Abstract: 【Objective】 The purpose of the test was to identify the primary pathogen causing recurrent pneumonia in intensive cattle farms in Shihezi region of Xinjiang,and in vitro drug sensitivity analysis of this pathogen was performed simultaneously.【Method】 Nasal swabs from 10 cattle with suspected symptoms and 1 lung tissue from dead cattle were collected and tested by PCR using primers specific for Mycoplasma bovis (M.bovis),and the positive samples were cultured and purified.Morphological observation,Dienes staining,biochemical test,16S rRNA sequencing and evolutionary analysis were carried out on the colony of the purified isolate.The growth curve of the isolate was determined by measuring the color change unit (CCU),and drug sensitivity test was carried out on the isolate.【Result】 The PCR results indicated that 7 M.bovis positive samples were detected in the 10 nasal swabs,and 1 of the lung of dead cattle collected was also positive.Needle like colonies grew on PPLO solid medium coated with lung tissue grinding fluid culture medium,after purification,the colony morphology of the isolates was typical fried egg like.A distinct dark blue central umbilicus was visible with Danies’ staining.Biochemical experiments showed that the isolate did not hydrolyze gelatin,arginine,esculin,nonfermentable lactose,glucose and mannitol,did not break down urea,and could reduce triphenyltetrazolium chloride.16S rRNA sequencing showed that the isolate shared 99.7% identity with the M.bovis international standard strain PG45 and 99.9% identity with the domestic M.bovis endemic strains XBY01,Ningxia-1,NM2012 and Tibet-10.The results of growth curve measurement showed that the isolate entered the logarithmic growth period after 6 h in the medium,reached the peak at 54 h and entered the stable period,and entered the decline period after 72 h.Susceptibility test revealed that the isolate was sensitive to nitrofurantoin,tetracycline and doxycycline,gentamycin and kanamycin,and moderately sensitive to norfloxacin and ofloxacin,whereas resistance developed to ciprofloxacin,lomefloxacin,azithromycin and erythromycin.【Conclusion】 In this study,a strain of M.bovis isolate was successfully identified from lung tissue,and 54 h was the optimal harvest time for this isolate,which was less different from most of the endemic strains of M.bovis in China,the overall genetic evolution was relatively stable,and there was certain drug resistance.The result of this study provided a reference for local prevention and control of M.bovis,and also laid a foundation for the research on the pathogenic mechanism of M.bovis and the development of vaccines.

Key words: Mycoplasma bovis; isolate and identification; drug susceptibility testing