In silico and in vivo Immunological Characterization of Aquaporin 1 Peptide and Ferritin2 Recombinant Protein of Brown Dog Tick Rhipicephalus sanguineus (Acari: Ixodidae) for Vaccine Development

The brown dog tick, Rhipicephalus sanguineus (R. sanguineus), is a significant vector of pathogens causing canine monocytic ehrlichiosis and babesiosis. While several emerging tick proteins show promise as vaccine targets, few have been rigorously tested. Given the crucial roles of aquaporin 1 (RsAQP1) in blood feeding and ferritin 2 (RsFer2) in hemoglobin processing for tick physiology, these two proteins represent potential targets for disrupting essential biological functions. However, traditional in vivo vaccine trials are resource intensive. This study employed an immunoinformatics approach to predict and compare the immunological properties of a synthetic peptide derived from RsAQP1 and a recombinant RsFer2 protein as potential vaccine candidates against R. sanguineus. The evaluation considered their potential as individual vaccines and as a cocktail in Baladi (native Egyptian) dogs. In silico analysis predicted that both the RsAQP1 peptide and the RsFer2 protein contain several promising B-cell and T-cell epitopes with high antigenicity and immunogenicity scores. The cocktail vaccine was predicted to induce a humoral immune response for the entire duration of the experiment. Consistently, from the animal experiment, all vaccinated dogs developed significant antibody levels and were notably higher in the case of injection with rFer2 protein or in the case of mixed antigens (AQP1&rFer2) vaccination. Western blot analysis indicated the recognition of the native tick antigen by the vaccinated dog serum either in the case of individual or combined vaccination. In conclusion. the favorable in silico immunological profiles of the RsAQP1 peptide and rRsFer2 protein, both individual rRsFer2 protein and as a cocktail with RsAQP1 peptide, suggest their potential as effective subunit vaccine candidates against R. sanguineus. These findings warrant further in vivo tick investigation to validate their protective efficacy in dogs.