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Browsing by Subject "Bakterien"

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Bio-effectors for improved growth, nutrient acquisition and disease resistance of crops
(2017) Weinmann, Markus; Neumann, Günter
Recent scientific approaches to sustain agricultural production in face of a growing world food demand, limited natural resources, and ecological concerns have been focusing on biological processes to support soil fertility and healthy plant growth. In this context, the use of “bio-effectors”, comprising living (micro-) organisms and active natural compounds, has been receiving increasing attention. In contrast to conventional fertilizers and pesticides, the effectiveness of “bio-effectors” is essentially not based on the substantial direct input of mineral plant nutrients, neither in inorganic nor organic forms, nor of a-priori toxic compounds. Their direct or indirect effects on plant performance are rather based on the functional implementation or activation of biological mechanisms, in particular those interfering with soil-plant-microbe interactions. The general objective of the present research work was to improve the empirical and conceptual understanding concerning the utilization of bio-effectors in agricultural practice, following the principles of plant growth stimulation, bio-fertilization and bio-control. One main aspect of investigation was the application of bio-effectors to improve the efficiency of phosphorus (P) acquisition by the plant. Promising bio-preparations based on microbial inoculants (e.g. Bacillus, Pseudomonas, Trichoderma species) as well as natural compounds (e.g. algae extracts, humic acids) were tested in screening assays, greenhouse, and field experiments to characterize their potential effectiveness under varying environmental conditions. The most significant effects on plants appeared under severely low phosphate availability, but even under controlled conditions, bio-effectors required a narrow range of conductive environmental settings to reveal their potential effectiveness. Another focus of research was the application of bio-effectors to control soil borne pathogens, which typically appear in unsound crop rotations. Emphasis was set on take-all disease in wheat induced by the fungus Gaeumannomyces graminis. While the effectiveness of oat precrops to control take-all in subsequent wheat has been attributed to microbial changes and enhanced manganese (Mn) availability in soils, the take-all fungus is known to decrease the availability of Mn by oxidation. Against this background, the effectiveness of oat precrops and alternative crop management strategies to improve the Mn status and suppress the severity of take-all in wheat was investigated under controlled and field conditions. In conclusion, none of the tested supplemental treatments, such the application of microbial bio-effectors, stabilized ammonium or manganese fertilizers, could fully substitute for the multiple effectiveness of oat precrops, which was further confirmed by the results of a field experiment. Finally, some general conclusions and perspectives are summarized. Selected bio-effectors showed a strong capacity to improve the nutrient acquisition and healthy growth of crop plants under controlled conditions, but not in field experiments. However, even under controlled conditions the strongest effects occurred when plants were exposed to abiotic or biotic stresses, such as severely limited P availability or pathogen infestation of the soil substrate, still restricting plant growth to unproductive levels. Facing this situation, there is no perspective to improve the field efficiency of promising bio-effectors applications as a stand-alone approach. The only chance to develop viable alternatives to the conventional use of fertilizers or pesticides, for an ecological intensification of agriculture that maintains high yield levels, seems to be a reasonable integration of bio-effectors into the whole crop management of sound agricultural practice.
Bio-effectors for improved growth, nutrient acquisition and disease resistance of crops.- 2nd unrevised edition
(2019) Weinmann, Markus; Madora GmbH, Luckestr.1, D-79539 Lörrach; Raupp, Manfred G.
Recent scientific approaches to sustain agricultural production in face of a growing world food demand, limited natural resources, and ecological concerns have been focusing on biological processes to support soil fertility and healthy plant growth. In this context, the use of “bio-effectors”, comprising living (micro-) organisms and active natural compounds, has been receiving increasing attention. In contrast to conventional fertilizers and pesticides, the effectiveness of “bio-effectors” is essentially not based on the substantial direct input of mineral plant nutrients, neither in inorganic nor organic forms, nor of a-priori toxic compounds. Their direct or indirect effects on plant performance are rather based on the functional implementation or activation of biological mechanisms, in particular those interfering with soil-plant-microbe interactions. The general objective of the present research work was to improve the empirical and conceptual understanding concerning the utilization of bio-effectors in agricultural practice, following the principles of plant growth stimulation, bio-fertilization and bio-control. One main aspect of investigation was the application of bio-effectors to improve the efficiency of phosphorus (P) acquisition by the plant. Promising bio-preparations based on microbial inoculants (e.g. Bacillus, Pseudomonas, Trichoderma species) as well as natural compounds (e.g. algae extracts, humic acids) were tested in screening assays, greenhouse, and field experiments to characterize their potential effectiveness under varying environmental conditions. The most significant effects on plants appeared under severely low phosphate availability, but even under controlled conditions, bio-effectors required a narrow range of conductive environmental settings to reveal their potential effectiveness. Another focus of research was the application of bio-effectors to control soil borne pathogens, which typically appear in unsound crop rotations. Emphasis was set on take-all disease in wheat induced by the fungus Gaeumannomyces graminis. While the effectiveness of oat precrops to control take-all in subsequent wheat has been attributed to microbial changes and enhanced manganese (Mn) availability in soils, the take-all fungus is known to decrease the availability of Mn by oxidation. Against this background, the effectiveness of oat precrops and alternative crop management strategies to improve the Mn status and suppress the severity of take-all in wheat was investigated under controlled and field conditions. In conclusion, none of the tested supplemental treatments, such the application of microbial bio-effectors, stabilized ammonium or manganese fertilizers, could fully substitute for the multiple effectiveness of oat precrops, which was further confirmed by the results of a field experiment. Finally, some general conclusions and perspectives are summarized. Selected bio-effectors showed a strong capacity to improve the nutrient acquisition and healthy growth of crop plants under controlled conditions, but not in field experiments. However, even under controlled conditions the strongest effects occurred when plants were exposed to abiotic or biotic stresses, such as severely limited P availability or pathogen infestation of the soil substrate, still restricting plant growth to unproductive levels. Facing this situation, there is no perspective to improve the field efficiency of promising bio-effectors applications as a stand-alone approach. The only chance to develop viable alternatives to the conventional use of fertilizers or pesticides, for an ecological intensification of agriculture that maintains high yield levels, seems to be a reasonable integration of bio-effectors into the whole crop management of sound agricultural practice.
Biochar amendment for C sequestration in a temperate agroecosystem : implications for microbial C- and N-cycling
(2018) Bamminger, Chris; Kandeler, Ellen
Climate warming will have great impact on terrestrial ecosystems. Different soil properties such as temperature and moisture will be altered, thereby influencing C- and N-cycles, microbial activity as well as plant growth. This may contribute to the observed increase in soil greenhouse gas (GHG) emissions under climate change. Therefore, new options are needed to mitigate theses projected consequences. Biochar is primarily suggested to be effective in long-term C sequestration in agricultural soils due to its long-term stability. In addition, it could be applied to improve various soil properties, plant growth and to reduce soil GHG emissions. To date, knowledge about such beneficial biochar effects in soil under predicted warming climate is extremely scarce. In the first study, a slow-pyrolysis biochar from Miscanthus x giganteus feedstock (600 °C, 30 Min.) was incubated for short time (37d) under controlled laboratory conditions in agricultural soil in the presence of earthworms and N-rich litter (Phacelia tanacetifolia Benth.). Biochar increased microbial abundances and the fungal-to-bacterial PLFA ratio after 37 days in arable soil applied with litter suggesting improved living conditions for microorganisms with biochar. Fungi may benefit most from newly created habitats due to colonizable biochar pores and surfaces. Additionally, fungi could have also mineralized small amounts of recalcitrant biochar-C during plant litter decomposition. Without litter, biochar led to interactions between earthworms and soil microorganisms resulting in enhanced bacterial and fungal abundances. This indicates better growth habitats for soil microbes in earthworm casts containing biochar. Soil respiration and metabolic quotients (qCO2) and N2O emissions (in litter treatments) were decreased after biochar application suggesting a more efficient microbial community and underscoring the GHG mitigation potential of the used biochar. The field experiment, investigated in the second and third study, focused on the stability and long-term soil C sequestration potential of comparable Miscanthus biochar (850 °C, 30 Min.). Related effects on soil GHG emissions, physical, chemical and microbiological soil properties as well as plant growth were determined in an agroecosystem at year-round elevated soil temperature (+2.5 °C, since 2008). The second study investigated the short-term effects of biochar on microbial abundances and growth of winter rapeseed during the first year after field application to a warmed temperate arable soil. It was found that fungal biomass and the fungal-to-bacterial ratio were increased in the warmed biochar plots only after three months in the presence of spring barley litter from the previous growing season. The disappearance of this effect points to an overall high stability of the investigated biochar. Moreover, biochar proved to be effective in mitigating negative effects of seasonal dryness on microbial abundances and early plant growth in the dry spring period in 2014. However, biochar had no effect on final aboveground biomass of winter rapeseed at harvest in the first growing season. As shown in the third study, after two vegetation periods of winter rapeseed and spring wheat, the assumption that plant productivity in already fertile temperate arable soils is unlikely to be further enhanced with biochar amendment, was confirmed. Total CO2 emissions after two years were not reduced by biochar and remained unchanged even under warming suggesting a high degradation stability of the used biochar. N2O emissions were increased in biochar-amended soil at elevated soil temperature, presumably due to enhanced water and fertilizer retention with biochar. By using the global warming potential (GWP100) of total soil GHG emissions, the storage of biochar-C in soil was estimated to compensate warming-induced elevated soil GHG emissions for 20 years. To conclude, this thesis revealed that biochar may have only minor influence on soil microorganisms and crop growth in temperate, fertile arable field soils. However, it was shown that biochar could be a valuable tool for C sequestration in temperate arable soils, thus potentially offsetting a warming-induced increase in GHG emissions. In order to face climate change impacts, more long-term studies on microbiological effects and the C sequestration potential of biochar in cultivated soil under warming are urgently needed.
