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• Microbial Carbohydrate Esterases Deacetylating Plant Polysaccharides.

  Authors: Peter Biely

  Biotechnology advances.

  Several plant polysaccharides are partially esterified with acetic acid. One of the roles of this modification is protection of plant cell walls against invading microorganisms. Acetylation ofSeveral plant polysaccharides are partially esterified with acetic acid. One of the roles of this modification is protection of plant cell walls against invading microorganisms. Acetylation of glycosyl residues of polysaccharides prevents hydrolysis of their glycosidic linkages by the corresponding glycoside hydrolases. In this way the acetylation also represents an obstacle of enzymatic saccharification of plant hemicelluloses to fermentable sugars which appears to be a hot topic of current research. We can eliminate this obstacle by alkaline extraction or pretreatment leading to saponification of ester linkages. However, this task has been accomplished in a different way in the nature. The acetyl groups became targets of microbial carbohydrate esterases that evolved to overcome the complexity of the plant cell walls and that cooperate with glycoside hydrolases in plant polysaccharide degradation. This article concentrates on enzymes deacetylating plant hemicelluloses excluding pectin. They are currently grouped in at least 8 families, specifically in CE families 1-7 and 16, originally assigned as Acetylxylan esterases, the enzymes acting on hardwood acetyl glucuronoxylan and its fragments generated by endo-β-1,4-xylanases. There are esterases deacetylating softwood galactoglucomannan, but they have not been classified yet. The enzymes present in CE families 1-7 differ in structure and substrate and positional specificity. There are families behaving as endo-type and exo-type deacetylates, i.e. esterases deacetylating internal sugar residues of partially acetylated polysaccharides and also esterases deacetylating non-reducing end sugar residues in oligosaccharides. With one exception, the enzymes of all mentioned CE families belong to serine type esterases. CE family 4 harbours enzymes that are metal-dependent aspartic esterases. Three-dimensional structures have been solved for members of the first seven CE families, however, there is still insufficient knowledge about their substrate specificity and real physiological role. Current knowledge on catalytic properties of the selected families of CEs is summarized in this review. Some of the families are emmerging also as new biocatalysts for regioselective acylation and deacylation of carbohydrates.

• Perspectives of plant-associated microbes in heavy metal phytoremediation.

  Authors: M Rajkumar, S Sandhya, M N V Prasad, H Freitas

  Biotechnology advances.

  "Phytoremediation" know-how to do-how is rapidly expanding and is being commercialized by harnessing the phyto-microbial diversity. This technology employs biodiversity to remove/contain pollutants"Phytoremediation" know-how to do-how is rapidly expanding and is being commercialized by harnessing the phyto-microbial diversity. This technology employs biodiversity to remove/contain pollutants from the air, soil and water. In recent years, there has been a considerable knowledge explosion in understanding plant-microbes-heavy metals interactions. Novel applications of plant-associated microbes have opened up promising areas of research in the field of phytoremediation technology. Various metabolites (e.g., 1-aminocyclopropane-1-carboxylic acid deaminase, indole-3-acetic acid, siderophores, organic acids, etc.) produced by plant-associated microbes (e.g., plant growth promoting bacteria, mycorrhizae) have been proposed to be involved in many biogeochemical processes operating in the rhizosphere. The salient functions include nutrient acquisition, cell elongation, metal detoxification and alleviation of biotic/abiotic stress in plants. Rhizosphere microbes accelerate metal mobility, or immobilization. Plants and associated microbes release inorganic and organic compounds possessing acidifying, chelating and/or reductive power. These functions are implicated to play an essential role in plant metal uptake. Overall the plant-associated beneficial microbes enhance the efficiency of phytoremediation process directly by altering the metal accumulation in plant tissues and indirectly by promoting the shoot and root biomass production. The present work aims to provide a comprehensive review of some of the promising processes mediated by plant-associated microbes and to illustrate how such processes influence heavy metal uptake through various biogeochemical processes including translocation, transformation, chelation, immobilization, solubilization, precipitation, volatilization and complexation of heavy metals ultimately facilitating phytoremediation.

• Valorization of biodiesel derived glycerol as a carbon source to obtain added-value metabolites: Focus on polyunsaturated fatty acids.

