also observed. Three of the four rabbits inoculated with the heterologous genotype 4 swine HEV also became infected, showing similar levels of anti-HEV antibody to that generated following infection with the homologous virus isolate. The duration of both viremia and fecal shedding of virus was however shorter following infection with the heterologous virus and there was no significant elevation of liver function biomarkers. These results suggest that rabbit HEV infection may cause more severe hepatitis and prolong the course of the disease, with a possible chronic trend of hepatitis in SPF rabbits.

Figure 1. Dynamic seroconversion of anti-HEV, HEV RNA,ALT and AST observed in rabbits.

(A) Rabbit C1 in group 1 inoculated with sterile PBS. (B) Rabbit R1 in group 2 inoculated with rabbit HEV strain. (C) Rabbit R3 in group 2 1inoculated with rabbit HEV strain. (D) Rabbit S2 in group 3 inoculated with a genotype 4 swine HEV at 0wpi, and rabbit HEV at 25wpi. (E) Rabbit S4 in group 3 inoculated with a genotype 4 swine HEV at 0wpi, and rabbit HEV at 25wpi. Note: ↑ indicates group 3 rabbits were inoculated with rabbit HEV strain at 25wpi (when recovered from initial infection).

Figure 2. Liver histology. A–D (H & E stain, original magnification, ×10), E–G (Masson’s trichrome stain, original magnification, ×10), H–I (Immunohistochemistry, original magnification, ×40).

(A) Liver section from a control rabbit with no visible pathological signs of HEV infection. (B)-(C) Lymphocytes distributed focal or scattered in hepatic lobule, the inflammatory cells gathered along blood vessel walls. (D) Chronic inflammatory cells infiltrate the portal area, blood vessel walls thickening associated with fibrosis, local hyaline degeneration. (E) No histopathological changes with minimal staining limited to areas immediately adjacent to portal structures. (F) Artery wall thickening associated with moderate to severe fibrosis. (G) More advanced portal and periportal fibrosis with short fibrous septa. (H) Negative immunohistochemistry result for HEV antigen in liver sections from the control rabbits. (I) Positive results for HEV antigen in liver sections of experimental groups.

Figure 3. Extrahepatic tissue histology. A–E (Immunohistochemistry, original magnification, ×40), F–J (H & E stain, original magnification, ×10).

(A) Negative immunohistochemistry result for HEV antigen in extrahepatic tissue sections from the control rabbits. (B)–(E) Positive results for HEV antigen in brain, stomach, duodenum and kidney. (F) Duodenum section from a control rabbit with no visible pathological signs of inflammation. (G) A large number of lymphocytes infiltrate mucosal interstitial, focal lymph follicles formed in duodenum sections. (H) Kidney section from a control rabbit with no visible pathological signs of HEV infection. (I) Multifocal lymphocytes and mononuclear cells infiltrate in renal interstitial.