Publications (49)110.92 Total impact
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Article: Formation of ethyl acetate from whey by Kluyveromyces marxianus on a pilot scale.
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ABSTRACT: Whey arising in huge amounts during milk processing is a valuable renewable resource in the field of White Biotechnology. Kluyveromyces marxianus is able to converted whey-borne lactose into ethyl acetate, an environmentally friendly solvent. Formation of ethyl acetate as a bulk product is triggered by iron (Fe). K. marxianus DSM 5422 was cultivated aerobically in whey-borne medium originally containing 40μg/L Fe, supplemented with 1, 3 or 10mg/L Fe in the pre-culture, using an 1-L or 70-L stirred reactor. The highest Fe content in the pre-culture promoted yeast growth in the main culture causing a high sugar consumption for growth and dissatisfactory formation of ethyl acetate, while the lowest Fe content limited yeast growth and promoted ester synthesis but slowed down the process. An intermediate Fe dose (ca. 0.5μg Fe/g sugar) lastly represented a compromise between some yeast growth, a quite high yield of ethyl acetate and an acceptable duration of the process. The mass of ethyl acetate related to the sugar consumed amounted to 0.113, 0.265 and 0.239g/g in the three processes corresponding to 21.9, 51.4 and 46.3% of the theoretically maximum yield. The performance on a pilot scale was somewhat higher than on lab scale.Journal of biotechnology 10/2012; · 2.88 Impact Factor -
Article: Formation of ethyl acetate by Kluyveromyces marxianus on whey during aerobic batch and chemostat cultivation at iron limitation.
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ABSTRACT: The ability of Kluyveromyces marxianus to convert lactose into ethyl acetate offers a chance for an economic reuse of whey. Former experiments with K. marxianus DSM 5422 proved limitation of growth by iron (Fe) or copper as a precondition for significant ester synthesis. Several aerobic batch and chemostat cultivations were done with whey-borne media of a variable Fe content for exploring the effect of Fe on growth, the Fe content of biomass, and metabolite synthesis. At low Fe doses, Fe was the growth-limiting factor, the available Fe was completely absorbed by the yeasts, and the biomass formation linearly depended on the Fe dose governed by a minimum Fe content in the yeasts, x (Fe,min). At batch conditions, x (Fe,min) was 8.8 μg/g, while during chemostat cultivation at D = 0.15 h(-1), it was 23 μg/g. At high Fe doses, sugar was the growth-limiting factor, Fe was more or less absorbed, and the formed biomass became constant. Significant amounts of ethyl acetate were only formed at Fe limitation while high Fe doses suppressed ester formation. Analysis of formed metabolites such as glycerol, pyruvate, acetate, ethanol, ethyl acetate, isocitrate, 2-oxoglutarate, succinate, and malate during chemostat cultivation allowed some interpretation of the Fe-dependent mechanism of ester synthesis; formation of ethyl acetate from acetyl-SCoA and ethanol is obviously initiated by a diminished metabolic flux of acetyl-SCoA into the citrate cycle and by a limited oxidation of NADH in the respiratory chain since Fe is required for the function of aconitase, succinate dehydrogenase, and the electron-transferring proteins.Applied Microbiology and Biotechnology 06/2012; 96(3):685-96. · 3.42 Impact Factor -
Article: Formation of ethyl acetate by Kluyveromyces marxianus on whey during aerobic batch cultivation at specific trace element limitation.
