References of "Delvigne, Frank"
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See detailDynamic single-cell analysis of Saccharomyces cerevisiae under process perturbation: comparison of different methods for monitoring the intensity of population heterogeneity
Delvigne, Frank ULg; Baert, Jonathan ULg; Gofflot, Sébastien et al

in Journal of Chemical Technology & Biotechnology (in press)

BACKGROUND: Single cell biology has attracted a lot of attention these past few years and has led to numerous fundamental results pointing out the heterogeneity of clonal cell populations. In this context ... [more ▼]

BACKGROUND: Single cell biology has attracted a lot of attention these past few years and has led to numerous fundamental results pointing out the heterogeneity of clonal cell populations. In this context, microbial phenotypic heterogeneity under bioprocessing conditions needs to be further investigated. In this study, yeast based processes have been investigated by using on-line flow cytometry in combination with a fluorescent transcriptional reporter (GFP) and viability fluorescence tags (propidium iodide, PI). Methods aiming at expressing the dispersion of these fluorescence tags among the yeast populations have been investigated for different bioreactor operating conditions. RESULTS: Yeast viability was determined on the basis of PI uptake. Segregation between PI negative and positive subpopulations could be efficiently quantified on the basis of the mean-to-median ratio or the amplitude of the interquartile range. On the other hand, the same quantification could not be made for the segregation occurring at the level of GFP synthesis. Indeed, when cells were exposed to sub-lethal or mild stresses (such as in scale-down reactors) two GFP subpopulations could be visualized by real-time FC, but quantification by one of the above-mentioned methods was not possible. CONCLUSIONS: Yeast population heterogeneity was observed in representative bioreactor operating conditions. Difficulties for the determination of segregation at the level of GFP synthesis point out the fact that one needs to understand the segregation mechanisms for the applied fluorescent reporters, to judge whether simple mathematical tools may be applied or if more sophisticated computational tools are needed for the quantification of the microbial population segregation. [less ▲]

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See detailFungal biofilm reactor improves the productivity of hydrophobin HFBII
Khalesi, Mohammadreza; Zune, Quentin ULg; Telek, Samuel ULg et al

in Biochemical Engineering Journal (in press)

Production and purification of hydrophobin HFBII has recently been the subject of intensive research, but the yield of production needs to be further improved for a generic use of this molecule at ... [more ▼]

Production and purification of hydrophobin HFBII has recently been the subject of intensive research, but the yield of production needs to be further improved for a generic use of this molecule at industrial scale. In a first step, the influence of different carbon sources on the growth of Trichoderma reesei and the production of HFBII was investigated. The optimum productivity was obtained by using 40 g/L lactose. Carbon starvation and excretion of extracellular enzyme were determined as two main conditions for the production of HFBII. In the second phase, and according to the physiological mechanisms observed during the screening phase, a bioreactor set up has been designed and two modes of cultures have been investigated, i.e. the classical submerged fermentation and a fungal biofilm reactor. In this last set-up, the broth is continuously recirculated on a metal packing exhibiting a high specific surface. In this case, the fungal biomass was mainly attached to the metal packing, leading to a simplification of downstream processing scheme. More importantly, the HFBII concentration increased up to 48.6 ± 6.2 mg/L which was 1.8 times higher in this reactor configuration and faster than the submerged culture. X-ray tomography analysis shows that the biofilm overgrowth occurs when successive cultures are performed on the same packing. However, this phenomenon has no significant influence on the yield of HFBII, suggesting that this process could be operated in continuous mode. Protein hydrolysis during stationary phase was observed by MALDI-TOF analysis according to the removal of the last amino acid from the structure of HFBII after 48 h from the beginning of fermentation in biofilm reactor. Hopefully this modification does not lead to alternation of the main physicochemical properties of HFBII. [less ▲]

