References of "Zune, Quentin"
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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 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 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 (2016), 26(5), 617-629

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 detailDesign of a single-species biofilm reactor based on metal structured packing for the production of high added value biomolecules
Zune, Quentin ULg

Doctoral thesis (2015)

Thesis summary In the last decade, numerous single-species biofilm reactors of various configurations have been implemented at lab and pilot scale for the production of chemicals and biological products ... [more ▼]

Thesis summary In the last decade, numerous single-species biofilm reactors of various configurations have been implemented at lab and pilot scale for the production of chemicals and biological products. Compared to their counterparts in submerged cultures, these processes benefit from the specific physiology of biofilms, i.e. high robustness of the microbial system, long-term activity, continuous implementation and low ratio size / productivity. However, the risks of biofouling and the lack of analytical tools for the control and the monitoring of biofilms are obstacles for scale-up strategies. Up to now, single-species biofilm reactors have been mainly confined to the production of metabolites ranging from low (bulk chemicals) to medium (fine chemicals) added values. In this way, there is a need to design efficient single-species biofilm reactors exhibiting good scalability potentials and intended for the production of high added value compounds. In this work, an experimental single-species biofilm reactor has been designed for the production of target molecules derived from metabolic pathways involved in biofilm physiology. On the basis of these criteria, three biological models having good abilities of biofilm formation and secretion performances were selected : - the gram positive bacterium Bacillus subtilis for the production of surfactin, a surface active metabolite involved in biofilm formation. - the filamentous fungus Trichoderma reesei for the production of hydrophobin (HFBII), a surface active protein (7kDa) involved in adhesion process of spores and mycelium on solid surface. - the filamentous fungus Aspergillus oryaze (engineered strain) for the production of a recombinant protein (Gla::GFP) under the control of the glaB promoter specifically activated in solid-state fermentation. The proposed experimental biofilm reactor has the configuration of a trickle-bed bioreactor. The agitation axis of a stirred tank reactor has been removed and replaced by a stainless steel structured packing filling the top of the vessel. The liquid medium, located in the bottom of the vessel is continuously recirculated on the packing element thanks to a peristaltic pump. An ascending air flow is performed above the liquid phase just under the packing element. This thesis reports the screening of the three biological models in the experimental biofilm reactor. The results include the characterization of process performances in terms of biofilm formation and secretion of the target molecule under different operating conditions. An original methodology based on high energy X-ray tomography has been developed to non-invasively visualize and quantify the biofilm colonization inside the packing element. This technique has highlighted that biofilm colonization and liquid phase distribution across the packing are strongly interrelated phenomena. The biofilm of B. subtilis occurring by cell aggregation preferentially developed on solid areas wetted by the liquid. Accordingly, optimal operating conditions improving liquid phase distribution have been defined for biofilm colonization. The fungal biofilm of A. oryzae and T. reesei occuring by cell filamentation equally colonize submerged and aerial surfaces of the packing element. Consequently, another configuration of biofilm reactor comprising a packing element totally immersed in the liquid medium has been investigated. The production yields of surfactin and hydrophobin in the experimental biofilm reactor are respectively 1.25 and 2.64 times greater than those of a submerged culture in a stirred tank reactor. This suggests that surface-active molecules involved in biofilm formation have a real interest for the design of single-species biofilm reactors. Although the Gla::GFP fusion protein is greater produced in the stirred tank culture, its integrity was preserved in the biofilm reactor despite the presence of proteases. This suggests that the quality and the stability of heterologous proteins produced in a fungal biofilm reactor are improved compared with a submerged culture. Finally, the implementation of the biofilm reactor has led to technological progresses including low energy consumption, no foam formation, continuous processing and simplification of downstream process operations. Further experiments should deepen the understanding of structured phenotypic heterogeneity impact on secretion performances in the biofilm reactor. These experiments should consider development of operating conditions allowing for the growth of a thin biofilm homogeneously distributed on the whole surface provided by the packing element in order to optimize nutrients and metabolites mass transfers. The scale-up and the continuous implementation of the process should be also investigated. [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 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 detailBiopesticides: quand les biotechs se mettent au service d’une agriculture durable
Zune, Quentin ULg; Ongena, Marc ULg

Scientific conference (2014, December 10)

Présentation des différents outils de la biotechnologie (Omics et bioprocédés) pour mieux comprendre le fonctionnement des biopesticides microbiens afin d'améliorer leur efficacité lors de leurs ... [more ▼]

Présentation des différents outils de la biotechnologie (Omics et bioprocédés) pour mieux comprendre le fonctionnement des biopesticides microbiens afin d'améliorer leur efficacité lors de leurs applications sur les cultures. [less ▲]

