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Master thesis

Caractérisation des déterminants moléculaires de virulence de souches européennes ou méditerranéennes du virus West Nile

Abstract : West Nile virus (WNV) is a neurotropic arbovirus transmitted by mosquito bites that can infect a wide variety of animal species. The natural reservoir of the WNV is composed by the wild bird fauna. The aim of my work was to characterize the molecular determinants of virulence of European or Mediterranean strains of the West Nile virus. To achieve this objective, I relied on the laboratory's previous work, namely the production of an infectious clone of the strain IS-98-ST1. We also used the strain Italy 2008 (It08) (strain 229892, GENBANK: FJ483549) which is considered to be pathogenic. I constructed chimeric viruses (by exchange of genomic fragments between the WNV Is98 and It08 strains) or mutants (integration of the NS3-P249T mutation) from the infectious clone Is98 and ensured their amplification on Vero cells. I have characterized their biological properties in in vitro cell lines and in vivo models (mouse, mammalian model for West Nile virus virulence, and birds: 1-day chick in partnership with CERVA CODA, Belgium). In vitro I observed that the viruses Is98, It08 and Chimera 3 behave differently from those of chimera 4 and the mutant Thr (small lysis areas). Kinetics of infections in VERO cells suggest that chimera 4 and mutant NS3 P249T do not exhibit a replication defect in this cell model. For the NS3 mutant, our in vivo results suggest that virulence of WNV is strongly impacted in the murine or avian host by the presence of the Proline residue at position 249 on the NS3 reading frame in a more global Is98 genomic context . For chimeric viruses, the survival curves in the murine model are comparable for the different viruses tested (Is98, It08, Chimera 3 and 4), while the RT-qPCR analyzes of the different samples show lower viral loads after infection with The chimera 4. Differences in the behavior of chimera 4 suggest that it induces a viremia shifted in time compared to other viruses (a priori later). Infections in the avian model demonstrated statistically significant differences in viral loads measured in different peripheral compartments and in oral swabs after infection with different viruses. Comparable survival curves and viral loads were observed after infection of SPF chicks of 1J respectively between the virus It08 and the chimera 3 (viral loads in the different samples taken, low mortality) and between the virus Is98 and the chimera 4 (oral excretion and high peripheral viral loads, higher mortality). The set of results I obtained allowed me to objectify the role of three different determinants for WNV virulence, NS3 zone 249, NS4A / B and NS5. However, it seems that their impacts are different depending on the target host and that in order to have a real idea of ​​the virulence of a WNV strain, the part of the mutated genome and the target host species must be taken into account. These determinants appear to be "major" determinants of the replicative capacity or "fitness" of WNV to its host. That is, they play a preponderant role in the intrinsic ability of the virus to replicate and escape the host-specific immune system. We can think for the future that the discovery of the virulence factors of this virus will allow us to determine the virulence factors of other types of flaviviruses. This will probably enable us to rapidly characterize the epizootic or epidemic potential of a given strain and to consider ways of fighting these arboviruses more effectively (vaccines, antivirals, etc.)
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Master thesis
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Submitted on : Monday, July 3, 2017 - 4:42:26 PM
Last modification on : Wednesday, November 17, 2021 - 12:31:52 PM
Long-term archiving on: : Friday, December 15, 2017 - 12:19:02 AM

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Steeve Lowenski. Caractérisation des déterminants moléculaires de virulence de souches européennes ou méditerranéennes du virus West Nile. Sciences du Vivant [q-bio]. 2017. ⟨hal-01553679⟩

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