A key prerequisite to achieve a deeper understanding of microbial communities and to engineer synthetic ones is to identify the individual metabolic interactions among key species and how these interactions are affected by different environmental factors. Deciphering the physiological basis of species–species and species–environment interactions in spatially organized environments requires reductionist approaches using ecologically and functionally relevant species. To this end, we focus here on a defined system to study the metabolic interactions in a spatial context among the plant-beneficial endophytic fungus Serendipita indica, and the soil-dwelling model bacterium Bacillus subtilis. Focusing on the growth dynamics of S. indica under defined conditions, we identified an auxotrophy in this organism for thiamine, which is a key co-factor for essential reactions in the central carbon metabolism. We found that S. indica growth is restored in thiamine-free media, when co-cultured with B. subtilis. The success of this auxotrophic interaction, however, was dependent on the spatial and temporal organization of the system; the beneficial impact of B. subtilis was only visible when its inoculation was separated from that of S. indica either in time or space. These findings describe a key auxotrophic interaction in the soil among organisms that are shown to be important for plant ecosystem functioning, and point to the potential importance of spatial and temporal organization for the success of auxotrophic interactions. These points can be particularly important for engineering of minimal functional synthetic communities as plant seed treatments and for vertical farming under defined conditions.

This paper reports on the use of scanning ion conductance microscopy (SICM) to locally map the ionic properties and charge environment of two live bacterial strains: the Gram-negative Escherichia coli and the Gram-positive Bacillus subtilis. SICM results find heterogeneities across the bacterial surface and significant differences among the Gram-positive and Gram-negative bacteria. The bioelectrical environment of the B. subtilis was found to be considerably more negatively charged compared to E. coli. SICM measurements, fitted to a simplified finite element method (FEM) model, revealed surface charge values of −80 to −140 mC m −2 for the Gram-negative E. coli. The Gram-positive B. subtilis show a much higher conductivity around the cell wall, and surface charge values between −350 and −450 mC m −2 were found using the same simplified model. SICM was also able to detect regions of high negative charge near B. subtilis, not detected in the topographical SICM response and attributed to the extracellular polymeric substance. To further explore how the B. subtilis cell wall structure can influence the SICM current response, a more comprehensive FEM model, accounting for the physical properties of the Gram-positive cell wall, was developed. The new model provides a more realistic description of the cell wall and allows investigation of the relation between its key properties and SICM currents, building foundations to further investigate and improve understanding of the Gram-positive cellular microenvironment.

The last five decades of molecular and systems biology research have provided unprecedented insights into the molecular and genetic basis of many cellular processes. Despite these insights, however, it is arguable that there is still only limited predictive understanding of cell behaviours. In particular, the basis of heterogeneity in single-cell behaviour and the initiation of many different metabolic, transcriptional or mechanical responses to environmental stimuli remain largely unexplained. To go beyond the status quo , the understanding of cell behaviours emerging from molecular genetics must be complemented with physical and physiological ones, focusing on the intracellular and extracellular conditions within and around cells. Here, we argue that such a combination of genetics, physics and physiology can be grounded on a bioelectrical conceptualization of cells. We motivate the reasoning behind such a proposal and describe examples where a bioelectrical view has been shown to, or can, provide predictive biological understanding. In addition, we discuss how this view opens up novel ways to control cell behaviours by electrical and electrochemical means, setting the stage for the emergence of bioelectrical engineering.

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Metabolism is generally considered as a neatly organised system of modular pathways, shaped by evolution under selection for optimal cellular growth. This view falls short of explaining and predicting a number of key observations about the structure and dynamics of metabolism. We highlight these limitations of a pathway-centric view on metabolism and summarise studies suggesting how these could be overcome by viewing metabolism as a thermodynamically and kinetically constrained, dynamical flow system. Such a systems-level, first-principles based view of metabolism can open up new avenues of metabolic engineering and cures for metabolic diseases and allow better insights to a myriad of physiological processes that are ultimately linked to metabolism. Towards further developing this view, we call for a closer interaction among physical and biological disciplines and an increased use of electrochemical and biophysical approaches to interrogate cellular metabolism together with the microenvironment in which it exists.

Microbial communities present the next research frontier. We argue here that understanding and engineering microbial communities requires a holistic view that considers not only species-species, but also species-environment interactions, and feedbacks between ecological and evolutionary dynamics (eco-evo feedbacks). Due this multi-level nature of interactions, we predict that approaches aimed soley at altering specific species populations in a community (through strain enrichment or inhibition), would only have a transient impact, and species-environment and eco-evo feedbacks would eventually drive the microbial community to its original state. We propose a higher-level engineering approach that is based on thermodynamics of microbial growth, and that considers specifically microbial redox biochemistry. Within this approach, the emphasis is on enforcing specific environmental conditions onto the community. These are expected to generate higher-level thermodynamic bounds onto the system, which the community structure and function can then adapt to. We believe that the resulting end-state can be ecologically and evolutionarily stable, mimicking the natural states of complex communities. Toward designing the exact nature of the environmental enforcement, thermodynamics and redox biochemistry can act as coarse-grained principles, while the use of electrodes-as electron providing or accepting redox agents-can provide implementation with spatiotemporal control.

Although the identification of the multigene family encoding mammalian olfactory receptors were identified more than 20 years ago, we are far from understanding olfactory perception because of the difficulties in functional expression of these receptors in heterologous cell systems. Cell-free (CF) or in vitro expression systems offer an elegant alternative route to cell based protein expression, as the functional expression of membrane proteins can be directly achieved from the genetic template without the need of cell cultivation and protein isolation. Here we investigated in detail the cell-free expression and membrane insertion of the olfactory receptor OR5 in dependence of different experimental conditions like probing different origins of the cell-free expression system (from bacteria, via plants and insects toward mammalian system) and lipid composition of the respective extracts. We provided substantial biochemical indications by radioactive labeling based on [(35)S]-methionine, followed by proteolytic digestion, and we found that the insertion of the olfactory receptor OR5 into liposomes resulted in an unidirectional orientation with the binding side exposed into the aqueous space, resembling the native orientation in the cilia of the olfactory neurons. We report the different results in synthesis capacity for the different in vitro systems employed as we like to demonstrate the first in vitro kit toward and ex situ and ex vivo odorant receptor array.

Silica-mineralizing organisms such as diatoms manage several aspects of silica chemistry when polymerizing monomeric silicic acid into amorphous silica. Silicic acid is undersaturated in the diatoms' habitats and mechanisms of enrichment and prevention of uncontrolled mineralization are not well understood. Diatom-biosilica is associated with organic compounds, including polycationic, post-translationally modified peptides termed silaffins, which induce the condensation of silicic acid under supersaturated conditions. Here, we report the pleiotropic action of the designed silaffin-like peptide P5S3, which (i) stabilizes 4-8x silicic acid (in supersaturated conditions of 8.3 mm), (ii) decelerates silica hydrolysis in non-saturated conditions (1 mm) and (iii) enhances silica precipitation at 15-30x supersaturation (30 mm), forming a composite precipitate. Abundant cationic amino acids in P5S3 can interact with silica, though blocking experiments with selected anions also indicate a role of non-electrostatic (e. g. amide-silica) interactions. This shows how silaffin-like peptides could potentially contribute to both keeping silica soluble, and forming peptide-silica precipitates.