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Use of a Vero Cell-Based Fluorescent Assay to Assess Relative Toxicities of Shiga Toxin 2 Subtypes from Escherichia coli

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Book cover Microbial Toxins

Part of the book series: Methods in Molecular Biology ((MIMB,volume 739))

Abstract

Shiga toxin-producing Escherichia coli is a leading cause of human gastroenteritis from food and waterborne sources worldwide. Shiga toxins 1 and 2 are important virulence factors linked to severe human illness. In particular, Shiga toxin 2 is composed of a diverse and heterogeneous group of subtypes with differential cytotoxicities in mammalian cells. In this chapter, we describe the use of the Vero-d2EGFP fluorescent assay to examine the relative toxicities of Stx2 and Stx2 subtypes expressed by strains of Shiga toxin-producing E. coli.

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References

  1. Rangel JM, Sparling PH, Crowe C, Griffin PM, Swerdlow DL (2005) Epidemiology of Escherichia coli O157:H7 outbreaks, United States, 1982–2002. Emerg Infect Dis 11:603–609

    Article  PubMed  Google Scholar 

  2. Karmali MA (2004) Infection by Shiga toxin-producing Escherichia coli: an overview. Mol Biotechnol 26:117–122

    Article  PubMed  CAS  Google Scholar 

  3. Bettelheim KA (2007) The non-O157 Shiga-toxigenic (verocytotoxigenic) Escherichia coli; under-rated pathogens. Crit Rev Microbiol 33:67–87

    Article  PubMed  Google Scholar 

  4. O’Brien AD, Kaper JD (1998) Shiga Toxin-Producing Escherichia coli: yesterday, today, and tomorrow. In: Kaper JD, O’Brien AD (eds) Escherichia coli O157:H7 and other Shiga Toxin-producing E. coli strains. ASM Press, Washington, DC, pp 1–11

    Google Scholar 

  5. Serna A, Boedeker EC (2008) Pathogenesis and treatment of Shiga toxin-producing Escherichia coli infections. Curr Opin Gastroenterol 24:38–47

    Article  PubMed  CAS  Google Scholar 

  6. Boerlin P, McEwen SA, Boerlin-Petzold F, Wilson JB, Johnson RP, Gyles CL (1999) Associations between virulence factors of Shiga toxin-producing Escherichia coli and disease in humans. J Clin Microbiol 37:497–503

    PubMed  CAS  Google Scholar 

  7. Ostroff SM, Tarr PI, Neill MA, Lewis JH, Hargrett-Bean N, Kobayashi JM (1989) Toxin genotypes and plasmid profiles as determinants of systemic sequelae in Escherichia coli O157:H7 infections. J Infect Dis 160:994–998

    Article  PubMed  CAS  Google Scholar 

  8. Müthing J, Schweppe CH, Karch H, Friedrich AW (2009) Shiga toxins, glycosphingolipid diversity, and endothelial cell injury. Thromb Haemost 101:252–264

    PubMed  Google Scholar 

  9. Scheutz F, Strockbine NA (2005) Genus I. Escherichia. In: Garrity GM, Brenner DJ, Krieg NR, Staley JT (eds) Bergey’s manual of systematic bacteriology, 2nd edn. Springer, New York, pp 607–624

    Google Scholar 

  10. Schmidt H, Scheef J, Morabito S, Caprioli A, Wieler LH, Karch H (2000) A new Shiga toxin 2 variant (Stx2f) from Escherichia coli isolated from pigeons. Appl Environ Microbiol 66:1205–1208

    Article  PubMed  CAS  Google Scholar 

  11. Schmitt CK, McKee ML, O’Brien AD (1991) Two copies of Shiga-like toxin II-related genes common in enterohemorrhagic Escherichia coli strains are responsible for the antigenic heterogeneity of the O157:H- strain E32511. Infect Immun 59:1065–1073

    PubMed  CAS  Google Scholar 

  12. Ito H, Terai A, Kurazono H, Takeda Y, Nishibuchi M (1990) Cloning and nucleotide sequencing of Vero toxin 2 variant genes from Escherichia coli O91:H21 isolated from a patient with the hemolytic uremic syndrome. Microb Pathog 8:47–60

    Article  PubMed  CAS  Google Scholar 

  13. Melton-Celsa AR, Kokai-Kun JF, O’Brien AD (2002) Activation of Shiga toxin type 2d (Stx2d) by elastase involves cleavage of the C-terminal two amino acids of the A2 peptide in the context of the appropriate B pentamer. Mol Microbiol 43:207–215

    Article  PubMed  CAS  Google Scholar 

  14. Piérard D, Muyldermans G, Moriau L, Stevens D, Lauwers S (1998) Identification of new verocytotoxin type 2 variant B-subunit genes in human and animal Escherichia coli isolates. J Clin Microbiol 36:3317–3322

