Growth rate effects during starvation-survival of a marine psychrophilic vibrio Public Deposited

http://ir.library.oregonstate.edu/concern/graduate_thesis_or_dissertations/fn107142s

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  • Cell populations of the marine bacterium ANT-300, from either batch culture or continuous culture with dilution rates ranging from D = 0.170 h⁻¹ (fast) to D = 0.015 h⁻¹ (slow) were monitored for 98 days under starvation. Viability (CFU), acridine orange direct counts (AODC), and optical density were measured. DNA, RNA, and protein concentrations were also estimated on a total and viable cell basis. A new method for the assay of nucleic acids was developed for this study. Three viability patterns of starvation-survival were observed for each of the cell populations. AODC cells remained at 2-3 x 10⁷ cells per ml throughout the starvation period. Large fluctuations occurred in cell viability during stage 1 (0 to 14 days) of starvation-survival. Stage 2 (14 to 70 days) involved an overall decrease in viability for each of the cell populations, with the rate of viability loss being dependent upon the growth rate. Cell viability stabilized at approximately 0.3% of the AODC cell number in stage 3 (70 to 98 days). Long-term starvation was the prolongation of stage 3 starvation-survival. Biovolumes for each of the cell populations decreased with the length of the starvation period. However, the biovolume of starved cells depended on growth rate more than the length of time starved. DNA, RNA, and protein concentrations on a total and viable cell basis fluctuated corresponding to the three stages of starvation-survival. During stage 1 of starvation-survival, 2-3 peaks in the concentration levels for all three macromolecules were characteristic. DNA per total cell in stage 2 dropped to 4.2 - 8.3% for all of the cell populations examined, and then stabilized throughout stage 3. The decrease in DNA per cell was also observed in electron micrographs of cellular DNA in starved and unstarved ANT-300 cells. The fluctuations of RNA and protein concentrations observed during stage 2 and 3 were parallel in all cases except for the cells from D = 0.015 h"1. This ANT-300 cell population showed a decrease in RNA to 29.2% and an increase in protein to 129.7% of original concentrations. RNA and protein concentrations also stabilized in stage 3. The cells from the faster growth rate cell populations of D = 0.170 h⁻¹ and batch culture had elevated protein concentrations, which remained mostly residual during the starvation period. Macromolecule ratio data indicate two phenomena: (i) The D = 0.015 h⁻¹ cells demonstrate the production and maintenance of protein relative to diminished RNA levels, (ii) The D = 0.057 h⁻¹ cells suggest the existence of a transitory state between the high efficiency of the D = 0.015 h⁻¹ cells and the high residuals of the D = 0.170 h⁻¹ l and batch culture cells. It has been demonstrated that three stages of starvation-survival exist as indicated by cell counts and the estimation of DNA, RNA, and protein, for all the cell populations examined. Slow growth rate (D = 0.015 h⁻¹ ) cells indicate enhanced viability during the starvation period, and the capability to produce and maintain relatively high protein levels per cell. These cells exhibit low concentrations of RNA per cell, thus yielding more efficient translation and maintenance capabilities under starvation conditions. Cells from this population also have the lowest biovolume, corresponding to the greatest surface area during the starvation period. Therefore, it is hypothesized that under starvationsurvival conditions the ANT-300 population of slow growth rate cells (D = 0.015 h⁻¹ ) are the closest representatives to cells found in the marine environment at in situ growth rates.
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