表皮葡萄球菌耐氟喹诺酮机制的初步探讨
发布时间：2018-12-14 16:14
【摘要】： 目的:初步探讨耐氟喹诺酮类药物的表皮葡萄球菌与其主动外排蛋白的相关性,以揭示表皮葡萄球菌对氟喹诺酮类药物的耐药机制,为临床耐药性表皮葡萄球菌的防治提供实验依据。 方法:(1)收集临床分离表皮葡萄球菌菌株,用纸片扩散法(Kirby-Bauer)检测50株表皮葡萄球菌对氟喹诺酮药物的敏感性,筛选出耐药菌株和敏感菌株,以标准菌株ATCC12228作为对照;(2)PCR法扩增norA基因并测序。(3)提取表皮葡萄球菌的总RNA,应用实时荧光定量PCR检测表皮葡萄球菌多药转运蛋白norA的mRNA表达水平,分析norA在敏感菌株和耐药菌株中的表达情况。(4)应用荧光法检测氧氟沙星在表皮葡萄球菌内的累积量,以及加入叠氮化钠后对氧氟沙星累积量的影响。 结果:(1)表皮葡萄球菌耐药率较高,80%以上对一种或一种以上氟喹诺酮类药物产生耐药。(2)50株临床分离表皮葡萄球菌有48株检测到NorA基因,阳性率为98%;对norA基因启动子区测序分析发现,5株耐药菌株中有3株在386位点有碱基C的缺失。(3)临床分离耐药菌株norA基因mRNA表达水平明显高于敏感菌株,差异有显著性(P＜0.05)。(4)氧氟沙星在表皮葡萄球菌内的累积量耐药组低于敏感组,差异有统计学意义(P＜0.05);加入能量抑制剂叠氮化钠后氧氟沙星在菌体内的累积量有所增加,菌株SE40尤为明显。 结论:表皮葡萄球菌对氟喹诺酮类药物存在明显的交叉耐药,临床医生在应用该类药物时应予以注意,减少耐药菌株的产生;表皮葡萄球菌nora基因的转录水平的增加使norA基因介导的主动外排增强使氟喹诺酮类药物在表皮葡萄球菌菌体内蓄积量减少,可能是表皮葡萄球菌对氟喹诺酮类耐药的重要原因;norA基因启动子区的碱基C的缺失与norA基因转录水平的增高相关。
[Abstract]:Objective: to explore the relationship between Staphylococcus epidermidis resistant to fluoroquinolones and its active efflux protein in order to reveal the mechanism of resistance of Staphylococcus epidermidis to fluoroquinolones. To provide experimental basis for the prevention and treatment of Staphylococcus epidermidis. Methods: (1) Clinical isolates of Staphylococcus epidermidis were collected. The susceptibility of 50 strains of Staphylococcus epidermidis to fluoroquinolone was detected by disk diffusion method (Kirby-Bauer). The standard strain ATCC12228 was used as control. (2) norA gene was amplified by PCR and sequenced. (3) Total RNA, extracted from Staphylococcus epidermidis was used to detect the mRNA expression of multidrug transporter norA in Staphylococcus epidermidis by real-time fluorescence quantitative PCR. The expression of norA in sensitive and resistant strains was analyzed. (4) the accumulation of ofloxacin in Staphylococcus epidermidis and the effect of sodium azide on the accumulation of ofloxacin were detected by fluorescence method. Results: (1) the resistance rate of Staphylococcus epidermidis to one or more fluoroquinolones was higher than 80%. (2) NorA gene was detected in 48 strains of 50 clinical isolates of Staphylococcus epidermidis and the positive rate was 98%. The sequence analysis of promoter region of norA gene showed that 3 of the 5 drug-resistant strains had base C deletion at 386.3.The mRNA expression level of norA gene in clinical isolates was significantly higher than that in sensitive strains. The cumulative dose of ofloxacin in Staphylococcus epidermidis group was significantly lower than that in sensitive group (P < 0. 05). The accumulation of ofloxacin in the bacteria was increased with the addition of energy inhibitor sodium azide, especially the strain SE40. Conclusion: Staphylococcus epidermidis has obvious cross resistance to fluoroquinolones. The increase of transcription level of nora gene in Staphylococcus epidermidis makes the active efflux mediated by norA gene enhance the accumulation of fluoroquinolones in Staphylococcus epidermidis, which may be an important reason for the resistance of Staphylococcus epidermidis to fluoroquinolones. The deletion of base C in the promoter of norA gene was associated with the increase of transcription level of norA gene.
【学位授予单位】：中南大学
【学位级别】：硕士
【学位授予年份】：2009
【分类号】：R378
[Abstract]:Objective: to explore the relationship between Staphylococcus epidermidis resistant to fluoroquinolones and its active efflux protein in order to reveal the mechanism of resistance of Staphylococcus epidermidis to fluoroquinolones. To provide experimental basis for the prevention and treatment of Staphylococcus epidermidis. Methods: (1) Clinical isolates of Staphylococcus epidermidis were collected. The susceptibility of 50 strains of Staphylococcus epidermidis to fluoroquinolone was detected by disk diffusion method (Kirby-Bauer). The standard strain ATCC12228 was used as control. (2) norA gene was amplified by PCR and sequenced. (3) Total RNA, extracted from Staphylococcus epidermidis was used to detect the mRNA expression of multidrug transporter norA in Staphylococcus epidermidis by real-time fluorescence quantitative PCR. The expression of norA in sensitive and resistant strains was analyzed. (4) the accumulation of ofloxacin in Staphylococcus epidermidis and the effect of sodium azide on the accumulation of ofloxacin were detected by fluorescence method. Results: (1) the resistance rate of Staphylococcus epidermidis to one or more fluoroquinolones was higher than 80%. (2) NorA gene was detected in 48 strains of 50 clinical isolates of Staphylococcus epidermidis and the positive rate was 98%. The sequence analysis of promoter region of norA gene showed that 3 of the 5 drug-resistant strains had base C deletion at 386.3.The mRNA expression level of norA gene in clinical isolates was significantly higher than that in sensitive strains. The cumulative dose of ofloxacin in Staphylococcus epidermidis group was significantly lower than that in sensitive group (P < 0. 05). The accumulation of ofloxacin in the bacteria was increased with the addition of energy inhibitor sodium azide, especially the strain SE40. Conclusion: Staphylococcus epidermidis has obvious cross resistance to fluoroquinolones. The increase of transcription level of nora gene in Staphylococcus epidermidis makes the active efflux mediated by norA gene enhance the accumulation of fluoroquinolones in Staphylococcus epidermidis, which may be an important reason for the resistance of Staphylococcus epidermidis to fluoroquinolones. The deletion of base C in the promoter of norA gene was associated with the increase of transcription level of norA gene.
【学位授予单位】：中南大学
【学位级别】：硕士
【学位授予年份】：2009
【分类号】：R378
【相似文献】
相关期刊论文 前10条
1 王家学;朱涛;娄强;吴e，
本文编号：2378927
本文链接：https://www.wllwen.com/yixuelunwen/shiyanyixue/2378927.html
最近更新
教材专著
