Establishment Of Real-time Fluorescence PCR Methods For Diagnosing Therlieria Equi And It’s Molecular Biology Epidemiological Studies From Ili

Equine piroplasmosis is a tick-borne protozoan parasite that causes piroplasmosis in equids. It hasbeen reported that pathogens of Equine piroplasmosis is including Theileria equi and Babesia caballi.Equine piroplasmosis will be columned animal disease list by the office international des epizooties,whichis a statutory reporting epidemics in our country is classified as Class II animal diseases. T.equi occur moreserious and widespread than B.caballi, and it has occurred in many countries and regions, which will befailed to timely diagnosis and treatment, can cause significant damage to the horse industry. Currently, toprevent and control of coccidiosis Theileria equine, research for the diagnostic methods have been carriedout in many countries, but these methods can be directly used for clinical diagnosis, especially lesstechnical methods of early diagnosis, and therefore it is very necessary to carry out research work in this.(Ⅰ)In this study, we established a real-time PCR method to detect T.equi, the assay utilized a pair ofspecific primers and a TaqMan probe which was based on the conserved region of the18S rRNA of T.equi.The results showed that the sensitivity of the detection limit was1×101copies/μL, which was1,000timeshigher sensitivity than conventional PCR, and the assay was specific for T.equi but not for B.caballi,B.bovis, B.bigemina and T.annulata. The coefficients of variations were less than3%for both intra-assayand inter-assay. Among50clinical samples, the positive rate was40%detected by real-time PCR and34%by conventional PCR. These results showed that the real-time PCR approach provides a diagnostic toolwith high sensitivity and specificity for the identification and quantitation of T.equi(Ⅱ) In this paper,906horses blood samples were collected from peripheral part of the Ili Valley todetect T.equi using blood smear staining and real-time PCR method developed.The results suggested thatthe positive rate of the latter was34.44%, which was significantly higher than20%by microscopicexamination of Giemsa-stained blood smears. The results demonstrated that the use of quantitative PCRdetection of positive (29.93%) was significantly higher than the blood smear staining (20%). The resultswere analyzed by statistical methods including different pastures, age, grazing environment.We found thatdifferent regions of T.equi infection rates is significantly different (P＜0.05), and the infection of differentseasons is also significantly different (P＜0.05), while ages were same. This is a first we used real-time PCR detection method to carry out the experimental study and molecular epidemiology researchparameters, can provide technical support for the prevention and control of Ili horse tick-borne bloodparasite.