The Comparison Of Staphylococcus Aureus Biofilm Formation Ability And The Synergistic Effect Of Ambroxol With Vancomycin On Its Biofilm

Staphylococcus aureus(S.aureus) is the common condition of Gram stain-positive pathogens.They are also the common pathogens, which to result in medical materials-related infections,ventilator-associated pneumonia, bronchiectasis, pulmonary abscess and so on. In the course of acute infections, S.aureus resistant to commonly used antibiotics, often with its own characteristics of the properties; in the course of chronic infections, its resistance is concerned with bacterial biofilm (BF) formation. S.aureus BF infiltration barriers, low oxygen and low bacterial metabolism, the release of passivation enzymes and hydrolytic enzymes played a important role on the generation of the drug-resistant strains,which can not be ignored. Our group's pre-research has been typed 21 S.aureus, isolated from the First Affiliated Hospital of Guangxi Medical University bacterial Room,used resistant spectrum of sub-type and protein A gene (spa) sub-type. On this basis, this topic through the establishment of S.aureus BF mode in vitro.First of all, compared their biofilm formation ability;then,explore ambroxol if or not could in coordination with vancomycin destruct S.aureus BF.PARTⅠThe comparison of Staphylococcus aureus biofilm formation abilityOBJECTIVES To establish the S.aureus BF mode in vitro, then compare with the 21 clinical isolates of S.aureus differences in biofilm formation ability.METHODS (1) using Medical Materials (PVC) as a carrier, using TSB (tryptone soy broth containing glucose 2.5g/L) as medium, put it aside for the establishment of S.aureus BF model in vitro. Respectively, the 3 days and 7 days, using scanning electron microscope (SEM) observe the surface of vector formed BF.(2) After successful establishment of S.aureus BF model in vitro, then to explore the effect of the culture medium with different concentrations of glucose (2.5 g/L,5.0 g/L,7.5 g/L,10 g/L) on the adhesion and BF formation of S.aureus.(3) Firstly,use of Congo red agar method to distinguish which strains could form BF;then to be able to form BF's S.aureus by crystal violet staining BF semi-quantitative comparison of clinical isolates of S.aureus differences in adhesion and BF formation ability.RESULTS (1) Cultured 3 days, Sliver staining,SEM observe the carrier surface, could be seen early BF; cultured 7 days, could be seen the three-dimensional structure of mature BF.(2) Containing 5.0g/L glucose's TSB can significantly promote the S.aureus adhesion and its BF formation;but by adding higher concentrations (7.5g/L, 10g/L) of glucose,their efforts to promote adhesion and the formation of BF capacity does not increase.(3) 21 S.aureus of clinical isolates, for the Congo red agar method,there are 20 BF formation-positive strains, the positive rate is 95.2% (20/21).(4) MRSA adhesion and early BF-forming ability are stronger than MSSA; but with the culture time extended, until to form the mature BF, MSSA BF-forming ability are stronger than MRSA.(5)Among the homologous strains, their adhesion and BF-forming ability are differences:in adhesion and early BF stage,the number of 17546 (MRSA) strain adhesion and BF forming ability is the strongest, compared with 17642 (MRSA) strain, the difference is not statistically significant; 17422 strain's adhesion and BF forming ability is the weakest, compared with other strains, the difference is statistically significant; in the mature BF stage, the number of 17546 strain is still the strongest BF-forming ability strain, but compared with the number of 17642 strain, the difference is no statistically significant.17422 strain has the weakest biofilm-forming ability, but compared with the number of 17431,18541-2,18558,18565,18719,there are no statistically significant difference, P>0.05.CONCLUSIONS 1.using PVC as the carrier, TSB as the medium can be successfully establish S.aureus BF model in vitro;2. The detection rate of Our hospital's biofilm-positive S.aureus is high. Between MRSA and MSSA, and among the homologous strains, their BF formation ability are different.PARTⅡThe effect of Ambroxol and Vancomycin on destruction Staphylococcus aureus biofilm structure and synergistic bactericidalOBJECTIVES to explore the effect of Ambroxol and Vancomycin on destruction Staphylococcus aureus biofilm structure and synergistic bactericidal.METHODS (1) Detection the MIC,MBC of Vancomycin on S.aureus.(2) Divided into blank control group, ambroxol group, vancomycin group, ambroxol + vancomycin group. Cultured 3 days,7 days later, the above-mentioned drug and their combination are added the medium.24 hours later, SEM and silver staining observe the surface of the carrier's BF; drug for 24 hours, using MTT method to count viable cells on the surface of the carrier.RESULTS (1) The MIC,MBC of vancomycin on S.aureus is 1μg/mL, 2μg/mL.(2) Whether cultured 3 days or 7 days, after drug for 24 hours, SEM observation, the vancomycin and ambroxol + vancomycin group's BF, their structure and size were destroyed and reduced,compared with blank control group.Further more,the ambroxol + vancomycin group's BF were smaller and less then the vancomycin group. (3) Whether cultured 3 days or 7 days, MTT method to count viable cells, results showed that after drug for 24 hours, the viable cells of ambroxol + vancomycin group and vancomycin group were less than the blank control group and ambroxol group, and ambroxol + vancomycin group less than vancomycin group (P<0.001).CONCLUSIONS Vancomycin and Ambroxol could destroy had already formed early or mature S.aureus biofilm, and ambroxol could in coordinate with vancomycin play the destruction role on reduce the number of viable cells within the BF.