First Serological Evidence of West Nile Virus in Horses and Dogs from Corsica Island,France

West Nile virus (WNV) is widely distributed over the world, including Europe, Africa, and Asia and spread over the past two decades to North and South America (Petersen et al. 2013). Its circulation in Europe is known since 1960 in France (Camargue), Portugal, and Cyprus. The virus widely emerged since the 1990s, being responsible for sporadic cases and outbreaks in Europe (Calistri et al. 2010). These five past years we observed a huge expansion of the WNV circulation area. New territories were affected especially in Italy, where the virus is now endemic (Magurano et al. 2012), Greece, and the Balkans with emergence of both lineages 1 and 2, suggesting an increase in the virus introductions and a better dispersion ability of lineage 2 (Barzon et al. 2015). Moreover, the hypothesis of winter persistence is more and more suggested, especially in Italy, Greece, and Romania (Calistri et al. 2010). In the south of France, sporadic cases are frequently described and two epidemics occurred in Camargue in 2010 (Bahuon and Lecollinet 2015) and 2015 (Bournez et al. 2015), a region highly concentrated in marshland, mosquitoes, wild birds, and horses. The last epidemic occurred in 2015 in this region and involved 48 horses and 1 symptomatic human case (Bournez et al. 2015).

Corsica Island is a French island in the Mediterranean Sea close to the southeast French coast, Sardinia, and the west Italian coast. To date, no circulation of WNV has been described. The aim of this study was to identify a possible WNV circulation in Corsica in sheep, horses, and dogs as sentinel animals for the virus surveillance. In September 2014, 386 blood samples were collected from 219 sheep, 96 horses not vaccinated against WNV, and 71 dogs, in 12 localities in Corsica, all in the oriental coast of the island known for its frequent ponds, with an average altitude of 37 meters. Among the 71 dogs, 12 were military dogs previously sampled in 2011 (Davoust et al. 2014). Animal origin was assessed to evaluate the real local circulation of the virus in the island (Table 1). Horses and sheep are all free-range raised and protected from the rain by a shelter. All the dogs sampled are military dogs or hunter dogs, living outside with a kennel protection. Origin of continental dogs is unknown. They can also come from Hungary where they could have been infected.

The serum samples were obtained by centrifugation of the collected peripheral blood, frozen at −20°C, and sent to the laboratory of the French National Reference Centre for Arbovirus (Marseille, France). Each sample was systematically tested for WNV immunoglobulin G (IgG) using an in-house enzyme-linked immunosorbent assay (ELISA) with inactivated WNV as antigen as described by Cabre et al. (2006). This ELISA was adapted to sheep and dogs IgG detection using specific antibodies (commercial peroxidase rabbit antisheep IgG and sheep anticanine IgG). Owing to antigenic cross-reactivity among flaviviruses, all positive samples were confirmed as true positive by serum neutralization test as described by Cabre et al. (2006), using a strain of WNV lineage 2 isolated in 2008 from an Austrian patient. Serum neutralization test was also performed with a strain of Usutu virus isolated in 2010 from a mosquito in Senegal to rule out any other flavivirus circulation in this area.

The result of the ELISA for the WNV antibody test on the sheep sera was all negative, whereas 9 of 96 horses (9.4%) and 6 of 71 dogs (8.4%) presented WNV antibodies. All the positive samples from horses and dogs were confirmed by serum neutralization test and did not present neutralizing antibodies against Usutu virus. Among the nine WNV-positive horses, six (66.6%) were native of Corsica, two (22.2%) originated from continental France, and one from Spain (11.1%); however, the three imported horses have been living on the island for more than 3 years. Among the six positive dogs, three (50%) were native of Corsica and three (50%) originated from continental France, two of which were living in the island for more than 3 years. One of the six positive dogs seroconverted as it tested negative for WNV antibodies in 2011.

Among the positive cases, the origin of the WNV infection could not be firmly determined as the animals in question have lived both in Corsica and in continental France or Spain where the virus also circulates. Nevertheless, 50% of the WNV-positive dogs and 66.6% of the WNV-positive horses detected were native of Corsica. Consequently, the origin of their infection can be related to a local circulation of the WNV over the Corsica Island.

This study is the first to evidence the spread of WNV into Corsica, although we cannot conclude about a current active circulation of the virus. Our data show that local animals or thus imported from continental France are WNV seropositive. We cannot speculate about the way of introduction of the WNV to Corsica Island and whether this virus is definitively established in this area. This study confirms that sheep do not play a role in WNV epidemiology (Davoust et al. 2016). The use of dogs and horses as sentinel animals for WNV surveillance in Corsica seems to be pertinent as already described in other countries such as Senegal by Davoust et al. (2016). Horses are particularly sensitive to the virus and are often used as indicators of the virus circulation (Leblond et al. 2007). Dating the WNV circulation is complicated as we only detect ancient infection. Nevertheless, one military dog seroconverted between 2011 and 2014. Surrounding countries and islands frequently reported outbreaks. Indeed, both lineages 1 and 2 were described recently in Sardinia (an Italian island close to Corsica) in 2011 and 2012 and in Italy and the south of France (Magurano et al. 2012). Trafficking between Corsica and these countries could have brought infected mosquitoes on the island. Moreover, the natural animal reservoir for this virus should include an unknown number of wild species of birds that develop sufficient viremia to be considered as good amplifiers and that may also play a key role in the virus dissemination from a country to another (Kilpatrick et al. 2007). Migratory birds could have brought the virus to the island during a stopover in Corsica.

Although no clinical case was reported to date in humans and horses, this report highlights the necessity to improve WNV surveillance in animals and humans, as well as in blood donors in Corsica.