  Authors: Sergi Abad, Xavier Turon

  Biotechnology advances. 30(3):733-41.

  The amount of glycerol derived from the biodiesel industry is exponentially increasing. The valorization of glycerol has acquired attention and resources with an obvious economic and environmentalThe amount of glycerol derived from the biodiesel industry is exponentially increasing. The valorization of glycerol has acquired attention and resources with an obvious economic and environmental interest. Glycerol has the potential to improve the profitability of biodiesel in a biorefinery scenario. Added-value metabolites obtained from glycerol-based fermentations are the target of multiple research studies, primarily chemicals and biopolymers. Pigments and polyunsaturated fatty acids are exceptional examples as they have market presence as nutraceuticals. Most of the studies reviewed have been based on microalgae cultures. Depending on the strain and the engineering aspects of such cultures the final yield suffers notable variations. This is an emerging field which shows great potential from the perspective of a byproduct usage and the increasing yields (value) obtained from the bioprocess.

• Role of biomaterials, therapeutic molecules and cells for hepatic tissue engineering.

  Authors: Kirthanashri Srinivasan Vasanthan, Anuradha Subramanian, Uma Maheswari Krishnan, Swaminathan Sethuraman

  Biotechnology advances. 30(3):742-52.

  Current liver transplantation strategies face severe shortcomings owing to scarcity of donors, immunogenicity, prohibitive costs and poor survival rates. Due to the lengthy list of patients requiringCurrent liver transplantation strategies face severe shortcomings owing to scarcity of donors, immunogenicity, prohibitive costs and poor survival rates. Due to the lengthy list of patients requiring transplant, high mortality rates are observed during the endless waiting period. Tissue engineering could be an alternative strategy to regenerate the damaged liver and improve the survival and quality of life of the patient. The development of an ideal scaffold for liver tissue engineering depends on the nature of the scaffold, its architecture and the presence of growth factors and recognition motifs. Biomimetic scaffolds can simulate the native extracellular matrix for the culture of hepatocytes to enable them to exhibit their functionality both in vitro and in vivo. This review highlights the physiology and pathophysiology of liver, the current treatment strategies, use of various scaffolds, incorporation of adhesion motifs, growth factors and stem cells that can stabilize and maintain hepatocyte cultures for a long period.

• Extraction of oil from microalgae for biodiesel production: A review.

  Authors: Ronald Halim, Michael K Danquah, Paul A Webley

  Biotechnology advances. 30(3):709-32.

  The rapid increase of CO(2) concentration in the atmosphere combined with depleted supplies of fossil fuels has led to an increased commercial interest in renewable fuels. Due to their high biomassThe rapid increase of CO(2) concentration in the atmosphere combined with depleted supplies of fossil fuels has led to an increased commercial interest in renewable fuels. Due to their high biomass productivity, rapid lipid accumulation, and ability to survive in saline water, microalgae have been identified as promising feedstocks for industrial-scale production of carbon-neutral biodiesel. This study examines the principles involved in lipid extraction from microalgal cells, a crucial downstream processing step in the production of microalgal biodiesel. We analyze the different technological options currently available for laboratory-scale microalgal lipid extraction, with a primary focus on the prospect of organic solvent and supercritical fluid extraction. The study also provides an assessment of recent breakthroughs in this rapidly developing field and reports on the suitability of microalgal lipid compositions for biodiesel conversion.

• From short peptides to nanofibers to macromolecular assemblies in biomedicine.

  Authors: Yihua Loo, Shuguang Zhang, Charlotte A E Hauser

  Biotechnology advances. 30(3):593-603.

  In the last few years, a variety of self-assembling short peptides that consist exclusively of simple amino acids have been designed and modified. These peptides exhibit self-assembling dynamicIn the last few years, a variety of self-assembling short peptides that consist exclusively of simple amino acids have been designed and modified. These peptides exhibit self-assembling dynamic behaviors. At the molecular structural level, they form α-helical, β-sheet and β-hairpins structures in water. These structures further undergo spontaneous assembly to form nanofibers which aggregate into supramolecular scaffolds that entrap large volumes of water. Furthermore, nanostructures and supramolecular structures that self-organized from these short peptides also have a broad spectrum of biotechnological applications. They are useful as biological materials for 2D and 3D tissue cell cultures, regenerative and reparative medicine, tissue engineering as well as injectable drug delivery matrices that gel in situ. We have endeavored to do a comprehensive review of short peptides that form nanofibrous hydrogels. In particular, we have focused on recent advances in peptide assembly motifs and applications.

