References of "Delvigne, Frank"
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See detailHistopathological effects of Aspergillus clavatus (Ascomycota: Trichocomaceae) on larvae of the southern house mosquito, Culex quinquefasciatus (Diptera: Culicidae)
Bawin, Thomas ULg; Seye, Fawrou; Boukraa, Slimane ULg et al

in Fungal Biology (in press)

Aspergillus clavatus (Ascomycota: Trichocomaceae) was previously found to be an opportunistic pathogen of mosquitoes (Diptera: Culicidae). In the present study, the mechanism leading to its insecticidal ... [more ▼]

Aspergillus clavatus (Ascomycota: Trichocomaceae) was previously found to be an opportunistic pathogen of mosquitoes (Diptera: Culicidae). In the present study, the mechanism leading to its insecticidal activity was investigated regarding histological damages on Culex quinquefasciatus larvae exposed to A. clavatus spores. Multiple concentration assays using spore suspensions (0.5 x 10^8 to 2.5 x 10^8 spores/ml) revealed 17.0% to 74.3% corrected mortalities after 48 h exposure. Heat-deactivated spores induced a lower mortality compared to non-heated spores suggesting that insecticidal effects are actively exerted. Spore-treated and untreated larvae were prepared for light microscopy as well as for scanning and transmission electron microscopy. Spores failed to adhere to the external body surface (except the mouth parts) of these aquatic immature stages but progressively filled the digestive tract where their metabolism seemed to activate. In parallel, the internal tissues of the larvae, i.e. the midgut wall, the skeletal muscles, and the cuticle-secreting epidermis, were progressively destroyed between 8 and 24 h of exposure. These observations suggest that toxins secreted by active germinating spores of A. clavatus in the digestive tract altered the larval tissues, leading to their necrosis and causing larval death. Fungal proliferation and sporulation then occurred during a saprophytic phase. A. clavatus enzymes or toxins responsible for these pathogenic effects need to be identified in further studies before any use of this fungus in mosquito control. [less ▲]

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See detailProduction of two entomopathogenic Aspergillus species and insecticidal activity against the mosquito Culex quinquefasciatus compared to Metarhizium anisopliae
Bawin, Thomas ULg; Seye, Fawrou; Boukraa, Slimane ULg et al

in Biocontrol Science & Technology (in press)

Entomopathogenic micro-organisms including fungi have become increasingly studied for integrated pest management. The spore productivity and insecticidal activity of two opportunistic insect pathogenic ... [more ▼]

Entomopathogenic micro-organisms including fungi have become increasingly studied for integrated pest management. The spore productivity and insecticidal activity of two opportunistic insect pathogenic Aspergillus species (namely: Aspergillus clavatus Desmazieres and Aspergillus flavus Link (Ascomycota: Eurotiales, Trichocomaceae)) were compared to Metarhizium anisopliae sensu lato (Metchnikoff) Sorokin (Ascomycota: Hypocreales, Clavicipitaceae) for mosquito (Diptera: Culicidae) control. The production of aerial spores on wheat bran and white rice was investigated in solid-, semi-solid-, and liquid-state media supplemented with a nutritive solution. Wheat bran-based media were suitable for spore production and increased the spore yield in solid-state from 3 to 7 fold: A. clavatus produced 48.4 ± 5.2 and 15.7 ± 1.6 x 10^8 spores/g, A. flavus produced 22.3 ± 4.1 and 3.1 ± 2.5 x 10^8 spores/g, and M. anisopliae produced 39.6 ± 6.5 and 13.1 ± 2.6 x 10^8 spores/g of wheat bran or white rice, respectively. A. clavatus, A. flavus and M. anisopliae spores harvested from wheat bran-based solid-state media showed lethal concentrations (LC50) of 1.1, 1.8, and 1.3 x 10^8 spores/ml against Culex quinquefasciatus Say larvae in 72 h. Because A. clavatus and M. anisopliae displayed similar features when cultured under these conditions, our results suggest that insect pathogenic Aspergillus species may be as productive and virulent against mosquito larvae as a well-recognized entomopathogenic fungus. Wheat bran could advantageously be used in large-scale fermentation for a possible cost-effective pest control using these fungi. [less ▲]

