The major aim of this study was to evaluate the relationships between the rates of microbial activities (phytoplankton primary production, bacterial secondary production, bacterial utilization of organic matter, enzymatic activities, protozoan grazing on bacteria), bacterial numbers, and dissolved organic carbon concentrations and the trophic state index (TSI) of lakes in the upper trophogenic water layer in the pelagial zone along the trophic gradient (from oligo/mesotrophy to hypereutrophy) in 19 lakes of the Mazurian Lake District (northeastern Poland). Multiple regression analysis (analysis of variance-ANOVA) on all collected data and the TSI along eutrophication gradient showed that all studied microbial processes and parameters were very tightly coupled to the trophic conditions of the studied lakes. All studied microbial processes involved in utilization and enzymatic degradation of organic matter were strongly positively dependent on the intensity and rates of photosynthetic organic matter production and exudation that markedly increased along the eutrophication gradient of lakes. V max of alkaline phosphatase, aminopeptidase, and nonspecific esterase showed significant correlation with the TSI of the studied lakes. Protozoans removed a significant portion of bacterial production, i.e., from ϳ20% to 75-85% of newly produced bacterial biomass was simultaneously consumed by protozoans along the eutrophication gradient. These observations suggest that the importance of protozoan grazing on bacteria on regulation of bacterial production depends on lake productivity. The general working hypothesis that the intensity of microbial processes of organic matter can be tightly coupled to increasing eutrophication was proven in these studies.

We present the results of an analysis of the 16S rRNA-based taxonomical structure of bacteria together with an analysis of carbon source utilization ability using EcoPlate (Biolog, USA) metabolic fingerprinting assessment against the backdrop of physicochemical parameters in fifteen interconnected lakes. The lakes exhibit a wide spectrum of trophic gradients and undergo different intensities of anthropopressure. Sequences of V3–V4 16S rRNA genes binned by taxonomic assignment to family indicated that bacterial communities in the highly eutrophicated lakes were distinctly different from the bacterial communities in the meso-eutrophic lakes (ANOSIM r = 0.99, p = 0.0002) and were characterized by higher richness and more diverse taxonomical structure. Representatives of the Actinobacteria , Proteobacteria , Cyanobacteria , Planctomycetes , Verrucomicrobia , Bacteroides phyla predominated. In most cases their relative abundance was significantly correlated with lake trophic state. We found no similar clear relationship of community-level physiological profiling with lake trophic state. However, we found some significant links between the taxonomic and metabolic structure of the microbes in the studied lakes (Mantel’s correlation r = 0.22, p = 0.006). The carbon source utilization ability of the studied microorganisms was affected not only by the taxonomic groups present in the lakes but also by various characteristics like a high PO 4 3− concentration inhibiting the utilization of phosphorylated carbon.

A method for determining enzymatically hydrolyzable phosphate (EHP) in natural waters is described, based on the determination of released inorganic phosphate after the hydrolysis of organophosphoric esters by free, dissolved phosphohydrolases (mainly phosphatase) produced by the biota. The method gives higher values in highly eutrophic waters than the classical procedure of Strickland and Parsons; in samples from less eutrophic habitats the two methods give similar results. The new method is simple, accurate, and can be used in both freshwater and marine studies. It is particularly recommended for determination in hypereutrophic waters, when Pi concentrations exceed 25 µg P liter−1. Representative field data from the euphotic zone of seven lakes and two marine habitats are presented.

This paper discusses the methodology for determination of dissolved DNA concentration by means of direct DAPI staining of water samples and compares it with the data obtained by the method of dissolved DNA precipitation (in 0.2 pm water filtrates) with the use of cetyltrimethyl-ammonium bromide (CTAB) and DAPI staining. The samples were collected from lakes of varying trophic states. Enzymatically hydrolysable DNA (EH-DNA) was estimated as the difference between the concentration of the DNA in samples without and with DNAse treatment. Concentrations of enzymatically hydrolysable DNA determined by the enzymatic method were 27 to 54 % lower than those measured by CTAB-DNA precipitation and DAPI staining. Enzymatically hydrolysable DNA concentrations increased with the trophic state of the lake and correlated positively with algal pigment concentrations and bacterial numbers. The contribution of phosphorus that can be enzymatically liberated from extracellular DNA to the total organic phosphorus concentration in lake water samples varied from 11 % (oligo/mesotrophic lake) to 27.6% (hypertrophic lake).