• Immobilized Candida antarctica lipase B: Hydration, stripping off and application in ring opening polyester synthesis.

  Authors: Ani Idris, Attaullah Bukhari

  Biotechnology advances. 30(3):550-63.

  This work reviews the stripping off, role of water molecules in activity, and flexibility of immobilized Candida antarctica lipase B (CALB). Employment of CALB in ring opening polyester synthesisThis work reviews the stripping off, role of water molecules in activity, and flexibility of immobilized Candida antarctica lipase B (CALB). Employment of CALB in ring opening polyester synthesis emphasizing on a polylactide is discussed in detail. Execution of enzymes in place of inorganic catalysts is the most green alternative for sustainable and environment friendly synthesis of products on an industrial scale. Robust immobilization and consequently performance of enzyme is the essential objective of enzyme application in industry. Water bound to the surface of an enzyme (contact class of water molecules) is inevitable for enzyme performance; it controls enzyme dynamics via flexibility changes and has intensive influence on enzyme activity. The value of pH during immobilization of CALB plays a critical role in fixing the active conformation of an enzyme. Comprehensive selection of support and protocol can develop a robust immobilized enzyme thus enhancing its performance. Organic solvents with a log P value higher than four are more suitable for enzymatic catalysis as these solvents tend to strip away very little of the enzyme surface bound water molecules. Alternatively ionic liquid can work as a more promising reaction media. Covalent immobilization is an exclusively reliable technique to circumvent the leaching of enzymes and to enhance stability. Activated polystyrene nanoparticles can prove to be a practical and economical support for chemical immobilization of CALB. In order to reduce the E-factor for the synthesis of biodegradable polymers; enzymatic ring opening polyester synthesis (eROPS) of cyclic monomers is a more sensible route for polyester synthesis. Synergies obtained from ionic liquids and immobilized enzyme can be much effective eROPS.

• Co-expression vs. co-infection using baculovirus expression vectors in insect cell culture: Benefits and drawbacks.

  Authors: Stanislav Sokolenko, Steve George, Andreas Wagner, Anup Tuladhar, Jonas M S Andrich, Marc G Aucoin

  Biotechnology advances. 30(3):766-81.

  The baculovirus expression vector system (BEVS) is a versatile and powerful platform for protein expression in insect cells. With the ability to approach similar post-translational modifications asThe baculovirus expression vector system (BEVS) is a versatile and powerful platform for protein expression in insect cells. With the ability to approach similar post-translational modifications as in mammalian cells, the BEVS offers a number of advantages including high levels of expression as well as an inherent safety during manufacture and of the final product. Many BEVS products include proteins and protein complexes that require expression from more than one gene. This review examines the expression strategies that have been used to this end and focuses on the distinguishing features between those that make use of single polycistronic baculovirus (co-expression) and those that use multiple monocistronic baculoviruses (co-infection). Three major areas in which researchers have been able to take advantage of co-expression/co-infection are addressed, including compound structure-function studies, insect cell functionality augmentation, and VLP production. The core of the review discusses the parameters of interest for co-infection and co-expression with time of infection (TOI) and multiplicity of infection (MOI) highlighted for the former and the choice of promoter for the latter. In addition, an overview of modeling approaches is presented, with a suggested trajectory for future exploration. The review concludes with an examination of the gaps that still remain in co-expression/co-infection knowledge and practice.

• Exploiting microRNAs for cell engineering and therapy.

  Authors: Tomaž Bratkovič, Gordana Glavan, Borut Strukelj, Marko Zivin, Boris Rogelj

  Biotechnology advances. 30(3):753-65.

