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Development of a Quantitative Real-time Polymerase Chain Reaction Method to Enumerate Total Bacterial Count in Ready-to- eat Fruits and Vegetables

Research Project

Project/Area Number 18580314
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field Applied veterinary science
Research InstitutionNational Institute of Health Sciences

Principal Investigator

YUKIKO Kodo  National Institute of Health Sciences, Div. Microbiology, Chief (50218632)

Project Period (FY) 2006 – 2007
Project Status Completed (Fiscal Year 2007)
Budget Amount *help
¥4,010,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥510,000)
Fiscal Year 2007: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000)
Fiscal Year 2006: ¥1,800,000 (Direct Cost: ¥1,800,000)
KeywordsFood / Hygiene / Bacterium / Microbiology / Veterinary
Research Abstract

A newly developed real-time PCR assay rapidly quantifies the total bacterial numbers in contaminating ready-to-eat vegetables and fruits as compared to the standard plate count method. Primers targeting the rpoB gene that encodes for the β-subunit of the bacterial RNA polymerase, common to most bacterial species was used instead of the 16S rRNA gene that has multiple copies and varies among bacterial species. A primer pair specific for rpoB was confirmed to amplify rpoB in wide-range of bacterial species by using 49 strains isolated from five kinds of fruits and vegetables. We purchased fruits and vegetables from retail shops and enumerated the bacteria associated with them using the real-time PCR and compared this to the number found by the culture method. We found a high correlation between the threshold PCR cycle numbers when compared to the plate count culture number. Further, the predominant bacterial flora detected by the culture method was compared with those detected by the real-time PCR assay using a denaturing gradient gel electrophoresis analysis. The results from the two methods were almost identical. The real-time PCR assay developed in this study can enumerate the dominant bacterial species in ready-to-eat fruits and vegetables.

Report

(3 results)
  • 2007 Annual Research Report   Final Research Report Summary
  • 2006 Annual Research Report
  • Research Products

    (4 results)

All 2007 2006

All Journal Article (3 results) (of which Peer Reviewed: 1 results) Presentation (1 results)

  • [Journal Article] Development of a quantitative real-time polymerase chain reaction method to enumerate total bacterial count in ready-to-eat fruits and vegetables2006

    • Author(s)
      Takahashi, H., Konuma, H., Hara-Kudo, Y.
    • Journal Title

      Journal of Food Protection 69

      Pages: 2504-2508

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2007 Final Research Report Summary
    • Peer Reviewed
  • [Journal Article] Development of a quantitative real-time polymerase chain reaction method to enumerate total bacterial count in ready-to-eat fruits and vegetables.2006

    • Author(s)
      Takahashi, H., Konuma, H., Hara-Kudo, Y.
    • Journal Title

      J. Food Prot. 69

      Pages: 2504-2508

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Journal Article] Development of a quantitative real-time polymerase chain reaction method to enumerate total bacterial count in ready-to-eat fruits and vegetables2006

    • Author(s)
      Takahashi, H., Konuma, H., Hara-Kudo, Y.
    • Journal Title

      Journal of Food Protection 69・10

      Pages: 2504-2508

    • Related Report
      2006 Annual Research Report
  • [Presentation] リアルタイムPCRによる野菜類の生菌数推定2007

    • Author(s)
      田中啓子、今井伸二郎、工藤由起子.
    • Organizer
      第28回日本食品微生物学会
    • Place of Presentation
      東京都内
    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2007 Annual Research Report 2007 Final Research Report Summary

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Published: 2006-04-01   Modified: 2016-04-21  

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