Comparison of five diagnostic methods for detecting bovine viral diarrhea virus infection in calves

Five diagnostic techniques performed on skin biopsies (shoulder region) and/or serum were compared for detection of bovine viral diarrhea virus infection in 224 calves 0-3 months of age, 23 calves older than 3 months but younger than 7 months, and 11 cattle older than 7 months. The diagnostic methods used were immunohistochemistry (IHC), 2 commercial antigen ELISAs, 1 commercial antibody ELISA, and real-time RT-PCR. Results of 249 out of 258 skin and serum samples were identical and correlated within the 3 antigen detection methods and the real-time RT-PCR used. Twenty-six of these 249 samples were BVDV-positive with all antigen detection methods and the real-time RT-PCR. Nine out of 258 samples yielding discordant results were additionally examined by RT-PCR, RT-PCR Reamplification (ReA), and antigen ELISA I on serum and by immunohistochemistry on formalin fixed and paraffin-embedded skin biopsies. Virus isolation and genotyping was performed as well on these discordant samples. In 3 cases, transiently infected animals were identified. Two samples positive by real-time RT-PCR were interpreted as false positive and were ascribed to cross-contamination. The antigen ELISA II failed to detect 2 BVDV-positive calves due to the presence of maternal antibodies; the cause of 2 false-positive cases in this ELISA remained undetermined. Only persistently infected animals were identified in skin samples by IHC or antigen ELISA I. The 3 antigen detection methods and the real-time RT-PCR used in parallel had a high correlation rate (96.5%) and similar sensitivity and specificity values

A novel panel of monoclonal antibodies against Schmallenberg virus nucleoprotein and glycoprotein Gc allows specific orthobunyavirus detection and reveals antigenic differences

International audienceA panel of monoclonal antibodies (mAbs) specific for the nucleocapsid (N) protein or the glycoprotein Gc of Schmallenberg virus (SBV), a novel member of the Simbu serogroup (genus Orthobunyavirus, family Bunyaviridae), was produced and used to analyze antigenic differences among members of this serogroup. Reactivity with various SBV-isolates and other Simbu serogroup viruses was assessed by an indirect immunofluorescence test and by immunoblotting. The Gc-specific mAbs detected different SBV isolates as well as two closely related members of the Simbu serogroup. In addition, one mAb showed a highly specific reactivity with the homologous SBV strain only. Based on their differing reactivity with different SBV-strains, these antibodies represent a valuable novel tool to rapidly determine the phenotype of new SBV isolates. In contrast, the N-specific mAbs showed a broad reactivity spectrum and detected not only all the tested SBV-isolates, but also several other viruses of the Simbu serogroup. One out of these mAbs even recognized all of the tested Simbu serogroup viruses in the indirect immunofluorescence assay. In order to further characterize the N-specific antibodies, PepScan analysis was performed and a specific epitope could be identified. In summary, the newly generated mAbs showed differing pan-Simbu virus-, pan-SBV- as well as SBV-isolate-specific reactivity patterns. Thus, they represent valuable tools for the development of novel antigen and antibody detection systems either specific for SBV or, in a broader approach, for the pan-Simbu serogroup diagnostics