Development and evaluation of a replicon particle vaccine expressing the E2 glycoprotein of bovine viral diarrhea virus (BVDV) in cattle.

BACKGROUND: Bovine viral diarrhea virus is one of the most significant and costly viral pathogens of cattle worldwide. Alphavirus-derived replicon particles have been shown to be safe and highly effective vaccine vectors against a variety of human and veterinary pathogens. Replicon particles are non-propagating, DIVA compatible, and can induce both humoral and cell mediated immune responses. This is the first experiment to demonstrate that Alphavirus-based replicon particles can be utilized in a standard prime/boost vaccination strategy in calves against a commercially significant bovine pathogen. FINDINGS: Replicon particles that express bovine viral diarrhea virus sub-genotype 1b E2 glycoprotein were generated and expression was confirmed in vitro using polyclonal and monoclonal antibodies specific to E2. Vaccine made from particles was generated in Vero cells and administered to BVDV free calves in a prime/boost regimen at two dosage levels. Vaccination resulted in neutralizing antibody titers that cross-neutralized both type 1 and type 2 BVD genotypes following booster vaccination. Additionally, high dose vaccine administration demonstrated some protection from clinical disease and significantly reduced the degree of leukopenia caused by viral infection. CONCLUSIONS: Replicon particle vaccines administered in a prime/boost regimen expressing BVDV E2 glycoprotein can induce cross-neutralizing titers, reduce leukopenia post challenge, and mitigate clinical disease in calves. This strategy holds promise for a safe and effective vaccine to BVDV.

Analysis of nucleotides 13-96 of the human parainfluenza virus type 3 antigenomic promoter reveals positive- and negative-acting replication elements.

During replication of human parainfluenza virus type 3 (HPIV3), the 96-nucleotide antigenomic promoter (AGP) of HPIV3 directs the synthesis of genomic RNA. Previous work showed that nucleotides 1-12 were critical in promoting replication of an HPIV3 minireplicon, but nucleotides 13-96 were not investigated. In this study, the role of nucleotides 13-96 in AGP function was analyzed by creating and assaying mutations in an HPIV3 minireplicon. A replication promoting element known as promoter element II (nt 79-96) was confirmed in the HPIV3 AGP. Additionally, nucleotides 13-39 were found to constitute an additional positive-acting cis-element. However, detailed analysis of the 13-39 element revealed a complicated control element with both stimulatory and repressing elements. Specifically, nucleotides 21-28 were shown to repress RNA replication, while flanking sequences had a stimulatory effect.