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Institute for Bioengineering and Biosciences
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Study of Legionella pneumophila Virulence in a Galleria mellonella Infection Model

Study of Legionella pneumophila Virulence in a Galleria mellonella Infection Model | iBB | Scoop.it

Legionella pneumophila is ubiquitous in freshwater environments and in man-made water systems. Most studies on L. pneumophila virulence focus on clinical strains and isolates from man-made environments, but little is known about the nature and extent of virulence  in strains isolated from natural environments. In a recent publication in Frontiers in Cellular and Infection Microbiology, Leonilde M. Moreira from BSRG-iBB, in collaboration with the group of Joana Costa from University of Coimbra, studied whether unrelated L. pneumophila strains, isolated from different environments, displayed differences in virulence, using the infection model Galleria mellonella. The work suggests that in water distribution systems, environmental filtering selection and biotic competition structure L. pneumophila populations by selecting more resilient and adapted strains that can rise to high concentration if unchecked. Click on title to learn more.

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Role of D-Lactate Dehydrogenase in Burkholderia Aggregation

Role of D-Lactate Dehydrogenase in Burkholderia Aggregation | iBB | Scoop.it
Cystic fibrosis patients often suffer from chronic respiratory infections caused by microorganisms. Among them are the Burkholderia cepacia complex bacteria, which cause progressive deterioration of lung function. Burkholderia pathogenesis is multifactorial as they express several virulence factors, form biofilms, and are highly resistant to antimicrobial compounds, making their eradication from the CF patients' airways very difficult. As Burkholderia is commonly found in CF lungs in the form of cell aggregates and biofilms, the need to investigate the mechanisms of cellular aggregation is obvious. In a recent study published in Applied and Environmental Microbiology, a team of researchers from iBB and Instituto Gulbenkian de Ciência led by Leonilde Moreira from BSRG-iBB has demonstrated the importance of a D-lactate dehydrogenase and LdhR regulator in controlling carbon overflow, planktonic cellular aggregates, and surface-attached biofilm formation. This not only enhances the understanding of Burkholderia pathogenesis but can also lead to the development of drugs to circumvent cell aggregation and biofilm formation (Click on title to learn more).
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Purification of M13 Bacteriophage With Ionic Liquids

Purification of M13 Bacteriophage With Ionic Liquids | iBB | Scoop.it

Bacteriophage M13 is widely used in phage display technology and bionanotechnology, and is considered a possible antibacterial therapeutic agent, among other applications. However, conventional phage purification involves many steps, with high costs and significant product loss (~ 60%). Researchers at BERG-iBB, in collaboration with Isabel Marrucho from Centro de Química Estrutural and João Gonçalves from iMed, propose a scalable M13 bacteriophage purification process using a novel stationary phase based on a polymeric ionic liquid (PIL) with a positively charged backbone structure. The work is part of Maria João Jacinto´s PhD thesis in Bioengineering. Click on title to learn more in Journal of Chromatography A.

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Stability of Aerobic Granules During Bioreactor Operation

Stability of Aerobic Granules During Bioreactor Operation | iBB | Scoop.it
Aerobic granular sludge technology is regarded as the upcoming new standard for biological treatment of wastewaters. Aerobic granules (AG) are dense, compact, self-immobilized microbial aggregates that allow better sludge-water separation and higher biomass concentrations in bioreactors than conventional activated sludge aggregates. This brings potential advantages in terms of investment cost, energy consumption and footprint. In a recent paper in Biotechnology Advances, BERG-researchers Helena Pinheiro, Nídia Lourenço and Rita Franca, and Mark van Loosdrecht from the University of Delft, provide an up-to-date review of the literature on AG stability. The identification of key factors for long-term stability of AG, the underlying mechanisms and operational conditions leading to granule disintegration are described, alongside with possible practical solutions to improve long-term stability. Click on title to learn more.
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The Prokaryotic Consortium of the Gorgonian Coral Eunicella labiata

The Prokaryotic Consortium of the Gorgonian Coral Eunicella labiata | iBB | Scoop.it
Microbial communities inhabiting gorgonian corals provide hosts with nutrient provision and chemical defence. BSRG-iBB researchers led by Rodrigo Costa have found that the microbiome of the gorgonian Eunicella labiata is diverse, distinct from seawater, and rich in specific bacterial phylotypes. Furthermore, they have shown that many dominant E. labiata symbionts can be cultivated. Click on tilte to learn more about the research in FEMS Microbiology and Ecology.
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Enzymatic Synthesis of Chiral Amino-Alcohols in a Continuous-Flow Microreactor System

