Symposium SB03—Smart Materials, Devices and Systems for Interface with Plants and Microorganisms

Genetic engineering of plants is at the core of sustainability efforts, natural product synthesis, and agricultural crop engineering. The plant cell wall is a barrier that limits the ease and throughput with which exogenous biomolecules can be delivered to plants. Current delivery methods either suffer from host range limitations, low transformation efficiencies, tissue regenerability, tissue damage, or unavoidable DNA integration into the host genome. Here, we demonstrate efficient diffusion-based biomolecule delivery into tissues and organs of intact plants of several species with a suite of pristine and chemically-functionalized high aspect ratio nanomaterials. Efficient DNA delivery and strong protein expression without transgene integration is accomplished in mature Nicotiana benthamiana, Eruca sativa (arugula), Triticum aestivum (wheat) and Gossypium hirsutum (cotton) leaves and arugula protoplasts [1]. We also demonstrate a second nanoparticle-based strategy in which small interfering RNA (siRNA) is delivered to mature Nicotiana benthamiana leaves and effectively silence a gene with 95% efficiency. We find that nanomaterials both facilitate biomolecule transport into plant cells, while also protecting polynucleotides such as RNA from nuclease degradation. DNA origami nanostructures further enable siRNA delivery to plants at programmable nanostructure loci [2]. Our work provides a tool for species-independent, targeted, and passive delivery of genetic material, without transgene integration, into plant cells for diverse plant biotechnology applications.


Demirer, G.S., Zhang, H., Matos, J., Goh, N., Cunningham, F.J., Sung, Y., Chang, R., Aditham, A.J., , Chio, L., Cho, M.J., Staskawicz, B., Landry, M.P. High Aspect Ratio Nanomaterials Enable Delivery of Functional Genetic Material Without DNA Integration in Mature Plants. Nature Nanotechnology (2019). DOI: 10.1038/s41565-019-0382-5NNANO-18081684

Zhang, H.*, Demirer, G.S.*, Zhang, H., Ye, T., Goh, N.S., Aditham, A.J., Cunningham, F.J., Fan, C., Landry, M.P. Low-dimensional DNA Nanostructures Coordinate Gene Silencing in Mature Plants. PNAS (2019). DOI: 10.1073/pnas.1818290116

As a consequence of their sessile nature, plants have developed complex signaling mechanisms to cope with the highly dynamic environment that they live in. In response to external stimuli, plants can utilize various signaling pathways for intercellular communication to engage different metabolic and genetic machineries involved in the plant defense system. Among the many signaling molecules recently uncovered, hydrogen peroxide (H₂O₂) represents one of the most versatile signaling molecule in plants in response to environmental stresses such as mechanical wounding. Here, we develop an optical nanosensor platform using DNA-wrapped single-walled carbon nanotubes (SWNT) to monitor the wound-induced H₂O₂ signaling pathway in plants. This approach enables non-destructive, real-time detection of changes in endogenous H₂O₂ level in plants at a remote distance with portable and inexpensive electronics. We demonstrate the versatility of this method to investigate the dynamics between H₂O₂ and electric signalling pathways in different vegetable crops and plant species. A mathematical model describing the self-propagating H₂O₂ wave mechanism will be presented. Our work provides a promising tool to elucidate the complex plant signaling pathways across species to aid botany research and agronomic applications.

In latest years the use of smart materials and devices started to attract attention for modulating the growth and functions of plant systems. Very recently, devices based on organic conductors have been object of increasing interest, both as actuators and as sensors. Here, we explore the opportunity to establish a functional interplay between light sensitive, organic semiconductors and plant systems. We chose Arabidopsis thaliana as a widely accepted plant model, and we study the effect of a work-horse photovoltaic conjugated polymer, poly-hexyltiophene, on guard cells phsyiology. Cell viability in presence of the conjugated polymer is evaluated. Effects of polymer optical excitation and cell metabolism are also preliminarily investigated and critically discussed.

The plant cell is protected by a complex polysaccharide cell wall composed primarily of cellulose, hemicellulose and pectin. The cell wall is followed by a semi-permeable barrier, the plasma membrane (PM) that serves two main roles: (1) transport of essential molecules in and out of the cell, and (2) sensory transduction of environmental stimuli as well as adaption to external influences. Considerable progress has been made over the past two decades to understand plant membrane proteins from a genetic perspective, but complementary tools for physiological, electrophysiological, biochemical characterization, and for studying protein function are facing unmet demand. This work highlights a new cell-free platform capturing the properties of the plant PM. This platform consists of a supported proteo-lipid bilayer (SPLB) derived from the plant PM. The main advantage of this platform is the compatibility with state-of-the-art surface, bioelectronic, and biophysical characterization tools. We demonstrates three straightforward ways to obtain proteoliposomes, from plant protoplasts. Three different plants (Arabidopsis thaliana, Nicotiana benthamiana and Zea mays) were successfully used for the formation of SPLBs. This plant based approach enables direct incorporation of transgenic membrane proteins into supported lipid bilayers without using detergents and reconstitution processes and provides a novel tool to isolate and study membrane properties, transport phenomena, membrane-proteins, and protein-lipid interactions. We illustrate here the single molecule tracking of plant membrane proteins using this platform. In the future, we anticipate that this platform will allow studies of the plant membrane characteristics that will enable design of the biotic/abiotic interfaces for new sensing applications that merge plant membranes with electro-active materials.

We vapor print conformal conjugated polymer electrodes directly onto living plants and use these electrodes to probe the health of actively-growing specimens. The conducting polymer PEDOT can be deposited on the surface of living organisms without damaging their health and self-sustenance. A bioimpedance signal can be portably recorded using these vapor printed electrodes, which can reveal detailed information about the health of the organism. Notably, polymer electrodes can be created directly on living plants, meaning that bioimpedance measurements can be performed on-demand, throughout the growth cycle of a plant without being damaged by environmental conditions such as wind, water, temperature changes and UV exposure. Vapor-printed polymer electrodes, unlike their adhesive thin-film counterparts, do not delaminate from microtextured living surfaces as the organism matures and do not observably attenuate the natural growth pattern and self-sustenance of the plants investigated herein. In particular, deep tissue damage caused by UVA irradiation and Ozone exposure can be reliably detected noninvasively. Long-term plant health monitoring can find strategic use in food farming, crop management and biohazard signaling.

To precisely monitor the physiology state of plants, this work presents a gentle non-invasive method to transfer functional liquid alloy circuits on living fragile plants using a water-soluble sacrificial layer. This process can print functional electrical circuits in high fidelity on complex irregularity morphology without introducing any external interventions such as heat flow, pressure, acid or base. The data from the circuits on flowers demonstrates this technique is well suitable for monitoring such kind of scenarios.
Internet of things is emerging in sorts of situations, but the access of live plants is rarely studied. With the internet connecting, people could obtain the real-time health sate and various behaviors of plants in the practical or stimulus environments. This can further lead to new customized strategies for crop management and even new approaches for botany research. Nevertheless, many of plant organisms, such as flowers and leaves, are fragile with irregular surface morphology, and their growth are easily affected by external constraints. These features make the integration of functional circuits challenging. Existing methods, such as smart patches, press-on buttons, 3D printing, roll-to-roll gravure, flexographic printing and vapor-printing, either impose external loads to the growing plants and hinder the normal growing process, or may easily harm the plants with external intervention during the complicated process.
In this work, a highly biocompatible polyvinyl alcohol (PVA) film was used as a substrate as well as an intermediate sacrificial layer to transfer the original 2-D liquid alloy (Galinstan) circuits to a vulnerable complex 3-D surface under water pressure. Firstly, the circuit was printed on a PVA film by masked deposition with a method developed by our group. Then the circuit with PVA was placed on water surface.When the PVA film was almost completed dissolved, it would become a PVA/water mixture with super flexibility, stretchability and stickiness. More importantly, this mixturecould maintain the shape of circuit, which make the transffering process predicatable and controllable. Concurrently, the plant organism was gradually immersed through the circuit with a given angle and immersing speed. Consequently, the liquid alloy circuit pattern was transferred and attached to the plant surface by water pressure and Van der Waals force. The whole process can be conducted in minutes, and there is no obviously interference on plants even after three months. To demonstrate the technique, the Galinstan circuits with resolution ~70 um was transferred on complex surfaces with prominent conformalbility, excellent electricity and repeatability. Further, water content sensors and LED circuits were fabricated on rose petals and bean sprouts by this technique. It was found that the measured results could precisely reflect the change of state in different water content, and the sensor could still work well even with an elongation of more than 200% during growth.

Organic bioelectronics is a unique platform of devices and materials that can serve as the actuator and recorder of signals at the technology-biology interface. In the e-Plant effort, we are exploring organic bioelectronics by developing devices that can selectively trigger or sense targeted physiological functions and signaling cascades, at high spatiotemporal resolution. With tailor-made organic electronic ion pumps (OEIP), we have managed to deliver phytohormones to regulate the growth rate of the root hairs of Arabidopsis and to provoke the closure of stomata of Tobaco leaves. With PEDOT-based electrodes we have derived efficient bio-photovoltaic electrochemical cells. And, with organic electrochemical transistors (OECT), including immobilized enzymes, we can monitor sugar concentrations from plants in real time. In the future, this sensor-actuator technology promise for a self-powered plant-area network possible to use in plant biology experiments and also in agriculture.

Despite their inspiration in improving synthetic photovoltaics, photosynthetic systems suffer from stabilities and efficiencies that limit their direct use in photovoltaics. While they benefit from autonomous repair and reproduction, photosynthetic systems contain light-harvesting machinery that is optimized for evolutionary purposes rather than photocurrent production. Herein we present bioengineered cyanobacteria capable of enhanced photocurrent production. We introduce genetically encoded exoelectronic pathways that serve as electron conduits for interfacing photosynthetic cells with nanomaterial scaffolds. These conduits further enable researchers to electronically rewire the cell metabolism and reprogram biological cells to perform as artificial electronic devices. The ability to improve photovoltaic performance through both nanomaterials engineering and synthetic biology unlocks new possibilities in achieving a viable technology.

Photosynthetic bio-solar energy conversion using algae or plant cells has been extensively investigated for the last decades. Photosynthesis occurs in the thylakoid membranes (TMs) that contain photosynthetic apparatus such as photosystem I (PS I) and photosystem II (PS II). In photosynthesis, water molecules are split by PS II and high energy photosynthetic electrons (PEs) are generated. Direct extraction of these PEs without a mediator has been investigated over the last decade to maximize the PE currents. Since PEs are transported along TMs, enlarging contact area between TMs and an electrode is a way to enhance PE extraction. Previously, several approaches have been investigated to maximize contact area using various three-dimensional (3D) electrode designs such as multi-wall carbon nanotubes(MWCNTs) modified electrode, sintered AuNPs on screen printed electrode, indium tin oxide (ITO) modified electrode, or micro-pillar electrodes. Then, TMs were usually drop cast on such 3D structured electrodes. However, it is difficult to have uniform and conformal deposition of TMs over the 3D electrodes by the drop casting method. This often results in unreliable performance of the TM-coated electrodes in particular for electrodes larger than a few mm². In this work, we aimed to enhance extraction of PEs from TM-coated 3D micro-pillar electrodes by electrospraying. We fabricated Si micro-pillar (MP) arrays of aspect ratio (AR) of 1, 1.8 with 1um diameter from metal-assisted chemical etching (MAC-Etching). Then, SU-8 MP arrays of 1 cm² size were replicated from the Si MP array as a master mold. To improve adhesion with a metal layer, the SU-8 MP arrays were immersed in 10 mM of 4-aminothiophenol (ATP) ethanol solution overnight. Au layer was sputter-coated on the 4-ATP-modified SU-8 MP arrays. Then, TMs were electrosprayed uniformly on the SU-8 MP array electrodes. The electrospraying solution contained 1 mg chl/ml of TMs, 3 mM of ammonium acetate, 10% (v/v) of ethanol as a surface tension reducing agent, and 3% (w/v) of sodium alginate as a viscosity modifier. The morphology of TMs, which were electrosprayed over SU-8 photo-anodes, was observed using confocal fluorescence microscopy and SEM to confirm their coating uniformity. TMs were electrosprayed for various spraying times of 0.5, 1, 2, 4 and 10 minutes in order to maximize extraction of PE currents. PE currents were measured using a three-electrode system, under 10 mW/cm² of halogen illumination and bias potential of 400mV vs. Ag/AgCl. The highest PE currents were measured at 4 minutes of TM-electrospraying time. The measured PE currents were 52, 79, and 109 nA/cm² for flat electrodes, MP electrodes with AR of 1, and AR of 1.8, respectively. It was demonstrated that electrosprayed TMs uniformly and conformally covered the entire electrode surface even for MP electrodes with high ARs. It was also shown that higher PE currents were measured from MP electrodes with higher AR values. This indicates larger contact between TMs and individual MPs were formed, resulting in enhanced direct electrochemical contacts.

