体外获得性人型支原体氟喹诺酮耐药株GyrA基因研究

The molecular mechanism of resistance of Mycoplasma hominis induced by fluoroquinolones in vitro

目的探讨GyrA基因突变与体外诱导人型支原体(Mh)氟喹诺酮耐药的相关关系。方法将Mh标准株PG21在分别含有4种次抑菌浓度氟喹诺酮类药物的液体培养基中传代培养3代和l2代后,筛选诱导株并分别检测其对4种药物的最低抑菌浓度(MIC)值;对所有诱导株GyrA基因PCR扩增后进行DNA测序分析。结果经体外诱导后,共筛选出8株诱导株。其中4株短期诱导株呈现低度的耐药和交叉耐药,仅SPX诱导株G3发生GyrA基因Ser83→Leu变异,4株长期诱导株呈现高度的耐药和交叉耐药,除CFX诱导株C12外,均发生GyrA基因Ser83→Leu变异。SPX诱导株G12同时还出现GyrA基因Ser84→Trp变异。结论体外长时间、次抑菌浓度的氟喹诺酮类药物刺激将诱导Mh产生耐药和交叉耐药。GyrA基因83位上丝氨酸(Ser)→亮氨酸(Leu)的变异可能是Mh体外获得性氟喹诺酮耐药的重要分子机制。

Objective To explore the correlation between GyrA gene mutations and resistance of Mycoplasrna hominis induced by fluoroquinolones in vitro. Methods The mycoplasma hominis standard strain PG21 were cultured stepwise for 3 and 12 passages respectively with four different fluoroquinolones at subinhibitory concentrations and minimum inhibition concentrations （MICs） to four induced drugs were detected. GyrA genes were amplified by PCR from the induced strains and then sequenced. Results 8 fluoroquinolone-resistant strains were selected after induced by fluoroquinolones in vitro. Four strains exhibited low lever resistance or cross-resistance to the four fluoroquinolones after 3-passage induction and only a GyrA mutation （Ser83→Leu） was detected in the strain G3 induced by SPX.Another four strains exhibited high resistance or cross-resistance to these fluoroquinolones after 12-passage induction and GyrA mutations（Ser83→Leu） were detected in three strains except for the strain C12 induced by CFX.Another GyrA mutation（Ser84→Trp） was detected in strain G12 induced by GAT. Conclusion The resistance and cross-resistance of Mycoplasma hominis to fluoroquinolones can be induced after long time and subinhibitory concentration stimulation with the fluoroquinolones in vitro. The Ser→Leu substitution at position 83 in GyrA gene may be the important molecular mechanism resulting in resistance of Mycoplasma hominis induced by fluoroquinolones in vitro.