Browsing by Subject "Ammoniumverbindungen"

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Developing indicators and characterizing direct and residual effects of biological nitrification inhibition (BNI) by the tropical forage grass Brachiaria humidicola
(2018) Karwat, Hannes; Cadisch, Georg
Nitrogen (N) losses from agroecosystems harm the environment via increased nitrate (NO3-) amounts in water-bodies and nitrous oxide (N2O) emissions to the atmosphere. Bacteria and archaea oxidize ammonium (NH4+) to NO3- under aerobic conditions. Furthermore, under mainly anaerobic conditions, microbial denitrification reduces NO3- to gaseous N forms. The tropical forage grass Brachiaria humidicola (Rendle) Schweick (Bh) has been shown to reduce soil microbial nitrification via root derived substances. Therefore, biological nitrification inhibition (BNI) by Bh might contribute to reduction of N losses from agroecosystems. The present doctoral thesis aimed at assessing the potential of the actual BNI by Bh, as well as the residual BNI effect with new developed methodologies. The overall research was based on the following major objectives: (1) characterization of the residual BNI effect by Bh on recovery of N by subsequent cropped maize (Zea mays L.) under different N fertilization rates; (2) investigate if low enzymatic nitrate reductase activity (NRA) in leaves of Bh is linked to reduced NO3- nutrition by effective BNI; (3) identify a possible link between plant delta 15N of Bh and the BNI effect of different Bh genotypes on nitrification, plant N uptake and NO3- leaching losses. The overall objective was to use and test new methodologies with a minimum of disturbance of the plant-soil system, to characterize BNI of different Bh genotypes in greenhouse and field studies. The first research study focused on the investigation of a potential residual BNI effect of a converted long-term Bh pasture on subsequent maize cropping, where a long-term maize monocrop field served as control. The residual BNI effect was characterized in terms of enhanced maize grain yield, total N uptake and 15N (labeled) fertilizer recovery. Furthermore, the impact of residual BNI effect on soil N dynamics was investigated. The residual BNI effect was confirmed for the first maize crop season after pasture conversion on the basis of lower nitrification in incubation soil, higher total N uptake and higher maize grain yields. However, the residual BNI effect did not result in higher 15N fertilizer uptake or reduced 15N fertilizer losses, nor in reduced N20 emissions. Applied N was strongly immobilized due to long-term root turnover effects, while a significant residual BNI effect from Bh prevented re-mineralized N from rapid nitrification resulting in improved maize performance. A significant residual Bh BNI effect was evident for less than one year only. In the second research study it was the aim to verify the potential of nitrate reductase activity (NRA) as a proxy for the detection of in vivo performance of BNI by selected Bh accessions and genotypes grown under contrasting fertilization regimes. NRA was detected in Bh leaves rather than in roots, regardless of NO3- availability. Leaf NRA correlated with NO3- contents in soils and stem sap of contrasting Bh genotypes substantiating its use as a proxy of in vivo performance of BNI. The leaf NRA assay facilitated a rapid screening of contrasting Bh genotypes for their differences in in vivo performance of BNI under field and greenhouse conditions; but inconsistency of the BNI potential by selected Bh genotypes was observed. The third research study emphasized to link the natural abundance of delta 15N in Bh plants with reduced NO3- losses and enhanced N uptake due to BNI. Increased leached NO3- was positively correlated to rising delta 15N in Bh grass, whereas the correlation between plant N uptake and plant delta 15N was inverse. Long-term field cultivation of Bh decreased nitrification in incubated soil, whereas delta 15N of Bh declined and plant N% rose over time. Delta 15N of Bh correlated positively with assessed nitrification rates in incubated soil. It was concluded that decreasing delta 15N of Bh over time reflects the long-term effect of BNI linked to lower NO3- formation and reduced NO3- leaching, and that generally higher BNI activity of Bh is indicated by lower delta 15N plant values. Within the framework of this thesis, a residual BNI effect by Bh on maize cropping could be confirmed for one season due to the combined methodological approaches of soil incubation and 15N recovery. The development of the NRA assay for sampled Bh leaves was validated as a rapid and reliable method linked to the actual soil nitrification after NH4+ fertilizer supply. Consequently, the assay could be used for both greenhouse and field studies as BNI proxy. The gathered data from the third study indicated that decreasing delta 15N of Bh over time reflects the long-term effect of BNI linked to lower NO3- formation and reduced NO3- leaching, and that generally higher BNI activity of Bh is indicated by lower delta 15N plant values. Consequently, it was suggested that delta 15N of Bh could serve as an indicator of cumulative NO3- losses. Overall, this doctoral thesis suggests the depressing effect on nitrification by Bh might be a combined effect by BNI and fostered N immobilization. Furthermore, BNI by Bh might be altered by different factors such as soil type, plant age and root morphology of the genotypes. Finally, future studies should consider that Bh genotypes express their respective BNI potential differently under contrasting conditions.
