Insights into the dynamics of endemic Coxiella burnetii infection in cattle by application of phase-specific ELISAs in an infected dairy herd. Veterinary Microbiology, Elsevier, 2011, 151 (3-4) This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.Page 1 are available, they are not yet used in ruminants as they are in humans. This study focuses on 14 phase-specific serology as a tool for analysis of the dynamics of infection in cattle. As a 15 prerequisite, sero-prevalence in Bavarian cattle (1) and sero-prevalences for age-groups (2) were 16 determined by ELISA (CHEKIT Q-Fever; mix of PhI/PhII-antigen). Subsequently, phase-17 specific antigens were coated onto ELISA plates individually and tests were simultaneously 18 applied in an endemically infected herd with about 90 dairy cows and 250 calves/heifers

No abstract

S u m m a r y A new ELISA kit was developed, based on highly purified whole-virus antigen derived from the Swiss maedi-visna virus strain OLV. The sensitivity, specificity and accuracy of this assay were compared with that of an established ELISA based on recombinant GAG (groupspecific antigens)-GST (glutathione S-transferase) fusion protein expressed in E. coli (GAG-GST ELISA). The whole-virus ELISA exhibits at least comparable specificity (99.3 Yo) but higher sensitivity (98.6 versus 86.3 Yo) and agreement with the 'true' status beyond chance in the detection of antiviral antibodies in serum from goats. Antibodies in milk samples are detected with higher specificity (98.9 versus 97.8 Yo) but lower sensitivity (91.4 versus 98.2 YO) than in GAG-GST ELISA. The specificity of the new ELISA in the detection of antibodies in serum might be superior, since a set of 40 samples falsely rated positive in GAG-GST ELISA in routine diagnostic work was negative in the new ELISA. In both assays, milk samples can be tested instead of serum, although with slightly reduced sensitivity in the new ELISA. The major advantage of the new test kit is the low number of equivocal samples needing confirmation in a supplementary test. Results obtained with sheep sera indicate that the new ELISA kit is also suitable for the detection of antibodies to maedi-visna virus. Recombinant GAG-GST fusion proteinRecombinant GAG-GST fusion protein derived from Dutch maedi-visna virus strain ZZV 1050 was produced in E . coli transformed with pGEX-2T/GAG(850) and purified as described by ZANONI et al. (1991b).ELISA and immunoblotting ELISA microtitre trays were coated monophasically with appropriate dilutions of highly purified virus in phosphate-buffered saline (PBS: 0.14 M NaC1, 2.6 mM KCI, 6.6 mM Na2HP04.12 H20, 1.5 mM KH2P0,1 p H 7.0), incubated overnight at 4 "C, blocked with 1 % chicken ovalbumin in PBS, washed, dried and stored at 4 "C after sealing with plastic foil. Coated microtitre trays and reagents were purchased as a kit from the manufacturer and ELISA was

BackgroundTick-borne encephalitis (TBE) is the most important viral tick borne zoonosis in Europe. In Germany, about 250 human cases are registered annually, with the highest incidence reported in the last years coming from the federal states Bavaria and Baden-Wuerttemberg. In veterinary medicine, only sporadic cases in wild and domestic animals have been reported; however, a high number of wild and domestic animals have tested positive for the tick-borne encephalitis virus (TBEV) antibody.Case presentationIn May 2015, a five-month-old lamb from a farm with 15 Merino Land sheep and offspring in Nersingen/Bavaria, a TBEV risk area, showed impaired general health with pyrexia and acute neurological signs. The sheep suffered from ataxia, torticollis, tremor, nystagmus, salivation and finally somnolence with inappetence and recumbency. After euthanasia, pathological, histopathological, immunohistochemical, bacteriological, parasitological and virological analyses were performed. Additionally, blood samples from the remaining, healthy sheep in the herd were taken for detection of TBEV antibody titres. At necropsy and accompanying parasitology, the sheep showed a moderate to severe infection with Trichostrongylids, Moniezia and Eimeria species. Histopathology revealed mild to moderate necrotising, lymphohistiocytic and granulocytic meningoencephalitis with gliosis and neuronophagia. Immunohistochemistry for TBEV was negative. RNA of a TBEV strain, closely related to the Kumlinge A52 strain, was detected in the brain by quantitative reverse transcriptase polymerase chain reaction (RT-qPCR) and subsequent PCR product sequencing. A phylogenetic analysis revealed a close relationship to the TBEV of central Europe. TBEV was cultured from brain tissue. Serologically, one of blood samples from the other sheep in the herd was positive for TBEV in an enzyme-linked immunosorbent assay (ELISA) and in a serum neutralisation test (SNT), and one was borderline in an ELISA.ConclusionTo the authors’ knowledge this is the first report of a natural TBEV infection in a sheep in Europe with clinical manifestation, which describes the clinical presentation and the histopathology of TBEV infection.

Recently, several so-called “atypical” Bluetongue virus (BTV) serotypes were discovered, including BTV-25 (Toggenburg virus), in Switzerland. Most “atypical” BTV were identified in small ruminants without clinical signs. In 2018, two goats from a holding in Germany tested positive for BTV-25 genome by RT-qPCR prior to export. After experimental inoculation of the two goats with the BTV-25 positive field blood samples for generation of reference materials, viremia could be observed in one animal. For the first time, the BTV-25-related virus was isolated in cell culture from EDTA-blood and the full genome of isolate “BTV-25-GER2018” could be generated. BTV-25-GER2018 was only incompletely neutralized by ELISA-positive sera. We could monitor the BTV-25 occurrence in the respective affected goat flock of approximately 120 goats over several years. EDTA blood samples were screened with RT-qPCR using a newly developed BTV-25 specific assay. For serological surveillance, serum samples were screened using a commercial cELISA. BTV-25-GER2018 was detected over 4.5 years in the goat flock with intermittent PCR-positivity in some animals, and with or without concomitantly detected antibodies since 2015. We could demonstrate the viral persistence of BTV-25-GER2018 in goats for up to 4.5 years, and the first BTV-25 isolate is now available for further characterization.