The results establish a communication system under controlled laboratory conditions, not yet a field-ready method for regulating microbes in agricultural soils. The researchers first installed the genetic code expansion machinery in B. subtilis PY79, a laboratory model strain. They used an engineered Methanocaldococcus jannaschii tyrosyl-tRNA synthetase and its corresponding suppressor tRNA, together with a fluorescent reporter containing an in-frame amber stop codon.

Without OMY, translation was expected to stop at that codon; with OMY, the reporter should be completed and become fluorescent. Exogenous OMY produced dose-dependent activation, including twofold induction at 1 micromolar and fivefold induction at 10 micromolar, although incomplete termination created substantial background signal. Adding a second amber codon reduced background almost 55-fold and yielded a 3,100-fold difference between conditions with and without OMY.

The number and position of stop codons therefore emerged as an important design variable rather than a universal setting. The system was not limited to a fluorescent marker. The researchers inserted an amber codon into the sfp gene, which encodes the enzyme required to activate the surfactin synthetase machinery.

Without OMY, the engineered B. subtilis strain produced no detectable surfactin; with 500 micromolar OMY, it produced approximately 0.18 grams per litre. OMY itself did not alter bacterial growth under the tested conditions. These experiments demonstrate that a plant-derived chemical can control bacterial protein production and a microbially synthesized compound.

They do not yet show that the platform can improve crop yield, control plant pathogens or safely deploy engineered organisms in an open agricultural environment. The result also remained dependent on the engineered translation machinery and the placement of amber codons in the targeted bacterial genes. On the plant side, the researchers engineered expression of the enzyme MfnG, which enables OMY biosynthesis.

Transient expression in Nicotiana benthamiana leaves produced a metabolite with the expected mass-to-charge ratio that coeluted with an OMY standard; no corresponding signal appeared in control leaves. They then generated stable Arabidopsis lines expressing an sfGFP–MfnG fusion under the constitutive UBQ10 promoter. Hydroponically grown transgenic seedlings released an average of 3.1 micromolar OMY into the surrounding medium, whereas none was detected from wild-type plants.

No optimization of metabolic flux, transport or tyrosine supplementation was required to reach this concentration in the laboratory setup. The finding establishes that a plant can manufacture and release the chemical signal under the tested growth conditions, without demonstrating how much would be available in a mature crop or a field rhizosphere. When the OMY-producing Arabidopsis plants were placed on agar with bacteria carrying an OMY-dependent NanoLuc reporter, luminescence appeared specifically around the transgenic roots.

In mixed plates containing transformed and wild-type seedlings, the signal remained concentrated around the OMY-producing plants, demonstrating genotype-specific activation rather than a general effect of seedling placement. OMY-independent bacteria also generated somewhat stronger signals near roots, which the researchers attributed to higher local bacterial density. The results nevertheless support spatial selectivity: chemical production by one plant could regulate nearby bacterial gene expression while producing a smaller signal around neighboring wild-type plants.

The researchers did not directly measure OMY concentration or bacterial cell density at the roots, so the relative contributions of the chemical gradient and local accumulation of cells remain unresolved. Constitutive OMY production carried a biological cost. Arabidopsis lines overexpressing MfnG had shorter primary roots, reduced lateral-root density, impaired gravitropism, delayed germination and a more rapid decline in seed viability with age.

Lateral-root density was the number of emerged lateral roots normalized by total primary-root length, rather than a direct count of lateral roots. Exogenous OMY inhibited wild-type root growth in a concentration-dependent manner, with a half-maximal inhibitory concentration of approximately 94–101 micromolar. The investigators found no evidence that the effect resulted from depletion of aromatic amino acid precursors or elevated misincorporation of OMY into native proteins.

Aromatic amino acid supplementation did not rescue the phenotype, and proteomics detected no significant increase in O-methylated tyrosine residues. The mechanism of toxicity therefore remained unresolved, although the severity of the measured effects was reduced when OMY production was restricted in space or time. The team next restricted when and where the signal was made.

