[go: up one dir, main page]
More Web Proxy on the site http://driver.im/
Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Protocol
  • Published:

Phylogenetic classification and identification of bacteria by mass spectrometry

Nature Protocols volume 4, pages 732–742 (2009)Cite this article

Abstract

Bacteria are a convenient source of intrinsic marker proteins, which can be detected efficiently by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The patterns of protein masses observed can be used for accurate classification and identification of bacteria. Key to the reliability of the method is a robust and standardized procedure for sample preparations, including bacterial culturing, chemical treatment for bacterial cell wall disruption and for protein extraction, and mass spectrometry analysis. The protocol is an excellent alternative to classical microbiological classification and identification procedures, requiring minimal sample preparation efforts and costs. Without cell culturing, the protocol takes in general <1 h.

Access through your institution
Buy or subscribe

This is a preview of subscription content, access via your institution

Access options

Access through your institution

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: A general overview of the procedure.
Figure 2: A mass spectrum of an E. coli sample.
Figure 3: Phylogenetic classification of a number of bacteria.

Similar content being viewed by others

References

  1. Sintchenko, V., Iredell, J.R. & Gilbert, G.L. Pathogen profiling for disease management and surveillance. Nat. Rev. Microbiol. 5, 464–470 (2007).

    Article  CAS  Google Scholar 

  2. Sauer, S. et al. Miniaturization in functional genomics and proteomics. Nat. Rev. Genet. 6, 465–476 (2005).

    Article  CAS  Google Scholar 

  3. Aebersold, R. & Mann, M. Mass spectrometry-based proteomics. Nature 422, 198–207 (2003).

    Article  CAS  Google Scholar 

  4. Claydon, M.A., Davey, S.N., Edwards-Jones, V. & Gordon, D.B. The rapid identification of intact microorganisms using mass spectrometry. Nat. Biotechnol. 14, 1584–1586 (1996).

    Article  CAS  Google Scholar 

  5. Holland, R.D. et al. Rapid identification of intact whole bacteria based on spectral patterns using matrix-assisted laser desorption/ionization with time-of-flight mass spectrometry. Rapid Commun. Mass Spectrom. 10, 1227–1232 (1996).

    Article  CAS  Google Scholar 

  6. Chong, B.E., Wall, D.B., Lubman, D.M. & Flynn, S.J. Rapid profiling of E. coli proteins up to 500 kDa from whole cell lysates using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Rapid Commun. Mass Spectrom. 11, 1900–1908 (1997).

    Article  CAS  Google Scholar 

  7. Williams, T.L., Andrzejewski, D., Lay, J.O. & Musser, S.M. Experimental factors affecting the quality and reproducibility of MALDI-TOF mass spectra obtained from whole bacteria cells. J. Am. Soc. Mass Spectrom. 14, 342–351 (2003).

    Article  CAS  Google Scholar 

  8. Lay, J.O. Jr. MALDI-TOF mass spectrometry of bacteria. Mass Spectrom. Rev. 20, 172–194 (2001).

    Article  CAS  Google Scholar 

  9. Baumann, S. et al. Standardized approach to proteome profiling of human serum based on magnetic bead separation and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Clin. Chem. 51, 973–980 (2005).

    Article  CAS  Google Scholar 

  10. Tiss, A. et al. Serum peptide profiling using MALDI mass spectrometry: avoiding the pitfalls of coated magnetic beads using well-established ZipTip technology. Proteomics 7 (Suppl 1): 77–89 (2007).

    Article  Google Scholar 

  11. Ryzhov, V. & Fenselau, C. Characterization of the protein subset desorbed by MALDI from whole bacterial cells. Anal. Chem. 73, 746–750 (2001).

    Article  CAS  Google Scholar 

  12. Hsieh, S.Y. et al. Highly efficient classification and identification of human pathogenic bacteria by MALDI-TOF MS. Mol. Cell. Proteomics 7, 448–456 (2008).

    Article  CAS  Google Scholar 

  13. Mead, P.S., Slutsker, L., Griffin, P.M. & Tauxe, R.V. Food-related illness and death in the United States reply to Dr. Hedberg. Emerg. Infect. Dis. 5, 841–842 (1999).

    Article  Google Scholar 

  14. Sauer, S. et al. Classification and identification of bacteria by mass spectrometry and computational analysis. PLoS ONE 3, e2843 (2008).

    Article  Google Scholar 

  15. Barbuddhe, S.B. et al. Rapid identification and typing of listeria species by matrix-assisted laser desorption ionization-time of flight mass spectrometry. Appl. Environ. Microbiol. 74, 5402–5407 (2008).

    Article  CAS  Google Scholar 

  16. Mellmann, A. et al. Evaluation of matrix-assisted laser desorption ionization-time-of-flight mass spectrometry in comparison to 16S rRNA gene sequencing for species identification of nonfermenting bacteria. J. Clin. Microbiol. 46, 1946–1954 (2008).

