Contamination of foods and sources of the emerging diarrheagenic Fec ericbia mallet foreshadowspandemics

• Project/Area Number: 18602001
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: 食の安全
• Research Institution: Osaka City University
• Principal Investigator: NISHIKAWA Yoshikazu Osaka City University, Graduate School of Human Life Science, Professor (60183539)
• Co-Investigator(Kenkyū-buntansha): OGASAWARA Jun Osaka City Institute of Public Health and Environmental Sciences, Department of Microbiology, Senior Researcher (00270760)
• Project Period (FY): 2006 – 2007
• Project Status: Completed (Fiscal Year 2007)
• Budget Amount: ¥3,980,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥480,000); Fiscal Year 2007: ¥2,080,000 (Direct Cost: ¥1,600,000、Indirect Cost: ¥480,000); Fiscal Year 2006: ¥1,900,000 (Direct Cost: ¥1,900,000)
• Keywords: darrheagenic / Escherichia coli / real-time PCR / multiplex / food / screening / epidemiology / contamination

Research Abstract

Diarrheagenic Escherichia coli (DEC) comprise a variety of pathotypes: enteropathogenic E. coli (EPEC); shigatoxin-producing E. coil (STEC); enterotoxigenic E. coli (ETEC); enteroinvasive E. coli (EIEC); enteroaggregative E. call (EAggEC); and diffusely adhering E. coil (DAEC). However, the source and routes of infection have not been clarified because these organisms are scarcely detected among numerous coliform bacteria. To exhaustively detect DEC in food, we have developed multiplex real-time PCR assays against enterovirulent genes (eae, stx1, stx2, elt, est, virB, aggR, astA and afaB). Primers and TaqMan probes were designed to amplify and quantify the genes in duplex or triplex reactions under the same PCR cycling conditions. Specificity was confirmed using 96 DEC strains, 2 strains of Vibrio cholerae El Tor Ogawa, and Shigella dysenteriae serotype 1. The fluorescence threshold cycle and DNA concentrations correlated with decision coefficients of more than 0.99. Subsequently, ground meat samples and enrichment broths were spiked with DEC. Detection limits varied from 3.6×10^2 to 6.0×10^3 CFU/ml, depending on the target genes. All meat samples spiked with a variety of DEC (more than 10 CFU/10g) were found to be positive by screening the enrichment broth using multiplex real-time PCR. As a preliminary test, 148 food samples were exhaustively assayed by multiplex real-time PCR; 41 samples (28%) were positive for DEC pathotypes. The present multiplex real-time PCR system allows the efficient and simultaneous determination of various DEC pathotypes in food.

Research Products

• [Journal Article] Prevalence and virulence gene profiles of Shiga toxin-producing Escbericbia coli and enteropathogenic Escbericbia coli from diarrhceie and healthy lambs in India (2008)
  • Author(s): Bhat, M.A., Nishikawa, Y:, and Wani, S.A.
  • Journal Title: Small Ruminant Res. 75 Pages: 65-70
  • Description: 「研究成果報告書概要(和文)」より
  • Peer Reviewed
• [Journal Article] Prevalence and virulence gene profiles of Shiga toxin-producing Escherichia coli and enteropathogenic Escherichia coil from diarrhoeic and healthy lambs in India (2008)
  • Author(s): Bhat, M. A, Nishikawa, Y, Wani, S. A
  • Journal Title: Small Ruminant Res 75 Pages: 65-70
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Enhancement of Shiga toxin production in enterohemorrhagic Escherichia coli serotype O157:H7 by DNase colicins (2007)
  • Author(s): Toshima, H., Yoshimura, A., Arikawa., K., Hidaka, A., Ogasawara, J., Hale, A., Masaki, H., and Nishikawa, Y.
  • Journal Title: Appl.Environ.Microbiol. 73 Pages: 7582-7588
  • Description: 「研究成果報告書概要(和文)」より
  • Peer Reviewed
• [Journal Article] Association of IL-8-Inducing strains of diffusely adherent Escherichia coli with sporadic diarrheal patients less than 5 years of age (2007)
  • Author(s): Meraz, I.M., Arikawa, K., Nakamura, H., Ogasawara, J., Hase, A., the Surveillance Team, and Nishikawa, Y.
