Schmallenberg Virus Infection among Red Deer, France, 2010–2012

Schmallenberg virus infection is emerging in European domestic and wild ruminants. We investigated the serologic status of 9 red deer populations to describe virus spread from September 2010 through March 2012 among wildlife in France. Deer in 7 populations exhibited seropositivity, with an average seroprevalence of 20%.

I n summer and fall 2011, an unidentified disease was reported in dairy cattle in Germany and the Netherlands, causing decreased milk production, fever, and diarrhea (1,2). The virus associated with these clinical signs was identified as a new member of the genus Orthobunyavirus of the Simbu serogroup and named Schmallenberg virus (SBV) (2). This virus was later associated with abortions and congenital malformations in calves, lambs, and kids in several European countries (3). Serologic testing among wild cervids in Belgium revealed antibodies against Schmallenberg virus in roe deer (Capreolus capreolus) and red deer (Cervus elaphus) (4). Seroprevalence was already high (27% on average) in wild cervids in October 2011 in Belgium, suggesting that the virus began circulating months earlier (before August 2011). It has recently been shown that SBV had already circulated in Culicoides vectors in Belgium during August and September 2011 (5). Although SBV has been closely monitored among domestic ruminants in France, suggesting that clinical cases and antibodies appeared almost at the same time during 2011-2012 (6), little is known about the geographic spread of SBV in wildlife. To correct this lack of data, we conducted a serologic study using serum specimens collected from red deer in different regions in France.

The Study
Blood samples from 502 red deer, which had been either killed by gunshot or captured, were collected within 9 French departments (administrative units) during 1 or 2 sampling seasons (i.e., during September 2010-January 2011 and September 2011-March 2012). The serum specimens were first screened by using an SBV indirect ELISA (i-ELISA) that was previously validated for the serum specimens from cattle, sheep, and goats (ELISA ID Screen Schmallenberg Virus Indirect, Bicupule; ID Vet, Montpellier, France) (7). The results were expressed as S/P values using the cutoff recommended for domestic species (S/P = [optical density sample (S)/optical density positive control (P)] × 100); S/P<60%, negative; S/P>70%, positive; and S/P 60-70%, doubtful result). Serum specimens were also tested with a new competitive ELISA (c-ELISA; ELISA ID Screen Schmallenberg Virus Competitive; ID Vet). Positive results by c-ELISA corresponded to a percentage of inhibition (PI) <50, doubtful result if 40>PI≤50, and negative when PI >50. The antigen used in both c-ELISA and i-ELISA is the same N recombinant protein. A subset of samples were also subjected to a seroneutralization test (SNT) as described (7).
Of 502 serum specimens, 492 could be tested by using i-ELISA and 486 by using c-ELISA. The 2 ELISA methods exhibited a 92% match (449/486). Because our samples (taken from dead animals in nonsterile conditions) generated bacterial contamination or cytotoxicity, conclusive SNT results were available from 114 animals only: 64 samples with positive or doubtful i-ELISA results and 50 samples (S/P>20) with negative i-ELISA results. A large part of the serum specimens that were positive or doubtful by ELISA methods were also positive for SBV by SNT, suggesting a good specificity of both methods, though slightly better for c-ELISA than for i-ELISA (Table 1). Many serum specimens that tested negative by i-ELISA or c-ELISA (all collected during 2011-2012) were positive by SNT (Table 1). Even though the c-ELISA kit appeared slightly more sensitive than the i-ELISA kit, these results suggest that SNT is the most sensitive technique for detecting antibodies against SBV in a recently infected population of red deer. These results are consistent with the fact that SNT and c-ELISA are able to detect IgG and IgM, whereas i-ELISA detects only IgG that appears after the IgM adaptive response (E. Breard, pers. comm.). Considering the performance of serologic methods in that study, seroprevalence was finally estimated as the proportion of positive or doubtful serum specimens by using the c-ELISA kit.
These results suggest that SBV was actively circulating during fall 2011 until mid-November or early December. In agreement with the findings of Linden et al. for Belgium (4), we consider that the mild temperature observed in France in fall 2011 may have favored a late activity of vectors (9). The date of first occurrence of seropositive red deer and the seroprevalence observed in each department (Table 2)

Conclusions
This study provides a preliminary view of SBV spread among wild cervids in France during 2010-2012. Our data suggest that SBV spread quickly from northeastern to southwestern France (≈800 km) between October and   December 2011. Our data also show the match of SBV spread among red deer and domestic flocks at the level of the department and highlight the perspective that red deer can be a sentinel of SBV spread for livestock. We also pinpointed the relevance of new competition ELISA for improving SBV surveillance in wildlife species, even though SNT remained the most reliable assay for SBV antibody detection in red deer. Further studies that encompass several years and include a larger number of species and localities would help provide a more complete picture of virus spread and risk factors in wildlife (11).