Fluorescence and Electron Microscopy Methods for Exploring Antimicrobial Peptides Mode(s) of Action

  • Ludovica Marcellini
  • Maria Giammatteo
  • Pierpaolo Aimola
  • Maria Luisa Mangoni
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 618)

Abstract

Due to the increasing resistance of microbial pathogens to the available drugs, the identification of new antimicrobial agents with a new mechanism of action is urgently needed. In this context, cationic antimicrobial peptides (AMPs) are considered promising candidates. Although there is evidence that, in contrast to conventional antibiotics, microbial membranes are the principal target of a large number of AMPs, thus making it difficult for the pathogen to acquire resistance, their mode(s) of action is not yet completely clear. Intense research is currently devoted to understand the effect(s) of AMPs on intact cells, either at sub-lethal or at lethal peptide concentrations, and fluorescence/electron microscopy techniques represent a valid tool to get insight into the damage caused by these molecules on the morphology and membrane structure of the target cell. We here present an overview of some microscopic methodologies to address this issue.

Key words

Antimicrobial peptides mode(s) of action fluorescence microscopy scanning electron microscopy transmission electron microscopy membrane permeation 

Notes

Acknowledgments

We thank Prof. Donatella Barra for careful reading the manuscript. Work reported here has been supported in part by grants from the Italian Ministry of Education, University and Research and from the Università di Roma “La Sapienza”.

References

  1. 1.
    Boman, H. G. (1995) Peptide antibiotics and their role in innate immunity. Annu. Rev. Immunol. 13, 61–92.PubMedCrossRefGoogle Scholar
  2. 2.
    Hancock, R. E. and Sahl, H. G. (2006) Antimicrobial and host-defense peptides as new anti-infective therapeutic strategies. Nat. Biotechnol. 24, 1551–1557.PubMedCrossRefGoogle Scholar
  3. 3.
    Zasloff, M. (2002) Antimicrobial peptides of multicellular organisms. Nature 415, 389–395.PubMedCrossRefGoogle Scholar
  4. 4.
    Jenssen, H., Hamill, P., and Hancock, R. E. (2006) Peptide antimicrobial agents. Clin. Microbiol. Rev. 19, 491–511.PubMedCrossRefGoogle Scholar
  5. 5.
    Nicolas, P. and Mor, A. (1995) Peptides as weapons against microorganisms in the chemical defense system of vertebrates. Annu. Rev. Microbiol. 49, 277–304.PubMedCrossRefGoogle Scholar
  6. 6.
    Mookherjee, N. and Hancock, R. E. (2007) Cationic host defence peptides: innate immune regulatory peptides as a novel approach for treating infections. Cell. Mol. Life Sci. 64, 922–933.PubMedCrossRefGoogle Scholar
  7. 7.
    Mangoni, M. L., Papo, N., Barra, D., Simmaco, M., Bozzi, A., Di Giulio, A., and Rinaldi, A. C. (2004) Effects of the antimicrobial peptide temporin L on cell morphology, membrane permeability and viability of Escherichia coli. Biochem. J. 380, 859–865.PubMedCrossRefGoogle Scholar
  8. 8.
    Rodriguez, G. G., Phipps, D., Ishiguro, K., and Ridgway, H. F. (1992) Use of a fluorescent redox probe for direct visualization of actively respiring bacteria. Appl. Environ. Microbiol. 58, 1801–1808.PubMedGoogle Scholar
  9. 9.
    Severin, E., Stellmach, J., and Nachtigall, H. M. (1985) Fluorimetric assay of redox activity in cells. Anal. Chim. Acta 170, 341–346.CrossRefGoogle Scholar
  10. 10.
    Nation, J. L. (1983) A new method using hexamethyldisilazane for preparation of soft insect tissues for scanning electron microscopy. Stain Technol. 58, 347–351.PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Ludovica Marcellini
    • 1
  • Maria Giammatteo
    • 2
  • Pierpaolo Aimola
    • 3
  • Maria Luisa Mangoni
    • 4
  1. 1.Istituto Pasteur-Fondazione Cenci Bolognetti, Dipartimento di Scienze BiochimicheUniversità “La Sapienza”RomaItaly
  2. 2.Centro di Microscopie, Università di L’AquilaL’AquilaItaly
  3. 3.Dipartimento di Biologia di Base ed ApplicataUniversità di L’AquilaL’AquilaItaly
  4. 4.Istituto Pasteur-Fondazione Cenci Bolognetti, Dipartimento di Scienze Biochimiche, Azienda Ospedaliera S. AndreaUniversità “La Sapienza”RomaItaly

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