| In the past decade, the epidemic of Bovine Mycoplasmosis is very serious in our country and the morbidity and mortality of infectious bovine mycoplasma pneumonia are very high especially in beef and dairy cow groups in some areas,that results in huge economic losses and hinders the healthy development of the cattle industry seriously. One Mycoplasma Bovis strain(NM2012) has been isolated from the Inner Mongolia region in 2012 by Inner Mongolia Agricultural University.In the early stage, the strain has been identified and its gene sequence has been detected.In order to fully understand and master the biological characteristics of NM2012 strain, cultural characteristics, immunogenicity and pathogenicity of different culture generation NM2012 were studied in our study. The results will lay the foundation for the further basic research and vaccine research to Bovine Mycoplasmosis.The Mycoplasma bovis strain NM2102 was inoculated in 6 kinds of special liquid medium, cultured in 37℃ for 5 days,and the viable organism was counted by colony forming unit detection method. The results showed that the liquid medium No.4 in our study was most suitable for the growth of NM2012 strain. The 6th,12th and 20th generation strains of Mycoplasma bovis strain NM2102 were inoculated into 100ml liquid medium, cultured in 37℃ for 5 days, the change of medium was observed daily, and the viable organism was counted by colony forming unit detection method respectively on the third day.The results showed that the situation of proliferation of the three generation strains in liquid culture medium was consistent basically and proliferation speed was similar, the concentration of the three generation strains seached 109/mL level after cultured for 3 days. The 6th,12th and 20th generation strains of Mycoplasma bovis strain NM2102 were cultured on solid medium in 37℃ for 3 days, and the colony Form were observed under a microscope. The results showed that the colony Form of the three generation of strains was similar basically.The 2 weeks to 2 months years old calves were infected with the 6th,12th and 20th strain of Mycoplasma Bovis NM2012 respectively through artificial trachea injection. The clinical symptoms of test calves were observed in 27 days, nasal secreta were detected by PCR for Mycoplasma Bovis, and Mycoplasma Bovis antibodies in the serum of the calves were detected by ELISA. half of calves were killed on the 14th day after challenge and all of the left calves were killed on the 27th days after challenge.The naked eye pathological changes and histological pathological changes of killed calves were observed. The results showed that the three generations of strain NM2012 could infect calves all and the infected calves showed obvious clinical symptoms and pathological changes of typical mycoplasma pneumonia.Mycoplasma Bovis were detected out in all nasal secreta of challenged calves and most of challenged calves discharged Mycoplasma Bovis intermittently. The antibodies of most challenged calves were detected out after the 14th day of challenge.The BALB/C mouse were infected with the 6th,12th and 20th strain of Mycoplasma Bovis NM2012 through nasal cavity. In the different days after infection,the serum were taken randomly for cytokines detection, and lung, spleen and thymus organ were taken for the organ index measurement. The results showed that IL-1β and IL-6 of some mouse in three infection groups were significantly higher than the blank control group,but TNF-a of infection groups and the blank control group had no significant difference.The lung index, spleen index and thymus index of the mice of infection groups generally higher than that of the blank control group. The above results and the result of pathogenicity test to calf showed that the 6th、12th and 20th strain of Mycoplasma Bovis NM2012 had immunogenicity. |
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