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The paper presents the current state of knowledge on the prevalence and risk of a new type of bovine viral diarrhea virus known as type 3 (BVDV-3). The first discovered atypical pestivirus was the isolate D32/00_’HOBI’, detected in fetal calf serum (FCS) originating from Brazil. The isolates CH-KaHO/cont, SVA/cont-08 and IZSPLV_To are further examples confirming the presence of BVDV type 3 in the FCS. This new species of pestiviruses (BVDV-3) is a problem not only for research laboratories using bovine serum, but also for cattle breeders. Natural infections with this virus have been reported in Brazil, Thailand and Italy, which may suggest that the new species of pestivirus is also present in European cattle. The methods used for routine diagnosis of BVDV infection are ineffective in detecting atypical pestiviruses, which may pose a risk of false negative results. It may also influence the safety of vaccines and biological products produced on the basis of contaminated batches of fetal bovine serum.
The aim of the study was to use real time RT-PCR for the detection of genetic material of bovine viral diarrhea virus (BVDV) type 1 and type 3 in serum and milk samples. Material tested included the fetal calf serum used for cell culture, serum samples from healthy calves and from calves experimentally inoculated with BVDV type 1 and type 3, and milk samples (pasteurized and treated with ultra high temperature). Sensitivity of the test was 200 copies of RNA per reaction (10⁵ viral RNA copies per ml) for both types of BVDV, using dedicated primers and standards. Detection limit was 10² tissue culture infectious dose 50 (TCID₅₀) and 1 TCID₅₀ for type 1 and type 3, respectively. Diagnostic specificity of the method was 100%. Out of 10 samples of milk, 3 were positive for BVDV type 1, while none was positive for type 3. On the other hand, BVDV type 3 was found in 6 out of 10 samples of fetal calf serum. Real Time RT-PCR for BVDV type 1 and type 3 proved to be a highly sensitive and highly specific technique, enabling the detection of viral genetic material in various samples, even when its detection by virus isolation or antigen ELISA tests is impossible because of virus inactivation by such processes as high temperature, gamma irradiation, or the presence of virus neutralizing antibodies.
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