Study On The PCR Assay, Cultivation And Drug Sensitivity Test Of Wild Boar Eperythrozoon Suis

Purpose:Mycoplasma suis (Eperythrozoonosis) is caused by M. Suis in rickettsiales, characterized by icteroanemia and fever. For intergrated therapeutic technique to wild boar,the study by wild boar Eperythrozoon isolation, culture, molecular biology and blood diagnostics, drug susceptibility testing and treatment effect analysis, showed that infected wild boar and domestic pigs Eperythrozoon have high homology, and screened out doxycycline drugs Eperythrozoon infected wild boar has a good therapeutic effect for the treatment of wild boar Eperythrozoon provide theoretical and technical support.Method:(1)Blood collected from wild boars with inspected Eperythrozoonosis was used to extract genomic DNA, PCR primers were designed according to the conserved region of M. Suis16S rRNA genome published by GenBank. The DNA fragment was amplified and cloned to pUC-18vector. The enzyme fingerprinting assay, specificity test and sequence analysis.(2) Basal mediums to culture wild boar eperythrozoon was prepared by PPLO, inactivated calf serum mixed at a specific ratio with mycoplasma, M-199medium, RPMI-1640,meanwhile the supernatant of culture media were refreshed regularly and put into incubator containing5%CO2to analysis the prons and cons of culture condition by observing the magnitude of infection, the size of granulosis, movement feature of M. Suis and to record the number of infected ones. The best condition of M. Suis culture was screened,according to different ratios of serous and continuous cell culture.(3) The antibiotic susceptibility test in vitro was carried out to determine the susceptibility of M.suis to berenil, oxytetracycline hydrochloride, sulfamonomethoxine sodium, erythromycin, doxycycline, acriflavine hydrochloride to get the best drug.Result:(1) The DNA fragment was960bp.It was proved that the targeted gene have high homogeneity with the gene of swine infected with M. Suis. As there were the same sequence in the wild boar and swine, the swine were the modle to research eperythrozoonosis. It also sets certain foundation to other gene testing of wild boar.(2) Compared with M-199、RPMI-1640, Mycoplasma culture medium is the best medium.(3)The results (shape, size and number of pathogen)of optimal concentration gradient tests were comprehensive assessed to conclude that doxycycline was the first-choice drug to cure eperythrozoonosis. The cure rates of doxycycline, oxytetracycline hydrochloride and sulfamonomethoxine sodium were87.5%,75%,25%, respectively.