A root-cap-specific promoter produced OMY in approximately one-fifth of root cells, while an estradiol-inducible system allowed production to be triggered by treatment. Both conditional designs produced less severe growth abnormalities than constitutive expression and brought root growth, lateral-root density, gravitropism and germination close to wild-type values. Root-cap-specific plants released 0.73 micromolar OMY in the tested hydroponic medium and activated the bacterial reporter in cocultivation, although their signal was weaker than that from constitutively expressing plants.

More sensitive flow-cytometry measurements detected reporter induction at OMY concentrations as low as 0.05 and 0.5 micromolar. The experiments therefore showed that spatially restricted production could preserve a detectable response without requiring the larger accumulation measured under constitutive expression. The inducible Arabidopsis system also produced a strong response after activation.

Plants treated with 20 micromolar estradiol generated approximately 18.7 times more maximum bacterial luminescence than uninduced plants. The peak-to-baseline ratio did not differ significantly between treated and untreated groups because the baseline did not decay equally between plant positions, showing why maximum intensity and spatial contrast must be interpreted separately. Collectively, the conditional experiments demonstrate a way to reduce the plant fitness costs while retaining measurable bacterial control.

They do not establish that either promoter would provide useful control in mature crops exposed to natural environmental and microbial variation. In particular, the estradiol system depends on an external chemical trigger, while the root-cap promoter is linked to a specific tissue and developmental context. To move beyond the model laboratory strain, the researchers engineered B. subtilis UD1022, a wild isolate known to colonize Arabidopsis roots.

Its response to OMY was dose-dependent, but its optimal reporter configuration differed from that of PY79: a single amber codon produced the best fold change in UD1022, whereas two codons were most effective in PY79. In autoclaved soil supplied with 500 micromolar OMY, UD1022 reporter fluorescence increased 41-fold by day three. When OMY-producing and wild-type Arabidopsis grew in the same sterilized soil, bacteria exposed to the engineered plants had a median fluorescence of 802 arbitrary fluorescence units, compared with 530 for wild-type plants, and the fluorescent bacterial fraction rose from 9.8% to 36.1%.

This experiment showed transfer of plant-derived OMY in soil, but its sterilized, single-strain setting excluded interactions with a natural microbial community. The platform was also tested in agriculturally relevant plants through a rapid hairy-root transformation method. Composite tomato plants carrying MfnG in transgenic hairy roots and composite poplar plants with the same construct produced luminescence from OMY-dependent B. subtilis reporters specifically at fluorescent-marker-positive roots.

These results show that OMY biosynthesis and bacterial activation can extend beyond Arabidopsis to tomato and poplar, but the shoots in the composite plants remained wild type, and the work did not produce stable, whole-plant transgenic crops. Field use will require testing in nonsterile soils, where OMY may be degraded or transformed by resident organisms, engineered bacteria may face competition, and unintended activation of native organisms must be assessed. The study’s central achievement is therefore a cross-kingdom proof of concept: plants can synthesize a chemical signal that selectively activates engineered bacterial genes, while tissue-specific and inducible production can limit the plant-level costs.

Subject of Research: Genetic code expansion for plant-directed control of engineered soil bacteria

Article Title: Genetic code expansion enables plant-directed control of bacterial activity

Article References: Zhong, V., Jones, M. A., Cabales, A., Gevorgyan, A., Inckemann, R., Johnson, A. A., Karunadasa, S. S., Forti, A. M., Xu, S.-L., Kunjapur, A. M., & Brophy, J. A. N. (2026). Genetic code expansion enables plant-directed control of bacterial activity. Nature Biotechnology. https://doi.org/10.1038/s41587-026-03360-5

Image Credits: AI Generated

DOI: 10.1038/s41587-026-03360-5

Keywords: genetic code expansion, noncanonical amino acids, plant-microbe signaling, synthetic biology, Bacillus subtilis, Arabidopsis, plant biotechnology, soil bacteria, OMY, surfactin, Genetic, code

Anthracnose, driven by fungal species in the genus Colletotrichum, is a persistent problem for sweet orange growers. The disease scars leaves and fruit, depresses yields, and degrades the commercial quality of harvests, and conventional management relies heavily on fungicides whose repeated use raises environmental and resistance concerns. Bacillus subtilis has long been recognized as a biocontrol agent capable of suppressing pathogens directly and of inducing host defense responses, but the host-side transcriptional mechanisms that accompany its protective effect in sweet orange remained poorly defined. The new study set out to close that gap by asking which members of the WRKY transcription factor family respond when the pathogen attacks and how those responses change when B. subtilis is applied as an intervention.

