Previous research and current efforts at control of babesiosis fall short of meeting the needs of countries where the disease is endemic, such as Israel, as well as the needs of exporting countries and countries bordering on endemic areas, such as the U.S. Our long-term goal is to develop improved methods of immunization against bovine babesiosis based on an understanding of the molecular mechanisms of immune protection and parasite targets of a protective immune response. In our previous BARD project, we established the basis for focusing on rhoptry antigens as components of a subunit vaccine against bovine babesiosis, and for additional research to better characterize rhoptry associated protein-1 (RAP-1) as a target of protective immunity. In this continuation BARD project, our objectives were to [1] optimize the immune response against RAP-1, and [2] identify additional rhoptry candidate vaccine antigens. The entire locus encoding B. bovis RAP-1 was sequenced, and the rap-1 open reading frame compared among several strains. Unlike B. bigemina, in which multiple gene copies with variant domains encode RAP-1, the B. bovis RAP-1 locus contains only two identical genes which are conserved among strains. Through testing of multiple truncated constructs of rRAP-1, one or more immunodominant T cell epitopes were mapped to the amino terminal half of RAP-1. At least one linear and one conformational B cell epitope have been demonstrated in the same amino terminal construct, which in B. bigemina RAP-1 also contains an epitope recognized by neutralizing antibody. The amine terminal half of the molecule represents the most highly conserved part of the gene family and contains motifs conserved broadly among the apicomplexa. In contrast, the carboxy terminal half of B. bovis RAP-1 is less well conserved and contains multiple repeats encoding a linear B cell epitope potentially capable of inducing an ineffective, T cell independent, type 2 immune response. Therefore, we are testing an amino terminal fragment of RAP-1 (RAP-1N) in an immunization trial in cattle. Cattle have beer immunized with RAP-1N or control antigen, and IL-12 with Ribi adjuvant. Evaluation of the immune response is ongoing, and challenge with virulent B. bovis will occur in the near future. While no new rhoptry antigens were identified, our studies did identify and characterize a new spherical body antigen (SBP3), and several heat shock proteins (HSP's). The SBP3 and HSP21 antigens stimulate T cells from immune cattle and are considered new vaccine candidates worthy of further testing. Overall, we conclude that a single RAP-1 vaccine construct representing the conserved amino terminal region of the molecule should be sufficient for immunization against all strains of B. bovis. While results of the ongoing immunization trial will direct our next research steps, results at this time are consistent with our long term goal of designing a subunit vaccine which contains only the epitopes relevant to induction of protective immunity. Parallel studies are defining the mechanisms of protective immunity. Apicomplexan protozoa, including babesiosis and malaria, cause persistent diseases for which control is inadequate. The apical organelles are defining features of these complex protozoa, and have been conserved through the evolutionary process, Past and current BARD projects on babesiosis have established the validity and potential of exploiting these conserved organelles in developing improved control methods applicable to all apicomplexan diseases.
Mycoplasma pneumoniae Respiratory Tract Infection Prevailing among Infants at a Nursery School
