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High Resolution FESEM and TEM Reveal Bacterial Spore Attachment

Published online by Cambridge University Press:  16 July 2007

Barbara J. Panessa-Warren ,
George T. Tortora  and
John B. Warren
Barbara J. Panessa-Warren
Affiliation:
Department of Condensed Matter Physics and Materials Science, Building 480, Brookhaven National Laboratory, Upton, NY 11973, USA
George T. Tortora
Affiliation:
Clinical Microbiology Laboratory, University Hospital, SUNY Stony Brook, Stony Brook, NY 11794, USA School of Health Technology & Management, Stony Brook University, Stony Brook, NY 11794, USA
John B. Warren
Affiliation:
Division of Instrumentation, Building 535B, Brookhaven National Laboratory, Upton, NY 11973, USA
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Abstract

Transmission electron microscopy (TEM) studies in the 1960s and early 1970s using conventional thin section and freeze fracture methodologies revealed ultrastructural bacterial spore appendages. However, the limited technology at that time necessitated the time-consuming process of imaging serial sections and reconstructing each structure. Consequently, the distribution and function of these appendages and their possible role in colonization or pathogenesis remained unknown. By combining high resolution field emission electron microscopy with TEM images of identical bacterial spore preparations, we have been able to obtain images of intact and sectioned Bacillus and Clostridial spores to clearly visualize the appearance, distribution, resistance (to trypsin, chloramphenicol, and heat), and participation of these structures to facilitate attachment of the spores to glass, agar, and human cell substrates. Current user-friendly commercial field emission scanning electron microscopes (FESEMs), permit high resolution imaging, with high brightness guns at lower accelerating voltages for beam sensitive intact biological samples, providing surface images at TEM magnifications for making direct comparisons. For the first time, attachment structures used by pathogenic, environmental, and thermophile bacterial spores could be readily visualized on intact spores to reveal how specific appendages and outer spore coats participated in spore attachment, colonization, and invasion.

Keywords

field emission scanning electron microscopyClostridium and Bacillus sporesspore attachmenthuman cellstransmission electron microscopy
Type
BIOLOGICAL APPLICATIONS
Information
Microscopy and Microanalysis , Volume 13 , Issue 4 , August 2007 , pp. 251 - 266
Copyright
© 2007 Microscopy Society of America

