Antiviral activity of brequinar against African swine fever virus infection in vitro

Research Areas: VirologyAfrican swine fever virus (ASFV) is a double-stranded DNA virus that causes an acute and hemorrhagic disease in domestic swine, resulting in significant economic losses to the global porcine industry. The lack of vaccines and antiviral drugs highlights the urgent need for antiviral studies against ASFV. Here, we report that brequinar (BQR), which is a specific inhibitor of dihydroorotate dehydrogenase, robustly inhibits ASFV replication in Vero cells, as well as in porcine macrophages. We demonstrate that BQR exerts its antiviral activity in a dosedependent manner through the depletion of pyrimidine pool. Although BQR does not affect the synthesis of an early viral protein, pI215L, the synthesis of late viral proteins, p17 and p72, is suppressed in the presence of BQR. We also show that BQR is able to induce cellular antiviral response in ASFV-infected macrophages by enhancing the expression of interferon-stimulated genes. Taken together, our study reveals that targeting nucleotide biosynthesis represents a promising strategy for developing antiviral agents against ASFV.info:eu-repo/semantics/publishedVersio

Pathology of porcine peripheral white blood cells during infection with African swine fever virus

<p>Abstract</p> <p>Background</p> <p>African swine fever virus (ASFV) is the causative agent of African swine fever (ASF) that is the significant disease of domestic pigs. Several studies showed that ASFV can influence on porcine blood cells in vitro. Thus, we asked ourselves whether ASFV infection results in changes in porcine blood cells in vivo. A series of experiments were performed in order to investigate the effects of ASFV infection on porcine peripheral white blood cells. Nine pigs were inoculated by intramuscular injection with 10⁴50% hemadsorbing doses of virus (genotype II) distributed in Armenia and Georgia. The total number of fifteen cell types was calculated during experimental infection.</p> <p>Results</p> <p>Although band-to-segmented neutrophils ratio became much higher (3.5) in infected pigs than in control group (0.3), marked neutropenia and lymphopenia were detected from 2 to 3 days post-infection. In addition to band neutrophils, the high number of other immature white blood cells, such as metamyelocytes, was observed during the course of infection. From the beginning of infection, atypical lymphocytes, with altered nuclear shape, arose and became 15% of total cells in the final phase of infection. Image scanning cytometry revealed hyperdiploid DNA content in atypical lymphocytes only from 5 days post-infection, indicating that DNA synthesis in pathological lymphocytes occurred in the later stages of infection.</p> <p>Conclusion</p> <p>From this study, it can be concluded that ASFV infection leads to serious changes in composition of white blood cells. Particularly, acute ASFV infection in vivo is accompanied with the emergence of immature cells and atypical lymphocytes in the host blood. The mechanisms underlying atypical cell formation remain to be elucidated.</p

A new microtubule-stabilizing agent shows potent antiviral effects against African swine fever virus with no cytotoxicity

© 2021 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.African swine fever virus (ASFV) is the causal agent of a fatal disease of domestic swine for which no effective antiviral drugs are available. Recently, it has been shown that microtubule-targeting agents hamper the infection cycle of different viruses. In this study, we conducted in silico screening against the colchicine binding site (CBS) of tubulin and found three new compounds with anti-ASFV activity. The most promising antiviral compound (6b) reduced ASFV replication in a dose-dependent manner (IC50 = 19.5 μM) with no cellular (CC50 > 500 μM) and animal toxicity (up to 100 mg/kg). Results also revealed that compound 6b interfered with ASFV attachment, internalization and egress, with time-of-addition assays, showing that compound 6b has higher antiviral effects when added within 2-8 h post-infection. This compound significantly inhibited viral DNA replication and disrupted viral protein synthesis. Experiments with ASFV-infected porcine macrophages disclosed that antiviral effects of the compound 6b were similar to its effects in Vero cells. Tubulin polymerization assay and confocal microscopy demonstrated that compound 6b promoted tubulin polymerization, acting as a microtubule-stabilizing, rather than a destabilizing agent in cells. In conclusion, this work emphasizes the idea that microtubules can be targets for drug development against ASFV.This work of E. A., A. H., and H. Z. was supported by the RA MESCS Science Committee, Armenia [grant number 19YR-1F039]; the work of F. F. was supported by the FCT – Fundação para a Ciência e a Tecnologia, Portugal [grant number UIDB/00276/2020].info:eu-repo/semantics/publishedVersio

Intracellular African swine fever virus DNA remains unmethylated in infected Vero cells

Aim: Sequence-specific CpG methylation of eukaryotic promoters is an important epigenetic signal for long-term gene silencing. We have now studied the methylation status of African swine fever virus (ASFV) DNA at various times after infection of Vero cells in culture. Methods & results: ASFV DNA was detectable throughout the infection cycle and was found unmethylated in productively infected Vero cells as documented by bisulfite sequencing of 13 viral DNA segments. Conclusion: ASFV DNA does not become de novo methylated in the course of infection in selected segments spread across the entire genome. Thus DNA methylation does not interfere with ASFV genome transcription. Lack of de novo methylation has previously been observed for free intracellular viral DNA in cells permissively infected with human adenoviruses, with human papillomaviruses and others

Evidence of hemolysis in pigs infected with highly virulent African swine fever virus

Aim: The research was conducted to understand more profoundly the pathogenetic aspects of the acute form of the African swine fever (ASF). Materials and Methods: A total of 10 pigs were inoculated with ASF virus (ASFV) (genotype II) in the study of the red blood cells (RBCs), blood and urine biochemistry in the dynamics of disease. Results: The major hematological differences observed in ASFV infected pigs were that the mean corpuscular volume, mean corpuscular hemoglobin, and hematocrits were significantly decreased compared to controls, and the levels of erythropoietin were significantly increased. Also were detected the trends of decrease in RBC count at terminal stages of ASF. Analysis of blood biochemistry revealed that during ASF development, besides bilirubinemia significantly elevated levels of lactate dehydrogenase, and aspartate aminotransferase were detected. Analysis of urine biochemistry revealed the presence of bilirubinuria, proteinuria during ASF development. Proteinuria, especially at late stages of the disease reflects a severe kidney damage possible glomerulonefritis. Conclusion: The results of this study indicate the characteristics of developing hemolytic anemia observed in acute ASF (genotype II)