SpringerLink
Log in

Rapid Detection of Diarrheagenic Escherichia coli by a New Multiplex Real-Time Quantitative PCR Assay

  • Published:
Applied Biochemistry and Microbiology Aims and scope Submit manuscript

Abstract

Diarrheagenic Escherichia coli (DEC) is a group of important foodborne pathogens that can spread and cause infection in humans. As a significant public health issue, DEC infection may lead to a series of diseases such as abdominal pain, diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome. Therefore, its rapid and accurate detection has become an important concern for food safety issues and healthcare-associated infections. We have developed a multiplex real-time quantitative PCR assay (qPCR), DEC Sensor Test, to simultaneously recognize 5 major DEC pathotypes in a single tube, enterotoxigenic, enterohemorrhagic, enteropathogenic, enteroaggregative and enteroinvasive E. coli. In this assay, 13 DEC virulence genes, stp, sth, lt, stx1, stx2, eae, escV, bfpB, aggR, astA, pic, invE, and ipaH, can be simultaneously detected along with a spiked-in internal amplification control to avoid false negative results. All the constituent PCR for the 13 DEC virulence genes had high analytical sensitivity and specificity. Detection of all virulence genes by DEC Sensor Test can be much easier, cheaper and faster to perform than conventional tests. It also has added advantages of being able to distinguish live or dead bacteria as well as eliminating the possibility of switching partial results between different samples associated with multi-tube tests. Therefore, the application of this assay may significantly enhance our ability to prevent and fight DEC-associated diseases and to improve public health.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Subscribe and save

Springer+ Basic
EUR 32.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or Ebook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1.
Fig. 2.

Similar content being viewed by others

REFERENCES

  1. Scallan, E., Hoekstra, R.M., Angulo, F.J., Tauxe, R.V., Widdowson, M.A., Roy, S.L., et al., Emerging Infect. Dis., 2011, vol. 17, no. 1, pp. 7–15.

    Article  Google Scholar 

  2. Brandal, L.T., Lindstedt, B.A., Aas, L., Stavnes, T.L., Lassen, J., and Kapperud, G., J. Microbiol. Methods, 2007, vol. 68, no. 2, pp. 331–341.

    Article  CAS  Google Scholar 

  3. Chowdhury, F., Khan, I.A., Patel, S., Siddiq, A.U., Saha, N.C., Khan, A.I., et al., PLoS One, 2015, vol. 10, no. 6. e0130105.

    Article  Google Scholar 

  4. Patzi-Vargas, S., Zaidi, M.B., Perez-Martinez, I., Leon-Cen, M., Michel-Ayala, A., Chaussabel, D., and Estrada-Garcia, T., PLoS Neglected Trop. Dis., 2015, vol. 9, no. 3. e0003510.

    Article  Google Scholar 

  5. Yang, S.C., Lin, C.H., Aljuffali, I.A., and Fang, J.Y., Arch. Microbiol., 2017, vol. 199, no. 6, pp. 811–825.

    Article  CAS  Google Scholar 

  6. Nyachuba, D.G., Nutr. Rev., 2010, vol. 68, no. 5, pp. 257–269.

    Article  Google Scholar 

  7. McPherson, M., Kirk, M.D., Raupach, J., Combs, B., and Butler, J.R., Foodborne Pathog. Dis., 2011, vol. 8, no. 1, pp. 55–62.

    Article  Google Scholar 

  8. Toljander, J., Dovarn, A., Andersson, Y., Ivarsson, S., and Lindqvist, R., Scand. J. Public Health, 2012, vol. 40, no. 3, pp. 294–302.

    Article  Google Scholar 

  9. Nataro, J.P., and Kaper, J.B., Am. Soc. Microbiol., 1998, vol. 11, no. 1, pp. 142–201.

    CAS  Google Scholar 

  10. Toma, C., Lu, Y., Higa, N., Nakasone, N., Chinen, I., Baschkier, A., et al., J. Clin. Microbiol., 2003, vol. 41, no. 6, pp. 2669–2671.

