氨溴索对铜绿假单胞菌生物膜群体感应系统的影响

Effect of ambroxol on biofilm quorum-sensing system of Pseudomonas aeruginosa

目的探讨氨溴索对铜绿假单胞菌生物膜群体感应(QS)系统的影响。方法平板法培养成熟的铜绿假单胞菌PAO1野生型菌株和QS系统缺陷株△lasR△rhlR基因缺陷型、△lasI△rhlI基因缺陷型菌株生物膜,分别予生理盐水和氨溴索1.875 g·L^(-1)、3.75 g·L^(-1)作用8 h。通过荧光探针SYTO9/PI标记,激光共聚焦显微镜(CLSM)观察不同浓度氧溴索作用前后不同菌株生物膜的结构变化,利用生物膜图象结构分析软件(ISA)对氨溴索作用前后的生物膜结构参数进行定量分析。结果 PAO1野生型菌株可见大片状生物膜,细菌密集。2个基因缺陷型菌株的生物膜菌落较稀疏,氨溴索作用后不同菌株生物膜厚度、平均扩散距离和结构熵均显著减少,区域孔率增加,有剂量相关性(P<0.05),PAO1野生型菌株生物膜的变化幅度比QS系统缺陷株更大(P<0.05)。结论氨溴索破坏铜绿假单胞菌生物膜的结构,具有拮抗QS系统的特性,高浓度氨溴索(3.75 g·L^(-1))效果更显著。

AIM To investigate the effect of ambroxol on biofilm quorum-sensing system of Pseudomonas aeruginosa. METHODS In vitro biofilm models of Pseudomonas aeruginosa PAO1 wild-type, △lasR △rhlRand △lasI △rhlI isolate strain were established. The effects of normal saline and ambroxol 1.875 g·L^-1 and 3.75 g·L^-1 on biofilm were monitored by confocal laser scanning microscopy （CLSM） respectively, based on nucleic acid SYTO9/PI stained. Quantitative parameters describing biofilm spatial structure were acquired after the image information and calculated by Image Structure Analyzer （ISA） software. RESULTS The PAO1 wild-type strain produced a significantly thicker biofilm compared to biofilms produced by the △rhlR and △lasl △rhll isolates which appeared more diffuse, respectively. Ambroxol treatment reduced biofilm thickness, increased areal porosity, decreased the average diffusion distance and the textual entropy of wild-type and the two doublemutant strains （P 〈 0.05）. However, compared to the double mutant strains, the changes observed for the wildtype strain were more clearly defined （P 〈 0.05）. CONCLUSION Ambroxol destroys the structure of biofilm and exhibits significant antagonistic quorum-sensing property, and its high concentration （3.75 g·L^-1） has more significant effect.