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See detailDevelopment of entomopathogenic fungi in mosquito control: which kind of production for which efficiency?
Bawin, Thomas ULg; Delvigne, Frank ULg; Francis, Frédéric ULg

Poster (2014, August 06)

Mosquitoes (Diptera: Culicidae) are zoonotic vectors responsible for numerous infectious diseases of medical and veterinary importance such as filariasis, malaria and encephalitis. As part of an ... [more ▼]

Mosquitoes (Diptera: Culicidae) are zoonotic vectors responsible for numerous infectious diseases of medical and veterinary importance such as filariasis, malaria and encephalitis. As part of an integrated vector control, entomopathogenic fungi could be developed as biopesticides in two ways: spores and metabolites recognized as effective virulence factors. Solid-state fermentation enhances spore production and induces the secretion of metabolites quantitatively and qualitatively different from submerged fermentation, which impairs fungal metabolic efficiency. In this context, we showed high spore productivity of solid-state media based on agro-industrial substrates as wheat bran. Spores remained pathogenic, as revealed by classical toxicity tests and electron microscopy. However, the absence of free water makes culture parameter variations difficult to control in large-scale. Recently, we performed a bioreactor design intended for simultaneous spore and metabolite production, combining the technological advantages of submerged and solid-state fermentations. Biofilm fermentation (i.e. growth of fungal biomass on an inert support immerged in a nutrient medium) is a tremendous production system favouring the secretion of insecticidal metabolites in the liquid medium as we showed recently. This is also an interesting tool to provide an overview of the complexity of the metabolic pathways involved in the regulation of extracellular metabolites secretion because corresponding genes are reported to be differentially expressed from classical fermentation systems. Researches in vector control are currently intensified. In this context, the identification of genes and metabolites specifically expressed during biofilm fermentation will help to develop new technologies related both to the design of bioreactor and the production of insecticidal proteins. [less ▲]

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See detailEffect of entomopathogenic Aspergillus strains against the pea aphid, Acyrthosiphon pisum (Hemiptera: Aphididae)
Seye, Fawrou; Bawin, Thomas ULg; Boukraa, Slimane ULg et al

in Applied Entomology and Zoology (2014)

Aphids (Homoptera: Aphididae) are sap-sucking insect pests that feed on several plants of agronomical importance. Entomopathogenic fungi are valuable tools for potential aphid control. As part of a ... [more ▼]

Aphids (Homoptera: Aphididae) are sap-sucking insect pests that feed on several plants of agronomical importance. Entomopathogenic fungi are valuable tools for potential aphid control. As part of a selection process, laboratory bioassays were carried with five different concentrations of Aspergillus clavatus (Desmazières), Aspergillus flavus (Link) and Metarhizium anisopliae ((Metschnikoff) Sorokin) spores against the pea aphid, Acyrthosiphon pisum (Harris). Aspergillus isolates induced higher mortalities than M. anisopliae which is a well-known entomopathogen in the literature. Lethal concentrations (LC50 and LC90) were 1.23 x 10^3 and 1.34 x 10^7 spores/ml for A. flavus, 4.95 x 10^2 and 5.65 x 10^7 spores/ml for A. clavatus, and 3.67 x 10^3 and 9.71 x 10^7 spores/ml for M. anisopliae five days after treatment. Mycelia development and sporulation on adult cadavers was observed 48 hours after incubation. The intrinsic growth rate of A. pisum decreased with increased spore concentration for all fungal strains suggesting an increase in pathogen fitness related to a consumption of host resources. In conclusion, Aspergillus species could be useful in aphid control as pest control agents despite their saprophytic lifestyle. This is also in our knowledge the first report of A. clavatus and A. flavus strains pathogenic to aphids. [less ▲]

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See detailBiorefine: Recovery of nutrients and metallic trace elements from different wastes by chemical and biochemical processes
Tarayre, Cédric ULg; Fischer, Christophe ULg; De Clercq, Lies et al

Conference (2014, June 05)

