Effects Of Andrographolide On QS System And Virulence Factors Of Streptococcus Suis Biofilm

In order to investigate the inhibitory effect of andrographolide on the formation of Streptococcus.suis biofilm and its related mechanisms,Streptococcus suis type II was used to establish in vitro biofilm models using coverslip fragmentation,crystal violet staining?CV?and scanning electron microscopy?SEM?.Detection of Streptococcus suis biofilm.Vitro cell culture plates Streptococcus suis,double dilution method for the determination of the minimum inhibitory concentration of andrographolide Streptococcus suis?MIC?and minimum bactericidal concentration?MBC?in 96-well plates.Gram negative and positive bacteria,a widespread system is called Quorum Sensing LuxS/type AI-2 quorum sensing systems that produce a signal inducing molecule can be used for communication between bacterial species,this types of signaling molecules in a variety of physiological functions have played an important role;detection of AI-2 signal by using of Vibrio harveyi bioluminescence;by qPCR,detection of quorum sensing?QS system?related genes lasR rhlR,regulatory genes and virulence genes pvdQ,gapdh,sly,fbps,ef,gdh,mrp,cps2J and luxS mRNA levels.Result:?1?The results of crystal violet staining showed that andrographolide was able to reduce the formation of bacterial biofilms in sub-microbial concentrations?1/2 MIC,1/4 MIC,1/8 MIC?in a dose-dependent manner.Electron microscopic observation showed that the bacteria in the negative control group were embedded in the biofilm,which was consistent with the crystal violet staining results;?2?The MIC and MBC of Andrographolide against Streptococcus.suis were 0.25 mg·mL^-1and 0.5 mg·mL^-1,respectively;?3?Vibrio harveyi bioluminescence assay of bacterial cultures showed andrographolide AI-2 signaling molecules at subinhibitory concentrations?1/2 MIC,1/4MIC,1/8 MIC?against S.suis The production has inhibitory effects in a dose-dependent manner;?4?qPCR detection of LuxS gene and virulence gene showed that sub-inhibitory concentrations?1/2MIC,1/4 MIC?of andrographolide significantly decreased transcription of gapdh,sly,fbps,ef,and luxS genes,significantly increased cps2J,mrp and gdh gene transcription;Andrographolide can increase the lasR gene transcription at 1/2 MIC concentration,1/4 MIC,1/8 MIC concentration when its transcription level decreases,andrographolide a concentration of 1/2 MIC,1/4 MIC,and1/8 MIC,the transcription levels of rhlR and pvdQ genes can be decreased.Conclusion:This study successfully constructed biofilm model in vitro of Streptococcus suis by using coverslip fragments;crystal violet staining and scanning electron microscopy demonstrated the intervention of andrographolide forming a biofilm on Streptococcus.suis;qPCR technology employed to verify the level of transcription from Effects of Andrographolide on Biofilm,QS System and Virulence Factors.The results showed that andrographolide at subinhibitory concentrations?1/2 MIC,1/4 MIC?significantly reduced the transcription levels of gapdh,sly,fbps,ef and luxS genes,and significantly increased the transcription of cps2J,mrp and gdh genes;Lactone increased lasR gene transcription at 1/2 MIC concentration,and its transcript level decreased at 1/4MIC and 1/8 MIC concentrations.Andrographolide can make rhlR at 1/2 MIC,1/4 MIC,and 1/8MIC concentrations.The level of gene transcription pvdQ decline.