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Type of Document Dissertation Author LaClair, Darcy Danielle Author's Email Address dlaclair@nd.edu URN etd-04212006-162550 Title Optimization and Characterization of the Growth of the Photosynthetic Bacterium Blastochloris Viridis and a Brief Survey of Its Potential as a Remediative Tool Degree Doctor of Philosophy Department Chemical Engineering Advisory Committee
Advisor Name Title Agnes E. Ostafin Committee Chair Andre Palmer Committee Member David Leighton Committee Member Jennifer Woertz Committee Member Mark Stadtherr Committee Member Keywords
- absorbance measurement
- spin plating device
- spin plating
- plate count
- Blastochloris viridis
- bacteria
- procaryote
- waste remediation
- Toxin degradation
- anaerobic purple photosynthetic bacteria
- media optimization
- growth characterization for pigmented bacteria
Date of Defense 2006-04-12 Availability unrestricted Abstract The growth of B. viridis was characterized in an undefined rich medium anda well-defined medium, which was later selected for further experimentation
to insure repeatability. This medium presented a significant problem in obtaining either multigenerational or vigorous growth because of metabolic limitations; therefore optimization of the medium was undertaken. A primary requirement to obtain good growth was a shift in the pH of the medium from 6.9 to 5.9. Once this shift was made, it was possible to obtain growth in subsequent generations, and the media formulation was optimized. A response curve suggested optimum concentrations of 75 mM carbon, supplemented as sodium malate, 12.5 mM nitrogen, supplemented as ammonium sulfate, and 12.7 mM phosphate buffer. In addition, the vitamins p-Aminobenzoic acid, Thiamine, Biotin, B12, and Pantothenate were important to achieving good growth and good pigment formation. Exogenous carbon dioxide, added as 2.5 g sodium bicarbonate per liter media also enhanced growth and reduced the lag time. The optimized medium enhanced the total growth of cells slightly, and increased the culture optical density and pigmentation by a factor of 2 and 3, respectively, bringing the culture performance in the defined medium above the performance in the rich, undefined medium.
A total of 31 chemicals from 17 chemical families were screened at either 10 mM or 2 mM concentration to determine their effect on the growth and pigment formation of B. viridis. Most alcohols surveyed caused significant enhancement to growth rate, and the primary carbon in a secondary alcohol was proposed as the site of attack allowing usage of these compounds. Aromatic compounds were not as successfully cultured, and potential mechanisms for cell damage and compound degradation were discussed. B. viridis demonstrated the ability to grow unimpaired in the presence of MEK, methyl chloride, mimethyl sulfoxide, dimethyl formamide, chloroethane, acetonitrile, and dioxane.
Several methods for working with bacteria were presented. The Two-Point method, a rapid and reliable method to compare cell growth and health, was demonstrated. A mechanized device for spin plating was also introduced, both significantly reducing operator fatigue and injury, and increasing the consistency of plate count data.
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