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United States Department of Agriculture

Agricultural Research Service

Research Project: HANDLING AND TRANSPORT STRESS INTERACTIONS WITH PATHOGEN BIOLOGY IN SWINE AND CATTLE Title: Photonic plasmid stability of transformed Salmonella typhimurium: A comparison of three unique plasmids

Authors
item Moulton, K - MISSISSIPPI STATE UNIV
item Ryan, P - MISSISSIPPI STATE UNIV
item Lay, Jr, Donald
item Willard, S - MISSISSIPPI STATE UNIV

Submitted to: BMC Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: January 12, 2009
Publication Date: July 27, 2009
Citation: Moulton, K., Ryan, P., Lay Jr, D.C., Willard, S. 2009. Photonic plasmid stability of transformed Salmonella typhimurium: A comparison of three unique plasmids. BMC Microbiology. 9:152-159.

Interpretive Summary: Researchers are increasingly interested in observing biological processes in real-time. The development of reporter systems such as fluorescence and bioluminescence allows for imaging and measuring of various biological activities. The objective of this study was to determine the stability of Salmonella typhimurium (S. typh-lux) which was transformed using a plasmid to emit light. In addition, we sought to determine the respective photonic properties of plasmids using different imaging platforms and by varying concentrations of S. typh-lux bacteria. These data characterize the photon stability properties for Salmonella typhimurium transformed with three different photon generating plasmids. Salmonella typhimurium that is transformed with two of three plasmids were more stable and have better correlations with actual bacterial concentration than a third plasmid. However for short-term evaluations of 1 to 6 days, all three plasmids may permit real-time Salmonella tracking using in vivo or in situ biophotonic paradigms where antibiotic selective pressure to maintain plasmid incorporation may not be feasible.

Technical Abstract: Acquiring a highly stable photonic plasmid in transformed Salmonella typhimurium for use in biophotonic studies of bacterial tracking in vivo is critical to experimental paradigm development. The objective of this study was to determine stability of transformed Salmonella typhimurium (S. typh-lux) using three different plasmids and characterize their respective photonic properties. In presence of ampicillin (AMP), S. typh-lux with pCGLS-1, pAK1-lux and pXEN-1 plasmids exhibited 100% photon-emitting colonies over a 10-d study period. Photon emitters of S. typh-lux with pCGLS-1, pAK1-lux and pXEN-1 without AMP selection decreased over time (P < 0.05), representing only 11 ± 1%, 35 ± 1% and 43 ± 1%, respectively, of original photon emitting properties of the bacterial population by d 10. Photonic emissions were positively correlated with bacterial concentration (P < 0.05) for pAK1-lux, pCGLS-1 and pXEN-1 (r = 0.96, 0.98 and 0.82, respectively). When stratified by high, medium and low density bacteria concentrations, photonic emissions for high density populations containing pAK1-lux, pCGLS-1 and pXEN-1 resulted in differences of photonic emissions across a range of bacterial concentrations (1×10*7 to 1×10*9 CFU, P < 0.05) with positive correlations (P < 0.05) of (r = 0.72, 0.46 and 0.72, respectively). The correlation of photonic emissions with bacterial concentrations for samples with medium and low density bacteria (pAK1-lux, pCGLS-1, and pXEN-1 plasmids) imaged in tubes were also positively correlated (medium; r=0.69, 0.49, 0.46, low; r=0.90, 0.71, 0.68, respectively; P > 0.05); although photonic emissions across a range of bacterial concentrations were not different (1×10*4 to 1×10*6 CFU, P > 0.05). For very low density bacterial concentrations imaged in 96 well plates photonic emissions were positively correlated with bacterial concentration (P < 0.05) for pAK1-lux, pCGLS-1, and pXEN-1 plasmids (r=0.99, 0.99, and 0.96, respectively), and photonic emissions across a range of bacterial concentrations (1×10*3 to 1×10*5 CFU) low to high were different in the 96-well plate format (P < 0.05). These data characterize photon stability properties for S. typh-lux transformed with three different photon generating plasmids that may facilitate real-time Salmonella tracking using in vivo or in situ biophotonic paradigms. [USDA-ARS Biophotonics Initiative # 58-6402-3-0120].

Last Modified: 10/20/2014
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