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See detailDesign of a fungal biofilm reactor for recombinant protein production from Aspergillus oryzae
Zune, Quentin ULg; Delepierre, Anissa ULg; Bauwens, Julien ULg et al

Poster (2014, September 07)

Fungi are microorganisms exhibiting high secretive power of various metabolites and have the ability to perform post-translational modifications during protein synthesis. In the field of fermentation ... [more ▼]

Fungi are microorganisms exhibiting high secretive power of various metabolites and have the ability to perform post-translational modifications during protein synthesis. In the field of fermentation industry, they are ideal hosts for secondary metabolites and recombinant protein production. At the industrial-scale, equipments usually required for solid-state or submerged fermentation of filamentous fungi have demonstrated their limitations in terms of productivity, mass transfers or products recovery (1, 2). Recently, fungal biofilm reactors were designed to combine advantages from submerged and solid-state culture and reveal their usefulness for greater secondary metabolites production relative to submerged culture conditions (3). In our work, we propose the design of a fungal biofilm reactor for a recombinant protein production from an Aspergillus oryzae strain containing a GFP reporter gene system under the control of a promoter specifically induced in solid-state conditions. The fungal biofilm reactor is composed of a metal structured packing, having the function of inert support for biofilm growth, immerged or aspersed by a liquid medium. Whereas recombinant protein production is not significantly different at the flask-scale between submerged and biofilm conditions, productivity is higher in the submerged conditions at the bioreactor-scale. Presence of recombinant proteins entrapped in the biofilm matrix highlights a diffusion constraint and a lower mass transfer in our fungal biofilm reactor. However, persistence of a free liquid biomass of low viscosity and fungal biomass retention on the support are attractive for the implementation of a continuous process in our fungal biofilm reactor. Further studies will consider a 2-D proteomic comparison of the extracellular medium from fungal biofilm reactor and submerged culture conditions in order to better understand proteins secretion and identify over-expressed proteins in biofilm conditions. [less ▲]

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See detailImplications of microbial phenotypic heterogeneity in large-scale bioprocessing conditions
Delvigne, Frank ULg; Gorret, Nathalie; Molina-Jouve, Carole et al

Conference (2014, September)

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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 detailHigh-energy X-ray tomography analysis of a metal packing biofilm reactor for the production of lipopeptides by Bacillus subtilis
Zune, Quentin ULg; Soyeurt, Delphine; Toye, Dominique ULg et al

in Journal of Chemical Technology & Biotechnology (2014), 89

BACKGROUND: Whereas multi-species biofilm reactors are commonly used for the treatment of liquid and solid wastes, new strategies are progressing for the development of single species biofilm for the ... [more ▼]

BACKGROUND: Whereas multi-species biofilm reactors are commonly used for the treatment of liquid and solid wastes, new strategies are progressing for the development of single species biofilm for the production of high-value metabolites. Technically, this new concept relies on the design of bioreactors able to promote biofilm formation and on the identification of the key physico-chemical parameters involved in biofilm formation. RESULTS: An experimental setting comprising a liquid continuously recirculated on a metal structured packing has been used to promote Bacillus subtilis GA1 biofilm formation. The colonization of the packing has been visualized non-invasively by X-ray tomography. This analysis revealed an uneven, conical, distribution of the biofilm inside the packing. Compared with a submerged culture carried out in a stirred tank reactor, significant modification of the lipopeptide profile has been observed in the biofilm reactorwith the disappearance of fengycin and iturin fractions and an increase of the surfactin fraction. In addition, considering the biofilm reactor design, no foam formation has been observed during the culture. CONCLUSIONS: The configuration of this biofilm reactor set-up allows for a higher surfactin production by comparison with a submerged culture while avoiding foam formation. Additionally, scale-up could easily be performed by increasing the number of packing elements. [less ▲]

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See detailUse of on-line flow cytometry to detect segregation in the microbial population
Brognaux, Alison ULg; Zune, Quentin ULg; Han, Shanshan et al

Poster (2013, October 08)

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See detailBiofilm formation on metal structured packing for the production of high added value biomolecules
Zune, Quentin ULg; Toye, Dominique ULg; Delvigne, Frank ULg et al

in Récents Progrès en Génie des Procédés (2013)

Many white biotechnology bioprocesses apply techniques from chemical engineering based on bioreactors with mechanical stirring system commonly employed in pharmaceutical sector, food industry or energy ... [more ▼]

