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A novel Ebola virus expressing luciferase allows for rapid and quantitative testing of antivirals.

ABSTRACT: Ebola virus (EBOV) causes a severe hemorrhagic fever with case fatality rates of up to 90%, for which no antiviral therapies are available. Antiviral screening is hampered by the fact that development of cytopathic effect, the easiest means to detect infection with wild-type EBOV, is relatively slow. To overcome this problem we generated a recombinant EBOV carrying a luciferase reporter. Using this virus we show that EBOV entry is rapid, with viral protein expression detectable within 2 h after infection. Further, luminescence-based assays were developed to allow highly sensitive titer determination within 48 h. As a proof-of-concept for its utility in antiviral screening we used this virus to assess neutralizing antibodies and siRNAs, with significantly faster screening times than currently available wild-type or recombinant viruses. The availability of this recombinant virus will allow for more rapid and quantitative evaluation of antivirals against EBOV, as well as the study of details of the EBOV life cycle.

SUBMITTER: Hoenen T

PROVIDER: S-EPMC3787978 | biostudies-literature | 2013 Sep

REPOSITORIES: biostudies-literature

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A novel Ebola virus expressing luciferase allows for rapid and quantitative testing of antivirals.

Hoenen Thomas T Groseth Allison A Callison Julie J Takada Ayato A Feldmann Heinz H

Antiviral research 20130607 3

Ebola virus (EBOV) causes a severe hemorrhagic fever with case fatality rates of up to 90%, for which no antiviral therapies are available. Antiviral screening is hampered by the fact that development of cytopathic effect, the easiest means to detect infection with wild-type EBOV, is relatively slow. To overcome this problem we generated a recombinant EBOV carrying a luciferase reporter. Using this virus we show that EBOV entry is rapid, with viral protein expression detectable within 2 h after ...[more]

PMID: 23751367

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Project description:Ebola virus (EBOV) is the causative agent of the often-fatal Ebola virus disease (EVD) characterized by hemorrhagic fever in humans and non-human primates. Sexual transmission from male survivors has been at the origin of multiple outbreak flare-ups between 2015 and 2021. However, this route is still poorly understood and the resulting EVD from it is also understudied. To support epidemiological studies documenting sexual transmission to women, and as a transition from previously using monolayer vaginal epithelial cells (VK2/E6E7), we first determined the biological relevance of two similar air-liquid interface models of the human vaginal epithelium (VEC and VLC Epivaginal™) and then characterized their susceptibility to EBOV and virus-induced inflammation. Finally, we evaluated toxicity of Polyphenylene Carboxymethylene (PPCM) microbicide in VLC and reassessed its antiviral effect. As expected, the VEC, but also VLC model showed stratified layers including a lamina propria under an epithelial structure similar to the full thickness of the human vaginal epithelium. However, we could not detect the immune cells featured in the most relevant model (VLC) of the vaginal epithelium using the dendritic cell CD1a and CD11c markers. Consistent with our previous work using the VK2/E6E7 cell line, infectious virus was detected from the apical side of both primary human cell systems, but only when using a high infective dose, with titers remaining at a constant level of 103-4 pfu/ml over 7 days suggesting lasting infectious virus shedding. In addition, infection caused disruption of the epithelium of both models and virus antigen was found from the apical superficial layers down to the lamina propria suggesting full virus penetration and overall confirming the susceptibility of the human vaginal tissue for EBOV. Just like previously seen in VK2/E6E7 cells, VLC infection also caused significant increase in inflammatory markers including IL-6, IL-8, and IP-10 suggesting vaginitis which is again consistent with tissue lesions seen in non-human primates. Finally, both virus infection and virus-induced inflammatory response in VLC could be prevented by a single 5-min PPCM microbicide treatment prior infection.

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A novel Ebola virus expressing luciferase allows for rapid and quantitative testing of antivirals.

Publications

A novel Ebola virus expressing luciferase allows for rapid and quantitative testing of antivirals.

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