scholarly journals Small interfering RNAs are highly effective inhibitors regarding Crimean-Congo hemorrhagic fever virus replication in vitro

2020 ◽  
Author(s):  
Fanni Földes ◽  
Mónika Madai ◽  
Henrietta Papp ◽  
Gábor Kemenesi ◽  
Brigitta Zana ◽  
...  
Keyword(s):  
Rna Interference ◽  
Hemorrhagic Fever ◽  
Fever Virus ◽  
World Health ◽  
Dependent Manner ◽  
Small Interfering Rnas ◽  
Crimean Congo Hemorrhagic Fever ◽  
Hemorrhagic Fever Virus ◽  
Antiviral Therapies

AbstractCrimean-Congo hemorrhagic fever virus (CCHFV) is one of the prioritized diseases of World Health Organization, considering its potential to create a public health emergency and more importantly, the absence of efficacious drugs and/or vaccines regarding treatment. The highly lethal nature characteristic to CCHFV restricts research to BSL-4 laboratories, which complicates effective research and developmental strategies. In consideration of antiviral therapies, RNA interference can be used to suppress viral replication by targeting viral genes. RNA interference uses small interfering RNAs (siRNAs) to silence genes. The aim of our study was to design siRNAs that inhibit CCHFV replication and can serve as a basis for further antiviral therapies. A549 cells were infected with CCHFV after transfection with the siRNAs. Following 72 hours, nucleic acid from the supernatant was extracted for Droplet Digital PCR analysis. Among the investigated siRNAs we identified four effective candidates against all three segments of CCHF genome: one for the S and M segments, whilst two for the L segment. Consequently, blocking any segment of CCHFV leads to changes in the virus copy number that indicates an antiviral effect of the siRNAs in vitro. The most active siRNAs were demonstrated a specific inhibitory effect against CCHFV in a dose-dependent manner. In summary, we demonstrated the ability of specific siRNAs to inhibit CCHFV replication in vitro. This promising result can be used in future anti-CCHFV therapy developments.

scholarly journals Small Interfering RNAs Are Highly Effective Inhibitors of Crimean-Congo Hemorrhagic Fever Virus Replication In Vitro

Molecules ◽  
2020 ◽  
Vol 25 (23) ◽  
pp. 5771
Author(s):  
Fanni Földes ◽  
Mónika Madai ◽  
Henrietta Papp ◽  
Gábor Kemenesi ◽  
Brigitta Zana ◽  
...  
Keyword(s):  
Rna Interference ◽  
A549 Cells ◽  
Hemorrhagic Fever ◽  
Fever Virus ◽  
World Health ◽  
Small Interfering Rnas ◽  
Crimean Congo Hemorrhagic Fever ◽  
Hemorrhagic Fever Virus ◽  
Antiviral Therapies

Crimean-Congo hemorrhagic fever virus (CCHFV) is one of the prioritized diseases of the World Health Organization, considering its potential to create a public health emergency and, more importantly, the absence of efficacious drugs and/or vaccines for treatment. The highly pathogenic characteristic of CCHFV restricts research to BSL-4 laboratories, which complicates effective research and developmental strategies. In consideration of antiviral therapies, RNA interference can be used to suppress viral replication by targeting viral genes. RNA interference uses small interfering RNAs (siRNAs) to silence genes. The aim of our study was to design and test siRNAs in vitro that inhibit CCHFV replication and can serve as a basis for further antiviral therapies. A549 cells were infected with CCHFV after transfection with the siRNAs. Following 72 h, nucleic acid from the supernatant was extracted for RT Droplet Digital PCR analysis. Among the investigated siRNAs we identified effective candidates against all three segments of the CCHF genome. Consequently, blocking any segment of CCHFV leads to changes in the virus copy number that indicates an antiviral effect of the siRNAs. In summary, we demonstrated the ability of specific siRNAs to inhibit CCHFV replication in vitro. This promising result can be integrated into future anti-CCHFV therapy developments.

Hepatocyte pathway alterations in response to in vitro Crimean Congo hemorrhagic fever virus infection

2014 ◽  
Vol 179 ◽  
pp. 187-203 ◽  
Author(s):  
Christophe Fraisier ◽  
Raquel Rodrigues ◽  
Vinh Vu Hai ◽  
Maya Belghazi ◽  
Stéphanie Bourdon ◽  
...  
Keyword(s):  
Virus Infection ◽  
Hemorrhagic Fever ◽  
Fever Virus ◽  
Crimean Congo Hemorrhagic Fever ◽  
Hemorrhagic Fever Virus

scholarly journals Antagonistic Antiviral Activity between IFN-Lambda and IFN-Alpha against Lethal Crimean-Congo Hemorrhagic Fever Virus In Vitro

PLoS ONE ◽  
2015 ◽  
Vol 10 (2) ◽  
pp. e0116816 ◽  
Author(s):  
Licia Bordi ◽  
Eleonora Lalle ◽  
Claudia Caglioti ◽  
Damiano Travaglini ◽  
Daniele Lapa ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Hemorrhagic Fever ◽  
Fever Virus ◽  
Crimean Congo Hemorrhagic Fever ◽  
Hemorrhagic Fever Virus ◽  
Ifn Alpha ◽  
Ifn Lambda

scholarly journals The Crimean-Congo Hemorrhagic Fever Virus NSm Protein Is Dispensable for Growth In Vitro and Disease in Ifnar-/- Mice

2020 ◽  
Vol 8 (5) ◽  
pp. 775 ◽  
Author(s):  
Stephen R. Welch ◽  
Florine E. M. Scholte ◽  
Jessica R. Spengler ◽  
Jana M. Ritter ◽  
JoAnn D. Coleman-McCray ◽  
...  
Keyword(s):  
Clinical Signs ◽  
Hemorrhagic Fever ◽  
Fever Virus ◽  
Structural Proteins ◽  
Glycoprotein Precursor ◽  
Crimean Congo Hemorrhagic Fever ◽  
Virus Attachment ◽  
Hemorrhagic Fever Virus ◽  
Endoproteolytic Cleavage

