KIRISAWA Rikio Rakuno-gakuen Univ., School of Vet.Med., Associate Prof., 獣医学部, 助教授 (70153252)
TSUJI Masayoshi Rakuno-gakuen Univ., School of Vet.Med., Associate Prof., 獣医学部, 助教授 (10150088)
|Budget Amount *help
¥3,100,000 (Direct Cost : ¥3,100,000)
Fiscal Year 1998 : ¥1,000,000 (Direct Cost : ¥1,000,000)
Fiscal Year 1997 : ¥2,100,000 (Direct Cost : ¥2,100,000)
Diseases caused by hemoparasitic protozoa, such as Babesia and Theileria, still remain to be a major threat to livestock industry worldwide. In order to facilitate the experimental studies on those hemoparasites, we have previously developed an animal model using SCID mice whose circulating red blood cells were substituted with bovine ones (Bo-RBC-SCID mice). In the present study, we used this animal model system to obtain parasite clones. RBC samples containing high parasitemia (more than 40%) were prepared by infection of Bo-RBC-SCID mice with B.ovata or T.sergenti. Single RBCs were individually picked under microscopy with a microcapillary tube, and each of them was inoculated into a Bo-RBC-SCID mouse. Parasite clones were established by repeating this procedure twice. We were able to isolate multiple parasite clones, whose characteristics appeared to be different from each other. To further define the individual parasite clones, we searched genetic markers which show polymorphism among parasite clones From the clone 11E of Babesia ovata, several genes were cloned and sequenced, including SpS-7, small subunit ribosomal RNA, actine, beta-tubulin, roptory associated protein-1, spherical body protein, merozoite surface protein, and elongation factor-1. Many of those genes were found to show polymorphism when they were used as the probe for southern blot analysis.In some genes, significant sequence variations were also seen between parasite clones. We were able to differentiate the individual parasite clones using combinations of several marker genes.
The parasite clones and the polymorphic genetic markers obtained in the present study will serve as a useful experimental tool to study the diversified parasite population and possible genetic recombinational event occurring at the sexual stage in the vector ticks.