The Influence Of Macrolides On LasR Gene And Relative Causative Agent Associated With Quorum Sensing System Of Pseudomonas Aeruginosa

Objective: To study the biology characteristics of PAO1 and PAO-JP2 and to investigate the influence of quorum sensing on pathogenicity as well as drug tolerance to Pseudomonas aeruginosa; Comparing the biofilm morphology, causative agent and transcriptional level of AHLs between PAO1 and PAO-JP2 which were cultured with diffenent dosages of roxithromycin(ROM), Azithromycin. (AZM) and acetylspiramycin(ACM) and research the effect of QS system to Pseudomonas aeruginosa with the adding of ROM, AZM, ACM. Methods: The modified plate culture method was used to establish biofilm model in vitro; The morphology of PAO1 and PAO-JP2 biofilms was observed by the optical microscope after being stained by AgNO3 and analyzed by computer image manipulation system; The minimal inhibitory concentrations (MICs) of 3 Macrolides to PAO1 and PAO-JP2 were determined by agar plate dilution method; The influence of Macrolides on elastase, which one of virulence factor of Pseudomonas aeruginosa cultured with PTSB nutritive medium, was measured by OD after enzyme-substrate reaction; The influence of Macrolides on exotoxin A, which one of virulence factor of Pseudomonas aeruginosa cultured with PTSB nutritive medium, was measured by Western-Blot and SDS-PAGE; The influence of Macrolides on the transcriptional level of lasR was measured by RT-PCR after RNA extracted with TRizol method.Results: PAO1 and PAO-JP2 biofilms, identified by argentation and analyzed by computer image processing system, were gradually cleared with the increasing dosages of Azithromycin and Roxithromycin, and the number of bacteria was decreased, however, the effection to PAO1 was more obvious than that to PAO-JP2 at the same dosage of Azithromycin and Roxithromycin. The group disposed by Acetylspiramycin doesn't show parallel results; PAO1 and PAO-JP2's OD decreased gradually with the increasing dosages of Azithromycin and Roxithromycin, however, the effection to PAO1 was more obvious than that toPAO-JP2 at the same dosage of Azithromycin and Roxithromycin. The group disposed by Acetylspiramycin doesn't show parallel results; The strip gray scale of exotoxin A decreased gradually with the increasing dosages of Azithromycin and Roxithromycin, which doesn't show in group disposed by Acetylspiramycin and group PAO-JP2; The strip gray scale of lasR, detected by RT-PCR, decreased gradually with the increasing dosages of Azithromycin, which doesn't show in group PAO-JP2.Conclusions: There were obvious differences between PAO1 and PAO-JP2 in biology characteristics in vitro. The effect of sub-MICs of AZM and ROM that PAO1 or PAO-JP2 exposured to when culturing the bacteria showed that QS system could actively inhibit the formation of Pseudomonas aeruginosa bacterial biofilm, virulence factors such as elastase and exotoxin A, transcription of lasR gene, while sub-MICs of ACM didn't show parallel results.