CNRS UMR 5163 and Laboratoire Adaptation et Pathogénie des Micro-organismes, Université Joseph Fourier Grenoble 1, BP 170, F-38042 Grenoble Cedex 9, France.
J Antimicrob Chemother. 2010 Nov;65(11):2359-67. doi: 10.1093/jac/dkq315. Epub 2010 Sep 13.
Francisella tularensis subsp. holarctica strains are classified as biovars I and II, which are susceptible and naturally resistant to the macrolide erythromycin, respectively. The present study was aimed at both selecting biovar I strains with increased levels of erythromycin resistance and characterizing the underlying genetic mechanisms.
Serial cultures in the presence of increasingly high erythromycin concentrations were performed to select independent high- and intermediate-level erythromycin-resistant mutants from each of three different biovar I strains. The mutants were characterized for cross-resistance to several antibiotics, presence of mutations in the genes encoding the 23S rRNA and the L4 and L22 ribosomal proteins, and overexpression of efflux pumps.
Mutants displayed cross-resistance to all macrolide compounds tested but not to other classes of antibiotics. We found mutations in domain V of the 23S rRNA gene (G2057A, A2058G, A2058T and C2611T) and in the gene encoding L22, leading to either the G91D substitution or the M82K83R84 deletion. Analysis of mutants with intermediate resistance levels obtained over the course of the selection process revealed both a positive correlation between the number of mutated ribosomal operons and the resistance level, and an additional resistance mechanism in the early steps of selection.
We showed that high-level resistance to macrolides can be easily obtained in vitro in F. tularensis subsp. holarctica biovar I strains, thereby suggesting that in vivo selection for resistance may explain reported failures of antibiotic treatment. Ketolides were the most effective macrolides tested, which may limit the risk of selection for resistance.
弗氏志贺菌亚种土拉弗朗西斯菌被分类为生物型 I 和 II，分别对大环内酯类红霉素敏感和天然耐药。本研究旨在选择生物型 I 菌株，提高其对红霉素的耐药水平，并对其潜在的遗传机制进行表征。
在逐渐增加的红霉素浓度存在下进行连续培养，从 3 种不同的生物型 I 菌株中分别选择独立的高水平和中水平红霉素耐药突变株。对突变株进行交叉耐药性、编码 23S rRNA 和 L4 和 L22 核糖体蛋白的基因中的突变以及外排泵过表达的特征分析。
突变株表现出对所有测试的大环内酯化合物的交叉耐药性，但对其他类别的抗生素没有耐药性。我们发现 23S rRNA 基因的 V 结构域（G2057A、A2058G、A2058T 和 C2611T）和编码 L22 的基因中的突变，导致 G91D 取代或 M82K83R84 缺失。对选择过程中获得的中水平耐药突变株的分析表明，突变核糖体操纵子的数量与耐药水平之间存在正相关，并且在选择的早期步骤中存在另外的耐药机制。
我们表明，弗氏志贺菌亚种土拉弗朗西斯菌生物型 I 菌株在体外很容易获得高水平的大环内酯类耐药性，这表明体内对耐药性的选择可能解释了抗生素治疗失败的报告。酮内酯类是测试中最有效的大环内酯类药物，这可能限制了耐药性选择的风险。
