1887

Abstract

and are among the leading causes of hospital-acquired infections. Reliable and quick identification of and is important for accurate treatment and understanding their role in the pathogenesis of infections. Fluorescent hybridization (FISH) of whole bacterial cells with oligonucleotides targeted at the 16S rRNA molecule leads to a reduced time to identification. In clinical practice, FISH therefore can be used in situations in which quick identification is necessary for optimal treatment of the patient. Furthermore, the abundance, spatial distribution and bacterial cell morphology can be observed . This report describes the design of two fluorescent-labelled oligonucleotides that, respectively, detect the 16S rRNA of and the 16S rRNA of , , , and . Different protocols for the application of these oligonucleotides with FISH in different clinical samples such as faeces or blood cultures are given. Enterococci in a biofilm attached to a biomaterial were also visualized. Embedding of the biomaterial preserved the morphology and therefore the architecture of the biofilm could be observed. The usefulness of other studies describing FISH for detection of enterococci is generally hampered by the fact that they have only focused on one material and one protocol to detect the enterococci. However, the results of this study show that the probes can be used both in the routine laboratory to detect and determine the enterococcal species in different clinical samples and in a research setting to enumerate and detect the enterococci in their physical environment.

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2005-10-01
2024-04-18
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References

  1. Amann R. I., Binder B. J., Olson R. J., Chisholm S. W., Devereux R., Stahl D. A. 1990; Combination of 16S rRNA-targeted oligonucleotide probes with flow cytometry for analyzing mixed microbial populations. Appl Environ Microbiol 56:1919–1925
    [Google Scholar]
  2. Amann R. I., Ludwig W., Schleifer K. H. 1995; Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol Rev 59:143–169
    [Google Scholar]
  3. Behr T., Koob C., Schedl M. & 8 other authors; 2000; A nested array of rRNA targeted probes for the detection and identification of enterococci by reverse hybridization. Syst Appl Microbiol 23:563–572 [CrossRef]
    [Google Scholar]
  4. Beimfohr C., Krause A., Amann R. I., Ludwig W., Schleifer K. H. 1993; In situ identification of lactococci, enterococci and streptococci. Syst Appl Microbiol 16:450–456 [CrossRef]
    [Google Scholar]
  5. Betzl D., Ludwig W., Schleifer K. H. 1990; Identification of lactococci and enterococci by colony hybridization with 23S rRNA-targeted oligonucleotide probes. Appl Environ Microbiol 56:2927–2929
    [Google Scholar]
  6. Costerton J. W., Stewart P. S., Greenberg E. P. 1999; Bacterial biofilms: a common cause of persistent infections. Science 284:1318–1322 [CrossRef]
    [Google Scholar]
  7. Facklam R. R., Collins M. D. 1989; Identification of Enterococcus species isolated from human infections by a conventional test scheme. J Clin Microbiol 27:731–734
    [Google Scholar]
  8. Facklam R. R., Sahm D. F., Teixeira L. M. 1999; Enterococcus . In Manual of Clinical Microbiology pp 297–305 Edited by Murray P. R. and others Washington DC: American Society for Microbiology;
    [Google Scholar]
  9. Franks A. H., Harmsen H. J., Raangs G. C., Jansen G. J., Schut F., Welling G. W. 1998; Variations of bacterial populations in human feces measured by fluorescent in situ hybridization with group-specific 16S rRNA-targeted oligonucleotide probes. Appl Environ Microbiol 64:3336–3345
    [Google Scholar]
  10. Gilad J., Borer A., Riesenberg K., Peled N., Shnaider A., Schlaeffer F. 1998; Enterococcus hirae septicemia in a patient with end-stage renal disease undergoing hemodialysis. Eur J Clin Microbiol Infect Dis 17:576–577 [CrossRef]
    [Google Scholar]