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ABSTRACT: Kluyveromyces marxianus is able to transform lactose into ethyl acetate as a bulk product which offers a chance for an economical reuse of whey-borne sugar. Ethyl acetate is highly volatile and allows its process-integrated recovery by stripping from the aerated bioreactor. Extensive formation of ethyl acetate by K. marxianus DSM 5422 required restriction of yeast growth by a lack of trace elements. Several aerobic batch processes were done in a 1-L stirred reactor using whey-borne culture medium supplemented with an individual trace element solution excluding Mn, Mo, Fe, Cu, or Zn for identifying the trace element(s) crucial for the observed ester synthesis. Only a lack of Fe, Cu, or Zn restricted yeast growth while exclusion of Mn and Mo did not exhibit any effect due to a higher amount of the latter in the used whey. Limitation of growth by Fe or Cu caused significant production of ethyl acetate while limitation by Zn resulted in formation of ethanol. A lack of Fe or Cu obviously makes the respiratory chain inefficient resulting in an increased mitochondrial NADH level followed by a reduced metabolic flux of acetyl-SCoA into the citrate cycle. Synthesis of ethyl acetate from acetyl-SCoA and ethanol by alcoholysis is thus interpreted as an overflow metabolism.Applied Microbiology and Biotechnology 05/2012; · 3.42 Impact Factor -
Article: Constitutive expression of hydrophobin HFB1 from Trichoderma reesei in Pichia pastoris and its pre‐purification by foam separation during cultivation
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ABSTRACT: Hydrophobins are small surface-active proteins that have considerable potential for use in applications ranging from medical and technical coatings, separation technologies, biosensors, and personal care. Their wider use would be facilitated by the availability of recombinant tailor-made hydrophobins. We successfully expressed the class II hydrophobin HFB1 from Trichoderma reesei in Pichia pastoris under the control of the constitutive GAP (glyceraldehyde 3-phosphate dehydrogenase) promoter. Avoiding the use of the AOX1 (alcohol oxidase 1) promoter prevents the costs and risks associated with the storage and delivery of methanol used as an inducer. Efficient secretion of hydrophobin was achieved using either the alpha-factor prepro-peptide or the native secretion signal of HFB1. The secreted hydrophobins have been isolated with a purity of up to 70% using in situ foam separation during the cultivation process. Coating experiments and surface pressure measurements demonstrated the activity of the hydrophobins. An immunodot assay showed the accessibility of carboxyterminally fused tags of the hydrophobin, which is necessary for potential applications using functionalized hydrophobins. The presented data show that Pichia pastoris is a suitable system for production of constitutively expressed and secreted active hydrophobin, allowing for in situ pre-purification using foam separation.Engineering in Life Sciences 01/2012; 12(2):162 - 170. · 1.92 Impact Factor -
Article: Bioprocessing of differentiated plant in vitro systems
Engineering in Life Sciences 01/2012; · 1.92 Impact Factor -
Chapter: Bioreactors for the Cultivation of Red Beet Hairy Roots Red Beet Biotechnology
01/2012: pages 251-281; , ISBN: 9781461434580 -
Article: Substrate utilization by recombinant Yarrowia lipolytica growing on sucrose.
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ABSTRACT: We report the study of the dynamics of substrate utilization by the genetic modified strain Yarrowia lipolytica H222-S4(p67ICL1) T5. In contrast to its wild-type equivalent, this recombinant strain is able to excrete the sucrose cleaving enzyme invertase. Both the sucrose degradation rate and the glucose and fructose consumption rate have been investigated. In all experiments, satisfied amounts of invertase were produced so that all sucrose was cleaved into its monomers. While glucose and fructose as sole carbon sources were consumed with the same uptake rate, a clear preference for glucose uptake was detected in cultivations with sucrose as sole carbon source or mixed substrates when compared with fructose. Nevertheless, no real diauxie could be observed because of partly simultaneous consumption of both monosaccharides. Fructose being present in the cultivation medium at the beginning of the fermentation led to the retardation of glucose uptake. This effect was observed for various fructose starting concentrations in the range of 5-85 g/l.Applied Microbiology and Biotechnology 11/2011; 93(4):1695-702. · 3.42 Impact Factor -
Article: Solid‐state fermentation of lignocellulotic materials for the production of enzymes by the white‐rot fungus Trametes hirsuta in a modular bioreactor
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ABSTRACT: Trametes hirsuta, a filamentous basidiomycete, was successfully cultivated in solid-state fermentation (SSF) on a mixture of pine wood chips and orange peel in a novel bioreactor that allows mixing of the inoculated solid material during the fermentation. Copper sulfate or xylidine (2,5-dimethylaniline) were added to enhance the production of enzymes, especially laccases. For comparison, Trametes hirsuta was also cultivated in submerged conditions. The effect of additives in SSF was low, whereas the choice of the solid material and the rotation of the reactor vessel showed a significant influence on the enzyme production. The space-time yields for the fermentations were calculated and showed that SSF on low-cost substrates can effectively produce extracellular enzymes at a sufficient rate by a cultivation method that requires only low technology equipment.Engineering in Life Sciences 08/2011; 11(4):395 - 401. · 1.92 Impact Factor -
Article: Correlation of community dynamics and process parameters as a tool for the prediction of the stability of wastewater treatment.
