Urban Planning and Corona Spaces – Scales, Walls and COVID-19 Coincidences

This study focuses on the role and responsibility of urban planning in mitigating the COVID-19 pandemic ́s impact. The far-reaching social and economic consequences of this threat are counteracted by organisational and constructional measures to prevent lockdowns and finally illnesses and deaths. Corona spaces and voids are introduced as a consistent multi scale approach concerning the pandemics spatial implications and respective measures. These terms are operationalised for urban planning and can be used as an overarching concept to be communicated within cross-sectoral planning tasks. A comparison of Taiwan's and the USA's responses to the outbreak suggests that the coincidence at the beginning of a pandemic can be controlled by institutional precautions. On an urban scale, organisational measures (e.g. contact tracing, quarantine, and lockdown) can be supported by constructional ones concerning e.g. transport, public spaces, urban agriculture, and offices aimed at crowding reduction. If appropriate measures are applied, urban density does not seem to increase spreading the virus, whereby a regression analysis based on data of districts in Germany shows no relationship between population density and COVID-19 deaths. Lockdown prevention should be a planning goal and multifunctional approaches that integrate aspects of virus resilience should be favoured over the monothematic urban development approach (Leitbild) of a virus resilient city. Urban planning can contribute to the mitigation of future outbreaks by including pandemic preparedness in planning frameworks

Miniature bioreactors: current practices and future opportunities

This review focuses on the emerging field of miniature bioreactors (MBRs), and examines the way in which they are used to speed up many areas of bioprocessing. MBRs aim to achieve this acceleration as a result of their inherent high-throughput capability, which results from their ability to perform many cell cultivations in parallel. There are several applications for MBRs, ranging from media development and strain improvement to process optimisation. The potential of MBRs for use in these applications will be explained in detail in this review. MBRs are currently based on several existing bioreactor platforms such as shaken devices, stirred-tank reactors and bubble columns. This review will present the advantages and disadvantages of each design together with an appraisal of prototype and commercialised devices developed for parallel operation. Finally we will discuss how MBRs can be used in conjunction with automated robotic systems and other miniature process units to deliver a fully-integrated, high-throughput (HT) solution for cell cultivation process development

Simplified immobilisation method for histidine-tagged enzymes in poly(methyl methacrylate) microfluidic devices

Article in press. Kulsharova, G., New BIOTECHNOLOGY (2017), https://doi.org/10.1016/j.nbt.2017.12.004Poly(methyl methacrylate) (PMMA) microfluidic devices have become promising platforms for a wide range of applications. Here we report a simple method for immobilising histidine-tagged enzymes suitable for PMMA microfluidic devices. The 1-step-immobilisation described is based on the affinity of the His-tag/Ni-NTA interaction and does not require prior amination of the PMMA surface, unlike many existing protocols. We compared it with a 3-step immobilisation protocol involving amination of PMMA and linking NTA via a glutaraldehyde cross-linker. These methods were applied to immobilise transketolase (TK) in PMMA microfluidic devices. Binding efficiency studies showed that about 15% of the supplied TK was bound using the 1-step method and about 26% of the enzyme was bound by the 3-step method. However, the TK-catalysed reaction producing l-erythrulose performed in microfluidic devices showed that specific activity of TK in the device utilising the 1-step immobilisation method was approximately 30% higher than that of its counterpart. Reusability of the microfluidic device produced via the 1-step method was tested for three cycles of enzymatic reaction and at least 85% of the initial productivity was maintained. The device could be operated for up to 40 h in a continuous flow and on average 70% of the initial productivity was maintained. The simplified immobilisation method required fewer chemicals and less time for preparation of the immobilised microfluidic device compared to the 3-step method while achieving higher specific enzyme activity. The method represents a promising approach for the development of immobilised enzymatic microfluidic devices and could potentially be applied to combine protein purification with immobilisation.Peer reviewe

Klebsiella pneumoniae as cell factory for chemicals production

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Characterization and multi-step transketolase-ω- transaminase bioconversions in an immobilized enzyme microreactor (IEMR) with packed tube

