Browsing by Subject "Denitrifikation"

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Community Structure and Activity of Nitrate-Reducing Microorganisms in Soils under Global Climate Change
(2006) Deiglmayr, Kathrin; Kandeler, Ellen
Since the beginning of the Industrial Revolution, atmospheric carbon dioxide concentrations have been steadily increasing and, thus, contributed to a warming of the climate and altered biogeochemical cycles. To study the response of soil microorganisms to altered environmental conditions under global climate change, the nitrate-reducing community was regarded as a model community in the present thesis. This functional group, which performs the first step in the denitrification pathway, was selected because it is phylogenetically very diverse. In particular rising levels of atmospheric carbon dioxide as the most important catalyst of temperature rise and the retreat of glaciers in the Alps as one of the most evident consequences of climate change were investigated. The behaviour of nitrate reducers was investigated in a biphasic approach: (i) at the level of its enzyme activity of the nitrate reductase and (ii) at the level of community structure, which was characterised by RFLP (Restriction Fragment Length Polymorphism)-fingerprints using the functional gene narG. The effect of elevated atmospheric carbon dioxide concentrations on nitrate-reducing micro-organisms was studied in the Swiss FACE (Free Air Carbon dioxide Enrichment) experiment including the rhizosphere of two functional plant types (Lolium perenne and Trifolium repens), two N fertilisation levels and two sampling dates (June and October 2002). Whereas in June no significant treatment effect was observed, the nitrate reductase activity proved to be significantly reduced under elevated atmospheric carbon dioxide at the autumn sampling date. Simultaneously, elevated enzyme activities were recorded under Trifolium repens and high N fertilisation pointing to a control of nitrate reductase activity by nitrate availability at the time of sampling. The community structure of nitrate reducers, however, showed a different response pattern with sampling date and the strongly varying pH of the different experimental plots constituting the main driving factors. With respect to the three experimental factors atmospheric carbon dioxide, plant type and N fertilisation the composition of the nitrate reducers revealed a high stability. The microbial succession of nitrate-reducing microorganisms was studied in the rhizosphere of Poa alpina across the glacier foreland of the Rotmoosferner/Oetz valley. Sampling was performed in August and at the end of the short period of vegetation in September. The nitrate reductase activity increased significantly with progressing successional age, whereas organic carbon together with nitrate concentrations in the soils explained the major part of this effect. The microbial community of nitrate reducers revealed a significant shift across the glacier foreland, with pH and organic carbon representing the most important environmental factors inducing this shift. A detailed analysis of the clone libraries that were constructed for the youngest and the oldest site in the glacier foreland pointed to the tendency of lower diversity in the late succession compared to the young succession. Possibly an increasing selective pressure due to higher densities of microorganisms and, hence, a higher competition for limited resources contributed to the decline in diversity. In conclusion, the functional group of nitrate reducers responded to changing environmental conditions under global climate change particularly through altered enzyme activities. The amount and the direction of this response depended strongly on the nitrate availability and the organic carbon content in soils. The community structure of nitrate-reducing microorganisms, however, proved to be resilient towards short-term substrate fluctuations. This indicates that the genetic pool of this group of soil microorganisms possesses a high functional stability characterized by a relatively persistent composition and an independent modulation of enzyme activity.
