摘要
本文研究了苯乳酸在亚抑菌浓度条件下对创伤弧菌(Vibrio vulnificus)生物膜形成的抑制作用.首先构建创伤弧菌生物膜模型,然后利用结晶紫染色法和XTT还原法分析苯乳酸对创伤弧菌生物膜形成量和代谢活性的抑制作用,并以胞外多糖、胞外蛋白和胞外DNA为指标,评价苯乳酸对创伤弧菌胞外聚合物的影响,进一步借助光学和激光共聚焦显微镜探究苯乳酸对创伤弧菌生物膜结构的影响.结果表明:创伤弧菌生物膜的聚集期、成熟期和脱落期分别在 16～32、32～56 和 56～72 h.苯乳酸浓度为 0.5 MIC(minimum inhibitory concentration,MIC)时,对创伤弧菌成熟期生物膜抑制作用较好,对生物膜形成量、代谢活性、胞外多糖、胞外蛋白、胞外DNA的抑制率分别为51.04％、33.62％、61.97％、32.70％、52.25％,且破坏了生物膜的致密性和完整性,使细菌呈松散分布.苯乳酸可通过减少创伤弧菌生物膜胞外聚合物的形成,从而抑制其生物膜的形成.本研究可为控制水产品中创伤弧菌生物膜提供理论基础.
Abstract
In this study,the inhibitory effect of phenyllactic acid(PLA)on Vibrio vulnificus biofilm formation at sub-inhibitory concentration was studied.Firstly,the biofilm model of V.vulnificus was established.Then the inhibitory effect of PLA on the formation mass and metabolic activity of V.vulnificus biofilm was analyzed using crystal violet staining method and XTT reduction assay.Using extracellular polysaccharides,extracellular proteins,and extracellular DNA as indicators,the effect of PLA on the extracellular polymers of V.vulnificus was evaluated.The effect of PLA on the structure of V.vulnificus biofilm was investigated by optical microscopy and laser confocal microscopy.The results showed that the aggregation,maturation and shedding periods of V.vulnificus biofilm were at 16～32,32～56,and 56～72 h,respectively.PLA at 0.5 MIC inhibited the biofilm of V.vulnificus in the maturation stage better,and the inhibition rates of biofilm formation mass,metabolic activity,extracellular polysaccharides,extracellular proteins,and extracellular DNA were 51.04％ ,33.62％ ,61.97％ ,32.70％ ,and 52.25％ ,respectively.Additionally,PLA disrupted the density and integrity of V.vulnificus biofilms,resulting in a dispersed distribution of bacteria.Therefore,PLA inhibited the biofilm formation of V.vulnificus by reducing the formation of extracellular polymers.This study provides a theoretical basis for the control of V.vulnificus biofilm in aquatic products.
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