Fachbereich Veterinärmedizin



    Erste Untersuchungen zu Vorkommen und Zusammensetzung von Biofilmen bei Hunden, Katzen und Pferden mit chronischen Wundnahtinfektionen (2016)

    König, Lydia Marie (WE 12)
    Berlin: Mensch und Buch Verlag, 2016 — VII, 51 Seiten
    ISBN: 978-3-86387-710-1
    URL (Volltext): http://www.diss.fu-berlin.de/diss/receive/FUDISS_thesis_000000102025
    Institut für Tierpathologie

    Robert-von-Ostertag-Str. 15
    Gebäude 12
    14163 Berlin
    +49 30 838 62450

    Abstract / Zusammenfassung

    Resistant bacteria are an increasing problem for the medical treatment of chronically infected wounds. This is caused by resistance mechanisms bacteria developed for their protection. Living as a community, bacteria are able to develop biofilms which are composed of bacteria and an extracellular polymeric matrix (EMP). Those biofilms work as a barrier against antimicrobial factors including traditional antibiotic therapy of bacterial infections. Especially suture material and medical devices such as implants constitute an interface for bacterial colonization and the development of biofilms. The first part of the study was a retrospective investigation of archive material from the Department of Veterinary Pathology, Freie Universität Berlin, to assess the incidence of biofilms. Ninety-one samples were analyzed. Sixty-eight of them originated from dogs, 15 from cats, and eight from horses. Fifty-three tissue samples were collected from skin, mucosa, or mammary gland. Three originated from skeletal muscle, seven from intestine and 28 from urogenital tract. By using a combination of different histopathological stains, the components of a biofilm could be visualized so that bacteria, interface and matrix could be detected under the light microscope. A hematoxylin and eosin staining (HE) was used to identify the interface between the suture material and the tissue. A periodic-acid-Schiff reaction (PAS) was used to visualize the EMP. Giemsa and Gram stains identified nucleic acid or Gram-positive bacterial organisms, respectively, in the EPM. The investigation identified biofilms in two of 91 samples. Both positive samples included polyfilic suture material from dog tissue. The first one was a biopsy from a skin wound and showed a severe chronic-active suppurative and granulomatous inflammation. The second one was a fragment of an ovariohysterectomized uterus stump associated with a severe chronic-active lymphoplasmacytic and granulomatous inflammation. The literature on the subject of biofilms is dominated by human medicine studies and describes a much higher incidence than shown in this investigation. These differences might be caused by the following reasons: First, there are hints in the literature that the potential for biofilm-building of animal bacterial is lower than of human bacteria; second, the data regarding pre- and postoperative antimicrobial treatment, surgical methods and hygienic management during the surgeries are incomplete. Because these factors have probably influenced the infection rate during the implantation of foreign body material, it is difficult to compare these results with recent investigations of human tissues. A further problem is the lack of a gold standard for biofilm detection which makes it even more difficult to compare the results. The second part of the study consisted of the identification of bacterial in the biofilms by means of Next Generation Genome Sequencing (NGGS). Subsequent to the selection of samples and the DNA isolation this was realized by Dr. Dirk Höper, Friedrich-Loeffler-Institute, Island Riems. In total, three samples underwent NGGS. Two of them were taken from the first part of the study while the third one was included from a different project. The third sample was also dog tissue and it also contained polyfilic suture in a surgical wound of the skin. The genetic analysis identified typical biofilm-associated bacteria like Enterobacteriaceae as well as bacteria like Deinococcaceae which have not been identified in infected wounds so far. The three samples showed overlapping results of bacterial families but they also had differences in their composition. Overlapping bacterial families in all of the three samples were Fusobacteriaceae and Porphyromonadaceae. The three most prominent families in the first analysed biofilm were Porphyromonadaceae, Fusobacteriaceae and Peptostreptococcaceae, in the second biofilm Deinococcaceae, Methylobacteriaceae and Nocardiaceae and in the third sample Porphyromonadaceae, Alteromonadaceae and Fusobacteriaceae. The NGGS-analysis has advantages over other methods used for retrospective studies. In this method it is possible to use formalin fixed and paraffin embedded material and a small sample quantity is sufficient. A further advantage is the clear assignment of DNA sequences to bacterial families and species based on a database matching. However, it is a sophisticated method with complicated procedures and special knowledge to evaluate the data is necessary. Furthermore, the method provides no information about the vitality of the detected bacteria and the causality among these bacteria and the biofilm formation. Summarizing the results of this study, it could be shown that (1.) biofilms occur in infected wounds associated with suture material in animals, (2.) the incidence of biofilms seems to be lower than reported in human medicine, and (3.) the number of detected bacterial families was surprisingly high and there were differences among the samples and to the results generated in previous studies.