The concept of de novo metabolic engineering through novel synthetic pathways offers new directions for multi-step enzymatic synthesis of complex molecules. This has been complemented by recent progress in performing enzymatic reactions using immobilized enzyme microreactors (IEMR). This work is concerned with the construction of de novo designed enzyme pathways in a microreactor synthesizing chiral molecules. An interesting compound, commonly used as the building block in several pharmaceutical syntheses, is a single diastereoisomer of 2-amino-1,3,4-butanetriol (ABT). This chiral amino alcohol can be synthesized from simple achiral substrates using two enzymes, transketolase (TK) and transaminase (TAm). Here we describe the development of an IEMR using His6-tagged TK and TAm immobilized onto Ni-NTA agarose beads and packed into tubes to enable multi-step enzyme reactions. The kinetic parameters of both enzymes were first determined using single IEMRs evaluated by a kinetic model developed for packed bed reactors. The Km(app) for both enzymes appeared to be flow rate dependent, while the turnover number kcat was reduced 3 fold compared to solution-phase TK and TAm reactions. For the multi-step enzyme reaction, single IEMRs were cascaded in series, whereby the first enzyme, TK, catalyzed a model reaction of lithium-hydroxypyruvate (HPA) and glycolaldehyde (GA) to l-erythrulose (ERY), and the second unit of the IEMR with immobilized TAm converted ERY into ABT using (S)--methylbenzylamine (MBA) as amine donor. With initial 60 mM (HPA and GA each) and 6 mM (MBA) substrate concentration mixture, the coupled reaction reached approximately 83% conversion in 20 min at the lowest flow rate. The ability to synthesize a chiral pharmaceutical intermediate, ABT in relatively short time proves this IEMR system as a powerful tool for construction and evaluation of de novo pathways as well as for determination of enzyme kinetics

Methodology to rapidly assess enzyme cascades in aid of metabolic engineering of host cells

Chiral amino alcohols are compounds of pharmaceutical interest as they are building blocks of sphingolipids, antibiotics, and antiviral glycosidase inhibitors. Due to the challenges of chemical synthesis we recently developed two TK-TAm reaction cascades using natural and low cost feedstocks as substrates: a recycling cascade comprising of 2 enzymes and a sequential 3-step enzyme cascade yielding 30% and 1% conversion, respectively. In order to improve the conversion yield and aid the host strain engineering we used a combination of microscale experiments and statistical experimental design. For this we implemented a full factorial design to optimise pH, temperature and buffer type, followed by the implementation of Response Surface Methodology (RSM) for the optimisation of substrates and enzymes concentrations. We achieved 60% conversion for the recycling cascade and 3-fold improvement on the sequential pathway. Based on the results, limiting steps and individual requirements for host cell metabolic integration were identified expanding the understanding of the cascades without implementing extensive optimisation modelling. Therefore, the approach described here is ideal for exploratory work or when the interest is in defining the enzymatic expression levels required for microbial cell factories development

Single-use bioreactor technologies for early stage development of microalgae cultivation processes

This work describes the engineering characterization and evaluation of two novel, single-use photobioreactor technologies for microalgal cultivation. There is currently considerable interest in microalgae as alternative expression systems for the production of value-added chemicals as well as therapeutic proteins and vaccines. In contrast to mammalian expression systems, however, bioreactor platforms to address early stage development challenges are not well established particularly for phototrophic and mixotrophic cultivation strategies. To support early stage cell line selection and process characterization a single-use, 24-well micro photo-bioreactor (mPBr) was established together with an illuminated and environmentally controlled shaker platform [1]. The same orbital shaker platform was also used for microalgae cultivation up to 10L scale, in single-use bags usually used for mammalian cell cultivation on rocked platforms [2]. Particularly for small and micro-companies this single-use photobioreactor (SUPBr) provides a route for rapid process optimization and materialization of novel products from microalgae. Both bioreactor technologies were characterized in terms of their fluid hydrodynamics, mixing and gas-liquid mass transfer. The influence of these factors as well as light path length and light intensity on growth and pigment formation in Chlorella sorokiniana was also studied. Successful scale translation between the mPBr and the SUPBr was demonstrated illustrating the complementarity of the two approaches to help reduce microalgal bioprocess development timelines