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See detailPhenotypic variability in bioprocessing conditions can be tracked on the basis of on-line flow cytometry and fits to a scaling law
Baert, Jonathan ULg; Kinet, Romain ULg; Brognaux, Alison ULg et al

in Biotechnology Journal (in press)

Noise in gene and protein expression is a major cause for bioprocess deviation. However, this phenomenon has been only scarcely considered in real bioprocessing conditions. In this work, a scaling-law ... [more ▼]

Noise in gene and protein expression is a major cause for bioprocess deviation. However, this phenomenon has been only scarcely considered in real bioprocessing conditions. In this work, a scaling-law derived from a genome-scale studies based on GFP reporter systems has been calibrated to an on-line flow cytometry device, allowing thus to get an insight at the level of promoter activity and associated noise during a whole microbial culture carried out in bioreactor. We show that most of the GFP reporter systems investigated and thus corresponding genes could be included inside the area covered by the scaling-law. The experimental results suggest that this scaling-law could be used to predict the dynamics of promoter activity, as well as the associated noise, in bioprocessing conditions. The knowledge acquired throughout this work could be used for the design of more robust expression systems. [less ▲]

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See detailScalable temperature induced stress for the large-scale production of functionalized Bifidobacteria
Nguyen, Huu Thanh ULg; Razafindralambo, Hary; Richel, Aurore ULg et al

in Journal of Industrial Microbiology & Biotechnology (in press)

The application of sub-lethal stresses is known to be an efficient strategy to enhance survival of probiotic bacteria during drying processes. In this context, we previously showed that the application of ... [more ▼]

The application of sub-lethal stresses is known to be an efficient strategy to enhance survival of probiotic bacteria during drying processes. In this context, we previously showed that the application of heat stress upon the entry into stationary phase increased significantly the viability of Bifidobacterium bifidum. However, this heat shock has been considered only in small scale bioreactor and no information is available for a possible scaling-up strategy. Five different operating scales (0.2 L, 2 L, 20 L, 200 L and 2000 L) have thus been tested and the results showed that the viability of B. bifidum increases from 3.15 to 6.57 folds, depending on the scale considered. Our observations pointed out the fact that the heat stress procedure is scalable according to the main outcome, i.e. increases in cell viability, but other factors have to be taken into account. Among these factors, dissolved carbon dioxide seems to play a significant role since it explain the differences observed between the test performed at lab-scale and in industrial conditions. [less ▲]

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See detailMetabolic variability in bioprocessing : implications of microbial phenotypic heterogeneity
Delvigne, Frank ULg; Zune, Quentin ULg; Lara, Alvaro et al

in Trends in Biotechnology (in press)

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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 detailFungal biofilm reactor improves the quality of a fusion protein GLA::GFP produced by Aspergillus oryzae
Zune, Quentin ULg; Delepierre, Anissa ULg; Bauwens, Julien ULg et al

Poster (2015, October)

Fungal biofilm is known to promote the excretion of secondary metabolites, in accordance with solid-state related physiological mechanisms. In this work, the potentialities of fungal biofilm will be ... [more ▼]

Fungal biofilm is known to promote the excretion of secondary metabolites, in accordance with solid-state related physiological mechanisms. In this work, the potentialities of fungal biofilm will be investigated in the context of the production of a Gla::GFP fusion protein by Aspergillus oryzae. Since the production of this protein is under the control of the promoter glaB, specifically induced in solid-state fermentation, biofilm mode of culture is expected to enhance the global productivity. However, we found that the glaB promoter is also activated in submerged bioreactor and the fusion protein production is higher in this mode of culture. This result is related to the high shear stress leading to biomass autolysis and leakage of intracellular fusion protein into the extracellular medium. Moreover, 2D-gel electrophoresis highlights preservation of the fusion protein integrity produced in biofilm conditions whereas proteolysis strongly affects fusion protein recovery in the submerged cultures performed at high stirring rate. Fungal biofilm reactor design was then further investigated and the scale-up potentialities were evaluated. Indeed, the specific design investigated in this work involves the use of metal structured packing exhibiting a high specific area and that can be easily expanded to large-scale bioprocessing conditions. [less ▲]