  MicroRNAs (miRNAs) form a large class of non-coding RNAs that function in repression of gene expression in eukaryotes. By recognizing short stretches of nucleotides within the untranslated regions ofMicroRNAs (miRNAs) form a large class of non-coding RNAs that function in repression of gene expression in eukaryotes. By recognizing short stretches of nucleotides within the untranslated regions of mRNAs, miRNAs recruit partner proteins to individual transcripts, leading to mRNA cleavage or hindering of translation. Bioinformatic predictions and a wealth of data from wet laboratory studies indicate that miRNAs control expression of a large proportion of protein-coding genes, implying involvement of miRNAs in regulation of most biologic processes. In this review we discuss the biology of miRNAs and present examples of how manipulation of miRNA expression or activity can be exploited to attain the desired phenotypic traits in cell engineering as well as achieve therapeutic outcomes in treatment of a diverse set of diseases.

• Evaluating metabolic stress and plasmid stability in plasmid DNA production by Escherichia coli.

  Authors: Filomena Silva, João A Queiroz, Fernanda C Domingues

  Biotechnology advances. 30(3):691-708.

  In the context of recombinant DNA technology, the development of feasible and high-yielding plasmid DNA production processes has regained attention as more evidence for its efficacy as vectors forIn the context of recombinant DNA technology, the development of feasible and high-yielding plasmid DNA production processes has regained attention as more evidence for its efficacy as vectors for gene therapy and DNA vaccination arise. When producing plasmid DNA in Escherichia coli, a number of biological restraints, triggered by plasmid maintenance and replication as well as culture conditions are responsible for limiting final biomass and product yields. This termed "metabolic burden" can also cause detrimental effects on plasmid stability and quality, since the cell machinery is no longer capable of maintaining an active metabolism towards plasmid synthesis and the stress responses elicited by plasmid maintenance can also cause increased plasmid instability. The optimization of plasmid DNA production bioprocesses is still hindered by the lack of information on the host metabolic responses as well as information on plasmid instability. Therefore, systematic and on-line approaches are required not only to characterise this "metabolic burden" and plasmid stability but also for the design of appropriate metabolic engineering and culture strategies. The monitoring tools described to date rapidly evolve from laborious, off-line and at-line monitoring to online monitoring, at a time-scale that enables researchers to solve these bioprocessing problems as they occur. This review highlights major E. coli biological alterations caused by plasmid maintenance and replication, possible causes for plasmid instability and discusses the ability of currently employed bioprocess monitoring techniques to provide information in order to circumvent metabolic burden and plasmid instability, pointing out the possible evolution of these methods towards online bioprocess monitoring.

• Recent advances in synthesis and surface modification of lanthanide-doped upconversion nanoparticles for biomedical applications.

  Authors: Min Lin, Ying Zhao, Shuqi Wang, Ming Liu, Zhenfeng Duan, Yongmei Chen, Fei Li, Feng Xu, Tianjian Lu

  Biotechnology advances.

  Lanthanide (Ln)-doped upconversion nanoparticles (UCNPs) with appropriate surface modification can be used for a wide range of biomedical applications such as bio-detection, cancer therapy,Lanthanide (Ln)-doped upconversion nanoparticles (UCNPs) with appropriate surface modification can be used for a wide range of biomedical applications such as bio-detection, cancer therapy, bio-labeling, fluorescence imaging, magnetic resonance imaging and drug delivery. The upconversion phenomenon exhibited by Ln-doped UCNPs renders them tremendous advantages in biological applications over other types of fluorescent materials (e.g., organic dyes, fluorescent proteins, gold nanoparticles, quantum dots, and luminescent transition metal complexes) for: (i) enhanced tissue penetration depths achieved by near-infrared (NIR) excitation; (ii) improved stability against photobleaching, photoblinking and photochemical degradation; (iii) non-photodamaging to DNA/RNA due to lower excitation light energy; (iv) lower cytotoxicity; and (v) higher detection sensitivity. Ln-doped UCNPs are therefore attracting increasing attentions in recent years. In this review, we present recent advances in the synthesis of Ln-doped UCNPs and their surface modification, as well as their emerging applications in biomedicine. The future prospects of Ln-doped UCNPs for biomedical applications are also discussed.

• Symposium BIOTECH 2011 in Prague and its postsymposium in Olomouc, Czech Republic joined with the 5th Czech-Swiss Symposium.