  • Journal Title: Braz.J.Infect.Dis. 11 Pages: 44-49
  • Description: 「研究成果報告書概要(和文)」より
  • Peer Reviewed
• [Journal Article] Variants of eae and stx genes of atypical enteropathogenic Escherlchie coli and non-O157 Shiga toxin-producing Escherichia coli from calves. (2007)
  • Author(s): Wani, S.A., Hussain, I., Nabi, A., Fayaz, I., and Nishikawa, Y.
  • Journal Title: Lett.Appl.Microbiol. 45 Pages: 610-615
  • Description: 「研究成果報告書概要(和文)」より
  • Peer Reviewed
• [Journal Article] Prevalence of enteric bacteria that inhibit growth of enterohaemorrhagic Escherichia coli 0157 in humans (2007)
  • Author(s): Toshima, H., Y., Hachio, M., Ikemoto, Y., Ogasawara, J., Hase, A., Takahashi, K., Masaki, N., and Nishikawa, Y.
  • Journal Title: Epidemiol. Infect. 135 Pages: 110-117
  • Description: 「研究成果報告書概要(和文)」より
  • Peer Reviewed
• [Journal Article] Enhancement of Shiga toxin production in enterohemorrhagic Escherichia coil serotype 0157:H7 by DNase colicins (2007)
  • Author(s): Toshima, H, Yoshimura, A, Arikawa, K, Hidaka, A, Ogasawara, J, Hase, A, Masaki, H, Nishikawa, Y
  • Journal Title: Appl. Environ. Microbiol 73 Pages: 7582-7588
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Association of IL-8-Inducing strains of diffusely adherent Escherichia coli with sporadic diarrheal patients less than 5 years of age (2007)
  • Author(s): Meraz, I. M, Arikawa, K, Nakamura, H, Ogasawara, J, Hase, A, the Surveillance, Team, Nishikawa, Y
  • Journal Title: Braz. J. Infect. Dis. 11 Pages: 44-49
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Variants of eae and stx genes of atypical enteropathogenic Escherichia coli and non-O157 Shiga toxin-producing Escherichia coli from calves (2007)
  • Author(s): Wani, S. A, Hussain, I, Nabi, A, Fayaz, I, Nishikawa, Y
  • Journal Title: Lett. Appl. Microbiol. 45 Pages: 610-615
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Prevalence of enteric bacteria that inhibit growth of enterohaemorrhagic Escherichia coil ○ 157 in humans (2007)
  • Author(s): Toshima, H., Y. Hachio, M., Ikemoto, Y. Ogasawara, J. Hase, A. Takahashi, K. Masaki, H. and Nishikawa, Y
  • Journal Title: Epidemiol. Infect. 135 Pages: 110-117
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Association of IL-8-Inducing strains of diffusely adherent Eseheriehia coli with sporadic diarrheal patients less than 5 years of age (2007)
  • Author(s): Meraz, I. M, Arikawa, K, Nakamura, H., Ogasawara, J., Hase, A., the Surveillance Team, and Nishfkawa, Y.
  • Journal Title: Braz. J. Infect. Dis. 11 Pages: 44-49
  • Peer Reviewed
• [Journal Article] Enhancement of Shiga toxin production in enterohemorrhagic Escherichia coli semtype O157:H7 by DNase cohcins (2007)
  • Author(s): Toshima, H., Yoshimura, A, Arikawa, K, Hidaka, A, Ogasawara, J., Hase, A., Masaki, H., and Nishikawa, Y.
  • Journal Title: Appl. Environ. Microbiol. 73 Pages: 7582-7588
  • Peer Reviewed
• [Journal Article] Variants of eae and stx genes of atypical enteropathogenic Escherichia coli and non-O157 Shiga toxin-producing Escheriehia coli from calves. (2007)
  • Author(s): Wani, S. A., Hussain, I., Nabi, A., Fayaz, I., and Nishikawa, Y.
