Molecular Genotyping of Microbes by Multilocus PCR and Mass Spectrometry: A New Tool for Hospital Infection Control and Public Health Surveillance

  • David J. Ecker
  • Christian Massire
  • Lawrence B. Blyn
  • Steven A. Hofstadler
  • James C. Hannis
  • Mark W. Eshoo
  • Thomas A. Hall
  • Rangarajan Sampath
Protocol
Part of the Methods in Molecular Biology™ book series (MIMB, volume 551)

Abstract

We describe a new technology for the molecular genotyping of microbes using a platform known commercially as the Ibis T5000. The technology couples multilocus polymerase chain reaction (PCR) to electrospray ionization/mass spectrometry (PCR/ESI-MS) and was developed to provide rapid, high-throughput, and precise digital analysis of either isolated colonies or original patient specimens on a platform suitable for use in hospital or reference diagnostic laboratories or public health settings. The PCR/ESI-MS method measures digital molecular signatures from microbes, enabling real-time epidemiological surveillance and outbreak investigation. This technology will facilitate understanding of the pathways by which infectious organisms spread and will enable appropriate interventions on a time frame not previously achievable.

Key words

Molecular genotyping mass spectrometry PCR/ESI-MS base composition Ibis T5000 

References

  1. 1.
    Ecker, D. J., Drader, J., Gutierrez, J., Gutierrez, A., Hannis, J., Schink, A., et al (2006). The Ibis T5000 universal biosensor—an automated platform for pathogen identification and strain typing. J. Assoc. Lab. Automat. 11, 341–351.CrossRefGoogle Scholar
  2. 2.
    Ecker, D. J., Sampath, R., Blyn, L. B., Eshoo, M. W., Ivy, C., Ecker, J. A., et al (2005). Rapid identification and strain-typing of respiratory pathogens for epidemic surveillance. Proc. Natl. Acad. Sci. U S A. 102, 8012–8017.PubMedCrossRefGoogle Scholar
  3. 3.
    Hofstadler, S. A., Sampath, R., Blyn, L. B., Eshoo, M. W., Hall, T. A., Jiang, Y., et al (2005). TIGER: the universal biosensor. Int. J. Mass. Spectrom. 242, 23–41.CrossRefGoogle Scholar
  4. 4.
    Sampath, R., Hofstadler, S. A., Blyn, L., Eshoo, M. W., Hall, T. A., Massire, C., et al (2005). Rapid identification of emerging pathogens: coronavirus. Emerg. Infect. Dis. 11, 373–379.PubMedCrossRefGoogle Scholar
  5. 5.
    Sampath, R., Russell, K. L., Massire, C., Eshoo, M. W., Harpin, V., Blyn, L. B., et al (2007). Global surveillance of emerging influenza virus genotypes by mass spectrometry. PLoS ONE 2, e489.PubMedCrossRefGoogle Scholar
  6. 6.
    Blyn, L. B., Hall, T. A., Libby, B., Ranken, R., Sampath, R., Rudnick, K., et al (2008). Rapid detection and molecular serotyping of adenovirus by the use of PCR followed by electrospray ionization mass spectrometry. J. Clin. Microbiol. 46, 644–651.PubMedCrossRefGoogle Scholar
  7. 7.
    Eshoo, M. W., Whitehouse, C. A., Zoll, S. T., Massire, C., Pennella, T. T., Blyn, L. B., et al (2007). Direct broad-range detection of alpha­viruses in mosquito extracts. Virology 368, 286–295.PubMedCrossRefGoogle Scholar
  8. 8.
    Urwin, R., and Maiden, M. C. J. (2003). Multi-locus sequence typing: a tool for global epidemiology. Trends Microbiol. 11, 479–487.PubMedCrossRefGoogle Scholar
  9. 9.
    Farlow, J., Smith, K. L., Wong, J., Abrams, M., Lytle, M., and Keim, P. (2001). Francisella tularensis strain typing using multiple-locus, variable-number tandem repeat analysis. J. Clin. Microbiol. 39, 3186–3192.PubMedCrossRefGoogle Scholar
  10. 10.
    Van Ert, M. N., Easterday, W. R., Huynh, L. Y., Okinaka, R. T., Hugh-Jones, M. E., Ravel, J., et al (2007). Global genetic population structure of Bacillus anthracis. PLoS ONE 2, e461.PubMedCrossRefGoogle Scholar
  11. 11.
    Skuce, R. A., McCorry, T. P., McCarroll, J. F., Roring, S. M., Scott, A. N., Brittain, D., et al (2002). Discrimination of Mycobacterium tuberculosis complex bacteria using novel VNTR-PCR targets. Microbiology 148, 519–528.PubMedGoogle Scholar
  12. 12.
    Enright, M. C., Spratt, B. G., Kalia, A., Cross, J. H., and Bessen, D. E. (2001). Multilocus sequence typing of Streptococcus pyogenes and the relationships between emm type and clone. Infect. Immun. 69, 2416–2427.PubMedCrossRefGoogle Scholar
  13. 13.
    Crum, N. F., Hale, B. R., Bradshaw, D. A., Malone, J. D., Chun, H. M., Gill, W. M., et al (2003). Outbreak of group A streptococcal pneumonia among marine corps recruits—California, November 1–December 20, 2002. MMWR Morb. Mortal. Wkly Rep. 52, 106–109.Google Scholar
  14. 14.
    Tong, M. J. (1972). Septic complications of war wounds. J. Am. Med. Assoc. 219, 1044–1047.CrossRefGoogle Scholar
  15. 15.
    Ecker, J. A., Massire, C., Hall, T. A., Ranken, R., Pennella, T. T., Agasino Ivy, C., et al (2006). Identification of Acinetobacter species and genotyping of Acinetobacter baumannii by multilocus PCR and mass spectrometry. J. Clin. Microbiol. 44, 2921–2932.PubMedCrossRefGoogle Scholar
  16. 16.
    van Dessel, H., Dijkshoorn, L., van der Reijden, T., Bakker, N., Paauw, A., van den Broek, P., et al (2004). Identification of a new geographically widespread multiresistant Acinetobacter baumannii clone from European hospitals. Res. Microbiol. 155, 105–112.PubMedCrossRefGoogle Scholar
  17. 17.
    Dijkshoorn, L., Aucken, H., Gerner-Smidt, P., Janssen, P., Kaufmann, M. E., Garaizer, J., et al (1996). Comparison of outbreak and nonoutbreak Acinetobacter baumannii strains by genotypic and phenotypic methods. J. Clin. Microbiol. 34, 1519–1525.PubMedGoogle Scholar
  18. 18.
    Wortmann, G., Weintrob, A., Barber, M., Scott, P., Zoll, S. T., Eshoo, M. W., et al (2008). Genotypic evolution of Acinetobacter baumannii strains in an outbreak associated with war trauma. Infect. Control Hosp. ­Epidemiol. 29, 553–555.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • David J. Ecker
    • 1
  • Christian Massire
    • 1
  • Lawrence B. Blyn
    • 1
  • Steven A. Hofstadler
    • 1
  • James C. Hannis
    • 1
  • Mark W. Eshoo
    • 1
  • Thomas A. Hall
    • 1
  • Rangarajan Sampath
    • 1
  1. 1.Ibis BiosciencesCarlsbadUSA

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