The light reactions of natural photosynthesis can be manipulated towards electricity generation in electrochemical devices called biological solar cells or photo-bioelectrochemical cells (PBEC) that resemble conventional fuel cells in principle without the requirement of a chemical fuel. The device uses a photo-bio anode and a enzymatic cathode that could convert light energy into electricity by photosynthesis assisted oxidation of water. The photosynthetic reactions occur in the thylakoid membrane and involve a complex set of redox reactions that result in the transfer of electrons between protein complexes. The electrons from the photosynthetic reactions can be channeled towards an external electrode for electricity generation by inhibiting the photosynthetic protein complexes in order to divert the flow of electrons towards to desired electron transport path within the photosynthetic membrane. Investigation using site-specific photosynthesis inhibitors revealed that the electrons generated through water oxidation reaction from photosystem II complex can be diverted to the external electrode for photocurrent generation primarily through the plastoquinone pool. Besides site specific inhibition, other approaches to increase photocurrent generation were investigated. One of those approaches is to engineer photosynthetic organisms such as Synechococcus elongatus PCC7942 with an outer membrane cytochrome (OmcS) derived from dissimilatory metal reducing bacteria which increase the photocurrent generation at an external electrode by two fold. This presentation will summarize some of our recent work on multiple different approaches to enhance photocurrent generation through an understanding of electron transport reactions in photosynthetic membranes.

Anaerobic bacteria routinely interface with abiotic electrodes to catalyze the interconversion of electrical and chemical energy. Integration of materials at the biotic/abiotic interface has improved electron transfer efficiency and cell adhesion, but charge injection/extraction is still largely restricted to the immediate proximity between bacteria and the external electrode. A three-dimensional organization whereby electronic contact extends beyond the confines of the electrode surface would be desirable. Here, we introduce bioelectronic composites comprised of the self-p-doped conjugated polyelectrolyte CPE-K and Shewanella oneidensis MR-1. These composites are simple to assemble and biocompatible. Furthermore, testing in three-electrode electrochemical cells show that the composites can generate ~200-fold greater biocurrent density than control S. oneidensis MR-1 biofilms. The impact of CPE-K concentration on bioelectronic composite performance will be discussed, along with an evaluation of electroactive surface area and charge transfer resistance.

Extracellular electron transfer, in which electrons are transferred from the metabolism of a microbe to an extracellular substrate is an intriguing process that may have applications in living electronics, bioremediation, chemical synthesis, sensing, and microbial fuel cells. Although a number of anode-respiring bacteria can exchange electrons with extracellular electrodes, many genetically tractable model microorganisms, especially those used in synthetic biological applications, cannot. Furthermore, the mechanisms by which microbes interact with extracellular electrodes and the biological components required remain unclear.
Among the hundreds of c-type cytochromes found in Geobacter sulfurreducens, OmcZs has been shown to have the largest impact on extracellular electron transfer to electrode surfaces. Herein, we overexpressed OmcZs in E. coli and defined its function in vitro. UV-vis absorption and florescence assays show that it can reduce riboflavin. Furthermore, electrochemical measurements show that E. coli cells expressing OmcZs are capable of extracellular electron transfer, a function not native to E. coli. The activity can be enhanced by inclusion of riboflavin in the experiment. Currents produced are comparable to other, more complex E. coli circuits reported by others. We conclude that OmcZs and riboflavin alone are sufficient to create a minimal, functional, synthetic pathway in E. coli for extracellular electron transfer.

Photoactive electrodes based on redox-active films are a promising approach to increasing the active area in which enzymes can be deposited and electrochemically accessed via mediated electron transfer. Beside this, these films are able to entrap the deposited enzyme. A great challenge to increase the efficiency arises from different charge recombination pathways, intrinsically connected to the process of the light-induced electron transfer chain from the electrode to the final solar fuel formation or to the charge carrier for electricity production [1]. Experimental investigation based on electrochemical methods and/or product quantification often have the disadvantage, that they cannot distinguish between the catalytic process and the detrimental effect of different short-circuiting processes. We present a general model constructed from diffusion and reaction processes describing redox-active film based biophotoelectrodes [2]. The model predicts the resulting photocurrent-time curve and distinguishes between the main catalytic process and the contribution of the two proposed short-circuiting pathways. It describes the corresponding time dependent concentration profiles of all involved species which can be analyzed to identify bottlenecks present in the solar fuel or photocurrent generation of a given system. The equations consist of dimensionless groups for a straightforward identification of possible limiting processes. The importance of the model for quantitative understanding of biophotoelectrochemical processes is highlighted with an example of simulation results predicting the effect of the diffusion coefficient of the charge carrier on photocurrent generation for different charge recombination kinetics.


References:
[1] M. M. Nowaczyk, N. Plumeré, Adv Biochem Eng Biotechnol. 158 (2016) 111-136.
[2] D. Buesen, T. Hoefer, H. Zhang, N. Plumeré, Faraday Discussion. (2019) accepted

Biofuel cells are one of the most attractive power sources for implantable and microscale devices because of their biocompatibility and ability to operate at near neutral pH and ambient temperature. A typical enzymatic glucose biofuel cell comprises two enzyme-modified electrodes, with Glucose Oxidase (GOx) catalyzing glucose oxidation into gluconolactone at the anode, and Bilirubin Oxidase (BOD) catalyzing oxygen reduction at the cathode. For these enzymes to be recycled, specific redox mediators are necessary, to insure an efficient electron transfer between their active sites and the electrodes.
During the process of photosynthesis, vegetal cells use energy from sunlight to convert carbon dioxide and water into glucose and oxygen, which could be both used in a glucose biofuel cell as fuel and oxidant, respectively. Our idea was therefore to integrate the energy harvesting system directly within the structure of a living plant. To achieve this, the direct functionalization of the vascular system containing the plants’ endogenous sugars and oxygen is necessary in order to operate the biofuel cell. The electrodes of such bio-hybrid systems should encompass conductive materials, enzymes and redox mediators.
Our approach is based on the growth of a conductive material (functionalized with enzymes and redox mediators) inside the vascular system of the plant, either through the roots or the leaves. The design, synthesis and characterization of such smart materials will be described and discussed.
We believe that the integration of energy harvesting and storage systems within the living plants could be a major breakthrough for energy applications.

Reference:
Stavrinidou E. et al. PNAS, 114, 2807-2812 (2017)

Photosynthesis produces high energy photosynthetic electrons (PEs) with internal quantum efficiency (IQE) of nearly 100%. Due to this high IQE, bioenergy harvesting using algal cells or photosynthetic organelles such as photosystems (PS I, II) and thylakoid membranes (TMs) has been studied extensively. In the absence of a mediator, the magnitude of harvested PE currents directly depends on an electrical contact between the photosynthetic organelles and an electrode. Thus, thicker deposition of the photosynthetic organelles on an electrode does not increase PE currents despite larger quantity of PEs generated from the thicker photosynthetic film. Even in the presence of a mediator, although a mediator can diffuse between a spot of PE generation and an electrode, PE transportation through the thicker film becomes more difficult.
In this study, we propose formation of electrical percolation networks for PE transport from the surface of the film to an electrode. Electrical percolation network was formed by mixing highly-conductive single-wall carbon nanotubes (CNTs) and thylakoid membranes (TMs). Since CNTs have no charge on the surface, we coated mussel-adhesive protein (MAP) on CNTs to have a positive surface potential due to the exposed amine groups of the MAP. The positive zeta potential resulted in well-dispersed CNTs in an aqueous solution. The exposed amine groups of MAP-CNTs also formed peptide bonds with the carboxyl groups of TMs such that CNTs and TMs were firmly connected. The percolation network in MAP-CNT/TM film was analyzed by cyclic voltammetry (CV), chronoamperometry (CA), electrical impedance spectroscopy (EIS), and linear sweep voltammetry (LSV). CNT percolation networks were achieved when the CNT weight was near 30% of the weight of chlorophyll a (Chl a) in the MAP-CNT/TM composites. Moreover, when the CNT weight was 50% of the weight of Chl a, the composites achieved the highest electrical conductivity without a mediator. Finally, when we increased the amount of TM deposition on an electrode, larger difference of PE current magnitudes was observed between the MAP-CNT/TM and TM-only films. The PE currents from the MAP-CNT/TM composite film became about 100 times larger at the Chl a concentration of 300 μg/cm².

Photosynthetic bacteria can be envisaged as a source of active materials for photoconversion or light-induced processes in bioelectronic devices [1].
The Reaction Centers (RCs) from photosynthetic bacteria, such as Rhodobacter sphaeroides,are photoenzymes capable to convert solar energy into charge separated states with efficiency close to 100% [2], thus being promising biomolecules to design a new generation of photoactive materials for light-responsive bioelectronics [3].
We recently demonstrated the possibility to increase the light harvesting capability of the native RC isolated from Rhodobacter sphaeroides bacteriumby covalently affixing different tailored molecular antennas, thus obtaining hybrid systems which outperform the native protein in light absorption and photo-generation[4,5]. We have also built up smart supramolecular architectures combining multiple enzymes assembled with tailored linkers, which are able to perform manifold functions. Finally, we have explored approaches for integration of RC as the light-responsive material in bio-optoelectronic and electrochemical devices. In particular, we have developed protocols for addressing the RC on thin films of molecular or polymeric semiconductors deposited onto metal electrodes by covalent [6] or weak interactions. Biomimetic polymers, such as polydopamine, have been also investigated as efficient electronic interfaces of RC with electrodes in photoelectrochemical cells.
The lecture will discuss the logic behind designing and synthesizingmaterials for sunlight photoconversion and for light-triggered bioelectronics, by combination of tailored functional molecules with highly efficient bacterial photoenzymes optimized in billions of years of evolution.

