Effects of cell culture and laboratory conditions on type 2 dengue virus infectivity.

A full-length infectious cDNA clone (ic) was constructed from the genome of the dengue virus type 2 (DENV-2) Jamaica83 1409 strain, pBAC1409ic, by using a bacterial artifical chromosome plasmid system. Infectious virus was generated and characterized for growth in cell culture and for infection in Aedes aegypti mosquitoes. During construction, an isoleucine to methionine (Ile→Met) change was found at position 6 in the envelope glycoprotein sequence between low- and high-passage DENV-2 1409 strains. In vitro-transcribed genomic RNA of 1409ic with E6-Ile produced infectious virions following electroporation in mosquito cells, but not mammalian cells, while 1409ic RNA with an E6-Met mutation produced virus in both cell types. Moreover, DENV-2 1409 with the E6-Ile residue produced syncytia in C6/36 cell culture, whereas viruses with E6-Met did not. However, in vitro cell culture-derived growth-curve data and in vivo mosquito-infection rates revealed that none of the analysed DENV-2 strains differed from each other.

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Australia Ae. aegypti mosquitoes are susceptible to a highly divergent and sylvatic dengue virus type 2 strain infection but are unlikely to transmit

10.21203/rs.2.18361/v1 ◽

2019 ◽

Author(s):

Paul Pickering ◽

Leon Hugo ◽

Gregor J Devine ◽

John G Aaskov ◽

Wenjun Liu

Keyword(s):

Cell Culture ◽

Dengue Virus ◽

Dengue Fever ◽

Vector Competence ◽

Immunosorbant Assay ◽

Infectious Dose ◽

Fever Patient ◽

A Cell ◽

Divergent Virus

Abstract Background: Humans are the primary hosts of the dengue virus; However, sylvatic cycles of transmission can occur among non-human primates and human encroachment to forested regions can be a source of emergence of new strains. We reported the isolation of a highly divergent and sylvatic DENV-2 strain (QML22) from a dengue fever patient returning Australia from Borneo. The objective of the present study was to evaluate the vector competence of Australian Ae. aegypti mosquitoes for this virus. Methods: Four-day old mosquitoes from two strains of Ae. aegypti from Queensland, Australia, were feed sheep blood meal containing 108 50% cell culture infectious dose per ml (CCID50/ml) of either QML22 or an Australian epidemic DENV serotype 2 strain (QML16) isolated from a dengue fever patient in 2015. Mosquitoes were maintained at 28°C, 75% relative humidity and sampled at 7, 10 and 14 days post-infection (DPI). Live virions in mosquito bodies (abdomen/thorax), legs and wings and saliva expectorates from individual mosquitoes were quantified using a Cell Culture Enzyme-linked Immunosorbant Assay (CCELISA) to determine infection, dissemination and transmission rates. Findings: The infection and dissemination rates of the sylvatic DENV2 strain, QML22, within mosquitoes were significantly lower than that for QML16. While the titres of virus in the bodies of mosquitoes infected with either of these viruses were similar, titres in legs and wings were significantly lower in mosquitoes infected with QML22 at most time points although they reached similar levels by 14 DPI. QML16 was detected in 16% (n = 25) and 28% (n = 25) of saliva expectorates at 10 and 14 DPI, respectively. In contrast, no virus was detected in the saliva expectorates of QML22 infected mosquitoes. Conclusions: Australia urban/peri-urban Ae.aegypti species are susceptible to infection by the sylvatic and highly divergent DENV-2 virus QML22. However, our results indicate that replication of QML22 is attenuated relative to the contemporary strain QML16 and/or a salivary gland infection or escape barrier acts to prevent infection of saliva, potentially preventing onward transmission of this highly divergent virus in Australia.

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ANTIVIRAL ACTIVITY OF COPPER(II)CHLORIDE DIHYDRATE AGAINST DENGUE VIRUS TYPE-2 IN VERO CELL

Indonesian Journal of Tropical and Infectious Disease ◽

10.20473/ijtid.v6i4.3806 ◽

2017 ◽

Vol 6 (4) ◽

pp. 84 ◽

Cited By ~ 5

Author(s):

Teguh Hari Sucipto ◽

Siti Churrotin ◽

Harsasi Setyawati ◽

Tomohiro Kotaki ◽

Fahimah Martak ◽

...

Keyword(s):

Cell Culture ◽

Dengue Virus ◽

Vero Cell ◽

Virus Type ◽

Inhibitory Concentration ◽

Enzyme Linked Immunosorbent Assay ◽

Antiviral Compound ◽

Chloride Dihydrate ◽

Dengue Virus Type 2

Infection of dengue virus (DENV) was number of globally significant emerging pathogen. Antiviral dengue therapies ar importantly needed to control emerging dengue. Dengue virus (DENV) is mosquito-borne arboviruses responsible for causing acute systemic diseases and grievous health conditions in humans. To date, there is no clinically approved dengue vaccine or antiviral for humans, even though there have been great efforts towards this end. Copper and copper compounds have more effective in inactivation viruses, likes an influenza virus and human immunodeficiency virus (HIV). Purpose in this project was investigated of Copper(II)chloride Dihydrate antiviral compound were further tested for inhibitory effect on the replication of DENV-2 in cell culture. DENV replication was measures by Enzyme linked Immunosorbent Assay (ELISA) with selectivity index value (SI) was determined as the ratio of cytotoxic concentration 50 (CC50) to inhibitory concentration 50 (IC50) for compound. The maximal inhibitory concentration (IC50) of Copper(II)chloride Dihydrate against dengue virus type-2 was 0.13 μg/ml. The cytotoxic concentration (CC50) of compound against Vero cell was 5.03 μg/ml. The SI values for Copper(II)chloride Dihydrate 38.69. Result of this study suggest that Copper(II)chloride Dihydrate demonstated significant anti-DENV-2 inhibitory activities and not toxic in the Vero cells. Copper mechanisms play an important role in the prevention of copper toxicity, exposure to excessive levels of copper can result in a number of adverse health effects, as a result increased reactive oxygen species and oxidative damage to lipid, DNA, and proteins have been observed in human cell culture models or clinical syndromes of severe copper deficiency and inhibition was attributed to released cupric ions which react with cysteine residues on the surface of the protease.