    Article  CAS  Google Scholar 

  17. Lasch, P. et al. MALDI-TOF mass spectrometry compatible inactivation method for highly pathogenic microbial cells and spores. Anal. Chem. 80, 2026–2034 (2008).

    Article  CAS  Google Scholar 

  18. Sauer, S. Matrix-assisted laser desorption/ionization mass spectrometry: principles and applications in life sciences. in Lasers in Chemistry, Vol. 1, 593–616 (Wiley-VCH, Weinheim, Germany, 2008).

    Google Scholar 

  19. Sauer, S., Reinhardt, R., Lehrach, H. & Gut, I.G. Single-nucleotide polymorphisms: analysis by mass spectrometry. Nat. Protoc. 1, 1761–1771 (2006).

    Article  CAS  Google Scholar 

  20. Fenselau, C. & Demirev, P.A. Characterization of intact microorganisms by MALDI mass spectrometry. Mass Spectrom. Rev. 20, 157–171 (2001).

    Article  CAS  Google Scholar 

  21. Villmann, T., Schleif, F.M., Kostrzewa, M., Walch, A. & Hammer, B. Classification of mass-spectrometric data in clinical proteomics using learning vector quantization methods. Brief. Bioinform. 9, 129–143 (2008).

    Article  CAS  Google Scholar 

  22. Keys, C.J. et al. Compilation of a MALDI-TOF mass spectral database for the rapid screening and characterisation of bacteria implicated in human infectious diseases. Infect. Genet. Evol. 4, 221–242 (2004).

    Article  CAS  Google Scholar 

  23. Teramoto, K. et al. Phylogenetic classification of Pseudomonas putida strains by MALDI-MS using ribosomal subunit proteins as biomarkers. Anal. Chem. 79, 8712–8719 (2007).

    Article  CAS  Google Scholar 

  24. Hettick, J.M. et al. Discrimination of intact mycobacteria at the strain level: a combined MALDI-TOF MS and biostatistical analysis. Proteomics 6, 6416–6425 (2006).

    Article  CAS  Google Scholar 

  25. Arnold, R.J. & Reilly, J.P. Fingerprint matching of E. coli strains with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry of whole cells using a modified correlation approach. Rapid Commun. Mass Spectrom. 12, 630–636 (1998).

    Article  CAS  Google Scholar 

  26. Bright, J.J., Claydon, M.A., Soufian, M. & Gordon, D.B. Rapid typing of bacteria using matrix-assisted laser desorption ionisation time-of-flight mass spectrometry and pattern recognition software. J. Microbiol. Methods 48, 127–138 (2002).

    Article  CAS  Google Scholar 

  27. Carbonnelle, E. et al. Rapid identification of Staphylococci isolated in clinical microbiology laboratories by matrix-assisted laser desorption ionization-time of flight mass spectrometry. J. Clin. Microbiol. 45, 2156–2161 (2007).

    Article  CAS  Google Scholar 

  28. Chen, P., Lu, Y. & Harrington, P.B. Application of linear and nonlinear discrete wavelet transforms to MALDI-MS measurements of bacteria for classification. Anal. Chem. 80, 7218–7225 (2008).

    Article  CAS  Google Scholar 

  29. Chen, P., Lu, Y. & Harrington, P.B. Biomarker profiling and reproducibility study of MALDI-MS measurements of Escherichia coli by analysis of variance-principal component analysis. Anal. Chem. 80, 1474–1481 (2008).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank Drs Markus Kostrzewa and Thomas Maier (Bruker Daltonics), Dr Michael Kube (MPI for Molecular Genetics) and Prof. Klaus Geider (JKI Dossenheim) for valuable help in setting up the system described in this protocol, fruitful scientific discussions and the provided test samples. We also thank Dr Chung-Ting Han (MPI for Molecular Genetics) and Drs Kostrzewa and Maier for critical reading of the paper. This work was supported by the German Ministry for Education and Research (BMBF, Grant no. 0315082), the National Genome Research Net (NGFN, Grant no. 01 GS 0828), the European Union (FP7/2007-2013, under grant agreement no. HEALTH-F4-2008-201418 entitled READNA) and the Max Planck Society.

Author information

Authors and Affiliations

  1. Max Planck Institute for Molecular Genetics, Otto-Warburg-Laboratory, Berlin, Germany

    Anja Freiwald & Sascha Sauer

Authors
  1. Anja Freiwald
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Sascha Sauer
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Sascha Sauer.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Freiwald, A., Sauer, S. Phylogenetic classification and identification of bacteria by mass spectrometry. Nat Protoc 4, 732–742 (2009). https://doi.org/10.1038/nprot.2009.37

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nprot.2009.37

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing