Wu Cong-Ming, Wu Huiming, Ning Yibao, Wang Jianguo, Du Xiangdang, Shen Jianzhong
Department of Pharmacology and Toxicology, College of Veterinary Medicine, China Agricultural University, Beijing 100094, China.
FEMS Microbiol Lett. 2005 Jun 15;247(2):199-205. doi: 10.1016/j.femsle.2005.05.012.
Antibiotic-resistant mutants of Mycoplasma gallisepticum were selected in vitro from the susceptible strains S6 and BG44T by serial passages in stepwise concentrations of erythromycin, tylosin, or tilmicosin. High resistance to erythromycin or tilmicosin developed readily, whereas resistance to tylosin developed only after greater numbers of passages. Three mutants selected by each selector antibiotic were cloned and detected, and all cloned mutants exhibited cross-resistance to the three selector antibiotics as well as to lincomycin. Portions of the genes encoding domain V of 23S rRNA of the cloned mutants were amplified by PCR, and their nucleotide sequences were compared to those of the susceptible parent strains. Five of the six mutants selected by erythromycin harbored an A2058G (Escherichia coli numbering) mutation in one of the two 23S rRNA. One of the six mutants selected by erythromycin harbored a G2057A mutation and an A2059G mutation in the other 23S rRNA. In tilmicosin-selected mutants, two mutations, A2058G and A2503U, occurred in one of the two 23S rRNA. No mutation was detected in the two 23S rRNA of tylosin-selected mutants with low-level resistance. Mutations at homologous locations in the 23S rRNA of other macrolide-resistant bacteria indicate that the phenotype of macrolide resistance occurring in M. gallisepticum is strongly associated with point mutations in domain V of 23S rRNA.
通过在逐步增加浓度的红霉素、泰乐菌素或替米考星中连续传代，从鸡毒支原体的敏感菌株S6和BG44T中体外筛选出抗生素抗性突变体。对红霉素或替米考星的高抗性很容易产生，而对泰乐菌素的抗性只有在传代次数更多后才会出现。对每种选择抗生素筛选出的三个突变体进行克隆和检测，所有克隆突变体均对三种选择抗生素以及林可霉素表现出交叉抗性。通过PCR扩增克隆突变体编码23S rRNA结构域V的基因部分，并将其核苷酸序列与敏感亲本菌株的序列进行比较。在红霉素筛选出的六个突变体中，有五个在两个23S rRNA之一中存在A2058G（大肠杆菌编号）突变。在红霉素筛选出的六个突变体中，有一个在另一个23S rRNA中存在G2057A突变和A2059G突变。在替米考星筛选出的突变体中，两个23S rRNA之一中发生了A2058G和A2503U两个突变。在低水平抗性的泰乐菌素筛选出的突变体的两个23S rRNA中未检测到突变。其他大环内酯类抗性细菌23S rRNA同源位置的突变表明，鸡毒支原体中出现的大环内酯类抗性表型与23S rRNA结构域V中的点突变密切相关。
