大环内酯类药物对肺炎链球菌的防耐药突变浓度的研究

Study of mutant prevention concentrations of macrolide-resistant Streptococcus pneumoniae

目的测定大环内酯类药物对肺炎链球菌的防耐药突变浓度(mutant prevention concentration,MPC),扩增突变选择窗(mutant selection window,MSW)内筛选的耐药突变体的耐药基因,以了解肺炎链球菌对大环内酯类药物耐药的发生机制。方法肉汤法富集1010CFU/ml ATCC49619,采用平板二倍稀释法测定罗红霉素和阿奇霉素对ATCC49619的最低抑菌浓度(minimal inhibitory concentration,MIC)、MIC99、初测MPC(Provisional MPC,MPCpr)以及MPC。PCR法扩增不同药物筛选出的耐药突变株的耐药基因ermB和mefA并测序。结果罗红霉素和阿奇霉素对ATCC49619的MPC分别为0.80μg/ml和0.51μg/ml,细菌耐药选择指数(MPC/MIC99)为5.0和3.9。筛选出的耐药突变株中扩增出ermB耐药基因。结论通过调整药物剂量,可以限制对大环内酯类耐药的肺炎链球菌突变体的富集。肺炎链球菌对大环内酯类药物耐药机制可能与携带ermB基因有关。

Objective To detect the mutant prevention concentration （MPC） of macrolide resistant Streptococcus pneumoniae, to amplify the resistance gene of Streptococcus pneumoniae mutants selected in the mutant selection window （MSW） , and to investigate the mechanism in the resistance of Streptococcus pneumoniae to macrolides. Methods The Streptococcus pneumoniae strain ATCC49619 was enriched in broth, and the bacterial concentrations were adjusted to 10^10 colony forming units per milliliter. The minimal inhibitory concentration （ MIC ）, MIC for 99 % of input cells （ MIC99 ）, provisional MPC （ MPCpr）, and MPC of roxithromycin and azithromyein for Streptococcus pneumoniae were determined by agar plate dilution method. The ermB and mefA of Streptococcus pneumoniae mutants selected in the MSW were obtained by PCR method and sequencing. Results The MPCs of roxithromycin and azithromyein for ATCC49619 were 0.80μg/ml and 0.51μg/ml, and the MPC/MIC99 were 5.0 and 3.9. The ermB gene was detected in Streptococcus pneumoniae mutants selected in the MSW. Conclusion Adjustment of the drug dose may limit the enrichment of macrolide-resistant Streptococcus pneumoniae mutant. The mechanism in the resistance of Streptococcus pneumonia to macrolides may be associated with ermB genes carried.