Bioeffector products for plant growth promotion in agriculture : modes of action and the application in the field
(2021) Weber, Nino Frederik; Neumann, Günter
Modern agriculture faces a conflict between sustainability and the demand for a higher food production. This conflict is exacerbated by climate change and its influence on vegetation, ecology and human society. To reduce land use, the reduction of yield losses and food waste is crucial. Moreover a sustainable intensification is necessary to increase yields, while at the same time input of limited resources such as drinking water or fertilizer should be kept as low as possible. This might be achieved by improving nutrient recycling and plant resistance to abiotic or biotic stress. Bioeffectors (BE) comprise seaweed or plant extracts and microbial inoculums that may stimulate plant growth by phytohormonal changes and increase plant tolerance to abiotic stress (biostimulants), solubilize or mobilize phosphorus from sparingly soluble sources such as Al/Fe or Ca-phosphates in the soil, rock phosphates, recycling fertilizer or organic phosphorus sources like phytate (biofertilizer), or improve plant resistance against pathogens by induced-systemic resistance (ISR) or antibiosis (biocontrol). For this study, in total 18 BE products were tested in germination, pot and field experiments for their potential to improve plant growth, cold stress tolerance, nutrient acquisition and yield in maize and tomato. Additionally, a gene expression analysis in maize was performed using whole transcriptome sequencing (RNA-Seq) after the application of two potential plant growth promoting rhizobacteria (PGPR), the Pseudomonas sp. strain DSMZ 13134 “Proradix” and the Bacillus amyloliquefaciens strain FZB42. Seaweed products supplemented with high amounts of the micronutrients Zn and Mn were effective in reducing detrimental cold stress reactions in maize whereas microbial products and seaweed extracts without micronutrient supplementation failed under the experimental conditions. At optimal temperature the product containing the Pseudomonas sp. strain was repeatedly able to stimulate root and shoot growth of maize plants whereas in tomato only in heat-treated soil substrate significant effects were observed. Results indicate that the efficacy of the product was mainly attributed to stimulation or shifts in the soil microbial community. Additionally, the FZB42 strain was able to stimulate root and plant growth in some experiments whereas the effects were less reproducible and more sensitive to environmental conditions. Fungal BE products were less effective in plant growth stimulation and showed detrimental effects in some experiments. Under the applied experimental conditions BE-derived plant growth stimulation mainly was attributed to biostimulation but aspects of biofertilization or biocontrol cannot be excluded, as all experiments were conducted in non-sterile soil substrates. Root and shoot growth are stimulated in response to hormonal shifts. In the gene expression analysis only weak responses to BE treatments were observed, as previously reported from other studies conducted under non-sterile conditions. Nevertheless, some plant stress responses were observed that resembled in some aspects those reported for phosphorus (P) deficiency in others those reported for ISR/SAR. Especially the activation of plant defence mechanisms, such as the production of secondary metabolites, ethylene production and reception and the expression of several classes of stress-related transcription factors, including JA-responsive JAZ genes, was observed. It also seems probable that in plants growing in PGPR-drenched soils, especially at high application rates, a sink stimulation for assimilates triggers changes in photosynthetic activity and root growth leading to an improved nutrient acquisition. Nevertheless, due to the complexity of interactions in natural soil environments as well as under practice conditions, a designation of a distinct mode of action for plant growth stimulation by microbial BEs is not realistic. A comparison of the overall results with those reported in literature or other working groups in a common research project (“Biofector”) supported the often-reported low reproducibility of plant growth promotion effects by BE products under applied conditions. Factors that influenced BE efficacy were application time and rates, temperature, soil buffer capacity, phosphorus sources and nitrogen fertilization, light conditions and the soil microbial community. Results indicate that in maize cultivation seed treatment is the most economic application technique for microbial products whereas for vegetable or high-value crops with good economic benefit soil drenching is recommended. For seaweed extracts foliar application seems to be the most economic and efficient choice. Furthermore, results emphasize the importance of a balanced natural soil microflora for plant health and yield stability.
Biological regulation of subsoil C-cycling
(2019) Preußer, Sebastian; Kandeler, Ellen
Soils are the largest terrestrial reservoir of organic carbon (OC). Substantial proportions of the stored OC are found in stabilized form in deeper soil layers. Beside the quality and quantity of C input from plant biomass, C storage in soil is primarily controlled by the microbial decomposition capacity. Various physical, chemical and biological factors (e.g., substrate availability, temperature, water content, pH, texture) vary within soil profiles and directly or indirectly influence the abundance, composition and activity of microbial communities and thus the microbial C turnover. While soil microbiological research has so far focused mainly on processes in topsoil, the mechanisms of C storage and turnover in subsoil are largely unknown. The objective of the present thesis was therefore to investigate the specific influence of substrate availability and different environmental factors as well as their interactions on microbial communities and their regulatory function in subsoil C-cycling. This objective was addressed in three studies. In the first and second study, one-year field experiments were established in which microbial communities from different soil depths were exposed to altered habitat conditions to identify crucial factors influencing the spatial and temporal development of microbial abundance and substrate utilization within soil profiles. This was achieved by reciprocal translocation of soils between subsoil horizons (first study) and topsoil and subsoil horizons (second study) in combination with addition of 13C-labelled substrates and different sampling dates. In the third study, a flow cascade experiment with soil columns from topsoil and subsoil horizons and soil minerals (goethite) coated with 13C-labelled organic matter (OM) was established. This laboratory experiment investigated the importance of exchange processes of OM with reactive soil minerals for the quality and quantity of dissolved OM and the influence of these soil micro-habitats on microbial abundance and community composition with increasing soil depth. In the first study, the reciprocal translocation of subsoils from different soil depths revealed that due to comparable micro-climatic conditions and soil textures within the subsoil profile, no changes in microbial biomass, community composition and activity occurred. Moreover, increasing microbial substrate utilization in relation to the quantity of added substrate indicated that deep soil layers exhibit high potential for microbial C turnover. However, this potential was constrained by low soil moisture in interplay with the coarse soil texture and the resulting micro-scale fragmentation of the subsoil environment. The bacterial substrate utilization was more affected by this spatial separation between microorganisms and potentially available substrate than that of fungi, which was further confirmed by the translocation experiment with topsoil and subsoil in the second study. While the absolute substrate utilization capacity of bacteria decreased from the more moist topsoil to the drier subsoil, fungi were able to increase their substrate utilization and thus to partially compensate the decrease in C input from other sources. Furthermore, the addition of root litter as a preferential C source of fungal decomposer communities led to a pronounced fungal growth in subsoil. The third study demonstrated the high importance of reactive soil minerals both in topsoil and in subsoil for microbial growth due to extensive exchange processes of OM and the associated high availability of labile C. In particular copiotrophic bacteria such as Betaproteobacteria benefited from the increased C availability under non-limiting water conditions leading to a pronounced increase in bacterial dominance in the microbial communities of these soil micro-habitats. In conclusion, this thesis showed that subsoil exhibits great potential for both bacterial and fungal C turnover, albeit this potential is limited by various factors. This thesis, however, allowed to determine the specific effects of these factors on bacteria and fungi and their function in subsoil C-cycling and thus to identify those factors of critical importance. The micro-climate in subsoil, in particular soil moisture, was the primary factor limiting bacterial growth and activity, whereas fungi were more strongly restricted by substrate limitations.