References
[1] F. Milano, A. Punzi, R. Ragni, M. Trotta, and G. M. Farinola, Adv. Funct. Mater., 1805521, 1-17 (2018)
[2] R. R. Tangorra, A. Antonucci, F. Milano, S. La Gatta, G. M. Farinola, A. Agostiano, R. Ragni and M. Trotta, In Handbook of Photosynthesis(3^rdEd, M. Pessarakli, Ed. CRC Press), 201-220 (2016)
[3] A. Operamolla, R. Ragni, F. Milano, R.-R. Tangorra, A. Antonucci, A. Agostiano, M. Trotta and G. M. Farinola, J. Mater. Chem. C,3(25), 6471-6478 (2015)
[4] F. Milano, R.-R. Tangorra, O. Hassan Omar, R. Ragni, A. Operamolla, A. Agostiano, G. M. Farinola and M. Trotta, Angew. Chem. Int. Ed.,51(44), 11019-11023 (2012).
[5] O. Hassan Omar, S. La Gatta, R. R. Tangorra, F. Milano, R. Ragni, A. Operamolla, R. Argazzi, C. Chiorboli, A. Agostiano, M. Trotta and G. M., Farinola, Bioconj. Chem., 27(7), 1614-1623 (2016).
[6] E. D. Glowacki, R. R. Tangorra, H. Coskun, D. Farka, A. Operamolla, Y. Kanbur, F. Milano, L. Giotta and N. S. Sariciftci, J. Mater. Chem. C, 3(25), 6554-6564 (2015).

Plants convert radiative energy from the sun into carbohydrates being the primary resource in our ecosystem. Recently, the use of organic electronic materials allowed interaction with plants in a new way: using the vascular system of plant to form electrical circuits and devices targeting bio-hybrid devices. Firstly, a water-soluble conducting polymer, PEDOT-S has been uptaken by the xylem of a rose, forming self-organized conducting wires in the stem. As a next step a conjugated oligomer, ETE-S was developed with bigger versatility in distribution within the plant due to its small size. The ETE-S in-vivo polymerized within the living tissue forming a network of conducting wires from the stem to the flower. It is well known that plant has developed mechanisms for stability and defense that rely on the polymerization of compounds within the cell wall. The major actor in these processes is called the type III peroxidase. This enzyme can proceed with two different catalytic cycles to reduce or increase the cell wall density. The increase of the cell wall density is achieved through the peroxidative cycle that uses peroxide to crosslink phenolic compounds on the cell wall creating a complex network of various polysaccharides such as lignin or extensin. In this work, we demonstrate the involvement of this peroxidative cycle in the polymerization of the ETE-S oligomer by reproducing the reaction in-vitro. We then follow the kinetics of the polymerization reaction while changing the enzyme or substrate concentrations to understand deeper the underlying mechanism. By understanding the mechanism of in-vivo polymerization we can rationally design materials for plant functionalization as well as optimize targeted functionalization.

Organic bioelectronic devices use conducting polymers as active materials because of their soft nature and mixed ionic and electronic conductivity. Electrode arrays, transistors, and drug delivery devices have taken advantage of the conducting polymers in order to improve their performance for the purpose of diagnosis and treatment. However, the aforementioned devices are commonly being developed by using expensive microfabrication techniques. Moreover, conducting polymers occupy only a part of the whole device geometry, while the rest acts as support. Biology, however, builds biological devices by using its own machinery. Recently, the machinery in plants has shown to polymerize trimers resulting in conducting polymer wires, electrodes, and supercapacitors. Here, we show in vivo polymerization of thiophene based molecules by the machinery in the brain in Zebrafish. Localized areas with conducting polymers exist in different parts of the brain. This process paves the way for the development of self-organized organic electronic devices.

There is an urgent need to develop a sustainable battery technology that is cheap, environmentally friendly, easy to fabricate and to dispose of, especially to tackle the world-wide increase in illegally dumped electronic wastes. Microbial biophotovoltaic (BPV) technology is a renewable bioenergy system currently being developed at the laboratory scale. It generates electricity from the photosynthetic metabolism of cyanobacteria and microalgae and exploits their ability to convert light energy into electrical current using water as the source of electrons. Innovative approaches are needed to solve scale-up issues such as cost, ease of fabrication (particularly the fabrication of the inorganic and biological (microbes) parts).
In this talk, I will report the feasibility of the thermal-inkjet method to fabricate a thin-film paper-based BPV cell consisting of a layer of cyanobacterial cells on top of a carbon nanotube conducting surface on plain copy paper. The digitally printed thin-film BPV system produced electricity both in the light and dark, with a maximum electrical power output of 0.38 mW m-² in one system and the sustained electrical current production over 100 hours in another more fully printed system. I will discuss latest efforts in development of the proof of concept and address limitations and challenges as well possible applications in the area of printed bioelectronics.

The concept of a light-emitting plant, or plant exhibiting chemiluminescence powered from its own stored chemical energy, offers promise to advance off grid illumination and other autonomous photonic applications. In this work, introduce and investigate an additional nanoparticle designed to augment plant light emission in the form of silica coated strontium aluminate nanoparticles as nanophosphore elements. These nanoparticles can adsorb and re-emit generated light at longer times, increasing the duration of light emission, where the particles sizes and concentration are the key parameters of plants brightness tunability. Moreover, such nanophosphores can also scavenge additional energy from solar fluence, increasing and augmenting total light emission from the plant. Infiltrated strontium aluminate particles showed homogeneous distribution inside plants leaves in spongy mesophyll region without penetration inside plants cell, preserving their intact structure, as well as efficient particles infiltration deep into the plant’s stem.

The intersection between synthetic biology and materials science is an underexplored area with great potential to positively affect our daily lives, with applications ranging from manufacturing to medicine. Inorganic-biological hybrid systems have potential to besustainable, efficient, and versatilechemical synthesis platforms by integrating the light-harvesting properties of semiconductors with the synthetic potential of biological cells. We have developed a modularbioinorganic hybrid platformthat consists of highly efficient light-harvesting indium phosphide nanoparticles and genetically engineered Saccharomyces cerevisiae: a workhorse microorganism in biomanufacturing. The InP particles are adhered to the yeast surface using a versatile polyphenol-based chemistry. The yeast harvests photo-generated electrons from the illuminated nanoparticles and uses them for the cytosolic regeneration of redox cofactors. This enables the decoupling of biosynthesis and cofactor regeneration, facilitating a carbon- and energy-efficient production of the metabolite shikimic acid – a common precursor for several drugs and fine chemicals. We observed higher per-cell production of shikimic acid for biohybrids compared to their unmodified counterparts, in a light-dependent manner, and a global metabolic shift toward shikimic acid from other fermentative byproducts, like glycerol and ethanol. Our work provides a platform for the rational design of biohybrids for efficientbiomanufacturing processeswith higher complexity and functionality. The biohybrid polyphenol-based fabrication scheme can easily be adapted to virtually any combination of particle and cell type, greatly expanding the range of accessible systems and facilitating the exploration of structure-function relationships. Additionally, the vast toolbox of available genetically modified yeast strains will enable further probing of electron transport mechanisms and applicability to a wide variety of biochemical targets.

Semi-artificial photosynthesis interfaces biological catalysts with synthetic materials and aims to overcome the limitations of natural and artificial photosynthesis.¹ It also provides an underexplored strategy to study the functionality of biological catalysts on synthetic scaffolds through a range of techniques. This presentation will summarise our progress in integrating biocatalysts in bespoke hierarchical 3D electrode scaffolds and photoelectrochemical circuits.² The fundamental insights gained into the function of the water oxidation Photosystem II will be discussed, where (i) unnatural charge transfer pathways have been revealed at the enzyme-electrode interface, and (ii) O₂ reduction that short-circuit the water-oxidation process has been discovered.^3-4
The wiring of Photosystem II to a H₂ evolving hydrogenase or a CO₂ reducing formate dehydrogenase has subsequently enabled the in vitro re-engineering of natural photosynthetic pathways. We have assembled efficient H₂ evolution and CO₂ reduction systems that are driven by enzymatic water oxidation using semi-artificial Z-scheme architectures.^5-8 In contrast to natural photosynthesis, these photoelectrochemical cells allow panchromic light absorption by using complementary biotic and abiotic light absorbers. As opposed to low-yielding metabolic pathways, the electrochemical circuit provides effective electronic communication without losses to competing side-reactions. Progress in the integration of robust live cyanobacteria in 3D structured electrodes will also be discussed.⁹

Selected References
(1) Kornienko et al., Nature Nanotech., 2018, 13, 890–899
(2) Mersch et al., J. Am. Chem. Soc., 2015, 137, 8541–8549
(3) Zhang et al., Nature Chem. Biol., 2016, 12, 1046–1052
(4) Kornienko et al., J. Am. Chem. Soc., 2018, 140, 17923–17931
(5) Sokol et al., Nature Energy, 2018, 3, 944–951
(6) Nam et al., Angew. Chem. Int. Ed., 2018, 57, 10595–10599
(7) Sokol et al., J. Am. Chem. Soc., 2018, 140, 16418–16422
(8) Miller et al., Angew. Chem. Int. Ed., 2019, 58, 4601–4605
(9) Zhang et al., J. Am. Chem. Soc., 2018, 140, 6–9

Microbial electrochemical systems provide an environmentally-friendly means of energy conversion between chemical and electrical forms, with applications in wastewater treatment, biofuel production, and environmental biosensing. However, a major challenge to further development, miniaturization, and deployment is the limited thickness of biofilms, necessitating large anodes to achieve sufficient signal-to-noise ratios. In this talk, we present our recent demonstrations of embedding living electroactive bacteria in electrodes based on organic conducting polymers, enabling truly hybrid bioelectronics with a well-defined biological-to-electronic signal pathway. Our method embeds a large, dense biomass of current-producing bacteria inside a three-dimensional PEDOT:PSS matrix, which we call a multilayer conductive bacterial film. The resulting bio-hybrid electrode provides a conformable electrode interface to each bacterium, provides nutrient-permeability, and permits electron transfer via multiple bacterial mechanisms. As a result, it produces 20× more current than a native biofilm grown on the same substrate. We will discuss how this work can both expand our mechanistic understanding of interfacial electron transfer and enable miniaturization and direct incorporation of living biological materials into electronic devices, such as organic electrochemical transistors and microbial biosensors.

Conducting polymers offer several key advantages over their inorganic counterparts, such as mechanical flexibility, transparency, and material abundance, which can enable low-cost fabrication and novel applications such as printed and flexible electronics. The conducting polymer poly(3,4-ethylenedioxythiophene), PEDOT, is one material which displays (among others) high electrical conductivity [1], enhanced thermal conductivity [2], good electrocatalytic performance [3], as well as thermoelectric behaviour [4]. Enhancing the properties of PEDOT has been achieved through Vapour Phase Polymerisation (VPP), an oxidative polymerization process under vacuum conditions. We report that VPP PEDOT shows interesting interactions with anions, in some examples almost specific ion effects [5]. For example, the uptake of anions from an aqueous solution into electrochemically reduce PEDOT is highly dependent on the anion itself. More interestingly is nitrate is specifically absorbed by PEDOT when present in a mixed electrolyte solution [6]. This selective absorption is hypothesised to originate from anion-π interactions, in part validated by MD simulations [5]. Such interactions, relating to both chemistry and structure, lead to interesting opportunities in energy storage as well as (agricultural) sensing.

