Analysis on protective antigen of Babesia parasites by developmental engineering

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1. Babesia microti produces a self-limiting infection in mice, and recovered mice are resistant to reinfection. In the present study, the protective mechanisms against B.microti were examined using macrophage scavenger receptor (SR), inducible nitric oxide synthase (iNOS), and gamma interferon (IFN-γ) knock-out mice. Decrease of packed cell volume and increase of spleen weight were less obvious in SR-/- mice than in SR+/+ mice. Parasitemia in iNOS-deficient mice increased more rapidly than those of control mice during the early stage of infection, although they recovered from the infection. IFN-γ knock-out mice could not control primary infection, nor protect against challenge infection.2. B.rodhaini antigen p26 was expressed in Escherichia coli and in insect cells infected with a recombinant baculovirus. BALB/c mice immunized with both recombinant antigens and Freund's adjuvants showed 40- 100% survival rate against challenge infection. MAb 1-5H recognized a 58-kDa protein of B.microti and was found to cross-react with a 60-kDa protein of B.rodhaini. The complete nucleotide sequence encoding p58 protein contained an open reading frame of 1,629-base pair (bp) nucleotides that consists of 542 amino acid residues and the gene was named as p58 gene. A protein homology search showed significant amino acid identities to the η subunit of the chaperonin.3. MAb BEG3 recognized 19kDa antigen of B.equi and inhibited the multiplication of B.equi in vitro. MAb BCllD recognized a 48-kDa protein rhoptry protein of B.caballi merozoite. A cDNA encoding a 50kDa protein of B.gibsoni was cloned and the complete nucleotide sequence was determined. Recombinant proteins of B.equi (MEA-1), B.caballi (BC48) and B.gibsoni(p50) were found to be useful as antigens for serodiagnostic methods such as ELISA or latex agglutination test.