Effects of resource availability and quality on soil microorganisms and their carbon assimilation
(2014) Kramer, Susanne; Kandeler, Ellen
Soil microorganisms play a pivotal role in decomposition processes and therefore influence nutrient cycling and ecosystem function. Availability and quality of resources determines activity, growth and identity of substrate users. In agricultural systems, availability of resources is dependent on, for example, crop type, management, season, and depth. At depth substrate availability and microbial biomass decrease. However, there remain gaps in our understanding of C turnover in subsoil and how processes in the topsoil may influence abundance, activity, and function of microorganisms in deeper soil layers. With respect to substrate quality it is thought that bacteria are the dominant users of high quality substrates and more labile components whereas fungi are more important for the degradation of low quality and more recalcitrant substrates (i.e. cellulose, lignin). Therefore, this thesis was designed to increase our understanding of C turnover and the influence of both availability and quality of substrates on microorganisms in an agricultural soil. In the first and second studies, a recently established C3-C4 plant exchange field experiment was used to investigate the C flow from belowground (root) and aboveground (shoot litter) resources into the belowground food web. Maize plants were cultivated to introduce a C4 signal into the soil both by plant growth (belowground / root channel) and also by applying shoot litter (aboveground litter channel). To separate C flow from the shoot litter versus the root channel, maize litter was applied on wheat cultivated plots, while on half of the maize planted plots no maize litter was returned. Wheat cultivated plots without additional maize litter application served as a reference for the calculation of incorporated maize-C into different soil pools. Soil samplings took place in two consecutive years in summer, autumn and winter. Three depths were considered (0-10 cm: topsoil, 40-50 cm: rooted zone beneath the plough layer, 60-70 cm: unrooted zone). In the third study a microcosm experiment with substrates of different recalcitrance and complexity was carried out to identify primary decomposers of different plant litter materials (leaves and roots) during early stages of decomposition (duration of 32 days) and to follow the C flow into the next higher trophic level (protozoa).
Evaluation and method development for the biosynthesis of microbial lipopeptides by bacillus species
(2023) Vahidinasab, Maliheh; Hausmann, Rudolf
Microbial lipopeptides are secondary metabolites produced by bacteria and single-celled microorganisms. They consist of a cyclic or linear peptide chain linked to a lipid residue. Due to their high-foaming biosurfactant properties, they have various industrial applications such as in detergents, food emulsifiers, bioremediation, and enhanced oil recovery. Additionally, they possess other functional properties such as antifungal activity, making them an environmentally friendly alternative to synthetic fertilizers and fungicides. Bacillus species produce cyclic lipopeptides known for their potent antifungal activity, which makes them a potential source of bio-fungicides in agriculture. However, the production titer of wild-type Bacillus species does not meet industrial needs. Thereby, genetic modification of producer strains and bioprocess engineering can help increase the production of lipopeptides. Nevertheless, the regulation and basis of biosynthesis for Bacillus lipopeptides are still not completely understood, and ongoing research aims to enhance their production. In general, three main lipopeptide families, including surfactins, iturins, and fengycins are produced by different Bacillus species. Among these, surfactin as the strong biosurfactant is the most extensively studied lipopeptide produced by Bacillus species. The focus of this doctoral thesis was mainly to evaluate the biosynthesis of iturin and fengycin families, which are strong antimicrobial lipopeptides produced by Bacillus subtilis and Bacillus velezensis. This involved developing strains through genetic engineering and enhancing the lipopeptide titer by evaluating the cultivation medium. Initially, the entire genome of the bacteria used in this thesis was examined in terms of lipopeptide biosynthesis, and the structure and yield of the different produced lipopeptides were analyzed. Regarding the lipopeptide producer derivatives of the domesticated laboratory model strain B. subtilis 168 and B. subtilis 3NA, a spore deficient strain appropriate for bioreactor cultivation, surfactin is the lipopeptide with the highest yield, while plipastatin which is a member of fengycin family, is produced in lower quantities. In the present thesis, the biosynthesis of plipastatin by B. subtilis BMV9 as the lipopeptide producer derivative of strain 3NA was evaluated. The study aimed to convert BMV9 to a constitutive plipastatin mono-producer strain. In this sense, overexpressing plipastatin biosynthesis operon using the stronger constitutive Pveg promoter led to a five-fold increase in plipastatin production. Interestingly, it was observed that deletion of srfAA-AD operon in BMV9 and the constructed constitutive plipastatin producer strain has not improved plipastatin production. Therefore, it can be stated that presumably the biosynthesis of plipastatin may be positively influenced in a post-transcriptional manner by the surfactin synthetase or some of its subunits. However, the regulatory mechanism behind this effect remained unknown and requires further research. Another attempt to enhance the plipastatin biosynthesis in strain BMV9 was repairing the degQ expression. One main genome characterization of strains with B. subtilis 168 and 3NA background is that the pleiotropic degQ gene expression, which is known to have a positive effect on plipastatin biosynthesis, is silenced due to a mutation in the promoter area. However, while repair of degQ expression in BMV9 increased the plipastatin production, combination of both repaired degQ expression and promoter exchange (Ppps::Pveg) has not significantly increased the plipastatin yield. To further evaluate the impact of degQ expression on surfactin and plipastatin biosynthesis, two strains of B. subtilis were selected: JABs24, a lipopeptide producer derived from the 168 strain, and DSM10T, the wild-type strain expressing native degQ. The findings demonstrated that surfactin biosynthesis is negatively affected by DegQ-associated DegU regulation, while increased plipastatin biosynthesis is achieved in the presence of native degQ expression. In addition to production of lipopeptides, the DegU regulatory system also plays a role in the formation of secretory proteases. A comparison of extracellular protease activities between JABs24 and DSM10T showed that degQ expression led to DSM10T having five times higher protease activity than JABs24. Interestingly, production of extracellular proteases has not affected the stability of both plipastatin and surfactin during cultivation, suggesting that lipopeptides are less targeted by extracellular proteases. The identification of proficient wild-type strains is critical to the advancement of bio-fungicide in agriculture. Therefore, the subsequent approach of this thesis centered on the production of microbial lipopeptide by wild-type B. velezensis strains. Here, the lipopeptide productivity and antifungal ability of B. velezensis UTB96 was higher than B. velezensis FZB42, as a well-established strain for biocontrol of plant pathogens in agriculture. Furthermore, addition of certain amino acids stimulated lipopeptide production, and using a bioreactor system resulted in enhancement of lipopeptide production, especially iturin A by UTB96. Overall, the doctoral thesis evaluates the biosynthesis of antimicrobial lipopeptides produced by B. subtilis and B. velezensis. The study involves genetic engineering such as promoter exchange, deletion of genes involved in competing biosynthetic pathways and cultivation medium development with amino acid supplementation to enhance the lipopeptide titer. The thesis also identifies B. velezensis UTB96 as a promising candidate for further research to be used as a wild-type antifungal agent in agriculture.