References:
[1] Fabretto MV, Evans DR, Mueller M, Zuber K, Hojati-Talemi P, Short RD, Wallace GG, Murphy PJ. "Polymeric material with metal-like conductivity for next generation organic electronic devices", Chem. Mater. 24, pp. 3998-4003, 2012.
[2] Weathers A, Khan ZU, Brooke R, Evans D, Pettes MT, Andreasen JW, Crispin X, Shi L. "Significant Electronic Thermal Transport in the Conducting Polymer Poly(3,4-ethylenedioxythiophene)", Adv. Mater. 27, pp. 2101-2106, 2015.
[3] Vucaj N, Quinn MDJ, Baechler C, Notley SM, Cottis P, Hojati-Talemi P, Fabretto MV, Wallace GG, Murphy PJ, Evans DR. "Vapor Phase Synthesis of Conducting Polymer Nanocomposites Incorporating 2D Nanoparticles", Chem. Mater. 26, pp. 4207-4213, 2014.
[4] Bubnova O, Khan ZU, Wang H, Braun S, Evans DR, Fabretto M, Hojati-Talemi P, Dagnelund D, Arlin J-B, Geerts Y, Desbief S, Breiby D, Andreasen JW, Lazzaroni R, Chen W, Zozoulenko I, Fahlman M, Murphy P, Berggren M, Crispin X. "Semi-metallic polymers", Nat. Mater. 13, pp. 190-194, 2014.
[5] Rudd S, Franco-Gonzalez JF, Kumar Singh S, Khan ZU, Crispin X, Andreasen JW, Zozoulenko I, Evans DR. “Charge transport and structure in semimetallic polymers”, J. Polym. Sci. B 56, pp. 97-104, 2018.
[6] Rudd S, Dalton M, Buss P, Treijs A, Portmann M, Ktoris N, Evans DR, “Selective uptake and sensing of nitrate in poly(3,4-ethylenedioxythiophene)”, Sci. Rep. 7, pp 16581, 2017.

Relative humidity (RH) detection and quantification play a major role in both agriculture and floriculture, as the water content in air and soil has a strong impact on the plant’s growth, flowering and fruiting.¹ Nowadays, the crop and floral industry are facing unprecedented challenges, especially because of the combination of the rapid increase of the world’s population with the unpredictable effects of global warming: atmospheric and soil humidity monitoring has thus become crucial to avoid production insecurity problems.² Research on air and soil RH sensors for agricultural and floriculture applications is now focusing on the development of devices that are low-cost, low power-consumption and easily portable.³ These needs may be met by utilising printing techniques to fabricate simple capacitive sensors on flexible, plastic foils.
The devices presented in our work are fabricated on polyimide foils (Kapton®) by inkjet-printing a capacitive structure consisting in coplanar, interdigitated silver electrodes; cellulose acetate butyrate is subsequently printed onto the interdigitated comb and used as humidity sensing layer.
Impedance spectroscopy (at fixed RH) was used to identify the frequency range where our sensors exhibit the most stable capacity values; the sensors’ response to different RH levels was fully characterised by performing both static and dynamic measurements.
A particularly innovative aspect of our work is the sensors characterisation in soil: the printed devices were dipped inside potting soil at different humidity levels (measured by means of a commercial electronic sensor) and again both statically and dynamically characterised. Their lifetime inside the soil was also investigated.
Finally, we present the preliminary results of a protocol to fabricate the aforementioned printed RH humidity sensors using directly plant leaves as substrate.
Our results could pave the way for the fabrication of the next generation of low-cost, flexible and environmentally-friendly humidity sensors.
References
[1] M. Lee and H. Yoe, BioMed Research International (2015), 2015, 292543.
[2] I. Ali Lakhiar et al., Journal of Sensors (2018), 2018, 8672769.
[3] M. D. Steven, J. A. Clark, Elsevier, 2013, ISBN: 1483161781.

Food and agricultural commodities contamination by various toxigenic molds (fungi) is a serious neglected problem, affecting ~25% of the world’s crops with staggering economical loses [1]. Regardless of decades of extensive research, mycotoxins continue to penetrate the food chain trough food and feed crops and pose health risks both to humans and livestock [2]. Mycotoxins may lead to adverse health effects, including cancer, gene mutation, as well as estrogenic, gastrointestinal and kidney disorders [3]. Some mycotoxins are also immunosuppressant which reduce resistance to infectious diseases [4, 5]. Mycotoxins are secondary metabolites produced by specific filamentous fungi species usually belong to Aspergillus, Penicillium or Fusarium that infect crops at the field level and endure on foods during storage. Due to the increased toxicological effects, many countries have set stringent limits for mycotoxins residues in food and feedstuffs particularly with respect to international trade, including developed and developing countries [1]. Thus, it is rather essential to have an accurate, rapid and sensitive method for multiplex detection of mycotoxins in unprocessed food and feed. An oxidized porous silicon (PSi) nanostructures, a Fabry-Pérot thin film, is synthesized and used as an optical transducer element for the detection and quantification of Fumonisin B1 (FB1, target molecule). The nanostructure is modified with specific robust bioprobes for the validation of fungal contamination and its secreted toxins by indirect immunoassay amplified by FB1-horseradish peroxidase (HRP) conjugates. The catalytic activity is monitored in real-time by reflective interferometric Fourier transform spectroscopy (RIFTS). This technique is sensitive to small changes in the average refractive index of the thin-film induced by enzymatic activity products infiltrating the pores. Optical studies reveal high specificity and selectivity towards FB1, presenting LOD of 0.38 ppm. The main advantage of the presented biosensing concept is the ability to detect fumonisin contamination, at environmentally relevant concentrations, using a simple and portable experimental setup.

[1] Deng, G., et al., High sensitive immunoassay for multiplex mycotoxin detection with photonic crystal microsphere suspension array. Anal. Chem., 2013. 85(5): 2833-2840.
[2] Gallo, A., et al., Review on mycotoxin issues in ruminants: occurrence in forages, effects of mycotoxin ingestion on health status and animal performance and practical strategies to counteract their negative effects. Toxins, 2015. 7(8): 3057-3111.
[3] Wu, F., J.D. Groopman, and J.J. Pestka, Public health impacts of foodborne mycotoxins. Annu. Rev. Food Sci. Technol., 2014. 5: 351-372.
[4] Antonissen, G., et al., The impact of Fusarium mycotoxins on human and animal host susceptibility to infectious diseases. Toxins, 2014. 6(2): 430-452.
[5] Wild, C.P. and Y.Y. Gong, Mycotoxins and human disease: a largely ignored global health issue. Carcinogenesis, 2010. 31(1): 71-82.

Low cost biodegradable sensors for environmental monitoring can enable the collection of large data sets useful for resource optimization in precision agriculture. Therefore, we have developed wireless soil moisture and nitrogen sensors that are printed from solutions of functional biodegradable materials using scalable manufacturing techniques. These sensors will be placed in the field and remotely interrogated, controllably degrading into the soil over time when no longer needed.

The development of biodegradable sensors presents a complex design problem wherein sensor performance must be constant for several months until a prescribed end of life is reached. We leveraged the complex ecosystem of soil in order to more carefully tune degradation rates of electronic devices. We combined enzymatically degradable polymers, such as poly(lactic acid), polycaprolactone, and polyhydroxybutyrate, with benign conductive materials such as magnesium and carbon, to print conductive lines. Similar polymers were also employed as substrates and encapsulant materials. Techniques such as solvent aging and the addition of various composite fillers were explored and optimized for electronic characteristics and degradation rates. We then fabricated wirelessly readable and fully degradable capacitive soil moisture sensors and report on performance shifts due to degradation.

By integrating and modifying existing enzymatic biodegradation techniques, and developing new accelerated testing approaches, we compared degradation rates in continuous flow with traditional batch techniques. We then compared this data with soil-compost mixtures in incubator conditions. Component materials, such as polymers and metals, were tested independently for mass loss and topological changes; and then printed composites and entire devices were tested to investigate performance loss during degradation. In order to ensure the degraded byproducts have a benign impact on the crops being monitored at the local level and on streams and water tables at the systems level, we analyzed the effluent from these accelerated degradation techniques for ecotoxicity.

Technologies that allow manipulation of cellular processes with high spatiotemporal resolution can lead to engineering plants that are tolerant of harsh conditions, which is one of the primary goals of modern plant biology. The organic electronic ion pump (OEIP) provides unique means for the electronically-mediated delivery of phytohormones without fluid flow at the cellular scale. Here the OEIP is used to deliver phytohormones at very high accuracy with respect to rate and spatial resolution directly into leaf tissue. Owing to the capillary form factor and the sub-100 µm diameter of the device, there is no significant wound response from the plant benchmarking the OEIP as minimal invasive tool for regulating plant physiology. The delivery of ABA, the phytohormone that mediates plant’s tolerance to stress, induces closure of stomata, the microscopic pores in leaf epidermis that play a vital role in both photosynthesis and transpiration. High performing OEIP mediated delivery revealed surprising kinetics of the ABA-induced signal. This technology offers a new versatile tool for basic plant research but also potential application in agriculture.

Hydrogels with Fe(III) ions and carboxylate-containing polysaccharides such as alginate and pectin were photoresponsive and showed photoreduction to Fe(II) and oxidative decarboxylation of the polysaccharide¹. Interestingly, these Fe(III)-polysaccharide hydrogels showed phosphate uptake capability from aqueous solutions for pH range 4.8 – 11.5. Different hydrogel beads prepared by changing the polysaccharide combination showed a phosphate uptake more than 1 mgg^-1 at neutral pH of 7. Reclaiming the phosphate from chemical fertilizers and animal manure applied in agriculture is a challenge, and thus we explored the use of these hydrogles to capture phosphate from waste. The phosphate uptake from 100 ppm solutions at pH = 7 was 99% and and~80% from 800 ppm solutions. For raw manure solutions around same pH and with a phosphate content of 340 ppm, the phosphate uptake was 99%. This showed that the uptake behavior was unaffected by the presence of other ions and dissolved solid particles in manure.
These phosphate-loaded hydrogels were also photoresponsivie, and the Fe(III)- carboxylate photochemistry slowly degraded the hydrogel network while releasing the phosphate ions. In addition, the amount of phosphate release could be controlled with light intensity. Nutrient uptake by plants (kale and tomato) from the fertilizer-loaded hydrogels was significant. Compared to the controls, plants treated with our fertilizer hydrogel beads showed enhanced plant growth and fruit formation showing the photo controlled nutrient release mechanism. These results show a promising system for reclamation of phosphate from waste solutions and reusing them as a controlled release fertilizer for sustainable agriculture.

The enzymatic degradation of biomacromolecules is critical to the sustenance of life, whether at the cellular or the ecological size scale. Thus, measurement of hydrolytic enzyme activities is often used to characterize an ecosystem’s capacity to support life, particularly in agriculture, where this rate portends to the long-term return of nutrients and overall health of the soil. While soil hydrolytic activity assays exist, they are often confined to dedicated lab spaces and suffer inconsistencies related to in-house preparation of unstable reagents and variable dynamic ranges. In this work, we present a modular sensor that can be used to evaluate modification of different large, agriculturally-relevant substrates. By suspending semiconducting single-walled carbon nanotubes (SWNT) in an amphiphilic enzyme substrate, we observe enzymatic modification of the substrate through modulation of SWNT fluorescent signal. These substrates may be inherently amphiphilic or may be chemically modified to exhibit hydrophilic and hydrophobic domains. Herein, we demonstrate the throughput and modularity of this nanosensor by measuring proteolytic, cellulytic, and lignin-modifying activity in soil to evaluate health of the soil microbiome. We explore variables including type of crop rotation, microbial carbon content, and soil categorization for each type of enzyme. We also demonstrate this assay’s utility for on-site testing using a custom, portable, battery-powered fluorimeter.

