The effects of natural and anthropogenic factors on microbes decomposing the emerging macrophyte Scirpus lacustris in prairie aquatic systems
Emergent macrophytes, like Scilpus lacustris, are the foundation of the very high biological productivities of wetlands in the Northern Prairies of North America. Fungi and bacteria are the primary organisms that sequester carbon and nutrients from these macrophytes. Almost nothing is known about the process of microbial decomposition and how natural environmental factors and anthropogenic pollutants may impact the microbes associated with Scirpus as it decays while standing and after it falls into the water. Pond 50, located at the St. Denis National Wildlife area, Saskatchewan, Canada, was chosen as the study site because it is a wetland that typifies the prairie ecozone. First, a procedure to extract ergosterol, a molecule used to estimate living fungal biomass, from various environmental matrices was developed. Ergosterol was detected by high-pressure liquid chromatography with a UV detector (HPLC-UV) and corroborated by mass spectrometry analysis. Seasonal variations in fungal biomass and fungal production, the latter measured by the incorporation of [1_14C] acetate into ergosterol, associated with SCilPUS stems, both above and below water, were determined. Changes in fungal biomass and productivities on freshly cut green Scirpus stems decaying in the water under either natural solar radiation (UV +) or protected from ultraviolet (UV -) radiation, were followed over the summer. Several experiments measuring the impact of temperature on fungal biomass and production on Scirpus decaying in pond water were conducted in the laboratory. In view of antibiotics being detected in surface waters, tetracycline was chosen to determine its effects on the microbes that decompose Scilpus. Prior to this experiment, tetracycline's adsorptive characteristics in distilled, river and pond water were measured. Tetracycline's photolytic and microbial rates of degradation were determined in the three mentioned waters in the laboratory in the light and dark and also in natural sunlight (UV +) or protected (UV -) from UV radiation in non-sterile waters. The effects of tetracycline on protein production in planktonic bacteria in river and pond water were experimentally measured. Lastly, confocal laser microscopy (CLSM) was used to study the microbial colonization on Scirpus stems immediately after submersion in river and pond water, in the absence or presence of 500 /-Lg L-1 and 4000 /-Lg L-1 tetracycline. Saponification in a hot water bath for 30 mins followed by toluene extraction was the most ideal method to extract ergosterol from environmental samples. HPLCUV detections of ergosterol, compared to detections by mass spectrometry, were found to be reliable with all matrices except water. Fungal decomposition began and was the greatest in the spring despite low water temperatures. There was no significant difference in biomass or production on aerial versus submerged portions of Scirpus. Water temperature was correlated to fungal production (r = 0.7, P < 0.005) for aerial stem pieces but not for submerged pieces. However, in laboratory experiments water temperature had a measurable effect on both biomass and production in submerged stem pieces. With respect to Scirpus decaying under UV+ or UV- in pond water, there was no significant difference in either fungal biomass (P = 0.76) or production (P = 0.96) between the two treatments. There were significant differences (P<0.05) in the adsorption of tetracycline between distilled, river and pond waters. Half-lives of tetracycline were significantly shorter (P<0.05) in the light than in the dark and in sterile waters compared to nonsterile waters (P<0.05) and also were significantly different (P<0.05) between the three waters in all experiments. Tetracycline photolysis experiment conducted in natural sunlight showed there were significant differences (P<0.05) between the UV+ and UVlight treatments as well as between the three different waters (P<0.05) in both light treatments. Tetracycline bound to the matrix did not undergo photolysis suggesting tetracycline will probably persist in aquatic environments. Tetracycline significantly (P<0.05) inhibited protein production in planktonic bacteria at concentrations ~ 10 Ilg L-1 in river water but the inhibition was considerable (P<0.05) only at 4000 Ilg L-1 in pond water. Submersion of dry Scirpus stems in river and pond water resulted in dramatic increases in microbial biomass on the stems that peaked about 10 days after immersion. The increase in biomass coincided with the colonization of the external surface of the stem and the decrease with the exhaustion of this substrate. In treatments where tetracycline was added there was a significant decreases (P<0.05) in microbial biomass between the control and other treatments in the river experiment but not in pond water due to large variances in data (P>0.05). Biofilms tended to be thicker in the 4000 Ilg L-1 tetracycline treatments in both water experiments suggesting the high concentration of tetracycline may have induced polymer formation so as to limit the toxicity of tetracycline. Fungal biomass tended to be higher at 4000 Ilg L-1 compared to the control treatments in both water experiments suggesting that in normal conditions bacteria exerted an inhibitory influence on fungi. Total microbial biomass on the stems was greater in the pond water experiment compared to the river water experiment, possibly because of the higher DOC and POM in the pond water, but fungal biomass was greater in the river system compared to the wetland system indicating that fungal dominance may vary in different water ecosystems.