Plant ammonium transporter (AMT) integration in regulatory networks
(2016) Straub, Tatsiana; Ludewig, Uwe
Ammonium is a ubiquitous key nutrient in agricultural soils and the preferred nitrogen source for plants. However, excessive ammonium accumulation represses plant growth and development. Ammonium is taken up by plant cells via high-affinity ammonium transporters (AMTs). Six AMT genes were identified in Arabidopsis, which are separated in two distinct clades, five AMT1s and one AMT2. In the plasma membrane, AMT proteins form homo- and heterotrimers with extra-cytoplasmic N-termini and cytoplasmic C-termini. In addition to transcriptional and post-transcriptional control of AMTs by ammonium, phosphorylation in the C-terminus serves as a rapid allosteric switch of the AMT activity and prevents further internal ammonium accumulation. In a physiological screen, a kinase (CIPK23) was identified, which directly regulates ammonium transport activity under high-NH4+ conditions. Interestingly, CIPK23 is already known to regulate nitrate and potassium uptake in roots. Lesion of the CIPK23 gene significantly increased ammonium uptake, but caused growth inhibition. As expected, cipk23 plants were also limited in potassium accumulation, but high potassium availability failed to rescue the cipk23 phenotype. Furthermore, cipk23 plants were more susceptible to methylammonium (MeA), a non-metabolizable analogue of ammonium. The sensitivity to MeA was lost upon genetic suppression of AMT1 genes in the cipk23 background. The data suggest that CIPK23 directly phosphorylates AMT1s in a complex with CBL1 (calcineurin B-like protein) and thereby regulates transport activity. The expression of the CIPK23 and the CBL1 genes were ammonium-dependent and increased when N-starved plants were resupplied with ammonium. Furthermore, cbl1 mutants had enhanced NH4+ accumulation; this phenocopies the larger ammonium uptake in the cipk23 loss-of-function mutant. In vivo experiments demonstrated bimolecular interaction between CIPK23, AMT1;1, and AMT1;2, but not with AMT2;1, suggesting direct phosphorylation of AMT1-type ammonium transporters by CIPK23. However, Western blot analysis with the cipk23 mutant suggested that the loss of the kinase was not sufficient to completely abolish AMT1;1 and AMT1;2 phosphorylation, indicating several independent pathways to regulate ammonium transport activity in AMT trimers. The data identify complex post-translational regulation of ammonium transporters via the CBL1–CIPK23 pathway, which ensures reduction of AMT1 activity and suppression of ammonium uptake under high external NH4+ concentrations.
Transcriptional and proteomic responses towards early nitrogen depletion in Arabidopsis thaliana
(2016) Menz, Jochen; Ludewig, Uwe
Plant roots acquire nitrogen predominantly as ammonium and nitrate, which besides serving as nutrients, also have signaling roles. Re-addition of nitrate to starved plants rapidly and di-rectly transcriptionally re-programs the metabolism and induces root architectural changes, but the earliest responses to nitrogen deprivation are unknown. In this thesis, the early transcriptional response of developed roots to nitrate or ammonium deprivation were analyzed in two Arabidopsis ecotypes contrasting in their nitrogen use efficiency: the inefficient genotype Col-0 and the efficient Tsu-0. The rapid transcriptional repression of known nitrate-induced genes proceeded the tissue NO3- concentration drop, with the transcription factor genes LBD37/38 and HRS1/HHO1 among those with earliest significant change. Some transcripts were stabilized by nitrate, but similar rapid transcriptional repression occurred in loss-of-function mutants of the nitrate response factor NLP7. In contrast, an early transcriptional response to ammonium deprivation was almost completely absent. In Col-0, the analysis was extended with the proteome and phospho-proteome resulting in a rapid and transient perturbation of the proteome induced by ammonium deprivation and a differential phosphorylation pattern in proteins involved in adjusting the pH and cation homeostasis, plasma membrane H+, NH4+, K+ and water fluxes. Fewer differential phosphorylation patterns in transporters, kinases and other proteins occurred with nitrate deprivation. The deprivation responses are not just opposite to the resupply responses, identify NO3--deprivation induced mRNA decay and signaling candidates potentially reporting the external nitrate status to the cell. Transcrip-tome comparison revealed only few N-nutrition related genes between both ecotypes contributing the increased NUE of Tsu-0, which probably relies on higher biomass accumulation. Besides, Tsu-0 confirmed the transcriptional depletion response of Col-0.