    Article  CAS  Google Scholar 

  11. Eigner, U., Hiergeist, A., Veldenzer, A., Rohlfs, M., Schwarz, R., and Holfelder, M., Diagn. Microbiol. Infect. Dis., 2017, vol. 88, no. 1, pp. 12–16.

    Article  CAS  Google Scholar 

  12. Hazen, T.H., Michalski, J., Luo, Q., Shetty, A.C., Daugherty, S.C., Fleckenstein, J.M., and Rasko, D.A., Sci. Rep., 2017, vol. 7, no. 1, pp. 3513.

    Article  Google Scholar 

  13. Gomes, T.A., Elias, W.P., Scaletsky, I.C., Guth, B.E., Rodrigues, J.F., Piazza, R.M., et al., Braz. J. Microbiol., 2016, vol. 47, suppl. 1, pp. 3–30.

    Article  CAS  Google Scholar 

  14. Sanchez, J., and Holmgren, J., Curr. Opin. Immunol., 2005, vol. 17, no. 4, pp. 388–398.

    Article  CAS  Google Scholar 

  15. Kendall, M.M., Gruber, C.C., Rasko, D.A., Hughes, D.T., and Sperandio, V., J. Bacteriol., 2011, vol. 193, no. 24, pp. 6843–6851.

    Article  CAS  Google Scholar 

  16. Jerse, A.E., Yu, J., Tall, B.D., and Kaper, J.B., Proc. Natl. Acad. Sci. U. S. A., 1990, vol. 87, no. 20, pp. 7839–7843.

    Article  CAS  Google Scholar 

  17. HALE, T.L., Microbiol. Rev., 1991, vol. 55, no. 2, pp. 206–224.

    Article  CAS  Google Scholar 

  18. Uber, A.P., Trabulsi, L.R., Irino, K., Beutin, L., Ghilardi, A.C., Gomes, T.A., et al., FEMS Microbiol. Lett., 2006, vol. 256, no. 2, pp. 251–257.

    Article  CAS  Google Scholar 

  19. Kaper, J.B., Nataro, J.P., and Mobley, H.L., Nat. Rev. Microbiol., 2004, vol. 2, no. 2, pp. 123–140.

    Article  CAS  Google Scholar 

  20. Menard, L.P., and Dubreuil, J.D., Crit. Rev. Microbiol., 2002, vol. 28, no. 1, pp. 43–60.

    Article  CAS  Google Scholar 

  21. Wang, J., Lin, C.-W., Zhao, X., and Oh, D.H., J. Microbiol. Biotechnol., 2014, vol. 24, no. 3, pp. 297–312.

    Article  Google Scholar 

  22. Kambire, O., Adingra, A.A., Yao, K.M., and Koffi-Nevry, R., Int. J. Microbiol., 2017, vol. 2017, no. 9532170, p. 8.

    Article  Google Scholar 

  23. Yokoyama, E., Uchimura, M., and Ito, K., J. Food Prot., 2010, vol. 73, no. 6, pp. 1064–1072.

    Article  CAS  Google Scholar 

  24. Li, Y., Liu, D., Cao, B., Han, W., Liu, Y., Liu, F., et al., J. Clin. Microbiol., 2006, vol. 44, no. 12, pp. 4376–4383.

    Article  CAS  Google Scholar 

  25. Chen, Q., Shi, X., Li, Y., Jiang, Y., Lin, Y., Qiu, Y., et al., Ann. Clin. Microbiol. Antimicrob., 2014, vol. 13, no. 1, pp. 30.

    Article  Google Scholar 

  26. Law, J.W., Ab Mutalib, N.S., Chan, K.G., and Lee, L.H., Front. Microbiol., 2014, vol. 5, no. 770.

  27. Calvó, L., Martínez-Planells, A., Pardos-Bosch, J., and Garcia-Gil, L.J., Food Anal. Method, 2008, vol. 1, no. 4, pp. 236–242.

    Article  Google Scholar 

  28. Blanco-Abad, V., Ansede-Bermejo, J., Rodriguez-Castro, A., and Martinez-Urtaza, J., Int. J. Food Microbiol., 2009, vol. 129, no. 3, pp. 229–236.

