1887

Abstract

Purpose. Infective endocarditis is a severe and potentially fatal disease. Nearly a third of all cases remain culture-negative, making a targeted and effective antibiotic therapy of patients challenging. In the past years, fluorescence in situ hybridization (FISH) has proven its value for the diagnosis of infective endocarditis, particularly when it is caused by fastidious bacteria. To increase the number of infective endocarditis causing agents, which can be identified by FISH, we designed and optimized a FISH-probe for the specific detection of Coxiella burnetii in heart valve tissue.

Methodology. Even with specific probes the detection and identification of bacteria can be complicated by the high autofluorescence due to calcification of the analysed tissue. To overcome this problem, we developed a protocol to detect C. burnetii by hybridizing, stripping and reprobing the identical section with different species-specific probes repeatedly.

Results/Key findings. The newly designed specific FISH probe and the developed protocol exemplarily allowed us to unequivocally identify C. burnetii in tissue sections of a patient with infective endocarditis.

Conclusion. This method provides an add-on to existing protocols for the unambiguous diagnosis of bacteria directly within tissues or other difficult tissue samples in cases with small sample size and limited sections.

Loading

Article metrics loading...

/content/journal/jmm/10.1099/jmm.0.000704
2018-02-20
2019-10-20
Loading full text...

Full text loading...

/deliver/fulltext/jmm/67/4/537.html?itemId=/content/journal/jmm/10.1099/jmm.0.000704&mimeType=html&fmt=ahah

References

  1. Brouqui P, Raoult D. Endocarditis due to rare and fastidious bacteria. Clin Microbiol Rev 2001;14:177–207 [CrossRef][PubMed]
    [Google Scholar]
  2. Brouqui P, Raoult D. New insight into the diagnosis of fastidious bacterial endocarditis. FEMS Immunol Med Microbiol 2006;47:1–13 [CrossRef][PubMed]
    [Google Scholar]
  3. Geissdörfer W, Moos V, Moter A, Loddenkemper C, Jansen A et al. High frequency of Tropheryma whipplei in culture-negative endocarditis. J Clin Microbiol 2012;50:216–222 [CrossRef][PubMed]
    [Google Scholar]
  4. Mallmann C, Siemoneit S, Schmiedel D, Petrich A, Gescher DM et al. Fluorescence in situ hybridization to improve the diagnosis of endocarditis: a pilot study. Clin Microbiol Infect 2010;16:767–773 [CrossRef][PubMed]
    [Google Scholar]
  5. Gescher DM, Mallmann C, Kovacevic D, Schmiedel D, Borges AC et al. A view on Bartonella quintana endocarditis - confirming the molecular diagnosis by specific fluorescence in situ hybridization. Diagn Microbiol Infect Dis 2008;60:99–103 [CrossRef][PubMed]
    [Google Scholar]
  6. Frickmann H, Zautner AE, Moter A, Kikhney J, Hagen RM et al. Fluorescence in situ hybridization (FISH) in the microbiological diagnostic routine laboratory: a review. Crit Rev Microbiol 2017;43:263–293 [CrossRef][PubMed]
    [Google Scholar]
  7. Melenotte C, Million M, Audoly G, Gorse A, Dutronc H et al. B-cell non-Hodgkin lymphoma linked to Coxiella burnetii. Blood 2016;127:113121 [CrossRef][PubMed]
    [Google Scholar]
  8. Jensen TK, Montgomery DL, Jaeger PT, Lindhardt T, Agerholm JS et al. Application of fluorescent in situ hybridisation for demonstration of Coxiella burnetii in placentas from ruminant abortions. APMIS 2007;115:347–353 [CrossRef][PubMed]
    [Google Scholar]
  9. Daims H, Stoecker K, Wagner M. Fluorescence In Situ Hybridization for the Detection of Prokaryotes Abingdon, UK: Bios-Garland; 2005
    [Google Scholar]
  10. Quast C, Pruesse E, Yilmaz P, Gerken J, Schweer T et al. The SILVA ribosomal RNA gene database project: improved data processing and web-based tools. Nucleic Acids Res 2013;41:D590–D596 [CrossRef][PubMed]
    [Google Scholar]
  11. Amann RI, Binder BJ, Olson RJ, Chisholm SW, Devereux R et al. Combination of 16S rRNA-targeted oligonucleotide probes with flow cytometry for analyzing mixed microbial populations. Appl Environ Microbiol 1990;56:1919–1925[PubMed]
    [Google Scholar]
  12. Kumpf O, Dohmen P, Ertmer M, Knebel F, Wiessner A et al. Rapid molecular diagnosis of infective aortic valve endocarditis caused by Coxiella burnetii. Infection 2016;44:813–817 [CrossRef][PubMed]
    [Google Scholar]
  13. Ludwig W, Strunk O, Westram R, Richter L, Meier H et al. ARB: a software environment for sequence data. Nucleic Acids Res 2004;32:1363–1371 [CrossRef][PubMed]
    [Google Scholar]
  14. Fuchs BM, Glöckner FO, Wulf J, Amann R. Unlabeled helper oligonucleotides increase the in situ accessibility to 16S rRNA of fluorescently labeled oligonucleotide probes. Appl Environ Microbiol 2000;66:3603–3607 [CrossRef][PubMed]
    [Google Scholar]
  15. Daims H, Lücker S, Wagner M. daime, a novel image analysis program for microbial ecology and biofilm research. Environ Microbiol 2006;8:200–213 [CrossRef][PubMed]
    [Google Scholar]
  16. Moter A, Leist G, Rudolph R, Schrank K, Choi BK et al. Fluorescence in situ hybridization shows spatial distribution of as yet uncultured treponemes in biopsies from digital dermatitis lesions. Microbiology 1998;144:2459–2467 [CrossRef][PubMed]
    [Google Scholar]
  17. Wallner G, Amann R, Beisker W. Optimizing fluorescent in situ hybridization with rRNA-targeted oligonucleotide probes for flow cytometric identification of microorganisms. Cytometry 1993;14:136–143 [CrossRef][PubMed]
    [Google Scholar]
  18. Manz W, Amann R, Ludwig W, Wagner M, Schleifer K-H. Phylogenetic oligodeoxynucleotide probes for the major subclasses of proteobacteria: problems and solutions. Syst Appl Microbiol 1992;15:593–600 [CrossRef]
    [Google Scholar]
  19. Stoecker K, Dorninger C, Daims H, Wagner M. Double labeling of oligonucleotide probes for fluorescence in situ hybridization (DOPE-FISH) improves signal intensity and increases rRNA accessibility. Appl Environ Microbiol 2010;76:922–926 [CrossRef][PubMed]
    [Google Scholar]
  20. Hoshino T, Yilmaz LS, Noguera DR, Daims H, Wagner M. Quantification of target molecules needed to detect microorganisms by fluorescence in situ hybridization (FISH) and catalyzed reporter deposition-FISH. Appl Environ Microbiol 2008;74:5068–5077 [CrossRef][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jmm/10.1099/jmm.0.000704
Loading
/content/journal/jmm/10.1099/jmm.0.000704
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