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See detailFluorescent reporter libraries as useful tools for optimizing microbial cell factories: a review of the current methods and applications
Delvigne, Frank ULg; Pêcheux, Hélène; Tarayre, Cédric ULg

in Frontiers in Bioengineering and Biotechnology (2015), 3

The use of genetically encoded fluorescent reporters allows speeding up the initial optimization steps of microbial bioprocesses. These reporters can be used for determining the expression level of a ... [more ▼]

The use of genetically encoded fluorescent reporters allows speeding up the initial optimization steps of microbial bioprocesses. These reporters can be used for determining the expression level of a particular promoter, not only the synthesis of a specific protein but also the content of intracellular metabolites. The level of protein/metabolite is thus proportional to a fluorescence signal. By this way, mean expression profiles of protein/ metabolites can be determined non-invasively at a high-throughput rate, allowing the rapid identification of the best producers. Actually, different kinds of reporter systems are available, as well as specific cultivation devices allowing the on-line recording of the fluorescent signal. Cell-to-cell variability is another important phenomenon that can be integrated into the screening procedures for the selection of more efficient microbial cell factories. [less ▲]

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See detailEuler–Lagrange approach to model heterogeneities in stirred tank bioreactors – Comparison to experimental flow characterization and particle tracking
Delafosse, Angélique ULg; Calvo, Sébastien ULg; Collignon, Marie-Laure ULg et al

in Chemical Engineering Science (2015), 134

The aim of this work is the validation of an Euler–Lagrange modeling approach coupling a CFD-based compartment model (Eulerian approach) and a stochastic model based on a Continuous-Time Markov Chain ... [more ▼]

The aim of this work is the validation of an Euler–Lagrange modeling approach coupling a CFD-based compartment model (Eulerian approach) and a stochastic model based on a Continuous-Time Markov Chain (Lagrangian approach). The turbulent flow structure and the mixing process in a bioreactor stirred by an axial Mixel TT impeller is characterized by PIV and tracer experiments. Comparison between experimental and numerical data shows that the CFD-based compartment model is able to reproduce accurately the spatial heterogeneities inside the bioreactor. The trajectory of a small tracer particle which perfectly follows the fluid flow is measured by optical trajectography. It is then simulated by a stochastic model which is either based on an homogeneous or on an inhomogeneous Continuous Time Markov Chain (CTMC). Comparison of residence and circulation time distributions in three zones defined inside the bioreactor shows that the inhomogeneous CTMC model predicts with an excellent accuracy the particle trajectories inside the bioreactor. The modeling approach proposed here could be an useful tool to design scale-down bioreactors in order to reproduce at lab-scale the stress levels encountered in large-scale production bioreactors and to characterize and compare different bioreactor configurations. [less ▲]

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See detailLa lutte contre les moustiques (Diptera: Culicidae): diversité des approches et application du contrôle biologique
Bawin, Thomas ULg; Seye, Fawrou; Boukraa, Slimane ULg et al

in Canadian Entomologist (2015), 147(4), 476-500

Many mosquito (Diptera: Culicidae) species are zoonotic vectors responsible for numerous infectious diseases of medical and veterinary importance. Currently, changes in the vectors’ geographical ... [more ▼]

Many mosquito (Diptera: Culicidae) species are zoonotic vectors responsible for numerous infectious diseases of medical and veterinary importance. Currently, changes in the vectors’ geographical distribution induced chiefly by anthropogenic factors are accompanied by emerging and reemerging infectious diseases in Europe and North America. Since the advent of synthetic insecticides during the Second World War, mosquitoes are the object of considerably expanded and deepened research. In an integrated pest management context, means of control are now mainly classified as: (1) environmental management and physical control, (2) chemical control, (3) genetic control, and (4) biological control by means of entomophagous predators and entomopathogenic microorganisms. In this context, these last have significant potential because of their ability to infect and kill their host with more or less targeted selectivity. This article proposes to emphasize biological control among other techniques in mosquito control, and to assess the potential and the opportunities offered by entomopathogenic bacteria, viruses and fungi. Finally, their use as biopesticides is discussed. [less ▲]

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See detailImpact of phenotypic heterogeneity and metabolic specialisation on metabolic engineering strategies: case of study of E.coli as a representative microbial cell factory
Brognaux, Alison ULg; Delepierre, Anissa ULg; Pecheux, Hélène ULg et al

