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United States Department of Agriculture

Agricultural Research Service

Research Project: MINING THE GENOME OF RHIPICEPHALUS MICROPLUS TO DEVELOP NOVEL CONTROL TECHNOLOGY AND VACCINES

Location: Tick and Biting Fly Research

Title: Reverse vaccinology approach to identify novel cattle tick protective antigens

Authors
item Lew-Tabor, Ala -
item Valle, Manuel -
item Moolhuijzen, Paula -
item Guerrero, Felix
item Andreotti, Renato -
item Bellgard, Matthew -

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: June 8, 2012
Publication Date: N/A

Technical Abstract: A reverse vaccinology genomic approach was developed to identify novel antigens for the protection of cattle from the tick: Rhipicephalus (Boophilus) microplus. This ectoparasite causes extensive economic losses to cattle production in tropical and sub-tropical regions of the world through tick burdens and the transmission of tick fever (babesiosis and anaplasmosis). Ticks rapidly develop resistance to acaricides; thus the development of a vaccine with long duration of immunity is an industry priority. Such a vaccine needs to be capable of producing an anamnestic ("remembered") response following natural tick infestations. This research commenced with the analysis of available expressed sequence tags (ESTs) as the R. microplus genome is large (7.1Gb) and consists of ~80% repeat sequences. Subsequent steps included bioinformatics screening of existing (USDA database V.2 ~13,643 ESTs) and new targeted R. microplus sequences (subtraction library analysis, and microarray analysis of ticks from resistant vs susceptible cattle~300 ESTs). Identified candidates were further scrutinized using expression analysis (localization to stages and organs) and informatics (membrane and secretion predictions; domain analysis and B cell epitope analysis). B cell epitopes (748 representing 240 sequences from 95 candidate protein families and singletons) were screened against pools of sera collected from tick susceptible and resistant cattle following tick challenge. Eighty peptides representing those recognized by resistant cattle were used to prepare antibodies for in vitro adult female tick feeding experiments. Simultaneously, 6 polypeptide proteins were constructed from peptides shown to be recognized by resistant cattle. In vitro feeding resulted in groupings of 'effectivity' calculated by measuring the capacity of each antibody to: kill adult ticks, lay eggs, and emerge as larvae in comparison to naive serum/blood fed tick controls. Successful cattle vaccinations with strong protection from tick challenge were observed using a mix of 6 peptides (selected from the strongest top 9 tick feeding effectivities), a polypeptide mixture (9 antigens with peptides recognized by tick resistant cattle) and a single recombinant (selected from the top 9 feeding effectivities). Current research is focusing on vaccine product development. This approach can be easily adapted for the identification of vaccine candidates for other parasites with large genomes.

Last Modified: 10/1/2014
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