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Objective To observe the destructive and scavenging effect of ambroxol (AMB) on the biofilm (BF) of Pseudomonas aeruginosa (P.a).To evaluate the synergistically bactericidal effect of AMB along with levofloxacin (LFX) on BF of P.a.Methods The early model (cultured for 3 d) and mature model (cultured for 7 d) of P.a wild strain (PAO1) BF were established,in vitro,respectively.The models were randomly (random number) divided into control group,AMB group and AMB + LFX group.The concentrations of AMB were 256 μg/ml and 512 μg/ml,respectively.When the early BF model and mature BF model were made,different concentrations of AMB were added in AMB group and AMB + LFX (1μg/ml) was added in AMB + LFX group.The number of viable P.a on the BF carrier was counted with the continuous dilution method 24 h after AMB or/and LFX added.Then,the BF morphological changes on the carrier surface were observed by using scanning electron microscopy (SEM).Measured data were analyzed with single factor analysis of variance (One-Way ANOVA).Results Both in the early BF model and in the mature BF model,the SEM examination showed that the BF in AMB group was significantly reduced compared to the control group,and this reduction of BF was in dose-dependent manner.LFX 1 μg/ml could reduce the number of viable bacterial in BF in both early model and mature model (P ＜ 0.05).LFX with addition of different concentrations of AMB showed stronger bactericidal effect than LFX used alone identified by more significant reduction in the number of colonv within the BF (P ＜ 0.05).Furthermore,the LFX combined with 512 μg/ml AMB reduced more significant number of colony apparently than the LFX combined with 256 μg/ml AMB (P ＜ 0.05).Conclusions AMB can destroy the early BF or mature BF partly,and LFX alone can partly reduce the number of viable P.a within BF.When LFX combined with AMB,they exert a synergistically bactericidal effect.
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Objective To evaluate the effect of azithromycin on Pseudomonas aeruginosa PAO1 biofilm formation and virulence factors production. Methods Detect the minimum inhibitory concentration of azithromycin against PAO1 by 2-fold dilution method. Crystal violet staining assay was used for initial adhesion assays. The PAO1 biofilm was established in vitro and observed by scanning electron microscope. Viable bacterial counts were determined by serial dilution. LasB elastolytic activity was determined by using Elastin-Congo Red. Protease activity was determined by Azo-casein. Chloroform extraction method was used for pyoverdine assay . The orcinol assay was used to directly assess the amount of rhamnolipids . Results Scanning electron microscope biofilm and viable bacterial counts of PAO1 adhered to the surface of catheter in PAO1 azithromycin group were less than the PAO1 control group after incubated for 3 d and 7 d ( P ＜0.05), and the initial adhesion was weaker ( P ＜ 0. 05 ). The virulence factors production were obviously decreased (P ＜0.01 ). LasB elastolytic activity and pyoverdine were even reduced to the same as with the PA-JP3 group ( P ＞ 0.05 ), but the protease activity and the rhamnolipids concentration were higher than the PA-JP3 group ( P ＜ 0.05 ). Conclusion Azithromycin can inhibit PAO1 bioflim formation in vitro and virulence factors production.