牛支原体的分离鉴定及对大环内酯类抗生素耐药机制研究

发布时间：2018-05-08 09:38

  本文选题：牛支原体 + 大环内酯类类抗生素　； 参考：《吉林农业大学》2015年硕士论文

【摘要】：牛支原体(Mycoplasma bovis,M.bovis)是引起肉牛和奶牛多种疾病的重要病原体,其中以引发呼吸系统疾病(Bovine respiratory disease,BRD)最为常见。2008年,我国首次从患肺炎的犊牛肺脏中分离到M.bovis。由于M.bovis不具有细胞壁,因此β-内酰胺类抗生素对其治疗无效,且近年来抗生素的广泛及不合理使用,使M.bovis逐渐出现了耐药株,其产生的耐药性不仅给控制和治疗该病带来困难,而且加剧了抗生素在牛体内的残留,严重的危害动物食品安全。对吉林省售出至他省后患BRD的病牛进行病料采集,经过对分离菌进行纯化、形态观察及分子生物学鉴定共获得32株M.bovis。采用改良的微量二倍稀释法对临床治疗M.bovis有效的9种抗菌药物进行抗菌药物敏感性实验,结果显示,所分离的32株M.bovis有13株对红霉素、阿奇霉素、吉他霉素、泰乐菌素耐药(MIC≥64μg/m L),对克林霉素耐药(MIC≥4μg/m L),占40.63%,且这13株M.bovis同时对泰乐菌素、克林霉素和十四元环、十五元环、十六元环大环内酯类抗生素耐药,有1株对氟苯尼考耐药(MIC≥8μg/m L);部分菌株对红霉素、泰乐菌素、环丙沙星、恩诺沙星、氟苯尼考中度敏感;所有菌株均对多西环素敏感,多数菌株对恩诺沙星较敏感。对M.bovis上与大环内酯类抗生素耐药相关的靶位包括23S rRNA结构域Ⅱ,23S rRNA结构Ⅴ,L4、L22核糖体蛋白的基因序列进行比对分析,发现13株对大环内酯类药物高度耐药的M.bovis,其23S rRNA rrnA 2058位的碱基都发生了A—G的突变,23S rRNA结构域Ⅱ、核糖体蛋白L4、L22均未发现其有耐药相关的基因突变或氨基酸置换。对编码甲基化酶的ermA、ermB、ermC基因及大环内酯类钝化酶基因mphA、mphC基因和调控药物主动外排系统的mefA、mefE、msrA、msrB基因进行检测,所分离32株M.bovis上均未检测到上述基因。采用微量稀释法检测敏感株Mb9、Mb10及耐药株Mb1和Mb12加入CCCP、VP和SOV后MIC值的变化,对其体内存在的药物主动外排系统进行初步确证,发现应用EPI前后抗菌药物对M.bovis MIC值无明显变化,提示M.bovis体内可能不存在对大环内酯类药物的主动外排系统,但还有待于进一步确证。
[Abstract]:Mycoplasma bovis-M.bovis) is an important pathogen causing many diseases in beef cattle and dairy cattle, among which the respiratory system disease caused by bovine respiratory disease (BRD) is the most common. In 2008, M. bovisis was first isolated from the lungs of calf infected with pneumonia in China. Because M.bovis does not have cell wall, 尾 -lactam antibiotics are not effective in its treatment, and the widespread and unreasonable use of antibiotics in recent years has led to the emergence of drug-resistant strains of M.bovis. The drug resistance produced by M.bovis not only makes it difficult to control and treat the disease. Moreover, the residues of antibiotics in cattle are aggravated, which seriously endangers the safety of animal food. A total of 32 strains of M.bovisis were collected from infected cattle sold to his province in Jilin province. After purification, morphological observation and molecular biological identification, 32 strains of M.bovisis were obtained. A modified microdilution method was used to test the antimicrobial susceptibility of 9 effective antimicrobial agents for M.bovis. The results showed that 13 of the 32 strains of M.bovis isolated were resistant to erythromycin, azithromycin, and guitar mycin, and the results showed that 13 of the 32 strains of M.bovis were resistant to erythromycin, azithromycin and guitar mycin. The MIC of tylosin resistance was 鈮，

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