含新的mucA突变基因黏液型铜绿假单胞菌生物被膜特性的研究

Mucoid Pseudomonas aeruginosa Containing a Newly Discovered Mutation of mucA Gene:Its Biofilm Characterization

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 learn the effect of the newly discovered mution in mucA on the biofilm of the Pseudo- monas aeruginosa by observing the biofilm of the mucoid P. aeruginosa PAE17 which contains a newly discovered mutation of mucA gene under different growth conditions. METHODS Using PIA agar to observe the Alg~ phenotype, applying tryptophan reaction to quantitate the glycoealyx. Employing Congo red stain method and stand culture to observe biofilms in solid and liquid conditions respectively. Applying modified plate method to establish biofilm model in vitro, using confocal laser scanning microscope to further observe the morphology of the biofilm with different ages by electroporating the GFP-expressing plasmid （pGFPuv） into the above three strains. RESULTS PAE17 and PDO300 displayed the obvious mucoid phenotype and both had much higher production of glycocalyx （101. 6 μg/ml and 118. 3 μg/ml） than PAEO1 （56. 3μg/ml）, however PAE17 displayed totally different morphology of biofilm from PDO300 in different conditions, inversely which was similar to nomucoid strain PAEO1. CONCLUSIONS The newly discovered mutation of mucA might have other pathways other than alginate to regulate the formation of biofilm.