  Authors: Jan Káš

  Biotechnology advances.

• The insulin-like androgenic gland hormone in crustaceans: From a single gene silencing to a wide array of sexual manipulation-based biotechnologies.

  Authors: Tomer Ventura, Amir Sagi

  Biotechnology advances.

  Due to the over-harvesting and deterioration of wild populations, the ever-growing crustacean market is increasingly reliant on aquaculture, driving the need for better management techniques. SinceDue to the over-harvesting and deterioration of wild populations, the ever-growing crustacean market is increasingly reliant on aquaculture, driving the need for better management techniques. Since most cultured crustacean species exhibit sexually dimorphic growth patterns, the culture of monosex populations (either all-male or all-female) is a preferred approach for gaining higher yields, with the ecological benefit of reducing the risk of invasion by the cultured species. Sexual manipulations may also render sustainable solutions to the environmental problems caused by the presence of invasive crustacean species with detrimental impacts ranging from aggressive competition with native species for food and shelter, to affecting aquaculture facilities and harvests and causing structural damage to river banks. Recent discoveries of androgenic gland (AG)-specific insulin-like peptides (IAGs) in crustaceans and the ability to manipulate them and their encoding transcripts (IAGs) have raised the possibility of sexually manipulating crustacean populations. Sexual manipulation is already a part of sustainable solutions in fish aquaculture and in the bio-control of insect pest species, and attempts are also being made to implement it with crustaceans. As recently exemplified in a commercially important prawn species, IAG silencing, a temporal, non-genetically modifying and non-transmissible intervention, has enabled the production of non-breeding all-male monosex populations that are the progeny of sexually reversed males ('neo-females'). IAG manipulations-based biotechnologies therefore have the potential to radically transform the entire industry. We review here how this proof of concept could be broadened to meet both aquacultural and environmental needs. We include the major cultured decapod crustacean groups and suggest a sustainable solution for the management of invasive and pest crustacean species. We also review the key considerations for devising a biotechnological approach that specifically tailors the molecular technological abilities to the management of each target group.

• Flow cytometry for the development of biotechnological processes with microalgae.

  Authors: P Hyka, S Lickova, P Přibyl, K Melzoch, K Kovar

  Biotechnology advances.

  The current interest in microalgae as a sustainable source of next generation biofuels and other valuable substances is driving exploration of their use as unique biotechnological production systems.The current interest in microalgae as a sustainable source of next generation biofuels and other valuable substances is driving exploration of their use as unique biotechnological production systems. To design and optimise appropriate production strategies, the behaviour of particular microalgal species should be well characterised under different culture conditions. Thus, flow cytometric (FCM) methods, which are already well established in environmental and toxicological studies of microalgae, are also useful for analysing the physiological state of microalgae, and have the potential to contribute to the rapid development of feasible bioprocesses. These methods are commonly based on the examination of intrinsic features of individual cells within a population (such as autofluorescence or size). Cells possessing the desired physiological or morphological features, which are detectable with or without fluorescent staining, are counted or isolated (sorted) using an FCM device. The options for implementation of FCM in the development of biotechnological processes detailed in this review are (i) analysing the chemical composition of biomass, (ii) monitoring cellular enzyme activity and cell viability, and (iii) sorting cells to isolate those overproducing the target compound or for the preparation of axenic cultures.

• Imaging cells and sub-cellular structures with ultrahigh resolution full-field X-ray microscopy.

  Authors: C C Chien, P Y Tseng, H H Chen, T E Hua, S T Chen, Y Y Chen, W H Leng, C H Wang, Y Hwu, G C Yin, K S Liang, F R Chen, Y S Chu, H I Yeh, Y C Yang, C S Yang, G L Zhang, J H Je, G Margaritondo

  Biotechnology advances.