Evaluation of bio-oil produced from fast pyrolysis of lignocellulosic biomass as carbon source for bacterial bioconversion
(2020) Arnold, Stefanie; Hausmann, Rudolf
Scarcity of fossil resources, climate change and growing world population demand the transition from a fossil-based economy towards a bioeconomy – a knowledge-based strategy which relies on the efficient and sustainable integration of bio-based resources into value-added process chains. As lignocellulosic biomass is an abundant renewable resource which does not directly compete with food and feed, its deployment in biorefineries is of special interest for a sustainable bioeconomy. Owing to its compact and complex structure, suitable conversion techniques need to be selected. Combinations of thermochemical and biochemical conversion technologies are considered to be a promising approach regarding a fast and efficient conversion of lignocellulosic biomass into value-added products. Bio-oil derived from fast pyrolysis of lignocellulosic biomass is a complex mixture and composed of water and a wide variety of organic components. Among these components pyrolytic sugars and small organic acids are particularly interesting as potential carbon sources for microbial processes. However, bio-oil also comprises many unidentified substances, as well as components which are known to display adverse effects on microbial growth. To evaluate the potential and challenges of bio-oil as an alternative and sustainable carbon source for bacterial bioconversion this thesis was divided into three parts (Figure 1). In Part I different pretreatment strategies were applied and evaluated regarding their effect on stability and detoxification of bio-oil fractions. For this purpose, the organic solvent tolerant bacterial strain Pseudomonas putida KT2440 was applied as a reference system and cultivated on different pretreated bio-oil fractions. It was shown that solid phase extraction is a suitable tool to obtain bio-oil fractions with significantly increased stability along with less inhibitory substances. Part II is focused on the evaluation of small organic acids mainly present in bio-oil with respect to their suitability as feedstock for bacterial growth. Four biotechnological production hosts Escherchia coli, Pseudomonas putida, Bacillus subtilis and Corynebacterium glutamicum were cultivated on different concentrations of acetate, mixtures of small organic acids, as well as pretreated bio-oil fractions as carbon source for their growth. Results reveal that P. putida, as well as C. glutamicum metabolizes acetate – the major small organic acid generated during fast pyrolysis of lignocellulosic biomass – as sole carbon source over a wide concentration range and grow on mixtures of small organic acids present in bio-oil. Moreover, both strains show a distinct potential to tolerate inhibitory substances within bio-oil. Part III describes the growth behavior of a genetically engineered, nonpathogenic bacterium Pseudomonas putida KT2440 and its simultaneous heterologous production of rhamnolipid biosurfactants on bio-oil derived small organic acids and pretreated fractions. Results suggest that both maximum achievable productivities and substrate-to-biomass yields are in a comparable range for glucose, acetate, as well as the mixture of acetate, formate and propionate. Similar yields were obtained for a pretreated bio-oil fraction, although with significantly lower titers. In conclusion, this thesis shows that microbial valorization of bio-oil is a challenging task due to its highly complex and variable composition, as well as its adverse effects on microbial growth and issues with analytical procedures. This work depicts a proof of concept by outlining a potential biorefinery route for microbial valorization of pretreated bio-oil and its unexploited side streams. It provides a step in search of suitable bacterial strains for bioconversion of lignocellulosicbased feedstocks into value-added products and thus contributes to establishing bioprocesses within a future bioeconomy.
Fertilizer placement and the potential for its combination with bio-effectors to improve crop nutrient acquisition and yield
(2016) Nkebiwe, Peteh Mehdi; Müller, Torsten
Even when total nitrogen (N) and phosphorus (P) concentrations in most agricultural soils are high, the concentrations of plant-available N and P fractions are often inadequate for acceptable yield. In comparison to conventional fertilizer application by homogenous broadcast over the soil surface (with or without subsequent incorporation), fertilizer placement in defined soil areas/volumes close to seeds or crop roots is a more effective application method to enhance the plant-availability of applied fertilizers. Nevertheless, considerable root growth in subsurface nutrient patches or around concentrated fertilizer-depots (and/or improved nutrient influx rates in roots) is a prerequisite for improved uptake of placed nutrients. Furthermore, zones with intense rooting around placed fertilizer depots (“rhizosphere hotspots”) with high concentrations of organic nutrients released as root exudates may be favorable for the survival and establishment of inoculated plant-growth-promoting microorganisms (PGPMs), which mobilize nutrients in soil to favor plant growth. In the last three decades, several published field studies comparing fertilizer placement to fertilizer broadcast arrived at different and often conflicting results regarding their effects on yield and nutrient status of various crops. For this reason, the first task was to conduct a Meta-analysis on data in published peer-reviewed field studies on fertilizer placement that met a set of pre-defined criteria for inclusion. We investigated the relative effect of fertilizer placement for specific fertilizer formulations (e.g. NH4+ and CO(NH2)2 without or in combination with soluble P (HPO42-; H2PO4-); soluble K; solid or liquid manure) in a precise restricted area on surface or subsurface soil in comparison to fertilizer broadcast on yield, nutrient concentration and content in above-ground plant parts. We utilized data from a total of 40 field studies published between 1982 and 2015 (85% of studies published from 2000) that met our criteria. We used the method of “baseline contrasts” to compare different fertilizer placement treatments to fertilizer broadcast as a common control or baseline treatment. Results showed that overall, fertilizer placement led to +3.7% higher yields, +3.7% higher concentrations of nutrients in above-ground plant parts and +11.9% higher contents of nutrients also in above-ground plant parts than fertilizer broadcast application. Placement depth had a strong effect of the outcome of fertilizer placement because relative placement effects increased with increasing fertilizer placement depth. Composition of fertilizer formulations was also an important factor. High yields of fertilizer placement relative to fertilizer broadcast application were obtained for CO(NH2)2 in combination with soluble P (HPO42-; H2PO4-) (+27%) or NH4+ in combination with HPO42-; H2PO4- (+15%) (Nkebiwe et al., 2016 a: Field Crops Research 196: 389–401). The next aim was to investigate the effect of fertilizer placement in subsurface soil in combination with application of bio-effectors (BEs) (PGPMs and natural active substances such as humic acids and seaweed extracts) on root growth of crop plants, establishment of inoculated PGPM in the rhizosphere, grain and biomass production as well as plant nutrient status for maize (Zea mays L) and wheat (Triticum aestivum L) cultures. Through various pot and rhizobox experiments, we observed that placement of a subsurface concentrated NH4+-fertilizer depot stabilized with the nitrification inhibitor DMPP (3,4-di-methylpyrazolphosphate) induced dense rooting around the depot contributing to more efficient exploitation of the depot. For this, it was crucial the N persisted in the depot mainly as poorly mobile NH4+, in order to induce localized depot-zone root-growth as well as favorable chemical and biological changes in the rhizosphere to improve N and P uptake by crop plants. Through in vitro culture experiments on solid and liquid media, we could show that via acidification of the growth media, several selected microbial BEs were capable to solubilize sparingly soluble inorganic phosphates and also that these BEs showed considerable tolerance to high concentrations of NH4+ und DMPP. The latter indicated a potential for the BEs to colonize plant roots in NH4+-rich well rooted soil zones around a subsurface NH4+-fertilizer depot (Nkebiwe et al., 2016 c: Manuscript submitted). Through further pot experiments and four others experiments as Bachelor and Master theses conduction under my supervision, we observed that certain BEs that readily solubilized tri-calcium phosphates in vitro were able to mobilize rock phosphate (RP) applied in soil-based substrates when N was supplied as stabilized NH4++DMPP, thereby contributing to enhanced P uptake and growth of maize and wheat plants. The bacterial BE Pseudomonas sp. DSMZ 13134 and BE consortia products containing bacteria and fungi such as CombiFectorA were good candidates. BE-induced RP-solubilzation occurred mainly in substrates with low CaCO3 contents indicating low P sorption capacity for neutral and moderately alkaline soils. With CombiFectorA, maize P-acquisition from sewage sludge ash could be enhanced, thus increasing the efficiency of a sparingly soluble fertilizer based of recycled wastes. Possible explanations for the beneficial effects of best performing BEs to improve plant growth were enhanced solubility of sparingly soluble P fertilizers via acidification of the rhizosphere and release of nutrient-chelating substances as well as improvement of root growth for better spatial interception of nutrients (Nkebiwe et al., 2016 d: Manuscript in preparation). Alongside, more greenhouse and two field experiments (grain maize 2014 and maize silage 2015) were designed, planned, conducted and evaluated. A peer-reviewed paper from this work has already been published (Nkebiwe et al., 2016 b: Chemical and Biological Technologies in Agriculture 3:15). In the greenhouse and experiments, placement of a concentrated stabilized NH4+-fertilizer depot led to improved root and shoot growth, and increased shoot N and P contents. Through intense root growth of maize around the NH4+-depot, increased root-colonization by Pseudomonas sp. DSMZ 13134 close to seeds could be observed. In the field, many weeks after subsurface placement of the concentrated stabilized NH4+-depot, it could be shown that N considerably persisted in the depot-zone as NH4+, which strongly induced depot-zone root growth. Placement of the NH4+-depot led to +7.4 % increase in grain yield of maize (2014) and +5.8% increase in maize silage yield (2015) in comparison to fertilizer broadcast. Placement of Pseudomonas sp. DSMZ 13134 inoculum in the sowing row let to +7.1% increase in yield of maize silage (2015) in comparison to the non-inoculated control. In total, these results showed that precise placement of specific fertilizer formulations in combination with the application of selected PGPMs can lead to improved plant growth, improved N and P uptake with a potential to save resources.
