Validation of flavivirus infectious clones carrying fluorescent markers for antiviral drug screening and replication studies

Frontiers in Microbiology

Microbiology

Matthieu Villeneuve¹  Mathilde Hénaut¹  Alice Trausch¹  Sébastien Lyonnais¹  Nathalie Gros¹  Aymeric Neyret¹  Gregor Dubois¹  Christine Chable-Bessia¹  Delphine Muriaux²  Liubov Cherkashchenko³  Andres Merits⁴ 

[1] CEMIPAI UAR3725 CNRS, University of Montpellier, Montpellier, France;CEMIPAI UAR3725 CNRS, University of Montpellier, Montpellier, France;IRIM UMR9004 CNRS, University of Montpellier, Montpellier, France;CEMIPAI UAR3725 CNRS, University of Montpellier, Montpellier, France;Institute of Technology, University of Tartu, Tartu, Estonia;Institute of Technology, University of Tartu, Tartu, Estonia;

关键词: flaviviruses;    infectious clone;    Zika virus;    dengue virus;    Kunjin virus;    fluorescent marker and reporter genes;    reverse genetics;   

DOI  :  10.3389/fmicb.2023.1201640

 received in 2023-04-06, accepted in 2023-08-17,  发布年份 2023

来源: Frontiers

【 摘 要 】

Flaviviruses have emerged as major arthropod-transmitted pathogens and represent an increasing public health problem worldwide. High-throughput screening can be facilitated using viruses that easily express detectable marker proteins. Therefore, developing molecular tools, such as reporter-carrying versions of flaviviruses, for studying viral replication and screening antiviral compounds represents a top priority. However, the engineering of flaviviruses carrying either fluorescent or luminescent reporters remains challenging due to the genetic instability caused by marker insertion; therefore, new approaches to overcome these limitations are needed. Here, we describe reverse genetic methods that include the design and validation of infectious clones of Zika, Kunjin, and Dengue viruses harboring different reporter genes for infection, rescue, imaging, and morphology using super-resolution microscopy. It was observed that different flavivirus constructs with identical designs displayed strikingly different genetic stabilities, and corresponding virions resembled wild-type virus particles in shape and size. A successful strategy was assessed to increase the stability of rescued reporter virus and permit antiviral drug screening based on quantitative automated fluorescence microscopy and replication studies.

【 授权许可】

Unknown   
Copyright © 2023 Cherkashchenko, Gros, Trausch, Neyret, Hénaut, Dubois, Villeneuve, Chable-Bessia, Lyonnais, Merits and Muriaux.