Abstract
Baculoviruses are large DNA virus of insects principally employed in recombinant protein expression. Its ability to form occlusion bodies (OBs), which are composed mainly of polyhedrin protein (POLH), makes them biotechnologically attractive, as these crystals (polyhedra) can incorporate foreign peptides and can be easily isolated. On the other hand, peptide microarrays allow rapid and inexpensive high-throughput serological screening of new candidates to be incorporated to OBs. To integrate these 2 biotechnological approaches, we worked on Babesia bovis, one of the causative agents of bovine babesiosis. Current molecular diagnosis of infection with B. bovis includes enzyme-linked immunosorbent assay (ELISA) techniques, which use merozoite lysate obtained from infected bovine erythrocytes. However, it is important to produce recombinant antigens that replace the use of crude antigens. Here, we describe a new biotechnological platform for the design of indirect ELISAs based on 5 antigenic peptides of 15 amino acid residues of B. bovis (ApBb), selected from a peptide microarray and expressed as a fusion to POLH. An Sf9POLHE44G packaging cell line infected with recombinant baculoviruses carrying POLH-ApBb fusions yielded higher levels of chimeric polyhedra, highlighting the advantage of a trans-contribution of a mutant copy of polyhedrin. Finally, the use of dissolved recombinant polyhedra as antigens was successful in an ELISA assay, as B. bovis-positive sera recognized the fusion POLH-ApBb. Thus, the use of this platform resulted in a promising alternative for molecular diagnosis of relevant infectious diseases.
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Acknowledgements
We acknowledge Dr. Julia Sabio y García for the English language editing.
Funding
This work was funded by the Agencia Nacional de Promoción Científica y Tecnológica (ANPCyT), grant number PICT 2013-1048, and the Instituto Nacional de Tecnología Agropecuaria (INTA), grant number PNBIO 1131032.
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This article does not contain studies with human participants performed by any of authors. The protocol for animal handling and venipuncture was performed following the guidelines of the Institutional Committee for the Use and Care of Experimentation Animals (protocol approval No. 025/2011). All samples came from privately owned herds and were sampled with the approval of the owners.
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López, M.G., Pallarés, H.M., Alfonso, V. et al. Novel biotechnological platform based on baculovirus occlusion bodies carrying Babesia bovis small antigenic peptides for the design of a diagnostic enzyme-linked immunosorbent assay (ELISA). Appl Microbiol Biotechnol 102, 885–896 (2018). https://doi.org/10.1007/s00253-017-8662-1
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DOI: https://doi.org/10.1007/s00253-017-8662-1