Graduate Thesis Or Dissertation
 

Nutritional factors associated with the presence and growth of Klebsiella in redwood environments

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  • Bacteria of the genus Klebsiella have been found to be associated with redwood water storage reservoirs, redwood chips, and liquid expressed from within living redwood. A 1,000 gallon experimental redwood reservoir was employed to determine how to control the presence of Klebsiella and Enterobacter in drinking water stored in these tanks. Over a twenty month period of operation, these coliforms were not detected in tank water. Control of Klebsiella and Enterobacter was dependent on three factors: maintenance of a free chlorine residual of 0.2-0.4 ppm in the tank water, maximum retention times of four to seven days, and the use of a separate overhead inlet water pipe. Klebsiella multiplied to 10⁶ per ml in dechlorinated water samples from the experimental tank, utilizing nutrients leached from the redwood. The predominant metabolizable carbon sources in aqueous extracts of redwood are cyclitol compounds, of which myo-inositol is the parent compound. Pinitol and sequoyitol, two other cyclitols, are also encountered in redwood. 100%, 97%, and 68% of the combined Klebsiella isolates from clinical and environmental origins fermented inositol, sequoyitol, and pinitol, respectively. These compounds can also be used as a sole source of carbon and energy by Klebsiella. Similar results were obtained with Enterobacter isolates, but most other enteric bacteria tested could not metabolize cyclitols. Strains of Klebsiella multiplied to levels exceeding 10⁵/ml in aqueous extracts of non-sterile redwood within 6 days. Most other enteric bacteria did not grow in these extracts. Cyclitol metabolism correlated well with the ability to multiply in redwood extract in the presence of cyclitol-negative indigenous bacteria. Other experiments were conducted with three small redwood tanks in order to assess the effects of chlorine residuals, retention times, and cyclitol concentrations in redwood tank Water on coliforms and on Klebsiella. As chlorine residuals were increased, coliform and total bacterial counts decreased. Longer retention times resulted in the accumulation of cyclitol compounds and a decrease in chlorine residuals. A decrease in cyclitol concentration was found to correlate with a drop in coliform and Klebsiella densities. No cyclitol degradation by indigenous coliforms was observed in water taken from the tanks, which partially explained why water flow is required to completely remove cyclitols from a redwood tank. In another study, gas chromatographic analysis was employed to demonstrate in situ cyclitol utilization in redwood extracts by isolates of Klebsiella, Enterobacter, and other gramnegative bacteria. In aqueous redwood extracts, all of the Klebsiella tested reached densities exceeding 5.0x10⁶ cells/ml within 4 days and all utilized pinitol and sequoyitol, as did Enterobacter. Other enteric bacteria did not utilize cyclitols in this extract. A defined minimal medium, containing the carbohydrates and cyclitols in redwood, was used to determine which carbon sources are preferentially utilized by Klebsielleae and other bacteria. It was found that D-glucose and L-arabinose were consumed by Klebsiella before the cyclitols were utilized. Pinitol utilization proceeded more slowly than that of sequoyitol and myoinositol. Cyclitol utilization in the defined medium was also observed for strains of Yersinia, Erwinia, and Salmonella. E. coli isolates did not utilize cyclitols. The ability to use cyclitols as a carbon source can explain the presence of Klebsiella and Enterobacter in redwood water storage reservoirs and in redwood lumber. This ability may also be related to their presence and growth in other botanical material containing cyclitol compounds. Other experiments demonstrated the utilization of myo-inositol hexaphosphate (phytic acid) by Klebsielleae, and the transfer of antibiotic resistance between strains of Klebsiella in aqueous extracts of redwood sawdust with cell densities of Klebsiella likely to occur in nature.
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