Conference (2015, July 21)

The goal of this research is to directly highlight the simultaneous occurrence of several phenotypes with distinct metabolic function among clonal population of E. coli., often used for recombinant ... [more ▼]

The goal of this research is to directly highlight the simultaneous occurrence of several phenotypes with distinct metabolic function among clonal population of E. coli., often used for recombinant protein and pDNA production in bioprocesses. This phenotypic heterogeneity, first due to the noise, is reinforced by environmental heterogeneities occurring at large scale during fed-batch processes. This phenotypic heterogeneity has been tracked according to GFP reporter strains (biosensors) that circulate in the bioreactor and encounter environmental heterogeneities. First, we have highlighted the simultaneous occurrence of several phenotypes with distinct metabolic functions. Indeed, a diversity of glucose uptake strategies has recently been noticed with the PtsG and MglABC transporters. Moreover, when E.coli encounters zones of glucose excess, acetate is produced through the overflow metabolism. Only the sub-population with high acs expression (acetate transporter) could consume this acetate. GFP reporter strains have thus been constructed for PtsG, MgIABC and acs genes. In addition of the stable GFP, two destabilized GFP variants for each gene have been used to obtain more instantaneous responses. The response of these biosensors have been followed by on-line flow cytometry. In the end, this experimental strategy for direct phenotyping at the single cell level will also be used to investigate the impact of metabolic engineering strategies on phenotypic heterogeneity, robustness and fitness of microbial population in industrial conditions. [less ▲]

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See detailBIOREFINE Project – Recovery of useful nutrients from organic wastes produced in five European countries (3AV.3.55)
Tarayre, Cédric ULg; De Clercq, Lies; Michels, Evi et al

Poster (2015, June 01)

At this time, many wastes are used or eliminated through processes that do not really consider their potential applications. Such wastes contain useful nutrients (nitrogen, phosphorus and potassium), the ... [more ▼]

At this time, many wastes are used or eliminated through processes that do not really consider their potential applications. Such wastes contain useful nutrients (nitrogen, phosphorus and potassium), the importance of which has been demonstrated in agriculture for many years. The composition of wastes is highly heterogeneous, which makes treatment techniques more difficult to apply on a large scale. Sewage sludge is usually used as a fertilizer in agriculture, in energy production or in the field of construction. The main use of manure is agriculture, although considerable amounts of nutrients are lost and cause pollution. Digestate is also used in agriculture, but other alternatives have been proposed. Ashes should also be highlighted, although they do not contain nitrogen, which is lost into the atmosphere during the combustion process. Finally, household and industrial wastes are resources that should be considered as well. Those different types of wastes could be recycled to produce environment-friendly fertilizers. Here, we propose to investigate these opportunities inside five European countries (Belgium, France, Germany, United Kingdom and The Netherlands) through five work packages with the BioRefine Project. [less ▲]

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See detailMultiple analyses of microbial communities applied to the gut of the wood-feeding termite Reticulitermes flavipes fed on artificial diets
Tarayre, Cédric ULg; Bauwens, Julien ULg; Mattéotti, Christel et al

in Symbiosis (2015)

The purpose of this work was the observation of the differences between the microbial communities living in the gut of the termite Reticulitermes flavipes fed on different diets. The termites were fed on ... [more ▼]

The purpose of this work was the observation of the differences between the microbial communities living in the gut of the termite Reticulitermes flavipes fed on different diets. The termites were fed on poplar wood (original diet) and artificial diets consisting of crystalline cellulose (with and without lignin), α-cellulose (with and without lignin) and xylan. The termites were then dissected and the protist communities were analyzed through microscopy, leading to the conclusion that protist species are strongly influenced by diets. BIOLOG ECO Microplates® were used to assess the metabolic properties of the different types of consortia, highlighting strong differences on the basis of principal component analysis and calculation of similarity rates. The microorganisms were cultivated in liquid media corresponding to the artificial diets before being characterized through a metagenetic analysis of gut microbiota (16S ribosomal DNA). This analysis identified several phyla: Acidobacteria, Actinobacteria, Bacteroidetes, Cyanobacteria, Fibrobacteres, Firmicutes, Nitrospirae, OP9, Planctomycetes, Proteobacteria, Spirochaetes, TM6, Tenericutes, Verrucomicrobia and WS3. The OTUs were also determined and confirmed the abundance of Proteobacteria, Bacteroidetes, Firmicutes and Verrucomicrobia. It was possible to isolate several strains from the liquid media, and one bacterium and several fungi were found to produce interesting enzymatic activities. The bacterium Chryseobacterium sp. XAvLW produced α-amylase, β-glucosidase, endo-1,4-β-D-glucanase, endo-1,4-β-D-xylanase and filter paper-cellulase, while the fungi Sarocladium kiliense CTGxxyl and Trichoderma virens CTGxAviL generated the same activities added with endo-1,3-β-D-glucanase. [less ▲]