  • Journal Title: Lett. Appl. Microbiol. 45 Pages: 610-615
  • Peer Reviewed
• [Journal Article] Prevalence of enteric bacteria that inhibit growth of enterohaemorrhagic Escherichia coli 0157 in humans (2007)
  • Author(s): Toshima, H., Y., Hachio, M., Ikemoto, Y., Ogasawara, J., Hase, A., Takahashi, K., Masaki, H., Nishikawa, Y.
  • Journal Title: Epidemiology and Infection 135・1 Pages: 110-117
• [Journal Article] Epithelial cells secreteinterleukin-8 in response to adhesion and invasion of diffusely adhering Escherichia coli lacking Afa/Dr genes (2006)
  • Author(s): Meraz, I.M., Arikawa, K., Ogasawara, J., Hase, A., and Nishikawa, Y.
  • Journal Title: Microbiol.Imnunol. 50 Pages: 159-169
  • Description: 「研究成果報告書概要(和文)」より
  • Peer Reviewed
• [Journal Article] Investigation of diarrhoeic faecal samples for enterotoxigenic,Shiga toxin-producing and typical or atypical enteropathogenic Escherichia coli in Kashmir,India (2006)
  • Author(s): Wani, S.A., Nabi, A., Fayaz, I., Ahmad, I., Nishikawa, Y., Qureshi, K., Khan, M.A., and Chowdhary, J.
  • Journal Title: FEMS Microbiol.Lett. 261 Pages: 238-244
  • Description: 「研究成果報告書概要(和文)」より
  • Peer Reviewed
• [Journal Article] Shiga toxin-producing Escherichia coli and enteropathogenic Escherichia coli in healthy goats in India:occurrence and virulence properties (2006)
  • Author(s): Wani, S.A., Samanta, I., Munshi, Z.H., Bhat, M.A., and Nishikawa, Y.
  • Journal Title: J.App1.Microbiol. 261 Pages: 108-113
  • Description: 「研究成果報告書概要(和文)」より
  • Peer Reviewed
• [Journal Article] Epithelial cells secrete interleukin-8 in response to adhesion and invasion of diffusely adhering Escherichia coli lacking Afa/Dr genes (2006)
  • Author(s): Meraz, I. M, Arikawa, K, Ogasawara, J, Hase, A, Nishikawa, Y
  • Journal Title: Microbiol. Immunol 50 Pages: 159-169
  • NAID: 10017327442
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Investigation of diarrhoeic faecal samples for enterotoxigenic, Shiga toxin-producing and typical or atypical enteropathogenic Escherichia coli in Kashmir, India (2006)
  • Author(s): Wani, S. A, Nabi, A, Fayaz, I, Ahmad, I. Nishikawa, Y, Qureshi, K, Khan, M. A, Chowdhary, J
  • Journal Title: FEMS Microbiol. Lett. 261 Pages: 238-244
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Shiga toxin-producing Escherichia coli and enteropathogenic Escherichia coli in healthy goats in India: occurrence and virulence properties (2006)
  • Author(s): Wani, S. A, Samanta, I, Munshi, Z. H, Bhat, M. A, Nishikawa, Y
  • Journal Title: J. Appl. Microbiol. 100 Pages: 108-113
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Epithelial cells secrete interleukin-8 in response to adhesion and invasion of diffusely adhering Escherichia coli lacking Afa/Dr genes (2006)
  • Author(s): Meraz, I.M., Arikawa, K., Ogasawara, J., Hase, A., Nishikawa, Y.
  • Journal Title: Microbiology and Immunology 50・3 Pages: 159-169
  • NAID: 10017327442
• [Journal Article] Investigation of diarrhoeic faecal samples for enterotoxigenic Shiga toxin-producing and typical or atypical enteropathogenic Escherichia coli in Kashmir, India (2006)
  • Author(s): Wani, S.A., Nabi, A., Fayaz, I., Ahmad, I., Nishikawa, Y., Qureshi, K., Khan, M.A., Chowdhary, J.
  • Journal Title: FEMS Microbiology Letters 261・2 Pages: 238-244
• [Journal Article] Shiga toxin-producing Escherichia coli and enteropathogenic Escherichia coli in healthy goats in India : occurrence and virulence properties (2006)
  • Author(s): Wani, S.A., Samanta, I., Munshi, Z.H., Bhat, M.A., Nishikawa, Y.