During photosynthesis, plants convert light into chemical energy, building sugar molecules, using water and carbon dioxide. Glucose is one of the main products of photosynthesis and it holds an essential role in plants function being the fuel for respiration, growth and development. In order to meet the plants energetic demand sugars are transported from source to sink tissues through the phloem vascular tissue, mainly in the form of sucrose. Although it is well known that sucrose is transported via the phloem, there is evidence of sucrose transport in the xylem vascular tissue, that is adjacent to phloem and it is mainly responsible for water transport. Monitoring sugar transport in xylem, has gain interest in the scientific community since sugars can play a vital role in the wood development; however the current understanding is limited. Apart from being energy source sugars are also important signalling molecules, involved in regulation of physiology including stress responses.
Currently there are no methods that allow real time monitoring of sugar transport in plants. Sugar detection is based on enzymatic assays, chromatography and/or mass spectrometry; all of these methods require sample collection and preparation. Organic electrochemical transistors (OECTs) are ideal tools for interfacing with organisms, since they can translate complex biological input to an electronic readout signal. OECT sensors can operate in complex media and they can directly detect products from the biological unit. Recently we demonstrated that OECTs are able to measure the export of glucose in real time, from isolated chloroplasts, with a temporal resolution of 1min.
In this work we develop implantable OECT-based sugar sensors for in-vivo, real time monitoring of sugar transport in poplar trees. OECTs are developed with standard microfabrication techniques, with multienzyme functionalized gate for specificity to glucose and sucrose. The OECTs sensors show high device to device reproducibility, stability during the operation in the in-vivo environment and most importantly they do not cause a significant wound response in the plant. We demonstrate that OECT-based sugar sensors are attractive tools for studying transport kinetics in plants, in vivo and real time.

Solid-state nanopore is a physical sensor capable of detecting single-particles and molecules. The sensing mechanism involves measurements of the temporal ionic current blockade called resistive pulses by an individual object passing through a nanopore. Combing with machine learning to examine difference in multiple features in each ionic current profile, it enables discriminations of analytes by the size, shape, and surface charge states (M. Tsutsui et al., Sci. Rep. 7, 17371 (2017)). In my presentation, I will present our recent research on peptide nanopore approach for adding bioselectivity to the multimodal single-particle analysis. We synthesized recognition probes of various amino acid sequences designed to possess specific affinity to surface proteins of bacteria and/or viruses. We adhered the peptides on Au nanopore wall so as to render antigen-specific perturbation to single-particle translocation dynamics. We found positive correlation between the single-amino-acid-sensitive peptide-analyte affinity and the single-bacteria and -virus translocation time, which was ascribed to longer residence duration of the bioparticle inside the nanochannel with stronger peptide-protein interaction at the wall surface. Multiphysics simulations revealed the intermolecular-interaction-mediated single-antigen translocation mechanism to involve tensile deformations of the soft envelope via electrophoretic forces. Most importantly, we demonstrated multimodal discriminations of flagellated bacteria (Tsutsui et al., Anal. Chem. 90, 1511 (2018)) and influenza types (Arima et al., J. Am. Chem. Soc. 140, 16834 (2018)) in a high-dimensional feature space that revealed the resistive pulse apex as an effective characteristic for distinguishing the intermolecular-interaction-derived difference in the in-pore translocation motions of the two viruses. As the amino-acid sequence degrees of freedom can potentially offer variety of recognition ability to the molecular probes, the peptide nanopore approach can be a versatile single-particle immonosensor that promises wide applications in bioanalysis including environmental monitoring to infectious disease diagnosis.

Probing cell adhesion forces is one of the approaches used to evaluate bacterial motility and surface interactions, which are crucial to the understanding of microorganism behavior. This work presents the application of Indium Phosphide (InP) nanowire arrays as force sensors to investigate the inhibitory effect of N-acetylcysteine (NAC) on adhered cells of the phytopathogenic bacteria Xylella fastidiosa.^1,2
These bacteria produce extracellular polymeric substances (EPS), which allow them to adhere on a surface and to form biofilms.² NAC is a cysteine analog assumed to disrupt disulfide bonds and therefore remove EPS, among other antibacterial effects. However, the NAC action mechanisms on adhered bacteria are not yet validated experimentally³. In this work we analyzed in real time the effect of NAC on cells attached to the top of InP nanowires. Adhesion forces are measured by optical detection of nanowire deflections due to the forces exerted by bacterial cells, using Confocal Laser Scanning Microscopy. Nanowire positions are determined using a 2D Gaussian fitting of the laser reflection profile. Adhesion forces can be calculated from the measured displacements within the limits of linear theory of elasticity.²
Changing bacterial growth time and NAC concentrations in the liquid cell housing the nanowire sensor, we investigated the NAC action on single cells and biofilms. We observed that the measured forces decreased with increasing NAC concentration. Release of cells and biofilms upon NAC addition to the culture are observed in real time; force values on the nanowires decrease once the cells are free. Furthermore, our results suggest a different behavior for the elongated cells bridging neighboring clusters⁴; higher adhesion forces keep them attached to the surface after NAC addition, even after biofilm release.
References
¹P. Sahoo et al., Nano Letters (2016), 16, 7, 4656-4664.
²Silva et al., Small Methods (2018), 2, 1700411.
³Muranaka, L. S.et al., PLoS ONE (2013), 8(8), e72937
⁴Janissen, R. et al., Sci. Rep. (2015) 5, 9856

Programmable release fertilizers have been intensively investigated in the last decades to regularly deliver nutrients to plants. The most used approach is to coat the nutrients using a polymeric matrix to slow or/and control their release on the environment ^[1,2]. However, the matrix may affect the soil properties, as conductivity and pH, depending on its composition, as a result bio-based and biodegradable polymer had been applied to avoid the harm ^[3]. In response to this, nanofibrillated cellulose (NFC) has been considered due to the presence of hydroxyl groups on its chemical structure. Those groups allow chemical functionalization of the NFC's surface, giving it an anionic or cationic charge, and enable the interaction with ionic compounds, e.g. the potassium nitrate nutrient (KNO₃). As an effect, it should hold the nutrient and delay its release depending on the nutrient charge. Also, the spray drying technique was used because of its advantages as versatility, reproducibility, and flexibility of the process. In this work we use two types of nozzles to understand the relation of the produced particle (capsule or sphere ^[4]) with the release mechanism in water. First, the NFC has been chemically modified using TEMPO-mediated oxidation ^[5] and quaternary ammonium salts ^[6]. Then, the encapsulation was done on a Mini Spray Dryer B-290-BUCH (inlet temperature: 180°C, aspirator: 100%) using two-fluid (flowrate: 9ml/min) and three-fluid nozzle (inner flowrate: 1 ml/min, outer flowrate: 10ml/min, approx.) to attain microsphere and microcapsule (core-shell) structure, respectively. The materials were characterized and compared by their structure (Z-potential, conductivity titration, FTIR and XRD), morphology (SEM), thermal properties (TGA) and the nutrient release profile. The nutrient encapsulated by nanocellulose has shown a lower released percentage (in water) as compared to non-encapsulated nutrient (KNO₃). Also, the two- and three-fluid nozzles have formed different structure, as well as the three-fluid nozzle provided a delayed release of nutrients compared to two-fluid nozzle microparticles. Furthermore, the releasing rate dropped from 100% to 45% for the nutrient encapsulate by positively charged nanofibrillated cellulose. The reasons for the slower released are attributed to the charge of the NFC and to the core-shell structure. The content of the positive charges of NFC was determined by conductometric titration of chloride ions with AgNO₃ and found to be 0.119 mmol g^-1 corresponding to a degree of substitution of 0.02 (AGU). For the negative charges, the value of 1.46 mmol g^-1 was obtained by conductometric titration of acid groups with NaOH. The Z-potential of anionic NFC was found to be -60.1 ± 2.2 mV and for the cationic NFC, 36.5 ± 0.8 mV (proofs of positive charges were grafted on NFC). After the encapsulation, the charges of cationic NFC decrease to -8.4 ± 0.8 mV and 6.6 ± 0.6 mV for microspheres and microcapsules, respectively; confirming the interaction between NFC cationic and the KNO₃. As a result, the nutrient is bond with the cationic NFC in the core and also surrounded by the additional layer of NFC (shell) postponing its release. Our approach has showed the great potential of functionalized NFCs and core-shell structure in the preparation of environmentally-friend systems for programmable release fertilizers.

References:
1. Trenkel, M.E. (2010) Slow- and Controlled-Release and Stabilized Fertilizers: An Option for Enhancing Nutrient Use Efficiency in Agriculture, IFA, Paris.
2. Milani et al (2017). Polímeros, 27 (3), 256–266.
3. França et al (2019). Polym. Agri-Food Appl., 29–44.
4. França et al (2018). Carbohydr. Polym., 196, 47–55.
5. Weishaupt et al (2015). Biomacromolecules, 16 (11), 3640–3650.
6. Pei et al (2013). Soft Matter, 9 (6), 2047–2055.

Transparent wood is an emerging material with the combination of optical functions and mechanical performances, showing promising prospect in energy-saving buildings, smart windows, and photonics applications. [1] Transparent wood is usually prepared by removing the light-absorbing components followed by successful impregnation with a refractive-index-matching polymer such as polymethyl methacrylate (PMMA).[2]–[4] During the process, the hierarchical wood structure is preserved. The synergy between wood and PMMA is observed for mechanical properties. Optical properties of TW are tunable via control of the fabrication process and doping the host wood matrix with various additives. Wood nanotechnology is an efficient approach to fabricate multi-functional transparent wood. Through wood-PMMA interface modulation, transparent wood with superior high transmittance and low haze is obtained, paving the way for centimeter thick transparent wood preparation.[5] Wood cell wall modification with organic dye Rhodamine 6G (Rh6G) before PMMA infiltration results in luminescent transparent wood.[6] Lasing effect was observed with the dye modified transparent wood, extending the application of transparent wood in the photonic area. By incorporating functional particles or polymers into the wood structure, hybrid transparent wood materials could be obtained, such as luminescent transparent wood, thermal energy storage transparent wood etc.[7], [8]
[1] Y. Li, E. Vasileva, I. Sychugov, S. Popov, and L. Berglund, “Optically Transparent Wood: Recent Progress, Opportunities, and Challenges,” Adv. Opt. Mater., vol. 6, no. 14, p. 1800059, Jul. 2018.
[2] Y. Li, Q. Fu, S. Yu, M. Yan, and L. Berglund, “Optically Transparent Wood from a Nanoporous Cellulosic Template: Combining Functional and Structural Performance,” Biomacromolecules, vol. 17, no. 4, pp. 1358–1364, Apr. 2016.
[3] M. Zhu, J. Song, T. Li, A. Gong, Y. Wang, J. Dai, Y. Yao, W. Luo, D. Henderson, and L. Hu, “Highly Anisotropic, Highly Transparent Wood Composites,” Adv. Mater., vol. 28, no. 26, pp. 5181–5187, Jul. 2016.
[4] S. Fink, “Transparent Wood - a New Approach in the Functional-Study of Wood Structure,” Holzforschung., vol. 46, no. 5, pp. 403–408, 1992.
[5] Y. Li, X. Yang, Q. Fu, R. Rojas, M. Yan, and L. A. Berglund, “Towards centimeter thick transparent wood through interface manipulation,” J. Mater. Chem. A, vol. 6, no. 3, pp. 1094–1101, 2017.
[6] E. Vasileva, Y. Li, I. Sychugov, M. Mensi, L. Berglund, and S. Popov, “Lasing from Organic Dye Molecules Embedded in Transparent Wood,” Adv. Opt. Mater., vol. 5, no. 10, p. 1700057, 2017.
[7] Y. Li, S. Yu, J. G. C. Veinot, J. Linnros, L. Berglund, and I. Sychugov, “Luminescent Transparent Wood,” Adv. Opt. Mater., vol. 5, no. 1, p. 1600834, Jan. 2017.
[8] C. Montanari, Y. Li, H. Chen, M. Yan, and L. A. Berglund, “Transparent Wood for Thermal Energy Storage and Reversible Optical Transmittance,” ACS Appl. Mater. Interfaces, p. acsami.9b05525, May 2019.

Additive engineering is a well-known effective strategy to obtain high quality perovskite films, which is critical for successful perovskite-based device technology^[1]. Some challenges of unclear mechanism, limited material selection still exist for these bulk-incorporated materials. In addition, the toxicity and cost of the reported additives are often ignored, in fact being one barrier for the future large-scale fabrication.