Many white biotechnology bioprocesses apply techniques from chemical engineering based on bioreactors with mechanical stirring system commonly employed in pharmaceutical sector, food industry or energy field (Dasilva, 2004). As in chemical engineering, scale-up of these bioprocesses induces physicochemical constraints that affect physiological pathways and decrease performances. In this context, it is essential to think new bioprocesses better suited to physiology of microorganisms, minimizing physicochemical constraints. The aim of this work consists to use stainless steel structured packing (SSP) with high specific area (500-750 m²/m³) as inert support for biomass immobilization in order to produce high added value biomolecules. These bioreactors are biocatalysts in which microbial system is immobilized biomass on the form of a biofilm performing bioconversion of a substrate into a specific product (Rosche, 2009). In this study, an experimental setting containing a SSP reproduces solid-state fermentation (SSF) like conditions. Two well known microorganisms for their ability to form biofilm and secrete metabolites are tested in the experimental setting : Bacillus subtilis for its lipopeptides and Aspergillus oryzae for its glucoamylase. Effectiveness of the bioprocess in term of dynamic of the excretion of the target biomolecule is compared with a classical submerged culture (SmF). For lipopeptides production from B. subtilis, SSP is located in a 20L bioreactor continuously aspersed by liquid medium required to the growth of the biofilm. In the case of A. oryzae, the SSP is partially immerged in a 250 mL shake flask. X-ray tomography of the SSP allows non-invasive visualization and quantification of biofilm repartition inside the support. Implementation of SSP permits almost total immobilization of biomass on the form of a mono-species biofilm to the detriment of the liquid phase. Processing of images obtained by X-ray tomography of the SSP provides relevant information for the optimization of the bioprocess. For both microorganism species, results indicate the influence of parameters such as hydrodynamics, aeration rate and microorganism specificity, on the biofilm morphology inside the support and the performances of the bioprocess. SSF-like conditions in the experimental setting lead to technologic progress, such as absence of foam formation, persistence of the microbial system, and improve the dynamic of metabolites excretion compared with conditions imposed by the submerged culture. Further experiment will consider hydrodynamics aspects and amount of carbon source on effectiveness of the bioprocess. [less ▲]

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See detailImplementation of structured metal packing for the design of biofilm reactor : analysis by high energy X-ray tomography and application to the production of lipopeptides by Bacillus subtilis
Zune, Quentin ULg; Soyeurt, Delphine; Ongena, Marc ULg et al

Poster (2012, October 08)

1. Whereas multi-species biofilm reactors are commonly used for treatments of water and gas effluents, new strategies are arising for the development of mono-species biofilm reactors in order to produce ... [more ▼]

1. Whereas multi-species biofilm reactors are commonly used for treatments of water and gas effluents, new strategies are arising for the development of mono-species biofilm reactors in order to produce high added value molecules. Thus, it is required to design new bioreactors able to promote the growth of the biomass on the form of a biofilm and to identify the key physico-chemical parameters involved in order to optimize the bioprocess. 2. Aim of this study was to investigate a pilot-scale biofilm reactor comprising a metal structured packing promoting growth of Bacillus subtilis as a biofilm for the production of lipopeptides, high added value compounds with high surface active properties. 3. In this work, the mechanical stirring system of a 20L stirred tank bioreactor has been removed and replaced by a metal structured packing positioned in the headspace of the vessel above a liquid phase. The culture medium is continuously recirculated on the packing thanks to a peristaltic pump and air supply is performed just above the liquid phase under the packing. High energy X-ray tomography was used to estimate non-invasively the biofilm distribution inside the packing and permitted to define parameters that affect scale-up. Performances of the biofilm reactor were compared with a submerged culture in a stirred tank reactor in terms of lipopeptides production. 4. After 72 hours of fermentation, 94 % of the total biomass adheres onto the metal packing on the form of a biofilm. The colonization of this latter has been visualized non-invasively by X-ray tomography directly inside the packing and shows a conical repartition of the biofilm mass (about 25% of the total volume of the packing) as well as the presence of clogging. However, unlike the submerged culture, no foam formation appeared during fermentation and surfactin yield reaches 345,4 ± 32,8 mg / L for the biofilm reactor against 277,3 ± 34,4 mg / L in the stirred tank reactor. 5. In conclusion, this experimental setting leads to a major technological progress avoiding foam formation and increasing surfactin production. Nevertheless, significant improvements are required at the level of the biofilm distribution in thin layers inside the packing in order to increase mass transfer and lipopeptides recoveries. Further investigations will be devoted to the optimization of the physico-chemical parameters involved in biofilm distribution. [less ▲]

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