Impacts of the fungal bio-control agent Fusarium oxysporum f.sp. strigae on plant beneficial microbial communities in the maize rhizosphere
(2016) Musyoki, Mary Kamaa; Cadisch, Georg
Striga hermonthica causes severe yield reduction in cereal crop production in Sub-Saharan Africa. Intergrated Striga management has been proposed as one of the best options to control striga. Along this line, the use of biocontrol agent (BCA) Fusarium oxysporum f.sp. strigae (Foxy-2) has been proven as an effective and environmental friendly management strategy. It is well established that a prerequisite for a successful BCA is sufficient risk assessment analysis. Towards this direction, Foxy-2 was assessed for its potential non-target impacts on the abundance, community structure of bacterial and archaeal nitrifying prokaryotes as well as enzymatic activities of proteolytic bacteria. Maize rhizosphere soils treated with or without Foxy-2, Striga and high quality organic residues (i.e., Tithonia diversifolia) as N source were evaluated by quantitative polymerase chain reaction (qPCR) and terminal restriction fragment length polymorphism (TRFLP). It was observed that Foxy-2 had a promoting effect on archaeal abundance under controlled conditions in sandy soils. Furthermore, crop growth stage, seasonality and soil type had a strong effect on abundance and community structure of nitrifying prokaryotes over Foxy-2 inoculation. In addition proteolytic enzymatic activities analysis showed that Foxy-2 did not affect their activities. Correlation analysis also showed that abundance and community structure on nitrifying communities positively correlated with extractable organic carbon, extractable organic nitrogen and soil pH, while proteolytic enzymatic activities correlated with extractable organic nitrogen and soil ammonium. It was concluded that Foxy-2 is compatible with nitrifying prokaryotes and proteolytic enzymatic activities.
Interaktion des Photosensors Ppr aus Rhodocista centenaria mit Proteinkomponenten der chemotaktischen Signaltransduktion
(2008) Kreutel, Sven; Kuhn, Andreas
Rhodocista centenaria (previously known as Rhodospirillum centenum) is a photosynthetic alpha-proteobacterium which exhibits a unique phototactic response in respect of the direction of light. In this work, the focus is on the potential photoreceptor Ppr and its C-terminal histidine kinase Pph to identify putative binding partners in the signal transduction pathway. The results of overexpression experiments with the Ppr-receptor or the Pph-domain in E. coli indicated that there may be an interaction between the photosensor and the chemotactic signalling pathway. Even cells expressing only small amounts of the R. centenaria proteins showed no chemotactic response at all, whereas uninduced cells exhibited normal chemotactic response on swarm plates as well as in capillary assays. To investigate whether the receptor interacts with components of the chemotactic pathway, the Ppr-protein and the Pph-domain as well as the chemotactic proteins CheW and CheAY of R. centenaria were heterologously expressed in E. coli and purified to homogeneity by affinity chromatography. Binding experiments were carried out by using an IAsys biosensor cuvette system. The results indicated that the kinase domain Pph binds to the chemotactic linker protein CheW with a dissociation constant of 0.13 ± 0.026 μM. Pull-down experiments were made to verify this finding and to investigate the role of ATP in the binding process. The results confirmed our previous observations but in contrast to the complex formation in the E. coli chemotactic pathway, the binding of the C-terminal histidine kinase to CheW was ATP-dependent. To study whether the kinase domain also binds CheW in vivo, expression and coelution experiments with tagged Pph-protein were carried out in R. centenaria. The findings suggest that the complex formation occurs in vivo as well as in vitro. From the data of the E. coli chemotactic complex formation it is well known that CheA is part of the trimeric complex which consists of MCP-receptors, CheW and CheA. To analyse this, pulldown experiments with all three proteins (Pph, CheW and CheAY) were performed. The results clearly showed a participation of CheAY in the formation of the complex with CheW and the kinase Pph. It is known that the photoreceptor Ppr is autophosphorylated during its light induced photocycle. We therefore examined whether the kinase domain is sufficient for this autophosphorylation reaction and whether CheAY could function as a phosphate acceptor. Our results confirmed the hypothesis that the kinase domain is sufficient for autophosphorylation and that the Pph-protein assists CheAY to take over phosphate groups. Taken together, the results in combination with data from the literature lead to a detailed working model for the function of the photoreceptor Ppr and the signal transduction pathway.
Microbial community structure and function is shaped by microhabitat characteristics in soil
(2016) Ditterich, Franziska; Kandeler, Ellen
Soil microorganisms play a key role in degradation processes in soil, such as organic matter decomposition and degradation of xenobiotics. Microbial growth and activity and therefore degradation processes are influenced by different ecological factors, such as substrate availability, pH and temperature. During soil development different microhabitats are formed which differ in their physiochemical properties. There is some evidence that mineral composition is a driver for specific microbial colonization. Thereby, the heterogeneity of soils with differences in mineral composition and substrate availability can lead to a spatial distribution of soil microorganisms. At the soil-litter interface, a biogeochemical hot spot in soil, the abundance and activity of soil microorganisms increases due to high substrate availability, and degradation processes such as pesticide degradation are enhanced. This thesis aimed to clarify the influence of habitat properties on the structure and function of the microbial community in soil. In particular, focus was on mineral-microbe interactions that result from the mineral composition and substrate availability in an artificial soils system. Furthermore this thesis was designed to increase our understanding of the bacterial and fungal roles in pesticide degradation at the soil-litter interface using 4-chloro-2-methylphenoxyacetic acid (MCPA) as a model xenobiotic. These two aspects of the thesis were examined in three studies. The first study focused on the succession of microbial communities and enzyme activities in an artificial soils system with varying mineral composition and substrate availability over a period of 18 months. In the second study a microcosm experiment was used to study the bacterial pathway of MCPA degradation at the soil-litter interface. Over a period of 27 days the succession of bacterial degraders was followed. The third study focused on the degradation of MCPA in soil by nonspecific fungal enzymes, through the addition of fungal laccases as well as litter during 42 days of incubation. Both studies indicated the involvement of fungi in MCPA degradation and the importance of the ecological behavior of different degraders as a function of substrate availability. Results of the first study indicated that the microbial community was affected by mineral properties under high substrate availability and by the availability of beneficial nutrients at the end of incubation when substrate had become limited. The measured enzyme activities provided clear evidence that microbial community structure was driven by nutrient limitation during incubation. In the presence of easily available organic substrates at the beginning of the experiment, the soil microbial community was dominated by copiotrophic bacteria (e.g. Betaproteobacteria), whereas under substrate limitation at the end of incubation, more recalcitrant compounds became important to oligotrophic bacteria (e.g. Acidobacteria), which then became dominant. The results of the second study indicated that the contribution of the potential degraders to degradation of MCPA differed, and this was also seen in the succession of specific bacterial MCPA degraders. Added litter stimulated MCPA degradation due to the availability of litter-derived carbon and induced a two-phase response of fungi. This was seen in the development of pioneer and late stage fungal communities. Both fungal communities were probably involved in MCPA degradation. Therefore, the third study focused on the fungal pathway. These results indicated that the fungal laccases used had no direct influence on degradation and were as efficient as litter in providing additional nutrient sources, increasing MCPA degradation by bacteria and fungi. The observed differences between litter and enzyme addition underscored the observation that the enzyme effect was short-lived and that substrate quality is an important factor in degradation processes. In conclusion, this thesis demonstrated that soil microbial communities and therefore degradation processes are driven by mineral composition as well as substrate availability and quality. In addition, this thesis extends our understanding of degradation processes such as the degradation of xenobiotics, with MCPA as model compound, in soil. The combined insights from all three studies suggest that the use of a simple system such as the artificial soil system can increase our understanding of complex mechanisms such as degradation of pesticides.