  11. Harmsen H. J. M., Elfferich P., Schut F., Welling G. W. 1999; A 16S rRNA-targeted probe for detection of lactobacilli and enterococci in faecal samples by fluorescent in situ hybridization. Microb Ecol Health Dis 11:3–12 [CrossRef]
    [Google Scholar]
  12. Holdeman L. V., Cato E. P., Moore W. E. C. 1977 Anaerobe Laboratory Manual Edited by Holdeman L. V. and others Blacksburg, VA: Virginia Polytechnic Institute and State University;
    [Google Scholar]
  13. Huycke M. M., Sahm D. F., Gilmore M. S. 1998; Multiple-drug resistant enterococci: the nature of the problem and an agenda for the future. Emerg Infect Dis 4:239–249 [CrossRef]
    [Google Scholar]
  14. Jansen G. J., Mooibroek M., Idema J., Harmsen H. J., Welling G. W., Degener J. E. 2000; Rapid identification of bacteria in blood cultures by using fluorescently labeled oligonucleotide probes. J Clin Microbiol 38:814–817
    [Google Scholar]
  15. Kaufhold A., Ferrieri P. 1991; Isolation of Enterococcus mundtii from normally sterile body sites in two patients. J Clin Microbiol 29:1075–1077
    [Google Scholar]
  16. Low D. E., Keller N., Barth A., Jones R. N. 2001; Clinical prevalence, antimicrobial susceptibility, and geographic resistance patterns of enterococci: results from the SENTRY antimicrobial surveillance program, 1997–1999. Clin Infect Dis 32:Suppl 2S133–S145
    [Google Scholar]
  17. Ludwig W., Strunk O., Klugbauer S., Klugbauer N., Weizenegger M., Neumaier J., Bachleitner M., Schleifer K. H. 1998; Bacterial phylogeny based on comparative sequence analysis. Electrophoresis 19:554–568 [CrossRef]
    [Google Scholar]
  18. Maidak B. L., Olsen G. J., Larsen N., Overbeek R., McCaughey M. J., Woese C. R. 1997; The RDP (Ribosomal Database Project). Nucleic Acids Res 25:109–111 [CrossRef]
    [Google Scholar]
  19. Monstein H. J., Quednau M., Samuelsson A., Ahrne S., Isaksson B., Jonasson J. 1998; Division of the genus Enterococcus into species groups using PCR-based molecular typing methods. Microbiology 144:1171–1179 [CrossRef]
    [Google Scholar]
  20. Munson E. L., Diekema D. J., Beekmann S. E., Chapin K. C., Doern G. V. 2003; Detection and treatment of bloodstream infection: laboratory reporting and antimicrobial management. J Clin Microbiol 41:495–497 [CrossRef]
    [Google Scholar]
  21. Murray B. E. 1997; Vancomycin-resistant enterococci. Am J Med 102:284–293 [CrossRef]
    [Google Scholar]
  22. Neut D., van Horn J. R., van Kooten T. G., van der Mei H. C., Busscher H. J. 2003; Detection of biomaterial-associated infections in orthopaedic joint implants. Clin Orthop 413:261–268 [CrossRef]
    [Google Scholar]
  23. Pascual A. 2002; Pathogenesis of catheter-related infections: lessons for new designs. Clin Microbiol Infect 8:256–264 [CrossRef]
    [Google Scholar]
  24. Patel R., Piper K. E., Rouse M. S., Steckelberg J. M., Uhl J. R., Kohner P., Hopkins M. K., Cockerill F. R. 3rd, Kline B. C. 1998; Determination of 16S rRNA sequences of enterococci and application to species identification of nonmotile Enterococcus gallinarum isolates. J Clin Microbiol 36:3399–3407
    [Google Scholar]
  25. Poulsen L. K., Licht T. R., Rang C., Krogfelt K. A., Molin S. 1995; Physiological state of Escherichia coli BJ4 growing in the large intestines of streptomycin-treated mice. J Bacteriol 177:5840–5845
    [Google Scholar]
  26. Pryce T. M., Wilson R. D., Kulski J. K. 1999; Identification of enterococci by ribotyping with horseradish-peroxidase-labelled 16S rDNA probes. J Microbiol Methods 36:147–155 [CrossRef]
    [Google Scholar]
  27. Ruoff K. L., de la Maza L., Murtagh M. J., Spargo J. D., Ferraro M. J. 1990; Species identities of enterococci isolated from clinical specimens. J Clin Microbiol 28:435–437
    [Google Scholar]
  28. Vancanneyt M., Snauwaert C., Cleenwerck I. & 8 other authors; 2001; Enterococcus villorum sp.nov., an enteroadherent bacterium associated with diarrhoea in piglets. Int J Syst Evol Microbiol 51:393–400
    [Google Scholar]
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