Syracuse

A comparative study of two Toscana virus strains pathogenicity and replication capacity

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Thiesson, Adrien | Roy, Marlène | Lacôte, Sandra | Desloire, Sophie | Aurine, Noémie | Mathieu, Cyrille | Pain, Bertrand | Marianneau, Philippe | Arnaud, Frédérick | Ratinier, Maxime

Edité par CCSD -

International audience. In recent decades, the number of emerging arthropod-borne viruses has increased drastically. Toscana virus (TOSV) is a zoonotic virus transmitted by several sandflies species. It was isolated for the first time 50 years ago, in Italy, from infected sandflies (P. perniciosus et perfiliewi) and is now circulating in most countries around the Mediterranean basin. TOSV infection in humans is mostly asymptomatic or produces mild symptoms. However, some patients develop neurological diseases due to viral invasion of the central nervous system (CNS). Thus, TOSV is one of the leading causes of meningitis and encephalitis in humans during summer. Nevertheless, it is still an understudied virus and its biology and epidemiology remain poorly understood. Among isolated strains, two distinct genetic lineages have been identified based on phylogenetic analysis (named lineages A and B). More recently, a third lineage (named lineage C) has been identified based on partial viral sequences only, and no virus has been isolated so far. Until now, the relationship between the TOSV inter-lineage genetic diversity and viral pathogenicity or vector competence remains unknown.In this context, we conducted a comparative study of two TOSV strains from lineages A (TOSV-A) and B (TOSV-B). Using 129/Sv IFNAR-/- mice as a model of infection, our team unveiled striking differences between these two strains in term of pathogenicity. More precisely, the mice survival rate infected by TOSV-A is lower than those infected by TOSV-B (56% and 89% respectively). Moreover, TOSV-A is detected in the brain of 10 mice whereas TOSV-B is found in this organ of only one mouse (n=18). By testing the replicative capacity of these two viruses in (i) human induced pluripotent stem cells (hiPS) differentiated in the neural pathway as glial and neuronal cells and (ii) organotypic cultures of mouse brain slice, we demonstrated that the difference in mice brain infection was not due to a defect of TOSV-B entry or replication in CNS cells.Using a newly developed reverse genetic system for TOSV (Alexander et al., 2020), we generated two viruses, rTOSV-A and rTOSV-B, representatives of the strains previously characterized in vivo. We first showed that rTOSV-A is replicating at a higher rate in A549 and A549 Npro cells than rTOSV-B. We then demonstrated that rTOSV-B displays a slower internalization during the viral entry process and produces fewer infectious viral particles in the infected cells' supernatant compared to rTOSV-A. Using reassortant and chimeric viruses, we further determined that the genetic diversity within the M segment was responsible of the differences observed between the two studied strains. Overall, the development of new experimental models and molecular tools combined with a comparative approach between two strains of TOSV enables us to demonstrate that the genetic variability of TOSV is highly impacting its biological properties. As no vaccine against TOSV has been developed yet, it is essential to strengthen the characterisation of circulating TOSV strains in order to identify viral determinants involved in pathogenicity and transmission. Ultimately, these works will help us to evaluate more efficiently the epidemiologic risk of a given strain.

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