Complete Genome Sequence of Border Disease Virus Genotype 3 Strain Gifhorn

The complete genome sequence of the genotype 3 border disease virus strain Gifhorn has been determined; this strain was originally isolated from pigs. This represents the consensus sequence for the virus used to produce the bacterial artificial chromosome (BAC) cDNA clone pBeloGif3, which yields a virus that is severely attenuated in cell culture

Detection and Typing of Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus by Multiplex Real-Time RT-PCR

Porcine reproductive and respiratory syndrome (PRRS) causes economic losses in the pig industry worldwide, and PRRS viruses (PRRSV) are classified into the two distinct genotypes “North American (NA, type 2)” and “European (EU, type 1)”. In 2006, a highly pathogenic NA strain of PRRSV (HP-PRRSV), characterized by high fever as well as high morbidity and mortality, emerged in swine farms in China. Therefore, a real-time reverse transcription polymerase chain reaction (RT-qPCR) assay specific for HP-PRRSV was developed and combined with type 1- and type 2-specific RT-qPCR systems. Furthermore, an internal control, based on a heterologous RNA, was successfully introduced. This final multiplex PRRSV RT-qPCR, detecting and typing PRRSV, had an analytical sensitivity of less than 200 copies per µl for the type 1-assay and 20 copies per µl for the type 2- and HP assays and a high diagnostic sensitivity. A panel of reference strains and field isolates was reliably detected and samples from an animal trial with a Chinese HP-PRRS strain were used for test validation. The new multiplex PRRSV RT-qPCR system allows for the first time the highly sensitive detection and rapid differentiation of PRRSV of both genotypes as well as the direct detection of HP-PRRSV

Pea-derived vaccines demonstrate high immunogenicity and protection in rabbits against rabbit haemorrhagic disease virus

Vaccines against rabbit haemorrhagic disease virus (RHDV) are commercially produced in experimentally infected rabbits. A genetically engineered and manufactured version of the major structural protein of RHDV (VP60) is considered to be an alternative approach for vaccine production. Plants have the potential to become an excellent recombinant production system, but the low expression level and insufficient immunogenic potency of plant-derived VP60 still hamper its practical use. In this study, we analysed the expression of a novel multimeric VP60-based antigen in four different plant species, including Nicotiana tabacum L., Solanum tuberosum L., Brassica napus L. and Pisum sativum L. Significant differences were detected in the expression patterns of the novel fusion antigen cholera toxin B subunit (CTB)::VP60 (ctbvp60(SEKDEL)) at the mRNA and protein levels. Pentameric CTB::VP60 molecules were only detected in N. tabacum and P. sativum, and displayed equal levels of CTB, at approximately 0.01% of total soluble protein (TSP), and traces of detectable VP60. However, strong enhancement of the CTB protein content via self-fertilization was only observed in P. sativum, where it reached up to 0.7% of TSP. In rabbits, a strong decrease in the protective vaccine dose required from 48-400 microg potato-derived VP60 [Castanon, S., Marin, M.S., Martin-Alonso, J.M., Boga, J.A., Casais, R., Humara, J.M., Ordas, R.J. and Parra, F. (1999) Immunization with potato plants expressing VP60 protein protects against rabbit hemorrhagic disease virus. J. Virol. 73, 4452-4455; Castanon, S., Martin-Alonso, J.M., Marin, M.S., Boga, J.A., Alonso, P., Parra, F. and Ordas, R.J. (2002) The effect of the promoter on expression of VP60 gene from rabbit hemorrhagic disease virus in potato plants. Plant Sci. 162, 87-95] to 0.56-0.28 microg antigenic VP60 (measured with VP60 enzyme-linked immunosorbent assay) of crude CTB::VP60 pea extracts was demonstrated. Rabbits immunized with pea-derived CTB::VP60 showed anti-VP60-specific antibodies, similar to RikaVacc((R))-immunized rabbits, and survived RHDV challenge

Comparison of Porcine Epidemic Diarrhea Viruses from Germany and the United States, 2014

Since 2013, highly virulent porcine epidemic diarrhea virus has caused considerable economic losses in the United States. To determine the relation of US strains to those recently causing disease in Germany, we compared genomes and found that the strain from Germany is closely related to variants in the United States

Detection and Identification of the Atypical Bovine Pestiviruses in Commercial Foetal Bovine Serum Batches