WRKY transcription factors are one of the largest families of plant-specific regulators, named for a highly conserved amino acid motif in their DNA-binding domain, and they are famous for orchestrating responses to stress, including pathogen attack. They bind promoter sequences known as W-boxes, which are enriched in the regulatory regions of many defense-related genes. To map this regulatory landscape in sweet orange, the researchers systematically searched the reference genome DVS_A1.0 and identified 55 CsWRKY family members. They then characterized the structural features of these genes and analyzed how their expression changed across an experimental system that compared healthy controls, plants infected with the anthracnose pathogen, and infected plants that also received B. subtilis treatment.

The phenotypic results were striking in their clarity. When the researchers measured lesion diameters on leaves, the infected plants that had not received the bacterium, designated the BT group, developed lesions averaging 8.72 plus or minus 0.84 millimeters. In contrast, plants in the BST group, which were infected and then treated with B. subtilis, showed lesions averaging 5.47 plus or minus 0.66 millimeters. That difference corresponds to a 37.3 percent reduction in lesion size attributable to the bacterial intervention, a magnitude of protection that is biologically meaningful for a disease that otherwise spreads aggressively through orchards.

Behind that visible symptom reduction lay an extensive transcriptional reprogramming event. Transcriptomic analysis revealed that 28 CsWRKY genes were differentially expressed when infected plants were compared with healthy controls, and 27 were differentially expressed when B. subtilis-treated infected plants were compared with infected plants alone. This near-overlapping set of responsive genes indicates that the WRKY family is broadly engaged during anthracnose infection and that the biocontrol bacterium substantially reshapes that engagement rather than acting on a single isolated pathway. The scale of the response suggests that B. subtilis does not simply kill the pathogen at the leaf surface but actively reorganizes the host’s internal defense signaling network.

To move from a list of responsive genes to a candidate regulator, the team integrated multiple lines of evidence. They examined expression patterns across treatments, constructed a predicted protein-protein interaction network to see which WRKY proteins occupied central positions in the inferred regulatory web, and validated key expression changes using reverse transcription quantitative PCR. This convergence of approaches prioritized CsWRKY40 as a candidate regulatory node. Its expression was downregulated under infection alone but restored and upregulated when B. subtilis was applied, a pattern consistent with a gene whose activity supports the protective state induced by the beneficial bacterium.

Beyond its expression dynamics, CsWRKY40 carries molecular features that hint at how its function might be controlled in the cell. The researchers used the NetPhos 3.1 tool to predict phosphorylation sites across the protein and found multiple potential sites, suggesting that CsWRKY40 could be regulated post-translationally through phosphorylation, a common mechanism by which plant cells rapidly modulate transcription factor activity during immune signaling. While the study stops short of functional validation, the presence of these predicted sites provides a concrete starting point for future experiments aimed at determining how the protein’s activity is switched on or off during the interaction between the pathogen, the bacterium, and the host.

Perhaps the most ambitious component of the study was its genome-wide promoter scan. The researchers searched the entire sweet orange genome for genes containing W-box motifs in their promoter regions and cross-referenced those candidates with the differential expression data from the BT versus CK and BST versus BT comparisons. This analysis identified 5,143 W-box-containing candidate genes that were responsive in at least one of the treatment contrasts. The team then integrated Gene Ontology, KEGG and KO orthology, NR protein database, Swiss-Prot, and protein-domain annotations to classify these candidates functionally. Of the total, 1,736 were classified as defense-associated candidates, falling into categories that include reactive oxygen species and redox homeostasis, general defense responses, hormone signaling, MAPK signaling cascades, cell wall remodeling, secondary metabolism, and pathogenesis-related functions.

