The bacteria Pseudomonas fragi and Pseudomonas aeruginosa pose a major challenge due to their ability to form biofilms, which enable them to survive even in the unfavorable conditions of healthcare and industrial environments. Biofilms provide bacteria with additional protection against external influences, making them difficult to remove. In master's thesis, we established a method for the detection and quantification of P. fragi bacteria using fluorescent proteins. First, we prepared plasmids pBBR1MCS-2_mTagBFP2 and pBBR1MCS-2_mCherry, which carry genes for the expression of fluorescent proteins mTagBFP2 and mCherry, and performed bacterial transformation using electroporation. The obtained transformed P. fragi and P. aeruginosa bacteria, which expressed fluorescent proteins, were used to identify bacteria in planktonic form and in the form of biofilms. Measurements of fluorescent light intensity showed a correlation between bacterial growth and the intensity of fluorescent marker expression in P. fragi bacteria, confirming the reliability of the proposed method. The established method is fast, accurate, and reproducible, enabling accurate quantification of bacterial populations under various experimental conditions. In addition, the transformed P. fragi bacteria retained their ability to form biofilms, confirming the usefulness of fluorescent markers for studying bacterial growth and their attachment to surfaces. The established method represents a new way of detecting bacteria in the form of biofilms, opening up new possibilities for the development of strategies for the prevention and control of biofilms and the reduction of the risks of bacterial contamination and infection.
|
