Study of Pseudomonas aeruginosa and different wound dressing products Master of Science Thesis

An injury to the skin resulting in a wound starts the complex healing machinery of the body, normally leading to rapid wound closure. In some cases the normal wound healing process is disturbed and prolonged; which can lead to chronic, non-healing wounds such as venous leg ulcers. Chronic wounds in general are estimated to affect 1 – 2 % of the world’s population, and about 1 % is suffering from venous leg ulcers. The persistency of chronic wounds is often caused by bacterial infections, where the invading microorganisms cause significantly impaired wound healing. There is now increasing knowledge that infections in humans often involve bacterial biofilms, where bacteria form microcolonies embedded in a protective extracellular polysaccharide matrix. Bacteria growing in mature biofilm are very hard to eradicate due to development of several resistance mechanisms. One of the most common pathogens in chronic wounds is Pseudomonas aeruginosa, a very problematic microbe due to its ability to form resistant biofilms. The bacteria produce several virulence factors that allows for establishment in host tissue, where one important substance is the blue pigment pyocyanin. Pyocyanin has several toxic effects on host cells, including depletion of NADH, glutathione and other antioxidants in the host cell, an alteration of the redox status, and generation of oxygen radicals. Chronic wounds, especially venous leg ulcers, are often treated with foam dressings capable of absorbing high amounts of wound exudates. This project is an attempt to elucidate if polyurethane foam dressings can promote wound infection by P. aeruginosa. This was investigated by in vitro studies of bacterial growth, biofilm formation and production of pyocyanin in the absence and presence of wound dressing samples. P. aeruginosa PAO1 was cultured together with wound dressing pieces. Viable cell counts showed that growth of P. aeruginosa was not affected by the presence of any wound dressing product included in the study. The presence of polyurethane foam dressings in P. aeruginosa cultures caused an altered environment, which was observed as a change in color from green to yellow due to reduction of pyocyanin. Measurement by absorbance spectrophotometry also showed that the presence of wound dressing products resulted in lower concentration of pyocyanin in the solution, compared to the control. The amount of pyocyanin in P. aeruginosa cultures was found to increase over time, where the control culture showed the largest increase. In cultures containing polyurethane foam dressings the pyocyanin content increased in the solutions only between day 1 and day 3. Between day 3 and day 7 the increase in pyocyanin content instead occurred inside the dressing products. Dressings were also placed in a cell-free pyocyanin solution derived from stationary phase culture. Absorbance measurements of the pyocyanin solutions showed pyocyanin adherence to the dressing material polyurethane, even in the absence of bacteria. The polyurethane foam dressing products included in this study all show similar results in the experiments. The wound dressings do not seem to influence the number of viable cells in the bacterial suspensions. The presence of dressings in cultures seems to have a decreasing effect on the level of the virulence factor pyocyanin in the bacterial suspensions, due to accumulation and adherence inside the wound dressing.