In this paper we d~s c u s s the sultablllty of the cetyltrimethylammonium bromlde (CTAB) precipitation method for the determination of dissolved DNA (dDNA) In lake water. Factors affecting dDNA recovery from the precipitate and further DAPI fluorometr~c assay were also examined. The proposed method was particularly suitable for routine determination of dDNA concentration in low and moderately eutrophicated freshivater environments The detection 11m1t of the method v a n e d from 1.5 pg dDNA 1. ' for mesotrophlc waters to 2 5 pg dDNA I-' for e u t r o p h~c lakes. Preliminary observatlons on the concentration of dDNA in the surface water of 8 southern German lakes showed that dDNA concentrations were generally related to their trophic status and varied between 2.5 and 46.0 pg I-' in mesotrophlc and from 11.5 to 72 pg I-' in eutrophlc waters KEY WORDS. Dissolved D N A . Lake w a t e r . CTAB . DAPI

ABSTRACT. Concentrations and origin of dissolved DNA (dDNA) were studied in 14 lakes in the Mazurian Lake District (Northern Poland) and 7 lakes in Southern Germany during the summer stratification period. dDNA concentration varied markedly (from 0.5 to 70 p g 1.' ) in the studied lakes. We suggest that this dissolved organic matter fraction can be potentially one of the most important P and N sources for planktonic microorganisms. Laboratory experiments and field observations suggested that eukaryotic microorganisms (algae) are the most lrnportant sources of dDNA in lake water, whereas bacteria mainly decompose the dDNA pool.

This study analyzes proteolytic enzyme persistence and the role of dead (or metabolically inactive) aquatic bacteria in organic matter cycling. Samples from four lakes of different trophic status were used. Irrespective of the trophic status of the examined lakes, bacterial aminopeptidases remained active even 72 h after the death of the bacteria that produced them. The total pool of proteolytic enzymes in natural lake water samples was also stable. We found that the rates of amino acid enzymatic release from proteinaceous matter added to preserved lake water sample were constant for at least 96 h (r(2)  = 0.99, n = 17, P ≤ 0.0001, V(max)  = 84.6 nM h(-1) ). We also observed that proteases built into bacterial cell debris fragments remained active for a long time, even after the total destruction of cells. Moreover, during 24 h of incubation time, about 20% of these enzymatically active fragments adsorbed onto natural seston particles, becoming a part of the 'attached enzymes system' that is regarded as the 'hot-spot' of protein degradation in aquatic ecosystems.

Growing anthropopressure over the last several decades has resulted in rapid progressive eutrophication of the Great Masurian Lakes (GML) system located in northeastern Poland. In our studies, we investigated whether there is a relationship between the occurrence of pathogenic bacteria: Legionella spp. and Aeromonas spp., not explored so far in the waters of GML system, and the trophic status of the studied lakes. The GML system of glacial origin includes lakes connected by natural and artificial channels, and it extends from north to south for approximately 100 kilometers. Water samples were taken during the summer, subsequently spring and autumn seasons from 15 lakes in land-water ecotones. At all sampling sites, basic in situ measurements of physicochemical parameters were recorded. The amounts of chlorophyll a, nitrogen, phosphorus, dissolved organic carbon were also measured. The trophic state index (TSI) of the sampling sites was also estimated. The real-time PCR technique enabled the determination of the presence and abundance of Legionella spp. and Aeromonas spp. The results clearly showed that several environmental water quality parameters, associated with eutrophication, and among them: nitrogen, phosphorus, chlorophyll, ammonium concentration, conductivity, turbidity, water transparency, highly affected the presence and abundance of the detected pathogenic bacteria in the studied lakes. Special attention should be paid to the high impact of water eutrophication on the number of pathogenic microorganisms, which result both from human activities in lakes and climate change.