1887

Abstract

is the causative agent of the majority of antibiotic associated diarrhoea cases. spores are recognized as the persistent and infectious morphotype as well as the vehicle of transmission of CDI. However, there is a lack of knowledge on how spores interact with the host’s epithelial surfaces. In this context, we have characterized the ability of spores to adhere to human Caco-2 cells. Despite the similarities in spore-surface hydrophobicity between spores of and (another enteric pathogen that also sporulates in the gut), spores of adhere better to Caco-2 cells. Adherence to Caco-2 cells was significantly reduced when spores were treated with trypsin. Sonication of spores altered the ultrastructure of the outermost exosporium-like structure, releasing two protein species of ~40 kDa and significantly reduced spore hydrophobicity and adherence to Caco-2 cells. Using a trifunctional cross-linker, we were able to co-immunoprecipitate four protein species from the surface of Caco-2 cells. In conclusion, this study provides evidence that spores adhere to human intestinal enterocyte-like cells through spore- and enterocytic-surface-specific ligand(s) and/or receptor(s).

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2012-09-01
2022-01-18
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References

  1. Agerer F., Waeckerle S., Hauck C. R. 2004; Microscopic quantification of bacterial invasion by a novel antibody-independent staining method. J Microbiol Methods 59:23–32 [View Article][PubMed]
    [Google Scholar]
  2. Akerlund T., Persson I., Unemo M., Norén T., Svenungsson B., Wullt M., Burman L. G. 2008; Increased sporulation rate of epidemic Clostridium difficile type 027/NAP1. J Clin Microbiol 46:1530–1533 [View Article][PubMed]
    [Google Scholar]
  3. Andersson A., Granum P. E., Rönner U. 1998; The adhesion of Bacillus cereus spores to epithelial cells might be an additional virulence mechanism. Int J Food Microbiol 39:93–99 [View Article][PubMed]
    [Google Scholar]
  4. Auwerx J. 1991; The human leukemia cell line, THP-1: a multifaceted model for the study of monocyte-macrophage differentiation. Experientia 47:22–31 [View Article][PubMed]
    [Google Scholar]
  5. Barbut F., Richard A., Hamadi K., Chomette V., Burghoffer B., Petit J. C. 2000; Epidemiology of recurrences or reinfections of Clostridium difficile-associated diarrhea. J Clin Microbiol 38:2386–2388[PubMed]
    [Google Scholar]
  6. Brahmbhatt T. N., Janes B. K., Stibitz E. S., Darnell S. C., Sanz P., Rasmussen S. B., O’Brien A. D. 2007; Bacillus anthracis exosporium protein BclA affects spore germination, interaction with extracellular matrix proteins, and hydrophobicity. Infect Immun 75:5233–5239 [View Article][PubMed]
    [Google Scholar]
  7. Calabi E., Calabi F., Phillips A. D., Fairweather N. F. 2002; Binding of Clostridium difficile surface layer proteins to gastrointestinal tissues. Infect Immun 70:5770–5778 [View Article][PubMed]
    [Google Scholar]
  8. Cerquetti M., Serafino A., Sebastianelli A., Mastrantonio P. 2002; Binding of Clostridium difficile to Caco-2 epithelial cell line and to extracellular matrix proteins. FEMS Immunol Med Microbiol 32:211–218 [View Article][PubMed]
    [Google Scholar]
  9. Collie R. E., McClane B. A. 1998; Evidence that the enterotoxin gene can be episomal in Clostridium perfringens isolates associated with non-food-borne human gastrointestinal diseases. J Clin Microbiol 36:30–36[PubMed]
    [Google Scholar]
  10. Denève C., Janoir C., Poilane I., Fantinato C., Collignon A. 2009; New trends in Clostridium difficile virulence and pathogenesis. Int J Antimicrob Agents 33:Suppl. 1S24–S28 [View Article][PubMed]
    [Google Scholar]
  11. Dingle T., Mulvey G. L., Humphries R. M., Armstrong G. D. 2010; A real-time quantitative PCR assay for evaluating Clostridium difficile adherence to differentiated intestinal Caco-2 cells. J Med Microbiol 59:920–924 [View Article][PubMed]
    [Google Scholar]
  12. Dingle T. C., Mulvey G. L., Armstrong G. D. 2011; Mutagenic analysis of the Clostridium difficile flagellar proteins, FliC and FliD, and their contribution to virulence in hamsters. Infect Immun 79:4061–4067 [View Article][PubMed]
    [Google Scholar]
  13. Drudy D., O’Donoghue D. P., Baird A., Fenelon L., O’Farrelly C. 2001; Flow cytometric analysis of Clostridium difficile adherence to human intestinal epithelial cells. J Med Microbiol 50:526–534[PubMed]
    [Google Scholar]
  14. Duncan C. L., Strong D. H. 1968; Improved medium for sporulation of Clostridium perfringens. Appl Microbiol 16:82–89[PubMed]