  Our experimental results demonstrate that full-field hard-X-ray microscopy is finally able to investigate the internal structure of cells in tissues. This result was made possible by three mainOur experimental results demonstrate that full-field hard-X-ray microscopy is finally able to investigate the internal structure of cells in tissues. This result was made possible by three main factors: the use of a coherent (synchrotron) source of X-rays, the exploitation of contrast mechanisms based on the real part of the refractive index and the magnification provided by high-resolution Fresnel zone-plate objectives. We specifically obtained high-quality microradiographs of human and mouse cells with 29nm Rayleigh spatial resolution and verified that tomographic reconstruction could be implemented with a final resolution level suitable for subcellular features. We also demonstrated that a phase retrieval method based on a wave propagation algorithm could yield good subcellular images starting from a series of defocused microradiographs. The concluding discussion compares cellular and subcellular hard-X-ray microradiology with other techniques and evaluates its potential impact on biomedical research.

• Lipoxygenases: Potential starting biocatalysts for the synthesis of signaling compounds.

  Authors: Young-Chul Joo, Deok-Kun Oh

  Biotechnology advances.

  Lipoxygenases (LOXs) have attracted a great deal of attention as potential starting biocatalysts for synthesizing signaling compounds. Significant advances during the past decade include theLipoxygenases (LOXs) have attracted a great deal of attention as potential starting biocatalysts for synthesizing signaling compounds. Significant advances during the past decade include the discovery of regiospecific LOXs and structural investigation for their diverse regiospecificity. Seven regiospecific (5-, 8-, 9-, 10-, 12-, 13-, and 15-) LOXs catalyze positional-specific dioxygenation of polyunsaturated fatty acids, forming positional-specific hydroperoxy fatty acids that are further metabolized into signaling compounds. The LOX-derived signaling compounds can be applied not only for clinical uses but also for industrial uses. For example, animal lipoxin LXA(4), plant jasmonic acids, plant green leaf volatiles, and bacterial lactones have been used as anti-inflammatory agents, anti-pest agents, flavors, and food additives, respectively. Prostaglandins, as controllers of hormone regulation and cell growth, are also suggested as LOX-derived compounds in corals.

• Recent advances in microbial production of δ-aminolevulinic acid and vitamin B(12).

  Authors: Zhen Kang, Junli Zhang, Jingwen Zhou, Qingsheng Qi, Guocheng Du, Jian Chen

  Biotechnology advances.

  δ-aminolevulinate (ALA) is an important intermediate involved in tetrapyrrole synthesis (precursor for vitamin B(12), chlorophyll and heme) in vivo. It has been widely applied in agriculture andδ-aminolevulinate (ALA) is an important intermediate involved in tetrapyrrole synthesis (precursor for vitamin B(12), chlorophyll and heme) in vivo. It has been widely applied in agriculture and medicine. On account of many disadvantages of its chemical synthesis, microbial production of ALA has been received much attention as an alternative because of less expensive raw materials, low pollution, and high productivity. Vitamin B(12), one of ALA derivatives, which plays a vital role in prevention of anaemia has also attracted intensive works. In this review, recent advances on the production of ALA and vitamin B(12) with novel approaches such as whole-cell enzyme-transformation and metabolic engineering are described. Furthermore, the direction for future research and perspective are also summarized.

• Omega-3 biotechnology: Errors and omissions.

  Authors: Colin Ratledge

  Biotechnology advances.

• Advantages of intermediate X-ray energies in Zernike phase contrast X-ray microscopy.

  Authors: Zhili Wang, Kun Gao, Jian Chen, Youli Hong, Xin Ge, Dajiang Wang, Zhiyun Pan, Peiping Zhu, Wenbing Yun, Chris Jacobsen, Ziyu Wu

  Biotechnology advances.

  Understanding the hierarchical organizations of molecules and organelles within the interior of large eukaryotic cells is a challenge of fundamental interest in cell biology. Light microscopy is aUnderstanding the hierarchical organizations of molecules and organelles within the interior of large eukaryotic cells is a challenge of fundamental interest in cell biology. Light microscopy is a powerful tool for observations of the dynamics of live cells, its resolution attainable is limited and insufficient. While electron microscopy can produce images with astonishing resolution and clarity of ultra-thin (<1μm thick) sections of biological specimens, many questions involve the three-dimensional organization of a cell or the interconnectivity of cells. X-ray microscopy offers superior imaging resolution compared to light microscopy, and unique capability of nondestructive three-dimensional imaging of hydrated unstained biological cells, complementary to existing light and electron microscopy. Until now, X-ray microscopes operating in the "water window" energy range between carbon and oxygen k-shell absorption edges have produced outstanding 3D images of cryo-preserved cells. The relatively low X-ray energy (<540eV) of the water window imposes two important limitations: limited penetration (<10μm) not suitable for imaging larger cells or tissues, and small depth of focus (DoF) for high resolution 3D imaging (e.g., ~1μm DoF for 20nm resolution). An X-ray microscope operating at intermediate energy around 2.5keV using Zernike phase contrast can overcome the above limitations and reduces radiation dose to the specimen. Using a hydrated model cell with an average chemical composition reported in literature, we calculated the image contrast and the radiation dose for absorption and Zernike phase contrast, respectively. The results show that an X-ray microscope operating at ~2.5keV using Zernike phase contrast offers substantial advantages in terms of specimen size, radiation dose and depth-of-focus.