Enzymatic Synthesis of Chiral Amino-Alcohols in a Continuous-Flow Microreactor System | iBB | Scoop.it

Chiral amino-alcohols are key building blocks in the production of optically pure pharmaceuticals. While many of these compounds can be synthesized enzymatically, the rapid process development and intensification continues to be challenging. A recent contribution co-authored by BERG-iBB researchers Pedro Fernandes and Filipe Carvalho with UCL colleagues led by Nicolas Szita presents a novel microreactor-based approach based on free enzymes for the synthesis of the chiral building block ABT. The system couples a transketolase- and a transaminase-catalyzed reaction and is operated continuously. The presented approach illustrates how continuous-flow microreactors can be applied for the design and optimization of biocatalytic processes. Click on title to learn more about the work on Biotechnology and Bioengineering. Photo: 3D model of transketolase from Escherichia coli (pdb 1QGD).

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Alginate Encapsulation of Human Mesenchymal Stem/Stromal Cells

Alginate Encapsulation of Human Mesenchymal Stem/Stromal Cells | iBB | Scoop.it
Mesenchymal stem/stromal cells (MSC) can stimulate tissue regeneration during wound healing. SCERG-iBB researchers Cláudia Lobato Silva, Frederico Ferreira and Joaquim Cabral, working in collaboration with Todd McDevittt from UCSF, propose the use of alginate encapsulation as a strategy to assess the activity of 3D- and 2D-cultured human bone marrow MSC in the setting of wound repair and regeneration processes. The study, published in Journal of Biotechnology, shows that encapsulation within alginate matrices protects MSC from oxidative stress and extends its therapeutic potential. Click on title to learn more.
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Colorimetric Detection of D-dimer in a Paper-based Device

Colorimetric Detection of D-dimer in a Paper-based Device | iBB | Scoop.it
Microfluidic paper-based analytical devices (μPADs) have emerged in the last years as an ideal technology to develop Point-Of-Care testing. In an article just published in Analytical Biochemistry, Sónia Ruivo, Ana Azevedo and Miguel Prazeres from BERG-iBB describe a μPADs immunoassay for detection of D-dimer, a blood native biomarker helpful in diagnosing diseases like disseminated intravascular coagulation, deep vein thrombosis and pulmonary embolism. The μPADs were successfully used to detect concentrations of D-dimer in simulated plasma samples as low as 15 ng/mL. The tests were further adapted to yield positive results only for samples with D-dimer concentrations above the clinically relevant threshold range of 250–500 ng/mL. Click on title to learn more.
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Production of P(3HB), Xylitol and Xylonic Acid by Burkholderia sacchari

Production of P(3HB), Xylitol and Xylonic Acid by Burkholderia sacchari | iBB | Scoop.it
Lignocellulosic materials have been suggested as alternative sustainable carbon sources for bioproduction of biofuels and biomaterials. In a recent publication in New Biotechnology, BERG-iBB researchers led by Teresa Cesário and Manuela Fonseca, in collaboration with Catarina Almeida from ISCSEM and Conceição Oliveira from CQE-IST,  describe the efficient production of poly-3-hydroxybutyrate (P(3HB)) by Burkholderia sacchari using glucose-xylose mixtures that simulate different types of lignocellulosic hydrolysate. The production of xylitol and xylonic acid by the bacterium under specific substrate concentrations is also reported for the first time. Click on title to learn more.

Photo details: section of a poly-hydroxybutyrate filament, Neptilo, wikipedia, public domain.
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Using Biotechnology to Solve Engineering Problems

Using Biotechnology to Solve Engineering Problems | iBB | Scoop.it

In an increasingly miniaturized technological world, reliable, economical, and ecologic non-destructive testing (NDT) techniques are necessary to detect unprecedented micro defects that conventional NDT cannot perceive. Dr. Carla CCR de Carvalho (BERG-iBB) and colleagues from FCT-UNL, led by Prof. Telmo Santos, used bacterial cells to identify micro and nano defects in aluminium, steel and copper alloys. The properties of bacterial cells, such as hydrophobicity, production of biosurfactants and response to electric and magnetic fields, were further exploited, allowing the detection of defects in e.g. microscrews for dental implants and engraved jewels. Click on title to learn more.

