穿心莲内酯影响猪链球菌生物被膜形成

Effect of Andrographolide on the Biofilm Formation of Streptococcus suis

旨在探究穿心莲内酯对猪链球菌生物被膜形成的抑制作用,为猪链球菌病的耐药性提供替代疗法。试验采用微量稀释法测定穿心莲内酯对猪链球菌的最小抑菌浓度(MIC)和最小杀菌浓度(MBC);利用结晶紫染色(CV)和扫描电镜(SEM)检测穿心莲内酯对猪链球菌生物被膜形成的影响。结果表明:1)穿心莲内酯对猪链球菌的MIC和MBC分别为0.25和0.5mg·mL^(-1);2)扫描电镜观察可见阴性对照组细菌镶嵌于生物被膜中,而穿心莲内酯在亚抑菌浓度下(1/2MIC、1/4MIC、1/8MIC)能使细菌生物被膜中的细菌数量和生物被膜形成量均减少,并呈剂量依赖性;3)luxS基因和毒力基因的RT-PCR检测显示亚抑菌浓度(1/2MIC、1/4MIC)的穿心莲内酯明显降低gapdh、sly、fbps、ef和luxS基因的转录,显著增加cps2J、mrp和gdh基因的转录;4)哈维氏弧菌生物发光法测定细菌培养液显示穿心莲内酯在亚抑菌浓度下(1/2MIC、1/4MIC、1/8MIC)对猪链球菌AI-2信号分子的产生有抑制作用,呈剂量依赖性。穿心莲内酯亚抑菌浓度的干预可抑制猪链球菌生物被膜的形成,并且可致毒力基因表达水平差异。

The aim of this study was to evaluate the inhibitory effect of Andrographolide on the Streptococcussuis.The dilution method was conducted to determine the minimum inhibitory biofilm formation of S.suis and to provide an alternative therapy for the drug resistance of concentration（MIC）and minimum bactericidal concentration（MBC）of Andrographolide on S.suis;The effects of Andrographolide on the biofilm formation of S.suis were detected by crystal violet staining（CV）and scanning electron microscopy（SEM）.Result were as follows：1）The MIC and MBC of Andrographolide on S.suis were 0.25 and 0.5 mg·mL^（-1）,respectively.2）Scanning electron microscopy showed that the bacteria were embedded in the biofilm in the blank control group.Andrographolide（1/2 MIC,1/4 MIC,1/8 MIC）can make the number of bacteria in the bacterial biofilm and the morphology of the biofilm changed greatly.According to the dose of the trend changed,bacterial biofilm gradually reduced.The results showed that Andrographolide had inhibitory effect on the formation of S.suis biofilm in a dose-dependent manner.3）Through quantitatively detected the expression of luxSgene and virulence gene in S.suis by RT-PCR,we can see the sub-MICs（1/2 MIC,1/4 MIC）of Andrographolide can respectively reduce the expression of gapdh,sly,fbps,efand luxSgenes and significantly increase the expression of cps2 J,mrp,and gdhgenes.4）Detected the bacterial culture medium by Bioluminescence of Vibrio harveyi showed that Andrographolide（1/2 MIC,1/4 MIC,1/8 MIC）had inhibitory effect on the production of AI-2 signal molecule in S.suis,in a dose-dependent manner.Intervention of Andrographolide can inhibit the formation of biofilm in S.suis,and can cause the difference of virulence gene expression levels.