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In this work we model the glomerular filtration barrier, the structure responsible for filtering the blood and preventing the loss of proteins, using human podocytes and glomerular endothelial cells seeded into microfluidic chips. In long-term cultures, cells maintain their morphology, form capillary-like structures and express slit diaphragm proteins. This system recapitulates functions and structure of the glomerulus, including permselectivity. When exposed to sera from patients with anti-podocyte autoantibodies, the chips show albuminuria proportional to patients’ proteinuria, phenomenon not observed with sera from healthy controls or individuals with primary podocyte defects. We also show its applicability for renal disease modeling and drug testing. A total of 2000 independent chips were analyzed, supporting high reproducibility and validation of the system for high-throughput screening of therapeutic compounds. The study of the patho-physiology of the glomerulus and identification of therapeutic targets are also feasible using this chip. The glomerular filtration barrier is a complex structure in charge of renal ultrafiltration. Here the authors present a glomerulus-on-a-chip for disease modelling and high-throughput drug screening where human podocytes and human glomerular endothelial cells are separated by an extracellular matrix resembling the in vivo basement membrane.

Immobilization has been reported as an efficient technique to address the bacterial vulnerability for application in bio self-healing concrete. In this study, for the first time, magnetic iron oxide nanoparticles (IONs) are being practically employed as the protective vehicle for bacteria to evaluate the self-healing performance in concrete environment. Magnetic IONs were successfully synthesized and characterized using different techniques. The scanning electron microscope (SEM) images show the efficient adsorption of nanoparticles to the Bacillus cells. Microscopic observation illustrates that the incorporation of the immobilized bacteria in the concrete matrix resulted in a significant crack healing behavior, while the control specimen had no healing characteristics. Analysis of bio-precipitates revealed that the induced minerals in the cracks were calcium carbonate. The effect of magnetic immobilized cells on the concrete water absorption showed that the concrete specimens supplemented with decorated bacteria with IONs had a higher resistance to water penetration. The initial and secondary water absorption rates in bio-concrete specimens were 26% and 22% lower than the control specimens. Due to the compatible behavior of IONs with the concrete compositions, the results of this study proved the potential application of IONs for developing a new generation of bio self-healing concrete.

When the freshwater microalga Chlorella sorokiniana and the plant growth-promoting bacterium Azospirillum brasilense were deployed as free suspensions in unsterile, municipal wastewater for tertiary wastewater treatment, their population was significantly lower compared with their populations in sterile wastewater. At the same time, the numbers of natural microfauna and wastewater bacteria increased. Immobilization of C. sorokiniana and A. brasilense in small (2-4 mm in diameter), polymer Ca-alginate beads significantly enhanced their populations when these beads were suspended in normal wastewater. All microbial populations within and on the surface of the beads were evaluated by quantitative fluorescence in situ hybridization combined with scanning electron microscopy and direct measurements. Submerging immobilizing beads in wastewater created the following sequence of events: (a) a biofilm composed of wastewater bacteria and A. brasilense was created on the surface of the beads, (b) the bead inhibited penetration of outside organisms into the beads, (c) the bead inhibited liberation of the immobilized microorganisms into the wastewater, and (d) permitted an uninterrupted reduction of ammonium and phosphorus from the wastewater. This study demonstrated that wastewater microbial populations are responsible for decreasing populations of biological agents used for wastewater treatment and immobilization in alginate beads provided a protective environment for these agents to carry out uninterrupted tertiary wastewater treatment. [PUBLICATION ABSTRACT]

Polymerases that synthesize artificial genetic polymers hold great promise for advancing future applications in synthetic biology. However, engineering natural polymerases to replicate unnatural genetic polymers is a challenging problem. Here we present droplet-based optical polymerase sorting (DrOPS) as a general strategy for expanding polymerase function that employs an optical sensor to monitor polymerase activity inside the microenvironment of a uniform synthetic compartment generated by microfluidics. We validated this approach by performing a complete cycle of encapsulation, sorting and recovery on a doped library and observed an enrichment of ∼1,200-fold for a model engineered polymerase. We then applied our method to evolve a manganese-independent α- L -threofuranosyl nucleic acid (TNA) polymerase that functions with >99% template-copying fidelity. Based on our findings, we suggest that DrOPS is a versatile tool that could be used to evolve any polymerase function, where optical detection can be achieved by Watson–Crick base pairing. Droplet-based optical polymerase sorting employs a fluorescent sensor to monitor polymerase activity inside the microenvironment of uniform water-in-oil emulsions. Here, the authors use this technique to select and isolate single cells for evolution of an unnatural nucleic acid polymerase.

We developed a bioelectronic communication system that is enabled by a redox signal transduction modality to exchange information between a living cell-embedded bioelectronics interface and an engineered microbial network. A naturally communicating three-member microbial network is ‘plugged into’ an external electronic system that interrogates and controls biological function in real time. First, electrode-generated redox molecules are programmed to activate gene expression in an engineered population of electrode-attached bacterial cells, effectively creating a living transducer electrode. These cells interpret and translate electronic signals and then transmit this information biologically by producing quorum sensing molecules that are, in turn, interpreted by a planktonic coculture. The propagated molecular communication drives expression and secretion of a therapeutic peptide from one strain and simultaneously enables direct electronic feedback from the second strain, thus enabling real-time electronic verification of biological signal propagation. Overall, we show how this multifunctional bioelectronic platform, termed a BioLAN, reliably facilitates on-demand bioelectronic communication and concurrently performs programmed tasks. Living electrodes enable signalling into a microbial community, coordinating behaviour.