• Microbial transformation of antimalarial terpenoids.

  Authors: Igor A Parshikov, Alexander I Netrusov, John B Sutherland

  Biotechnology advances.

  The fungal and bacterial transformation of terpenoids derived from plant essential oils, especially the sesquiterpenoid artemisinin from Artemisia annua, has produced several new candidate drugs forThe fungal and bacterial transformation of terpenoids derived from plant essential oils, especially the sesquiterpenoid artemisinin from Artemisia annua, has produced several new candidate drugs for the treatment of malaria. Obtaining new derivatives of terpenoids, including artemisinin derivatives with increased antimalarial activity, is an important goal of research in microbial biotechnology and medicinal chemistry.

• FTIR spectro-imaging of collagens for characterization and grading of gliomas.

  Authors: Razia Noreen, Michel Moenner, Yeukuang Hwu, Cyril Petibois

  Biotechnology advances.

  Collagens are a family of at least 30 protein types organized as networks. They constitute the main support material of cells under the form of extracellular matrix as well as for membranes inCollagens are a family of at least 30 protein types organized as networks. They constitute the main support material of cells under the form of extracellular matrix as well as for membranes in vessels, organs, and tissue compartments. Collagen network abnormalities are at the origin of many diseases, including myopathies and fibroses. The characterization of collagens remains an analytical challenge due to the insolubility of these molecules and the difficulty encountered in isolating given types without altering their structure or in maintaining network organization, which is critical to diagnosing related pathologies. We have proposed using a vibrational spectroscopy based imaging technique, namely Fourier-transform infrared (FTIR) imaging, for a spatially-resolved analysis of secondary structure of different collagen types in complex samples, and more specifically for characterizing gliomas. With newly developed spectral data treatments and chemometrics using secondary structure parameters of collagen proteins, FTIR imaging is now able to distinguish between several types. On this basis, gliomas have been investigated as specific collagen-rich tissues developing in a non-collagenous environment, providing high specificity to this FTIR imaging utilization. Here, we review the recent advances in this imaging approach for understanding glioma development, with FTIR imaging now being proposed as a molecular histopathology tool for clinicians.

• Marine biotechnology advances towards applications in new functional foods.

  Authors: Ana C Freitas, Dina Rodrigues, Teresa A P Rocha-Santos, Ana M P Gomes, Armando C Duarte

  Biotechnology advances.

  The marine ecosystem is still an untapped reservoir of biologically active compounds, which have considerable potential to supply food ingredients towards development of new functional foods. WithThe marine ecosystem is still an untapped reservoir of biologically active compounds, which have considerable potential to supply food ingredients towards development of new functional foods. With the goal of increasing the availability and chemical diversity of functional marine ingredients, much research has been developed using biotechnological tools to discover and produce new compounds. This review summarizes the advances in biotechnological tools for production of functional ingredients, including enzymes, for the food industry. Tools involving biotechnological processes (bioreactors, fermentations, bioprocessing) and those involving genetic research designated as molecular biotechnology are discussed highlighting how they can be used in the controlled manipulation and utilization of marine organisms as sources of food ingredients, as well as discussing the most relevant shortcomings towards applications in new functional foods.

• Carbohydrate synthesis and biosynthesis technologies for cracking of the glycan code: Recent advances.

  Authors: Hynek Mrázek, Lenka Weignerová, Pavla Bojarová, Petr Novák, Ondřej Vaněk, Karel Bezouška

  Biotechnology advances.