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API Degenotoxification with Adenine-Functionalized Polymers

API Degenotoxification with Adenine-Functionalized Polymers | iBB | Scoop.it
Sulfonate genotoxic impurity (GTIs) pose a high risk for human health due to their ability to alkylate DNA. Removal of such GTIs from pharmaceutical formulations (<1.5 µg/day) is thus a regulatory requirement. In a recent study performed by Teresa Esteves and Frederico Ferreira from BERG-iBB, in collaboration with researchers from Faculty of Pharmacy (UL), propose using a polybenzimidazole (PBI) polymer modified with a DNA base to remove sulfonated GTIs. The polymer was able to efficiently remove the GTI from solutions prepared in dichloromethane, removing more than 96% of the impurity as compared to only about 10% for the original non-modified polymer. Click on title to learn more. 
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3D Differentiation of Human Oligodendrocyte Precursors from Pluripotent Stem Cells

3D Differentiation of Human Oligodendrocyte Precursors from Pluripotent Stem Cells | iBB | Scoop.it
Oligodendrocyte precursor cells (OPCs) offer considerable potential for the treatment of demyelinating diseases and injuries of the CNS. However, generating large quantities of high-quality OPCs remains a challenge that impedes their therapeutic application. In collaboration with researchers at the Stem Cell Engineering Research Group (SCERG), researchers at the Department of Bioengineering, University of California, Berkeley, have recently developed a novel defined and scalable culture system for differentiation of OPCs from Human Pluripotent Stem Cells, under 3D conditions, using a fully-defined thermoresponsive biomaterial. This system enabled the generation of large quantities of high-quality OPCs that were able to engraft, migrate and mature into myelinating oligodendrocytes upon transplantation into the brains of NOD/SCID mice.
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Genome-wide Search for Candidate Genes for Yeast Robustness Improvement Against Formic Acid

Genome-wide Search for Candidate Genes for Yeast Robustness Improvement Against Formic Acid | iBB | Scoop.it

Synthetic pathway engineering for robustness improvement of industrial strains envisaging their use in lignocellulosic biorefineries requires the understanding of the complex molecular mechanisms underlying Saccharomyces cerevisiae tolerance to inhibitors present in  hydrolysates. This study, recently published in Biotechnology for Biofuels, was led by Isabel Sá-Correia and part of Silvia F. Henriques PhD thesis in Biotechnology and Biosciences and also co-authored by Nuno P. Mira. The paper reports the identification, at a genome-wide scale, of genes whose expression confers protection or susceptibility to formic acid. This chemogenomic analysis allowed the identification of 172 determinants of tolerance and 41 determinants of susceptibility, among them HAA1, encoding the main transcriptional regulator of yeast transcriptome reprograming in response to acetic acid, and genes of the Haa1-regulon. TRK1, encoding the high-affinity potassium transporter and genes encoding positive regulators of Trk1 activity were surprisingly found among the genes that when deleted lead to increased tolerance to formic acid. It was hypothesized that Trk1 facilitates formic acid uptake into the yeast cell and demonstrated that at a saturating K+ concentrations, trk1Δ mutant is more tolerant than the parental strain. The list of genes resulting from this study provides potentially valuable information to guide improvement programs for the development of more robust strains against formic acid.

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LYTAG-driven Purification Strategies for Monoclonal Antibodies

LYTAG-driven Purification Strategies for Monoclonal Antibodies | iBB | Scoop.it

Monoclonal antibodies are becoming a leading class of biopharmaceuticals but to increase their accessibility to the general population, new production processes must be developed in particular for the downstream processing. In this publication, researchers from BERG-iBB (Ana Azevedo and Raquel Aires-Barros) in collaboration with the biotechnology companies Biomedal S.L. (Seville, Spain) and ChiPro GmbH (Bremen, Germany) propose an alternative and innovative affinity chromatographic purification method using quaternary amine anion-exchange matrices. Separation is driven by the dual affinity ligand LYTAG-Z, composed of a choline binding polypeptide (LYTAG) and the synthetic antibody binding Z domain. Click on title to learn more.

