Almeida, Carina

Azevedo, N. F.

Gomes, C.

Jardim, T.

Keevil, C. W.

Melo, A.

Vieira, M. J.

Universidade do Minho: RepositoriUM

865 DETECTION OF STAPHYLOCOCCUS EPIDERMIDIS ON BIOFILMS BY FLUORESCENCE IN SITU HYBRIDIZATION (FISH) USING A NOVEL PEPTIDE NUCLEIC ACID (PNA) PROBE N.F. Azevedo1, T. Jardim1, C. Almeida1, A. Melo2, C. Gomes2, C.W. Keevil3, M.J. Vieira1 1IBB - Institute for Biotechnology and Bioengineering, Centre of Biological Engineering, Universidade do Minho, Braga, Portugal, 2Hospital da Santa Casa da Misericórdia de Vila Verde, Serviço de Anestesiologia, Vila Verde, Portugal, 3Environmental Healthcare Unit, School of Biological Sciences, University of Southampton, Southampton, United Kingdom Background: Contamination of catheters by Staphyloccus epidermidis might cause life-threatening complications such as septicaemia. Here, we report the development and application of a novel peptide nucleic acid (PNA) probe for the specific detection of this bacterium by fluorescence in situ hybridization (FISH).  Methods: The probe was shown to be highly specific and sensitive, as it was able to detect all S. epidermidis strains tested but not closely related species such as Staphyloccus aureus. The PNA FISH technique was adapted for the pathogen detection in catheters, and applied to screen 52 indwelling catheters that had been inserted in patients. Neither of these patients exhibited complications associated with infections derived from the insertion of the catheter.  Results: Most catheters had levels of 103-105 total cells/cm2 (as assessed by DAPI), and in a small percentage of them, the presence of S. epidermidis was detected by PNA FISH. However, no slime matrix enclosing the cells was observed and detection by standard culture methods was negative for most catheters (84.6%).  Conclusions: This work provides a rapid molecular biology method to assess infection of S. epidermidis in catheters and confirms the presence (albeit in low numbers) of this bacterium in asymptomatic patients that were tested negative by cultivation techniques. As such, PNA FISH will not indicate a potentially hazardous contamination of S. epidermidis simply based on the presence/absence of the bacterium. Further studies will attempt to correlate infection with the numbers of S. epidermidis cells detected by PNA FISH or the presence of a biofilm matrix. 

Detection of staphylococcus epidermidis on biofilms by fluorescence in situ hybridization (FISH) using a novel peptide nucleic acid (PNA) probe

http://repositorium.sdum.uminho.pt/bitstream/1822/8644/1/865.pdf

Detection of staphylococcus epidermidis on biofilms by fluorescence in situ hybridization (FISH) using a novel peptide nucleic acid (PNA) probe

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Universidade do Minho: RepositoriUM