Cardiopathology in acute African Swine Fever

The present study describes the gross, histopathologic lesions of the heart arising in pigs infected with acute African Swine Fever (ASF) and their biochemical profile. Ten pigs were infected by intramuscular injection of ASF virus (Georgia 2007). Selected heart samples were submitted for histopathological examination and Hematoxylin-Basic Fuchsin-Picric Acid (HBFP) staining. Enzymatic abnormalities were evaluated by measurement of main cardiac markers, whose activity increased during the early stage of infection, with histopathological changes occurring later. Minor myocardial haemorrhages were first observed at four days post infection (dpi), and were noted in all pigs by six dpi. Early vascular response to infection was manifested as increased capillary permeability leading to diapedesis and the retention of blood cells in myocardial tissue. The terminal stage of the disease was characterised by massive haemorrhages caused by the rupture of large vessels. Substantial ischemic areas were detected by HBFP staining at the terminal stages of ASF.

Association of hemophagocytic lymphohistiocytosis with kidney lesions in acute African swine fever virus infection

Glomerulonephritis due to African swine fever (ASF) is well documented. However, there is absence of good understanding of mechanisms involved in the development of pathology development. This study examines glomerulonephritis in association with acute infection induced by II genotype (Georgia 2007) of ASF virus. Taken together, the results of urinary analysis and the renal histological analysis led to the diagnosis of diffuse endocapillary proliferative glomerulonephritis with severe tubular injury associated with acute ASF (Georgia 2007). According to the pathogenesis, we have found that the diffuse endocapillary proliferative glomerulonephritis associated with the acute ASF develops with a delay of one to two days compared to development of hemophagocytic lymphohistiocytosis. The diagnosis of endocapillary proliferative glomerulonephritis confirms the characteristic of pathological changes in the composition of urine and urine sediment. The development of acute proliferative glomerulonephritis begins at 3 dpi, and finished at 4­6 dpi with the development of tubular necrosis. Our study demonstrates local macrophage proliferation. Local proliferation may be an important mechanism for amplifying macrophage-mediated renal injury. We have shown that the development of diffuse acute proliferative glomerulonephritis during ASF does not coincide with the presence of the virus in the blood or kidney tissues, but coincides with the developmental of ASFV derived hemophagocytic lymphohistiocytosis. The development of hemophagocytic lymphonocytosis also begins at least at 2­3 dpi and continues up to the terminal stage of the disease.

Evaluation of hemostaseological status of pigs experimentally infected with African swine fever virus.

African swine fever is a highly contagious hemorrhagic disease of pigs caused by African swine fever virus (ASFV). Hemorrhages are the most frequently reported lesions in acute and subacute forms of ASF. Hemorrhagic lesions are accompanied by impaired hemostasis, which includes thrombocytopenia and changes in the coagulation system. In the present study, experimental infection was conducted to elucidate whether a highly virulent ASFV genotype II circulating in the Trans-Caucasus and Eastern Europe affects the hemostasis of infected pigs. Platelet count changes and platelet size, as well as coagulation parameters were evaluated upon experimental infection. In contrast to other ASFV strains, ASFV genotype II showed a significant decrease in the number of platelets from 3rd dpi onwards. Furthermore, a decrease in platelet size was observed throughout the entire period of experiment. A significant increase in the number of platelet aggregates was observed from the beginning of infection. Unlike other ASFV strains, ASFV genotype II induced a slight shortening of an activated partial thromboplastin time (aPTT) throughout the experiment. Thrombin time (TT) was prolonged from day 5 onwards, whereas no changes in prothrombin time (PT) were found upon infection. The level of d-dimers was permanently higher than in control with a peak on day 3 post-infection. ASFV induced a significant decrease in the level of fibrinogen from day 5 till the end of experiment. Thus, it can be concluded that ASFV genotype II isolated in Armenia affects the hemostasis of infected pigs and causes changes that differ from that of other ASFV strains described previously.

African swine fever virus infection induces tumor necrosis factor alpha production: implications in pathogenesis.

