Ruminant pestiviruses genetic diversity: The key to detection and control

Ruminant pestiviruses are worldwide problems for the bovine industry. We reported the genotyping of a BVDV-2 from dromedary camels that can also infect cattle and goats. Nucleotide and amino acid sequence analysis of several regions of the genome of the camel pestivirus in comparison of other ruminant pestiviruses indicated the presence of a continuum of genetic and antigenic mutations within pestiviruses across genotypes. This demonstrated the need for a pestivirus vaccine than can be use to broaden the capacity for immune response across the three pestivirus genotypes affecting ruminants. Genetic analysis and reports of genetic recombinations between vaccinal and field strains of pestiviruses were use to determine the isolates more suitable for inclusion in the new vaccine. A series of mammalian expression plasmids containing the C and Erns genes of BVDV-1, BVDV-2, and BDV were constructed. We demonstrated pestivirus protein expression in NIH/3T3 cells. A pool of the DNA plasmids was used to immunize Balb/c mice. Compared to control groups, DNA vaccinates exhibited a significantly higher cell-mediated and humoral responses to the three ruminant pestivirus genotypes. Overall, our results warrant future protection studies in farm ruminants. Nucleotide sequence analysis data was also used to develop pestivirus-NS3-type specific molecular beacons. This approach allowed simultaneous detection and typing of single-type as well as mixed BVDV-1/BVDV-2 RNA in a single tube RT-PCR using a single set of universal pestivirus NS3 primers. We recommend using NS3 typing as an anchor for pestivirus genotyping in diagnostic settings and for developing of virus-characteristic genetic profiles to allow tracking vaccinal and field isolates in the environment. The pestivirus-NS3-type-specific molecular beacons were also used to identify the effect of mixed BVDV-1 BVDV-2 mixed infection in bovine and ovine cells. Our data demonstrated that replication of superinfecting cytopathic BVDV-1 RNA is inhibited in the presence of prior noncytopathic BVDV-2 RNA replication however; there was little effect on the RNA replication of simultaneously co-infecting viruses. Our work provides a different perspective of some aspects of the pestivirus genetic diversity that can be used in developing effective control and monitoring tools.