Library construction of cDNA fragments of R. microplus proteolytic enzymes (peptidases) and serpins on the surface of filamentous phages

Abstract

Rhipicephalus (Boophilus) microplus is a tick species present in Brazil and well known to parasite cattle. High levels of parasitism can seriously affect the animals, resulting in reduced milk and meat production and causing substantial economic losses to the livestock. In addition, this species can transmit different etiologic agents to cattle, including those that cause babesiosis and anaplasmosis, contributing to animal productivity decreasing and mortality levels increasing in herds. The control of ticks in animals is carried out mainly by using chemical acaricides from different families, such as pyrethroids, organophosphates and macrocyclic lactones. However, the indiscriminate use of these agents can be responsible to the emergence of resistant tick populations and the environmental contamination. Thus, it is necessary to search for alternative control methods that could be employed in herds and contribute to the reduction of parasitism levels by ticks. The main goal of this project is to identify antigenic targets that could be useful in control of R. microplus through the phage display technique. Firstly, from the tick's midgut cDNA, a library of sequences related to R. microplus peptidases and serpins will be developed in the vector pCANTAB5E. Then, using the phage display technique, a selection of the most reactive M13 phages against resistant bovines or previously tick immunized bovines antibodies will be performed. The selected phages will be sequenced and their peptides/proteins shall be produced in bacteria or yeast system. In parallel, a similar library will be developed, presented in T7 phages, and the technique will be performed according to the same parameters in collaboration with Dr. Albert Mulenga of University of Texas. The targets selected from both libraries and produced as recombinant proteins will be tested in cattle immunization tests in collaboration with Dr. Itabajara da Silva Vaz Jr of Federal University of Rio Grande do Sul. Therefore, it is expected that the results of this project can generate new information contributing in the development of R. microplus parasitism alternative control method. (AU)