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References

REFERENCES

Baird-Parker, A.C. (1969). Medical and veterinary significance of spore-forming bacteria and their spores. In The Bacterial Spore, Gould, G.W. & Hurst, A. (Eds.), pp. 517–541. New York: Academic Press.
Becker, M.L., Fagan, J.A., Gallant, N.D., Bauer, B.J., Bajpai, V., Hobbie, E.K., Lacerda, S.H., Migler, K.B. & Jakupciak, J.P. (2007). Length dependent uptake of DNA-wrapped single walled carbon nanotubes. Adv Mater 19, 939–945.Google Scholar
Bowen, W.R., Fenton, A.S., Lovitt, R.W. & Wright, C.J. (2001). The measurement of Bacillus mycoides spore adhesion using atomic force microscopy, simple counting methods and a spinning disk technique. Biotechnol & Bioengin 79, 171–179.Google Scholar
Bozzola, J. & Russell, L. (1992). Metal shadowing technique. In Electron Microscopy, Principles and Techniques for Biologists. p. 120. Boston, MA: Jones and Bartlett Publishers.
Davis, B.D. (1990). Bacterial architecture. In Microbiology, Davis, B., Dulbecco, R., Eisen, H. & Ginsberg, H. (Eds.), p. 47. Philadelphia, PA: J.B. Lippincott.
Driks, A. (1999). Bacillus subtilis spore coat. Microbiol Mol Biol Rev 63, 1–20.Google Scholar
Eveillard, M.V., Fourel, V., Barc, M., Kerneis, S., Colonnier, M., Karjalainen, T., Bourlioux, P. & Servin, A. (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.Google Scholar
Finlay, B. & Cossart, P. (1997). Exploitation of mammalian host cell functions by bacterial pathogens. Science 276, 718–725.Google Scholar
Finlay, B. & Falkow, S. (1989). Common themes in microbial pathogenicity. Microbiol Rev 53, 210–220.Google Scholar
Gerhardt, P. & Ribi, E. (1964). Ultrastructure of the exosporium enveloping spores of Bacillus cereus. J Bacteriol 88, 1774–1789.Google Scholar
Ginocchio, C.C. & Galan, J.E. (1995). Functional conservation among members of the Salmonella typhimurium InvA family of proteins. Infect Immun 63, 729–732.Google Scholar
Gould, G.W. (1969). Germination. In The Bacterial Spore, Gould, G.W. & Hurst, A. (Eds.), pp. 397–444. New York: Academic Press.
Gould, G.W. (1977). Recent advances in understanding of resistance and dormancy in bacterial spores. J Appl Bacteriol 42, 297–309.Google Scholar
Guidi-Rontani, C. (2002). The alveolar macrophage: The Trojan horse of Bacillus anthracis. Trends Microbiol 10, 405–409.Google Scholar
Guidi-Rontani, C., Pereira, Y., Ruffie, S., Sirard, J.C., Weber-Levy, M. & Mock, M. (1999). Identification and characterization of a germination operon on the virulence of plasmid pXO1 of Bacillus anthracis. Mol Microbiol 33, 407–414.Google Scholar
Hachisuka, Y., Kojima, K. & Sato, T. (1966). Fine filaments on the outside of the exosporium of Bacillus anthracis spores. J Bacteriol 91, 2382–2384.Google Scholar
Helgason, E., Okstad, O., Caugant, D., Johansen, H., Fouet, A., Mock, M., Hegna, I. & Kolsto, A. (2000). Bacillus anthracis, Bacillus cereus and Bacillus thurengiensis—One species on the basis of genetic evidence. J Appl Environ Microbiol 66, 2627–2630.Google Scholar
Hoeniger, J. & Headley, C. (1969). Ultrastructural aspects of spore germination and outgrowth in C. sporogenes. Can J Microbiol 15, 1061–1066.Google Scholar
Keynan, A. & Evenchik, Z. (1969). Activation. In The Bacterial Spore, Gould, G.W. & Hurst, A. (Eds.), pp. 359–396. New York: Academic Press.
Luft, J.H. (1971). Ruthenium red and violet. II. Fine structural localization in animal tissues. Anat Rec 171, 369–416.Google Scholar
Olsen, I., Johnson, J., Moore, L. & Moore, W. (1995). Rejection of Clostridium putrificum and conservation of Clostridium botulinum and Clostridium sporogenes. Int J Syst Bacteriol 45, 414.Google Scholar
Panessa-Warren, B., Tortora, G. & Warren, J. (1994). Electron microscopy of C. sporogenes endospore attachment and germination. Scanning 16, 227–240.Google Scholar
Panessa-Warren, B., Tortora, G. & Warren, J. (1997). Exosporial membrane plasticity of Clostridium sporogenes and Clostridium difficile. Tissue Cell 29, 449–461.Google Scholar
Panessa-Warren, B., Tortora, G., Wong, S.S., Ghebrehiwet, B. & Warren, J. (2003). Bacillus and Clostridium spore attachment/entry of human colon cells. Microsc Microanal 9(Suppl. 2), 1378–1379.Google Scholar
Pope, L., Yolton, D. & Rode, L. (1967). Appendages of Clostridium bifermentans spores. J Bacteriol 94, 1206–1215.Google Scholar
Ross, J.M. (1957). The pathogenesis of anthrax following administration of spores by the respiratory route. J Path Bacteriol 73, 485–494.Google Scholar
Roth, I.L. & Williams, R.P. (1963). Comparison of the fine structure of virulent and avirulent spores of Bacillus anthracis. Tex Rep Biol Med 21, 394–399.Google Scholar
Russell, A. (1990). Bacterial spores and chemical sporocidal agents. Clin Micrbiol Rev 3, 99–119.Google Scholar
Sadoff, H.L. (1970). Heat resistance of spore enzymes. J Appl Bacteriol 33, 130–140.Google Scholar
Samsonoff, W., Hashimoto, T. & Conti, S. (1970). Ultrastructural changes associated with germination and outgrowth of an appendage bearing Clostridial spore. J Bacteriol 101, 1038–1045.Google Scholar
Schaeffer, A. & Fulton, D. (1933). A simplified method for staining spores. Science 77, 194–195.Google Scholar
Shvedova, A., Kisin, E., Mercer, R., Murray, A., Johnson, V., Potapovich, A., Tyurina, Y., Gorelik, O., Arepalli, S., Schwegler-Berry, D., Hubbs, A., Antonini, J., Evans, D., Ku, B.-K., Ramsey, D., Maynard, A., Kagan, V., Castranova, V. & Baron, P. (2005). Unusual inflammatory and fibrogenic pulmonary responses to single-walled carbon nanotubes in mice. Am J Physiol Lung Cell Mol Physiol 289, 698–708.Google Scholar
Strange, R.E. & Hunter, J.R. (1969). Outgrowth and the synthesis of macromolecules. In The Bacterial Spore, Gould, G.W. & Hurst, A. (Eds.), pp. 445–483. New York: Academic Press.
Sylvestre, P., Couture-Tosi, E. & Mock, M. (2002). A collagen-like surface glycoprotein is a structural component of the Bacillus anthracis exosporium. Mol Microbiol 45, 169–178.Google Scholar
Tipper, D. & Gauthier, T. (1972). Structure of the bacterial spore. In Spores V, Halvorson, H., Hanson, R. & Campbell, L. (Eds.), pp. 3–12. Washington, DC: American Society for Microbiology.
Walker, P., Short, J., Roper, G. & Hodgkiss, W. (1976). The structure of clostridial spore. In Microbial Ultrastructure, Fuller, R. & Lovelock, D. (Eds.), pp. 117–123. London: Academic Press.
Yolton, D., Pope, L., Williams, M. & Rode, L. (1968). Electron microscope characterization of Clostridium bifermentans. J Bacteriol 95, 231–238.Google Scholar