铜绿假单胞菌Arr基因突变对生物膜和绿脓菌素合成的影响

Effects of Arr Mutation on Biofilm and Pyocyanin Biosynthesis in Pseudomonas aeruginosa

摘要:

为了研究铜绿假单胞菌Arr基因对生物膜和绿脓菌素合成的影响, 采用抗庆大霉素基因序列(Gentamycin resistance cassette, aacC1)插入失活的策略构建了铜绿假单胞菌Arr基因突变株PA-AG, 通过96孔板静止培养、结晶紫染色的方法检测其生物膜的形成量, 利用抽提的方法检测绿脓菌素的合成量。结果在KMB或LB培养基中, 突变株PA-AG形成生物膜的量均有所减少, 野生株约是突变株的2倍, 然而突变株合成绿脓菌素的能力却明显加强, 约为野生株的2.5倍。由此推测, 铜绿假单胞菌Arr基因在一定程度上促进了生物膜的形成, 抑制了绿脓菌素的合成。

Abstract:

As an opportunistic pathogen, Pseudomonas aeruginosa can produce biofilm and pyocyanin, which play a critical role in its pathogenesis. This study aims to elucidate the function of the Arr gene in formation of biofilm and production of pyocyanin in Pseudomonas aeruginosa. Using the chromosome DNA of Pseudomonas aeruginosa as template, we first cloned the Arr gene by PCR. With the insertion of gentamycin resistance cassette (aacC1), the mutant PA-AG has then been constructed by homologous recombination. The formation of biofilm is determined by staining with crystal violet, and production of pyocyanin is detected with spectrophotometric method. In KMB or LB medium, biofilm formation of the parental strain PAO1 is about 2 folds higher than that of the Arr mutant PA-AG. The biosynthesis of pyocyanin in the Arr mutant PA-AG is about 2.5 folds higher than that in the parental strain PAO1. These results indicate that the Arr gene negatively controls the pyocyanin biosynthesis, but Arr could exert some positive effect on the biofilm formation. It is suggested that regulation mediated by the Arr gene on biofilm and biosynthesis of pyocyanin in Pseudomonas aeruginosa is specific and different.