Effect of reduced nitrogen deposition on microbial activity, abundance and diversity in forest soils
(2012) Enowashu, Esther Eneckeh; Kandeler, Ellen
The deposition of nitrogen has increased many-fold due to anthropogenic activities. Since forest ecosystems are often limited by N availability, elevated N inputs from the atmosphere can have a fertilization effect but in the long-term, excess N can influence above- and below-ground production. One of the consequences of N deposition and increased N inputs is a shift in microbial community structure and function as ecosystems move towards N saturation. Soil microorganisms through the action of enzymes play an important role in N dynamics. Thus, the availability and turnover of N depends strongly on microbial abundance, diversity and activity which are in turn influenced by soil properties. Studies on the effects of high nitrogen inputs and the response of forest ecosystems to nitrogen saturation are many and well understood. However, the reversibility of N-induced shifts in forest ecosystem processes is largely unknown. This thesis was therefore designed to study the response of soil microorganisms to reduced N deposition. A biphasic approach was employed to look into (i) the general microbial functional status of the Solling forest site as well as (ii) the microbial community structure which may be a key regulator of two important processes of N transformation: denitrification and proteolysis. The goal of the present thesis was addressed in three studies. Denitrification is considered sensitive to environmental changes and the response of nitrate-reducers and denitrifiers to reduced N deposition was determined in the first study. The goal of the second study was to investigate the overall microbial activity of the Solling forest profiles especially focussing on enzymes involved in the N cycle. This revealed a pronounced activity of peptidases whereby a set of novel pepN primers encoding alanine aminopeptidase enzyme was designed in the third study to determine the group of bacteria involved in proteolysis in forest as well as agricultural and grassland soils. The Solling experimental station was established more than two decades ago and it gave the opportunity to study the N cycle in a natural forest ecosystem at different sampling dates and depths. A combination of classical biological methods and modern molecular techniques were used in the studies. Soil physico-chemical parameters (OC, Nt, NO3-, NH4+, pH, % Water content) were analysed to gain more information on mineralization and immobilization of N in the soil profiles. The analysis of microbial biomass, ergosterol content and the activity of several enzymes of the N, C and P cycles as well as enzyme activity of nitrate reducers was determined in order to interpret microbial functions. The abundance of nitrate reducers and denitrifiers were determined by quantitative PCR of 16S rRNA, nitrate reductase (narG and napA) and denitrification (nirK, nirS and nosZ) genes. The diversity of peptide degrading bacteria was analysed by PCR, cloning and sequencing and the construction of pepN gene libraries. The results of the first study indicated that time and space were the main drivers influencing the abundance and activity of the nitrate reducers and denitrifier communities in the forest soil profiles. Reduced N deposition had a of minimal effect. Interestingly, the ratios of nosZ to16S rRNA gene and nosZ to nirK increased with soil depth thereby tempting to conclude that the size of denitriﬁers capable of reducing N2O into N2 might be bigger in the mineral horizons. In the second study, a stronger response of N cycling enzymes to reduced N deposition could be seen. However, these responses especially that of specific peptidases differed in magnitude which could be indicative of a modification of the reaction rates of the different N cycling enzymes. Correlation of nutrients (N, C, P) with microbial biomass and enzyme activities in the soil profiles revealed that substrate availability was the main factor influencing microbial activity. In the third study, analyses of gene libraries from extracted DNA from forest, agricultural and glacier soil samples revealed a high diversity of pepN sequences related to mainly α-Proteobacteria. A majority of the sequences showed similarity to published data revealing that the amplified region of pepN might be conserved. Linking diversity and enzymatic data, lowest diversity was observed in the agricultural soil where activity levels of alanine aminopeptidase were lowest indicating the importance of diversity studies for ecosystem functioning. In conclusion, this thesis offers valuable contributions to understanding the impact of N deposition. The approach used was suitable to assess the response of the different microbial communities to reduced N deposition. The magnitude of the response depended strongly on space, time and substrate availability in soils as well as their interactions.
Influence of land use on abundance, function and spatial distribution of N-cycling microorganisms in grassland soils
(2015) Keil, Daniel; Kandeler, Ellen
This thesis focuses on the influence of land use on the abundance, function and spatial distribution of N-cycling microorganisms in grassland soils, but also on soil biogeochemical properties, as well as on enzyme activities involved in the carbon-, nitrogen-, and phosphorous cycle. The objective of this thesis was tackled in three studies. All study sites that were investigated as part of this thesis were preselected and assigned according to study region and land use within the framework of the “Exploratories for Functional Biodiversity Research – The Biodiversity Exploratories” of the Deutsche Forschungsgemeinschaft priority program 1374. The first study addressed the question whether land-use intensity influences soil biogeochemical properties, as well as the abundance and spatial distributions of ammonia-oxidizing and denitrifying microorganisms in grasslands of the Schwäbische Alb. To this end, a geostatistical approach on replicated grassland sites (10 m × 10 m), belonging to either unfertilized pastures (n = 3) or fertilized mown meadows (n = 3), representing low and high land-use intensity, was