1887

Abstract

Sepsis is a major cause of mortality in hospitalized patients worldwide, with lethality rates ranging from 30 to 70 %. Sepsis is caused by a variety of different pathogens, and rapid diagnosis is of outstanding importance, as early and adequate antimicrobial therapy correlates with positive clinical outcome. In recent years, matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) fingerprinting has become a powerful tool in microbiological diagnostics. The direct identification of micro-organisms in a positive blood culture by MALDI-TOF MS can shorten the diagnostic procedure significantly. Therefore, the aim of the present study was to evaluate whether identification rates could be improved by using the new Sepsityper kit from Bruker Daltonics for direct isolation and identification of bacteria from positive blood cultures by MALDI-TOF MS compared with the use of conventional separator gel columns, and to integrate the MALDI-TOF MS-based identification method into the routine course of blood culture diagnostics in the setting of a microbiological laboratory at a university hospital in Germany. The identification of Gram-negative bacteria by MALDI-TOF MS was significantly better using the Sepsityper kit compared with a separator gel tube-based method (99 and 68 % correct identification, respectively). For Gram-positive bacteria, only 73 % were correctly identified by MALDI-TOF with the Sepsityper kit and 59 % with the separator gel tube assay. A major problem of both methods was the poor identification of Gram-positive grape-like clustered cocci. As differentiation of from coagulase-negative staphylococci is of clinical importance, a PCR was additionally established that was capable of identifying directly from positive blood cultures, thus closing this diagnostic gap. Another benefit of the PCR approach is the possibility of directly detecting the genes responsible for meticillin resistance in staphylococci and for vancomycin resistance in enterococci, which is of high importance for early adequate treatment. Both of the described methods were finally integrated into a protocol for fast and effective identification of bacteria from positive blood cultures.

Loading

Article metrics loading...

/content/journal/jmm/10.1099/jmm.0.035550-0
2012-03-01
2020-07-04
Loading full text...

Full text loading...

/deliver/fulltext/jmm/61/3/323.html?itemId=/content/journal/jmm/10.1099/jmm.0.035550-0&mimeType=html&fmt=ahah

