Epitope mapping of tick salivary antigens for bovine B cells: a comparative analysis of in silico and in vitro methods

Author(s)

Luiz Gustavo Nogueira de Almeida

• Document type: Master's Dissertation
• Press: Ribeirão Preto.
• Institution: Universidade de São Paulo (USP). Faculdade de Medicina de Ribeirão Preto (PCARP/BC)
• Defense date: 2019-04-30

Examining board members

Isabel Kinney Ferreira de Miranda Santos

• Advisor: Isabel Kinney Ferreira de Miranda Santos

Abstract

Ticks are the most important vectors for the transmission of diseases to cattle, and for this reason, the efficient control of infestations is of great interest. Multi-antigenic vaccines based on recombinant salivary antigens have demonstrated good anti-tick activity, but their development is complex. This process can be facilitated by the use of in silico tools for the prediction of epitopes on antigens of interest. However, there is no data available on the usefulness of these methods, constructed with human and murine databases, for the identification of epitopes recognized by bovine antibodies. In this work, we investigated in tick antigens whether there is a correspondence between the in silico prediction and the in vitro mapping for B cell epitopes. For this, we submitted nine antigen protein sequences present in the saliva of the Rhipicephalus microplus tick to three-dimensional modeling and prediction of continuous and discontinuous epitopes by the Bebipred and Epitopia tools, respectively. At the same time, we immunized bovine with the same recombinant antigens to produce hyperimmune serum to the antigen pool, and verified the reactivity of this serum with mimotopes identified by phage display using IgY-specific antigen in biopanning. Bioinformatic predictions indicated the occurrence of 46-94% of antigenic amino acids in the analyzed sequences, while in vitro mapping identified about 20% to 42% of antigenic amino acids for the different antigens. We observed the occurrence of a large overlap of the antigenic amino acids identified in vitro with those predicted in silico, but most of the predicted amino acids were not identified in vitro. Thus, it is necessary to adjust the parameters used in the prediction, since its optimization can result in a more accurate analysis by the bioinformatics tools. However, with the current methodology, in silico tools are useful for the exclusion of sequences that are not characteristic of epitopes. This approach can be used for the identification and restriction of regions of the antigens with little predicted antigenicity, a strategy that can reduce the number of sequences to be evaluated as vaccine candidates and also facilitate the construction of multi-antigenic vaccines. (AU)

• FAPESP's process: 17/14297-6 - Epitope mapping of salivary proteins of Rhipicephalus microplus for the development of a multicomponent vaccine
• Grantee: Luiz Gustavo Nogueira de Almeida
• Support Opportunities: Scholarships in Brazil - Master