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Updated: Sep 18, 2025
Rescue and Characterization of Recombinant Virus from a New World Zika Virus Infectious Clone
Published on: June 7, 2017
Construction and application of infectious cDNA clone, subgenomic replicon and packaging system for Zika virus and
Yu He1, Yibin Tang2, Xiaoli Wang2
1Institute of Veterinary Medicine and Immunology, Sichuan Agricultural University, Chengdu, Sichuan 611130, China; Research Center of Avian Disease, College of Veterinary Medicine, Sichuan Agricultural University, Chengdu, Sichuan 611130, China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Chengdu, Sichuan 611130, China; Key Laboratory of Agricultural Bioinformatics, Ministry of Education, Chengdu, Sichuan 611130, China.
Abstract:
Flaviviruses pose a significant global health threat due to their rapid spread and potential to cause severe clinical manifestations. Comprehending the mechanisms of replication of these pathogens and developing effective antiviral strategies are essential for combating these pathogens. In the present study, full-length infectious cDNA clones were generated for Zika virus (ZIKV) and Dengue virus (DENV), respectively. Recombinant viruses were successfully produced by transfecting cDNA clone plasmids. Using the infectious clone, ZIKV and DENV subgenomic replicons were also generated, which lack the prM-E gene and instead expressing a luciferase or fluorescent marker (OXGFP or mCherry). The replicons exhibited efficient replication in BHK-21 cells. Through the utilization of ZIKV and DENV-2 replicons that express luciferase, three potential antiviral agents were identified. These agents demonstrated activity against DENV-2 and ZIKV, while not inducing significant cytotoxic effects. This demonstrates the significance of these replicons in the screening of antiviral agents. Moreover, DENV-2 and ZIKV single-round infectious particles (SRIPs) were produced by co-transfecting packaging plasmids and replicons into BHK-21 cells. The packaging assay demonstrated that flaviviruses can utilize the prM-E protein from other species to generate SRIPs. The specific binding of the nucleocapsid (NC) to the prM-E protein varies among different flaviviruses. The reverse genetics tools established in this study will facilitate research on DENV-2 and ZIKV virus replication and the development of antiviral medications targeting these two arboviruses.
Insights
Researchers developed new reverse genetics tools for Zika virus (ZIKV) and Dengue virus (DENV). These tools enabled the identification of novel antiviral agents effective against ZIKV and DENV-2 without causing significant cytotoxicity.
Area of Science:
- Virology
- Molecular Biology
- Drug Discovery
Background:
- Flaviviruses like Zika virus (ZIKV) and Dengue virus (DENV) represent significant global health concerns due to their rapid transmission and severe disease potential.
- Understanding flavivirus replication mechanisms is crucial for developing effective antiviral therapies.
Purpose of the Study:
- To establish reverse genetics tools for ZIKV and Dengue virus (DENV).
- To utilize these tools for screening and identifying potential antiviral agents against ZIKV and DENV-2.
- To investigate the cross-species utilization of prM-E proteins in flavivirus particle production.
Main Methods:
- Generation of full-length infectious cDNA clones for ZIKV and DENV.
- Construction of ZIKV and DENV subgenomic replicons expressing reporter genes (luciferase, OXGFP, mCherry).
- Production of single-round infectious particles (SRIPs) using replicons and packaging plasmids.
Main Results:
- Recombinant viruses and replicons demonstrated efficient replication in BHK-21 cells.
- Three potential antiviral agents were identified using luciferase-expressing replicons, showing activity against DENV-2 and ZIKV with low cytotoxicity.
- Packaging assays confirmed cross-species utilization of prM-E proteins and highlighted variations in nucleocapsid-prM-E binding.
Conclusions:
- The established reverse genetics systems provide valuable tools for studying ZIKV and DENV replication.
- These systems facilitate the discovery and development of novel antiviral medications targeting flaviviruses.
- The findings contribute to a better understanding of flavivirus assembly and interspecies protein interactions.