    [Google Scholar]
  15. Eveillard M., Fourel V., Bare M.-C., Kernéis S., Coconnier M.-H., Karjalainen T., Bourlioux P., Servin A. L. 1993; Identification and characterization of adhesive factors of Clostridium difficile involved in adhesion to human colonic enterocyte-like Caco-2 and mucus-secreting HT29 cells in culture. Mol Microbiol 7:371–381 [View Article][PubMed]
    [Google Scholar]
  16. Goulding D., Thompson H., Emerson J., Fairweather N. F., Dougan G., Douce G. R. 2009; Distinctive profiles of infection and pathology in hamsters infected with Clostridium difficile strains 630 and B1. Infect Immun 77:5478–5485 [View Article][PubMed]
    [Google Scholar]
  17. Hennequin C., Janoir C., Barc M. C., Collignon A., Karjalainen T. 2003; Identification and characterization of a fibronectin-binding protein from Clostridium difficile. Microbiology 149:2779–2787 [View Article][PubMed]
    [Google Scholar]
  18. Henriques A. O., Moran C. P. Jr 2007; Structure, assembly, and function of the spore surface layers. Annu Rev Microbiol 61:555–588 [View Article][PubMed]
    [Google Scholar]
  19. Hookman P., Barkin J. S. 2009; Clostridium difficile associated infection, diarrhea and colitis. World J Gastroenterol 15:1554–1580 [View Article][PubMed]
    [Google Scholar]
  20. Kang T. J., Fenton M. J., Weiner M. A., Hibbs S., Basu S., Baillie L., Cross A. S. 2005; Murine macrophages kill the vegetative form of Bacillus anthracis. Infect Immun 73:7495–7501 [View Article][PubMed]
    [Google Scholar]
  21. Kuehne S. A., Cartman S. T., Heap J. T., Kelly M. L., Cockayne A., Minton N. P. 2010; The role of toxin A and toxin B in Clostridium difficile infection. Nature 467:711–713 [View Article][PubMed]
    [Google Scholar]
  22. Kuijper E. J., Coignard B.and Tüll P.on behalf of ESCMID Study Group for Clostridium difficile (ESGCD) and EU Member Statesthe European Centre for Disease Prevention and Control 2006; Emergence of Clostridium difficile-associated disease in North America and Europe. Clin Microbiol Infect 12:Suppl. 62–18 [View Article][PubMed]
    [Google Scholar]
  23. Lawley T. D., Croucher N. J., Yu L., Clare S., Sebaihia M., Goulding D., Pickard D. J., Parkhill J., Choudhary J., Dougan G. 2009; Proteomic and genomic characterization of highly infectious Clostridium difficile 630 spores. J Bacteriol 191:5377–5386 [View Article][PubMed]
    [Google Scholar]
  24. Lyras D., O’Connor J. R., Howarth P. M., Sambol S. P., Carter G. P., Phumoonna T., Poon R., Adams V., Vedantam G. other authors 2009; Toxin B is essential for virulence of Clostridium difficile. Nature 458:1176–1179 [View Article][PubMed]
    [Google Scholar]
  25. McEllistrem M. C., Carman R. J., Gerding D. N., Genheimer C. W., Zheng L. 2005; A hospital outbreak of Clostridium difficile disease associated with isolates carrying binary toxin genes. Clin Infect Dis 40:265–272 [View Article][PubMed]
    [Google Scholar]
  26. McFarland L. V. 2005; Alternative treatments for Clostridium difficile disease: what really works?. J Med Microbiol 54:101–111 [View Article][PubMed]
    [Google Scholar]
  27. Merrigan M., Venugopal A., Mallozzi M., Roxas B., Viswanathan V. K., Johnson S., Gerding D. N., Vedantam G. 2010; Human hypervirulent Clostridium difficile strains exhibit increased sporulation as well as robust toxin production. J Bacteriol 192:4904–4911 [View Article][PubMed]
    [Google Scholar]
  28. Naaber P., Lehto E., Salminen S., Mikelsaar M. 1996; Inhibition of adhesion of Clostridium difficile to Caco-2 cells. FEMS Immunol Med Microbiol 14:205–209 [View Article][PubMed]
    [Google Scholar]
  29. Nicholson W. L., Setlow P. 1990; Sporulation, germination and outgrowth. In Molecular biological methods for Bacillus pp. 391–450 Edited by Harwood C. R., Cutting S. M. Chichester: John Wiley and Sons;
    [Google Scholar]
  30. Novak J. S., Juneja V. K., McClane B. A. 2003; An ultrastructural comparison of spores from various strains of Clostridium perfringens and correlations with heat resistance parameters. Int J Food Microbiol 86:239–247 [View Article][PubMed]
    [Google Scholar]
  31. O’Neill G. L., Beaman M. H., Riley T. V. 1991; Relapse versus reinfection with Clostridium difficile. Epidemiol Infect 107:627–635 [View Article][PubMed]
    [Google Scholar]
  32. Oka K., Osaki T., Hanawa T., Kurata S., Okazaki M., Manzoku T., Takahashi M., Tanaka M., Taguchi H. other authors 2012; Molecular and microbiological characterization of Clostridium difficile isolates from single, relapse, and reinfection cases. J Clin Microbiol 50:915–921 [View Article][PubMed]
