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目的探究铜绿假单胞菌在生物膜状态与浮游状态时对常见抗菌药物的耐药性差异,为临床合理用药提供依据。方法在体外培养基中对铜绿假单胞菌进行孵育处理,并分为浮游液对照培养组(A组)、放置生物膜培养后的培养液组(B组)和生物膜培养硅胶片粘附组(C组)。取浮游培养(培养液)、生物膜培养(培养液)以及生物膜培养(硅胶片粘附)铜绿假单胞菌进行银染色、胞外糖染色并且扫描电镜鉴定。向3组培养基中加入不同抗菌药物,利用K-B方法观察细菌耐药性情况。结果 B组与A组药物耐药率和最低抑菌浓度(MIC)值差异均无统计学意义(P>0.05);C组菌株与A组比较,对美洛培南(50.00%vs 26.47%)、头孢吡肟(53.52%vs 25.00%)、头孢他啶(60.94%vs 20.59%)、头孢曲松(70.31%vs 61.76%)、左氧氟沙星(39.84%vs 2.94%)及哌拉西林(48.05%vs 8.82%)的耐药性差异均有统计学意义(P<0.01)。C组最低生物膜清除浓度(MBEC)值是A组和B组MIC值的100倍左右。结论体外建立细菌生物膜简便可行,通过银染法联合电镜可观察生物膜形成情况;生物膜会改变细菌的理化特性,提高对抗菌药物的耐药性,增加临床治疗难度。
Objective To investigate the difference of drug resistance of Pseudomonas aeruginosa to common antibacterials in the biofilm state and the planktonic state and provide the basis for clinical rational drug use. Methods Pseudomonas aeruginosa was incubated in in vitro culture medium and divided into floating culture control group (group A), culture medium group (group B) and biofilm culture silica film adhesion Group (Group C). Planktonic culture (medium), biofilm culture (culture medium) and biofilm culture (silica film adhesion) Pseudomonas aeruginosa were performed silver staining, extracellular sugar staining and scanning electron microscopy. Different antibacterials were added into the three groups of culture media, and bacterial resistance was observed by K-B method. Results There was no significant difference in drug resistance rate and minimum inhibitory concentration (MIC) between group B and group A (P> 0.05). Compared with group A, the susceptibility to meropenem (50.00% vs 26.47% ), Cefepime (53.52% vs 25.00%), ceftazidime (60.94% vs 20.59%), ceftriaxone (70.31% vs 61.76%), levofloxacin (39.84% vs 2.94%) and piperacillin (48.05% vs 8.82% %) Of the drug resistance differences were statistically significant (P <0.01). The lowest MBEC value in group C was about 100 times of that in group A and group B. Conclusion The establishment of bacterial biofilm in vitro is simple and feasible. The biofilm formation can be observed by silver staining and electron microscopy. The biofilm can change the physical and chemical properties of bacteria, improve the antibiotic resistance and increase the difficulty of clinical treatment.