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Quantification of viable Brochothrix thermosphacta in cooked shrimp and salmon by real-time PCR

机译：实时PCR定量分析煮熟的虾和鲑鱼中的嗜热溴化拟南芥

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摘要

Brochothrix thermosphacta, a Gram-positive bacterium, is considered as the predominant spoilage microbiota of modified atmosphere packing (MAP) shrimp and fish. Traditional methods currently used to detect B. thermosphacta in foods are time-consuming and labour-intensive. The aim of this study was to develop a real-time PCR quantification method combined with a propidium monoazide (PMA) sample treatment step to monitor the population of B. thermosphacta in cooked shrimp and salmon. The specificity of the two primers MO405 and M0404 used to amplify a 70 bp fragment of the 16S rRNA gene was demonstrated by using purified DNA from 30 strains, among 21 bacterial species including 22 reference strains. Using these primers for real-time PCR and in pure culture, a good correlation was obtained between real-time PCR and the conventional plating method. Quantification was linear over 7-log units using artificially inoculated samples. The method performed successfully when tested on naturally contaminated cooked shrimp and fresh salmon, with a minimum threshold of 1.9 × 10~2 CFU/g for accurate quantification of B. thermosphacta. The correlation between the B. thermosphacta counts obtained by real-time PCR and plate counts on naturally contaminated shrimp and salmon was high (R~2 = 0.895). Thus, this study presents a rapid tool for producing reliable quantitative data on B. thermosphacta in cooked shrimp and fresh salmon.
机译：革兰氏阳性细菌嗜热气单胞菌（Brochothrix thermosphacta）被认为是气调包装（MAP）虾和鱼的主要腐败菌群。当前用于检测食物中嗜热芽孢杆菌的传统方法既费时又费力。这项研究的目的是开发一种实时PCR定量方法，结合单叠氮化丙锭（PMA）样品处理步骤，以监测熟虾和鲑鱼中嗜热芽孢杆菌的种群。通过使用来自21个细菌物种（包括22个参考菌株）中的30个菌株的纯化DNA，证明了用于扩增16S rRNA基因的70 bp片段的两个引物MO405和M0404的特异性。使用这些引物进行实时PCR和纯培养，在实时PCR和常规铺板方法之间获得了良好的相关性。使用人工接种的样品，定量以7对数单位为线性。该方法在对天然污染的煮熟虾和新鲜鲑鱼进行测试时成功进行，最低阈值为1.9×10〜2 CFU / g，可准确定量热芽孢杆菌。实时PCR获得的嗜热芽孢杆菌计数与天然污染的虾和鲑鱼的平板计数之间的相关性很高（R〜2 = 0.895）。因此，本研究提供了一种快速的工具，可用于生成熟虾和新鲜鲑鱼中嗜热芽孢杆菌的可靠定量数据。

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  • 来源
    《Food microbiology》 |2012年第1期|p.173-179|共7页
  • 作者

    Kelthoum Mamlouk; Sabrina Mace; Morgan Guilbaud; Emmanuel Jaffres; Mounir Ferchichi; Herve Prevost; Marie-France Pilet; Xavier Dousse;

  • 作者单位

    LUNAM Universite, Oniris. UMR1014 Secalim, Nantes, F-44307, France,INRA, Nantes, F-44307, France;

    LUNAM Universite, Oniris. UMR1014 Secalim, Nantes, F-44307, France,INRA, Nantes, F-44307, France;

    INRA, UMR 1319 M1CAL1S, F-7835 jouy-en-josas, France,AgroParisTech, UMR 1319 M1CAUS, F-9130 Massy, France;

    LUNAM Universite, Oniris. UMR1014 Secalim, Nantes, F-44307, France,INRA, Nantes, F-44307, France;

    LUNAM Universite, Oniris. UMR1014 Secalim, Nantes, F-44307, France,INRA, Nantes, F-44307, France;

    LUNAM Universite, Oniris. UMR1014 Secalim, Nantes, F-44307, France,INRA, Nantes, F-44307, France;

    LUNAM Universite, Oniris. UMR1014 Secalim, Nantes, F-44307, France,INRA, Nantes, F-44307, France;

    LUNAM Universite, Oniris. UMR1014 Secalim, Nantes, F-44307, France,INRA, Nantes, F-44307, France;

  • 收录信息 美国《科学引文索引》(SCI);美国《化学文摘》(CA);
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类
  • 关键词

    brochothrix thermosphacta; real-time PCR; 16S rRNA; seafood; spoilage; STAA medium; propidium monoazide;

    机译：嗜热胸膜炎实时PCR;16S rRNA;海鲜;损坏;STAA培养基;叠氮化丙锭;

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