The Development Of Nested-PCR Assay For Detection Of Eperythrozoon Suis And The Study Of Model Infection

Eperthrozoonsis is a parasitic disease caused by eperthrozoon parasitizing on the surface of the erythrocytes, plasma, tissue fluids and cerebrospinal flui- ds, which can cause anemia, icterus and fever.In recent years, the reports asso- ciated with this disease are increasing gradually. In order to determine the inf- ection status and morbidity of eperythrozoon suis , the epidemic, clinical signs and pathological observations for the disease were investigated; we establish the Nested-PCR assay for detection of Eperythrozoon suis, and a rat model of infection with eperythrozoon suis has been established inorder to approve the important of infection process.There are three experiments in this study:one is the development of Nested-PCR assay for detection of Eperythrozoon suis , another is that a mice model of infection infection with eperythrozoon suis has been established, the other is the study about cross-infection with Eperythrozoon suis in different animals.Experiment 1 The development of Nested-PCR assay for detection of Eperythrozoon suisPig blood with the clinical symptom and pathological feature of Epreythr- ozoonosis was got in aseptic operation, the template are extracted after Alsever's solution used as anti-coagulant.Two paris of E.suis specific primers were designed according to the 16S rRNA gene sequence of Eperythrozoon suis from GenBank, A rappid and exact Nested-PCR method for the detection of E.suis was established, products were assessed by electrophoresis in 0.85% agarose gel. A 498 bp specific fragment was amplified with Nested-PCR. It was verified by sequencing. The results showed that the Nested-PCR method was an effective tool for diagnosing Eperythrozoon.with high sensitivity and specificity The nested-PCR can be used in detection and epidemic investigate- on of E.suis.Experiment 2 A mice model of infection with eperythrozoon suis has been establishedMices were infected intraperitoneally, subcutaneously or intramuscularly with positive blood from infected swine. Blood samples were colleted and normal blood smear staining was done. The E.suis was purified and its DNA was extracted. Nested-PCR was done and the PCR products were sequenced.It was show that the specific fragment was amplified with Nested-PCR was we wanted. During the experiment no obvious clinical symptom was found in inf- ected mice.A mice model of infection with eperythrozoon has been establishedExperiment 3 the study about cross-infection with Eperythrozoon suis in different animals.Blood samples were colleted and normal blood smear staining was done. The E.suis was purified .We infected rat ,rabbit and chicken with purified Eperythrozoon suis and detected with Nested-PCR and other normal blood smear staining .The result showed that Eperythrozoon suis could infect that two kinds of animals.