Babesia bigemina enolase binds to plasminogen and contains conserved B-cell epitopes that induce neutralizing antibodies in cattle

Abstract

Babesia bigemina is an intraerythrocytic parasite that causes bovine babesiosis and is responsible for economic damage to livestock in tropical and subtropical countries. Enolase is a glycolytic enzyme with multiple functions. In many pathogens, enolase contributes to the conversion of plasminogen to plasmin, which helps to degrade the fibrin of blood clots and various elements of the extracellular matrix, supporting the pathogen invasion process. In addition, enolase’s antigenic and protective properties in several organisms, including the Apicomplexa parasites Plasmodium ssp, have been reported. This protein is considered a vaccine candidate. To determine whether B. bigemina enolase is antigenic and has some of the characteristics already described, we isolated the gene, cloned and expressed the recombinant protein, and performed different assays to evaluate its ability to bind to and activate plasminogen and induce neutralizing antibodies. First, we studied the conservation of the B. bigemina enolase protein (BbiEno) among Apicomplexa parasites; then, we determined that it is transcribed and expressed in erythrocytic stages and tick stages via RT–PCR. In addition, we expressed recombinant BbiEno (rBbiEno) and studied whether sera from naturally infected cattle recognized it via Western blotting. Besides, the ability of rBbiEno to bind and activate plasminogen was analyzed. Finally, we examined whether specific anti-rBbiEno antibodies could neutralize the merozoite invasion of erythrocytes. The results showed that BbiEno is a conserved and immunogenic protein; it is transcribed and expressed in erythrocytic stages, and rBbiEno is recognized by the sera of naturally infected cattle. In addition, rBbiEno binds and activates plasminogen, and the specific anti-rBbiEno antiserum was able to neutralize the invasion of erythrocytes in vitro significantly. These results show that BbiEno could be considered a potential vaccine candidate.