Serological testing of Schmallenberg virus in Swedish wild cervids from 2012 to 2016

When an emerging disease affects an animal or human population, there are often social, economic and welfare consequences. This was the case in 2011 and 2012, when Schmallenberg virus (SBV) was detected in Europe [1]. Initially, clinical signs were observed mainly in cattle, and included diarrhea, pyrexia, and a drop in milk yield. Infection during the first part of gestation was followed by stillbirths and birth defects in the offspring. No relation to any current and known circulating pathogen was identified [2]. Shortly thereafter, the causative agent was shown to be a novel Orthobunyavirus of the family Bunyaviridae [3] that is transmitted by biting midges (Culicoides spp.) [4]. The virus has the ability to cross the placental barrier and cause lesions in fetuses and neonates [5]. It infects a number of domestic and wild animal species including cattle, sheep, goats [6], alpaca [7] red deer, roe deer [8], fallow deer, moose, bison [9], wild boar [10], dogs [11], and a number of zoo animals [12]. In 2014 and 2015 the virus was still circulating in continental Europe [13, 14]. In Sweden, SBV was first detected in 2012 in domestic animals in the south. The virus spread rapidly north beyond the Arctic Circle, and occurred in high prevalence in tested animals [15]. However, it was not known if the virus did circulate (in 2012) or still is circulating in wildlife ruminant populations, and if they could act as reservoirs for the virus [16].

The aim of this study was to investigate if SBV is circulating among wild cervids in Sweden. Two hypotheses were tested: a) SBV-specific serum antibodies can be detected in Swedish wild cervids to the same extent and during the same time periods as SBV was diagnosed in domestic ruminants. b) SBV is still widely circulating in wild ruminant populations, despite likely being absent in the domestic ruminant population.

Sera from time period 2 were additionally analyzed for SBV-neutralizing antibodies. Such antibodies were detected in 16/16 SBV-N-antibody-positive sera, in 3/12 SBV-N-antibody negative and 2/2 doubtful sera. No SBV-specific antibodies were detected by any of the techniques in nine animals (Table 2). Seropositive animals were found in all sampling areas and in all tested cervid species (moose, roe deer, red deer, and fallow deer). The SBV-neutralizing antibody titers varied between species (Table 2), but low sample sizes precluded statistical analyzes with regards to inter- and intraspecies titer variation. These data show a very good agreement between both tests for negative samples or when the SBV-neutralizing antibody titers were >1/16. Five samples with an SBV-neutralizing antibody titer of 1/8 (n = 3) or 1/16 (n = 2) were found doubtful (n = 2) or negative (n = 3) by cELISA.

This is the first report of findings of SBV-specific antibodies in Scandinavian wildlife, showing that wild cervids in Sweden were exposed to the virus in the summer of 2012 and probably not after the vector season 2014. Moreover, this study indicates that SBV infection in wildlife and domesticated animals follows the same pattern in the same area in which cattle were seropositive in 2012 and 2013 but seronegative in a recent serological survey when born after the vector season 2013 (S. Zohari, personal communication). Schmallenberg virus does not seem to circulate in Sweden anymore, in contrast to other European countries such as Germany [13]. The difference between Sweden and countries in central and southern Europe is that Sweden has a long vector-free season. Virus transmission and spread is possible at temperatures around 15 °C with a temperature optimum between 18 °C and 19 °C due to vector limitations [20]. In Sweden, such daily mean temperatures are usually limited to May-August [21], but may occur in September as well. Virus persistence depends on the winter survival of adult midges which must have access to a naïve ruminant population. The absence of SBV antibodies in the tested animals suggests that the virus does not persist in ruminants (wild or domestic) or in the environment during the vector-free season. However, to completely verify that the infection is not present in Swedish cervid populations, a larger and predetermined number of animals need to be tested.

A seroneutralization test (SNT) was used at first to test wildlife following the emergence of SBV in Europe, since no cELISA was available to measure specific antibodies against SBV. Since then, a cELISA has been developed with high sensitivity and specificity for domestic ruminant samples [17], and in addition, is easier to perform. Our results indicate that the two methods are in agreement for most samples tested and the level of agreement increases with increasing titer. It is possible that none of the techniques perform well for samples with low titers. Our data suggest that the cELISA lacks sensitivity for sera with low antibody titers. The SNT, on the other hand, might lack specificity for sera at low dilution, as previously observed for sheep [22]. Nevertheless, in contrast to in the sera collected between November 2012 and February 2013, no specific antibodies were detected by cELISA in sera collected in 2015 from animals aged between 0.5 and 1.5 years of age, which suggest that the SBV has not recently circulated among wild cervids in the investigated area.

The effect of SBV in wildlife is still unknown. No clinical signs of SBV were reported in Swedish wildlife during the summer and early autumn of 2012 when the outbreak was detected. Possible signs of SBV infection in Swedish wild or fenced cervids, in the form of abortions and congenital malformations, were not reported during 2013 [23]. However, it is unlikely to make such observations in wild animals for a number of reasons such as scavenging birds and mammals. Furthermore, it requires a primary maternal infection and viremia during a particular period of pregnancy, i.e. after the first placentoma has developed and before the fetuses are immunocompetent [24]. The circulation of SBV most likely did not coincide with the placentoma formation in wild cervids in Sweden. Hence, delayed consequences of SBV infection in Swedish cervids are presumed to be minimal.

Schmallenberg virus was circulating among Swedish wild cervids during the vector season 2012, but no serological evidence of SBV was found during subsequent testing in 2015 and 2016. Based on Swedish climate conditions and the results of this study, we can assume that Sweden has an unfavorable climate for SBV-overwintering vectors. This might have contributed to the fact that the virus infection has seemingly not become endemic or possibly not reoccurred in Sweden. Midge activity and the reproductive season of Swedish wild cervids, is a seasonal and biological mismatch for the virus, which may explain that SBV has little impact on Swedish wild ruminant health. These animals are thus highly unlikely to be reservoirs of this virus.