The recently emerging atypical bovine pestiviruses have been detected in commercial foetal bovine serum (FBS) of mainly South American origin so far. It is unclear how widely the viruses are presented in commercial FBS of different geographic origins. To further investigate the possible pestivirus contamination of commercially available FBS batches, 33 batches of FBS were obtained from ten suppliers and analysed in this study for the presence of both the recognised and the atypical bovine pestiviruses. All 33 batches of FBS were positive by real-time RT-PCR assays for at least one species of bovine pestiviruses. According to the certificate of analysis that the suppliers claimed for each batch of FBS, BVDV-1 was detected in all 11 countries and BVDV-2 was detected exclusively in the America Continent. The atypical pestiviruses were detected in 13 batches claimed to originate from five countries. Analysis of partial 5′UTR sequences showed a high similarity among these atypical bovine pestiviruses. This study has demonstrated, for the first time that commercial FBS batches of different geographic origins are contaminated not only with the recognised species BVDV-1 and BVDV-2, but also with the emerging atypical bovine pestiviruses

Evaluation of a commercial E(rns)-capture ELISA for detection of BVDV in routine diagnostic cattle serum samples

BACKGROUND: Bovine viral diarrhoea virus (BVDV) is an important pathogen in cattle. The ability of the virus to cross the placenta during early pregnancy can result in the birth of persistently infected (PI) calves. These calves shed the virus during their entire lifespan and are the key transmitters of infection. Consequently, identification (and subsequent removal) of PI animals is necessary to rapidly clear infected herds from the virus. The objective of this study was to evaluate the suitability of a commercial E(rns)-capture ELISA, in comparison to the indirect immunoperoxidase test (IPX), for routine diagnostic detection of BVDV within a control programme. In addition, the effect of passive immunity and heat-inactivation of the samples on the performance of the ELISA was studied. METHODS: In the process of virus clearance within the Swedish BVDV control programme, all calves born in infected herds are tested for virus and antibodies. From such samples, sent in for routine diagnostics to SVA, we selected 220 sera collected from 32 beef herds and 29 dairy herds. All sera were tested for BVDV antigen using the E(rns )ELISA, and the results were compared to the results from the IPX used within the routine diagnostics. RESULTS: All 130 samples categorized as virus negative by IPX were tested negative in the ELISA, and all 90 samples categorized as virus positive were tested positive, i.e. the relative sensitivity and specificity of the ELISA was 100% in relation to IPX, and the agreement between the tests was perfect. CONCLUSION: We can conclude that the E(rns )ELISA is a valid alternative that has several advantages compared to IPX. Our results clearly demonstrate that it performs well under Swedish conditions, and that its performance is comparable with the IPX test. It is highly sensitive and specific, can be used for testing of heat-inactivated samples, precolostral testing, and probably to detect PI animals at an earlier age than the IPX

Cytopathic bovine viral diarrhea viruses (BVDV): emerging pestiviruses doomed to extinction

Bovine viral diarrhea virus (BVDV), a Flaviviridae pestivirus, is arguably one of the most widespread cattle pathogens worldwide. Each of its two genotypes has two biotypes, non-cytopathic (ncp) and cytopathic (cp). Only the ncp biotype of BVDV may establish persistent infection in the fetus when infecting a dam early in gestation, a time point which predates maturity of the adaptive immune system. Such fetuses may develop and be born healthy but remain infected for life. Due to this early initiation of fetal infection and to the expression of interferon antagonistic proteins, persistently infected (PI) animals remain immunotolerant to the infecting viral strain. Although only accounting for some 1% of all animals in regions where BVDV is endemic, PI animals ensure the viral persistence in the host population. These animals may, however, develop the fatal mucosal disease, which is characterized by widespread lesions in the gastrointestinal tract. Cp BVD virus, in addition to the persisting ncp biotype, can be isolated from such animals. The cp viruses are characterized by unrestrained genome replication, and their emergence from the persisting ncp ones is due to mutations that are unique in each virus analyzed. They include recombinations with host cell mRNA, gene translocations and duplications, and point mutations. Cytopathic BVD viruses fail to establish chains of infection and are unable to cause persistent infection. Hence, these viruses illustrate a case of “viral emergence to extinction” – irrelevant for BVDV evolution, but fatal for the PI host