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Organic matter composition in the subsoil: Contribution of root litter and microbial-derived compounds

The aim of P2 within the Research Unit 'The Forgotten Part of Carbon Cycling: Organic Matter Storage and Turnover in Subsoils (SUBSOM)' is to contribute to the understanding of the different sources and stabilization processes of subsoil organic matter. This will be achieved by the analysis of the soil organic matter composition in topsoil versus subsoil by 13C NMR spectroscopy in bulk soils as well as organo-mineral associations. This will be done on a number of soil profiles differing in parent material and mineralogy and therefore also in the relevance of organo-mineral associations for subsoil C stabilization. In addition, a specific sampling approach will allow to differentiate three zones associated with the dominating effect of (1) leaching of DOC (the 'bulk soil' between trees), (2) root litter decomposition (the 'root-affected zone'), and (3) direct rhizodeposition of root exudates (the 'rhizosphere' sensu strictu). The contribution of above-ground versus below-ground litter is differentiated by the analysis of cutin and suberin biomarkers. Organic matter derived from microbial sources will be identified by the microbial signature of polysaccharides in the subsoil through the analysis of neutral sugars and amino sugars. Organo-mineral associations will be further characterized by N2-BET analyses to delineate the coverage of the mineral phase with organic matter. With these analyses and our specific analytical expertise at the submicron scale (nanoSIMS) we will participate in selected joint experiments of the research unit.

Rehabilitation of Degraded Forests in Yunnan (German-Chinese Cooperation for Agrarian Research)

Background: An increasing frequency of massive flooding along the lower Yangtse River in China ended in a disastrous catastrophe in summer 1998 leaving several thousand people homeless, more than 3.600 dead and causing enormous economic damage. Inappropriate land-use techniques and large scale timber felling in the water catchment of the upper Yangtse and its feeder streams were stated to be the main causes. Immediate timber cutting bans were imposed and investigations on land use patterns were initiated by the Chinese Government. The Institute for World Forestry of the Federal Research Centre for Forestry and Forest Products was approached by the Yunnan Academy of Forestry in Kunming to exchange experiences and to cooperate scientifically in the design and application of appropriate afforestation and silvicultural management techniques in the water catchment area of the Yangtse. This cooperation was initiated in 1999 and is based on formal agreements in the fields of agrarian research between the German and Chinese Governments. Objectives: The cooperation was in the first step focussing on the identification of factors which caused the enormous floodings. After their identification measures of prevention were determined and put into practice. In this context experiences made in past centuries in the alpine region of central Europe served as an incentive and example for similar environmental problems and solutions under comparable conditions. Relevant key questions of the cooperation project were: - Analysis of forest related factors influencing the recent floodings of the Yangtse, - Analysis and evaluation of silvicultural management experiences from central Europe for know-how transfer, - Evaluation of rehabilitation measures for successful application in Yunnan, - Dissemination of knowledge through vocational training. Results: - Frequent wild grazing of husbandry is a key factor for forest degeneration beyond unsustainable timber harvests, forest fires and insect calamities leading to increased water run-off in the mountainous region of Yunnan; - Browsing of cattle interrupts succession thus avoiding natural regeneration and leaving a logging ban ineffective; - Mountain pasture in the Alps had similar effects in the past in central Europe. The introduction of controlled grazing has led to an ecologically compatible coexistence of pasture and ecology. Close-to-nature forestry can have positive effects in this sensitive environment. - Afforestation with site adopted broadleaves and coniferous tree species was implemented on demonstration level using advanced techniques in Yunnan.

