Molecular Analysis of Extrapolymeric Substances of Pseudomonas Aeruginosa and Staphylococcus Aureus Biofilms

Microbes are present virtually everywhere, including soil, water, inanimate and animate objects. The preferred mode of existence of most microbes is as biofilms. Microbial biofilms are a community of microbes in extracellular polymeric substances (EPS) that they synthesize themselves. Biofilms serve as protective, hydrated barriers between cells and the external environment. EPS are mainly composed of polysaccharides, however they may also contain proteins, lipids, and nucleic acids. This project focuses on characterizing the EPS components secreted by Pseudomonas aeruginosa and Staphylococcus aureus. Pseudomonas aeruginosa, and other members of its genus are plant pathogens, and are commonly found on the surface of plants and in soil. Staphylococcus aureus is normally found on the skin surface of humans and other organisms, but can also be found within the normal flora of soil. The EPS of single species biofilms were collected using the glutaraldehde extraction method. Concentrated films were analyzed for DNA, proteins, and lipids. Proteins were first analyzed using SDS-PAGE and Coomassie Blue staining techniques, however no proteins were detected. Silver staining, a more sensitive method has been used to confirm the presence of protein within the films. Thin layer chromatography has shown the presence of lipids in both biofilms. No DNA has been detected by agarose gel electrophoresis, however literature reports indicate the presence of DNA within these biofilms. Another method, using a sensitive UV spectrophotometer to confirm the presence and quantify the amount of DNA within the EPS, is being explored.