| Mycoplasma bovis,an important pathogenic pathogen of cattle,has been seriously endangering the healthy development of yak breeding industry for many years and has caused great economic losses to local herders and the development of related industries.In order to investigate whether chicken embryos and rabbits can be used as rejuvenation vectors for Mycoplasma bovis,to establish an animal model of bovine mycoplasmosis of yak origin in a highland environment,to obtain clinically effective drugs for the treatment of bovine mycoplasmosis,and to establish a set of integrated control measures for bovine mycoplasmosis.Firstly,the bovine mycoplasma was inoculated into chicken embryo,rabbit and yak calf respectively for the rejuvenation of the strains,and the regression test of the three rejuvenated strains was conducted in yak calves;the effect of modeling bovine mycoplasma disease was evaluated by clinical symptoms observation,excretion detection,serum antibody detection,organ index evaluation,pathological observation and pathogen detection,so as to clarify whether chicken embryo and rabbit can be used as rejuvenation vectors of bovine mycoplasma.Secondly,the successfully constructed bovine mycoplasma and animal models were treated with drugs,and the drug efficacy was evaluated by clinical symptoms and pathological changes,nasal excretion,and organ indices.Finally,the bovine mycoplasma disinfectants were screened by observing the killing efficacy of disinfectants and determining the disinfection effect of different disinfectants on the enclosures.In turn,integrated control measures for bovine mycoplasmosis were developed and further validated in the clinical setting.The details are subdivided as follows:1.The chicken embryo was inoculated with 1×10~9CCU/m L of T6resuscitated bacterial solution in the urinary bladder cavity at 9 days of age,and the chicken embryo was successfully isolated from the urinary bladder 5 d after inoculation.Yak calves were inoculated with 10 m L+5 m L of the above solution for 3 days,and nasal swabs were collected from yak calves 6～11 days after inoculation.Clinical signs such as elevated body temperature,cough and runny nose,and significant inhibition of body weight gain were observed in the calved yaks inoculated with the three different vector reinfection strains;their nasal discharge status was intermittent;antibodies to Mycoplasma bovis were detectable 7 d after the attack and persisted for more than 28 d.Autopsy revealed that the lungs were mainly characterized by diffuse hemorrhage,bruising and caseous necrotic lesions;pathological histological observations revealed alveoli with atrophy,collapse,alveolar wall thickening,parenchymalization of alveoli and alveolar walls,partial erythrocyte infiltration in the interstitium,marked vacuolization of interstitial cells,and partial inflammatory cell infiltration.The spleen index and lung index in the attack group were significantly different from those in the NC group(P<0.05).Mycoplasma bovis could be isolated from the lungs and trachea of the attacking group on the 28th day.Both the clinical symptoms and lesions caused by the Yak reovirus strain were more typical than those of the chicken embryo and rabbit reovirus strains,while the types of lesions were similar and not significantly different.Therefore,both chicken embryos and rabbits can be used as vectors for Mycoplasma bovis reassortment.2.Four drugs,enrofloxacin,ciprofloxacin,kanamycin and gentamicin,were selected to treat the successfully constructed animal models of bovine mycoplasmosis,and all were treated continuously for 14 d.The four treated groups no longer had cough and runny nose,shiny coat and increased appetite compared with the PC group,etc.Fluoroquinolones treatment could effectively increase the body weight gain and could lead to higher excretion rate of the animal models;the antibody absorbance increased after treatment in the four treated groups.Autopsy revealed that the lungs of the treated group showed only a small amount of bruising,and no pleural effusion,caseous necrosis and other symptoms were found;pathological histological observation revealed that the lung tissue lesions in the treated group were milder than those in the PC group,and only partial erythrocyte infiltration and inflammatory cell infiltration were seen in the interstitium of the treated group;there was slight atrophy and collapse of alveoli;the lung index was significantly different from that of the PC group(P<0.05);all four drugs could alleviate the bovine Mycoplasma damage to the lungs,but fluoroquinolones were more effective;there was a significant difference between the splenic index of fluoroquinolones and the PC group after treatment(P<0.05),and there was no significant difference between the indices of other organs(P>0.05).3.In this test,three commonly used livestock disinfectants(trichloroisocyanuric acid,glutaraldehyde,and compound phenol),which are inexpensive and have good disinfection effects,were selected for each disinfectant neutralizer identification test.The suspension quantitative disinfection test was conducted to ensure that the neutralizing agent had no effect on Mycoplasma bovis,and the disinfection effect of the three disinfectants was judged by color change units;and the disinfection effect was judged by screening the concentration of each disinfectant with good killing effect from each disinfectant to disinfect the enclosures of the animal model of Mycoplasma bovis disease.The results showed that 0.1%trichloroisocyanuric acid disinfectant solution was used for 30 min,0.15%trichloroisocyanuric acid disinfectant solution for 15 min and 30 min,0.5%glutaraldehyde disinfectant solution for 5 min,15 min and 30 min,1%glutaraldehyde disinfectant solution for 1 min,5 min,15 min and 30 min,0.05%compound phenol The disinfection solutions of 0.1%trichloroisocyanuric acid,0.5%glutaraldehyde and 0.05%compound phenol all had good effect in killing Mycoplasma bovis at 1 L/m~2for30 min.In conclusion,calved yaks showed typical clinical signs and pathological changes after inoculation with Mycoplasma bovis.It was confirmed that chicken embryo and rabbit could be used as a rejuvenation vector for Mycoplasma bovis strains,and the animal model of Mycoplasma bovis was successfully established.4 drugs were effective against Mycoplasma bovis,and kanamycin and gentamicin were more effective than ciprofloxacin and enrofloxacin.0.1%trichloroisocyanuric acid solution,0.5%glutaraldehyde solution,and 0.05%compound phenol solution were sprayed on diseased cattle at 1 L/m~2.The results of isolation and identification of Mycoplasma bovis in bedding sand were all negative after 30 min of action.According to the above conditions,a set of integrated control measures for bovine mycoplasmosis was established and clinically verified to be effective. |
