title: Development of a candidate DNA vaccine against Maedi-Visna virus.
abstract: DNA vaccine candidates against Maedi-Visna virus (MVV) infection in ovines were developed as an alternative to conventional vaccines. Candidates were constructed by cloning genes encoding the MVV gag polyprotein and gag proteins p16 and p25 fused to a beta-galactosidase reporter in a plasmid backbone. Transfection of different ovine cells showed a higher protein expression with plasmid lacZp16, which was hence further optimised by (i) removing a putative inhibitory sequence via reduction of the AU-content in the p16 gene or by (ii) introducing a secretory signal (Sc) to promote antigen secretion and increase its presentation to APCs. Unexpectedly, plasmids constructed on the basis of the first strategy by mutagenesis of lacZp16 (lacZp16mut(24)), led to a reduction in the expression of the antigen/reporter fusion in cultured ovine cells. This indicates that the high AU content in MVV does not inhibit protein expression. However, mice primed with lacZp16mut(24) and boosted with MVV protein displayed higher humoral response when compared with control lacZp16. The addition of the Sc signal (Sc-p16) led to lower amounts of intracellular antigen/reporter fusion in transfected ovine cells, thus confirming secretion. These findings correlate with in vivo experiments, which showed that mice primed with Sc-p16 and boosted with MVV exhibited stronger antibody responses when compared with control mice primed with lacZp16 and boosted with MVV. Stronger humoral responses were recorded by immunising mice with (i) Sc-p16 and lacZp16mut(24) plasmids together or with (ii) one plasmid containing both the mutations and the Sc signal.
authors.affiliation: Institute for Biotechnology and Bioengineering, Centre for Biological and Chemical Engineering, Lisboa, Portugal.
authors.forename: Ana M
authors.initials: AM
authors.lastname: Henriques
authors.forename: Miguel
authors.initials: M
authors.lastname: Fevereiro
authors.forename: Duarte M F
authors.initials: DM
authors.lastname: Prazeres
authors.forename: Gabriel A
authors.initials: GA
authors.lastname: Monteiro
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D000956:Antigens, Viral / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D005260:Female; D005814:Genes, Viral; D051379:Mice; D008807:Mice, Inbred BALB C; D011021:Pneumonia, Progressive Interstitial, of Sheep / Q000276:immunology* / Q000517:prevention & control; D012756:Sheep / Q000276:immunology* / Q000821:virology*; D013997:Time Factors; D019444:Vaccines, DNA / Q000276:immunology*; D014765:Viral Vaccines / Q000276:immunology*; D014790:Visna-maedi virus / Q000276:immunology*
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral; D019444:Vaccines, DNA; D014765:Viral Vaccines