PSA-Nutrition: Amino Acids and Vitamin/Mineral Nutrition I

Salmonella Typhimurium is attracting world wide attention as it continues to cause foodborne illness to human beings. The organism is resistant to a wide range of anti-infectious agents and as a result the illness is more difficult to treat. The objective of this study was to analyze the expression of specific virulence and stress genes (hilA and rpoS) in Salmonella Typhimurium at various nutritional stress and pH in a continuous culture system. Salmonella Typhimurium cells were propagated in continuous cultures with a total volume of 0.50 liter LB minimal medium at 98% turnover rate. Two chemostats were used for adjusting dilution rates and pH levels. Dilution rates were 0.0125 h-1, 0.025 h-1, , 0.05 h-1, , 0.1 h-1, , 0.27 h-1, , 0.54 h-1, , 1.08 h-1, and 1.44 h-1, while pH levels were 6.1 through 8.0. Results indicated that cell protein increases as dilution rates (D) increase (D 0.0125 h-1, =1184 g cell: 0.27 h-1, =1431g cell, however, at 0.54 h-1, there was a decline in the g/cell protein quantity. This suggests that when dilution rate reaches washout in the chemostat, the cell mass decreases. Changes in glucose yield (Yglc) also indicated that as the pH increases so does the Yglc(pH 7.29=243± 14g cell/mol glucose utilized: pH 6.15= 30±13g cell/mol glucose utilized (P>0.05)). To analyze gene expression, samples were stored in RNAprotect (Qiagen, Valencia, CA) in duplicate. Realtime PCR reactions were performed on an ABI Prism 7700 Sequence Detection System (Perkin-Elmer Applied Biosystems, Foster City, CA). Samples were analyzed using the Comparative Ct (∆∆Ct) method by normalizing to the endogenous control gene (rsmC ). Both hilA and rpoS expression appeared to change in response to corresponding pH and Yglc transitional changes in the continuous culture.