At present, most waste processing operations are not oriented towards the valorization of valuable reusable components such as nitrogen, phosphorus, potassium and even Metallic Trace Elements (MTEs ... [more ▼]

At present, most waste processing operations are not oriented towards the valorization of valuable reusable components such as nitrogen, phosphorus, potassium and even Metallic Trace Elements (MTEs). Currently, sewage sludge, for example is usually used as a fertilizer in agriculture, in energy production or in the field of construction. Ashes originating from sludge incineration contain heavy metals and minerals in large quantities. Manure is mainly used in agriculture, although considerable amounts of nutrients are lost and cause pollution. Digestate is also used in agriculture, but other alternatives have been proposed, such as the energetic valorization. Better valorization of these wastes in agriculture (or other sectors) is however largely constrained by a multitude of legal requirements. An important problematic point is the concentration in MTEs that is found in those wastes. Consequently, recovery of nutrients and MTEsmay be a key solution for optimal valorization of wastes. Many unit operations used in the field of chemical and biochemical engineering (mechanical operations on fluids, solids, mass and heat transfers, chemical reactions, etc.) could be used in order to achieve an efficient recovery yield of nutrients and trace elements. The aim of the BioRefine Project is to make an inventory of all recovery techniques of nutrients and MTEs in five countries: Belgium, France, Germany, United Kingdom and The Netherlands. Pilot plants will also be tested to assess the efficiency of new treatment techniques after which the most efficient processes will be chosen to be applied on a larger scale. In addition, the collected data will be used to propose exploitation scenarios taking into account legal constraints and optimized logistics.This work is supported through an INTERREG IVB NWE programme(ref. 320J-BIOREFINE). [less ▲]

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See detailResearch of phosphate accumulating microorganisms from WWTPs for the recovery of phosphorus from organic wastes (3BV.3.47)
Tarayre, Cédric ULg; Michels, Evi; Buysse, Jeroen et al

Poster (2014, June)

Many wastes containing reusable components, such as nitrogen, phosphorus and potassium, are not exploited through ideal processes. As an example, in Wallonia (Belgium), the main treatment applied to ... [more ▼]

Many wastes containing reusable components, such as nitrogen, phosphorus and potassium, are not exploited through ideal processes. As an example, in Wallonia (Belgium), the main treatment applied to sewage sludge consists in incinerating the material. Such a process is chosen when the heavy metals are too concentrated in the sludge, preventing an agricultural use. However, sewage sludge, as well as manure, slurry and digestate, contain notable amounts of nutrients (nitrogen, phosphorus and potassium). Some Waste Water Treatment Plants (WWTPs) are actually designed in order to promote Phosphate Accumulating Organisms (PAOs), able to store or release phosphorus in accordance with the environmental conditions. The aim of this work is to isolate PAOs from WWTPs and evaluate their applicability to phosphorus recovery from organic wastes. Metagenomics and metabolic properties are also considered. This work is supported by the BioRefine Project, a European project in which various member states focus on recovery of inorganics from organic wastestreams. [less ▲]

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See detailStochastic exposure to sub-lethal high temperature enhances exopolysaccharides (EPS) excretion and improves Bifidobacterium bifidum cell survival to freeze-drying
Nguyen, Huu Thanh ULg; Razafindralambo, Hary; Blecker, Christophe ULg et al

in Biochemical Engineering Journal (2014), 88

Exposure of microbial cells to sub-lethal stresses is known to increase cell robustness. In this work, a two-compartment bioreactor in which microbial cells are stochastically exposed to sub-lethal ... [more ▼]