Biopores in the subsoil: Formation, nutrient turnover and effects on crops with distinct rooting systems (BioFoNT)

Perennial fodder cropping potentially increases subsoil biopore density by formation of extensive root systems and temporary soil rest. We will quantify root length density, earthworm abundance and biopore size classes after Medicago sativa, Cichorium intybus and Festuca arundinacea grown for 1, 2 and 3 years respectively in the applied research unit's Central Field Trial (CeFiT) which is established and maintained by our working group. Shoot parameters including transpiration, gas exchange and chlorophyll fluorescence will frequently be recorded. Precrop effects on oilseed rape and cereals will be quantified with regard to crop yield, nutrient transfer and H2-release. The soil associated with biopores (i.e. the driloshpere) is generally rich in nutrients as compared to the bulk soil and is therefore supposed to be a potential hot spot for nutrient acquisition. However, contact areas between roots and the pore wall have been reported to be low. It is still unclear to which extent the nutrients present in the drilosphere are used and which potential relevance subsoil biopores may have for the nutrient supply of crops. We will use a flexible videoscope to determine the root-soil contact in biopores. Nitrogen input into the drilosphere by earthworms and potential re-uptake of nitrogen from the drilosphere by subsequent crops with different rooting systems (oilseed rape vs. cereals) will be quantified using 15N as a tracer.

Barley dwarfs acting big in agronomy. Identification of genes and characterization of proteins involved in dwarfism, lodging resistance and crop yield

Barley (Hordeum vulgare) is an important cereal grain which serves as major animal fodder crop as well as basis for malt beverages or staple food. Currently barley is ranked fourth in terms of quantity of cereal crops produced worldwide. In times of a constantly growing world population in conjunction with an unforeseeable climate change and groundwater depletion, the accumulation of knowledge concerning cereal growth and rate of yield gain is important. The Nordic Genetic Resource Center holds a major collection of barley mutants produced by irradiation or chemical treatment. One phenotypic group of barley varieties are dwarf mutants (erectoides, brachytic, semidwarf, uzu). They are characterized by a compact spike and high rate of yield while the straw is short and stiff, enhancing the lodging resistance of the plant. Obviously they are of applied interest, but they are also of scientific interest as virtually nothing is known about the genes behind the development of plant dwarfism. The aim of this project is to identify and isolate the genes carrying the mutations by using state of the art techniques for gene cloning at the Carlsberg Laboratory. The identified genes will be connected with the mutant phenotype to reveal the gene function in general. One or two genes will be overexpressed and the resulting recombinant proteins will be biochemically and structurally characterized. The insights how the mutation effects the protein will display the protein function in particular. Identified genes and their mutant alleles will be tested in the barley breeding program of the Carlsberg brewery.

Dissolved organic matter driven changes in minerals and organic-mineral interactions during paddy soil development

Previous studies indicated that the development and biogeochemistry of paddy soils relates to the parent material, thus the original soil paddies derive from. The proposed research focuses on redox-mediated changes in mineral composition and mineral-associated organic matter (OM) during paddy transformation of different soils. We plan to subject soil samples to a series of redox cycles, in order to mimic paddy soil formation and development. Soils with strongly different properties and mineral composition as well as at different states of paddy transformation; ranging from unchanged soils to fully developed paddy soils, are to be included. We hypothesize that dissolved organic matter is one key driver in redox-mediated transformations, serving as an electron donator as well as interacting with dissolved metals and minerals. The extent of effects shall depend on the parent soil's original mineral assemblage and organic matter and their mutual interactions. The experimental paddy soil transformation will tracked by analyses of soil solutions, of the (re-)distribution of carbon (by addition of 13C-labelled rice straw), of indicative biomolecules (sugars, amino sugars, fatty acids, lignin) and of minerals (including the redox state of Fe). For analyses of organic matter as well as of mineral characteristics we plan to utilize EXAFS and XPS, for Fe-bearing minerals also Mößbauer spectroscopy. This approach of experimental pedology seems appropriate to give insight into the major factors during paddy soil formation and development.

