Assessment of the effects of superoxide-generating agents on the growth and viability of Escherichia coli using traditional microbiological methods and fluorescence methods
- WCU Author/Contributor (non-WCU co-authors, if there are any, appear on document)
- Jennifer Lynn Patterson (Creator)
- Institution
- Western Carolina University (WCU )
- Web Site: http://library.wcu.edu/
- Advisor
- Lori Seischab
Abstract: The effects of superoxide-generating agents on the growth and viability of
Escherichia coli was investigated using two different strains, a lab strain (ER2566) and a
clinical isolate (ATCC 4157). Endogenous superoxide was generated using the redoxcycling
agent paraquat, while exogenous superoxide was generated using the
xanthine/xanthine oxidase (X/XO) enzyme system. Using optical density measurements
to monitor culture growth, the bacteriostatic effect of paraquat was tested in three
different growth media: Luria-Bertani broth, double strength Luria-Bertani broth, and
nutrient broth. For both strains, paraquat toxicity was greatest in nutrient broth, with
toxicity in each medium dependent upon the time of paraquat addition following
inoculation. Protection against paraquat toxicity by salts and yeast extract was suggested
by the differences observed between growth rates of ER2566 treated cultures in each
medium. Addition of the copper/zinc superoxide dismutase inhibitor
diethyldithiocarbamate decreased paraquat toxicity, consistent with its role in induction
of the superoxide response regulon (soxRS). Based on colony-forming unit (CFU)
counts, the toxicity of X/XO-generated superoxide on ER2566 was found to be altered by
pH, with cell viability lower at a pH of 6.5 than at 7.5. Using CFU counts obtained with the spread-plate method and total cell counts obtained with a hemacytometer, the
relationship between optical density and cell number was found to be different between
cultures of the two strains at both log and stationary phases. Reliable cell counts were
necessary to avoid the inner filter effect otherwise encountered during optimization of the
BacLight Bacterial Viability Kit for ATCC 4157. Assessment of cell viability following
X/XO treatment with the BacLight kit indicated this method was more sensitive than the
traditional spread plate method for determining cell viability.
Assessment of the effects of superoxide-generating agents on the growth and viability of Escherichia coli using traditional microbiological methods and fluorescence methods
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Created on 7/1/2010
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Additional Information
- Publication
- Thesis
- Language: English
- Date: 2010
- Subjects
- Escherichia coli -- Growth
- Superoxide dismutase