摘要目的：筛选、构建铜绿假单胞菌生物被膜体外模型，观察米卡芬净单用、联用美罗培南，对铜绿假单胞菌浮游相、被膜相细菌的体外作用效果。方法使用微孔板构建被膜体外模型、结晶紫比色法，筛选出中南大学湘雅医院2012年7月临床分离的10株铜绿假单胞菌成膜阳性株，并对被膜形成过程进行半定量分析及形态学观察。使用微量肉汤稀释法测定米卡芬净、美罗培南对铜绿假单胞菌（浮游相、被膜相）的最低抑菌浓度，观察成膜后细菌耐药性的改变。使用微量肉汤稀释法结合结晶紫比色法，观察米卡芬净单用、联用对菌株被膜形成的抑制和破坏成熟被膜的效果。使用SPSS 18．0软件分析，采用t 检验比较药物处理组与阳性生长对照组吸光度值的差异，P ＜0．05为差异有统计学意义。结果筛选出10株成膜阳性的铜绿假单胞菌临床菌株；菌株形成被膜后，美罗培南MIC 上升4～128倍，未能测得米卡芬净的 MIC；米卡芬净对9株菌株（PA1～PA9）被膜形成具有抑制作用，米卡芬净的最低有效浓度分别为：156．25、625、10000、2500、1250、2500、1250、625、10000 mg／L，最低有效浓度组与其阳性生长对照组的吸光度分别为：0．342±0．020与0．491±0．027、0．512±0．018与0．627±0．043、0．862±0．021与1．155±0．027、0．731±0．028 与0．863±0．017、0．311±0．003与0．447±0．021、0．435±0．021 与0．597±0．011、0．520±0．012 与0．605±0．027、0．611±0．059与0．734±0．017、0．223±0．011 与0．343±0．037， P 值分别是：0．02、0．03、0．00、0．01、0．01、0．00、0．03、0．01、0．03，差异具有统计学意义；米卡芬净对7株菌株（PA1、PA2、PA4～PA8）成熟被膜显示出破坏作用，米卡芬净的最低有效浓度分别为：2500、2500、5000、2500、5000、2500、5000 mg／L，最低有效浓度组与其阳性生长对照组的吸光度分别为：1．459±0．014与1．534±0．020、1．279±0．020与1．431±0．007、1．365±0．024与1．467±0．065、1．322±0．028与1．530±0．090、0．920±0．004与1．047± 0．013、1．860±0．005与1．953±0．055、1．407±0．005与1．553±0．045， P 值分别是：0．01、0．01、0．02、0．01、0．00、0．03、0．02，差异有统计学意义；米卡芬净与美罗培南联用可较好的抑制被膜的形成。结论米卡芬净具备抑制铜绿假单胞菌被膜形成、破坏成熟被膜的功效；米卡芬净联合美罗培南作用于多耐药铜绿假单胞菌，对被膜形成抑制能力较好。

abstractsObjectives Selecting and constructing the biofilm -model of Pseudomonas aeruginosa in vitro.Observing the antibacterial activity of using Micafungin alone , or combined with Meropenem against Pseudomonas aeruginosa ( plankton-grown and biofilm-grown ) . Methods Ten clinical isolates of Pseudomonas aeruginosa were collected in July 2012, constructing the biofilm-model by microwell plate from Xiangya Hospital, Central South University.The ability of biofilm-formation of these strains was estimated by crystal violet colorimetric method, and optical microscope was used to observe the shape of the biofilm .MICs of Micafungin and Meropenem against plankton -grown and biofilm-grown Pseudomonas aeruginosa were tested by broth microdilution method, and the changes of MICs were compared.Using broth microdilution method, and connecting with the crystal violet colorimetric method , to observe the antibacterial effect of using Micafungin alone, or combined with antibiotics in the inhibition of the biofilm formation and destruction of&nbsp;mature biofilm of Pseudomonas aeruginosa.SPSS18.0 and t-test were used in comparing the differences between both treatment group and control group.P <0.05 showed the difference was statistically significant . Results Ten strains of Pseudomonas aeruginosa were successful in forming biofilms.Comparing with their planktonic counterparts, biofilms became more resistant to Meropenem , with the MIC raised 4-128 times. However, MIC of Micafungin could not be measured.Micafungin can inhibit the formation of biofilm in 9 experimental strains (PA1-PA9), where the minimum effective concentration of Micafungin were 156.25, 625, 10 000, 2 500, 1 250, 2 500, 1 250, 625 and 10 000 mg/L respectively.The absorbance values of the minimum effective concentration group and its positive growth control group were 0.342 ±0.020 vs 0.491 ±0.027, 0.512 ±0.018 vs 0.627 ±0.043, 0.862 ±0.021 vs 1.155 ±0.027, 0.731 ±0.028 vs 0.863 ± 0.017, 0.311 ±0.003 vs 0.447 ±0.021, 0.435 ±0.021 vs 0.597 ±0.011, 0.520 ±0.012 vs 0.605 ± 0.027, 0.611 ±0.059 vs 0.734 ±0.017, 0.223 ±0.011 vs 0.343 ±0.037 respectively, where the P values were 0.02, 0.03, 0.00, 0.01, 0.01, 0.00, 0.03, 0.01 and 0.03 respectively.The differences are statistically significant.Micafungin can damage the mature biofilm of 7 strains (PA1, PA2, PA4 -PA8), where the minimum effective concentration of Micafungin were 2 500, 2 500, 5 000, 2 500, 5 000, 2 500, 5 000 mg/L respectively.The absorbance values of the minimum effective concentration group and its positive growth control group were 1.459 ±0.014 vs 1.534 ±0.020, 1.279 ±0.020 vs 1.431 ±0.007, 1.365 ±0.024 vs 1.467 ±0.065, 1.322 ±0.028 vs 1.530 ±0.090, 0.920 ±0.004 vs 1.047 ±0.013, 1.860 ±0.005 vs 1.953 ±0.055, 1.407 ±0.005 vs 1.553 ±0.045 respectively, where the P values were 0.01, 0.01, 0.02, 0.01, 0.00, 0.03, 0.02.The difference is statistically significant.Micafungin combined with Meropenem applied in multiple drug resistant strains , which can inhibit the formation of biofilm better.Conclusions Micafungin can inhibit the formation Pseudomonas aeruginosa biofilm and damage the mature biofilms.Micafungin combined with Meropenem can act on multiple drug resistant strain , which may get a higher inhibition rate of the biofilm.