We have analyzed the production of tumor necrosis factor alpha (TNF-alpha) induced by in vitro infection with African swine fever (ASF) virus (ASFV) and the systemic and local release of this inflammatory cytokine upon in vivo infection. An early increase in TNF-alpha mRNA expression was detected in ASFV-infected alveolar macrophages, and high levels of TNF-alpha protein were detected by ELISA in culture supernatants from these cells. When animals were experimentally infected with a virulent isolate (E-75), enhanced TNF-alpha expression in mainly affected organs correlated with viral protein expression. Finally, elevated levels of TNF-alpha were detected in serum, corresponding to the onset of clinical signs. TNF-alpha has been reported to be critically involved in the pathogenesis of major clinical events in ASF, such as intravascular coagulation, tissue injury, apoptosis, and shock. In the present study, TNF-alpha containing supernatants from ASFV-infected cultures induced apoptosis in uninfected lymphocytes; this effect was partially abrogated by preincubation with an anti-TNF-alpha specific antibody. These results suggest a relevant role for TNF-alpha in the pathogenesis of ASF.

Ultrastructural changes related to the lymph node haemorrhages in acute African swine fever.

In order to determine the pathogenic mechanisms involved in lymph node haemorrhages in acute African swine fever (ASF), eight pigs were inoculated with ASF virus, strain Malawi'83. Lymph node haemorrhages were observed from three days post infection (dpi) onwards, coinciding with ASF virus replication in monocytes and macrophages adjacent to stimulated endothelial cells, phagocytic stimulation of capillary and small-vessel endothelial cells, increase in the number of fenestrations of endothelial cells, and endothelial cell loss, as well as clusters of blood cells and necrotic material beneath the endothelium. Vascular lumina were blocked by platelet plugs and fibrin microthrombi. These phenomena became more marked as the disease progressed. At five dpi, virus replication was also found in circulating neutrophils. At seven dpi, lesions were more intense and were accompanied by virus replication in sinus and capillary endothelial cells, and in other cell populations including pericytes, fibroblasts, smooth muscle fibres and reticular cells. The results obtained in this study suggest that lymph node haemorrhages are related to endothelial stimulation and the onset of disseminated intravascular coagulation. Virus replication in vessel wall cells occurs only in the final stages of the disease and plays a secondary role.

African swine fever interference with foot-and-mouth disease infection and seroconversion in pigs.

Initial oral infection of pigs with either highly virulent (L-60) or moderately virulent (DR-2) African swine fever virus (ASFV), followed in 3 days with exposure to foot-and-mouth disease virus (FMDV) (tongue inoculation and contact), failed to cause FMDV infection or seroconversion in 18 of 22 L-60-infected pigs and 13 of 34 DR-2-infected pigs. Of the 13 DR-2-infected pigs remaining free of foot-and-mouth disease (FMD), 2 pigs survived to 24 days without antibody to FMDV, despite constant contact with clinically infected pigs with FMD. Three other DR-2-infected pigs never developed FMD lesions but did develop low levels of antibody to FMDV by day 17. A group of larger pig (in which DR-2 is less virulent) infected with DR-2 and then FMDV had a rapid but suppressed immune response to FMDV. Contact pigs introduced 3 days postinoculation and inoculated with FMDV only all became infected with ASFV by contact and died. This remarkably long lasting 1-way interference with FMD infection during acute and subacute African swine fever was not anticipated. Infection with ASFV may have blocked the initial target cells (possibly dendritic cells) necessary for establishment of FMDV infection.

The role of fibrinolysis in the pathogenesis of the haemorrhagic syndrome produced by virulent isolates of African swine fever virus.

The activity of several proteins involved in fibrinolysis and the morphological changes in the blood vessel walls of pigs infected with highly virulent (Malawi'83) and moderately virulent (Dominican Republic '78-DR'78) ASF virus isolates were determined. Pigs infected with the Malawi'83 virus developed an increased fibrinolytic activity due to high plasma levels of tissue-plasminogen activator (t-PA) of 71.3 +/- 22.8 IU/ml (mean +/- SD), which correlated well with an increased activation of interstitial capillary endothelial cells and high levels of 1150 +/- 73.6 nM of fibrin monomer in the circulation. Animals infected with DR'78 virus, in contrast, showed an inhibition of fibrinolysis in the late stages of disease with almost a 5-fold increase of plasminogen activator inhibitor (PAI) activity of 196.0 AU/ml. These results suggest that activation of the fibrinolytic system in pigs infected with the Malawi'83 virus is probably due to increased formation and deposition of fibrin in the circulation, contributing to an increased bleeding tendency and higher mortality. On the contrary, animals infected with DR'78 virus developed an inhibition of fibrinolysis and thus a reduction in bleeding.

Consumption coagulopathy associated with shock in acute African swine fever.

