Detection of Enteric Pathogen Bacteria with the PCR Method from Mixed Fecal Specimens of Food Handlers

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  • BABA Youichi
    Laboratories for Food & Environmental Science, Tokyo kenbikyoin Foundation
  • HAYASHIDA Mizuho
    Laboratories for Food & Environmental Science, Tokyo kenbikyoin Foundation
  • TAKAOKA Naoko
    Analytical & Measuring Instru ments Division, Shimadzu Corporation
  • NISHIMURA Naoyuki
    Analytical & Measuring Instru ments Division, Shimadzu Corporation
  • ITO Takeshi
    Laboratories for Food & Environmental Science, Tokyo kenbikyoin Foundation

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Other Title
  • 食品取り扱い従事者糞便からのPCR による 腸管系病原菌保菌者検索の検討
  • ショクヒン トリアツカイ ジュウジシャ フンベン カラ ノ PCR ニ ヨル チョウカンケイ ビョウゲンキン ホキンシャ ケンサク ノ ケントウ

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Abstract

We developed an assay for rapid, specific detection of Shigella, Salmonella, and verotoxin-producing Escherichia coli O157 using the direct PCR analysis of mixed human fecal specimens. In this study, the sensitivity of the direct PCR assay for 50 mixed human fecal specimen was found to be 1.3, 0.42, and 0.76 colonyforming units/kit for Shigella sonnei, Salmonella Typhimurium, and verotoxin-producing E. coli O157, respectively. We compared the efficiency of the direct PCR method with the conventional direct agar plate method for 5000 fecal specimens from food handlers. The 50 mixed fecal specimens were concentrated to approximately 2.5％in distilled water and were heated to 95℃for 5min. Then, 5μL of the specimen supernatant was added to 45μL of the PCR mixture. Direct PCR results were evaluated by melting curve analysis. Among the 5000 fecal specimens from food handlers, Salmonella Infantis was isolated from 1 specimen using the direct agar plate method, and it was positive for the Salmonella gene (stn), as confirmed with the direct PCR method. Shigella, Salmonella, or verotoxin-producing E. coli O157 were not detected in the remaining 4999 fecal specimens with the direct agar plate method. However, Salmonella Enteritidis isolated by the enrichment culture method from 1 fecal specimen was positive for stn. Non-O157 verotoxin-producing E. coli isolated using direct horse blood agar from another fecal specimen was positive for stx. Moreover, enteroinvasive E. coli isolated using direct BTB agar from one fecal specimen was positive for ipaH. We conclude that the direct PCR assay can be applied as a new rapid screening method for personal hygiene among food handlers.

Journal

  • Kansenshogaku Zasshi

    Kansenshogaku Zasshi 88 (5), 685-694, 2014

    The Japanese Association for Infectious Diseases

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