The Cloning Of The Major Surface Protein (P32) Of Theileria Sergenti And Establishment Of PCR On Theileria Sergenti

Theileria sergenti is a tick-borne protozoan parasite that multiplies in erythrocytes, causing mild hyperthermia and anemia. When infected calves are under stress, such as mixed infection with other parasites or viruses, they show severe symptoms and occasionally die. Theileriosis is one of the most economically important diseases of grazing cattle in jilin area.A pair of synthetic oligonucleotide primers, designed according to the gene published in genebank which encoding a 32-KDa intraerythrocytic piroplasm surface protein of T.sergenti ,were used to amplify parasite DNA from the blood of T.sergenti -infected cattle by means of the Polymerase chain reaction .The PCR fragment were ligated to the vector PMD-18T , sequenced and blast with the relative sequence of the Korea isolate and the Yanbian isolate. A predicted 875 bp DNA product were detected. The similarity of nucleotide and amino acid sequence between the Jilin isolate and the korea isolate were 99% and 98% respectively, While the similarity of nucleotide and amino acid sequence between the Jilin isolate and Yanbian isolate were 99% and 98% respectively. PCR were specific for T.sergenti, since no amplification was detected with DNA from Babesia bovis and leukocytes. This method was sensitive enough to detect about 4.5 organisms. Moreover,of 50 specimens from grazing cattle,18 were microscopically positive(36%), whereas PCR revealed 31 samples were positive(62%). Therefore, PCR provides a useful diagnostic tool for detecting T.sergenti infected cattle, and it is significantly more sensitive than the current methods. The PCR assay is a rapid method with high sensitivity and specificity, and it can be used as a diagnostic test for the detection of Theleria sergenti in clinical samples.