Here, we introduce an extremely low cost and green biomaterial, Ethyl Cellulose (EC) into the perovskite films. Versatile beneficial effects including trap passivation, crystallization kinetic modulation, balanced charge mobility, hydrogen bonding interaction, scaffold behavior, etc., are surprisingly observed in the EC modified perovskite layers^[2]. Due to these benefits, the resulting solar cells with EC additive exhibit a remarkably enhanced average power conversion efficiency from 17.11% to 19.27% with the increased of all device parameters including J_sc, V_oc and FF. The hysteresis index is found to decrease by 3 times. A pronounced retardation on degradation is also realized where the bare devices with EC retain 80% of their initial efficiencies after 30 days in ambient air under 45% relative humidity, whereas the pristine devices without EC are completely degraded under the same condition.

The work provides a promising way using simple, earth-abundant, but versatile cellulose-based biomaterials as additives to improve both efficiency and stability of the perovskite solar cells.

[1] Q. Jiang, et al. Nat. Photonics 2019, doi.org/10.1038/s41566-019-0398-2
[2] J. Yang , et al. ACS Appl. Mater. Interfaces 2019, 11, 13491

Lignin polymer represents a promising alternative of petroleum-derived polyacrylonitile (PAN) polymer as precursor to synthesize carbon fiber, because of the biorenewability, abundance, low cost, and high carbon content of lignin polymer. Lignin is one of the three main plant cell wall components and a major waste from both paper industry and lignocellulosic biorefinery. Converting lignin waste into carbon fiber has been sought after for several decades with attempt to make cheap carbon fiber and return economics and build sustainability for both biorefining and paper industry. Nevertheless, the poor mechanical performance remains to be a major barrier for commercial applications of lignin-based carbon fiber. We have established that the low mechanical performance was attributed to the chemical heterogeneity of lignin polymer, including diverse interunitary linkages, various functional groups, different molecular weight, and polydispersity. Recent advancements of our group in fractionation technologies showed the great potential to reduce lignin heterogeneity and thus enhance lignin carbon fiber performance. We have developed a series of fractionation technologies to process technical lignin, including 1) partly degradation of lignin using laccase-mediator system and autohydrolysis treatment; 2) solvent fractionation using different organic solvents with different solubility of lignin; 3) physical fractionation by dialysis tube according to lignin molecular weight; 4) lignin precipitation against water with gradient pH values. All these fractionation have trimmed lignin chemistry and reduced lignin heterogeneity by deriving lignin fraction with more β-O-4 linkage, less hydroxyl groups, higher molecular weight, and less polydispersity index (PDI). The resultant carbon fiber was found to have both enhanced mechanical performance and electrical conductivity. Mechanistic study have revealed that the improvement in lignin heterogeneity has improved the molecular interactions of lignin with guest molecules, and thus enhanced the growth of the crystallite structures in carbon fiber, resulting in carbon fibers with improved performances. Overall, fractionation of lignin represents a simple and efficient way to regulate lignin chemistry and boost the quality of lignin carbon fibers, which paved an avenue to transform the sustainability of carbon fiber industry and the bioeconomy of biomass refining.

The reinvention of structural biopolymers into technical materials has enabled the design of biomedical and optoelectronics devices with unique and compelling properties that can serve at the interface between the biotic and the abiotic worlds. For example, we have previously shown the use of silk fibroin bioinks for the detection of E.coli contamination in food. Silk fibroin, in fact, is an edible and biodegradable protein extracted from B. mori cocoons that can be engineered in several formats ranging from films to particles, foams, and gels. By exploiting silk fibroin essential properties (i.e. polymorphism, conformability, and hydrophobicity) it is possible to design a water-based protein suspension that assembles in nano- to centimeter thick materials that can be nanofabricated using advanced manufacturing techniques, including e-beam, soft lithography, replica molding, and printing. The so formed materials modulate mass transport (e.g. O₂ and CO₂ diffusion and water vapor permeability) and preserve the function of biological molecules and micro-organisms outside the cold chain. This is possible by controlling protein polymorphism (i.e. formation of random or beta-sheet structures) during and post material assembly. Here, we show how silk materials can be used to address technological challenges in precision agriculture and food preservation to enhance food security. We will present the design of edible coatings that prolong the shelf-life of perishable food by decreasing water evaporation and regulating O₂ and CO₂ diffusion, effectively reducing oxidative stresses and cell metabolism. The use of silk fibroin coating can also be expanded to seeds to engineer the inoculation of biofertilizers that can mitigate abiotic stress as saline and arid soils. Finally, we will discuss the possibility to use silk-based materials as a vehicle to deliver phytoactive payloads in plants vasculature to modify plant metabolism.

Recent developments in the fabrication of transparent wood (TW) have shown promise in the field on renewable energy. Impregnation of TW with silicon quantum dots (SiQDs) exhibits promise as light-harvesting composite. However, these advancements are hindered by the ability to selectivity control QD distribution and loading within the porous scaffold. Here, we demonstrate that by employing a variety of chemical methods, improvements in QD loading, distribution, and ultimately optical output are drastically improved.

As our understanding of the surroundings and technology advances so does the way we interact with our environment. Serving as food, improving our health, allowing us material extraction or just decorating our constructions, plants have played a vital role in helping humans thrive to adapt and survive. As new challenges arise, there is still room for creative ways to interact with plants. Recent research has focused on interfacing organic electronics with living plants. Although the plant by itself represents a complex system, the idea is to add an extra functionality.
Initially we demonstrated formation of conducting wires along the xylem vascular tissue of rose cuttings from stem to flower. The wires were formed spontaneously, when the cutting was immersed in aqueous solution of a thiophene based conjugated oligomer, ETE-S. ETE-S comprises key features such as water solubility and small molecular weight for plants to transport through its vascular system. The plant guided the polymerization reaction in the xylem acting as a template and catalyst for the polymerization reaction.
In this work we aim to advance the technology by functionalizing intact plants. We choose the bean as our model system and demonstrate electronic functionalization of roots. The ETE-S polymerizes effectively in the root epidermis forming polymer coating of thickness 1-3um. The functionalized roots exhibit conductivity in the order of 10S/cm and volumetric capacity of 50F/cm3. The plants ability to grow its not impaired from the electronic functionalization, making it an attractive system for biohybrid applications from sensing to energy

Conjugated oligoelectrolytes (COEs) are molecules that can spontaneously interact and modify microbial membrane properties. In this work, a three-dimensional COE containing a [2.2]paracyclophane unit (COE2-3-pCp) in a conjugated framework was synthesized to introduce more dimensionality to the core. Its biological activity was determined within the context of membrane perturbation and biocompatibility in lipid vesicles and microbial cells. Additionally, the effects of COE2-3-pCp in a bacterial membrane model were determined in silico using molecular dynamic simulations. The simulations indicate that, compared to its linear analog (COE2-3C), COE2-3-pCp introduces more lipid disorder with higher extent of membrane thinning. In microorganisms, COE2-3-pCp exhibits lower toxicity against E. coli K12 and yeast, while maintaining similar levels of membrane permeabilization. These findings suggest a new design of COEs as biocompatible cell permeabilizers.

Photoelectrochemistry is a relatively new tool for studying photosynthesis, and the knowledge and tools developed from this platform will form the foundation of emerging technologies utilizing photosynthetic materials for energy conversion. The photoelectrochemistry of purified photosystems have recently benefited enormously from an improved understanding of the interfacial wiring and improved electrode design. Here, I give an overview of this progress with a focus on lessons derived from photosystem II, and show how these lessons can be applied to the photoelectrochemistry thylakoids and whole cells. The systematic comparison of the different components, each being one step removed in metabolic complexity, allows the electron transfer processes at each level to be more effectively teased out.


References
1 Kornienko, N., Zhang, J. Z., Sakimoto, K. K., Yang, P., Reisner, E. Nature Nanotech., 2018, 13, 890−899.
2 Kato, M., Zhang, J. Z., Paul, N., Reisner, E. Chem. Soc. Rev., 2014, 43, 6485–6497.
3 Zhang, J. Z., Paul, N., Sokol, K. P., Romero, E., Grondelle, R-V., Reisner, E. Nature Chem. Bio. 2016, 12, 1046–1052.
4 Zhang, J. Z., Bombelli, P., Sokol, K., Fantuzzi, A., Rutherford, A. W., Howe, C., Reisner, E., 2017, J. Am. Chem. Soc., 2018, 140, 6-9.

Bioconjugation is a process useful for giving natural systems addictional artificial properties belonging to ad hoc newly synthetized organic small molecules or other biomolecules. Protein functionalization is usually achieved via click chemistry reactions or amide synthesis, and the partner molecule can be a lipid, or another protein, a nucleic acid, a polysaccharide or an organic molecule. During the bioconjugation processes, cysteine, tyrosine and lysine residues are tipically exploited for anchoring synthetic ligands¹. Moreover, if the protein is an enzyme, the creation of a protein-protein or protein-prosthetic assembly can finely modulate the enzymatic activity.
The photosynthetic reaction center (RC) is an ubiquitous protein. Among all the photosynthetic organisms, the RC obtained from the anoxygenic purple bacterium Rhodobacter sphaeroides is often used as a model system, due to its easy isolation and fully charactherized spectroscopic/electrochemical behaviour². RC is a trans-membrane photoenzyme which efficiently harvests sun-light and converts it in other viable forms of energy by generating charge separated states for fueling cell metabolism.
Biohybrid materials, based on the combination of RC and artificial organic antennas have been already produced to increase the spectral region of the harvested sun light, in order to ameliorate the charge separation aptitude and generation³. Now our research group is focusing on the possibility to covalently link RC with another non photosynthetic protein, the cytochrome c (Cyt C), which is the natural electron donor of these photosynthetic proteins. This is in principle done for ameliorating the electrical communication between RC and Cyt C^4,5. This RC-Cyt C bioconjugation is possible by developing a one-step “grafting-to” protocol exploiting specific lysine and organic aliphatic linkers. In a speculative perspective, we will use these biohybrids to better orient RC on devices for bioelectronic applications.

Biocatalytic oxyfunctionalization of inert C-H or C-C bonds is of special interest in organic synthesis. In particular, unspecific peroxygenases have gained much interests since they catalyze a wide range of selective oxyfunctionalization reactions; however, their broad application is hampered by oxidative degradation of the heme moiety in the presence of even low concentrations of H₂O₂. Typically, this challenge has been met by in situ provision of peroxygenases with suitable amounts of H₂O₂ by catalytic reduction of molecular oxygen using electrochemical^[1], photochemical^[2], or enzymatic^[3] methods. Photoelectrochemical (PEC) reduction of oxygen is an attractive approach to produce H₂O₂ with the use of abundant solar energy and water, avoiding the accumulation of byproducts in the reaction mixture. In this study, we demonstrated a novel, photoelectrochemically boosted peroxygenase catalysis through in situ generation of H₂O₂ via flavin-hybridized carbon single-walled nanotubes (SWNT) photoelectrodes. Flavins can be easily immobilized on the surface of SWNT electrodes via a simple hybridization process while maintaining their redox-active properties of flavins. The hybridization of flavins with SWNTs occurs through π-π interactions between aromatic isoalloxazine moities and graphitic carbons in addition to hydrogen bondings between adjacent flavin molecules. Compared to a pristine SWNT, the flavin-SWNT hybrid photoelectrode exhibited much higher photocurrent density and a marked anodic shift for oxygen reduction reaction, which is attributed to the facilitative effect on charge transfer to oxygen by flavin photosensitizers. As a result, H₂O₂ production rate is enhanced up to 0.15 mM h^-1 at -0.3 V (vs. Ag/AgCl) under illumination, which is 2.28- and 2.48-fold higher than control experiments without light or using a pristine SWNT cathode under light, respectively. We successfully coupled the flavin-SWNT-based PEC platform to peroxygenase-catalyzed reactions, such as ethylbenzene hydroxylation, 2-phenoxypropanoic acid hydroxylation, and indole oxidation.