Photosynthetic unicellular organisms, known as microalgae, are key contributors to carbon fixation on Earth. Their biotic interactions with other microbes shape aquatic microbial communities and influence the global photosynthetic capacity. So far, limited information is available on molecular factors that govern these interactions. We show that the bacterium Pseudomonas protegens strongly inhibits the growth and alters the morphology of the biflagellated green alga Chlamydomonas reinhardtii . This antagonistic effect is decreased in a bacterial mutant lacking orfamides, demonstrating that these secreted cyclic lipopeptides play an important role in the algal–bacterial interaction. Using an aequorin Ca 2+ -reporter assay, we show that orfamide A triggers an increase in cytosolic Ca 2+ in C . reinhardtii and causes deflagellation of algal cells. These effects of orfamide A, which are specific to the algal class of Chlorophyceae and appear to target a Ca 2+ channel in the plasma membrane, represent a novel biological activity for cyclic lipopeptides. Predatory or competitive interactions between microbes are poorly understood but likely influence global nutrient cycles. Here, the authors show that Pseudomonas bacteria could immobilize algal cells, potential prey, by releasing secondary metabolites that induce a Ca 2+ signal and algal deflagellation.

Pharmaceuticals and their metabolites are released into the environment by domestic, hospital, and pharmaceutical industry wastewaters. Conventional wastewater treatment technology does not guarantee effluents of high quality, and apparently clean water may be loaded with pollutants. In this study, we assess the performance and efficiency of free and immobilised cells of microalgae Nannochloropsis sp. in removing four pharmaceuticals, chosen for their occurrence or persistence in the environment. These are paracetamol, ibuprofen, olanzapine and simvastatin. The results showed that free microalgae cells remain alive for a longer time than the immobilised ones, suggesting the inhibition of cell proliferation by the polymeric matrix polyvinyl alcohol. Both cells, free and immobilised, respond differently to each pharmaceutical. The removal of paracetamol and ibuprofen by Nannochloropsis sp., after 24 h of culture, was significantly higher in immobilised cells. Free cells removed a significantly higher concentration of olanzapine than immobilised ones, suggesting a higher affinity to this molecule than to paracetamol and ibuprofen. The results demonstrate the effectiveness of Nannochloropsis sp. free cells for removing olanzapine and Nannochloropsis sp. immobilised cells for removing paracetamol and ibuprofen.

UV and gamma irradiation mutagenesis was applied on Aspergillus fumigatus and Alternaria tenuissima in order to improve their producing ability of paclitaxel. Among the screened mutants, two stable strains (designated TXD105–GM6 and TER995–GM3) showed the maximum paclitaxel production. Paclitaxel titers of the two respective mutants were dramatically intensified to 1.22- and 1.24-fold, as compared by their respective parents. Immobilization using five different entrapment carriers of calcium alginate, agar-agar, Na-CMC, gelatin, and Arabic gum was successfully applied for production enhancement of paclitaxel by the two mutants. The immobilized cultures were superior to free-cell cultures and paclitaxel production by the immobilized mycelia was much higher than that of the immobilized spores using all the tried carriers. Moreover, calcium alginate gel beads were found the most conductive and proper entrapment carrier for maximum production of paclitaxel. The feasibility of the paclitaxel production by the immobilized mycelia as affected by incubation period, medium volume, and number of beads per flask was adopted. Under the favorable immobilization conditions, the paclitaxel titers were significantly intensified to 1.31- and 1.88-fold by the respective mutants, as compared by their free cultures. The obtained paclitaxel titers by the immobilized mycelia of the respective mutants (694.67 and 388.65 μg L −1 ) were found promising in terms of fungal production of paclitaxel. Hence, these findings indicate the future possibility to reduce the cost of producing paclitaxel and suggest application of the immobilization technique for the biotechnological production of paclitaxel at an industrial scale.

Ethanol production from sweet sorghum juice (SSJ) using the thermotolerant Saccharomyces cerevisiae strain DBKKUY-53 immobilized in an alginate-loofah matrix (ALM) was successfully developed. As found in this study, an ALM with dimensions of 20 × 20 × 5 mm3 is effective for cell immobilization due to its compact structure and long-term stability. The ALM-immobilized cell system exhibited greater ethanol production efficiency than the freely suspended cell system. By using a central composite design (CCD), the optimum conditions for ethanol production from SSJ by ALM-immobilized cells were determined. The maximum ethanol concentration and volumetric ethanol productivity obtained using ALM-immobilized cells under the optimal conditions were 97.54 g/L and 1.36 g/L h, respectively. The use of the ALM-immobilized cells was successful for at least six consecutive batches (360 h) without any loss of ethanol production efficiency, suggesting their potential application in industrial ethanol production.

Enzyme-mediated synthesis of pharmaceutical compounds is a ‘green’ alternative to traditional synthetic chemistry, and microbial engineering opens up the possibility of using whole cells as mini-factories. Whole-cell biocatalysis reduces cost by eliminating expensive enzyme purification and cofactor addition steps, as well as resulting in increased enzyme stability. Haloferax volcanii is a model halophilic archaeon encoding highly salt and organic solvent tolerant enzymes such as alcohol dehydrogenase ( Hv ADH2), which catalyses the reduction of aldehydes and ketone in the presence of NADPH/NADH cofactor. A H. volcanii strain for constitutive Hv ADH2 expression was generated using a strong synthetic promoter (p. syn ). The strain was immobilised in calcium alginate beads and repeatedly used as a whole-cell biocatalyst. The reduction of acetophenone, used as test substrate, was very successful and high yields were detected from immobilised whole cells over repeated biotransformation cycles. The immobilised H. volcanii retained stability and high product yields after 1 month of storage at room temperature. This newly developed system offers halophilic enzyme expression in its native environment, high product yield, stability and reusability without the addition of any expensive NADPH/NADH cofactor. This is the first report of whole cell–mediated biocatalysis by the halophilic archaeon H. volcanii .