Pathogenic characteristics of three genotype II porcine reproductive and respiratory syndrome viruses isolated from China

Abstract

Background: We examined differences in pathogenicity in pigs from China that had been experimentally infected with porcine reproductive and respiratory syndrome virus (PRRSV).

Methods: We compared pathogenic characteristics of a field isolate (GX-1/2008F), two PRRSV isolates (HN-1/2008, YN-1/2008) propagated in cells, and GX-1/2008F that had been propagated in cells (GX-1/2008). The clinical courses, along with humoral and cell-mediated responses, were monitored for 21 days post-infection (DPI). Animals were sacrificed and tissue samples used for gross pathological, histopathological and ultrastructure examination.

Results: At 2-3 DPI, animals infected with cell-propagated viruses exhibited signs of coughing, anorexia and fever. However their rectal temperature did not exceed 40.5°C. Viremia was detectable as early as 3 DPI in animals infected with HN-1/2008 and YN-1/2008. Animals inoculated with GX-1/2008F displayed clinical signs at 6 DPI; the rectal temperature of two animals in this group exceeded 41.0°C, with viremia first detected at 7 DPI. Seroconversion for all challenged pigs, except those infected with GX-1/2008, was seen as early as 7 DPI. All of these pigs had fully seroconverted by 11 DPI. All animals challenged with GX-1/2008 remained seronegative until the end of the experiment. Innate immunity was inhibited, with levels of IFN-α and IL-1 not significantly different between control and infected animals. The cytokines IFN-γ and IL-6 transiently increased during acute infection. All virus strains caused gross lesions including multifocal interstitial pneumonia and hyperplasia of lymph nodes. Inflammation of the stomach and small intestine was also observed. Lesions in the group infected with GX-1/2008F were more serious than in other groups. Transmission electron microscopy revealed that alveolar macrophages, plasmacytes and lymphocytes had fractured cytomembranes, and hepatocytes had disrupted organelles and swollen mitochondria.

Conclusions: The pathogenicity of the PRRSV field isolate became attenuated when propagated in MARC-145 cells. Tissue tropism of highly pathogenic strains prevailing in China was altered compared with classical PRRSV strains. The observed damage to immune cells and modulation of cytokine production could be mechanisms that PRRSV employs to evade host immune responses.

Figure 1

Mean rectal temperatures following infection with HN-1/2008, YN-1/2008 and GX-1/2008. Rectal temperatures equal to or above 40.0°C were defined as fever. Fever lasting 3 or more days was defined as illness. There were an average of 6–7 days of high fever for pigs in groups B, C and E, and 2 days of high fever for pigs in group D.

Figure 2

Mean anti-PRRSV antibody levels. Development of PRRSV specific antibodies was monitored throughout the experiment following infection and reported as S/P ratios. An S/P ratio greater than 0.4 was considered positive.

Figure 3

Mean pro-inflammatory cytokines levels. The development of pro-inflammatory cytokines was monitored throughout the experiment after infection. Mean levels for IFN-α (a), IL-1 (b), IFN-γ (c) and IL-6 (d) are presented.

Figure 4

Histological analysis. Group E samples were examined by H&E staining at 21 DPI. (a) Alveolar septal thickening in macrophages indicated by a thick arrow. Hypertrophy of type II pneumocytes is indicated by a thin arrow. (b) Splenic corpuscles were diminished and the number of lymphocytes were decreased. (c) Swelling or destabilization in the structure of hepatocytes. (d) Kidney tubules were damaged. (e) The spaces between myocardial fibers were widened. (f) Gastric glands cells were swollen. (g) Necrosis and shedding of epithelial cells of the small intestine mucosa. (h) The number of lymphocytes were decreased (thin arrow) in mesenteric lymph nodes, with depletion of germinal centers (thick arrow).

Figure 5

TEM ultrastructural analysis. Group E animals were examined at 21 DPI. (a) The nuclear envelope of alveolar macrophages lacked integrity. (b) Autophagosomes were seen in alveolar macrophages. (c) Lymphocytes in mesenteric lymph nodes were decreased. (d) The chromatin in splenic lymphocytes had migrated towards the margin of cells. (e) Numbers of splenic lymphocytes were decreased. (f) Mitochondria in liver cells were swollen, with the number of mitochondrial crista decreased or absent. (g) The majority of organelles in hepatocytes were decayed. (h) Nuclei of glomerular vascular endothelial cells were pyknotic.