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ABSTRACT: Wastewater treatment often suffers from instabilities and the failure of specific functions such as biological phosphorus removal by polyphosphate accumulating organisms. Since most of the microorganisms involved in water clarification are unknown it is challenging to operate the process accounting for the permanent varying abiotic parameters and the complex composition and unrevealed metabolic capacity of a wastewater microbial community. Fulfilling the demands for water quality irrespective of substrate inflow conditions may emit severe problems if the limited management resources of municipal wastewater treatment plants are regarded. We used flow cytometric analyses of cellular DNA and polyphosphate to create patterns mirroring dynamics in community structure. These patterns were resolved in up to 15 subclusters, the presence and abundances of which correlated with abiotic data. The study used biostatistics to determine the kind and strength of the correlation. Samples investigated were obtained from a primary clarifier and two activated sludge basins. The stability of microbial community structure was found to be high in the basins and low in the primary clarifier. Despite major abiotic changes certain subcommunities were dominantly present (up to 80% stability), whereas others emerged only sporadically (down to 3% stability, both according to equivalence testing). Additionally, subcommunities of diagnostic value were detected showing positive correlation with substrate influxes. For instance blackwater (r(s) = 0.5) and brewery inflow (both r(s) = 0.6) were mirrored by increases in cell abundances in subclusters 1 and 6 as well as 4 and 8, respectively. Phosphate accumulation was obviously positively correlated with nitrate (r(s) = 0.4) and the presence of denitrifying organisms (Rhodacyclaceae). Various other correlations between community structure and abiotic parameters were apparent. The bacterial composition of certain subcommunities was determined by cell sorting and phylogenetic tools like T-RFLP. In essence, we developed a monitoring tool which is quick, cheap and causal in its interpretation. It will make laborious PCR based technique less obligatory as it allows reliable process monitoring and control in wastewater treatment plants.Environmental Science & Technology 08/2011; 46(1):84-92. · 4.80 Impact Factor -
Article: Hydrophobin signal sequence mediates efficient secretion of recombinant proteins in Pichia pastoris.
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ABSTRACT: Pichia pastoris is an important eukaryotic organism for the expression, processing, and secretion of recombinant proteins. Here, the secretion of enhanced green fluorescent protein (EGFP) in P. pastoris by using three novel secretion signals originating from the HFBI and HFBII class 2 hydrophobins of Trichoderma reesei was investigated. EGFP was fused to the carboxyl terminus of hydrophobin secretion signals and expressed under the control of the constitutive GAP promoter. In every case, recombinant EGFP entered the secretory pathway of P. pastoris. SDS-polyacrylamide gel electrophoresis, Western blot analysis of the cells' supernatant, and fluorescence measurements on single-cell level via flow cytometry confirmed the efficient secretion of EGFP mediated by the novel secretion sequences. In conclusion, the data clearly show that the secretion sequences derived from HFBI and HFBII of T. reesei have the potential to achieve an efficient secretion of heterologous proteins in P. pastoris. Due to the small size of the hydrophobin-derived secretion signals, their coding sequence can be easily introduced to the gene of interest by PCR.Applied Microbiology and Biotechnology 07/2011; 91(1):133-41. · 3.42 Impact Factor -
Article: Using a carbon-based ASM3 EAWAG Bio-P for modelling the enhanced biological phosphorus removal in anaerobic/aerobic activated sludge systems.
