Submitted to: Environmental Engineering Science
Publication Type: Peer reviewed journal
Publication Acceptance Date: 7/20/2009
Publication Date: 10/1/2009
Publication URL: http://hdl.handle.net/10113/43898
Citation: Gong, A.S., Bolster, C.H., Benavides, M., Walker, S.L. 2009. Extraction and Analysis of Extracellular Polymeric Substances (EPS): Comparison of Methods and EPS Levels in Salmonella pullorum SA 1685. Environmental Engineering Science. 26:1523-1532. Interpretive Summary: Bacteria have unique physical, chemical, and biological characteristics, one of which is the production of extracellular polymeric substances (EPS) that play an important role in cell adhesion and retention on mineral surfaces, thus impacting the fate and transport of microorganisms in groundwater environments. Therefore, in order to study the fate of microorganisms in the environment, the nature and content of the EPS must be characterized. Although numerous methods have been developed for the extraction and measurement of EPS, there have been few studies comparing the effectiveness of the different extraction methods under conditions representative of groundwater environments. In this study, systematic comparisons and statistical analyses were made between three extraction methods under a variety of ionic strengths and exposure times using Salmonella pullorum SA 1685. The results of this study showed the sensitivity of the three EPS extraction methods to various environmental conditions, thus providing insights on these three methods and helping to determine the capability of each method in detecting environmental changes in simulated or real subsurface environments.
Technical Abstract: The extracellular polymeric substances (EPS) production and composition for Salmonella pullorum SA 1685 exposed to artificial groundwater (AGW) has been examined utilizing three EPS extraction methods: lyophilization, ethanol, and sonication. Experiments were carried out to evaluate the robustness of the three EPS extraction methods and the sensitivity of each to subtle changes in solution ionic strength (IS) and duration of exposure. EPS extraction and analysis was conducted via sugar and protein analyses using the phenol sulfuric acid and Lowry methods, respectively, after 0, 6 h, 12 h, 18 h, and 24 h incubation times in AGW with 10-2.5, 10-2, and 10-1.5 M IS. The lyophilization and ethanol methods resulted in a greater amount of EPS extracted than the sonication method (mass of EPS/cell), yet these methods fluctuated to a greater extent in the total amount – or level – of EPS extracted under the various test conditions. Systematic comparisons and extensive statistical analyses were conducted between the various experimental conditions. To our knowledge, this is the first study systematically comparing EPS extraction techniques utilizing Salmonella. As we investigated the relative EPS content in Salmonella SA1685 under conditions simulating groundwater, our results provide insight into the suitability of each method for detection of environmentally induced changes in bacteria suspended in simulated or real subsurface aquatic systems.