That functional breadth is significant because it maps the potential downstream territory that CsWRKY40 and other responsive WRKY factors might govern. Reactive oxygen species bursts are among the earliest and most consequential events in plant immune signaling, and maintaining redox balance determines whether that burst is protective or damaging to host tissue. Hormone pathways, particularly those involving salicylic acid and jasmonic acid, cross-talk extensively to shape defense outcomes against different classes of pathogens. MAPK cascades relay signals from pathogen perception to transcriptional responses, and cell wall remodeling determines the physical barriers a pathogen must breach. Secondary metabolism supplies antimicrobial compounds, and pathogenesis-related proteins are canonical markers of induced defense. The fact that so many W-box-containing genes across these categories shifted expression in response to infection and to B. subtilis treatment suggests that the bacterium’s protective effect operates through a coordinated rewiring of multiple defense modules rather than a single linear pathway.

The authors are careful to frame CsWRKY40 as a candidate rather than a confirmed regulator, and that distinction matters. Correlation between expression patterns and phenotypic outcomes, even when supported by network predictions and qPCR validation, does not establish causation. Functional validation, whether through gene knockout, overexpression, or complementation in stable transgenic lines, will be needed to determine whether CsWRKY40 genuinely drives the protective response or simply correlates with it. Nevertheless, the study provides a well-defined starting point for such experiments and contributes a curated resource of 55 CsWRKY family members, their structural features, and their expression responses to both pathogen challenge and biocontrol intervention. For citrus growers and plant pathologists, the work adds molecular texture to a strategy that has long been used empirically in the field, and it points toward a future where beneficial bacteria might be deployed not just as living pesticides but as precision tools that engage specific host regulatory circuits to strengthen crop immunity from within.

Subject of Research: Identification of the WRKY transcription factor CsWRKY40 as a candidate regulator of Bacillus subtilis-mediated anthracnose suppression in sweet orange

Article Title: A genome-wide and transcriptome-integrated identification of CsWRKY40 as a candidate regulator associated with B. subtilis-mediated reduction of anthracnose symptoms in sweet orange

Article References: Wang, H., Ding, S., Wang, J., Pan, W., Wan, N., Sai, J., Li, Y., Zhang, L., Yao, W., Xu, R., Zhang, H., & Liu, X. (2026). A genome-wide and transcriptome-integrated identification of CsWRKY40 as a candidate regulator associated with B. subtilis-mediated reduction of anthracnose symptoms in sweet orange. BMC Plant Biology. https://doi.org/10.1186/s12870-026-10069-9

Image Credits: AI Generated

DOI: 10.1186/s12870-026-10069-9

Keywords: sweet orange, Citrus sinensis, anthracnose, Colletotrichum, Bacillus subtilis, WRKY transcription factors, CsWRKY40, biocontrol, transcriptomics, plant immunity, W-box, gene regulation

The research team started from a simple but consequential observation: aging is accompanied by progressive functional decline and morphological defects in cholinergic neurons, the nerve cells that use the neurotransmitter acetylcholine and whose degeneration is a hallmark of disorders such as Alzheimer’s disease. Because the human gut harbors between 10^13 and 10^14 bacteria comprising at least a thousand different species, and because bacterial metabolites can enter the circulation and cross the blood-brain barrier, the so-called microbiota-gut-brain axis has become a major focus of research into both neuroprotection and neurological disease. Lowered biodiversity and dysbiosis of the gut microbiota have been documented in elderly people and in patients with Alzheimer’s and Parkinson’s disease, raising the question of whether specific probiotic bacteria might counteract these changes.

Biofilms were central to the investigation. These are complex three-dimensional structures of adherent, cooperative bacterial communities embedded in a secreted extracellular matrix, and growth within them helps bacteria survive hostile environments. Previous work in rodent and C. elegans models had identified biofilm proficiency as a crucial determinant of both pro- and anti-neurodegenerative actions of gut bacteria. In particular, neuroprotective effects of probiotic B. subtilis strains have been linked to biofilm formation in the gut and the release of biofilm-derived metabolic and signaling factors. Some B. subtilis strains also secrete proteolytic enzymes, including the fibrin-degrading enzyme nattokinase first isolated from the Japanese food natto, which can degrade amyloid fibrils characteristic of Alzheimer’s disease.