References

  1. Avery R., Kalaycio M., Pohlman B., Sobecks R., Kuczkowski E., Andresen S., Mossad S., Shamp J., Curtis J. other authors 2005; Early vancomycin-resistant enterococcus (VRE) bacteremia after allogeneic bone marrow transplantation is associated with a rapidly deteriorating clinical course. Bone Marrow Transplant 35:497–499 [CrossRef][PubMed]
    [Google Scholar]
  2. Bartz H., Mendoza Y., Gebker M., Fischborn T., Heeg K., Dalpke A. 2004; Poly-guanosine strings improve cellular uptake and stimulatory activity of phosphodiester CpG oligonucleotides in human leukocytes. Vaccine 23:148–155 [CrossRef][PubMed]
    [Google Scholar]
  3. Bennett K., Sharp S. E. 2008; Rapid differentiation of methicillin-resistant Staphylococcus aureus and methicillin-susceptible Staphylococcus aureus from blood cultures by use of a direct cefoxitin disk diffusion test. J Clin Microbiol 46:3836–3838 [CrossRef][PubMed]
    [Google Scholar]
  4. Bochud P. Y., Bonten M., Marchetti O., Calandra T. 2004; Antimicrobial therapy for patients with severe sepsis and septic shock: an evidence-based review. Crit Care Med 32:Suppl.S495–S512 [CrossRef][PubMed]
    [Google Scholar]
  5. Christner M., Rohde H., Wolters M., Sobottka I., Wegscheider K., Aepfelbacher M. 2010; Rapid identification of bacteria from positive blood culture bottles by use of matrix-assisted laser desorption-ionization time of flight mass spectrometry fingerprinting. J Clin Microbiol 48:1584–1591 [CrossRef][PubMed]
    [Google Scholar]
  6. Costa A. M., Kay I., Palladino S. 2005; Rapid detection of mecA and nuc genes in staphylococci by real-time multiplex polymerase chain reaction. Diagn Microbiol Infect Dis 51:13–17 [CrossRef][PubMed]
    [Google Scholar]
  7. Dark P. M., Dean P., Warhurst G. 2009; Bench-to-bedside review: the promise of rapid infection diagnosis during sepsis using polymerase chain reaction-based pathogen detection. Crit Care 13:217 [CrossRef][PubMed]
    [Google Scholar]
  8. Emonet S., Shah H. N., Cherkaoui A., Schrenzel J. 2010; Application and use of various mass spectrometry methods in clinical microbiology. Clin Microbiol Infect 16:1604–1613 [CrossRef][PubMed]
    [Google Scholar]
  9. Ferreira L., Sánchez-Juanes F., Porras-Guerra I., García-García M. I., García-Sánchez J. E., González-Buitrago J. M., Muñoz-Bellido J. L. 2011; Microorganisms direct identification from blood culture by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Clin Microbiol Infect 17:546–551 [CrossRef]
    [Google Scholar]
  10. Funke G., Funke-Kissling P. 2004; Use of the BD PHOENIX Automated Microbiology System for direct identification and susceptibility testing of Gram-negative rods from positive blood cultures in a three-phase trial. J Clin Microbiol 42:1466–1470 [CrossRef][PubMed]
    [Google Scholar]
  11. Garnacho-Montero J., Ortiz-Leyba C., Herrera-Melero I., Aldabó-Pallás T., Cayuela-Dominguez A., Marquez-Vacaro J. A., Carbajal-Guerrero J., Garcia-Garmendia J. L. 2008; Mortality and morbidity attributable to inadequate empirical antimicrobial therapy in patients admitted to the ICU with sepsis: a matched cohort study. J Antimicrob Chemother 61:436–441 [CrossRef][PubMed]
    [Google Scholar]
  12. Ibrahim E. H., Sherman G., Ward S., Fraser V. J., Kollef M. H. 2000; The influence of inadequate antimicrobial treatment of bloodstream infections on patient outcomes in the ICU setting. Chest 118:146–155 [CrossRef][PubMed]
    [Google Scholar]
  13. Katayama Y., Ito T., Hiramatsu K. 2000; A new class of genetic element, Staphylococcus cassette chromosome mec, encodes methicillin resistance in Staphylococcus aureus. Antimicrob Agents Chemother 44:1549–1555 [CrossRef][PubMed]
    [Google Scholar]
  14. Klotz M., Opper S., Heeg K., Zimmermann S. 2003; Detection of Staphylococcus aureus enterotoxins A to D by real-time fluorescence PCR assay. J Clin Microbiol 41:4683–4687 [CrossRef][PubMed]
    [Google Scholar]
  15. Kumar A., Roberts D., Wood K. E., Light B., Parrillo J. E., Sharma S., Suppes R., Feinstein D., Zanotti S. other authors 2006; Duration of hypotension before initiation of effective antimicrobial therapy is the critical determinant of survival in human septic shock. Crit Care Med 34:1589–1596 [CrossRef][PubMed]
    [Google Scholar]
  16. Kumar A., Ellis P., Arabi Y., Roberts D., Light B., Parrillo J. E., Dodek P., Wood G., Kumar A. other authors 2009; Initiation of inappropriate antimicrobial therapy results in a fivefold reduction of survival in human septic shock. Chest 136:1237–1248 [CrossRef][PubMed]
    [Google Scholar]
  17. McGregor K. F., Young H.-K. 2000; Identification and characterization of vanB2 glycopeptide resistance elements in enterococci isolated in Scotland. Antimicrob Agents Chemother 44:2341–2348 [CrossRef][PubMed]
    [Google Scholar]
  18. Mellmann A., Bimet F., Bizet C., Borovskaya A. D., Drake R. R., Eigner U., Fahr A. M., He Y., Ilina E. N. other authors 2009; High interlaboratory reproducibility of matrix-assisted laser desorption ionization-time of flight mass spectrometry-based species identification of nonfermenting bacteria. J Clin Microbiol 47:3732–3734 [CrossRef][PubMed]
    [Google Scholar]
  19. Moussaoui W., Jaulhac B., Hoffmann A. M., Ludes B., Kostrzewa M., Riegel P., Prévost G. 2010; Matrix-assisted laser desorption ionization time-of-flight mass spectrometry identifies 90% of bacteria directly from blood culture vials. Clin Microbiol Infect 16:1631–1638 [CrossRef][PubMed]
    [Google Scholar]
  20. Stevenson L. G., Drake S. K., Murray P. R. 2010; Rapid identification of bacteria in positive blood culture broths by matrix-assisted laser desorption ionization-time of flight mass spectrometry. J Clin Microbiol 48:444–447 [CrossRef][PubMed]
    [Google Scholar]
  21. Szabados F., Michels M., Kaase M., Gatermann S. 2011; The sensitivity of direct identification from positive BacT/ALERT (bioMérieux) blood culture bottles by matrix-assisted laser desorption ionization time-of-flight mass spectrometry is low. Clin Microbiol Infect 17:192–195 [CrossRef]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jmm/10.1099/jmm.0.035550-0
Loading
/content/journal/jmm/10.1099/jmm.0.035550-0
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error