摘要
目的研究氧浓度变化对铜绿假单胞菌生物被膜产生量的影响,探讨密度感应系统在生物被膜调控中的作用以及Ⅲ型分泌系统表达和生物被膜形成的关系。方法将23株铜绿假单胞菌临床菌株在不同氧浓度环境下连续培养3 d,检测生物被膜和藻酸盐产生量,银染色法观察ATCC27853产生胞外多糖的时间窗,实时定量多聚酶链反应(PCR)测定密度感应系统中调控基因LasI和RhlI的表达,Western blot方法观察Ⅲ型分泌系统的代表毒素胞外酶S分泌情况。结果铜绿假单胞菌的生物被膜产生量(R=0.455,P=0.000)及藻酸盐产生量(R=0.367,P=0.000)均与氧浓度变化呈显著正相关。银染色法结果显示,高氧条件下胞外多糖合成时间缩短。实时定量PCR结果显示,LasI、RhlI的相对表达量与氧浓度(R=0.025,P=0.794;R=-0.044,P=0.653)、生物被膜产生量(R=0.001,P=0.990;R=0.011,P=0.909)和藻酸盐产生量(R=0.029,P=0.770;R=0.193,P=0.064)间无明显相关性。Western blot结果表明,铜绿假单胞菌在氧浓度为10%~30%条件下可分泌胞外酶S,在高氧条件下不分泌胞外酶S。结论高氧可促进铜绿假单胞菌生物被膜的产生。Las系统和Rhl系统可能因为菌群数量处于低水平而未在生物被膜形成早期阶段参与生物被膜的调控过程。生物被膜的产生增多可能抑制Ⅲ型分泌系统的表达,使该菌毒力发挥受到限制。
Objective To investigate the relationship among oxygen concentration,quorum sensing system,type Ⅲ secretion system,and biofilm production of Pseudomonas aeruginosa.Methods A total of 23 clinical strains of Pseudomonas aeruginosa were cultured at different levels of environmental oxygen for three days.Then biofilm mass and alginate were quantified.The expression levels of LasI and RhlI were detected by real time polymerase chain reaction(PCR).The secretion of exoenzyme S was examined by Western blot.Results Both the biofilm mass(R=0.455,P=0.000) and alginate(R=0.367,P=0.000) were positively correlated with oxygen concentration.Real time PCR showed that the expression levels of LasI and RhlI were not significantly correlated with oxygen concentration(R=0.025,P=0.794;R=-0.044,P=0.653),the production of biofilm(R=0.001,P=0.990;R=0.011,P=0.909),or alginate(R=0.029,P=0.770;R=0.193,P=0.064).Western blot showed that the optimal oxygen concentration range for exoenzyme S secretion of Pseudomonas aeruginosa ranged 10% to 30%.Conclusions Hyperoxia can promote the production of biofilm and alginate by Pseudomonas aeruginosa.Las/Rhl system may not participate in biofilm production at the early stage due to the low bacteria amount.The increased production of biofilm may inhibit the expression of Type Ⅲ Secretion system and thus inhibit bacterial virulence.
出处
《中国医学科学院学报》
CAS
CSCD
北大核心
2010年第3期310-314,共5页
Acta Academiae Medicinae Sinicae
关键词
铜绿假单胞菌
生物被膜
氧浓度
密度感应系统
Ⅲ型分泌系统
Pseudomonas aeruginosa
biofilm
oxygen concentration
quorum sensing system
type Ⅲ secretion system