To find the most promising candidate, the researchers screened several B. subtilis strains in vitro for biofilm production and fibrinolytic activity. The proprietary strain B4U 50, deposited at the German Collection of Microorganisms and Cell Cultures as DSM 34350, was grown for 24 hours at 37 degrees Celsius in a simulated-colonic-environment medium in 96-well plates, after which attached biofilms were stained with crystal violet and quantified spectrophotometrically. B4U 50 developed extensive biofilms under these sessile growth conditions, clearly outperforming the widely used laboratory strain NCIB3610, which showed only modest biofilm formation, while the strains 168 and JH642 produced none. On fibrin agar plates, B4U 50 also displayed the highest fibrinolytic activity of the four strains tested, generating the largest transparent hydrolysis halos, whereas strain 168 showed no detectable fibrinolytic activity at all. The biofilm- and protease-deficient strain 168 therefore served as the control in all subsequent feeding experiments.

The experimental animal was the transgenic nematode strain LX929, in which cholinergic neurons are labeled with green fluorescent protein, allowing researchers to watch neuronal aging unfold in living animals. C. elegans offers distinct advantages for this kind of work: the worms nourish on bacteria, some ingested bacteria survive to become part of their gut flora, key longevity and stress-response pathways are evolutionarily conserved between worm and human, the nervous system contains only 302 well-characterized neurons, and the animals’ transparency and short lifespan permit rapid, longitudinal evaluation of dietary interventions. In aging nematodes, cholinergic neurons undergo progressive damage without cell loss, a process that mirrors the progressive dendritic, axonal and synaptic degeneration of cholinergic neurons seen in aging humans.

Synchronized worms were fed from their first larval stage for up to 15 days on either B4U 50 or the control strain 168, and their neurons were imaged by fluorescence microscopy on days 3, 7, 11 and 15. Three blinded researchers classified the neurons into three categories: normal neurons showed a continuous, uniform GFP signal along the ventral nerve cord; mildly damaged neurons displayed scattered minor swellings or knots without breaks in fluorescence; and severely damaged neurons showed multiple large swellings or visible ruptures in the nerve cord. From these categories the team calculated a neurodegeneration index, a composite score obtained by weighting normal, mildly damaged and severely damaged neurons with values of 0, 1 and 2 respectively.

The results were striking. Neuronal damage increased progressively with age in all groups, but the decline was markedly more pronounced in worms kept on the control strain: by days 11 and 15 these animals showed a steep drop in the proportion of normal neurons and a higher incidence of both mild and severe defects. Worms continuously fed B4U 50 retained a greater proportion of morphologically intact neurons throughout the observation period, and their neurodegeneration index remained significantly lower. The team also ran dietary switch experiments, transferring worms initially raised on strain 168 to B4U 50 on day 4, day 8 or day 12. Switching early, on day 4, produced a neuroprotective effect comparable to continuous feeding with B4U 50; switching on day 8 still conferred a moderate benefit; but switching on day 12 no longer protected the neurons. This defines a window of early and middle adulthood for optimal dietary intervention.

The researchers also tested whether the bacterial diet could be combined with a known neuroprotective compound. They used TEBONIN intens, a standardized extract prepared from Ginkgo biloba leaves, which has been reported to enhance cognitive performance in patients with mild and moderate dementia and to extend lifespan and alleviate amyloid-beta-induced pathology in C. elegans models. Supplementation with the Ginkgo biloba preparation was neuroprotective in worms fed on either B. subtilis strain, reducing both the occurrence and the timing of mild and severe neuronal defects. Notably, the combination of B4U 50 and the Ginkgo biloba extract yielded the highest proportion of morphologically intact neurons by day 15 and the lowest neurodegeneration index values of any group, while worms on the control strain without the extract fared worst. Representative fluorescence images of day-15 animals made the difference visible: worms receiving the combined intervention displayed largely intact GFP-labeled ventral nerve cords, whereas those fed only the control strain exhibited pronounced swellings and discontinuities.

