Membrane-potential dynamics mediate bacterial electrical signaling at both intra- and intercellular levels. Membrane potential is also central to cellular proliferation. It is unclear whether the cellular response to external electrical stimuli is influenced by the cellular proliferative capacity. A new strategy enabling electrical stimulation of bacteria with simultaneous monitoring of single-cell membrane-potential dynamics would allow bridging this knowledge gap and further extend electrophysiological studies into the field of microbiology. Here we report that an identical electrical stimulus can cause opposite polarization dynamics depending on cellular proliferation capacity. This was demonstrated using two model organisms, namely Bacillus subtilis and Escherichia coli, and by developing an apparatus enabling exogenous electrical stimulation and single-cell time-lapse microscopy. Using this bespoke apparatus, we show that a 2.5-second electrical stimulation causes hyperpolarization in unperturbed cells. Measurements of intracellular K+ and the deletion of the K+ channel suggested that the hyperpolarization response is caused by the K+ efflux through the channel. When cells are preexposed to 400 ± 8 nm wavelength light, the same electrical stimulation depolarizes cells instead of causing hyperpolarization. A mathematical model extended from the FitzHugh–Nagumo neuron model suggested that the opposite response dynamics are due to the shift in resting membrane potential. As predicted by the model, electrical stimulation only induced depolarization when cells are treated with antibiotics, protonophore, or alcohol. Therefore, electrically induced membrane-potential dynamics offer a reliable approach for rapid detection of proliferative bacteria and determination of their sensitivity to antimicrobial agents at the single-cell level.

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Response to mechanical force is a well characterised phenomenon in eukaryotic organisms, helping to organise multicellular structures. Mechanotactic responses have only rarely been observed in prokaryotic taxa. This work reports on a morphological change due to variations in applied force and surface structure by Bacillus mycoides Flügge. B. mycoides is a ubiquitous soil organism well known among microbiologists for its characteristic spreading colony morphology. An apparent mechanotactic response is elicited by physical deformation of the gel media on which B.mycoides is growing, including applied forces of compression or tension. Variations in the surface such as curvature produced by casting the agar gel in the presence of curved objects also elicited the change. The morphological change in B.mycoides colonies associated with the application of force manifests as a pattern of parallel rhizoid filaments perpendicular to compressing force and parallel to stretching force in the agar medium. The phenomenon is most clearly demonstrated by reversible changes in the orientation of B. mycoides filaments during time-lapse microscopy.

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Metabolic interactions within microbial communities are essential for the efficient degradation of complex organic compounds, and underpin natural phenomena driven by microorganisms, such as the recycling of carbon-, nitrogen-, and sulfur-containing molecules. These metabolic interactions ultimately determine the function, activity and stability of the community, and therefore their understanding would be essential to steer processes where microbial communities are involved. This is exploited in the design of microbial fuel cells (MFCs), bioelectrochemical devices that convert the chemical energy present in substrates into electrical energy through the metabolic activity of microorganisms, either single species or communities. In this work, we analyzed the evolution of the microbial community structure in a cascade of MFCs inoculated with an anaerobic microbial community and continuously fed with a complex medium. The analysis of the composition of the anodic communities revealed the establishment of different communities in the anodes of the hydraulically connected MFCs, with a decrease in the abundance of fermentative taxa and a concurrent increase in respiratory taxa along the cascade. The analysis of the metabolites in the anodic suspension showed a metabolic shift between the first and last MFC, confirming the segregation of the anodic communities. Those results suggest a metabolic interaction mechanism between the predominant fermentative bacteria at the first stages of the cascade and the anaerobic respiratory electrogenic population in the latter stages, which is reflected in the observed increase in power output. We show that our experimental system represents an ideal platform for optimization of processes where the degradation of complex substrates is involved, as well as a potential tool for the study of metabolic interactions in complex microbial communities.

11Membrane-potential dynamics mediate bacterial electrical signaling at both intra-and inter-cellular 12 levels. Membrane potential is also central to cellular proliferation. It is unclear whether the cellular 13 response to external electrical stimuli is influenced by the cell's proliferative capacity. A new strategy 14 enabling electrical stimulation of bacteria with simultaneous monitoring of single-cell membrane 15 potential dynamics would allow bridging this knowledge gap and further extend electrophysiological 16 studies into the field of microbiology. Here we report that an identical electrical stimulus can cause 17 opposite polarization dynamics depending on cellular proliferation capacity. This was demonstrated 18using two model organisms, namely B. subtilis and E. coli, and by developing an apparatus enabling 19exogenous electrical stimulation and single-cell time-lapse microscopy. Using this bespoke apparatus, we 20show that a 2.5 sec electrical stimulation causes hyperpolarization in unperturbed cells. Measurements 21 of intracellular K + and the deletion of the K + channel suggested that the hyperpolarization response is 22caused by the K + efflux through the channel. When cells are pre-exposed to UV-violet light, the same 23 electrical stimulation depolarizes cells instead of causing hyperpolarization. A mathematical model 24 extended from the FitzHugh-Nagumo neuron model suggested that the opposite response dynamics are 25 due to the shift in resting membrane potential. As predicted by the model, electrical stimulation only 26induced depolarization when cells are treated with antibiotics, protonophore or alcohol. Therefore, 27electrically induced membrane potential dynamics offer a novel and reliable approach for rapid detection 28 of proliferative bacteria and determination of their sensitivity to antimicrobial agents at the single-cell 29 level. 30 31

Peer review is a widely accepted instrument for raising the quality of science. Peer review limits the enormous unstructured influx of information and the sheer amount of dubious data, which in its absence would plunge science into chaos. In particular, peer review offers the benefit of eliminating papers that suffer from poor craftsmanship or methodological shortcomings, especially in the experimental sciences. However, we believe that peer review is not always appropriate for the evaluation of controversial hypothetical science. We argue that the process of peer review can be prone to bias towards ideas that affirm the prior convictions of reviewers and against innovation and radical new ideas. Innovative hypotheses are thus highly vulnerable to being "filtered out" or made to accord with conventional wisdom by the peer review process. Consequently, having introduced peer review, the Elsevier journal Medical Hypotheses may be unable to continue its tradition as a radical journal allowing discussion of improbable or unconventional ideas. Hence we conclude by asking the publisher to consider re-introducing the system of editorial review to Medical Hypotheses.