    Article  CAS  Google Scholar 

  29. Sanchez, M.A., and Blower, S.M., Am. J. Epidemiol., 1997, vol. 145, no. 12, pp. 1127–1137.

    Article  CAS  Google Scholar 

  30. Postollec, F., Falentin, H., Pavan, S., Combrisson, J., and Sohier, D., Food Microbiol., 2011, vol. 28, no. 5, pp. 848–861.

    Article  CAS  Google Scholar 

  31. Chandra, M., Cheng, P., Rondeau, G., Porwollik, S., and McClelland, M., Int. J. Med. Microbiol., 2013, vol. 303, no. 4, pp. 210–216.

    Article  CAS  Google Scholar 

  32. Vendruscolo, J.W., Waldrich, T.L., Saikawa, G.I.A., Pelayo, J.S., Kobayashi, R.K.T., Nakazato, G., and Rocha, S.P.D., Genet. Mol. Res., 2017, vol. 16, no. 3, pp. 1–6.

    Article  Google Scholar 

  33. Bischoff, C., Luthy, J., Altwegg, M., and Baggi, F., J. Microbiol. Methods, 2005, vol. 61, no. 3, pp. 335–341.

    Article  CAS  Google Scholar 

  34. Guion, C.E., Ochoa, T.J., Walker, C.M., Barletta, F., and Cleary, T.G., J. Clin. Microbiol., 2008, vol. 46, no. 5, pp. 1752–1757.

    Article  CAS  Google Scholar 

Download references

Funding

This work was supported by “Hundred Talents Program” of Chinese Academy of Sciences (Grant no. Y421021102, Grant no. Y521051102), Key Technologies R & D Program for Social Development of Jiangsu Province (Grant no. BE2016685, Grant no. BE2015645), Suzhou Leading Talent Project (Grant no. ZXL2014128), New Medical Devices and Pharmaceuticals Fund of Suzhou (Grant no. ZXY201430).

Author information

Authors and Affiliations

  1. School of Life Sciences, University of Science and Technology of China, 230026, Hefei, Anhui, China

    J. Sun

  2. Suzhou Institute of Biomedical Engineering and Technology, Chinese Academy of Sciences, 215163, Suzhou, Jiangsu, China

    J. Sun, Y. Du, Z. Liu, H. Wang, Y. Ma & M. Zheng

  3. The First Affiliated Hospital of Jilin University, 130021, Changchun, Jilin, China

    Y. Shi

  4. School of Medical Technology, Xuzhou Medical University, 221004, Xuzhou, Jiangsu, China

    Z. Wang

  5. Zhejiang University Kunshan Biotechnology Laboratory, Zhejiang University Kunshan Innovation Institute, 215300, Kunshan, Jiangsu, China

    G. Zhao

Authors
  1. J. Sun
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Y. Shi
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Y. Du
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Z. Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Z. Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. H. Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. G. Zhao
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Y. Ma
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. M. Zheng
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

J. Sun, Y. Shi, M. Zheng, Y. Ma, and G. Zhao contributed to the conception and design of the study. J. Sun, Y. Shi, Y. Du, Z. Wang, Z. Liu, and H. Wang performed the experiments. J. Sun, Y. Shi, and Y. Du analyzed the data and performed the statistical analysis. J. Sun, M. Zheng, Y. Ma, and G. Zhao wrote this manuscript. All authors approved the manuscript submitted.

Corresponding authors

Correspondence to G. Zhao, Y. Ma or M. Zheng.

Ethics declarations

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interests. This article does not contain any studies involving animals or human participants performed by any of the authors.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Sun, J., Shi, Y., Du, Y. et al. Rapid Detection of Diarrheagenic Escherichia coli by a New Multiplex Real-Time Quantitative PCR Assay. Appl Biochem Microbiol 56, 748–757 (2020). https://doi.org/10.1134/S0003683820060174

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1134/S0003683820060174

Keywords:

Search

Navigation