Why might a biofilm-forming bacterium protect nerve cells? The authors point to a proposed model in which B. subtilis spores ingested with food survive gastric transit, germinate in the intestine, and grow in biofilms that produce and secrete substances spreading from the gut throughout the body, exerting anti-aging and neuroprotective effects. Previous studies have identified the quorum-sensing pentapeptide known as competence sporulation-stimulating factor and the gasotransmitter nitric oxide as key mediators of the benefits of B. subtilis biofilms, and probiotic biofilm-producing strains have been shown to counteract neurodegeneration in wild-type worms as well as in transgenic models of Alzheimer’s and Parkinson’s disease expressing human amyloid-beta and alpha-synuclein. Metabolic remodeling during biofilm development is complex, affecting cellular respiration, lipid metabolism and the production of secondary metabolites and signaling molecules.

The authors are careful about the limits of their work. They did not examine whether B4U 50 actually forms a biofilm inside the worm gut, nor did they identify the specific signaling or metabolic factors mediating the neuroprotective effect, and they note this as a limitation of the study. It is also worth noting that the research was funded by Evonik Operations GmbH, the company that maintains the proprietary B4U 50 strain, and that three of the co-authors are Evonik employees, though the other authors report no competing interests. Nevertheless, the study extends a promising line of research suggesting that B. subtilis strains with strong biofilm proficiency, whether laboratory strains like NCIB3610 or probiotic isolates from East Asian fermented foods, can extend lifespan and protect the nervous system in C. elegans. If the findings translate beyond the nematode, combining biofilm-proficient probiotic bacteria with established natural products such as standardized Ginkgo biloba extract could yield dietary supplements with enhanced neuroprotective capacity for the prevention and amelioration of neurodegenerative disorders.

Subject of Research: Neuroprotective effects of a biofilm-forming Bacillus subtilis probiotic strain on aging cholinergic neurons in C. elegans

Article Title: A Bacillus subtilis strain with high proficiency for biofilm formation delays the onset of aging-associated morphological defects in cholinergic neurons of C. elegans

Article References: Gong, W., Priebs, J., Ohse, V. A., Bremges, A., Dieck, H. T., Klotz, L.-O., Dargatz, M., & Steinbrenner, H. (2026). A Bacillus subtilis strain with high proficiency for biofilm formation delays the onset of aging-associated morphological defects in cholinergic neurons of C. elegans. GeroScience. https://doi.org/10.1007/s11357-026-02594-0

Image Credits: AI Generated

DOI: 10.1007/s11357-026-02594-0

Keywords: Bacillus subtilis, C. elegans, biofilm, cholinergic neurons, neurodegeneration, probiotics, Ginkgo biloba, microbiota-gut-brain axis, aging, nattokinase, neuroprotection, GeroScience

The 2011 milestone, when Kralj and colleagues imaged electrical spiking in E. coli at roughly one hertz, was field-shaping in two ways. First, the signal itself: electrical spikes in bacteria invited comparisons to neuronal action potentials, even though the underlying mechanisms and spatial scales differ profoundly. Second, the hypothesis space: the spikes coincided with rapid efflux of a small-molecule fluorophore, hinting that electrical state might couple directly to membrane transport machinery and potentially influence drug susceptibility or stress responses. That observation also seeded the field’s central controversy, which persists today: are membrane-potential transients a cause of downstream bacterial behavior, or merely an aftereffect of fluctuating metabolic conditions?

Conceptually, the review distinguishes three related but mechanistically distinct domains that are frequently conflated in the literature. Transmembrane voltage dynamics encompass fluctuations and excitability-like transients in individual cells. Ionic signaling in communities involves extracellular potassium waves that depolarize neighboring cells across millimeter scales. Electrochemical interactions at interfaces include electrode-based stimulation and impedance phenotyping. Bacterial membrane potential typically lies within a roughly seventy-millivolt range between minus seventy-five and minus one hundred forty millivolts, with strong dependence on growth medium, growth phase, and species. In neuroscience, excitability requires a stereotyped, amplified response to a threshold stimulus followed by a refractory period, and whether bacteria strictly fulfill this definition remains debated, which is why the authors deliberately use the term excitability-like.