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ABSTRACT: Modelling of activated sludge processes is a commonly used technique to design and optimize wastewater treatment processes. Since wastewater and activated sludge is characterized by chemical oxygen demand (COD) measurements, units of state variables describing organic matter are expressed as equivalent amounts of COD. However, current procedures for measuring it have several drawbacks, including the production of hazardous wastes, so the utility of other variables for characterizing the organic load in modelling, such as total organic carbon (TOC), warrant re-evaluation. Other advantages of TOC over COD are that it provides matrix-independent analytical results and it can be readily measured online. Proposals for TOC-based models were made in the 1990s, but they seem to have sunk into obscurity. To re-assess the value of TOC for this purpose, we have recalculated the EAWAG module for Bio-P removal coupled to the Activated Sludge Model No. 3 on a TOC basis, and tested it against data acquired in batch experiments with four single carbon sources (acetate, glucose, citrate and casein). The batch test-based calibrations showed a good match with experimental data, following modifications of the model to account for the anaerobic volumes and retention times applied in the tests.Bioprocess and Biosystems Engineering 03/2011; 34(3):287-95. · 1.81 Impact Factor -
Article: Repeated fed-batch fermentation using biosensor online control for citric acid production by Yarrowia lipolytica.
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ABSTRACT: Biosensor-controlled substrate feeding was used in a citric acid production process with the yeast strain Yarrowia lipolytica H222 with glucose as the carbon source. The application of an online glucose biosensor measurement facilitated the performance of long-time repeated fed-batch process with automated bioprocess control. Ten cycles of repeated fed-batch fermentation were carried out in order to validate both the stability of the microorganism for citric acid production and the robustness of the glucose biosensor in a long-time experiment. In the course of this fermentation with a duration of 553 h, a slight loss of productivity from 1.4 g/(L×h) to 1.1 g/(L×h) and of selectivity for citric acid from 91% to 88% was observed. The glucose biosensor provided 6,227 measurements without any loss of activity.Journal of biotechnology 03/2011; 153(3-4):133-7. · 2.88 Impact Factor -
Article: Applying dimorphic yeasts as model organisms to study mycelial growth: part 1. Experimental investigation of the spatio-temporal development of filamentous yeast colonies.
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ABSTRACT: Colony development of the dimorphic yeasts Yarrowia lipolytica and Candida boidinii on solid agar substrates under glucose limitation served as a model system for mycelial development of higher filamentous fungi. Strong differences were observed in the behaviour of both yeasts: C. boidinii colonies reached a final colony extension which was small compared to the size of the growth field. They formed cell-density profiles which steeply declined along the colony radius and no biomass decay processes could be detected. The stop of colony extension coincided with the depletion of glucose from the growth substrate. These findings supported the hypothesis that glucose-limited C. boidinii colonies can be regarded as populations of single cells which grow according to a diffusion-limited growth mechanism. Y. lipolytica colonies continued to extend after the depletion of the primary nutrient resource, glucose, until the populations covered the entire growth field which was accomplished by utilization of mycelial biomass.Bioprocess and Biosystems Engineering 01/2011; 34(1):13-20. · 1.81 Impact Factor -
Article: Applying dimorphic yeasts as model organisms to study mycelial growth: part 2. Use of mathematical simulations to identify different construction principles in yeast colonies.
