Development and evaluation of an indirect enzyme-linked immunosorbent assay for serological detection of Schmallenberg virus antibodies in ruminants using whole virus antigen

Abstract

BackgroundIn late 2011, a new Orthobunyavirus of the Simbu serogroup named Schmallenberg virus (SBV) emerged in continental Europe. The virus is transmitted by hematophagous arthropods, with the Culicoides species as, so far known, main vectors. Infection with the virus can cause clinical signs in adult ruminants including diarrhea, fever and reduced milk production. Transplacental infection of the developing fetus can lead to malformations of varying severity. To assess seroprevalence of SBV in Sweden an indirect enzyme-linked immunosorbent assay (ELISA) was established in connection with the surveys. Here, we describe the development and evaluation of the indirect ELISA, based on whole virus as the coating antigen and a monoclonal antibody for the detection of antibodies to SBV in ruminant sera. The evaluation includes comparison between the in-house ELISA, virus neutralization test and an indirect commercial ELISA.ResultsThe optimal working dilutions of antigens and conjugate were estimated with checkerboard titrations. Comparative studies, including ROC analyses, were used for the selection of an optimal cut-off (S/P value = sample value as percentage of positive control value). With an estimated S/P value of 15% the whole virus ELISA showed a specificity of 100% and a sensitivity of 99.19% compared to virus neutralization test (VNT) and with a good consistency as shown in reproducibility and variability experiments. Furthermore, the comparison of our whole virus indirect ELISA to an indirect ELISA with a SBV nucleoprotein antigen, demonstrated a higher sensitivity of our test.ConclusionThe indirect whole virus ELISA described in this paper is a readily available test for serological analysis of SBV antibodies. Since this in-house ELISA demonstrates a specificity and sensitivity comparable to virus neutralization test and also shows a higher sensitivity compared to commercially available indirect ELISA, it is a useful alternative for surveillance and screening purposes of SBV.

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

“…The neutralizing titer of a serum was determined as the highest dilution in which the cell monolayer was intact. A serum sample was considered negative if cpe was observed at and from a dilution of 1:8 [18, 19]. …”

Establishment and Evaluation of an Indirect ELISA for Detection of Antibodies to Goat Klebsiella Pneumonia

“…The antigenic cross-reactivity test results indicated that the indirect ELISA did not cross-react with common bacteria and was capable of detecting anti-sera to Klebsiella pneumoniae. Both the intra-assay and inter assay CVs were lower than 10%,which suggested that the method has good reproducibility [29]. To further evaluate the e cacy of the method, additional tests in clinical veterinary settings are essential.…”

Development of a Whole-Virus ELISA for Serological Evaluation of Domestic Livestock as Possible Hosts of Human Coronavirus NL63

“…The competitive ELISA technique has an added advantage of no requirement for sample clean-up and a high sensitivity to differences in composition of complex mixtures of different antigens even in the presence of relatively small quantities of the specific detection antibody [51,52]. Whole virus antigen preparations like the one used in this study have generally been found to be more sensitive than recombinant antigen targets [53,54] but tends to be less specific as a result of higher likelihood of non-specific binding of co-purified cellular proteins and non-target viral proteins [55,56].…”