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Identification of amino acids in the dengue virus type 2 envelope glycoprotein critical to virus infectivity

Journal of General Virology ◽

10.1099/vir.0.011486-0 ◽

2009 ◽

Vol 90 (10) ◽

pp. 2457-2461 ◽

Cited By ~ 11

Author(s):

Helga Kroschewski ◽

Jose-Luis Sagripanti ◽

Andrew D. Davidson

Keyword(s):

Amino Acids ◽

Dengue Virus ◽

Virus Type ◽

Envelope Glycoprotein ◽

Critical Role ◽

Virus Infectivity ◽

Virus Envelope ◽

Virus Attachment ◽

Dengue Virus Type 2

The dengue virus envelope glycoprotein mediates virus attachment and entry and is the major viral antigen. The identification of ‘critical’ amino acids in the envelope glycoprotein that cannot be altered without loss of infectivity could have a major impact on the development of dengue virus vaccines and diagnostics. In this context, we determined whether six amino acids, previously predicted by computational analysis to play a critical role in the virus life cycle, were essential for virus viability. The effects of mutating the six ‘critical’ amino acids and a further seven ‘neutral’ amino acids were analysed by using a dengue virus type 2 infectious cDNA clone. Of the six critical amino acids, three (Asp-215, Pro-217 and His-244) were found to be essential for virus viability in mammalian and mosquito cells.

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Review for "Glycogen synthetase kinase 3 inhibition drives MIC‐A/B to promote cytokine production by human natural killer cells in Dengue virus type 2 infection"

10.1002/eji.201948284/v1/review2 ◽

2019 ◽

Keyword(s):

Natural Killer Cells ◽

Dengue Virus ◽

Natural Killer ◽

Virus Type ◽

Cytokine Production ◽

Killer Cells ◽

Glycogen Synthetase ◽

Human Natural Killer Cells ◽

Dengue Virus Type 2

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Faculty Opinions recommendation of Engineering RNA interference-based resistance to dengue virus type 2 in genetically modified Aedes aegypti.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1030708.367881 ◽

2006 ◽

Author(s):

Vishvanath Nene

Keyword(s):

Rna Interference ◽

Aedes Aegypti ◽

Dengue Virus ◽

Virus Type ◽

Genetically Modified ◽

Dengue Virus Type 2

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Efﬁcacy of Catharanthus roseus Extract against Dengue Virus Type 2 Infection in Vitro

Indian Journal of Public Health Research & Development ◽

10.37506/v11/i2/2020/ijphrd/195051 ◽

2020 ◽

Vol 11 (2) ◽

pp. 1583

Author(s):

Noor Zarina Abd Wahab ◽

NorefrinaShafinaz Md. Nor ◽

Nazlina Ibrahim

Keyword(s):

Dengue Virus ◽

Virus Type ◽

Catharanthus Roseus ◽

Dengue Virus Type 2

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Reduced sphingosine kinase 1 activity in dengue virus type-2 infected cells can be mediated by the 3′ untranslated region of dengue virus type-2 RNA

Journal of General Virology ◽

10.1099/vir.0.055616-0 ◽

2013 ◽

Vol 94 (11) ◽

pp. 2437-2448 ◽

Cited By ~ 19

Author(s):

J. M. Carr ◽

T. Kua ◽

J. N. Clarke ◽

J. K Calvert ◽

J. R. Zebol ◽

...

Keyword(s):

Cell Death ◽

Dengue Virus ◽

Virus Type ◽

Elongation Factor ◽

Sphingosine Kinase ◽

Sphingosine Kinase 1 ◽

Diarrhea Virus ◽

Infected Cells ◽

Dengue Virus Type 2

Sphingosine kinase 1 (SphK1) is a lipid kinase with important roles including regulation of cell survival. We have previously shown reduced SphK1 activity in cells with an established dengue virus type-2 (DENV-2) infection. In this study, we examined the effect of alterations in SphK1 activity on DENV-2 replication and cell death and determined the mechanisms of the reduction in SphK1 activity. Chemical inhibition or overexpression of SphK1 after established DENV-2 infection had no effect on infectious DENV-2 production, although inhibition of SphK1 resulted in enhanced DENV-2-induced cell death. Reduced SphK1 activity was observed in multiple cell types, regardless of the ability of DENV-2 infection to be cytopathic, and was mediated by a post-translational mechanism. Unlike bovine viral diarrhea virus, where SphK1 activity is decreased by the NS3 protein, SphK1 activity was not affected by DENV-2 NS3 but, instead, was reduced by expression of the terminal 396 bases of the 3′ UTR of DENV-2 RNA. We have previously shown that eukaryotic elongation factor 1A (eEF1A) is a direct activator of SphK1 and here DENV-2 RNA co-localized and co-precipitated with eEF1A from infected cells. We propose that the reduction in SphK1 activity late in DENV-2-infected cells is a consequence of DENV-2 out-competing SphK1 for eEF1A binding and hijacking cellular eEF1A for its own replication strategy, rather than a specific host or virus-induced change in SphK1 to modulate viral replication. Nonetheless, reduced SphK1 activity may have important consequences for survival or death of the infected cell.

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