Exposure of microbial cells to sub-lethal stresses is known to increase cell robustness. In this work, a two-compartment bioreactor in which microbial cells are stochastically exposed to sub-lethal temperature stresses has been used in order to investigate the response of the stress sensitive Bifidobacterium bifidum THT 0101 to downstream processing operations. A stochastic model validated by residence time distribution experiments has shown that in the heat-shock configuration, a two-compartment bioreactor (TCB) allows the exposure of microbial cells to sub-lethal temperature of 42°C for a duration comprised between 100 and 300 seconds. This exposure resulted in a significant increase of cell resistance to freeze-drying by comparison with cells cultivated in conventional bioreactors or in the TCB in the cold shock mode (CS-TCB). The mechanism behind this robustness seems to be related with the coating of microbial cells with exopolysaccharide (EPS), as assessed by the change of the zeta potential and the presence of higher EPS concentration after heat shock. Conditioning of Bifidobacteria on the basis of the heat shock technique is interesting from the practical and economical point of view since this strategy can be directly implemented in the bioreactor during stationary phase preceding cell recovery and freeze-drying [less ▲]

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See detailAccumulation lipidique par Yarrowia lipolytica: un nouvel outil d'observation
Bouchedja, Doria Naïla; Delvigne, Frank ULg; Boudjellal, Abdelghani et al

Conference (2014, April 06)

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See detailUse of on-line flow cytometry for the characterization of microbial stress dynamics during the bioprocess
Brognaux, Alison ULg; Han, Shanshan; Sorensen, Soren et al

Conference (2014, April 03)

Microbial cell population heterogeneity is now recognized as a major source of issues for the development and optimization of bioprocesses. Flow cytometry is a very powerful tool for the follow up of ... [more ▼]

Microbial cell population heterogeneity is now recognized as a major source of issues for the development and optimization of bioprocesses. Flow cytometry is a very powerful tool for the follow up of physiological properties of microbial cells in process-related conditions at the single cell level, and can be used to study the dynamics of segregation directly in bioreactors. In this context, specific interfaces have been developed in order to connect flow cytometer (FC) directly on bioreactor for automated analyses. In this work, we propose a simplified version of such interface and demonstrated its usefulness for multiplexed experiments. This automated FC system has been tested for the follow up of the dynamics of an E. coli pfis::gfpAAV fluorescent bio-reporter and its PI uptake, correlated with membrane permeability. This bioreporter is composed of a fis promoter, a growth dependent promoter-indicator of the nutrient status of cells, fused to a gene expressing an unstable variant of GFP. The results obtained showed that the dynamics of the GFP synthesis is complex and can be attributed to a complex set of biological parameters. Segregation in the membrane permeability has been noticed. This work demonstrates that a simplified version of on-line FC can be used at the process level for the investigation of the dynamics of complex physiological mechanisms. [less ▲]

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See detailCFD-based Compartment model for description of mixing in bioreactors
Delafosse, Angélique ULg; Calvo, Sébastien ULg; Collignon, Marie-Laure ULg et al

in Chemical Engineering Science (2014), 106

In most bioprocesses, it is fundamental to accurately predict the hydrodynamics behavior of bioreactors of different size and its interaction with the biological reaction. Computational Fluid Dynamics can ... [more ▼]

In most bioprocesses, it is fundamental to accurately predict the hydrodynamics behavior of bioreactors of different size and its interaction with the biological reaction. Computational Fluid Dynamics can provide detailed modeling about hydrodynamics and mixing. However, it is computationally intensive, especially when reactions are taken into account. Another way to predict hydrodynamics is the use of “Compartment” or “Network-of-zones” model which are much less demanding in computation time than CFD. However, compartments and fluxes between them are often defined by considering global quantities not representative of the flow complexity. To overcome the limitations of these two methods, a solution is to combine compartment modeling and CFD simulations. The aim of this study is to propose a compartment model where the flow rates between two adjacent compartments are easily computed from the velocity fields obtained by CFD. The mixing evolution predicted by the CFD-based compartment model have been then compared with mixing experiment results. Unlike a CFD mixing simulation and a classical compartment model, the CFD-based compartment model proposed in this work reproduces with a good accuracy the spatial distribution of concentrations during the mixing process and this, without any adjustable parameters. [less ▲]