We studied the evolution of shock using a comprehensive array of haematological tests in pigs infected with the highly virulent strain Malawi '83 (Lilongwe 20/1). A sudden onset of illness was observed between day 5 and 7 after inoculation with development of flush, episodes of epistaxis and melaena. Prior to these clinical signs, initiation of a consumption coagulopathy was demonstrated with loss of antithrombin III and plasminogen activity. Our findings indicate that during infection with this highly virulent strain the development of a consumption coagulopathy precedes and possibly contributes to shock, which results in haemorrhage and death.

Glomerular pathology in surviving pigs experimentally infected with African swine fever virus.

Twelve miniature pigs were inoculated with an attenuated African swine fever virus to study glomerular involvement in surviving pigs. In acute phase, kidneys were severely affected and displayed a glomerular capillary thrombosis with fibrin deposition in vascular lumen, detected by immunofluorescence. Fibrin-positive deposits were progressively cleared between one to three months after infection in surviving pigs. The histological picture in kidneys of surviving pigs, up to one post-infection year, showed a focal and segmental glomerulonephritis with hyalinosis, and IgM and C3 deposition was detected by immunofluorescence. Its pathogeny as an evolutive stage of acute glomerular injury is pointed out.

Abortion in sows experimentally infected with African swine fever virus: pathogenesis studies.

Thirteen sows that were 38 to 92 days pregnant were experimentally infected with an African swine fever (ASF) virus strain of low virulence (Dominican Republic isolate). Seven of 11 sows that were not killed had aborted. The pathogenesis of the abortions was studied, using virus isolation, tissue immunofluoresence, and histopathologic techniques. African swine fever virus was recovered from 179 of 1,329 (13.5%) fetal tissues tested. The 3 fetal tissues most frequently yielding virus were the fetal placenta, amniotic fluid, and fetal heart blood. Virus was not recovered from fetal tissues obtained from 2 of the aborting sows. Direct immunofluorescent microscopy for ASF viral antigen was done on approximately 1,175 fetal tissues. Although brightly fluorescing cells were common in maternal tissues, specific immunofluorescence was present in only placental tissues from 2 sows. Microscopic lesions in fetal tissues were inconsistent and included mild focal placentitis, mild heptic degeneration and necrosis, and mild interstitial pneumonia. These changes were not considered to be sufficiently specific to have diagnostic significance. In marked contrast to these changes in the fetal tissues, maternal tissues had high titers of virus, with marked necrosis of lymphoid tissues, and contained many cells with ASF viral antigen. We conclude that specific diagnosis of abortion resulting from ASF infection should, therefore, be based on examination of maternal tissues, rather than fetal tissues. The pregnancy failure seems to result from the effects of the virus infection on the dam more so than from direct viral damage to the placenta or fetus.

Abortion in sows experimentally infected with African swine fever virus: clinical features.

Thirteen sows between 38 and 92 days of pregnancy were exposed oronasally to African swine fever virus (1979 Dominican Republic isolate) to determine whether infection would affect their reproductive performance. Abortions occurred in sows in all stages of pregnancy. Of 11 sows, 7 aborted 5 to 8 (6.1 +/- 0.90) days after inoculation or 0 to 3 (1.4 +/- 0.98) days after fever development. Abortions occurred soon after clinical signs of infection first developed and viremia titers peaked. Swelling of the vulva and treading preceded abortion in some sows. Ecchymotic placental and petechial cutaneous hemorrhages were present in 6 of 13 and 9 of 13 litters, respectively. African swine fever was transmitted to 2 healthy barrows by feeding fetal tissues collected from 1 inoculated sow. All inoculated sows not killed, died, precluding further study of neonates from infected sows.

[Pathomorphological changes in African swine fever in an African country]. / Patomorfologicni promeni pri afrikanskata chuma po svinete v edna afrikanska durzhava.

Studies were carried out on morphologic changes in 27 pigs experimentally infected with a virulent virus of the African swine fever and in 61 pigs that has spontaneously developed the acute form of the disease. A description and 11 colour plates are given of the characteristic gross and microscopic lesions, consisting of strongly mainfested hemorrhagic diathesis, two-to fourfold enlargement of the spleen, and two-to threefold enlargement of the stomach, liver, mesenterial, kidney, and sternal lymph nodes. The digestive tract was shown to be thoroughly involved, most pronounced being the changes in the stomach and the large intestine. It was demonstrated that the basic histologic changes were localized in the central lymphoreticular system--spleen and lymph nodes, consisting in dystrophic and necrotic lesions of the lymphocytes (karyorrhexis and karyopycnosis), the reticular cells, and the cell elements of the blood vessels. A comparison is made with the morphologic changes in the case of classic swine fever, stating the most essential differences between the lesions in the two forms of the disease.