Title page for ETD etd-08252008-162830
|Type of Document
||King, Laura Kathryn
||Response of indigenous heterotrophic groundwater bacteria to low organic substrate availability
||Master of Science
|Parker, Bruce C.
|Krieg, Noel R.
|Yousten, Allan A.
|Date of Defense
Groundwater is one of the least studied environments, yet many people rely
on groundwater for their sole drinking water supply. Little is known
about the indigenous microflora, but it is believed to be similar to
oceanic planktobacteria due to the low nutrient concentrations occurring
in both ecosystems. That is, groundwater microorganisms are atypically small, mostly Gram-negative cells. Also like the oceanic
planktobacteria, they may have no affinity for surface attachment and may rely on dissolved low molecular weight organic substrates in dilute solution for their nutrition. Periods of metabolic dormancy may occur when
natural substrate concentrations drop below the level required to sustain
vegetative cell function. In these studies total cells present were determined
by 4'6-diamidino-2-phenylindole (DAPI) epifluorescent counts.
The percentage of those bacteria which were metabolically active was
determined by a modification of the 2-(p-iodophenyl)-3-(p- nitrophenyl)-
5-phenyl tetrazolium chloride (INT) reduction method. Advantages of this
method over others include more specific fluorochrome staining, ease of
transfer of the cells to the slide, time saved, and ease of microscopic viewing. Heterotrophic uptake of aspartate, succinate, glucose and
fructose by indigenous bacteria was measured and calculations of maximum
uptake velocity (Vmax) and a constant (Kt + Sn) equalling the natural
substrate concentration (Sn) plus the half-saturation concentration (Kt )
were made based on net assimilation (cellular retention) of radiolabeled
substrate. Total counts by DAPI staining were 4-12.1 x 104 cells/ml of
which 17.4 to 20.85% were metabolically active (INT+). Mean maximum
uptake velocities ranged from 1.73 to 2000 nmol/l/hr with aspartate being
taken up at the highest rate followed by fructose, succinate and glucose.
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