Microbial regulation of pesticide degradation coupled to carbon turnover in the detritusphere
(2015) Pagel, Holger; Streck, Thilo
Many soil functions, such as nutrient cycling or pesticide degradation, are controlled by microorganisms. Dynamics of microbial populations and biogeochemical cycling in soil are largely determined by the availability of carbon (C). The detritusphere is a microbial “hot spot” of C turnover. It is characterized by a concentration gradient of C from litter (high) into the adjacent soil (lower). Therefore, this microhabitat is very well suited to investigate the influence of C availability on microbial turnover. My thesis aimed at the improved understanding of biochemical interactions involved in the degradation of the herbicide 4-chloro-2-methylphenoxyacetic acid (MCPA) coupled to C turnover. In the detritusphere gradients of organic matter turnover from litter into the adjacent soil could be identified. Increased C availability, due to the transport of dissolved organic substances from litter into soil, resulted in the boost of microbial biomass and activity as well as in the acceleration of MCPA degradation. Fungi and bacterial MCPA-degraders benefited most from litter-C input. Accelerated MCPA degradation was accompanied by increased incorporation of MCPA-C into soil organic matter. The experimental results show that the transport of dissolved organic substances from litter regulates C availability, microbial activity and finally MCPA degradation in the detritusphere. In general, litter-derived organic compounds provide energy and resources for microorganisms. The following possible regulation mechanisms were identified: i) Litter might directly supply the co-substrate alpha-ketoglutarate (or surrogates) required for enzymatic oxidation of MCPA by bacterial MCPA degraders. Alternatively it might provide additional energy and resources for production and regeneration of the needed co-substrate. ii) Additional litter-C might alleviate substrate limitation of enzyme production by bacteria and bacterial consortia resulting in an increased activity of specific enzymes attacking MCPA. iii) Litter-derived organic substances might stimulate MCPA degradation via fungal co-metabolism by unspecific extracellular enzymes, either directly by inducing enzyme production, or by supplying primary substrates that provide the energy consumed by co-metabolic MCPA transformation. A new biogeochemical model abstracts these regulation mechanisms in such a way that C availability controls physiological activity, growth, death and maintenance of microbial pools. Based on a global sensitivity analysis, 41% (n=33) of all considered parameters and input values were classified as “very important” and “important”. These mainly include biokinetic parameters and initial values. The calibration of the model allowed to validate the implemented regulation mechanisms of accelerated MCPA degradation. The Pareto-analysis showed that the model structure was adequate and the identified parameter values were reasonable to reproduce the observed dynamics of C and MCPA. The model satisfactorily matched observed abundances of gene-markers of total bacteria and specific MCPA degraders. However, it underestimated the steep increase of fungal ITS fragments, most probably because this gene-marker is only inadequately suited as a measure of fungal biomass. The model simulations indicate that soil fungi primarily benefit from low-quality C, whereas bacterial MCPA-degraders preferentially use high-quality C. According to the simulations, MCPA was predominantly transformed via co-metabolism to high-quality C. Subsequently, this C was primarily assimilated by bacterial MCPA-degraders. The highest turnover of litter-derived C occurred by substrate uptake for microbial growth. Input and microbial turnover of litter-C stimulated MCPA degradation mainly in a soil layer at 0-3 mm distance to litter. As a consequence of this, a concentration gradient of MCPA formed, which triggered the diffusive upward transport of MCPA from deeper soil layers into the detritusphere. The results of the three studies suggest: The detritusphere is a biogeochemical hot spot where microbial dynamics control matter cycling. The integrated use of experiments and mathematical modelling gives detailed insight into matter cycling and dynamics of microorganisms in soil. Microbial communities need to be explicitly considered to understand the regulation of soil functions.
Microbial regulation of soil organic matter decomposition at the regional scale
(2018) Ali, Rana Shahbaz; Kandeler, Ellen
The fate of soil organic carbon (SOC) is one of the greatest uncertainties in predicting future climate. Soil microorganisms, as primary decomposers of SOC, control C storage in terrestrial ecosystems by mediating feedbacks to climate change. Even small changes in microbial SOC decomposition rates at the regional scale have the potential to alter land-atmospheric feedbacks at the global scale. Despite their critical role, the ways in which soil microorganisms may change their abundances and functions in response to the climate change drivers of soil temperature and moisture is unclear. Additionally, most existing C models do not consider soil microorganisms explicitly as drivers of decomposition, one consequence of which is large variability in predicted SOC stock projections. This demonstrates the need for a better mechanistic understanding of microbial SOC decomposition at large scales. This thesis was designed to clarify the role of microbial SOC decomposition dynamics in response to climate change factors in two geographically distinct areas and land-use types. The hypothesis was that microbial communities would be adapted to climatic and edaphic conditions specific to each area and to the SOC organic quality in each land-use and would therefore exhibit distinct responses to soil temperature and moisture variations. Three studies were performed to address the goals of this thesis. The first study aimed to clarify temporal patterns of degradation in C pools that varied in complexity by modelling in situ potentials of microbially produced extracellular enzymes. Temperature and moisture sensitivity patterns of C cycling enzymes were followed over a period of thirteen months. The second study investigated group-specific temperature responses of bacteria and fungi to substrate quality variations through an additional incubation experiment. Here, complex environments were mimicked in order to determine the dependence of microbial responses not only on environmental conditions, but also under conditions of inter- and intra-specific community competition. Changes in microbial community composition, abundance, and function were determined at coarse (phospholipid fatty acid – PLFA, ergosterol) and relatively fine resolutions (16S rRNA, taxa-specific quantitative PCR, fungal ITS fragment). A third study investigated 1) the spatial variability of temperature sensitivity of microbial processes, and 2) the scale-specificity and relative significance of their biotic and physicochemical controls at landscape (two individual areas, each ca. 27 km2) and regional scales (pooled data of two areas). Strong seasonal dependency was observed in the temperature sensitivities (Q10) of hydrolytic and oxidative enzymes, whereas moisture sensitivity of β-glucosidase activities remained stable over the year. The range of measured enzyme Q10 values was similar irrespective of spatial scale, indicating a consistency of temperature sensitivities of these enzymes at large scales. Enzymes catalyzing the recalcitrant SOC pool exhibited higher temperature sensitivities than enzymes catalyzing the labile pool; because the recalcitrant C pool is relatively large, this could be important for understanding SOC sensitivity to predicted global warming. Response functions were used to model temperature-based and temperature and moisture-based in situ enzyme potentials to characterize seasonal variations in SOC decomposition. In situ enzyme potential explained measured soil respiration fluxes more efficiently than the commonly used temperature-respiration function, supporting the validity of our chosen modelling approach. As shown in the incubation experiment, increasing temperature stimulated respiration but decreased the total biomass of bacteria and fungi irrespective of substrate complexity, indicating strong stress responses by both over short time scales. This response did not differ between study areas and land-uses, indicating a dominant role of temperature and substrate quality in controlling microbial SOC decomposition. Temperature strongly influenced the responses of microbial groups exhibiting different life strategies under varying substrate quality availability; with soil warming, the abundance of oligotrophs (fungi and gram-positive bacteria) decreased, whereas copiotrophs (gram-negative) increased under labile C substrate conditions. Such an interactive effect of soil temperature and substrate quality was also visible at the taxon level, where copiotrophic bacteria were associated with labile C substrates and oligotrophic bacteria with recalcitrant substrates. Which physicochemical and biological factors might explain the observed alterations in microbial communities and their functions in response to climate change drivers at the regional scale was investigated in the third study. Here, it was shown that the soil C:N ratio exerted scale-dependent control over soil basal respiration, whereas microbial biomass explained soil basal respiration independent of spatial scale. Factors explaining the temperature sensitivity of soil respiration also differed by spatial scale; extractable organic C and soil pH were important only at the landscape scale, whereas soil texture as a control was independent of spatial scale. In conclusion, this thesis provides an enhanced understanding of the response of microbial C dynamics to climate change at large scales by combining field measurements with innovative laboratory assays and modelling tools. Component specific degradation rates of SOC using extracellular enzyme measurements as a proxy, group-specific temperature sensitivities of microbial key players, and the demonstrated scale-specificity of factors controlling microbial processes could potentially improve the predictive power of currently available C models at regional scale.
