RESUMEN

Bovine viral diarrhoea virus (BVDV) infection of cattle causes a diverse range of clinical outcomes from being asymptomatic, or a transient mild disease, to producing severe cases of acute disease leading to death. Four groups of calves were challenged with a type 1 BVDV strain, originating from a severe outbreak of BVDV in England, to study the effect of viral dose and immunosuppression on the viral replication and transmission of BVDV. Three groups received increasing amounts of virus: Group A received 10(2.55)TCID50/ml, group B 10(5.25)TCID50/ml and group C 10(6.7)TCID 50/ml. A fourth group (D) was inoculated with a medium dose (10(5.25)TCID50/ml) and concomitantly treated with dexamethasone (DMS) to assess the effects of chemically induced immunosuppression. Naïve calves were added as sentinel animals to assess virus transmission. The outcome of infection was dose dependent with animals given a higher dose developing severe disease and more pronounced viral replication. Despite virus being shed by the low-dose infection group, BVD was not transmitted to sentinel calves. Administration of dexamethasone (DMS) resulted in more severe clinical signs, prolonged viraemia and virus shedding. Using PCR techniques, viral RNA was detected in blood, several weeks after the limit of infectious virus recovery. Finally, a recently developed strand-specific RT-PCR detected negative strand viral RNA, indicative of actively replicating virus, in blood samples from convalescent animals, as late as 85 days post inoculation. This detection of long term replicating virus may indicate the way in which the virus persists and/or is reintroduced within herds.

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RESUMEN

Since bovine virus diarrhoea (BVD) was recognised as a unique disease complex, many different diagnostic methods have been used to detect the BVD virus (BVDV) itself, or immunity to BVDV. Of those that have evolved along with the current demands for accurate diagnostic tests, two categories are of interest for BVD control programmes. As reference assays, virus isolation and detection of virus neutralising antibodies are both carried out using cell cultures, which are time, resource and skill demanding. Enzyme immuno-assays are better suited for screening of large series of samples, and several variants of these have been developed for detection of both antibodies and viral antigens. Of other methods adapted for rapid diagnostic use are immunohistochemistry, flow cytometry and the reverse transcription-polymerase chain reaction. Basic properties of these and other methods are reviewed, with emphasis on the need for diagnostic assays in control programmes for BVD.

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In Europe, nationwide BVD control programs based on the TC principle have been running for up to 10 years in the Nordic countries. The results have shown that BVD eradication by removal of PI animals without use of vaccines is effective and that today's diagnostic tests, when used by experienced diagnosticians, are suitable for this task. Furthermore, to avoid control programs becoming Sisyphean tasks, adherence to strict biosecurity guide-lines to minimize infection of susceptible herds is a crucial additional measure. Efficient organization of testing, with sufficient capacity of diagnostic laboratories, is also important to minimize the period of overlap when remaining infected and recently emerged naive herds coexist close to each other. Control programs based on voluntary participation are possible, but when approaching final clearance of a national herd, significant delays can easily be suffered if any herd owners are allowed not to clear their herds. The control schemes used in the Nordic countries were tailored to fit the structure of the cattle production in each country. If BVD control programs based on the same principles are to be set up for other bovine populations,it is important to recognize and take into account for differences in the epidemiology of BVD and in the structure of the animal production,including cattle density and husbandry practices. To ensure optimal performance of the diagnostic tests, the diversity of BVDV in the region to be tested should also be considered.

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A selection of 43 bovine viral diarrhoea viruses isolated from mainly persistently infected cattle on 23 Slovenian farms between 1997 and 2001 were characterised genetically. Viral RNA was extracted from infected cell cultures, reverse transcribed and amplified by PCR with primers targeting the 5'-UTR and the N(pro) gene, followed by direct sequencing of purified PCR products obtained for both genomic regions. The N(pro) sequences provided the best genetic resolution, and gave also higher statistical support for phylogenetic classification of the viruses. Thirty-eight of the Slovenian isolates were of genetic subtypes 1d and 1f, four were 1b, and one subtype 1g. No BVDV type 2 viruses were found. This genetic prevalence matched those previously reported for neighbouring countries, as opposed to findings reported for more distant European countries, e.g. France, Spain and the UK. From eight cattle herds several virus isolates were analysed; with one exception all isolates from each herd were of the same genetic group. Extended sequencing of the N(pro) and part of the C gene of virus isolates with identical 5'-UTR sequences allowed differentiation between isolates obtained at different times from one herd.

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