References
[1] Horst, A. E. W.; Bormann, S.; Meyer, J.; Steinhagen, M.; Ludwig, R.; Drews, A.; Ansorge-Schumacher, M.; Holtmann, D. Electro-enzymatic hydroxylation of ethylbenzene by the evolved unspecific peroxygenase of Agrocybe aegerita. J. Mol. Catal. B: Enzym. 2016,133, S137– S142
[2] Zhang, W.; Fernández-Fueyo, E.; Ni, Y.; van Schie, M.; Gacs, J.; Renirie, R.; Wever, R.; Mutti, F. G.; Rother, D.; Alcalde, M.; Hollmann, F.Selective aerobic oxidation reactions using a combination of photocatalytic water oxidation and enzymatic oxyfunctionalizations. Nat. Catal. 2018, 1, 55– 62
[3] Ni, Y.; Fernández-Fueyo, E.; Baraibar, A. G.; Ullrich, R.; Hofrichter, M.; Yanase, H.; Alcalde, M.; van Berkel, W. J. H.; Hollmann, F. Peroxygenase-catalyzed oxyfunctionalization reactions promoted by the complete oxidation of methanol. Angew. Chem., Int. Ed. 2016, 55, 798– 801

The integration of photosynthetic proteins in biophotoelectrodes is envisioned for the production of electricity^[1] or chemical fuels. Redox hydrogels proved particularly suitable as matrices for the immobilization and electrical contacting of photosynthetic proteins to electrodes. We tuned the redox potentials of the electron relays and the properties of the polymeric supporting matrix to enable benchmark photocurrent densities (300 μA cm^-2 for PS1^[2] and up to 400 μA cm^-2 for PS2 ^[3]) at low overpotential ^[4]. In analogy to the working principle of dye sensitized solar cells, an important feature of biohybrid solar cells for conversion of light to electricity is the charge carriers needed for collection of the high-energy electron from the photosystem ^[5]. The main limitation in energy conversion efficiency is the recombination of this charge carrier at the photoelectrode, a process that decreases both the photocurrent and the open circuit voltage ^[6]. Moreover, this charge recombination process is suspected to induce degradation of the photosynthetic protein ^[7]. We demonstrate that the hydrogel film properties as well as the electrode surface chemistry can be tuned to minimize the various charge recombination pathways.

References:

[1] N. Plumeré. Nature Nanotechnology. 2012, 7, 616-617.
[2] T. Kothe, S. Pöller, F. Zhao, P. Fortgang, M. Rögner, W. Schuhmann, N. Plumeré Chem. Eur. J., 2014, 20, 11029 – 11034.
[3] K. Sokol, D. Mersch, V. Hartmann, J. Z. Zhang, M. M. Nowaczyk, M. Rögner, A. Ruff, W. Schuhmann, N. Plumeré, E. Reisner. Energy Environ. Sci., 2016, 9, 3698-3709.
[4] V. Hartmann, T. Kothe, S. Pöller, E. El-Mohsnawy, M. M. Nowaczyk, N. Plumeré, W. Schuhmann, M. Rögner Phys. Chem. Chem. Phys., 2014, 16, 11936 - 11941.
[5]. M. M. Nowaczyk, N. Plumeré, in Adv Biochem Eng Biotechnol, DOI: 10.1007/10_2016_7, Springer, 2016, “Biophotoelectrochemistry of Photosynthetic Proteins”.
[6] D. Buesen, T. Hoefer, H. Zhang, N. Plumeré. Faraday Discussion 2019, accepted
[7] M. M. Nowaczyk, N. Plumeré. Nature Chemical Biology 2016, 12, 990-991.

Funding: Financial support by the Cluster of Excellence RESOLV (EXC 1069), by the ERC starting grant 715900 and by the ANR-DFG project SHIELDS (PL 746/2-1) is gratefully acknowledged.

Many pathogens, such as Pseudomonas aeruginosa andEscherichia colibacteria can easily form biofilms. The formation of such biofilms in surfaces is known represents a problem for membrane-based water purification techniques, the area of biomedical devices among many others. Aiming to provide a plausible solution to this issue, herein we explore on an antimicrobial peptide (AMP’s) polyethylene glycol (PEG) conjugate that prevents biofouling formation. In this research, the anionic and hydrophobic peptide Maximin H5 (MH5, 2.4kDa), which is an integral part of the defense system of the frog Bombina maxima, is coupled to PEG of different sizes (i.e. 2 kDa and 5 kDa) followed by extensive characterization. Scanning Electron Microscopy (SEM) was employed to evaluate the formation of biofilms and its interaction with these conjugates, showing no biofilm formation. After physical and microbial characterization, MH5 was bio-conjugated to PEG followed by size exclusion purification. The reaction confirmation was achieved by SDS-PAGE and MALDI ToF. Moreover, the antimicrobial activity of the MH5-PEG conjugates was performed via the growth inhibition method and SEM. These conjugates have demonstrated potent antimicrobial activity with high inhibition of bacterial growth after 24 hours. Thus, the prepared polymer-peptide conjugates have the potential to be utilized on applications where an antimicrobial coating is necessary.

As a result of frequent outbreaks occurring due to poor hygiene and improper sanitation of processing environments, there has been an increasing demand for the development of material surfaces that intrinsically inhibit and reduce likelihood of potential microbial adherence and biofilm formation. Herein, we report the synergistic utilization of surface nanotexturing and chemical modifications with nonpolar functional groups on cotton surfaces to produce coatings having bacterial super-repellant characteristics. Using these coatings, the attachment of Staphylococcus aureus as pathogen surrogates was reduced more than 99.0%, compared to the bare cotton surfaces. Moreover, this method is both versatile and scalable, involving inert and biocompatible building blocks. Overall, this study contributes to the field of biomedical applications through the design and development of novel coatings for achieving improved healthcare and hygiene.

Staphylococcus aureus is a common pathogenic bacterium prevalent in a wide variety of diseases, including progressive blood and tissue infections such as chronic ulcers, sepsis and osteomyelitis. Additionally, there exist many S. aureus subtypes which have developed resistance to broad-spectrum antibiotics. Part of the acquired antibiotic resistance may be attributed to the capacity for S. aureus to thrive in colonies within a protective matrix of polymers, polysaccharides, extracellular DNA and water (biofilm). Biofilm residing bacteria are known to be many times more resistant to select antibiotics than their planktonic counterparts.

One mechanism proposed for mediating biofilm-associated infections is to disrupt the polymer structure of the biofilm, leading to the release of planktonic bacteria which are more susceptible to antibiotic therapy. Chemical agents have been evaluated for accomplishing this; however, physical means, such as heat or ultrasound can serve two purposes by first disrupting the biofilm structure while also altering individual bacterial cell walls. Physical disruption may be especially advantageous for eradicating biofilms associated with implanted medical devices, alleviating the need for their removal and maintaining device function.

Silicone is a biocompatible elastomer that can either compose an entire medical device (catheters) or be used as a pliable surface coating on metal or ceramic implants. For the generation of heat, we have included photothermal donor–acceptor conjugated polymer nanoparticles based on poly[4,4-bis(2-ethylhexyl)-cyclopenta[2,1-b;3,4-b’]dithiophene-2,6-diyl-alt-2,1,3-benzoselenadiazole-4,7-diyl] (PCPDTBSe). The composite materials were evaluated for the killing of planktonic and biofilm-residing S. aureus, in either the presence or absence of the antibiotic gentamicin. In the absence of near-infrared laser stimulation of the composite, addition of the nanoparticles exhibited an unexpected bactericidal effect. Stimulation of silicone alone with 612 J/cm2 of 800nm light had no effect on planktonic or biofilm S. aureus, whereas ablative temperatures in the nanoparticle doped composites resulted in a 95-100% reduction in viable bacteria. Sub-ablative temperatures resulted in a 75% reduction in planktonic bacteria treated with gentamicin and a 90% reduction in biofilm-residing S. aureus, compared to controls treated with gentamicin alone and no infrared stimulation. The PCPDTBSe nanocomposites were then evaluated in a murine model to simulate biofilm infections associated with subcutaneously placed silicone implants. A luminescent variant of S. aureus was used to monitor the bacterial burden in the mice before and after laser stimulation with and without antibiotics, and the burden was quantified by measuring colony forming units (CFU) following explantation of the nanocomposite. Depending upon the laser parameters used (1.2-3W/ cm² for 10-45s) there was a reduction of 33-94% CFUs following treatment with the heat-generating nanocomposite plus antibiotics. Our results demonstrate the potential for heat-generating biocompatible nano-composites to augment antibiotics used to treat S. aureus biofilms. These results offer a new avenue for the use of photothermal nanocomposites to eliminate challenging medical infections associated with in-dwelling medical devices.

Photosynthetic organisms are the primary producers of energy on planet Earth thanks to their ability to harvest and convert light into other forms of energy viable for virtually all the metabolic needs of any living organism. Exploiting this formidable attitude is a very intriguing possibility for developing innovative and efficient sustainable energy sources based on the conversion of solar energy.
Garnishing the photosynthetic enzymes [1,3] to produce hybrid systems formed by biological and organic moieties for energy transduction has been shown to be a promising route. On the other side, being able to use the whole, metabolically active, photosynthetic organisms is even more promising [4]. To this purpose, “wiring” the living organism with conductive systems, either organic or inorganic, is of paramount relevance [5] and, yet, has not been fully addressed.
Recently, the encapsulation of living yeasts in polydopamine (PDA)-based structures has been demonstrated [6], with the metabolic activity of the cells preserved in this confinement state because of the lack of any detrimental effect of the polymer.
We present here the preliminary results of the entrapping and confinement of active cells of the photosynthetic bacterium Rhodobacter sphaeroides into ad hoc engineered melanin-like polymers using a mild approach in water based buffers. Bacterial growth accompanied by cellular division are retained under photosynthetic anoxygenic condition, although some detrimental effects are registered in the case of some of the polymers investigated.

This work was funded by European Commission through the EU project 800926 - HyPhOE (Hybrid Electronics based on Photosynthetic Organisms)

[1] F. Milano, R. R. Tangorra, O. Hassan Omar, R. Ragni, A. Operamolla, A. Agostiano, G. M. Farinola, M. Trotta, Ang. Chemie Int. Ed., 51 (44), 11019-11023 (2012).
[2] S. La Gatta, F. Milano, G. M. Farinola, A. Agostiano, M. Di Donato, A. Lapini, P. Foggi, M. Trotta & R. Ragni, Biochem et Biophys Acta (BBA)-Bioen., 1860 (4), 350-359 (2019).
[3] A. Operamolla, R. Ragni, F. Milano, R.R. Tangorra, A. Antonucci, A. Agostiano, M. Trotta, G.M. Farinola, “Garnishing” the photosynthetic bacterial reaction center for bioelectronics, J. Mat. Chem. C, 25(3), 6471-6478 (2015)
[4] F. Milano, A. Punzi, R. Ragni, M. Trotta, and G. M. Farinola, Photosynthetic Bacteria: Photonics and Optoelectronics with Bacteria: Making Materials from Photosynthetic Microorganisms, Adv. Funct. Mater., 29(21), 1-17 (2019).
[5] E. Stavrinidou, R. Gabrielsson, K. P. R. Nilsson, S. K. Singh, J. F. Franco-Gonzalez, A. V. Volkov, M. P. Jonsson, A. Grimoldi, M. Elgland, I. V. Zozoulenko, D. Simon, M. Berggren, In vivo polymerization and manufacturing of wires and supercapacitors in plants, Proceedings of the National Academy of Sciences of the United States of America 114, 2807-2812 (2017).
[6] a). S. Hong, Y. S. Na, S. Choi, I. T. Song, W. Y. Kim & H. Lee, H., Non Covalent Self Assembly and Covalent Polymerization Co Contribute to Polydopamine Formation, Adv. Funct. Mater., 22(22), 4711-4717 (2012); b) D. R. Dreyer, D. J. Miller, B. D. Freeman, D. R. Paul & C. W. Bielawski, Perspectives on poly(dopamine), Chemical Science, 4 (10), 3796-3802 (2013).