Modeling microbial regulation of pesticide turnover in soils
(2022) Chavez Rodriguez, Luciana; Streck, Thilo
Pesticides are widely used for pest control in agriculture. Besides their intended use, their long-term fate in real systems is not well understood. They may persist in soils, thereby altering ecosystem functioning and ultimately affecting human health. Pesticide fate is assessed through dissipation experiments in the laboratory or the field. While field experiments provide a close representation of real systems, they are often costly and can be influenced by many unknown or uncontrollable variables. Laboratory experiments, on the other hand, are cheaper and have good control over the governing variables, but due to simplification, extrapolation of the results to real systems can be limited. Mechanistic models are a powerful tool to connect lab and field data and help us to improve our process understanding. Therefore, I used mechanistic, process-based models to assess key microbial regulations of pesticide degradation. I tested my model hypotheses with two pesticide classes: i) chlorophenoxy herbicides (MCPA (2-methyl-4-chlorophenoxyacetic acid) and 2,4-D (2,4-Dichlorophenoxyacetic acid)), and ii) triazines (atrazine (AT)), in an ideal scenario, where bacterial degraders and pesticides are co-localized. This thesis explores some potential controls of pesticide degradation in soils: i) regulated gene expression, ii) mass-transfer process across the bacterial cell membranes, iii) bioenergetic constraints, and iv) environmental factors (soil temperature and moisture). The models presented in this thesis show that including microbial regulations improves predictions of pesticide degradation, compared to conventional models based on Monod kinetics. The gene-centric models achieved a better representation of microbial dynamics and enable us to explore the relationship between functional genes and process rates, and the models that used transition state theory to account for bioenergetic constraints improved the description of degradation at low concentrations. However, the lack of informative data for the validation of model processes hampered model development. Therefore, in the fourth part of this thesis, I used atrazine with its rather complex degradation pathway to apply a prospective optimal design method to find the optimal experimental designs to enable us identifying the degradation pathway present in a given environment. The optimal designs found suggest to prioritize determining metabolites and biomass of specific degraders, which are not typically measured in environmental fate studies. These data will lead to more robust model formulations for risk assessment and decision-making. With this thesis, I revealed important regulations of pesticide degradation in soils that help to improve process understanding and model predictions. I provided simple model formulations, for example the Hill function for gene expression and transition state theory for bioenergetic growth constraints, which can easily be integrated into biogeochemical models. My thesis covers initial but essential steps towards a predictive pesticide degradation model usable for risk assessment and decision-making. I also discuss implication for further research, in particular how mechanistic process-based modeling could be combined with new technologies like omics and machine learning.
Novel bacterial species from the chicken gastrointestinal tract and their functional diversity
(2023) Rios Galicia, Bibiana; Seifert, Jana
The digestive system of chicken presents different physicochemical conditions along the gastrointestinal tract (GIT), shaping an individual microbial profile along sections with different metabolic capacities and divergence on the adaptations to the environment. Efforts to obtain cultivable bacteria originating from the upper region of chicken GIT enrich the reference genome database and provide information about the site- specific adaptations of bacteria colonizing such GIT sections allowing to understand the metabolic profile and adaptive strategies to the environment. However, the lack of sufficient reference genomes limits the interpretation of sequencing data and restrain the study of complex functions. In this study, 43 strains obtained from crop, jejunum and ileum of chicken were isolated, characterised and genome analysed to observe their metabolic profiles, adaptive strategies and to serve as future references. Eight isolates represent new species that colonise the upper gut intestinal tract and present consistent adaptations that enable us to predict their ecological role, expanding our knowledge on the adaptative functions. Strains of Limosilactobacillus were found to be more abundant in the crop, while Ligilactobacillus dominated the ileal digesta. Isolates from crop encode a high number of glycosidases specialised in complex polysaccharides compared to strains isolated from jejunum and ileum. While isolates from jejunum and ileum encode a higher number of genes that interact with the host such as collagenases and hyaluronidases, indicating preferential persistence and adaptations along the GIT. These results represent the first repository of bacteria obtained from the crop and small intestine of chicken using culturomics, improving the potential handling of chicken microbiome with biotechnological applications
Soil microbial assimilation and turnover of carbon depend on resource quality and availability
(2017) Müller, Karolin; Kandeler, Ellen
The decomposition of soil organic carbon (SOC), which is predominantly performed by soil microorganisms, is an important process in global carbon (C) cycling. Despite the importance of microbial activity to the global C budget, the effects of resource quality and availability on soil microorganisms are little understood. Most of this plant-derived C enters the soil organic C pool via incorporation into soil microorganisms, but the subsequent fate of C is rarely reported. Therefore, soil microbial biogeochemistry is still highly uncertain in earth system models. The study presented in Chapter 5 used a field experiment established in 2009 to investigate C flow at three soil depths over five consecutive years after a C3 to C4 crop exchange. Root-derived C (belowground pathway) was introduced by the cropping of maize plants, whereas shoot-derived C (aboveground pathway) was introduced by application of shoot litter to the soil surface. The proportion of maize-derived C varied between the different soil pools with lower incorporation into SOC and EOC (extractable organic C) and higher incorporation ratios of maize C into microbial groups. Although root-C input was three times higher than shoot-C input, similar relative amounts of maize-C were found in microorganisms. Both root and shoot C were transferred to a depth of 70 cm. At all three depths, fungi utilized the provided maize C to a greater extent than did either Gram-positive or Gram-negative bacteria. Fungal biomass was labeled with maize-C to 78% after the fifth vegetation period, indicating preferential utilization of litter-derived C by saprotrophic fungi. The second study investigated, in a microcosm experiment, the effects of decreasing resource quality on microorganisms during plant residue decomposition at the soil-litter interface. Reciprocal transplantation of labeled 13C and unlabeled 12C maize litter to the surface of soil cores allowed us to follow C transfer and subsequent C turnover from residues into microbial biomass of fundamental members (bacteria and fungi) of the detritivore food web during three stages of the litter decomposition process. Quality (i.e. age) of the maize litter influenced C incorporation into bacteria and fungi. Labile C from freshly introduced litter was incorporated by both groups of microorganisms, whereas saprotrophic fungi additionally used complex C in the intermediate stage of decomposition. Bacteria responded differentially to the introduced litter; either by turnover of litter C in their phospholipid fatty acids (PLFAs) over time, or by storage and/or reuse of previous microbially released C. Saprotrophic fungi, however, showed a distinct litter C turnover in the fungal PLFA. The mean residence time of C in the fungal biomass was 32 to 46 days; the same or shorter time than in bacterial PLFAs. In the third study, presented in Chapter 7, another field experiment was conducted to distinguish herbivore- from detritus-based food chain members over two consecutive years. Three treatments were established: maize as crop plant, maize shoot litter application, and fallow without C input. This provided root-derived C, shoot-derived C, and autochthonous organic matter, respectively, as the main C resource. The altered C supply due to plant removal had less severe effects on the micro-food web structure than expected. In the first growing season, nematode abundance under plant cultivation was similar to that under litter and fallow conditions. After the second harvest, the abundance of detritivore food chain members increased, reflecting the decomposition of root residues. Bacteria and fungi showed a marked resilience to changed C availability. Results of this experiment suggest considerable micro-food web resilience to altered C and nutrient availability, and indicate that organic matter from previous vegetation periods was successfully utilized to overcome C deprivation. In conclusion, this thesis provides new insights into microbially mediated decomposition processes at different time scales and at different soil depths. Stable isotope probing combined with biomarker analysis enabled us to study C fluxes between biotic and soil C pools to separate the contributions of bacteria and fungi to soil C cycling. These results can be used as a basis for an empirical model of C flow through the entire soil food web.