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See detailScale-down effect on the extracellular proteome of Escherichia coli: correlation with membrane permeability and modulation according substrate heterogeneities
Brognaux, Alison ULg; Francis, Frédéric ULg; Twizere, Jean-Claude ULg et al

in Bioprocess and Biosystems Engineering (2014)

Protein leakage is induced in well-mixed fed-batch bioreactor by comparison with cultures carried out in scale-down conditions. This effect is attributed to a progressive increase of cell membrane ... [more ▼]

Protein leakage is induced in well-mixed fed-batch bioreactor by comparison with cultures carried out in scale-down conditions. This effect is attributed to a progressive increase of cell membrane permeability and the synthesis of several outer-membrane components allowing to cope with substrate limitation commonly found in high-cell density culture. A comparative analysis of protein leakage has thus been performed in well-mixed bioreactors and in scale-down devices. The extracellular proteome of E.coli has been investigated by 2D-gel electrophoresis and identified by subsequent MALDI-TOF analysis. On 110 picked spots, 67 proteins have been identified and the sub-localisation and the molecular function of these proteins have been determined. A majority of the extracellular proteome was composed of outer-membrane and periplasmic proteins (64%) confirming the fact that leakage is involved in high-cell density cultures. About 50% of this extracellular proteome was composed of transport and binding proteins. Furthermore, the more abundant spots on the gel corresponded to porin proteins and periplasmic transporters. In particular, the OmpC porin was found to be very abundant. Moreover, the scale-down effect on this extracellular proteome has been investigated by 2D-DIGE analysis (2-Dimensional Differential in-Gel Electrophoresis) and significant differences have been observed by comparison with culture carried out in well-mixed systems. Indeed, since substrate limitation signal is alleviated in this kind of apparatus, cell permeability was lowered as shown by flow cytometry. In scale-down conditions, protein leakage was thus less abundant. [less ▲]

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See detailScale-down effect on the extracellular proteome of Escherichia coli: correlation with membrane permeability and modulation according substrate heterogeneities
Brognaux, Alison ULg; Francis, Frédéric ULg; Twizere, Jean-Claude ULg et al

in Bioprocess and Biosystems Engineering (2014)

Protein leakage is induced in well-mixed fed-batch bioreactor by comparison with cultures carried out in scale-down conditions. This effect is attributed to a progressive increase of cell membrane ... [more ▼]

Protein leakage is induced in well-mixed fed-batch bioreactor by comparison with cultures carried out in scale-down conditions. This effect is attributed to a progressive increase of cell membrane permeability and the synthesis of several outer-membrane components allowing to cope with substrate limitation commonly found in high-cell density culture. A comparative analysis of protein leakage has thus been performed in well-mixed bioreactors and in scale-down devices. The extracellular proteome of E.coli has been investigated by 2D-gel electrophoresis and identified by subsequent MALDI-TOF analysis. On 110 picked spots, 67 proteins have been identified and the sub-localisation and the molecular function of these proteins have been determined. A majority of the extracellular proteome was composed of outer-membrane and periplasmic proteins (64%) confirming the fact that leakage is involved in high-cell density cultures. About 50% of this extracellular proteome was composed of transport and binding proteins. Furthermore, the more abundant spots on the gel corresponded to porin proteins and periplasmic transporters. In particular, the OmpC porin was found to be very abundant. Moreover, the scale-down effect on this extracellular proteome has been investigated by 2D-DIGE analysis (2-Dimensional Differential in-Gel Electrophoresis) and significant differences have been observed by comparison with culture carried out in well-mixed systems. Indeed, since substrate limitation signal is alleviated in this kind of apparatus, cell permeability was lowered as shown by flow cytometry. In scale-down conditions, protein leakage was thus less abundant. [less ▲]