Measurement remains the field’s principal bottleneck and driver of controversy. Many classical voltage probes and calibration assumptions were developed for eukaryotic membranes and must be re-validated for bacterial envelopes and growth conditions. Commonly used Nernstian indicators such as thioflavin T, DiBAC, and TMRM tend to report steady-state potential rather than fast transients, and their signals can be confounded by membrane permeability changes, efflux pump activity, and other variations in membrane physiology independent of voltage. Even at low concentrations, these dyes can influence bacterial growth. Genetically encoded PROPS-like indicators avoid some of these pitfalls but require genomic modification, limiting clinical applicability. Recent synthetic reviews now explicitly call for bacterial-specific standardization, including careful reporting of medium composition, growth phase, probe concentration, illumination dose, and validation by multiple experimental approaches.

Non-optical approaches play a crucial complementary role, corroborating optical results while avoiding potential biases such as phototoxicity. Krasnopeeva and colleagues developed an electric-circuit analogue linking the proton motive force, membrane electrical properties, and catabolism, using the bacterial flagellar motor as a rapid single-cell readout under dynamic stresses. Their approach could separate stressors that directly change membrane potential, as an ionophore would, from those that cause physical membrane damage. Notably, they found that shorter-wavelength light can induce electrophysiological impairment, reinforcing the need for rigorous wavelength and dose controls in photobiology-driven experiments. Other physical voltmeters have achieved millisecond resolution in membrane-potential variations, and a 2025 study pushed toward absolute membrane-potential measurements in Bacillus subtilis using fluorescence lifetime, showing that a single strain can vary between minus sixty-five and minus one hundred twenty-seven millivolts when moving from rich to minimal medium.

The second inflection point came from biofilm studies in B. subtilis, where electrical and metabolic dynamics were shown to be coupled at community scale. In 2015, Liu and colleagues described long-range metabolic codependence between peripheral and interior biofilm cells as a mechanism resolving a growth-protection conflict, producing collective oscillations. The same year, Prindle and colleagues demonstrated that ion channels can conduct long-range electrical signals through spatially propagating potassium waves: a metabolic trigger induces potassium release, the increased extracellular potassium depolarizes neighboring cells, and the depolarization wave coordinates metabolic states across the biofilm. These findings extended beyond a single community, with two oscillating biofilms shown to synchronize through electrical signaling, an ecological phenomenon described as nutrient time-sharing.

Theory has matured in parallel. A percolation-based framework linked signal propagation to community heterogeneity and spatial organization, providing a quantitative language for when long-range transmission becomes possible and how near-critical organization can facilitate it. A complementary spatial reaction-diffusion model explicitly combined metabolic and electrical components, bridging the metabolism-first and channel-first schools of thought that often appear as competing intuitions. Preliminary coincident results have emerged in Gram-negative bacteria as well: membrane-potential oscillations have been demonstrated in E. coli biofilms, and collective polarization dynamics beyond a critical biofilm size have been observed in Neisseria gonorrhoeae. Electrical signals also reach outward: potassium-channel-mediated signaling generated by a biofilm can attract distant motile cells in a species-independent manner, and voltage transients in E. coli have been shown to induce calcium influx, supporting the idea that bacteria can treat calcium as a second messenger.