Spatial and temporal variations of microorganisms in grassland soils : influences of land-use intensity, plants and soil properties
(2019) Boeddinghaus, Runa S.; Kandeler, Ellen
Grassland ecosystems provide a wide range of services to human societies (Allan et al., 2015) and plants and soil microorganisms have been identified as key drivers of ecosystem functioning (Soliveres et al., 2016). Therefore, understanding soil microbial distributions and processes in agricultural grassland soils is crucial for characterizing these ecosystems and for predicting how they may shift in a changing environment. Yet we are only beginning to understand these complex ecosystems, which account for about 26% of the world’s terrestrial surface (FAOSTATS, 2018), making it especially urgent to gain better insights into the effects of land-use intensity on soil microbial properties and plant-microbe interactions. This thesis was conducted to evaluate the impact land-use intensity has on soil microbial biogeography of grasslands with respect to both spatial patterns and temporal changes in soil microbial abundance, function (in terms of enzyme activities), and community composition. It also investigated the relationships between plants and the spatial and temporal distributions of soil microorganisms. Thereby both, land-use intensity effects and plant-microbe interactions, were assessed in light of ecological niche and neutral theory. This thesis is based on three observational studies conducted on from one to 150 continuously farmed, un-manipulated grassland sites in three regions of Germany within the Biodiversity Exploratories project (DFG priority program 1374). The first study assessed the effects of land-use intensity and physico-chemical soil properties on the spatial biogeography of soil microbial abundance and function in 18 grasslands sites from two of the three regions, sampled at one time point. The second study analyzed spatial and temporal distributions of alpha- and beta-diversity of arbuscular mycorrhizal fungi in a low land-use intensity grassland with six sampling time points across one season. The third study investigated both legacy and short-term change effects of land-use intensity, soil physico-chemical properties, plant functional traits, and plant biomass properties on temporal changes in soil microbial abundance, function, and community composition in 150 grassland sites across three regions, with particular regard to direct and indirect land-use intensity effects. Although the three studies used different approaches and assessed different soil microbial properties, general patterns were detectable. Abiotic soil properties, namely pH, nitrogen content, texture, and bulk density played fundamental roles for spatial and temporal microbial biogeography. Since these factors were specific and unique for each investigated site, they formed the background based on which other processes occurred. In addition to abiotic soil properties, impacts of land-use intensity and plants were detected, though to various degrees in the three studies. Land-use intensity played a much smaller role than anticipated in the first and third study. No influence on the spatial distribution of soil microbial abundance and function could be detected in the first study. In the third study, short-term changes in and legacy effects of land-use intensity played a minor role with respect to short-term changes in soil microbial abundance, function, and community composition. Where detected, changes in land-use intensity had a direct and negative effect on soil microbial properties in structural equation modelling; i.e., increases in land-use intensity reduced, e.g., soil microbial enzyme activities, while legacy effects of land-use intensity were shown to act both directly and indirectly on soil microbial properties. Thereby indirect legacy effects were mediated via plant functional traits. Only one of the three studies detected minor plant diversity effects on soil microbial properties. Instead, functional properties of the plant communities, i.e., plant functional traits, biomass, and nutritional quality, were significantly related to spatial and temporal distributions of soil microorganisms. Finally, the findings of the three studies suggest that processes related to niche and neutral theory both drive spatial and temporal patterns of soil microbial properties at the investigated plot scale (up to 50 m × 50 m). This thesis concluded that in order to gain deeper insights into the complex functions and processes occurring in grassland ecosystems, a multidisciplinary approach investigating fundamental physico-chemical site characteristics, microbial soil properties, and plants is necessary. The results of the thesis suggest that focus be turned to functional properties of plant and microbial communities, as they are closely intermingled, provide more detailed insights into plant-microbe interactions, and are able to reflect effects of human impacts on grassland soils better than diversity measures.
UV-C-Behandlung von Traubenmost zur Inaktivierung von Mikroorganismen
(2018) Diesler, Kathrin; Scharfenberger-Schmeer, Maren
The development of new preservation process techniques to protect ingredients and maintain a high quality standard is always a main goal in the food industry. In course of this, microbial safety has top priority. UV-C technology is a modern, non-thermal process with high efficiency. It has been used for sterilization and treatment of drinking water for many years. Also, ultraviolet radiation for disinfection purposes is already being used in other areas of food production. To what extent this method can be successfully applied in the field of grape must production, will be investigated in this dissertation. For this purpose, several yeasts and bacteria, relevant in this area, were examined for their inactivation potential by UV-C treatment. To ensure the best possible microbial inactivation in must, it is essential to determine an ideal treatment dose for both yeasts and bacteria. The results have confirmed that bacteria are far more sensitive to UV C treatment, than yeasts. It was also shown that there are major differences in UV-C stability within the seven yeast species and six bacteria species used in this study. The analyses have identified Metschnikowia pulcherrima and Acetobacter aceti as the most UV-C stable and Brettanomyces custerianus and Pediococcus sp. as the most sensitive organisms. Furthermore, three morphologically different Brettanomyces strains were used to show that there are also strain-specific variances in the response to UV-C treatment. Using Saccharomyces cerevisiae as an example, a potential formation of UV-C resistance was also ruled out. For this purpose, yeast cells were exposed to a dose, that did not result in complete inactivation. The surviving cells were cultured and retreated. Even after repeating this process eight times, no change in the UV-C response of the yeast cells could be detected. For the application of UV-C technology in the juice and wine industry it has to be ensured, that microorganisms are killed directly and their enzymatic activities are directly inhibited. Yeasts and bacteria could further convert sugar to alcohol or form unwanted metabolic byproducts. Therefore, the enzymatic activity after the initial treatment and during the inactivation process of Saccharomyces cerevisiae was analyzed in more detail. HPLC was used to determine the content of glucose, fructose and ethanol. No enzymatic activity could be detected in the UV-C treated samples from the moment after the initial UV-C treatment, up to the day of complete destruction. However, the effectiveness of UV-C treatment of must and wine cannot be attributed solely to the responsiveness of the various microorganisms. Other product parameters such as grape variety, turbidity and optical density also play a decisive role. In this context, four different musts with different optical density and turbidity were treated and the inactivation kinetics of Saccharomyces cerevisiae were compared. In this work it could be proved, that with an increasing optical density and a higher turbidity, the efficiency of the UV-C treatment in must decreases strongly. The success of a treatment is also directly dependent on the initial contamination rate of the product. Tests with different starting cell numbers have shown, that the required inactivation dose also has to be increased, as the number of cells increases. In the winemaking process, however, not only yeasts and bacteria can be a potential source of danger. The fungal infection of grapes by Botrytis cinerea also carries a high risk. The polyphenol oxidase laccase, produced by the fungus, damages ingredients and leads to a colour change in must and wine. In the investigations it could be proven, that it is possible to strongly reduce or completely inactivate the enzymatic activity in Botrytis infected must, depending on the starting concentration. In summary, UV-C technology represents an effective alternative and extension for current oenological practice. It offers the possibility to inactivate a large number of wine relevant microorganisms without causing resistance. In addition, this work has created a new framework for the application of must specific parameters. The results for the inactivation of the enzyme laccase are also proved to be extremely promising.

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