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Analysis of emetic toxin production by Bacillus species using cellular cytotoxicity, molecular, and chromatographic assays

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Abstract

In this study, we analyzed production of the emetic toxin, cereulide, in 30 Bacillus strains by using a cellular cytotoxicity (3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide; MTT) assay, polymerase chain reaction (PCR), and micellar electrokinetic chromatographycapillary electrophoresis (MEKC-CE) analysis. The Bacillus cereus emetic strains produced 60 ∼ 227 µg/mL of cereulide when analyzed using MEKC-CE. Some other Bacillus species, including B. subtilis, B. pumilus, and B. megaterium, produced putative cereulide at levels similar to those produced by emetic B. cereus, whereas B. mycoides and B. thuringiensis did not produce the putative toxin or produced it at a concentration less than 2 µg/mL. Only B. cereus emetic strains were found to be toxic to bovine fibroblast cells, with the exception of one Bacillus diarrheal strain. The PCR results correlated with the MTT assay results, except in the case of one B. cereus diarrheal strain. This strain may produce either unusually heat-stable enterotoxins or is a co-producer of emetic and diarrheal toxins. Collectively, our results indicate that several Bacillus species produce toxin(s) with structures and properties similar to those of cereulide.

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References

  1. Granum, P. E. (1994) Bacillus cereus and its toxins. J. Appl. Microbiol. 76: 61S–66S.

    Google Scholar 

  2. Oh, M. and J. M. Cox (2009) Toxigenic bacilli associated with food poisoning. Food Sci. Biotechnol. 18: 594–603.

    CAS  Google Scholar 

  3. From, C., R. Pukall, P. Schumann, V. Hormazbal, and P. E. Granum (2005) Toxin-producing ability among bacillus spp. outside the Bacillus cereus group. Appl. Environ. Microbiol. 71: 1178–1183.

    Article  CAS  Google Scholar 

  4. Ehling-Schulz, M., M. Fricker, and S. Scherer (2004) Identification of emetic toxin producing Bacillus cereus strains by a novel molecular assay. FEMS Microbiol. Lett. 232: 189–195.

    Article  CAS  Google Scholar 

  5. Thorsen, L., B. M. Hansen, K. F. Nielsen, N. B. Hendriksen, R. K. Phipps, and B. B. Budde (2006) Characterization of emetic Bacillus weihenstephanensis, a new cereulide-producing bacterium. Appl. Environ. Microbiol. 72: 5118–5121.

    Article  CAS  Google Scholar 

  6. Lund, B. M. (1990) Foodborne disease due to Bacillus and clostridium species. The Lancet. 336: 982–986.

    Article  CAS  Google Scholar 

  7. Mikami, T., T. Horikawa, T. Murakami, T. Matsumoto, A. Yamakawa, S. Murayama, S. Katagiri, K. Shinagawa, and M. Suzuki (1994) An improved method for detecting cytostatic toxin (emetic toxin) of Bacillus cereus and its application to food samples. FEMS Microbiol. Lett. 119: 53–57.

    Article  CAS  Google Scholar 

  8. Hughes, S., B. Bartholomew, J. C. Hardy, and J. M. Kramer (1988) Potential application of a HEp-2 cell assay in the investigation of Bacillus cereus emetic-syndrome food poisoning. FEMS Microbiol. Lett. 52: 7–11.

    Article  Google Scholar 

  9. Andersson, M. A., R. Mikkola, J. Helin, M. C. Andersson, and M. Salkinoja-Salonen (1998) A novel sensitive bioassay for detection of bacillus cereus emetic toxin and related depsipeptide ionophores. Appl. Environ. Microbiol. 64: 1338–1343.

    CAS  Google Scholar 

  10. Finlay, W. J. J., N. A. Logan, and A. D. Sutherland (1999) Semiautomated metabolic staining assay for Bacillus cereus emetic toxin. Appl. Environ. Microbiol. 65: 1811–1812.

    CAS  Google Scholar 

  11. Toh, M., M. C. Moffitt, L. Henrichsen, M. Raftery, K. Barrow, J. M. Cox, C. P. Marquis, and B. A. Neilan (2004) Cereulide, the emetic toxin of Bacillus cereus, is putatively a product of nonribosomal peptide synthesis. J. Appl. Microbiol. 97: 992–1000.

