Characterization of the Mouse Neuroinvasiveness of Selected European Strains of West Nile Virus

Stephanie M Lim,Penelope Koraka,Sander Van Boheemen,Jouke M Roose,Albert D M E Osterhaus,David A M C Van De Vijver,Byron E E Martina,Dick Jaarsma,Robyn Klein

doi:10.1371/journal.pone.0074575

Abstract

West Nile virus (WNV) has caused outbreaks and sporadic infections in Central, Eastern and Mediterranean Europe for over 45 years. Most strains responsible for the European and Mediterranean basin outbreaks are classified as lineage 1. In recent years, WNV strains belonging to lineage 1 and 2 have been causing outbreaks of neuroinvasive disease in humans in countries such as Italy, Hungary and Greece, while mass mortality among birds was not reported. This study characterizes three European strains of WNV isolated in Italy (FIN and Ita09) and Hungary (578/10) in terms of in vitro replication kinetics on neuroblastoma cells, LD50 values in C57BL/6 mice, median day mortality, cumulative mortality, concentration of virus in the brain and spinal cord, and the response to infection in the brain. Overall, the results indicate that strains circulating in Europe belonging to both lineage 1 and 2 are highly virulent and that Ita09 and 578/10 are more neurovirulent compared to the FIN strain.

Highlights

West Nile virus (WNV) is a positive-sense single-stranded RNA virus, which belongs to the genus Flavivirus
The E protein of different WNV strains is not significantly affected by in vitro culture To determine whether generation of virus stocks, through one extra passage on insect cells, resulted in changes in the consensus sequence of the respective viruses, the E protein of the different virus stocks was sequenced using the Sanger method and sequences were compared to those deposited in GenBank
There were no amino acid changes found in Ita09 and 578/10 compared to the sequences deposited in Genbank

Summary

Introduction

West Nile virus (WNV) is a positive-sense single-stranded RNA virus, which belongs to the genus Flavivirus. Lineage 2 strains were restricted to sub-Saharan Africa This genetic classification has been used frequently to classify WNV obtained during outbreaks. In 2004, a lineage 2 strain was isolated from birds of prey in Hungary [9], which established itself in the region and largely spread throughout the country and into eastern Austria by 2008 [10]. During this outbreak, cases of human neuroinvasive disease were comparatively rare and rather mild with no deaths reported [10]. Up to October 4th, 2010, 192 cases of neuroinvasive disease in humans, including 32 deaths, had been laboratory diagnosed, all in the elderly

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