Transport of metabolic active bacteria through saturated quartz sand columns with and without substrate addition
Nowadays environmental protection not only refers to prevention of chemical pollution but also to biological contamination, which occurs due to manure or sludge on agricultural land. The released bacterial biomass is not negligible. The knowledge and the prediction of the behaviour of the released microorganisms in the upper soil as well as the deposition in lower soil compartments and the possible entry into the groundwater are of great interest, especially in case of pathogenic microorganisms. These transport pathways have been the subject of several investigations in the recent years. Experiments were carried out in different size scales: in columns, lysimeters, both filled with sand or soil, and outdoor tests. In all cases a so-called breakthrough curve (BTC) is calculated via analysis of the eluate. This BTC is characteristic for the chosen setup in means of conditions, e.g. column size, soil type, microorganism strain. At the moment a prediction of the transport behaviour is not possible, but there are competing theoretical models, from which calculations of microorganism distribution over a certain soil in the upper soil regions under the surface can be observed. But stringent boundary conditions (bacteria are treated as inactive particles) neglect the effects of bacterial properties on transport processes. Therefore predictions are only of limited value because they do not take into account effects of substrate, growth or stress. In this work the transport behaviour of metabolic active bacteria of Pseudomonas fluorescens in saturated quartz sand columns were investigated. The effect of the age of the used culture on the distribution of the bacteria between eluate and column was quantified. Furthermore the effect of substrate-addition under the same conditions was evaluated. For real-time measurements the used strain was genetically modified by chromosomal anchorage of the green fluorescent protein (gfp). Thus, in the eluate bacterial concentrations per time interval were detected. Prior to the bacterial breakthrough the breakthrough of D2O as a non reactive tracer was detected to control the package of the column. The bacterial BTC was compared to the one of D2O. After bacterial breakthrough the amount and morphology of bacteria in the single eluate fractions as well as in single column slices were determined. Both were carried out by visualization of the microorganisms by fluorescent dyes, digital photo imaging and digital picture processing. The obtained morphological date were saved in a database and via SQL examined in detail. For distinction definite features were acquired and a relationship between mobility and transport on the one hand and the age of a culture on the other hand were determined. The breakthrough of P. fluorescens-gfp occurred in the same time range like the one of D2O and was not retarded as observed from inactive microspheres. Without substrate addition the breakthrough shifted backwards with increasing age of the culture, i.e. fresh cells passed the column faster. Under this aspect the morphological difference between old, coccoid and fresh, rod-shaped cells played an important role. Furthermore the cell amount increased more than 100-times. This increase was not a result of real culture growth but due to the metabolic base respiration and therefore the resulting lack of oxygen inside the column. Consequently an increase of cell amount and a morphological change in parallel were detected. This change happened by dividing of rod-shaped into several coccoid cells. The morphological change decreased with increasing age of culture, at the same time the amount of cells which passed the column was increasing. Very old cells showed a nearly complete breakthrough, whereas the greatest part of log-phase cells was retarded inside the column. The distribution pattern inside the column was age-dependent, too: fresh cells were found at the column-inlet whereas older cells were uniformly spread all over the column. Addition of substrate also led to an age-dependent effect: the recovery time of cells to enter the log-phase was the main factor. Cells older than 70h showed no differences in the distribution between eluate and column. The expected breakthrough occurred earlier than the one of a non reactive tracer. The distribution inside a column was uniformly compared to the experiment without substrate. With substrate fresh cells showed a lower cell amount increase than without substrate, for old cells this was conversely.
info:eu-repo/classification/ddc/540, Bodenbakterien, Boden, Chemie, Transport, Mikroorganismen, microorganisms
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