    Article  CAS  Google Scholar 

  12. Hggblom, M. M., C. Apetroaie, M. A. Andersson, and M. S. Salkinoja-Salonen (2002) Quantitative analysis of cereulide, the emetic toxin of Bacillus cereus, produced under various conditions. Appl. Environ. Microbiol. 68: 2479–2483.

    Article  Google Scholar 

  13. Oh, M. and J. M. Cox (2010) Quantitative analysis of the bacillus cereus emetic toxin, cereulide, using micellar electrokinetic chromatography-capillary electrophoresis. J. Food Saf. 30: 652–665.

    CAS  Google Scholar 

  14. Suwan, S., M. Isobe, I. Ohtani, N. Agata, M. Mori, and M. Ohta (1995) Structure of cereulide, a cyclic dodecadepsipeptide toxin from Bacillus cereus and studies on NMR characteristics of its alkali metal complexes including a conformational structure of the K+ complex. J. Chem. Soc. Perkin Trans.1. 7: 765–775.

    Article  Google Scholar 

  15. Beattie, S. H. and A. G Williams (2000) Detection of toxins: Encyclopedia of food microbiology. pp. 141–149. Academic Press, A Harcourt Science and Technology Company, London, UK.

    Google Scholar 

  16. Beattie, S. H. and A. G. Williams (1999) Detection of toxigenic strains of Bacillus cereus and other Bacillus spp. with an improved cytotoxicity assay. Lett. Appl. Microbiol. 28: 221–225.

    Article  CAS  Google Scholar 

  17. Jenson, I., C. J. Moir, and A. D. Hocking (2003) Bacillus cereus and other Bacillus species: Foodborne microorganisms of public health significance. pp. 445–478. Academic Press, A Harcourt Science and Technology Company, London.

    Google Scholar 

  18. Toh, M. (2005) The emetic toxin, cereulide, produced by Bacillus cereus. Ph. D. thesis. Department of Food Science and Technology, The University of New South Wales, Sydney, Australia.

    Google Scholar 

  19. Taylor, J. M. W., A. D. Sutherland, K. E. Aidoo, and N. A. Logan (2005) Heat stable toxin production by strains of Bacillus cereus, Bacillus firmus, Bacillus megaterium, Bacillus simplex and Bacillus licheniformis. FEMS Microbiol. Lett. 242: 313–317.

    Article  CAS  Google Scholar 

  20. Peypoux, F., J. Bonmatin, and J. Wallach (1999) Recent trends in the biochemistry of surfactin. Appl. Microbiol. Biotechnol. 51: 553–563.

    Article  CAS  Google Scholar 

  21. From, C., V. Hormazabal, and P. E. Granum (2007) Food poisoning associated with pumilacidin-producing Bacillus pumilus in rice. Int. J. Food Microbiol. 115: 319–324.

    Article  CAS  Google Scholar 

  22. Oh, M., J. Ham, and J. M. Cox (2012) Diversity and toxigenicity among members of the bacillus cereus group. Int. J. Food Microbiol. 152: 1–8.

    Article  CAS  Google Scholar 

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Authors and Affiliations

  1. National Institute of Animal Science, Rural Development Administration, Suwon, 441-706, Korea

    Jae Gyu Yoo, Jin-Hee Chang, So-young Kim, Ju-Young Ji, Sung-Wook Hong, Beom-Young Park & Mi-Hwa Oh

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  1. Jae Gyu Yoo
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  2. Jin-Hee Chang
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  3. So-young Kim
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  7. Mi-Hwa Oh
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Correspondence to Mi-Hwa Oh.

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Yoo, J.G., Chang, JH., Kim, Sy. et al. Analysis of emetic toxin production by Bacillus species using cellular cytotoxicity, molecular, and chromatographic assays. Biotechnol Bioproc E 19, 978–983 (2014). https://doi.org/10.1007/s12257-014-0574-7

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  • DOI: https://doi.org/10.1007/s12257-014-0574-7

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