| 利用环介导等温扩增技术快速检测水产品中的副溶血弧菌 |
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| 引用本文： | 徐芊,孙晓红,赵勇,潘迎捷.利用环介导等温扩增技术快速检测水产品中的副溶血弧菌[J].农业生物技术学报,2008,16(5). |
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| 作者姓名： | 徐芊 孙晓红 赵勇 潘迎捷 |
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| 作者单位： | 上海水产大学食品学院,上海,200090 |
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| 基金项目： | 上海市科技兴农科技攻关项目，上海市重点学科建设项目，上海市高校优秀青年教师后备人选科研项目 |
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| 摘 要： | 副溶血弧菌(Vibrio parahaemolyticus)是一种重要的食源性致病菌。首次将环介导等温扩增技术（Loop-Mediated Isothermal Amplification, LAMP）应用于副溶血弧菌的快速检测。针对副溶血弧菌不耐热溶血毒素基因（tlh）设计四条特异性引物，建立了副溶血弧菌LAMP检测方法，1 h即可完成。对12种细菌共28株菌进行LAMP扩增，仅副溶血弧菌得到阳性结果，证明引物具有很高的特异性；副溶血弧菌基因组DNA和纯培养物的检测灵敏度分别为90 fg/LAMP反应体系和24 cfu/mL，对模拟食品样品进行直接检测，检测限为89 cfu/g；对40份水产样品进行检测，8份副溶血弧菌LAMP阳性，其中6份传统培养阳性。本研究表明，该方法检测副溶血弧菌特异性强、灵敏度高，并且操作简便、快捷、检测成本低，有望发展成为快速检测副溶血弧菌的有效手段。
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| 关 键 词： | 副溶血弧菌(Vibrio parahaemolyticus) 环介导等温扩增技术（LAMP） 不耐热溶血毒素基因（tlh） 快速检测 水产品 |
| 收稿时间： | 2007-12-25 |
| 修稿时间： | 2008-2-20 |
Rapid Detection of Vibrio parahaemolyticus in Seafood Using Loop-mediated Isothermal Amplification (LAMP) Method |
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| XU Qian,SUN Xiao-hong,ZHAO Yong,PAN Ying-jie.Rapid Detection of Vibrio parahaemolyticus in Seafood Using Loop-mediated Isothermal Amplification (LAMP) Method[J].Journal of Agricultural Biotechnology,2008,16(5). |
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| Authors: | XU Qian SUN Xiao-hong ZHAO Yong PAN Ying-jie |
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| Abstract: | Vibrio parahaemolyticus has been considered as one of the most important foodborne bacterial pathogens. In this paper, the loop-mediated isothermal amplification (LAMP) that amplifies DNA with high specificity and rapidity under an isothermal condition was applied for rapid detection of this pathogen for the first time. A set of four primers, two outer and two inner primers, was designed specifically to recognize the thermolabile hemolysin gene (tlh) of V. parahaemolyticus in this study. Genomic DNAs from 28 bacterial strains including 14 V. parahaemolyticus strains were amplified using LAMP, and no amplicon was observed in other bacterial strains. The detection limit of this LAMP assay was around 90 fg/LAMP Mixture of V.parahaemolyticus genomic DNA, 24 colony forming units for pure cultures, 89 cfu/g for non-cultured artificially contaminated food samples. In addition, 40 seafood samples were tested, and 8 samples were found V. parahaemolyticus positive by LAMP. Among the tested positive samples, 6 samples were detected positive by traditional culture methods. These results suggested that detection of V. parahaemolyticus by LAMP is an effective and low-cost procedure with high specificity and sensitivity that requires no specialized equipment. This assay is expected to become a valuable tool for rapid detection and identification of V. parahaemolyticus. |
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