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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: Immunoprotection of cattle vaccinated with rRmLTI against Rhipicephalus microplus

Authors
item Andreotti, Renato -
item Cunha, Rodrigo -
item Soares, Mariana -
item Guerrero, Felix
item Leivas Leite, Fabio -
item Perez De Leon, Adalberto

Submitted to: International Conference on Ticks and Tick-Borne Pathogens
Publication Type: Abstract Only
Publication Acceptance Date: May 28, 2011
Publication Date: N/A

Technical Abstract: Rhipicephalus microplus is arguably the most economically important cattle ectoparasite. There is interest in the development of second-generation anti-R. microplus vaccines with an improved product profile. Genes coding for trypsin inhibitors in R. microplus have diverse patterns of expression across life stages, and their products are involved in important physiological processes. Here, we report the ability of a recombinant larval trypsin inhibitor (rRmLTI) to elicit a protective immune response against R. microplus infesting cattle. Cattle ticks were obtained from a laboratory colony in Campo Grande, MS, Brazil. A partial DNA sequence coding for the TI gene identified in R. microplus larvae was optimized for Pichia pastoris codon usage and used to produce rRmLTI in P. pastoris. Mice were immunized to obtain polyclonal sera against larval extract and rRmLTI. Target proteins were detected by reactive mouse and bovine sera using SDS-PAGE and Western blotting. Each vaccine dose consisted of 2 ml containing 100 µg of rRmLTI adjuvated with Montanide ISA 61 VG. Yearling Holstein cattle assigned to two groups of six animals each were injected intramuscularly with one dose of rRmLTI or adjuvant alone at 0, 2, and 4 weeks. Cattle were challenged with 15,000 larvae 21 days after the last injection. Vaccine efficacy was determined as described previously. Vaccination with rRmLTI was 37% efficacious. A 28% reduction was observed in the number of engorged female ticks dropping from cattle in the experimental group relative to the control group. The antibody response against rRmLTI in vaccinates peaked 1 week after tick challenge. Cattle in the control group remained seronegative for anti-rRmLTI antibodies throughout the study. The estimated molecular weight of rRmLTI was 46 kDa. This observation corresponded with results where serum from mice and cattle inoculated with rRmLTI recognized it. Sera from mice inoculated with larval extract also recognized rRmLTI. Immunoprotection with rRmLTI was higher (37%) than that obtained with vaccination using a synthetic n-terminal fragment of BmTI-A (18%), which is a TI present in R. microplus larvae. Recognition of rRmLTI by serum of mice inoculated with this recombinant product reflects its immunogenicity. It is likely rRmLTI shares spatial epitopes with the native form of the TI as serum from mice immunized with larval extract recognized it. Seronegativity of the control animals during the study suggests that the antibody response in vaccinates was rRmLTI-specific. In conclusion, rRmLTI is a candidate antigen for use in a polyvalent vaccine to protect cattle against R. microplus. Embrapa Beef Cattle, CNPq, and Fundect are kindly acknowledged for financial support. F.D. guerrero and A.A. Perez de Leon are funded by USDA-ARS appropriated project 6205-32000-031-00D. The U.S. Department of Agriculture is an equal opportunity provider and employer.

Last Modified: 8/30/2014
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