Study On Gene Regulation And Drug Effects Of Pseudomonas Aeruginosa Biofilm Formation

Objective:l.to understand biofilm formation ability of the 100 strains of Pseudomonas aeruginosa,use CLSI recommended drug resistance analysis to compare the strains of stronger biofilm formation ability and the strains of weaker,molecular epidemiology analysis of two groups was determined by RAPD(random amplified polymorphic DNA).2 During the formation of Pseudomonas aeruginosa(Pseudomonas aeruginosa PA)biofilm,relative expression of polysaccharide biosynthesis genes,lasI/lasR quorum-sensing system and rhamnolipid transferase biosynthesis genes rhlA,and their relationship will be studied.3 To investigate the regulatory mechanism of gentamicin,ciprofloxacin,ceftazidime,piperacillin/tazobactam and erythromycin in the process of biofilm formation of Pseudomonas aeruginosa(Pseudomonas aeruginosa,PA).Methods:1.100 isolates of Pseudomonas aeruginosa were collected from two hospitals in Tianjin from January to December in 2010,100 isolats of Pseudomonas aeruginosa were tested for their biofilm formation with a modified microtiter test to classify.2.Antimicrobial suscepibility test was done by K-B test,when we analyze thirty-threes trains.3.RAPD technique is a molecular technique,which could detect the entire unknown genome sequence to analyze the polymorphism of gene.4.collect non-mucoid Pseudomonas aeruginosa PAO1 planktonic bacteria and biofilm bacteria,use real-time fluorescence quantitative RT-PCR method for relative gene expression quantitative analysis.5.96 well plate crystal violet staining method was used to quantitatively analysis the inhibition and scavenging effect of drugs on Pseudomonas aeruginosa biofilm.real-time fluorescence quantitative RT-PCR method was used for relative quantification of gene expression analysis.Results:1.Two patterns were identified in the 100 isolates of Pseudomonas aeruginosa.2.There are thirty-seven kinds of type in thirty-eight strains by RADP analysis.3.thirty-eight bacterial resistance results could be seen in table.4.In addition to relative expression of pslA gene of one day planktonic bacteria was slightly higher than its six days of biofilm bacteria,algD,pelA,LasI,LasR,RhlA biofilm bacteria mRNA relative expression were all higher than planktonic bacteria,and pslA,pelA,RhlA of mRNA expression of Id biofilm bacteria was consistent,expressed the highest.5.The effect of the four drugs for the treatment of Pseudomonas aeruginosa is very ideal when the formation of biofilm doesn’t occur,however there is a limit effect of ceftazidime in the presence of a mature biofilm.6.ceftazidime can promote the expression of biofilm formation related gene.Conclusion:1.100 isolates of Pseudomonas aeruginosa generally have the ability to form biofilms.2.Molecular epidemiology demonstrated that the strains belong to the same type may not seem aggregate and prevail tendency.3.Stronger biofilm formation ability of strains is more sensitive to the drugs than the weaker biofilm forming ability of strains.4.algD,pslA,pelA,LasI,LasR,RhlA expression is closely related to P.aeruginosa biofilm formation and in biofilm for mation algD,pslA,pelA,LasI,LasR,RhlA plays important roles,while the quorum-sensing system may be positive transcription regulation,involved pslA,pelA,RhlA.5.In this article we confirmed that ceftazidime can promote the biofilm formation of Pseudomonas aeruginosa at gene level for the first time.And therefore it should be used carefully in the treatment of Pseudomonas aeruginosa infection afterwards;At the same time,in this experiment we found that the effect of ciprofloxacin,piperacillin/tazobactam,and gentamicin was better than that of ceftazidime in the treatment of Pseudomonas aeruginosa biofilms.