The effect of elevated atmospheric CO2 concentration on gross nitrogen dynamics, plant N-uptake and microbial community dynamics in a permanent grassland

To predict ecosystem reactions to elevated atmospheric CO2 (eCO2) it is essential to understandthe interactions between plant carbon input, microbial community composition and activity and associated nutrient dynamics. Long-term observations (greater than 13 years) within the Giessen Free Air Carbon dioxide Enrichment (Giessen FACE) study on permanent grassland showed next to an enhanced biomass production an unexpected strong positive feedback effect on ecosystem respiration and nitrous oxide (N2O) production. The overall goal of this study is to understand the long-term effects of eCO2 and carbon input on microbial community composition and activity as well as the associated nitrogen dynamics, N2O production and plant N uptake in the Giessen FACE study on permanent grassland. A combination of 13CO2 pulse labelling with 15N tracing of 15NH4+ and 15NO3- will be carried out in situ. Different fractions of soil organic matter (recalcitrant, labile SOM) and the various mineral N pools in the soil (NH4+, NO3-, NO2-), gross N transformation rates, pool size dependent N2O and N2 emissions as well as N species dependent plant N uptake rates and the origin of the CO2 respiration will be quantified. Microbial analyses will include exploring changes in the composition of microbial communities involved in the turnover of NH4+, NO3-, N2O and N2, i.e. ammonia oxidizing, denitrifying, and microbial communities involved in dissimilatory nitrate reduction to ammonia (DNRA). Stable Isotope Probing (SIP) and mRNA based analyses will be employed to comparably evaluate the long-term effects of eCO2 on the structure and abundance of these communities, while transcripts of these genes will be used to target the fractions of the communities which actively contribute to N transformations.

Origin and fate of dissolved organic matter in the subsoil

Dissolved organic matter (DOM) is one major source of subsoil organic matter (OM). P5 aims at quantifying the impact of DOM input, transport, and transformation to the OC storage in the subsoil environment. The central hypotheses of this proposal are that in matric soil the increasing 14C age of organic carbon (OC) with soil depth is due to a cascade effect, thus, leading to old OC in young subsoil, whereas within preferential flowpaths sorptive stabilization is weak, and young and bioa-vailable DOM is translocated to the subsoil at high quantities. These hypotheses will be tested by a combination of DOC flux measurements with the comparative analysis of the composition and the turnover of DOM and mineral-associated OM. The work programme utilizes a DOM monitoring at the Grinderwald subsoil observatory, supplemented by defined experiments under field and labora-tory conditions, and laboratory DOM leaching experiments on soils of regional variability. A central aspect of the experiments is the link of a 13C-leaf litter labelling experiment to the 14C age of DOM and OM. With that P5 contributes to the grand goal of the research unit and addresses the general hypotheses that subsoil OM largely consists of displaced and old OM from overlying horizons, the sorption capacity of DOM and the pool size of mineral-associated OM are controlled by interaction with minerals, and that preferential flowpaths represent 'hot spots' of high substrate availability.

Ecotoxicology of Organotin compounds

Organotin and especially butyltin compounds are used for a variety of applications, e.g. as biocides, stabilizers, catalysts and intermediates in chemical syntheses. Tributyltin (TBT) compounds exhibit the greatest toxicity of all organotins and have even been characterized as one of the most toxic groups of xenobiotics ever produced and deliberately introduced into the environment. TBT is not only used as an active biocidal compound in antifouling paints, which are designed to prevent marine and freshwater biota from settlement on ship hulls, harbour and offshore installations, but also as a biocide in wood preservatives, textiles, dispersion paints and agricultural pesticides. Additionally, it occurs as a by-product of mono- (MBT) and dibutyltin (DBT) compounds, which are used as UV stabilizer in many plastics and for other applications. Triphenyltin (TPT) compounds are also used as the active biocide in antifouling paints outside Europe and furthermore as an agricultural fungicide since the early 1960s to combat a range of fungal diseases in various crops, particularly potato blight, leaf spot and powdery mildew on sugar beet, peanuts and celery, other fungi on hop, brown rust on beans, grey moulds on onions, rice blast and coffee leaf rust. Although the use of TBT and TPT was regulated in many countries world-wide from restrictions for certain applications to a total ban, these compounds are still present in the environment. In the early 1970s the impact of TBT on nontarget organisms became apparent. Among the broad variety of malformations caused by TBT in aquatic animals, molluscs have been found to be an extremely sensitive group of invertebrates and no other pathological condition produced by TBT at relative low concentrations rivals that of the imposex phenomenon in prosobranch gastropods speaking in terms of sensitivity. TBT induces imposex in marine prosobranchs at concentrations as low as 0,5 ng TBT-Sn/L. Since 1993, for the littorinid snail Littorina littorea a second virilisation phenomenon, termed intersex, is known. In female specimens affected by intersex the pallial oviduct is transformed of towards a male morphology with a final supplanting of female organs by the corresponding male formations. Imposex and intersex are morphological alterations caused by a chronic exposure to ultra-trace concentrations of TBT. A biological effect monitoring offers the possibility to determine the degree of contamination with organotin compounds in the aquatic environment and especially in coastal waters without using any expensive analytical methods. Furthermore, the biological effect monitoring allows an assessment of the existing TBT pollution on the basis of biological effects. Such results are normally more relevant for the ecosystem than pure analytical data. usw.

