1. Results Phenylalanine (Figure 1A) P. aeruginosa grew logarithmically in the presence of phenylalanine. P. aeruginosa increased 100 fold over 10 days in the presence of phenylalanine as the sole nutrient source. Control showed no growth over the same time period of 10 days. Tryptophan (Figure 1B) P. aeruginosa grew logarithmically in the presence of tryptophan. P. aeruginosa showed a 10 fold increase over the control after 6 days of growth; however, the control itself also grew, but to a lesser extent. Conclusions P. aeruginosa increased growth with either phenylalanine or tryptophan as its only source of nutrient. The phenylalanine control did not grow, but the tryptophan did to some extent. This growth could be due to the presence of residual nutrients on the cells caused by inadequate rinsing of the cells as they were being prepared for the experiment. Now that we have established this experimental system we can further test the question about the mesopore protection theory (Mayer, 1994). Due to the dramatic increase in growth compared to the control, phenylalanine is a good experimental system for examining this theory. Future studies may include retesting tryptophan and increasing the rinse process; if the controls don’t grow under these conditions then either phenylalanine or tryptophan can be used to test the hypothesis. Also, it could be beneficial to find a lower concentration of each of the amino acids to determine when the growth becomes dependent on the concentration. Introduction Bacteria play an important role at many levels in the environment. One of the functions of bacteria is the decomposition of organic matter which is important for the interactions that take place on the surface of minerals. These interactions affect the amount of carbon sequestered in various soils and sediments. They also affect carbon cycles related to such events as climatic changes and to the formation of fossil fuels which are produced by continued burial of organic matter without degradation by the bacterial population (Brantley et al., 2001). In the soil, these reactions occur on the surface of minerals within macroscopic divots called mesopores. In 1994, L. Mayer proposed a hypothesis related to this phenomenon which stated that organic matter may prevent degradation by adsorbing into small mesopores found along the surface area of soils and sediments. According to this theory, organisms such as bacteria, which can break down organic matter, are too large to fit into the mesopores and access the organic matter that has been adsorbed into these spaces. In order to test the hypothesis that sequestered nutrients in mesoporous rock are protected from microbial access, one must first find an organism whose growth is dependent on a single nutrient that can be adsorbed to mesoporous rock. Zimmerman has adsorbed both alumina and silica (common rock minerals) mesoporous samples with the amino acids tryptophan and phenylalanine. In addition, it has been shown that the bacteria Pseudomonas aeruginosa, which is a common bacterium in soil, can grow in minimal media such as tap water (Kooij et al., 1982). Lanyi (1969) and Kooij et al. (1982) found, through a study that was somewhat similar to ours, that P. aeruginosa has an increased growth rate in the presence of amino acids, particularly, phenylalanine and tryptophan. In order to determine if P. aeruginosa could be used to examine the Mayer hypothesis, my study was done to test if P. aeruginosa could grow with tryptophan or phenylalanine as the sole nutrient. Objectives The goal of this study was to develop an experimental system that could be used to examine the mesopore hypothesis theory. Our hypothesis is that P. aeruginosa will show increased growth in the presence of phenylalanine or in the presence of tryptophan as the sole nutrient source. Literature Cited Lanyi, B. 1969. Amino acid utilization by serologically grouped Pseudomonas aeruginosa strains. Acta Microbiologica Academiae Scientiarum Hungaricae 6:357-361 Mayer, L.M. 1994. Relationships between mineral surfaces and organic carbon concentrations in soils and sediments. Chemical Geology 114:347-363. Van der Kooij, D., Oranje, J. P. 1982. Growth of Pseudomonas aeruginosa in tap water in relation to utilization of substrates at concentrations of a few micrograms per liter. Applied and Environmental Microbiology 44:1086-1095. Zimmerman, Andrew R., Goyne, Keith W., Chorover, Jon, Komarneni, Sridhar, Brantley, Susan L. 2003. Mineral mesopore effects on nitrogenous organic mater adsorption. Submitted to Organic Geochemistry. Utilization of Phenylalanine or Tryptophan as a Single Nutrient Source for Growth of Pseudomonas aeruginosa Ronna Thomsen Department of Biological Sciences, York College of Pennsylvania Acknowledgements: Carolyn Mathur, PhD, Thesis Mentor; Ron Kaltreider, PhD; Karl Kleiner, PhD Figure 1. Growth pattern of P. aeruginosa in minimal media with different concentrations of phenylalanine or tryptophan as the sole nutrient source. Increased growth was observed in all concentrations of phenylalanine and tryptophan compared to the control. Standard plate counts (SPC) were done using nutrient agar and incubated overnight. Averages of the resulting SPC are shown with error bars indicating 95% confidence intervals of standard error. (A) Growth curve of P. aeruginosa in the presence of 0.64 mM and 1.28 mM phenylalanine. A total of eight SPC’s were performed over a period of ten days. (B) Growth curve of P. aeruginosa in the presence of 0.64 mM and 1.28 mM tryptophan. A total of five SPC’s were performed over a period of six days. A.B. Prepare overnight nutrient broth culture of P. aeruginosa at 35°C Wash 3x @ 4000 rpm with 0.02 M CaCl 2 Resuspend in CaCl 2 to use as innoculum Negative Control CaCl 2 1.28 mM (Tubes 4 & 5) 0.64 mM (Tubes 2 & 3) Standard Plate Count (SPC) on Day 0 Incubate @ 30ºC In a shaking water bath Sampled periodically to measure growth by SPC Harvest Cells Methods PhenylalanineTryptophan www.old.jccc.net/~pdecell/biochemistry/aminos.html Obtained from the collection of Dr. Carolyn Mathur