Analysis of variability and antigenic peptides prediction of E2 BVDV glycoprotein in a mucosal disease affected animal

The aim of our work was to study the genetic variability of BVDV and the quasispecies nature of the virus. Clones were obtained by cloning PCR products of E2 viral genome starting from two organs of a mucosal disease affected animal and a prediction of the antigenic sites of E2 protein portion was then performed. Two PCR products of E2 region of BVDV with genetic variability obtained from organs of a MD affected animal were cloned and screened with SSCP technique. Two or more clones with the same pattern in SSCP analysis were sequenced and the amino acid sequences were predicted and analyzed for theirs antigenic properties by several programs. Three different SSCP banding patterns were obtained from abomasums derived cloned. Eleven clones out of 31 obtained from abomasum were sequenced and a predicted fragment of 125 amino acids of E2 protein were subjected analysis. Nine clones out of 61 obtained from tonsils were sequenced and the predicted amino acid sequences were analyzed. The antigenic analysis by Kolaskar and Tongaonkar method evidenced the presence of 4 immunogenic peptides in 9 clones obtained from the abomasums and two clones showed another antigen site at the position 16-23. This result was confirmed by the antigenic analysis based on Jameson-Wolf approach. In particular clone 10 showed an additional antigenic peak, in the correspondent graphic, between the amino acids 20-25. Except clone 10 no significant differences in the antigenic profile was evidenced in the other clones whereas similar profiles were obtained with Emini method. About clones from tonsils the prediction of antigenic peptides by Kolaskar and Tongaonkar method evidenced in 8 samples the 4 immunogenic peptides observed in the abomasum; only one clone presented the antigenic site at the position 16-23. Also in this case the application of method Jameson and Wolf showed in this clone an antigenic peak between 20-25 amino acids. All clones demonstrated the same profile by Emini analysis. The presence of viral variants in the abomasum and tonsils were evidenced by SSCP analysis showing the existence of viral quasispecies in a subject with mucosal disease. The antigenic index analysis and the prediction of regions on the surface of the E2 protein confirmed the high level of immunogenicity of this region. The antigenic analysis of predicted protein showed different regions able to induce a humoral immune response from the host. Some viral variants evidenced an additional antigenic site; however seems to be possible the selection of a viral population with a lower antigenic degree able to elude the immune system of the host. These results demonstrate the high evolutionary ability of BVDV and the skill to evade the immune response by the selection of viral population with a lower antigenic property.