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Conference abstract2008

Low-level bovine viral diarrhoea virus contamination of a bovine coronavirus strain not detected by routine screening

Tråven, Madeleine; LeBlanc, Neil; Ståhl, Karl; Kampa, Jaruwan; Belak, Sandor; Alenius, Stefan

Abstract

Low-level bovine viral diarrhoea virus contamination of a bovine coronavirus strain not detected by routine screening Aim To detect and type bovine viral diarrhoea virus (BVDV) contaminating a bovine coronavirus (BCV) laboratory strain. Methods Seroconversion to BVDV was detected in 4 calves experimentally infected with BCV. The BCV strain Munich W270/83 was originally isolated in 1983 and serially propagated in bovine foetal turbinate cells using bovine foetal serum. Virus stock, cells and the bovine foetal serum tested negative for BVDV using RT-PCR (1). Sera from the experimentally infected calves were tested for antibody responses to BVDV in an indirect ELISA (Svanova Biotech, Uppsala, Sweden) and in virus neutralization tests (VNT) against viruses representing BVDV type 1, 2 and atypical Pestivirus strain Hobi. The sera were also tested for BVDV RNA using general and type-specific probes in an RT-PCR based oligonucleotide suspension microarray designed to detect and differentiate the recognised pestivirus species (Luminex microspere system, 2). Results and Discussion All four calves, seronegative to BVDV prior to inoculation, had seroconverted to BVDV 21 days after inoculation with the BCV preparation. All four calves had VNT titres ≥512 to BVDV type 1 in sera 35 days after inoculation. One calf also had a VNT titre ≥512 to BVDV type 2 and 2 calves had VNT titres ≥512 to Hobi. Sera from all four calves taken 4 and 7 days after inoculation were positive for BVDV type 1 RNA in the Luminex microarray. The detection of other BVDV types, if present, may have failed due to specificity of the RT-PCR primers or competition. Thus BVDV type 1 contamination of the BCV cell culture preparation was confirmed. The VNT results from the experimentally inoculated calves, however, indicate that other pestiviruses may have been present. Conclusions Low-level contamination of BVDV in established laboratory strains of various viruses may occur despite RT-PCR testing for BVDV of cell culture preparations and bovine foetal serum products. Such contamination may lead to distorted research results and there is also a risk for contamination of live vaccines produced using such viral strains. Experimental inoculation of seronegative calves is a highly sensitive method to detect low-level BVDV contamination and can be used to confirm the status of products that need to be strictly BVDV free. References 1. Elvander M, Baule C, Persson M, Egyed L, Ballagi-Pordány A, Belák S & Alenius S. An experimental study of concurrent primary infection with bovine respiratory syncytial virus (BRSV) and bovine viral diarrhoea virus (BVDV) in calves. Acta Vet Scand 1998,39, 251-64. 2. Deregt D, Gilbert SA, Dudas S, Pasick J, Baxi S, Burton KM & Baxi MK. A multiplex DNA suspension microarray for simultaneous detection and differentiation of classical swine fever virus and other pestiviruses. J Virol Methods 2006, 136, 17-23

Published in

Title: Proc. 7th ESVV Pestivirus Congress

Conference

7th ESVV Pestivirus Congress