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Determining Virulence in Candida glabrata Through Adaptation to Host Stress

Determining Virulence in Candida glabrata Through Adaptation to Host Stress | iBB | Scoop.it

Persistence and virulence of Candida glabrata infections are multifactorial phenomena, whose understanding is crucial for infection erradication. In this study, the multidrug transporter CgDtr1 was found to be a plasma membrane acetic acid exporter, relieving the stress induced upon C. glabrata cells within hemocytes, and thus enabling increased proliferation and virulence against G. mellonella larvae. These results, emerging from a collaboration between BSRG members and led by Miguel Cacho Teixeira, were just published in Frontiers in Cellular and Infection Microbiology and are expected to contribute to design more suitable therapeutic strategies. Click on title to learn more.

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Biomimetic Cues Improve the Functionality of Hematopoietic Stem/Progenitor Cells

Biomimetic Cues Improve the Functionality of Hematopoietic Stem/Progenitor Cells | iBB | Scoop.it
The adult bone marrow (BM) niche is a complex entity where a homeostatic hematopoietic system is maintained through a dynamic crosstalk between different cellular and non-cellular players. Different signaling mechanisms are involved in controlling quiescence, self-renewal, differentiation and migration of hematopoietic stem/progenitor cells (HSPC). A promising strategy to efficiently expand HSPC numbers and tune their properties ex-vivo is to mimic the hematopoietic niche through integration of adjuvant stromal cells, soluble cues and/or biomaterial-based approaches in HSPC culture systems. In a recent recview published in Biotechnology Journal, SCERG-researchers led by Joaquim Cabral and Cláudia Lobato Silva focus on depicting the characteristics of co-culture systems, as well as other bioengineering approaches to improve the functionality of HSPC ex-vivo. Click on title to access the review.
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Enzymatic Synthesis of Alkyl Esters in Miniemulsions

Enzymatic Synthesis of Alkyl Esters in Miniemulsions | iBB | Scoop.it

Oil-in-water miniemulsions display potential for application as a “green” system in the enzymatic alkyl ester synthesis. BERG-iBB researchers led by Luís Fonseca combined miniemulsion systems with medium engineering to increase the yields and produtivity of the cutinase-catalysed synthesis of medium-chain alkyl esters. Enzyme stability and reutilization were tuned by operating in a fed-batch mode, which allowed long-term utilization of the free enzyme. Click on title to learn more.


Photo details: 3D structure of cutinase from F. Solani pisi with catalytic triad in blue (PDB code: 1AGY).

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Biocatalytic Process Intensification in Flow Bioreactors

Biocatalytic Process Intensification in Flow Bioreactors | iBB | Scoop.it

Biocatalysis has widened its scope and relevance with the availability of new molecular tools, e.g. improved expression systems, protein and metabolic engineering, and rational immobilization techniques. However, applications are still hampered by low productivity and scale-up difficulties. A practical step to improve biocatalyst (enzymes, whole-cells) performance is to use flow reactors. In a recent review published in Trends in Biotechnology, Pedro Fernandes from BERG-iBB together with colleagues from the Universities of Milan and Nottingham describe the state of the art on the design and use of biocatalysis in flow reactors. The successes of the technology, new opportunities, problems and advances are critically discussed. Click on title to learn more.

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Chromatography-on-a-Chip

Chromatography-on-a-Chip | iBB | Scoop.it

Optimization of chromatography is a lengthy and demanding task that requires large volumes of reagents and costly equipment. Ana Azevedo and colleagues from BERG-iBB, in collaboration with João Pedro Conde from INESC-MN, describe an integrated, reusable and portable microfluidic platform for the rapid screening of chromatographic conditions. The platform was tested for the capture of fluorophore-labeled monoclonal antibodies from cell culture supernatants using a multimodal ligand (Capto MMC). The work is part of Inês Pinto´s PhD thesis in Biotechnology and Biosciences. Click on title to leran more in the journal Sensors and Actuators B.

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Process Simulation and Economic Analysis of mAbs Purification

Process Simulation and Economic Analysis of mAbs Purification | iBB | Scoop.it

Monoclonal antibodies are expensive to produce in part due to their purification process, which is centered in protein-A affinity chromatography. A recent publication by Ana Azevedo and co-workers from BERG-iBB highlights a novel purification strategy for mAbs comprising phenylboronic acid multimodal chromatography for capture followed by polishing by ion exchange monolithic chromatography and packed bed hydrophobic interaction chromatography. Process simulation and economical analysis show that the new process is worth investing in and could present a viable alternative to the platform process used by most industrial players. Click on title to learn more.

