? Immunogenicity of recombinant E2 protein of bovine viral diarrhea virus expressed in transgenic alfalfa plants

MARIA SOL PEREZ-AGUIRREBURUALDE; CRISTINA GOMEZ; GUILLERMO ALBANESI; ANDREA PECORA; FERNANDO ARDILA; ANDRES WIGDOROVITZ; MARIA JOSE DUS SANTOS

VERONA

Congreso; Plant-based bacines, antibodies and biologics; 2013

PBVA

Immunogenicity of recombinant E2 protein of bovine viral diarrhea virus expressed in transgenic alfalfa plants Aguirreburualde Perez, M.S.1, Gómez, C.2, Albanesi, G.1, Pecora, A.1, Ardila, F.2, Wigdorovitz, A.1, Dus Santos, M.J.1 1Institute of Virology, CICVyA, (Buenos Aires, Argentina)-INTA Castelar 2Institute of Genetics, CICVyA, (Buenos Aires, Argentina)-INTA Castelar Email: solperezburu@gmail.com Bovine viral diarrhea virus (BVDV) is a pestivirus of worldwide distribution and is a major cause of economic losses in livestock. The main control strategies are vaccination and segregation of persistently infected animals. As prevention tool subunit vaccines offer many advantages, however, the challenge is to generate a protective immune response at an affordable cost for veterinary applications. Previously, our group reported the development of a subunit experimental vaccine based on transgenic alfalfa plants expressing a truncated version of the E2 glycoprotein of BVDV without the transmembrane domain and fused to a single chain antibody against MHCII (APCH-E2T). In order to assess the immunogenic properties of the vaccine 18 (n = 6 each group) female pregnant cows were randomly divided into 3 groups. One group was immunized with 2 doses of APCH 3ug-E2T, with an interval of 30 days between doses. Following the same design, the positive control group was immunized with an inactivated virus vaccine with viral titer of 107 TDIC 50/ml (BVDV-1a) and the negative control group with a negative extract of a non transgenic alfalfa plant. Serum samples and PBMC s were collected at various time points for the measurement of BVDV-antibodies and cellular immune response. Also, after birth calves were sampled to evaluate the transference of passive immunity through colostrums. The humoral response was assessed by serum neutralization test against homologous and heterologous virus and by blocking and avidity ELISA assays. To assess the cellular immune response, mononuclear cells were purified by density gradient Ficoll-Paque Plus to proceed with LPDA (antigen-dependent lymphoproliferation) against the purified E2t protein expressed in mammalian cells. Cell proliferation was evaluated using the technique of thymidine incorporation and the results were expressed as stimulation index. In parallel to this test, CFSE staining was performed. The experimental trial involving animals was approved by the Institutional Committee for the Care and Use of Experimental Animals of INTA (CICUAE). Subunit vaccine induced neutralizing antibody titers against the homologous virus vaccine strain higher than 3 (evaluated by Reed-Muench´s method) being similar to those generated by the whole virus vaccine. Furthermore, the antibodies curves after vaccination showed a dynamic of increased avidity similar in both groups. In concern to the cellular response the two methodologies tested to evaluate T cell activation showed similar levels of proliferation, comparable to the control group, demonstrating efficient priming of the lymphocyte population by the antigen expressed in transgenic plants. Also, the evaluation of calves? serum samples showed an efficient transference of humoral immunity in the cows vaccinated with the experimental vaccine during the first two month of life. In conclusion, these results along with the efficiency afforded by the vaccine against viral challenge in the natural host support the foreseeable potential of the application of this vaccine as control strategy at herd level.