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Journal of Clinical Microbiology, August 2008, p. 2717-2722, Vol. 46, No. 8
0095-1137/08/$08.00+0     doi:10.1128/JCM.00501-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Distribution, Organization, and Ecology of Bacteria in Chronic Wounds{triangledown}

Klaus Kirketerp-Møller,1 Peter Ø. Jensen,2 Mustafa Fazli,3 Kit G. Madsen,4 Jette Pedersen,5 Claus Moser,2 Tim Tolker-Nielsen,3 Niels Høiby,2 Michael Givskov,3 and Thomas Bjarnsholt3*

Copenhagen Wound Healing Center, Bispebjerg Hospital, Copenhagen, Denmark,1 Department of Clinical Microbiology, Rigshospitalet, Copenhagen, Denmark,2 BioScience and Technology, BioCentrum-DTU, Technical University of Denmark, Lyngby, Denmark,3 AdvanDx A/S, Vedbaek, Denmark,4 Bartholin Institute, Copenhagen, Denmark5

Received 13 March 2008/ Returned for modification 27 April 2008/ Accepted 19 May 2008

Between 1 and 2% of the population in the developed world experiences a nonhealing or chronic wound characterized by an apparent arrest in a stage dominated by inflammatory processes. Lately, research groups have proposed that bacteria might be involved in and contribute to the lack of healing of these wounds. To investigate this, we collected and examined samples from chronic wounds obtained from 22 different patients, all selected because of suspicion of Pseudomonas aeruginosa colonization. These wound samples were investigated by standard culturing methods and peptide nucleic acid-based fluorescence in situ hybridization (PNA FISH) for direct identification of bacteria. By means of the culturing methods, Staphylococcus aureus was detected in the majority of the wounds, whereas P. aeruginosa was observed less frequently. In contrast, using PNA FISH, we found that a large fraction of the wounds contained P. aeruginosa. Furthermore, PNA FISH revealed the structural organization of bacteria in the samples. It appeared that P. aeruginosa aggregated as microcolonies imbedded in the matrix component alginate, which is a characteristic hallmark of the biofilm mode of growth. The present investigation suggests that bacteria present within these wounds tend to be aggregated in microcolonies imbedded in a self-produced matrix, characteristic of the biofilm mode of growth. Additionally, we must conclude that there exists no good correlation between bacteria detected by standard culturing methods and those detected by direct detection methods such as PNA FISH. This strongly supports the development of new diagnostic and treatment strategies for chronic wounds.

* Corresponding author. Mailing address: BioScience and Technology, BioCentrum-DTU, Technical University of Denmark, Lyngby 2800, Denmark. Phone: 45 45 25 25 98. Fax: 45 45 88 73 28. E-mail: tbj{at}biocentrum.dtu.dk

{triangledown} Published ahead of print on 28 May 2008.

Journal of Clinical Microbiology, August 2008, p. 2717-2722, Vol. 46, No. 8
0095-1137/08/$08.00+0     doi:10.1128/JCM.00501-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.



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