Zika virus recombination - from cultured cells to whole mosquitoes

Abstract

RNA viruses have the astonishing capacity to evolve rapidly due to their error-prone replication process and ability to undergo recombination which contribute to their genetic diversity. Recombination is a source of extensive genetic change that can yield hybrids of closely related viruses through a replicative process of template switching performed by the viral polymerase. An alternative non-replicative process has also been described which is poorly understood. Zika virus (ZIKV) is an arbovirus which belongs to the family Flaviviridae and has recently emerged as a threat to human health especially in the tropical regions of the globe. Prior to this work, there was no experimental evidence of ZIKV recombination. Mammalian and insect cell co-infection with two divergent ZIKV strains - MR766 and BeH819015 - was performed and a recombinant-specific PCR assay was developed to identify the first ZIKV hybrids in vitro. The viability of some of these recombinants was confirmed through construction of full-length infectious clones and transfection of viral RNA in Vero cells. ZIKV hybrids were shown to acquire adaptive changes upon multiple rounds of infection. The replication of ZIKV recombinants was also demonstrated in vivo - in live Aedes aegypti mosquitoes - and their transmission potential was assessed based on the presence of infectious virus in the mosquito saliva. Finally, next-generation sequencing was performed to analyse the mixed virus population in mosquitoes provided with infectious blood meal containing the two divergent ZIKV strains. Products of recombination were detected in co-fed mosquitoes as well as in co-infected mammalian and insect cells which were also subject to high-throughput sequencing analysis. This study is the first of its kind to address the role of recombination of ZIKV in the evolutionary process of virus emergence through in vitro and in vivo experimental systems.

Date of Award	14 Jun 2023
Original language	English
Awarding Institution	University of St Andrews
Supervisor	David Evans (Supervisor)