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This study aimed at estimating the Schmallenberg virus (SBV) seroprevalence in dairy heifers, non-dairy adult cattle, sheep and goats in the Netherlands after cessation of SBV transmission at the end of 2011. Archived serum samples from ruminants submitted to the GD Animal Health Service for monitoring purposes between November 2011 and March 2012 were selected and tested for presence of SBV-specific antibodies using an in-house ELISA. Animal seroprevalences were estimated at 63.4% in dairy heifers, 98.5% in adult non-dairy cattle, 89.0% in sheep and 50.8% in goats. Multivariable analyses were carried out to describe the relationship between potential risk factors and the ELISA outcome S/P%. The overall SBV seroprevalence in ruminants and ruminant herds in the Netherlands at the end of 2011 was high, with considerable differences between species and farm types. No gradient spatial pattern in final seroprevalence could be detected and therefore no suggestions about the site of introduction and spread of SBV in the Netherlands in 2011 could be made. In dairy heifers, it was shown that S/P% increased with age. In sheep, S/P% was lower in animals located in the coastal area. Whether herds were located near the German border did not affect the S/P% in sheep nor in dairy heifers. An attempt was made to gain insight in the spatiotemporal introduction of SBV in the Netherlands in 2011, by testing sheep serum samples from 2011. A seroprevalence of about 2% was found in samples from April, June and July 2011, but the ELISA positive samples could not be confirmed in a virus neutralization test. A clear increase in seroprevalence started at August 2011. From mid-August 2011 onwards, seropositive samples were confirmed positive by virus neutralization testing. This indicated the start of the epidemic, but without a clear spatial pattern., (Copyright © 2013 Elsevier B.V. All rights reserved.)

To detect Schmallenberg virus (SBV) infections in ruminants and to perform SBV epidemiological studies a cost-effective serological test is required. For these purposes an indirect whole virus Enzyme-linked Immunosorbent Assay (ELISA) for detection of SBV specific antibodies in ruminant blood samples was developed. Schmallenberg virus antigen was produced by propagation on Vero cells, partly purified and coated onto ELISA plates. The indirect ELISA procedure included the subsequent incubation of diluted samples, protein-G-HRP conjugate and TMB substrate solution. Net Optical Densities (OD) values were calculated and expressed as a sample to positive percentage (S/P%) by comparison of the average net OD with the OD of the positive control. Validation of this assay was performed using 633 samples from SBV-free sheep, goats and cattle, and 141 samples from SBV suspect ruminants. The diagnostic specificity was 98.8%. Test results of 86 ruminant serum samples using both the SBV-ELISA and an SBV virus neutralization test (VNT), designated as the gold standard serological test for SBV, showed good correlation: at an S/P cut-off of 15% only one VNT positive sample tested negative in the SBV ELISA. The diagnostic sensitivity of the ELISA, relative to the VNT, was 98.8% (95% CI: 93.3-100.0%). The ELISA showed a high repeatability (cv=6.5%) and reproducibility (100% agreement). It was concluded that this ELISA is a suitable test method for the detection of SBV antibodies in sera from cows, sheep and, possibly, goats., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