Treatment of benzene and ammonium contaminated groundwater using microbial electrochemical technology and constructed wetlands
(2018) Wei, Manman; Seifert, Jana
With the rapid development of modern industry, energy shortage and environmental pollution are getting more and more serious. Groundwater pollution is one of the most important problems. A multitude of remediation techniques in situ or ex situ have been used to treat contaminated groundwater. This thesis was to investigate whether groundwater contaminated mainly by benzene and ammonium can be remediated by constructed wetlands in combination with microbial electrochemical technology. The objectives of this thesis are (i) to develop and test systems for removing pollutants and simultaneously recovering energy from contaminated groundwater, (ii) to maximize the benefits of both constructed wetland and microbial electrochemical technology while treating contaminated groundwater, (iii) to elucidate the underlying electrochemical reactions and pollutant degradation pathways, and (iv) to investigate microbial active species and functional proteins involved in benzene degradation and ammonium removal. A microbial fuel cell (MFC) equipped with an aerated cathode and a control without aeration at the cathode were designed to remove benzene and ammonium from contaminated groundwater collected in the Leuna site (Saxony-Anhalt, Germany). The performance of pollutant removal and electricity generation was investigated and compared in the two reactors. Electrochemical processes occurring in the MFC were determined by benzene and ammonium spiking experiments as well as oxygen interruption experiments. Additionally, the biodegradation pathways and dominant organisms were elucidated by compound specific stable isotope analysis (CSIA) and Illumina sequencing. The results indicated the principal feasibility of treating benzene and ammonium contaminated groundwater by a MFC equipped with an aerated cathode. Benzene (~15 mg/L) was completely removed in the MFC, of which 80% disappeared already at the anoxic anode. Ammonium (~20 mg/L) was oxidized to nitrate at the cathode; this reaction was not directly linked to electricity generation. The maximum power density was 316 mW/m3 net anoxic compartment (NAC) at a current density of 0.99 A/m3 NAC. Coulombic and energy efficiencies of 14% and 4% were obtained based on the anodic benzene degradation. Benzene was initially activated by enzymatic monohydroxylation at the oxygen-limited anode; the further anaerobic oxidation of the intermediate metabolites released electrons accompanied by electrons transfer to the anode. Dominant phylotypes at the MFC anode revealed by 16S rRNA Illumina sequencing were affiliated to the Chlorobiaceae, Rhodocyclaceae and Comamonadaceae, presumably associated with benzene degradation. Nitrification took place at the aerated cathode of the MFC and was catalyzed by phylotypes belonging to the Nitrosomonadales and Nitrospirales. The control reactor failed to generate electricity, although phylotypes affiliated to the Chlorobiaceae, Rhodocyclaceae and Comamonadaceae were dominant as well; the control reactor can be thus regarded as a mesocosm in which granular graphite was colonized by benzene degraders, but showed a lower benzene removal efficiency compared to the MFC. In order to enhance benzene and ammonium removal while simultaneously harvesting energy, a constructed wetland integrated with microbial electrochemical technology (MET-CW) was established by embedding four anode modules into the sand bed and connecting it to a cathode placed in the open pond inside a bench-scale horizontal subsurface flow constructed wetland (HSSF-CW). Compared with the control CW, enhanced benzene and ammonium removal efficiencies were found in the MET-CW. The electrochemical performances of anode modules located at the four different depths were compared; the results showed that anode modules located in the deep layer (Module 3 and 4) had the relatively high power densities whereas the power densities located in the upper layer (modules 1 and 2) were extremely low. The initial activity mechanism of benzene degradation was analyzed by CSIA. Ammonium removal processes were assessed using nitrogen isotope fractionation of ammonium. Functional proteins and active microbial species involved in nitrogen transformation processes were detected using protein-based stable isotope probing (protein-SIP) with in situ feeding of 15N-NH4+. Additionally, potential denitrification and anammox rates were measured using Nitrogen isotope tracing. The results demonstrated that benzene and ammonium removal in a CW can be improved by combination with microbial electrochemical technology. The enhanced benzene removal was linked to the use of the anode modules as electron acceptor, whereas efficient ammonium removal was probably attributed to the elimination of inhibition effects by the co-contaminant benzene. Benzene was initially activated by monohydroxylation, forming intermediates which were subsequently oxidized accompanied by extracellular electron transfer, leading to current production. Partial nitrification accompanied by either heterotrophic denitrification or nitrifier-denitrification was mainly responsible for NH4+-N removal in the MET-CW, whereas anammox played a minor role. However, the contribution of anammox was markedly increased at the location near to the anode modules. In summary, this research indicated that microbial electrochemical technology can be used to improve the performance of pollutant removal while simultaneously harvesting energy from contaminated groundwater. Especially, the combination of MET with other traditional treatment approaches (e.g. constructed wetland) is a promising alternative to treat contaminated water.