Abstract
African swine fever is a viral haemorrhagic disease of domestic pigs and wild boar and is a serious threat to the economy of many countries. No vaccines are available to induce protection in pigs against this lethal disease. The most promising approach for vaccine development is the use of modified live attenuated vaccines (MLAV) and these can confer high levels of protection. One MLAV candidate is a gene-deleted genotype I Benin 97/1 strain (BeninΔDP148R), which lacks the DP148R gene. The BeninΔDP148R confers 100% protection in immunized pigs following challenge with parental pathogenic Benin 97/1, but adverse clinical signs associated with viremia and extended persistence of virus were observed. In this thesis, the mechanism by which the ASFV CD2v transmembrane protein, coded for by the EP402R gene, binds to red blood cells was investigated. Functional amino acid residues in CD2v that are involved in binding red blood cells (HAD) were identified by introducing single or multiple amino acid substitutions into the predicted ligand-binding domain on the CD2v protein. Mutations of the residues glutamic acid at position 99 or tyrosine at position 102 were shown to reduce HAD to the expressed CD2v protein. However, the expression of the ASFV membrane protein EP153R was shown to rescue the reduced HAD capacity of mutant CD2v proteins. This indicated that EP153R protein can augment this function of CD2v. To examine the effect of reducing the capacity of ASFV to cause HAD on virulence, levels and persistence of the virus in blood, two recombinant viruses were tested by immunisation of pigs. In the first virus deletion of the EP402R gene from the BeninDP148R virus genome was shown to dramatically reduce the period of virus persistence in blood, although moderate levels of virus and clinical signs were observed. In the second virus, the EP153R gene was deleted and the EP402R gene was replaced with a gene expressing a non-HAD CD2v with the amino acid residue glutamic acid at position 99 substituted with arginine in the genome of ASFV BeninΔDP148R. Immunisation of pigs showed the virus persistence in blood was reduced to 2 days, the onset and duration of clinical signs were decreased (1 day) and the viremia was also reduced by approximately 3 thousand-fold. This demonstrated that deletion of EP153R and the insertion of mutant CD2vE99R balanced the level of virus attenuation, and hence resulted in a reduced virus persistence, mild clinical signs for a reduced duration, a decrease in viremia and provided overall protection of 83.3%. This recombinant virus has an increased safety profile compared to the parental virus (BeninΔDP148R) and is a promising vaccine candidate.