At the end of 2011, a new Orthobunyavirus was discovered in Germany and named Schmallenberg virus (SBV). In the Netherlands malformations in new-born ruminants were made notifiable from the 20th of December 2011. After a notification, malformed new-borns were necropsied and brain tissue was sampled for reverse transcription-polymerase chain reaction (RT-PCR). In addition, blood samples from mothers of affected new-borns were tested for antibodies in a virus neutralization test (VNT). The aim of this study was to summarize and evaluate the diagnostic data obtained and to gain insight into the possible regional differences. In total 2166 brains were tested: 800 from lambs, 1301 from calves and 65 from goat kids. Furthermore 1394 blood samples were tested: 458 from ewes, 899 from cows and 37 from goats. Results showed that 29% of the lamb brains, 14% of the calf brains, and 9% of the goat kid brains were RT-PCR positive. The number of malformed and RT-PCR positive lambs decreased over time while the number of malformed and RT-PCR positive calves increased. In the VNT 92% of the ewes, 96% of the cows and 43% of the goats tested positive. Combining RT-PCR and VNT results, 18% of all farms tested positive in both the RT-PCR and VNT. The relative sensitivity and specificity of the RT-PCR are 19% and 97% respectively, and of the VNT 99% and 6%. The results show a widespread exposure to SBV and the regional evaluation seems to indicate an introduction of SBV in the central/eastern part., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The emergence of Schmallenberg virus (SBV), a novel orthobunyavirus, in ruminants in Europe triggered a joint veterinary and public health response to address the possible consequences to human health. Use of a risk profiling algorithm enabled the conclusion that the risk for zoonotic transmission of SBV could not be excluded completely. Self-reported health problems were monitored, and a serologic study was initiated among persons living and/or working on SBV-affected farms. In the study set-up, we addressed the vector and direct transmission routes for putative zoonotic transfer. In total, 69 sheep farms, 4 goat farms, and 50 cattle farms were included. No evidence for SBV-neutralizing antibodies was found in serum of 301 participants. The lack of evidence for zoonotic transmission from either syndromic illness monitoring or serologic testing of presumably highly exposed persons suggests that the public health risk for SBV, given the current situation, is absent or extremely low.

Background: At the end of 2011, a new orthobunyavirus, tentatively named Schmallenberg virus (SBV), was discovered in Germany. This virus has since been associated with clinical signs of decreased milk production, watery diarrhoea and fever in dairy cows, and subsequently also with congenital malformations in calves, lambs and goat kids. In affected countries, initial surveillance for the infection was based on examination of malformed progeny. These suspicions were followed up by real-time reverse transcription polymerase chain reaction (RT-PCR) on brain tissue. For epidemiological purposes, a serological assay was, however, needed., Results: A virus neutralisation test (VNT) was developed and optimized, and subsequently evaluated. This VNT has a specificity of >99% and the sensitivity is likely also very close to 100%. The assay is highly repeatable and reproducible. The final assay was used to test for antibodies in cows, ewes and does from herds known to be infected or suspected to be so. Targets for sampling in these herds were the mothers of malformed offspring. In herds with an RT-PCR confirmed SBV infection, more than 94% (190 out of 201) of the ewes and 99% (145 out of 146) of the cows were seropositive. In herds with suspicion of SBV infection based on birth of malformed offspring only (no or negative RT-PCR), more than 90% (231 out of 255) of the ewes and 95% (795 out of 834) of the cows were seropositive. In goats, on the other hand, only a low number of seropositives was found: overall 36.4%, being 16 out of 44 goats tested., Conclusions: Given the characteristics of this VNT, it can be used at a relative high throughput for testing of animals for export, surveillance, screening and research purposes, but can also be used as a confirmation test for commercially available enzyme-linked immunosorbent assays (ELISA's) and for (relative) quantification of antibodies.Suspicions of SBV infections that were confirmed by RT-PCR were almost always confirmed by serology in cows. Due to individual registration and identification of cows and calves, affected offspring could almost always be traced back to the mother. Ewes on the other hand were not always the mothers of affected lambs, but were in many cases herd mates with unaffected lambs. This indicated a high within-herd seroprevalence of antibodies against SBV.

Infections with Schmallenberg virus (SBV) are associated with congenital malformations in ruminants. Because reporting of suspected cases only could underestimate the true rate of infection, we conducted a seroprevalence study in the Netherlands to detect past exposure to SBV among dairy cattle. A total of 1,123 serum samples collected from cattle during November 2011-January 2012 were tested for antibodies against SBV by using a virus neutralization test; seroprevalence was 72.5%. Seroprevalence was significantly higher in the central-eastern part of the Netherlands than in the northern and southern regions (p<0.001). In addition, high (70%-100%) within-herd seroprevalence was observed in 2 SBV-infected dairy herds and 2 SBV-infected sheep herds. No significant differences were found in age-specific prevalence of antibodies against SBV, which is an indication that SBV is newly arrived in the country.