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ABSTRACT: The dimorphic yeasts Candida boidinii and Yarrowia lipolytica were applied as model organisms to study mycelial growth. A mathematical model of hybrid cellular automaton type was developed to analyze the impact of different biological assumptions on the predicted development of filamentous yeast colonies. The one-dimensional model described discrete cells and continuous distribution of nutrients. The simulation algorithm accounted for proliferation of cells, diffusion of nutrient, as well as biomass decay and recycling inside the mycelium. Simulations reproduced the spatio-temporal development of C. boidinii colonies when a diffusion-limited growth algorithm based on the growth of pseudohyphal cells was applied. Development of Y. lipolytica colonies could only be reproduced when proliferation was restricted to the colony boundary, and cell decay and biomass recycling were incorporated into the model. The results suggested that cytoplasm, which served as the secondary nutrient resource, had to be translocated inside the hyphal network.Bioprocess and Biosystems Engineering 01/2011; 34(1):21-31. · 1.81 Impact Factor -
Article: Phytochemical and flow cytometric analyses of Devil’s claw cell cultures
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ABSTRACT: A cell suspension culture of Devil’s claw (Harpagophytum procumbens), a South African plant with high medicinal value, cultivated under submerged conditions showed stable growth and accumulated high amounts of biomass (18.2gl−1). Flow cytometry analyses of the suspension’s cell cycle kinetics showed that proportions of cells in G0/G1 and S phases varied insignificantly (between 69–76% and 9–13%, respectively) during the cultivation, while the proportion of G2/M-phase cells increased until day 8 of cultivation, when the exponential phase of cell growth ended. Metabolite production in the culture was studied through simultaneous determination of three bioactive phenylethanoid glycosides (verbascoside, β-OH-verbascoside and leucosceptoside A) by high performance liquid chromatography. It was found that suspended Devil’s claw cells accumulated mainly verbascoside (517.3mgl−1), followed by leucosceptoside A (107.1mgl−1) and β-OH-verbascoside (80.3mgl−1). In addition, several fatty acids and β-sitosterol were identified in the cell suspension by gas chromatographic-mass spectrometry analysis. Comparison of the results with previously acquired data for Harpagophytum procumbens transformed roots indicate that cell suspensions cultures are more promising as potential commercial sources of metabolites such as phenylethanoid glycosides. KeywordsFlow cytometry–GC–MS– Harpagophytum procumbens cell culture–PhenylethanoidsPlant Cell Tissue and Organ Culture 01/2011; 105(1):79-84. · 3.09 Impact Factor -
Article: Formation of ethyl acetate by Kluyveromyces marxianus on whey: studies of the ester stripping.
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ABSTRACT: Kluyveromyces marxianus is capable of converting lactose into ethyl acetate offering a chance for an economical reuse of whey. The microbial formation of ethyl acetate as a bulk product calls for an aerobic process and, thus, the highly volatile ethyl acetate is discharged from the aerated bioreactor. This stripping process was modeled and investigated experimentally. The stripping rate was proportional to the gas flow and nearly independent of the stirring rate since the stripping was governed by the absorption capacity of the exhaust gas rather than the phase transfer. Cooling the exhaust gas did not noticeably influence the stripping. One batch experiment is presented in detail to demonstrate the formation of ethyl acetate by K. maxianus DSM 5422 on whey. Further batch experiments showed that a substantial formation of ethyl acetate only occurred when the yeast growth was limited by a lack of trace elements. The highest product yield observed was 0.25 g ethyl acetate per g lactose which is nearly 50% of the theoretical maximum.Bioprocess and Biosystems Engineering 12/2010; 34(5):547-59. · 1.81 Impact Factor -
Article: Bioactive metabolite production and stress-related hormones in Devil's claw cell suspension cultures grown in bioreactors.
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ABSTRACT: In a previous report, we showed that cell cultures of Harpagophytum procumbens, a South African plant with high medicinal value, accumulate high amounts of anti-inflammatory phenylethanoid glycosides during cultivation in shake-flasks. The aim of the present study was to transfer the phenylethanoid biosynthetic process to a 3-L stirred tank reactor and a 1-L glass-column bioreactor (operated with pulsed aeration). We found that, with stepwise increases in aeration, the stirred tank reactor yielded similar productivities of verbascoside (the major phenylethanoid glycoside in the cells) to those reported for shake-flask cultures (55.68 vs. 54.78 mg verbascoside/L/day, respectively). Transfer in the pulse-aerated column reactor resulted in 165.42 mg verbascoside/L/day, one of the highest yields reported to date. Further, to evaluate the physiological status of the suspended cells in the bioreactors cultures, we examined their hormone levels and compared them to those of cells in shake-flask cultures. While indole-3-acetic acid levels did not differ significantly between the bioreactor and shake-flask cultures, there were considerable differences in their levels of abscisic, jasmonic, and salicylic acids. These results are discussed with respect to relative stress levels in the different cultivation systems.Applied Microbiology and Biotechnology 11/2010; 89(6):1683-91. · 3.42 Impact Factor -
Article: From single cells to microbial population dynamics: modelling in biotechnology based on measurements of individual cells.