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See detailA fungal biofilm reactor based on metal structured packing improves the quality of a Gla::GFP fusion protein produced by Aspergillus oryzae
Zune, Quentin ULg; Delepierre, Anissa ULg; Gofflot, Sebastien et al

in Applied Microbiology and Biotechnology (2015)

Fungal biofilm is known to promote the excretion of secondary metabolites in accordance with solid-state related physiological mechanisms. This work is based on the comparative analysis of classical ... [more ▼]

Fungal biofilm is known to promote the excretion of secondary metabolites in accordance with solid-state related physiological mechanisms. This work is based on the comparative analysis of classical submerged fermentation with a fungal biofilm reactor for the production of a Gla::GFP fusion protein by Aspergillus oryzae. The biofilm reactor comprises a metal structured packing allowing the attachment of the fungal biomass. Since the production of the target protein is under the control of the promoter glaB, specifically induced in solid-state fermentation, the biofilm mode of culture is expected to enhance the global productivity. Although production of the target protein was enhanced by using the biofilm mode of culture, we also found that fusion protein production is also significant when the submerged mode of culture is used. This result is related to high shear stress leading to biomass autolysis and leakage of intracellular fusion protein into the extracellular medium. Moreover, 2D-gel electrophoresis highlights the preservation of fusion protein integrity produced in biofilm conditions. Two fungal biofilm reactor designs were then investigated further, i.e. with full immersion of the packing or with medium recirculation on the packing, and the scale-up potentialities were evaluated. In this context, it has been shown that full immersion of the metal packing in the liquid medium during cultivation allows for a uniform colonization of the packing by the fungal biomass and leads to a better quality of the fusion protein. [less ▲]

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See detailDetermination of Zinc, Cadmium and Lead Bioavailability in Contaminated Soils at the Single-Cell Level by a Combination of Whole-Cell Biosensors and Flow Cytometry
Hurdebise, Quentin ULg; Tarayre, Cédric ULg; Fischer, Christophe ULg et al

in Sensors (2015)

Zinc, lead and cadmium are metallic trace elements (MTEs) that are widespread in the environment and tend to accumulate in soils because of their low mobility and non-degradability. The purpose of this ... [more ▼]

Zinc, lead and cadmium are metallic trace elements (MTEs) that are widespread in the environment and tend to accumulate in soils because of their low mobility and non-degradability. The purpose of this work is to evaluate the applicability of biosensors as tools able to provide data about the bioavailability of such MTEs in contaminated soils. Here, we tested the genetically-engineered strain Escherichia coli pPZntAgfp as a biosensor applicable to the detection of zinc, lead and cadmium by the biosynthesis of green fluorescent protein (GFP) accumulating inside the cells. Flow cytometry was used to investigate the fluorescence induced by the MTEs. A curvilinear response to zinc between 0 and 25 mg/L and another curvilinear response to cadmium between 0 and 1.5 mg/L were highlighted in liquid media, while lead did not produce exploitable results. The response relating to a Zn2+/Cd2+ ratio of 10 was further investigated. In these conditions, E. coli pPZntAgfp responded to cadmium only. Several contaminated soils with a Zn2+/Cd2+ ratio of 10 were analyzed with the biosensor, and the metallic concentrations were also measured by atomic absorption spectroscopy. Our results showed that E. coli pPZntAgfp could be used as a monitoring tool for contaminated soils being processed. [less ▲]

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