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A Microfluidic Toolbox for the Optimization of Aqueous Two Phase Antibody Extraction

A Microfluidic Toolbox for the Optimization of Aqueous Two Phase Antibody Extraction | iBB | Scoop.it
The use of aqueous two-phase extraction (ATPE) is a viable option for the purification of antibodies and other vluable proteins, but these systems are difficult to model and optimize. In a publication in the Journal of Chmoratography A, researchers at BERG-iBB and INESC-MN describe a methodology for the rapid screening of antibody extraction conditions that relies on a microfluidic channel-based toolbox. A first microfluidic structure allows a simple negative-pressure driven rapid screening of up to 8 extraction conditions simultaneously, using less than 20 μL of each phase-forming solution per experiment, while a second microfluidic structure allows the integration of multi-step extraction protocols based on the results obtained with the first device. Click on title to learn more.
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Allogeneic Cell Therapy Manufacturing

Allogeneic Cell Therapy Manufacturing | iBB | Scoop.it
Commercial and large-scale production of allogeneic cell therapy products requires unique capabilities to develop technologies that generate safe and effective allogeneic cells/cell lines and their fully characterized master/working banks. In addition, it is necessary to design robust upstream and downstream manufacturing processes, and establish integrated, well-designed manufacturing facilities to produce high quality products in accordance with current GMP regulations. A recent publication by Joaquim Cabral from SCERG-iBB, in collaboration with researchers from the Royan Institute for Stem Cell Biology and Technology (Tehran), highlights the recent advances in the development of allogeneic products, the available options to develop robust manufacturing processes, and facility design considerations. Click on title to learn more.
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Sample Preparation With Aqueous Two-Phase Systems for On-site Immunodetection of Mycotoxins

Sample Preparation With Aqueous Two-Phase Systems for On-site Immunodetection of Mycotoxins | iBB | Scoop.it
Researchers at BERG-iBB and INESC-MN recently published a simple and versatile sample preparation methodology based on a green aqueous two-phase system for the simultaneous extraction and concentration of mycotoxins in multiple feeds, which are currently among the most relevant and widespread contaminants in the food and feed industry. This method was tailored for the subsequent rapid detection of ochratoxin A, aflatoxin B1 and deoxynivalenol at the relevant limits within the ng/g range, using a microfluidic multiplexed chemiluminescent immunoassay. Overall, sample preparation and detection were achieved in about 25 min using a “point-of-need” compatible process. Click on title to learn more.
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Structure of O-antigen and Hybrid Biosynthetic Locus in Burkholderia cenocepacia from a Cystic Fibrosis Patient

Structure of O-antigen and Hybrid Biosynthetic Locus in Burkholderia cenocepacia from a Cystic Fibrosis Patient | iBB | Scoop.it

Burkholderia cenocepacia is an opportunistic pathogen associated with chronic lung infections and increased risk of death in cystic fibrosis (CF) patients. This study, recently published in Frontiers in Microbiology, reports the chemical structure of the lipopolysaccharide (LPS) molecule of various sequential isolates and the identification of a novel hybrid O-antigen (OAg) biosynthetic cluster. The OAg repeating unit of the initial isolate LPS was not previously described in B. cenocepacia and none of the subsequent isolates express the OAg. This research, coordinated by Prof. Isabel Sá-Correia and with the BIOTECnico PhD student Amir Hassan and Dr. Sandra dos Santos as first co-authors, and also co-authored by Dr. Carla Coutinho from iBB BSRG and by our international collaborators in the UK, Italy and USA, provides support to the notion that OAg LPS modifications are important in B. cenocepacia adaptation to chronic infection of the CF lung.

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Fluorescence Correlation Spectroscopy of Protein:DNA Supramolecular Complexes

Fluorescence Correlation Spectroscopy of Protein:DNA Supramolecular Complexes | iBB | Scoop.it

Protein:DNA supramolecular complexes are at the heart of many molecular diagnostic systems. In a paper just published in Physical Chemistry Chemical Physics, Ana Rosa and Miguel Prazeres from BERG-iBB and Pedro Paulo from CQE present a fluorescence correlation spectroscopy (FCS) study of the complexation of oligonucleotides, antibodies and a chimeric protein of IgG-binding ZZ domains fused with a carbohydrate binding module. The FCS-derived information obtained (diffusion coefficients, association constants, nanometer-scale size of complexes) is useful to gain insight into molecular interactions involved in diagnostic assays. Click on title to learn more.

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