Hemoparasitic diseases remain endemic in most tropical and semitropical areas of the world. Development of effective vaccines for malarial and newly emerging babesial parasites has been partly constrained by the lack of relevant outbred animal models. Immunoregulatory mechanisms in cattle are much more like those of humans than mice, so this outbred large animal species provides an alternative system to study the mechanisms of protective immunity against protozoan parasites. The pathology of-Babesia bovis infection in cattle is very similar to that caused by Plasmodium falciparum infections in humans, and is characterized by a generalized circulatory disturbance and sequestration of parasitized erythrocytes in the capillary beds, especially in the brain. Cattle recovered from natural or experimental infection with viable tick- or blood-borne stages of B. bovis develop long-lived protective immunity against subsequent exposure to both homologous and heterologous parasite strains, which correlates with the in vitro development of a Th1 response. Immunization with killed parasites or fractionated merozoite antigen has also resulted in variable degrees of protective immunity against homologous and heterologous challenge, which does not correlate with a specific humoral response. Furthermore, attempts to select vaccine antigens based on serological immunodominance have failed to identify protective immunogens. Because cell-mediated immune effector mechanisms are crucial for the induction of protective immunity against many intracellular parasites, and induction of Type 1 (T1 or Th1) helper cells correlates with immunity to B. bovis and related malarial parasites, we propose an alternative method for identifying protective protozoan parasite antigens based on the capacity to induce T1 responses in immune animals. We hypothesize that antigens selected for in vitro stimulation of T1 responses will stimulate protective immunity in vivo. Th1 cells will be used as probes to identify potentially protective antigens of B. bovis. Th cell lines and clones derived from immune cattle and characterized for cytokine expression patterns will be used in proliferation assays to identify biochemically fractionated parasite antigens. Antisera raised against these partially purified proteins will be used to identify the genes encoding the stimulatory T cell proteins by screening a B. bovis expression library. Alternatively, T cell lines and clones will be used directly to screen the library. Selected recombinant proteins will then be tested for the capacity to induce protective immunity in cattle and to characterize the nature of the T cell, macrophage and antibody responses both in vitro and in vivo against the immunogen. These studies will provide insight into the cellular and molecular basis of protective immunity against antigens that naturally stimulate a Th1 response in immune cattle, which will be directly applicable to related babesial and malarial parasites of humans.
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