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See detailUse of on-line flow cytometry for the characterization of microbial stress dynamics during the bioprocess
Brognaux, Alison ULg; Han, Shanshan; Sorensen, Soren et al

Poster (2014, February 07)

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See detailIMPLEMENTATION OF A METAL STRUCTURED PACKING IN A FUNGAL BIOFILM REACTOR FOR THE PRODUCTION OF A RECOMBINANT PROTEIN BY ASPERGILLUS ORYZAE
Zune, Quentin ULg; Delepierre, Anissa; Toye, Dominique ULg et al

in Communications in Agricultural and Applied Biological Sciences (2014, February 07)

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See detailBiofilms from entomopathogenic fungi in mosquito control
Bawin, Thomas ULg; Boukraa, Slimane; Seye, Fawrou et al

Poster (2014, February 07)

Mosquitoes (Diptera: Culicidae) are zoonotic vectors of medical and veterinary importance. As part of an integrated vector control, metabolites secreted by entomopathogenic fungi could be developed as ... [more ▼]

Mosquitoes (Diptera: Culicidae) are zoonotic vectors of medical and veterinary importance. As part of an integrated vector control, metabolites secreted by entomopathogenic fungi could be developed as biopesticides. In this context, filamentous microorganisms growing on a support as biofilm in a liquid medium would offer several advantages in bioreactor regarding performances and metabolites recovery. The production of toxic metabolites by an entomopathogenic fungus Aspergillus flavus in such conditions was assessed. Three initial inoculum levels, i.e. 10^1, 10^3 and 10^6 spores/ml of PYG medium, have been tested in shake flask with or without support. Toxicity tests were performed on Culex quinquefasciatus larvae using dilutions of 1, 2, 4, 6, 8 and 10% of liquid cultures. The results indicated that A. flavus tends to form pellets in submerged culture; the size and the amount of pellets was affected by the initial inoculum level of spores. Under similar conditions, the filaments fixed on a support and didn’t appear in free form in the liquid. Toxicity tests revealed differences between both free and fixed forms. All combined conditions, LC50s ranging up to dilutions of 2.2 and 4.8% were observed within 48 hours. Secretomes could be compared between these culture conditions by proteomic and metabolomic approaches. [less ▲]

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See detailUsing micro-injection technique to assess fungal toxicity in mosquito control
Bawin, Thomas ULg; Boukraa, Slimane; Seye, Fawrou et al

in Communications in Agricultural and Applied Biological Sciences (2014, February 07), 79(1), 181-185

Topical application of insecticidal compounds allows directly exposing these substances on insect tissues and measuring their toxicity while ignoring many factors. However, this technique remains ... [more ▼]

Topical application of insecticidal compounds allows directly exposing these substances on insect tissues and measuring their toxicity while ignoring many factors. However, this technique remains difficult to apply on mosquito larvae considering their aquatic lifestyle. Micro-injection could be used for the direct deposition of toxic compounds in the larvae. Capillaries exhibiting an injection tip with an external diameter of 0.5 mm have been designed from silica tubes. For each treatment, a capillary is mounted on a pump connected to a flow rate regulator. Culex quinquefasciatus larvae were injected with 10^7 spores/ml of entomopathogenic fungi (Aspergillus clavatus, Metarhizium anisopliae, Metarhizium sp.). Mortalities were recorded daily during 72h. The distribution of spores stained with methylene blue and injected into the body of larvae was also observed according to the system described. Results showed that spores were distributed over the whole body. The injection of Aspergillus clavatus, Metarhizium anisopliae and Metarhizium sp spores induced corrected mortalities of 62%, 53% and 57% after 72h, and differed statistically from control groups. Finally, post-mortem emergences of filaments from dead larvae were observed in the case of the three fungal strains confirming spore viability. Injection of inactivated spores (or inert bodies of similar size) could help to reject the hypothesis of a response due to the presence of foreign bodies. [less ▲]

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