Protection provided by a recombinant ALVAC^®-WNV vaccine expressing the prM/E genes of a lineage 1 strain of WNV against a virulent challenge with a lineage 2 strain

Abstract

The emergence of lineage 2 strains of WNV in Europe as a cause of clinical disease and mortality in horses raised the question whether the existing WNV vaccines, all based on lineage 1 strains, protect against circulating lineage 2 strains of WNV. In the present paper we have determined the level of cross protection provided by the recombinant ALVAC^®-WNV vaccine in a severe challenge model that produces clinical signs of WNV type 2 disease. Ten horses were vaccinated twice at 4 weeks interval with one dose of the ALVAC-WNV vaccine formulated at the minimum protective dose. A further 10 horses served as controls. Two weeks after the second vaccination, all horses were challenged intrathecally with a recent neurovirulent lineage 2 strain of WNV. The challenge produced viraemia in 10 out of 10 and encephalitis in 9 out of 10 control horses. Three horses had to be euthanized for humane reasons. In contrast, none of the vaccinated horses developed WNV disease and only 1 vaccinated horse became viraemic at a single time point at low titre. The prevalence of WNV disease and viraemia were significantly lower in the vaccinated horses than in the control horses (P < 0.0001 for both). Based on these results, the ALVAC-WNV vaccine will provide veterinarians with an effective tool to control infections caused by lineage 1 and 2 strains of WNV.

Introduction

West Nile virus (WNV) is a flavivirus that is maintained and transmitted through an enzootic cycle involving birds as amplifying hosts and ornithophylic mosquitoes. Humans and horses are considered incidental, dead-end hosts that become infected from the bites of infected mosquitoes. Sequence analysis of WNV isolates show that they fall into two major phylogenetic lineages [1]. Lineage 1 viruses have a widespread distribution and historically have been associated with sporadic outbreaks with limited geographic extension in horses and humans in Africa, Europe, Middle East and Asia [2]. Since 1994, the number and severity of outbreaks have increased and the virus has emerged in areas traditionally free of WNV [3]. As an example, the virus was introduced in New York and environs in 1999 [4] and rapidly spread across the North American continent resulting in nearly 15,000 equine and over 4000 human clinical cases at the peak of the epidemic in 2002 [5].

Lineage 2 strains of WNV were traditionally found in sub-Saharan Africa and Madagascar. In 2004, the virus emerged for the first time in central Europe [6], [7] and established itself in Hungary. Sporadic cases of WNV disease were detected in the next year in birds of prey and the virus was also isolated from the brain of a sheep which died with encephalitis. In the summers of 2008 and 2009 outbreaks occurred all over Hungary and in the eastern part of Austria involving birds of prey (mainly goshawks), horses and humans. In July and August 2010, the virus was found in Greece causing neuroinvasive disease in 81 people [8]. Until recently, the lineage 2 strains of WNV have rarely been associated with severe disease in man and horse and its pathogenicity has been questioned [4], [9]. However, recent observations in Hungary [6], South Africa [10] and Greece [8] have changed this perception and indicate that highly neuro-invasive lineage 2 strains exist in horses and humans.

A variety of vaccines, all produced from lineage 1 strains of WNV, are commercially available to prevent WNV infection of equids, including conventional killed [11], DNA [12] and recombinant vectored vaccines [13], [14]. With the apparently evolving WNV situation, particularly in Europe, it is expected that foreseeable vaccination strategies will make use of vaccines which provide rapid onset of immunity. Both the ALVAC-WNV [13] and Flavivirus WNV chimera vaccine [14] have been shown to control WNV infection shortly after a single dose of vaccine.

The re-emergence of lineage 2 strains of WNV as a cause of clinical disease and mortality in horses raised the question whether the existing WNV vaccines cross protect against circulating lineage 2 strains of WNV. Here we determined the level of protection provided by the recombinant ALVAC-WNV vaccine against experimental infection with a virulent lineage 2 strain isolated from a horse during the Hungarian outbreak in 2008.