Virulence, pathology, and pathogenesis of Pteropine orthoreovirus (PRV) in BALB/c mice: Development of an animal infection model for PRV

Figures

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  Fig 1. Changes in the body weight (A) and survival rate (B) of BALB/c mice intranasally inoculated with 1.0 × 103–1.0 × 106 PFU of PRV-MB and the control mice. The mice infected with each dose of PRV-MB are shown (5 mice per group). Error bars in A indicate the standard error.

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  Fig 2. Viral RNA loads in each organ and the blood (A) and infectious virus titers in each organ (B) of BALB/c mice infected with PRV-MB. (A) The organs and blood samples were obtained from the PRV-MB-1.0×105 PFU mice on the 5th or 6th DPI (5 mice per group). The left and right panels indicate the viral RNA loads in the organs and the blood of the mice, respectively. (B) The organs were obtained from the PRV-MB-1.0×106 PFU mice on the 4th DPI (2 mice per group). UDL, under detection limit.

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  Fig 3. Histopathological findings of the lungs of BALB/c mice infected with PRV-MB. The lungs were obtained from PRV-MB-1.0×106 PFU mice on the 4th DPI. H&E staining (A, left panels, and B) and IHC analysis with an OCP antibody (A, middle panels) or with NRS (A, right panels) were performed. The H&E staining and IHC analysis with an OCP antibody or with NRS of the lung at 200× magnification (A, upper panels), of a bronchiole at 1000× magnification (A, middle panels), and of an alveolus at 1000× magnification (A, lower panels) are shown. The black boxes in the upper panels indicate the necrotic lesion, which was positive for PRV-MB antigen (A, upper-middle panel). These areas are shown at higher magnification in the middle panels. The black arrows in the middle panels indicate the area of bronchiolar epithelial cell necrosis. The black dotted boxes in the upper panels indicate the PRV-MB antigen-positive lesion in the alveolar area and the images are enlarged in the lower panels (A, lower-middle panel). The PRV-MB antigen-positive lesion did not show a positive reaction in the IHC analysis, in which NRS was used instead of the OCP antibody (A, right panels). The scale bars in A (upper…

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  Fig 4. Temporal changes in the viral RNA loads in the lungs of the BALB/c mice infected with PRV-MB. The lungs were obtained from PRV-MB-1.0×103 mice or PRV-MB-1.0×105 PFU mice on the 1st, 3rd, and 5th DPI (5 mice per group for each day). Error bars indicate the standard error. N.S., not significant. ***, statistically significant (p < 0.0001).

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  Fig 5. Histopathological findings in the lungs of BALB/c mice infected with PRV-MB on the 1st, 3rd, and 5th DPI. The lungs were obtained from the PRV-MB-1.0×103 PFU mice (A) and the PRV-MB-1.0×105 PFU mice (B) on the 1st, 3rd, and 5th DPI. IHC analysis of the lungs was performed using the OCP antibody for detection of PRV antigen. The right panels show enlarged views of the areas of interesting lesions (black squares in the left panels). The magnification levels of the left and right panels are 200× and 1000×, respectively. Black arrows indicate the pneumocytes that were positive for PRV-MB antigen. The scale bars in the left and right panels indicate 200 μm and 20 μm, respectively.

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  Fig 6. Histopathological findings in the terminal bronchiole of the BALB/c mice infected with a lethal dose of PRV-MB on the 1st DPI. The lungs from the PRV-MB-1.0×105 PFU mice were obtained on the 1st DPI. The lungs were H&E stained (left panels) and tested immunohistochemically using the OCP antibody (right panels). The lower panels show enlarged views of areas of interesting lesions (black squares in the upper panels). The black arrows indicate necrotic cells and cellular debris. The magnification levels of the upper and lower panels are 200× and 1000×, respectively. The scale bars in the upper and lower panels indicate 200 μm and 20 μm, respectively.

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  Fig 7. Changes in the body weight (A) and survival rate (B) in BALB/c mice challenged with a lethal dose of PRV-MB after non-lethal PRV-MB infection. BALB/c mice were pre-inoculated with a non-lethal dose (1.0 × 103 PFU) of PRV-MB or with mock solution, followed by infection with 1.0 × 105 PFU of PRV-MB on the 35th day after the first inoculation (5 mice per group). Error bars indicate the standard error.

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  Fig 8. Changes in the survival rate (upper panels) and body weight (lower panels) in BALB/c mice treated with antiserum after lethal PRV-MB infection. The PRV-MB-1.0×105 PFU mice were treated with the intraperitoneal administration of antiserum once daily during periods surrounded by the gray boxes (5 mice per group). Antiserum with a PRV-MB-specific serum neutralizing antibody titer of 1:10,240 was diluted 4-fold with PBS. The volume of the antiserum administered daily was 100 μL. The treatment was initiated at just after 1 h after the virus challenge (B) or from the 1st (C), 2nd (D), 3rd (E), and 4th (F) DPI. The control mice were administered the same volume of control serum, PRV antibody negative mouse serum diluted with PBS (A). Error bars indicate the standard error.

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  Fig 9. Viral RNA loads in organs and the blood (A) and infectious PRV titers in the organs (B) of BALB/c mice infected with PRV-Samal-24. (A) The organs and the blood samples were obtained from the PRV-Samal-24-1.0×105 PFU mice on the 5th DPI (5 mice per group). Four PRV-Samal-24-1.0×105 PFU mice were sacrificed on the 5th DPI due to their reaching the humane endpoint. Another mouse was found dead on the 5th DPI; therefore, a blood sample could not be obtained. Therefore, the viral RNA loads in each organ and those in the blood were determined for 5 and 4 mice, respectively. The left and right panels indicate the viral RNA in each organ and the blood of the PRV-Samal-24-1.0×105 PFU mice, respectively. (B) The organs were obtained from the PRV-Samal-24-1.0×106 PFU mice on the 4th DPI (2 mice per group). UDL, under detection limit.