    PubMed  Google Scholar 

  15. Persson S, Olsen KE, Ethelberg S, Scheutz F (2007) Subtyping method for Escherichia coli Shiga toxin (verocytotoxin) 2 variants and correlations to clinical manifestations. J Clin Microbiol 45:2020–2024

    Article  PubMed  CAS  Google Scholar 

  16. Samuel JE, Perera LP, Ward S, O’Brien AD, Ginsburg V, Krivan HC (1990) Comparison of the glycolipid receptor specificities of Shiga-like toxin type II and Shiga-like toxin type II variants. Infect Immun 58:611–618

    PubMed  CAS  Google Scholar 

  17. Waddell T, Head S, Petric M, Cohen A, Lingwood C (1988) Globotriosyl ceramide is specifically recognized by the Escherichia coli verocytotoxin 2. Biochem Biophys Res Commun 152:674–679

    Article  PubMed  CAS  Google Scholar 

  18. DeGrandis S, Law H, Brunton J, Gyles C, Lingwood CA (1989) Globotetraosylceramide is recognized by the pig edema disease toxin. J Biol Chem 264:12520–12525

    PubMed  CAS  Google Scholar 

  19. Keusch GT, Jacewicz M, Acheson DWK, Donohue-Rolfe A, Kane AV, McCluer RH (1995) Globotriaosylceramide, Gb3, is an alternative functional receptor for Shiga-like toxin 2e. Infect Immun 63:1138–1141

    PubMed  CAS  Google Scholar 

  20. Dean-Nystrom EA, Melton-Celsa AR, Pohlenz JFL, Moon HW, O’Brien AD (2003) Comparative pathogenicity of Escherichia coli O157 and intimin-negative non-O157 Shiga toxin-producing E. coli strains in neonatal pigs. Infect Immun 71:6526–6533

    Article  PubMed  CAS  Google Scholar 

  21. Zhang X, McDaniel AD, Wolf LE, Keusch GT, Waldor MK, Acheson DWK (2000) Quinolone antibiotics induce Shiga toxin-encoding bacteriophages, toxin production, and death in mice. J Infect Dis 181:664–670

    Article  PubMed  CAS  Google Scholar 

  22. de Sablet T, Bertin Y, Vareille M, Girardeau JP, Garrivier A, Gobert AP, Martin C (2008) Differential expression of stx2 variants in Shiga toxin-producing Escherichia coli belonging to seropathotypes A and C. Microbiology 154:176–186

    Article  PubMed  Google Scholar 

  23. Zhang W, Bielaszewska M, Friedrich AW, Kuczius T, Karch H (2005) Transcriptional analysis of genes encoding Shiga toxin 2 and its variants in Escherichia coli. Appl Environ Microbiol 71:558–561

    Article  PubMed  CAS  Google Scholar 

  24. Quiñones B, Massey S, Friedman M, Swimley MS, Teter K (2009) Novel cell-based method to detect Shiga toxin 2 from Escherichia coli O157:H7 and inhibitors of toxin activity. Appl Environ Microbiol 75:1410–1416

    Article  PubMed  Google Scholar 

  25. Cooley M, Carychao D, Crawford-Miksza L, Jay MT, Myers C, Rose C, Keys C, Farrar J, Mandrell RE (2007) Incidence and tracking of Escherichia coli O157:H7 in a major produce production region in California. PLoS ONE 2:e1159

    Article  PubMed  Google Scholar 

  26. Marques LRM, Peiris JSM, Cryz SJ, O’Brien AD (1987) Escherichia coli strains isolated from pigs with edema disease produce a variant of Shiga-like toxin II. FEMS Microbiol Lett 44:33–38

    Article  CAS  Google Scholar 

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Acknowledgments

The authors wish to thank Professor Ken Teter from the University of Central Florida, Orlando, Florida, for providing the mammalian cell lines Vero-d2EGFP and Vero, and Robert E. Mandrell from the USDA/ARS/Western Regional Research Center, Albany, California, for providing the STEC strains used in this study. This work was supported by the US Department of Agriculture, Agricultural Research Service CRIS project number 5325-42000-045.

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Authors and Affiliations

  1. United States Department of Agriculture/Agricultural Research Service, Produce Safety and Microbiology Research Unit, Western Regional Research Center, Albany, CA, USA

    Beatriz Quiñones

Authors
  1. Beatriz Quiñones
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  2. Michelle S. Swimley
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Editor information

Editors and Affiliations

  1. Leibniz-Zentrum fur Medizin und Biowisse, Forschungsinstitut Borstel, Parkallee 4a/c, Borstel, 23845, Germany

    Otto Holst

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Quiñones, B., Swimley, M.S. (2011). Use of a Vero Cell-Based Fluorescent Assay to Assess Relative Toxicities of Shiga Toxin 2 Subtypes from Escherichia coli . In: Holst, O. (eds) Microbial Toxins. Methods in Molecular Biology, vol 739. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-102-4_6

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  • DOI: https://doi.org/10.1007/978-1-61779-102-4_6

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-61779-101-7

  • Online ISBN: 978-1-61779-102-4

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