  • Journal Title: Journal of Applied Microbiology 100・1 Pages: 108-113
• [Journal Article] Molecular typing to trace Listeria monocytogenes isolated from cold-smoked fish to a contamination source in a processing plant (2006)
  • Author(s): Nakamura, H., Tokuda, Y., Sono, A., Koyama, T., Ogasawara, J., Hase, A., Haruki, K., Nishikawa, Y.
  • Journal Title: Journal of Food Protection 69・4 Pages: 835-841
• [Presentation] リアルタイムPCRを用いた食品中の下痢原性大腸菌の網羅的検出法 (2007)
  • Author(s): 日高 あゆみ、法橋 朋子、小笠原 準、長谷 篤、高鳥 浩介、西川 禎一
  • Organizer: 第28回日本食品微生物学会学術総会
  • Place of Presentation: 東京
  • Description: 「研究成果報告書概要(和文)」より
• [Presentation] Multiplex real-time PCR for exhaustive detection of diarrheagenic Escherichia coil in foods (2007)
  • Author(s): Hidaka, A, Hokyo, T, Ogasawara, J, Hase, A, Takatori, K, Nishikawa, Y
  • Organizer: The 28th Annual Meeting of Japanese Society of Food Microbiology
  • Place of Presentation: Tokyo
  • Description: 「研究成果報告書概要(欧文)」より
• [Presentation] リアルタイムPCRによる食品中の下痢原性大腸菌(EHEC, EPEC, ETEC)同時検出法 (2006)
  • Author(s): 日高 あゆみ、法橋 朋子、小笠原 準、長谷 篤、西川 禎一
  • Organizer: 第27回日本食品微生物学会学術総会
  • Place of Presentation: 大阪
  • Description: 「研究成果報告書概要(和文)」より
• [Presentation] Simultaneous detection of diarrheagenic Escherichia coli(EHEC, EPEC,ETEC)in foods by real-time PCR (2006)
  • Author(s): Hidaka, A, Hokyo, T, Ogasawara, J, Hase, A, Nishikawa, Y
  • Organizer: The 27th Annual Meeting of Japanese Society of Food Microbiology
  • Place of Presentation: Osaka
  • Description: 「研究成果報告書概要(欧文)」より
• [Book] 「食生活をおびやかす食中毒と感染症」 山口英昌編著『食環境科学入門 食の安全を環境問題の視点から』 (2006)
  • Author(s): 西川 禎一
  • Publisher: ミネルバ書房
  • Description: 「研究成果報告書概要(和文)」より
• [Book] 「EASTl遺伝子astA保有大腸菌の下痢原性に注意」「分散接着性大腸菌検査時のひと工夫」;渡辺治雄編著『食中毒検査・診療のコツと落とし穴』 (2006)
  • Author(s): 西川 禎一
  • Publisher: 中山書店
  • Description: 「研究成果報告書概要(和文)」より
• [Book] 「食生活をおびやかす食中毒と感染症」食環境科学入門 食の安全を環境問題の視点から(山口英昌編著) (2006)
  • Author(s): 西川禎一
  • Publisher: ミネルバ書房
• [Book] 「EAST1遺伝子astA保有大腸菌の下痢原性に注意」「分散接着性大腸菌検査時のひと工夫」;食中毒検査・診療のコツと落とし穴(渡辺治雄編著) (2006)
  • Author(s): 西川禎一
  • Publisher: 中山書店
• [Patent(Industrial Property Rights)] 下痢原性大腸菌検出方法 (2007)
  • Inventor(s): 西川 禎一
  • Industrial Property Rights Holder: 公立大学法人大阪市立大学、株式会社KALS、株式会社三共刃型工業
  • Filing Date: 2007-05-16
  • Description: 「研究成果報告書概要(和文)」より
• [Patent(Industrial Property Rights)] 下痢原性大腸菌検出方法 (2007)
  • Inventor(s): 西川 禎一
  • Industrial Property Rights Holder: 公立大学法人大阪市立大学、株式会社KALS、株式会社三共刄型工業
  • Filing Date: 2007-05-16