  The glycan code of glycoproteins can be conceptually defined at molecular level by the sequence of well characterized glycans attached to evolutionarily predetermined amino acids along theThe glycan code of glycoproteins can be conceptually defined at molecular level by the sequence of well characterized glycans attached to evolutionarily predetermined amino acids along the polypeptide chain. Functional consequences of protein glycosylation are numerous, and include a hierarchy of properties from general physicochemical characteristics such as solubility, stability and protection of the polypeptide from the environment up to specific glycan interactions. Definition of the glycan code for glycoproteins has been so far hampered by the lack of chemically defined glycoprotein glycoforms that proved to be extremely difficult to purify from natural sources, and the total chemical synthesis of which has been hitherto possible only for very small molecular species. This review summarizes the recent progress in chemical and chemoenzymatic synthesis of complex glycans and their protein conjugates. Progress in our understanding of the ways in which a particular glycoprotein glycoform gives rise to a unique set of functional properties is now having far reaching implications for the biotechnology of important glycodrugs such as therapeutical monoclonal antibodies, glycoprotein hormones, carbohydrate conjugates used for vaccination and other practically important protein-carbohydrate conjugates.

• Tissue engineering interventions for esophageal disorders - Promises and challenges.

  Authors: Purushothaman Kuppan, Swaminathan Sethuraman, Uma Maheswari Krishnan

  Biotechnology advances.

  The diseases of the esophagus include congenital defects like atresia, tracheoesophageal fistula as well as others such as gastro-esophageal reflux disease (GERD), Barrett's esophagus, carcinoma andThe diseases of the esophagus include congenital defects like atresia, tracheoesophageal fistula as well as others such as gastro-esophageal reflux disease (GERD), Barrett's esophagus, carcinoma and strictures. All esophageal disorders require surgical intervention and reconstruction with appropriate substitutes. Primary anastomosis is used to treat most cases but treatment of long gap atresia still remains a clinical challenge. Autologous graft therapies using tissues from colon, and small and large intestine or gastric transplantations have been attempted but have constraints like leakage, infection and stenosis at the implanted site, which leads to severe morbidity and mortality. An alternative for autologous grafts are allogenic and xenogenic grafts, which have better availability but disease transmission and immunogenicity limit their applications. Use of biodegradable and biocompatible scaffolds to engineer the esophagus promises to be an effective regenerative strategy for treatment of esophageal disorders. Nanotopography of the fibrous scaffolds mimics the natural extracellular matrix (ECM) of the tissue and incorporation of chemical cues and tailoring mechanical properties provide the right microenvironment for co-culture of different cell types. Scaffolds cultured with esophageal cells (epithelial cells, fibroblast and smooth muscle cells) might show enhancement of the biofunctionality in vivo. This review attempts to address the various strategies and challenges involved in successful tissue engineering of the esophagus.

• Research advances in plant-made flavivirus antigens.

  Authors: C A Martínez, A M Giulietti, J Rodríguez Talou

  Biotechnology advances.

  Outbreaks of flaviviruses such as dengue (DV), yellow fever (YFV), Japanese encephalitis (JEV), tick-borne encephalitis (TBEV) and West Nile (WNV) affect numerous countries around the world. The fastOutbreaks of flaviviruses such as dengue (DV), yellow fever (YFV), Japanese encephalitis (JEV), tick-borne encephalitis (TBEV) and West Nile (WNV) affect numerous countries around the world. The fast spread of these viruses is the result of increases in the human population, rapid urbanisation and globalisation. While vector control is an important preventive measure against vector-borne diseases, it has failed to prevent the spread of these diseases, particularly in developing countries where the implementation of control measures is intermittent. As antiviral drugs against flaviviruses are not yet available, vaccination remains the most important tool for prevention. Although human vaccines for YFV, TBEV and JEV are available, on-going vaccination efforts are insufficient to prevent infection. No vaccines against DENV and WNV are available. Research advances have provided important tools for flavivirus vaccine development, such as the use of plants as a recombinant antigen production platform. This review summarises the research efforts in this area and highlights why a plant system is considered a necessary alternative production platform for high-tech subunit vaccines.

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