Integrating Whole Cell Biotransformation of Aroma Compounds into a Novel Biorefinery Concept

The synthesis of aroma compounds that are utilized as precursors of multiple synthesis chains in the pharmaceutical industries and as ingredients in food and fragrance industries can be carried out using chemical processes, enzyme biocatalysis and whole cell biotransformation. Whole cell biotransformation has the potential of being more environmentally benign than chemical synthesis and more cost-effective as compared to enzyme catalysis. In a recently published study by the authors, the aroma compound Ethyl(3)hydroxybutyrate was produced by whole cell biotransformation under aerobic and anaerobic conditions. The yield of the anaerobic processes was similar to that of the aerobic processes, but additionally generated CO2 and ethanol as useful by-products. In this chapter we illustrate how the production process of Ethyl(3)hydroxybutyrate by whole cell biotransformation can be integrated into a novel biorefinery concept, based on the finding that the production of Ethyl(3)hydroxybutyrate under anaerobic conditions is efficient and environmentally friendly. CO2 may be converted to bio-methane together with H2 produced from excess regenerative power. A life cycle assessment confirmed that the anaerobic whole cell biotransformation process embedded into a biorefinery concept including bio-methane production has a lower environmental impact as compared to a concept based on the aerobic whole cell biotransformation

The aquaponic principle—It is all about coupling

The aquaponic principle is the coupling of animal aquaculture (e.g. fish) with plant production (e.g. vegetables) for saving resources. At present, various definitions of aquaponics exist, some bearing the risk of misinterpretation by dismissing the original meaning or being contradictory. In addition, there is no standard terminology for the aspects of coupling between the aquaponic subsystems. In this study, we addressed both issues. (1) We developed new or revised definitions that are summarised by: Aquaponic farming comprises aquaponics (which couples tank‐based animal aquaculture with hydroponics) and trans‐aquaponics, which extends aquaponics to tankless aquaculture as well as non‐hydroponics plant cultivation methods. Within our conceptual system, the term aquaponics corresponds to the definitions of FAO and EU. (2) A system analysis approach was utilised to explore different aquaponic setups aiming to better describe the way aquaponic subsystems are connected. We introduced the new terms ‘coupling type’ and ‘coupling degree’, where the former qualitatively characterises the water‐mediated connections of aquaponic subsystems. A system with on‐demand nutrient water supply for the independent operating plant cultivation is an ‘on‐demand coupled system’ and we propose to deprecate the counterintuitive term ‘decoupled system’ for this coupling type. The coupling degree comprises a set of parameters to quantitatively determine the coupling's efficiency of internal streams, for example, water and nutrients. This new framework forms a basis for improved communication, provides a uniform metric for comparing aquaponic facilities, and offers criteria for facility optimisation. In future system descriptions, it will simplify evaluation of the coupling's contribution to sustainability of aquaponics.Belmont ForumEuropean Commission via the CITYFOOD projectPeer Reviewe

The Analysis of Multiple Genome Comparisons in Genus Escherichia and Its Application to the Discovery of Uncharacterised Metabolic Genes in Uropathogenic Escherichia coli CFT073

A survey of a complete gene synteny comparison has been carried out between twenty fully sequenced strains from the genus Escherichia with the aim of finding yet uncharacterised genes implicated in the metabolism of uropathogenic strains of E. coli (UPEC). Several sets of adjacent colinear genes have been identified which are present in all four UPEC included in this study (CFT073, F11, UTI89, and 536), annotated with putative metabolic functions, but are not found in any other strains considered. An operon closely homologous to that encoding the L-sorbose degradation pathway in Klebsiella pneumoniae has been identified in E. coli CFT073; this operon is present in all of the UPEC considered, but only in 7 of the other 16 strains. The operon's function has been confirmed by cloning the genes into E. coli DH5α and testing for growth on L-sorbose. The functional genomic approach combining in silico and in vitro work presented here can be used as a basis for the discovery of other uncharacterised genes contributing to bacterial survival in specific environments