Molecular characterization of Theileria orientalis piroplasm protein encoded by an open reading frame (To ORF2) in a genomic fragment.
In the present study, a novel antigenic protein expressed in the piroplasm stage of Theileria orientalis was characterized. A 4,707 bp genomic fragment amplified by PCR contained two open reading frames (ORFs). The deduced amino acid sequence of the first ORF showed significantly high similarlity to the ubiquitin carboxy terminal hydrolases/proteases while the second ORF (To ORF2) showed homology to several surface antigens of plasmodia. To ORF2 was expressed to determine whether the protein product is expressed by the parasite. In western blot analysis, bovine antiserum from a T. orientalis-infected calf recognized the recombinant protein containing a C-terminal part of the ORF expressed by baculovirus system. Western blot analysis with the anti-To ORF2 mouse serum recognized a 48 kDa protein in T. orientalis piroplasm lysates. Indirect immunofluorescence antibody test by confocal scanning laser microscopic analysis showed that antisera against the recombinant protein recognized T. orientalis piroplasm in the infected erythrocyte. The results from this study indicate that To ORF2 protein is expressed at the piroplasm stage and is immunogenic. This novel antigenic To ORF2 protein could be exploited for vaccine development against bovine piroplasmosis.
Kim
Jung-Yeon
JY
National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Hokkaido, Japan.
Yokoyama
Naoaki
N
Kumar
Sanjay
S
Inoue
Noboru
N
Fujisaki
Kozo
K
Sugimoto
Chihiro
C
D000595:Amino Acid Sequence; D000818:Animals; D000953:Antigens, Protozoan / Q000235:genetics* / Q000378:metabolism; D016367:Baculoviridae / Q000378:metabolism; D001483:Base Sequence; D015153:Blotting, Western; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D019084:Fluorescent Antibody Technique, Indirect; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D016366:Open Reading Frames / Q000235:genetics; D015800:Protozoan Proteins / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017385:Sequence Homology; D016794:Theileria / Q000235:genetics* / Q000276:immunology*
D000953:Antigens, Protozoan; D017931:DNA Primers; D015800:Protozoan Proteins; D011994:Recombinant Proteins