Crimean-Congo hemorrhagic fever virus (CCHFV) is a tri-segmented, tick-borne nairovirus that causes disease of ranging severity in humans. The CCHFV M segment encodes a complex glycoprotein precursor (GPC) that undergoes extensive endoproteolytic cleavage, giving rise to two structural proteins (Gn and Gc) required for virus attachment and entry, and to multiple non-structural proteins (NSm, GP160, GP85, and GP38). The functions of these non-structural proteins remain largely unclear. Here, we investigate the role of NSm during infection by generating a recombinant CCHFV lacking the complete NSm domain (10200∆NSm) and observing CCHFV ∆NSm replication in cell lines and pathogenicity in Ifnar-/- mice. Our data demonstrate that the NSm domain is dispensable for viral replication in vitro, and, despite the delayed onset of clinical signs, CCHFV lacking this domain caused severe or lethal disease in infected mice.

scholarly journals Differential Growth Characteristics of Crimean-Congo Hemorrhagic Fever Virus in Kidney Cells of Human and Bovine Origin

Viruses ◽  
2020 ◽  
Vol 12 (6) ◽  
pp. 685
Author(s):  
Katalin Földes ◽  
Touraj Aligholipour Farzani ◽  
Koray Ergünay ◽  
Aykut Ozkul
Keyword(s):  
Cell Lines ◽  
Human Kidney ◽  
Hemorrhagic Fever ◽  
Fever Virus ◽  
Kidney Cells ◽  
Bovine Kidney ◽  
Crimean Congo Hemorrhagic Fever ◽  
Hemorrhagic Fever Virus ◽  
The Impact

Crimean-Congo hemorrhagic fever virus (CCHFV) causes a lethal tick-borne zoonotic disease with severe clinical manifestation in humans but does not produce symptomatic disease in wild or domestic animals. The factors contributing to differential outcomes of infection between species are not yet understood. Since CCHFV is known to have tropism to kidney tissue and cattle play an important role as an amplifying host for CCHFV, in this study, we assessed in vitro cell susceptibility to CCHFV infection in immortalized and primary kidney and adrenal gland cell lines of human and bovine origin. Based on our indirect fluorescent focus assay (IFFA), we suggest a cell-to-cell CCHF viral spread process in bovine kidney cells but not in human cells. Over the course of seven days post-infection (dpi), infected bovine kidney cells are found in restricted islet-like areas. In contrast, three dpi infected human kidney or adrenal cells were noted in areas distant from one another yet progressed to up to 100% infection of the monolayer. Pronounced CCHFV replication, measured by quantitative real-time RT-PCR (qRT-PCR) of both intra- and extracellular viral RNA, was documented only in human kidney cells, supporting restrictive infection in cells of bovine origin. To further investigate the differences, lactate dehydrogenase activity and cytopathic effects were measured at different time points in all mentioned cells. In vitro assays indicated that CCHFV infection affects human and bovine kidney cells differently, where human cell lines seem to be markedly permissive. This is the initial reporting of CCHFV susceptibility and replication patterns in bovine cells and the first report to compare human and animal cell permissiveness in vitro. Further investigations will help to understand the impact of different cell types of various origins on the virus–host interaction.

scholarly journals Independent Inhibition of the Polymerase and Deubiquitinase Activities of the Crimean–Congo Hemorrhagic Fever Virus Full-Length L-Protein

Proceedings ◽  
2020 ◽  
Vol 50 (1) ◽  
pp. 41
Author(s):  
Egor P. Tchesnokov ◽  
Ben A. Bailey-Elkin ◽  
Brian L. Mark ◽  
Matthias Götte
Keyword(s):  
Expression System ◽  
Hemorrhagic Fever ◽  
Fever Virus ◽  
Full Length ◽  
World Health ◽  
Rdrp Activity ◽  
Crimean Congo Hemorrhagic Fever ◽  
L Protein ◽  
Hemorrhagic Fever Virus ◽  
Research Activities

The Crimean–Congo hemorrhagic fever virus (CCHFV) is a segmented negative-sense RNA virus that can cause severe human disease. The World Health Organization (WHO) has listed CCHFV as a priority pathogen with an urgent need for enhanced research activities to develop effective countermeasures. We report on the expression, characterization, and inhibition of the CCHFV full-length L-protein that provides an important tool in this regard. The requirements for high biosafety measures hamper drug discovery and development efforts with infectious CCHFV. Hence, we decided to adopt a biochemical approach that targets the viral RNA-dependent RNA polymerase (RdRp). The CCHFV RdRp activity is part of a multifunctional L protein that is unusually large, with a molecular weight of ~450 kDa. The CCHFV L-protein also contains an ovarian tumor (OTU) domain that exhibits deubiquitinating (DUB) activity. Previous studies have shown that DUB activity interferes with innate immune responses and viral replication. Here, we utilized the baculovirus expression system and generated a full-length CCHFV L protein. RdRp activity was seen in the presence of divalent metal ions, and inhibition of RNA synthesis was demonstrated with nucleotide analogues. The ubiquitin analogue CC.4 inhibits the CCHFV-associated DUB activity of the full-length L protein and the isolated DUB domain to a similar extent. We have finally shown that RdRp and DUB activities are functionally independent. The full-length CCHFV L-protein provides an important tool for the discovery of antiviral agents. High-throughput screening (HTS) campaigns are now feasible. The same enzyme preparations can be utilized to identify polymerase and DUB inhibitors.

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