Evaluation of immunogenicity in bovines of Riphicephalus (Boophilus) microplus and Haemaphysalis longicornis recombinant calreticulins

Abstract

The ticks Riphicephalus (Boophilus) microplus and Haemaphysalis longicornis are blood-sucking ectoparasities of bovines, causing serious damages to the livestock production. The main method of control is based on the acaricides. However, the use of vaccines has been studied as a promising control method. The calreticulin (CRT) is a multifunctional protein present in almost all cells of animals. The secretion of CRT during feeding might be linked to the modulation of the parasite-host interaction. In the present study, recombinant CRTs of R. microplus (rBmCRT) cloned in pET-5b and H. longicornis (rHlCRT) cloned in pET-43a were expressed in Escherichia coli and purified by ion exchange chromatography and used for immunization of bovines. The fraction used for tests in vitro was purified by ion exchange and gel filtration chromatography. By ELISA, it was demonstrated that both CRTs are recognized by immunized bovines. The immunogenic and antigenic capacities of rBmCRT and rHlCRT were analyzed by two methods. In silico, despite the difference in amino acid sequences, antigenic index analysis of rHlCRT and rBmCRT with the Jameson-Wolf algorithm indicated that both proteins were very similar in the antigenicity index. Although six different regions between the tick CRTs have been determined. In vitro, this data were corroborated by competitive ELISA that suggests the presence of different epitopes between proteins. By Western blot, anti-native rBmCRT and rHlCRT bovine sera also recognized the native proteins in larvae extracts. These results demonstrate the presence of shared epitopes between recombinant and native proteins. In conclusion, the results suggest that the rBmCRT and rHlCRT could have a similar immunogenicity for bovines.