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目的探讨结核分枝杆菌耐药相关基因gyr A突变与氧氟沙星耐药水平之间的相关性。方法采用微孔板Alamar blue显色法检测经比例法确认的39株氧氟沙星耐药和32株氧氟沙星敏感的结核分枝杆菌临床分离株对氧氟沙星的最小抑菌浓度(minimun inhibitory concentration,MIC),基因DNA测序法检测氟喹诺酮耐药相关基因gyr A的突变情况,并用卡方检验检测gyr A基因突变与氧氟沙星耐药水平的相关性。结果 32株氧氟沙星敏感株MIC均<2.000μg/ml且gyr A未发现耐药相关位点突变。氧氟沙星耐药结核分枝杆菌氧氟沙星MIC≥4.000μg/ml的菌株gyr A 94位的突变率(18/24,75%)高于MIC≤2.000μg/ml(6/15,40%)的菌株(P=0.017);5/7的gyr A 91单突变菌株MIC均为2.000μg/ml;两株gyr A94和gyr A90或91位联合突变株氧氟沙星MIC明显高于其它菌株,分别为16.000μg/ml和64.000μg/ml。结论氟喹诺酮耐药菌株gyr A基因94位突变和双位点联合突变与氧氟沙星高水平耐药明显相关,gyr A 91突变与氧氟沙星低水平耐药有关。
Objective To investigate the relationship between the gyr A mutation of resistance-related gene in Mycobacterium tuberculosis and the drug resistance of ofloxacin. Methods The minimal inhibitory concentrations of ofloxacin against 39 ofloxacin-resistant and 32 ofloxacin-sensitive clinical isolates of Mycobacterium tuberculosis were determined by microplate Alamar blue assay. (MIC). The mutation of gyr A gene related to fluoroquinolone resistance was detected by DNA sequencing. The relationship between the mutation of gyr A gene and the level of ofloxacin resistance was tested by chi-square test. Results The MICs of ofloxacin 32 strains were all less than 2.000μg / ml and there was no mutation at gyr A site. The mutation rate (18/24, 75%) of the gyr A 94 locus of the ofloxacin-resistant Mycobacterium tuberculosis levofloxacin MIC≥4.000 μg / ml was higher than that of MIC≤2.000 μg / ml (6/15, 40%) (P = 0.017). MICs of gyr A 91 single mutant at 5/7 were both 2.000μg / ml. The MICs of ofloxacin of two strains gyr A94 and gyr A90 or 91 were significantly higher than those of gyr A 91 The other strains were 16.000 μg / ml and 64.000 μg / ml, respectively. Conclusion The mutation of gyr A 94 gene and double-site mutation of fluoroquinolone resistant strains were significantly associated with high level of ofloxacin. The gyr A 91 mutation was associated with low level of ofloxacin.