applied. Results of this study revealed that land-use intensity changed spatial patterns of both soil biogeochemical properties and N-cycling microorganisms at the plot scale. For soil biogeochemical properties, spatial heterogeneity decreased with higher land-use intensity, but increased for ammonia oxidizers and nirS-type denitrifiers. This suggests that other factors, both biotic and abiotic than those measured, are driving the spatial distribution of these microorganisms at the plot scale. Furterhmore, the geostatistical analysis indicated spatial coexistence for ammonia oxidizers (amoA ammonia-oxidizing archaea and amoA ammonia-oxidizing bacteria) and nitrate reducers (napA and narG), but niche partitioning between nirK- and nirS-type denitrifiers. The second study aimed at whether land-use intensity contributes to spatial variation in microbial abundance and function in grassland ecosystems of the Schwäbische Alb assigned to either low (unfertilized pastures, n = 3), intermediate (fertilized mown pastures, n = 3), or high (fertilized mown meadows, n = 3) land-use intensity. Plot-scale (10 m × 10 m) spatial heterogeneity and autocorrelation of soil biogeochemical properties, microbial biomass and enzymes involved in C, N, and P cycle were investigated using a geostatistical approach. Geostatistics revealed spatial autocorrelations (p-Range) of chemical soil properties within the maximum sampling distance of the investigated plots, while greater variations of p-Ranges of soil microbiological properties indicated spatial heterogeneity at multiple scales. An expected decrease in small-scale spatial heterogeneity in high land-use intensity could not be confirmed for microbiological soil properties. Finding smaller spatial autocorrelations for most of the investigated properties indicated increased habitat heterogeneity at smaller scales under high land-use intensity. In the third study, the effects of warming and drought on the abundance of denitrifier marker genes, the potential denitrification activity and the N2O emission potential from grassland ecosystems located in the Schwäbische Alb, the Hainich, and the Schorfheide region were investigated. Land use was defined individually for each grassland site by a land-use index that integrated mowing, grazing and fertilization at the sites over the last three years before sampling of the soil. It was tested if the microbial community response to warming and drought depended on more static site properties (soil organic carbon, water holding capacity, pH) in interaction with land use, the study region and the climate change treatment. It was further tested to which extent the N2O emission potential was influenced by more dynamic properties, e.g. the actual water content, the availability of organic carbon and nitrate, or the size of the denitrifier community. Warming effects in enhanced the potential denitrification of denitrifying microorganisms. While differences among the study regions were mainly related to soil chemical and physical properties, the land-use index was a stronger driver for potential denitrification, and grasslands with higher land use also had greater potentials for N2O emissions. The total bacterial community did not respond to experimental treatments, displaying resilience to minor and short-term effects of climate change. In contrast, the denitrifier community tended to be influenced by the experimental treatments and particularly the nosZ abundance was influenced by drought. The results indicate that warming and drought affected the denitrifying communities and the potential denitrification, but these effects are overruled by study region and site-specific land-use index. This thesis gives novel insights into the performance of N-cycling microorganisms in grassland ecosystems. The spatial distribution of soil biogeochemical properties is strongly dependent on land-use intensity, as in return is the spatial distribution of nitrifying and denitrifying microorganisms and the ecosystem services they perform. Yet, future work will be necessary to fully understand the interrelating factors and seasonal variability, which influence the ecosystem functioning and ecosystem services that are provided by N-cycling soil microorganisms at multiple scales.
Linking microbial abundance and function to understand nitrogen cycling in grassland soils
(2017) Regan, Kathleen Marie; Kandeler, Ellen
This thesis characterized spatial and temporal relationships of the soil microbial community, the nitrogen cycling microbial community, and a subset of the nitrogen cycling community with soil abiotic properties and plant growth stages in an unfertilized temperate grassland. Unfertilized perennial grasslands depend solely on soil-available nitrogen and in these environments nitrogen cycling is considered to be both highly efficient and tightly coupled to plant growth. Unfertilized perennial grasslands with high plant diversity, such as ours, have also been shown to have higher soil organic carbon, total nitrogen, and microbial carbon; greater food web complexity; and more complex biological communities than more intensively managed grasslands or croplands. This made the choice of study plot especially well-suited for characterizing the relationships we sought to identify, and made it possible to detect spatial and temporal patterns at a scale that has heretofore been under-examined. The first study used a combination of abiotic, plant functional group, and PLFA measurements together with spatial statistics to interpret spatial and temporal changes in the microbial community over a season. We found that its overall structure was strongly related to the abiotic environment throughout the sampling period. The strength of that relationship varied, however, indicating that it was not constant over time and that other factors also influenced microbial community composition. PLFA analysis combined with principal components analysis made it possible to discern changes in abundances and spatial distributions among Gram-positive and Gram-negative bacteria as well as saprotrophic fungi. Modeled variograms and kriged maps of the changes in distributions of exemplary lipids of both bacterial groups also showed distinct differences in their