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ABSTRACT: The development of dynamic modelling of microbial populations in bioprocesses is reviewed. In the 1960s Arnold Fredrickson established the theoretical basis of such models, and other researchers have subsequently advanced them. This review explores the relationships that describe cell proliferation and evaluates the importance of the application of flow cytometry to the fundamental parameterisation of the models for their use in bioprocess engineering. The section "Individual-Based Modelling" discusses recent theoretical developments. Delay-differential equations are demonstrated to describe accurately temporal variation of the cell proliferation cycle and specialised approaches and related iconography are applied to stochastic and deterministic modelling of stages of cellular development. Synchronised cultures of the bacterium Cupriavidus necator were prepared and monitored using a flow cytometer. The data obtained demonstrate that cell proliferation could be simulated quantitatively using the developed model.Advances in biochemical engineering/biotechnology 11/2010; 124:211-27. · 1.64 Impact Factor -
Article: Biosensor online control of citric acid production from glucose by Yarrowia lipolytica using semicontinuous fermentation
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ABSTRACT: Our study aimed at the development of an effective method for citric acid production from glucose by use of the yeast Yarrowia lipolytica. The new method included an automated bioprocess control using a glucose biosensor. Several fermentation methodologies including batch, fed-batch, repeated batch and repeated fed-batch cultivation were tested. The best results were achieved during repeated fed-batch cultivation: Within 3 days of cycle duration, approximately 100 g/L citric acid were produced. The yields reached values between 0.51 and 0.65 g/g and the selectivity of the bioprocess for citric acid was as high as 94%. Due to the elongation of the production phase of the bioprocess with growth-decoupled citric acid production, and by operating the fermentation in cycles, an increase in citric acid production of 32% was achieved compared with simple batch fermentation.Engineering in Life Sciences 07/2010; 10(4):311 - 320. · 1.92 Impact Factor -
Article: Ploidy levels in Beta vulgaris (red beet) plant organs and in vitro systems
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ABSTRACT: The ploidy levels of the cells in different organs (leaves, petioles and roots) of red beet (Beta vulgaris L.) plants of different ages, as well as of different in vitro systems (transformed hairy roots, calli derived from leaves and rhizogenic calli), were investigated using flow cytometry. Two callus lines with red and yellow phenotypes, derived by mechanical separation of the morphologically heterogeneous rhizogenic callus, were also examined. All investigated samples experienced several cycles of endoreduplication. The older organs exhibited higher levels of polysomaty than the young ones. The highest degree of endoreduplication was found in old petiole tissue and the lowest in the red callus line (cycle values of 1.81 and 0.55, respectively). Interestingly, the callus derived from leaves did not exhibit a 2Cx peak, but was tetraploid, probably due to genetic instability, which may have been caused by prolonged cultivation under in vitro conditions. Red and yellow calli showed significantly lower polysomaty (cycle values of 0.55 and 0.59, respectively) than the primary rhizogenic callus (cycle value of 1.09). The DNA profiles of the two phenotypes differed, possibly reflecting differences in their metabolism.Engineering in Life Sciences 02/2010; 10(2):139 - 147. · 1.92 Impact Factor
Top co-authors
Top Journals
Institutions
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2005–2012
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Technische Universität Dresden
- • Institut für Botanik
- • Makromolekulare Chemie
- • Institut für Lebensmittel- und Bioverfahrenstechnik
Dresden, Saxony, Germany
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2010
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The Stephan Angeloff Institute of Microbiology
Sofia, Oblast Sofiya-Grad, Bulgaria
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2009–2010
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Helmholtz-Zentrum für Umweltforschung
- Department Umweltmikrobiologie
Leipzig, Saxony, Germany -
Bulgarian Academy of Sciences
- Stefan Angelov Institute of Microbiology
Sofia, Oblast Sofiya-Grad, Bulgaria
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2008
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Forschungszentrum Jülich
Düren, North Rhine-Westphalia, Germany
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