Abstract
BackgroundBabesia bovis reproduces sexually in the gut of its tick vector Rhipicephalus microplus, which involves expression of 6cys A and 6cys B proteins. Members of the widely conserved 6cys superfamily are candidates for transmission blocking vaccines (TBV), but intricacies in the immunogenicity of the 6cys proteins in the related Plasmodium parasites required the identification of transmission blocking domains in these molecules for vaccine design. Hereby, the immunogenic efficacy of recombinant (r) B. bovis 6cys A and B proteins as a TBV formulation was studied.MethodsThe immunogenicity of r6cys A and 6cys B proteins expressed in a eukaryotic system was evaluated in a cattle immunization trial (3 immunized and 3 control calves). A B. bovis sexual stage induction in vitro inhibition assay to assess the ability of antibodies to block the production of sexual forms by the parasite was developed.ResultsImmunized cattle generated antibodies against r6cys A and r6cys B that were unable to block sexual reproduction of the parasite in ticks. Additionally, these antibodies also failed in recognizing native 6cys A and 6cys B and peptides representing 6cys A and 6cys B functional domains and in inhibiting the development of sexual forms in an in vitro induction system. In contrast, rabbit antibodies generated against synthetic peptides representing predicted B-cell epitopes of 6cys A and 6cys B recognized recombinant and native forms of both 6cys proteins as well as peptides representing 6cys A and 6cys B functional domains and were able to neutralize development of sexual forms of the parasite in vitro.ConclusionsThese data, combined with similar work performed on Plasmodium 6cys proteins, indicate that an effective 6cys protein-based TBV against B. bovis will require identifying and targeting selected regions of proteins containing epitopes able to reduce transmission.Graphical abstract
Highlights
Babesia bovis reproduces sexually in the gut of its tick vector Rhipicephalus microplus, which involves expression of 6cys A and 6cys B proteins
Antigenicity analysis of the B. bovis 6cys A and 6cys B recombinant proteins Full-length r6cys A and residues 1–594] 6cys B (r6cys B) proteins containing a His tag were expressed in human embryonic kidney (HEK) cells and purified using nickel columns
SDS-PAGE and western blot analysis using anti-His antibodies demonstrated bands at the expected size for r6cys A (66.55 kDa) and r6cys B (67.65 KDa) [12] (Fig. 1 and Additional file 2: Fig. S1). r6cys A and r6cys B proteins were recognized by rabbit antibodies generated against a cocktail of synthetic peptides A and B derived from 6cys A and 6cys B [12] respectively by western blot and ELISA (Fig. 1)
Summary
Babesia bovis reproduces sexually in the gut of its tick vector Rhipicephalus microplus, which involves expression of 6cys A and 6cys B proteins. The currently available live-attenuated B. bovis vaccines have proved to be effective in several countries that permit their use, critical drawbacks remain These limitations include the risk of reversion to virulence of the parasites in the vaccine, contamination with other pathogens during passage through live splenectomized calves for attenuation and the need for a cold chain for transportation and delivery [8, 9]. Effective control of bovine babesiosis requires the development of new alternative vaccines, including transmission blocking vaccines (TBVs) This can limit expansion of the parasite by ticks in Babesia-endemic areas, while avoiding the current drawbacks of current live vaccines
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