Carbon and Chorine Isotope Effect Study to Investigate Chlorinated Ethylene Dehalogenation Mechanisms

Chlorinated ethylenes are prevalent groundwater contaminants. Numerous studies have addressed the mechanism of their reductive dehalogenation during biodegradation and reaction with zero-valent iron. However, despite insight with purified enzymes and well-characterized chemical model systems, conclusive evidence has been missing that the same mechanisms do indeed prevail in real-world transformations. While dual kinetic isotope effect measurements can provide such lines of evidence, until now this approach has not been possible for chlorinated ethylenes because an adequate method for continuous flow compound specific chlorine isotope analysis has been missing. This study attempts to close this prevalent research gap by a combination of two complementary approaches. (1) A novel analytical method to measure isotope effects for carbon and chlorine. (2) A carefully chosen set of well-defined model reactants representing distinct dehalogenation mechanisms believed to be important in real-world systems. Isotope trends observed in biotic and abiotic environmental dehalogenation will be compared to these model reactions, and the respective mechanistic hypotheses will be confirmed or discarded. With this hypothesis-driven approach it is our goal to elucidate for the first timdehalogenation reactions.

Emmy Noether-Nachwuchsgruppen, Development and risk assessment of transgenic environmentally-friendly insect pest control methods for fruit flies and mosquitoes

Various species of pest insects cause substantial damage to agriculture every year, or transmit deadly diseases to animals and humans. A successful strategy to control pest insect populations is based on the Sterile Insect Technique (SIT), which uses the release of mass-reared, radiation sterilized male insects to cause infertile matings and thus reduce the pest population level. However, irradiation is not applicable to every insect species. Thus, new strategies based on genetic modifications of pest insects have been developed or are currently under investigation.The goal of the proposed research is to improve the development and ecological safety of genetically engineered (GE) insects created for enhanced biological control programs, including the SIT and new strategies based on conditional lethality. A major concern for GE insect release programs is transgene stability, and maintenance of their consistent expression. Transgene loss or intra-genomic movement could result in loss of strain attributes, and may ultimately lead to interspecies movement resulting in ecological risks. To address potential transgene instability, a new transposon vector that allows post-integration immobilization will be tested in the Mediterranean, Mexican and Oriental fruit fly tephritid pest species. In addition, the system will be established in the mosquito species Aedes and Anopheles - carriers of dengue and malaria.Random genomic insertion is also problematic for GE strain development due to genomic position effects that suppress transgene expression, and insertional mutations that negatively affect host fitness and viability. Diminished transgene expression could result in the unintended survival of conditional lethal individuals, or the inability to identify them. To target transgene vectors to defined genomic insertion sites having minimal negative effects on gene expression and host fitness, a recombinase-mediated cassette exchange (RMCE) strategy will be developed that. RMCE will also allow for stabilization of the target site, will be tested in tephritid and mosquito species, and will aid to the development of stabilized target-site strains for conditional lethal biocontrol. This will include a molecular and organismal evaluation of an RNAi-based lethality approach. Lethality based on an RNAi mechanism in the proposed insects would increase the species specificity and having multiple targets for lethality versus one target in existing systems. By seeking to improve transgene expressivity and stabilization of transposon-based vector systems, this proposal specifically addresses issues related to new GE insects by reducing their unintended spread after field release, and by limiting the possibilities for transgene introgression.

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