The Isolation And Identification Of Mycoplasma Bovis And Its Serological Surveys In Some Area In Xinjiang

In order to inspect the occurrence of Mycoplasma bovis disease,235cattle sera were detected by usingdouble-antibody sandwich ELISA method. The results showed that37cattle antibodies were positive andthe positive rate was15.7%. It can be conclude that the result that Mycoplasma bovis diseases dose exist inthe investigation area, and the disease’s rate of infection was higher in part of this area.Through collecting nasal swabs that belong to the corresponding positive serum,3generations wereblinded passage with PPLO cultural medium. Then cultured purified colonies after the shape of "Fried egg"in colony growth could be found. By using Gram staining method, the stained colors are found to beunqualified while the results of Giemsa staining was light purple; In addition, by using Dienes identifyingstaining method, we can find dark blue center showed. Therefore, it was initially suspected that thebacterium is Mycoplasma bovis.Then bio-chemical reactions identification was taken for the inspecting the isolation of the suspectedMycoplasma bovis bacteria, the results showed that it was consisted with the bio-chemical characteristicsof Mycoplasma bovis such as sensitive to digitonin、glucose、arginine and does not hydrolyzed urea. Thebacterial DNA was extracted by phenol chloroform method, It is expected amplified310bp fragment while296bp by using the universal primers amplification of the16S rRNA amplified. the PCR amplifiedproducts was sent to sequencing company and sequenced. Homology of16S rRNA sequence of the bacteriaand Mycoplasma bovis were used on GenBank DNA Star software.The results showed that the bacteria andMycoplasma bovis strain’s （GenBank accession number：NR-036946）、（accession number：AY526883）and （accession number：AF332753）16S rRNA sequence have a high homology, Respectively98.6%,98.2%,97.5%. Phylogenetic analysis showed that the isolated strain was an independent strain thatcombined into a same branch with mycoplasma bovis （accession number：NR-036946）.