Recent developments in the fabrication of transparent wood (TW) have shown promise in the field on renewable energy. Impregnation of TW with silicon quantum dots (SiQDs) exhibits promise as light-harvesting composite. However, these advancements are hindered by the ability to selectivity control QD distribution and loading within the porous scaffold. Here, we demonstrate that by employing a variety of chemical methods, improvements in QD loading, distribution, and ultimately optical output are drastically improved.

Conjugated oligoelectrolytes (COEs) are molecules that can spontaneously interact and modify microbial membrane properties. In this work, a three-dimensional COE containing a [2.2]paracyclophane unit (COE2-3-pCp) in a conjugated framework was synthesized to introduce more dimensionality to the core. Its biological activity was determined within the context of membrane perturbation and biocompatibility in lipid vesicles and microbial cells. Additionally, the effects of COE2-3-pCp in a bacterial membrane model were determined in silico using molecular dynamic simulations. The simulations indicate that, compared to its linear analog (COE2-3C), COE2-3-pCp introduces more lipid disorder with higher extent of membrane thinning. In microorganisms, COE2-3-pCp exhibits lower toxicity against E. coli K12 and yeast, while maintaining similar levels of membrane permeabilization. These findings suggest a new design of COEs as biocompatible cell permeabilizers.

Development of a clean and green renewable energy source without sacrificing a carbon footprint is one of most challenging issues that mankind faces in the modern society. Triboelectricity is a charge separation induced by friction through the physical contact of materials with different electron affinities. Recently, triboelectricity materials and devices have shown a promising pathway to harvest ubiquitous mechanical energy to charge mobile and portable electronic devices. M13 bacteriophage (phage) is a promising material to develop a clean and green triboelectric material. M13 phage is benign and non-harmful virus to infect only bacterial host cells. It has a filamentous shape with 880 nm in length and 6.6 nm in diameter. The M13 phage is covered by 2700 copies of major coat protein that encapsulates phage DNAs. Through the genetic modification, we can precise control the chemical and physical structures of the phage coat proteins. The resulting engineered phage can be easily amplified to produce identical copies of the phage in a large scale. Due to the anisotropic rod-shape, we can self-assembly the phage into various of liquid crystalline structures. Here we developed a novel triboelectric energy harvesting materials and devices using M13 phage. We first created a monolayer thin film of M13 phage and characterized triboelectric properties of the M13 phage. Through direct friction between Ti/Pt coated-AFM tip and M13 phages, we generated the triboelectric charge, and characterized them using Kelvin probe force microscopy (KPFM) measurement. Upon friction generation, we could successfully observe the triboelectric charge generation using the KPFM. We then modified the surface charge through addition of different number of glutamic acids on the M13 phage coat proteins. The phage with higher number of glutamic acids showed a higher triboelectric potential than that of fewer glutamic acids. We also characterized triboelectric charge generation depending on the friction conditions and their discharging behavior. We finally fabricated the large area thin film of M13 phage and developed a triboelectric thin film energy harvesting device. Upon the mechanical force application, the phage-based triboelectric devices can generate 3 V and 5x10-4 mA. The resulting device also could power the organic light emitting diode devices with thin film encapsulation. Our approach to develop a phage-based triboelectric material is useful to understand how friction can induce charges in a biological system in a molecular level and to develop clean and green energy harvesting solutions to charge portable electronic devices and biomedical applications in future.

The essential components of a sensor are receptors, signal converters and detectors. Here we propose an all-in-one bacterial biosensor in the form of supramolecular peptide assembly, which contains all three components by itself. The assembly consists of a double-layered octa-helical tetrameric coiled-coil structure. The outer surface of the assembly shows tetravalent helical bioreceptors for targeting bacteria and the inner core is composed of environment-sensitive fluorophores as detectors. At the connection between them are reversibly self-assembling peptides that play a role in signal transduction. The signal transduction exploits the reversible and non-covalent property of the self-assembled supramolecular system. The multivalent binding of bacteria activates supramolecular transformations, which are transduced into fluorescence changes. Our fluorescent supramolecular biosensors (fSBs) could selectively detect E. coli over other bacteria. Since the fSBs are not denatured by heating, they maintain their capability as a bacterial sensor even at high temperature (50 °C). Based on this research, a variety of responsive supramolecular sensors can be developed to detect diverse biomacromolecules and large biological objects.

By 2100, the population is expected to reach 11.2 billion people, and agriculture has the challenge to establish strategies to achieve sustainable production of food. Fertilizers are used to increase crop production by replacing the necessary nutrient for the plant and also are the major cause of contamination of underground water and rivers. About 30 to 50 % of the fertilizers applied are lost by lixiviation or volatilization, being necessary to use a higher quantity than the real need. The big question is how to increase fertilizer efficiency and crop yield without compromising natural resources? One of the answers is developing a material that controls the availability of fertilizer in the soil, a technology that enhanced efficiency fertilizer, reducing the loss, the amount of fertilizer applied and, consequently, production costs. To achieve this goal is relevant to use polymers from natural resources with wide availability, and potential to biodegrade, to act as a protection layer. Starch polymer fits all these characteristics and has a low cost. For this purpose, the aim of this work was developed microspheres of starch and micronutrients by spray drying technique. For starch gelatinization, 6 wt.%. of food-grade corn starch was added in 100 mL in distilled water and heat in an oil bath at 97 °C during 30 min. After, the gelatinized starch was homogenized in a Turrax for 5 min with 10,000 rpm and during the atomization process kept in magnetic stirring. For starch atomization, we use a 2 - fluid nozzle (0.5 mm diameter) at 130 °C and aspiration rate of 10 % in a Mini spray drying (B290/Buchi) yielding 47 %. The X-ray diffraction pattern of native corn starch exhibited peaks at 15 °, 18 ° and 23 ° (2θ), which indicates A-type starch crystallinity. After the atomization, the microspheres showed an amorphous pattern, which is a consequence of the drying process. The decrease of nutrient release in water demonstrates the potentiality of spray-dried starch microspheres as an enhanced efficiency fertilizer.

The development of microparticles based on natural polymers has attracted great interest for numerous applications, mainly as the release of compounds. It can offer a longer diffusional path, which prolonged release.^1,2 One of the most potential application of these systems of microparticles is in agriculture, where the main issue presented is the loss of fertilizers, like nitrogen (N), phosphorus (P) and potassium (K). Chitosan (CS) is a cationic polysaccharide frequently used for slow/controlled release due to its biodegradability and availability of functional groups (N-H, O-H) on its surface.³ Because of the reactivity of the protonated amino group (NH₃⁺), CS is able to interact with anionic compounds such as phosphate ions. In addition, cellulose nanocrystal (CNC), which is produced from acid hydrolysis of native cellulose fibers, has negatively charged groups due to the insertion of sulfate groups from hydrolysis.⁴ These groups can bind many cationic ions, such as potassium and ammonium. This work focus on the preparation of spray-dried microparticles and the surface interactions between of both polymers (CS and CNC) and the NPK fertilizer as well. First, the CNC was prepared by acid sulfuric hydrolysis (64% wt., 50°C, 120 min) from sugarcane bagasse cellulose. Then, 4 formulations based on 100 mL of Cs (1 wt.%) and two different mass ratios of CNC (0.1 and 1.0 g) and NPK (2.7 and 16.2 g) to CS were dried on a Mini Spray Dryer B-290-BUCH (inlet temperature: 180°C, aspirator: 100%) using two-fluid (flow rate: 7 ml/min). Microparticles were compacted using a hydraulic press (6 ton, 25°C for 1 min) to produce materials in pellet form. All produced materials were characterized using FTIR, XRD, SEM and releasing tests were carried out to evaluate the CNC effect on release property of the composites. XRD confirmed the presence of NPK in the spray-dried microparticles. The interactions between chitosan polymer and ions from fertilizer were confirmed by FTIR results. The non-encapsulated NPK had the fastest release due to high solubility of the fertilizer in water. SEM results showed that CS concentration used in this work was not able to encapsulate the high content of NPK (16.2 g). The fertilizer encapsulated by composites with 0.1 wt.% of CNC showed a slower release profile as compared those with 1.0 wt.% of CNC. This suggests that part of the NPK remains not encapsulated in the presence of CNC high content and can be quickly released from the system. Our work demonstrates one application of CNC and chitosan to bind fertilizers into microparticles that would improve nutrient use efficiency.

Single-cell genomics research has gained significant and rapidly growing interest due to its ability to characterize a population of cells at epigenomic, transcriptomic, or proteomic level with a single-cell resolution that is critical to understanding the genomic heterogeneity between individual cells comprising a population for its potential applications in industry and medicine. The existing single-cell techniques have mainly focused on multicellular, complex organisms such as primates, insects, rodents, and fish; however, understanding the heterogeneity in unicellular microbial organisms is important and will be critical in addressing some of the challenges in environment, medicine, and energy-related applications. This heterogeneity in microbes exists as a response to the ever-changing complex environment. Currently, there are no tools or technologies that allow the quantification of this variation. In the present work, a droplet-based approach is used to transcriptionally profile each microbial species at single-cell resolution and high throughput. Emphasis is laid on the lysis of the microbial cell wall which is one of the key challenges to overcome for the success of the project. A suite of different techniques (both physical and chemical) including the use of microelectromechanical systems [MEMS], laser ablation, acoustic waves, plasmon resonance either used independently or in combination are studied for the lysis of the cell wall. The single-cell lysates are then used as input for high-throughput single-cell RNA-Seq. This work will significantly enhance the current technologies and will enable generalized, cost-effective single-cell RNA sequencing in microbes at high throughput and therefore will be vital to advancing science from different fronts for applications in synthetic biology, environmental engineering, energy, and medicine.

As our understanding of the surroundings and technology advances so does the way we interact with our environment. Serving as food, improving our health, allowing us material extraction or just decorating our constructions, plants have played a vital role in helping humans thrive to adapt and survive. As new challenges arise, there is still room for creative ways to interact with plants. Recent research has focused on interfacing organic electronics with living plants. Although the plant by itself represents a complex system, the idea is to add an extra functionality.
Initially we demonstrated formation of conducting wires along the xylem vascular tissue of rose cuttings from stem to flower. The wires were formed spontaneously, when the cutting was immersed in aqueous solution of a thiophene based conjugated oligomer, ETE-S. ETE-S comprises key features such as water solubility and small molecular weight for plants to transport through its vascular system. The plant guided the polymerization reaction in the xylem acting as a template and catalyst for the polymerization reaction.
In this work we aim to advance the technology by functionalizing intact plants. We choose the bean as our model system and demonstrate electronic functionalization of roots. The ETE-S polymerizes effectively in the root epidermis forming polymer coating of thickness 1-3um. The functionalized roots exhibit conductivity in the order of 10S/cm and volumetric capacity of 50F/cm3. The plants ability to grow its not impaired from the electronic functionalization, making it an attractive system for biohybrid applications from sensing to energy