棘托竹荪提取物对副溶血性弧菌的抑菌作用机制  

Antimicrobial Mechanism of Active Substances from Dictyophora echinovolvata Against Vibrio parahaemolyticus

作　　者：孙晓红[1] 赵宏强[1] 蓝蔚青[1] 曹奕[1] 陈燕[1] 潘迎捷[1] Sun Xiaohong Zhao Hongqiang Lan Weiqing Cao Yi Chen Yan Pan Yingjie(College of Food Science and Technology Shanghai Ocean University, Shanghai Engineering Research Center of Aquatic Product Processing and Preservation, Shanghai 201306)

机构地区：[1]上海海洋大学食品学院上海水产品加工及贮藏工程技术研究中心,上海201306

出　　处：《中国食品学报》2017年第3期260-267,共8页Journal of Chinese Institute Of Food Science and Technology

基　　金：上海市教育委员会科研创新项目(11YZ159);上海市科技兴农重点攻关项目(沪农科攻字(2015)第4-12号);上海市科委工程中心能力提升项目(16DZ2280300)

摘　　要：目的:采用实时荧光定量PCR法结合细胞超微结构观察,分析棘托竹荪提取物对副溶血性弧菌的抑菌作用机制。方法:使用最小抑菌浓度的竹荪提取物分别对副溶血性弧菌菌液进行0.5,1.0与1.5 h处理,并分别提取对照组与处理组菌株的总RNA,随后通过反转录获得各自cDNA,选择相关基因合成引物作Real-time PCR分析。以本菌的16S rRNA作为内参基因,以nusA,tol C,fla,atpA,ef-Tu,fad L,dld为特征基因,分析棘托竹荪提取物对副溶血性弧菌相关基因表达量的影响,并结合扫描电镜与透射电镜的细胞超微结构观察,对提取物的抑菌效果与作用机制进行综合评价。结果:由荧光定量PCR法对不同处理时间的相关基因相对表达量分析得出:与对照组相比,处理组的tolC,atpA,dld表达量均下调,fad L表达量为上调趋势,表明副溶血性弧菌对竹荪提取物无明显抗性;nusA表达量的上调和ef-Tu表达量的下调说明棘托竹荪提取物能在转录翻译水平上抑制副溶血性弧菌的生命活动,fla表达量的下调表明细菌的黏附性在处理后有所降低。随着处理时间的延长,表达量持续上调的基因有nusA,持续下调的有atpA。结合细胞超微结构观察可知,副溶血性弧菌经棘托竹荪提取物处理后,细菌数量明显减少,菌体细胞膜开始破裂,内容物逐渐从细胞中释放出来。结论:棘托竹荪提取物对副溶血性弧菌细胞的转录过程和抗性产生显著影响,使菌体的细胞膜受损,细胞质溶出,从而导致菌体细胞死亡。Objective： The method of real-time fluorescent quantitative PCR and the observation of cell uhrastructure were used to evaluate the antimicrobial mechanism of Dictyophora echinovolvata extracts against Vibrio parahaemolyticus. Methods： the strain was treated with the MIC （minimum inhibitory concentration） of Dictyophora echinovolvata extracts under 0.5 h, 1.0 h and 1.5 h. Total RNA of the control and treated group strains were extracted, cDNA were obtained by reverse transcription respectively and the relevant gene primers were selected for Real-time PCR method. The strains of 16S rRNA as reference gene, nusA, tolC, fla, atpA, ef-Tu, fadL, dld as feature genes, the comprehensive evaluation of Dictyophora echinovolvata extracts against Vibrio parahaemolyticus were analyzed for its gene quantity and observed for cell microstructure by SEM （scanning electron microscope）, TEM （transmission electron microscope）. Results： When treated with Dictyophora echinovolvata extracts, the level of gene expression in tolC, atpA, did were down-regulated, and the level of fadL was up-regulated, which proved that Vibrio parahaemolyticus was no obvious resistance to the extracts, The results showed that the level of gene expression in nusA was down-regulated and ef-Tu was up-regulated, which proved that the extracts can inhibit the synthesis of RNA and protein in Vibrio parahaemolyticus, lower level of fla proved its lower adhesion. With the prolonging of treated time, the level of gene expression in nusA was rising and the level of atpA was falling continually. The uhrastrueture of cell observations showed that the number of Vibrio parahaemolyticus was declined after treated by Dictyophora echinovolvata extracts, the cell membrane of Vibrio parahaemolyticus began to break down and the contents released from cells gradually. Conclusion： Dictyophora echinovolvata extracts have a significant impact on Vibrio parahaemolyticus for the process of transcription and the resistant of bacteria, which can make the damage of

关 键 词：棘托竹荪提取物 副溶血性弧菌 抑菌机制 

分 类 号：TS202.3[轻工技术与工程—食品科学]