    [Google Scholar]
  33. Oliva C. R., Swiecki M. K., Griguer C. E., Lisanby M. W., Bullard D. C., Turnbough C. L. Jr, Kearney J. F. 2008; The integrin Mac-1 (CR3) mediates internalization and directs Bacillus anthracis spores into professional phagocytes. Proc Natl Acad Sci U S A 105:1261–1266 [View Article][PubMed]
    [Google Scholar]
  34. Orsburn B., Melville S. B., Popham D. L. 2008; Factors contributing to heat resistance of Clostridium perfringens endospores. Appl Environ Microbiol 74:3328–3335 [View Article][PubMed]
    [Google Scholar]
  35. Paidhungat M., Ragkousi K., Setlow P. 2001; Genetic requirements for induction of germination of spores of Bacillus subtilis by Ca2+-dipicolinate. J Bacteriol 183:4886–4893 [View Article][PubMed]
    [Google Scholar]
  36. Paredes-Sabja D., Sarker M. R. 2009; Clostridium perfringens sporulation and its relevance to pathogenesis. Future Microbiol 4:519–525 [View Article][PubMed]
    [Google Scholar]
  37. Paredes-Sabja D., Sarker M. R. 2011; Germination response of spores of the pathogenic bacterium Clostridium perfringens and Clostridium difficile to cultured human epithelial cells. Anaerobe 17:78–84 [View Article][PubMed]
    [Google Scholar]
  38. Paredes-Sabja D., Setlow B., Setlow P., Sarker M. R. 2008; Characterization of Clostridium perfringens spores that lack SpoVA proteins and dipicolinic acid. J Bacteriol 190:4648–4659 [View Article][PubMed]
    [Google Scholar]
  39. Pepin J., Alary M. E., Valiquette L., Raiche E., Ruel J., Fulop K., Godin D., Bourassa C. 2005; Increasing risk of relapse after treatment of Clostridium difficile colitis in Quebec, Canada. Clin Infect Dis 40:1591–1597 [View Article][PubMed]
    [Google Scholar]
  40. Rosenberg M., Perry A., Bayer E. A., Gutnick D. L., Rosenberg E., Ofek I. 1981; Adherence of Acinetobacter calcoaceticus RAG-1 to human epithelial cells and to hexadecane. Infect Immun 33:29–33 [View Article]
    [Google Scholar]
  41. Schwan C., Stecher B., Tzivelekidis T., van Ham M., Rohde M., Hardt W. D., Wehland J., Aktories K. 2009; Clostridium difficile toxin CDT induces formation of microtubule-based protrusions and increases adherence of bacteria. PLoS Pathog 5:e1000626 [View Article][PubMed]
    [Google Scholar]
  42. Sirard S., Valiquette L., Fortier L. C. 2011; Lack of association between clinical outcome of Clostridium difficile infections, strain type, and virulence-associated phenotypes. J Clin Microbiol 49:4040–4046 [View Article][PubMed]
    [Google Scholar]
  43. Sorg J. A., Sonenshein A. L. 2008; Bile salts and glycine as cogerminants for Clostridium difficile spores. J Bacteriol 190:2505–2512 [View Article][PubMed]
    [Google Scholar]
  44. Tam N. K., Uyen N. Q., Hong H. A., Duc H., Hoa T. T., Serra C. R., Henriques A. O., Cutting S. M. 2006; The intestinal life cycle of Bacillus subtilis and close relatives. J Bacteriol 188:2692–2700 [View Article][PubMed]
    [Google Scholar]
  45. Tasteyre A., Barc M. C., Collignon A., Boureau H., Karjalainen T. 2001; Role of FliC and FliD flagellar proteins of Clostridium difficile in adherence and gut colonization. Infect Immun 69:7937–7940 [View Article][PubMed]
    [Google Scholar]
  46. Uzal F. A., McClane B. A. 2011; Recent progress in understanding the pathogenesis of Clostridium perfringens type C infections. Vet Microbiol 153:37–43 [View Article][PubMed]
    [Google Scholar]
  47. Viswanathan V. K., Mallozzi M. J., Vedantam G. 2010; Clostridium difficile infection: An overview of the disease and its pathogenesis, epidemiology and interventions. Gut Microbes 1:234–242 [View Article][PubMed]
    [Google Scholar]
  48. Voth D. E., Ballard J. D. 2005; Clostridium difficile toxins: mechanism of action and role in disease. Clin Microbiol Rev 18:247–263 [View Article][PubMed]
    [Google Scholar]
  49. Waligora A. J., Hennequin C., Mullany P., Bourlioux P., Collignon A., Karjalainen T. 2001; Characterization of a cell surface protein of Clostridium difficile with adhesive properties. Infect Immun 69:2144–2153 [View Article][PubMed]
    [Google Scholar]
  50. Waters M., Savoie A., Garmory H. S., Bueschel D., Popoff M. R., Songer J. G., Titball R. W., McClane B. A., Sarker M. R. 2003; Genotyping and phenotyping of beta2-toxigenic Clostridium perfringens fecal isolates associated with gastrointestinal diseases in piglets. J Clin Microbiol 41:3584–3591 [View Article][PubMed]
    [Google Scholar]
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