distributions on the plot, especially at stages of most rapid plant growth. Although environmental properties were identified as the main structuring agents of the microbial community, components of those environmental properties varied over the season, suggesting that plant growth stage had an indirect influence, providing evidence of the complexity and dynamic nature of the microbial community in a grassland soil. The second study took the same analytical approach, this time applying it to abundances of key members of the soil nitrogen cycling community. Marker genes for total archaea and bacteria, nitrogen fixing bacteria, ammonia oxidizing archaea and bacteria, and denitrifying bacteria were quantified by qPCR. Potential nitrification activity and denitrifying enzyme activity were also determined. We found clear seasonal changes in the patterns of abundance of the measured genes and could associate these with changes in substrate availability related to plant growth stages. Most strikingly, we saw that small and ephemeral changes in soil environmental conditions resulted in changes in these microbial communities, while at the same time, process rates of their respective potential enzyme activities remained relatively stable. This suggests both short term niche-partitioning and functional redundancy within the nitrogen cycling microbial community. The seasonal changes in abundances we observed also provided additional evidence of a dynamic relationship between microorganisms and plants, an important mechanism controlling ecosystem nitrogen cycling. The third study determined spatial and temporal interactions between AOA, AOB and NOB. These steps are related in both space and time, as the ammonia-oxidizers provide the necessary substrate for nitrite-oxidizers. Using a combination of spatial statistics and phylogenetic analysis, our data indicated seasonally varying patterns of niche differentiation between the two bacterial groups, Nitrospira and Nitrobacter in April, but more homogeneous patterns by August which may have been due to different strategies for adapting to changes in substrate concentrations resulting from competition with plants. We then asked a further question: was the microbial structure at sampling sites with high NS gene abundances fundamentally different from those with low NS gene abundances? Using a phylogenetic approach, the operational taxonomic unit composition of NS was analyzed. Community composition did not change over the first half of the season, but by the second half, the relative proportion of a particular OTU had increased significantly. This suggested an intraspecific competition within the NS and the possible importance of OTU 03 in nitrite oxidation at a specific period of time. Observed positive correlations between AOA and Nitrospira further suggested that in this unfertilized grassland plot, the nitrification process may be predominantly performed by these groups, but is restricted to a limited timeframe.
Ein Vergleich zwischen Barometrischer Prozessseparation (BaPS) und 15N-Verdünnungsmethode zur Bestimmung der Bruttonitrifikationsrate im Boden
(2010) Schwarz, Ulrich; Streck, Thilo
Besides the carbon cycle, the nitrogen cycle plays a central role in soil. A key process of this cycle is nitrification. In practice, nitrification is measured as gross or net nitrification. Net nitrification rates are measured by determining the net change in the nitrate or ammonium pool over a period of time. Net rates are difficult to interpret, because the net nitrification rate is the sum of nitrate producing and consuming processes. In contrast, gross nitrification quantifies the total production of nitrate via nitrification. There are two methods for measuring gross nitrification: the 15N-Pool dilution technique and Barometric Process Separation (BaPS). In the 15N-Pool dilution technique, nitrate en-riched with the heavier isotope 15N is added to soil, and the dilution of the 15N pool and the change in the nitrate pool are measured over time. The BaPS method measures changes in pressure and the oxygen- and carbon dioxide concentration of the atmosphere in a closed chamber. The gross nitrification rate can then be computed by a step-by-step solution of the gas balance equations. In the present study, 15N enriched nitrate was added to soil and then put into the BaPS-incubation chamber. By this procedure gross nitrification rates were measured simultaneously with both the 15N-Pool dilution technique and the BaPS method. The aim of the present study was to find out under which conditions the two methods yield similar results and under which conditions different results. In the latter case, the thesis aimed at elucidating the cause for the disagreement between both methods. For this purpose extensive research on two agricultural soils from North China and three soils from Southwest Germany was undertaken. The two methods were compared under the following conditions: 1) application of ammonium fertilizer, 2) addition of nitrification inhibitors, 3) varying soil wa-ter contents, and 4) different soil temperatures. Moreover, a new methodological approach was tested: the 13CO2-Pool dilution technique. Combining this method with the 15N-Pool dilu-tion technique and the Barometric Process Separation made it possible to exactly determine the pH and respiration coefficient in situ. Both techniques corresponded well in soil with pH<6. In soil with higher pH, both methods led to very different results. The reason is that pH has a strong impact on the calculation of the nitrification rate in the BaPS method. In nearly all experiments with neutral to alkaline soils, the BaPS technique yielded higher nitrification rates than the 15N-Pool dilution technique if pH was determined in 0.01 M CaCl2. With pH determined in water, there was good agreement or nitrification rates were too low. Fertilization with ammonium did not in-duce an increase of nitrification in a sandy soil with pH<6. A decrease in nitrification to less than 60% was achieved by the application of the nitrification inhibitor DCD. For both techniques a positive correlation between temperature and nitrification rates was found. There was no correlation between water filled pore space and nitrification rate.