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目的研究含新的mucA突变基因黏液型铜绿假单胞菌(PAE17)在不同条件下形成的生物被膜,以了解新的mucA突变基因对铜绿假单胞菌生物被膜的影响。方法采用PIA平板法鉴定3种铜绿假单胞菌的黏液表型,色氨酸反应法测定其胞外多糖蛋白复合物的合成量;刚果红染色法和静态培养法分别观察其在固态及液态条件下形成的生物被膜;采用电转化法将绿色荧光蛋白表达质粒(pGFPuv)导入上述3种病原菌中,应用改良平板法建立体外生物被膜模型,激光共聚焦显微镜观察不同时间点生物被膜的形成。结果PAE17与PDO300呈现明显的黏液表型,其胞外多糖的产量(101.6μg/ml、118.3μg/ml)显著高于PAEO1(56.3μg/ml),但无论在固态、液态和改良平板法体外生物被膜模型中,PAE17形成的生物被膜均与PDO300迥异,而表现类似PAEO1。结论新的mucA突变基因可能存在藻酸盐以外的其他途径调节生物被膜的形成。
Objective To study the biofilm formation of mucA mutant mucosa-type Pseudomonas aeruginosa (PAE17) under different conditions to understand the influence of the new mucA gene on the biofilm of Pseudomonas aeruginosa. Methods The mucoid phenotypes of three Pseudomonas aeruginosa strains were identified by PIA plate assay and the synthesis of extracellular polysaccharide protein complex was determined by tryptophan reaction. Congo red staining and static culture were used to observe the mucoid phenotypes of the three Pseudomonas aeruginosa strains in solid and liquid (PGFPuv) was electroporated into the above three pathogenic bacteria. The biofilm formation model was established by modified plate method. The biofilm formation at different time points was observed by laser confocal microscopy. Results PAE17 and PDO300 showed obvious mucinous phenotype. The production of extracellular polysaccharides (101.6μg / ml, 118.3μg / ml) was significantly higher than that of PAEO1 (56.3μg / ml), but both in vitro and in vivo In the biofilm model, the biofilms formed by PAE17 are all distinct from PDO300 and behave like PAEO1. Conclusion The new mucA mutant gene may exist in other ways than alginate to regulate the formation of biofilm.