The field has now entered a perturb-and-measure phase, shifting from observation to intervention. A membrane-targeted azobenzene called Ziapin2 was shown to photomodulate membrane potential in B. subtilis, positioning photopharmacology-like strategies as practical tools for bacterial electrophysiology without genetic manipulation. Structured multiscale silicon has been developed as a nongenetic optical transducer capable of modulating single bacterial cells and biofilms at high spatiotemporal resolution. Electrical platforms are advancing too: identical electrical stimuli can produce opposite polarization dynamics depending on the proliferative capacity of the cells, emphasizing that bacterial electrical responses are state-dependent. At the translational end, a drug-free bioelectronic strategy exploited the selective excitability of Staphylococcus epidermidis and demonstrated reduced colonization on a porcine skin model using an electroceutical patch, while electrical impedance spectroscopy revealed that E. coli biofilms exhibit stable negative capacitances under small DC bias, implicating a specific voltage-gated potassium channel.

Among the more provocative post-2015 directions is the possibility that bacterial communities encode persistent electrical states. Transient optical perturbations have been shown to generate a long-lasting potassium-channel-mediated change in membrane potential within a biofilm, such that exposed cells subsequently responded out of phase relative to unexposed cells during oscillations, interpreted as a type of membrane-potential-based memory within a microbial community. This behavior is consistent with bistable dynamics within the metabolic-electrical network, and for photobiology it opens a methodological opportunity: optical perturbation can write states, optical readout can track them, and spatially structured illumination can test whether writing is local, propagative, or dependent on community connectivity.

Looking forward, the review identifies several frontiers. Nongenetic optical modulators point toward bacterial all-optical electrophysiology experiments where stimulation patterns are as controllable as in modern optogenetics. Supplementing cultures with tricarboxylic acid cycle intermediates such as glutamate, fumarate, or pyruvate can boost the proton motive force, amplify membrane potential, and increase the uptake of aminoglycoside antibiotics, suggesting a relevant clinical framework that might work synergistically with chemical phototransducers like Ziapin2. Standardized interrogation protocols pairing defined stimuli with time-resolved readouts could generate reproducible response fingerprints of bacterial physiological state, enabling diagnostics for antibiotic tolerance, persister formation, and dormancy. The relation between membrane potential and antibiotic tolerance still lacks consensus on how hyperpolarization versus depolarization affects antibiotic efficacy, requiring extensive investigation across bacterium-stimulus-antibiotic combinations. Whether bacterial spikes ultimately earn the name action potentials or demand bacteria-native descriptors anchored in metabolism and transport physiology, the next fifteen years promise to transform light-driven tools into causal instruments for reading, writing, and programmatically controlling the electrical lives of microbes.

Subject of Research: bacterial bioelectricity and membrane potential dynamics

Article Title: Bacterial bioelectricity, the first 15 years: developments, tools, and remaining questions

Article References: Cianflone, E., Keller, H. R., Subramaniam, N., & Paternò, G. M. (2026). Bacterial bioelectricity, the first 15 years: developments, tools, and remaining questions. Frontiers in Photobiology, 4, Article 1834373. https://doi.org/10.3389/fphbi.2026.1834373

Image Credits: AI Generated

DOI: 10.3389/fphbi.2026.1834373

Keywords: bacterial bioelectricity, membrane potential, electrophysiology, biofilms, potassium waves, voltage imaging, optogenetics, photobiology, antimicrobial control, ion channels, Bacillus subtilis, Escherichia coli

The work builds on a well-known fact of microbial life: microorganisms in nature rarely live alone. In the human gut, the Human Microbiome Project identified roughly 3,000 bacterial species coexisting in a single ecological niche, and their stability depends on constant chemical conversation. When antibiotics or dietary shifts disturb this balance, cross-feeding structures and nutrient competition shift with them, sometimes with severe health consequences. Understanding and eventually engineering such communication is therefore not just an academic curiosity — it is a prerequisite for designing stable synthetic microbial communities for industry, agriculture, and medicine.

Previous efforts had already pushed the boundaries of artificial cross-species signaling. In 2013, researchers showed that Escherichia coli could be made to talk to Bacillus megaterium using quorum-sensing peptides borrowed from Staphylococcus aureus. More recently, the Dresden group demonstrated communication in the opposite direction: yeast engineered to produce bacterial quorum-sensing peptides CSF and PhrF successfully activated reporter responses in B. subtilis. What remained missing was the reverse channel — bacteria signaling to yeast — and that is precisely what the new study delivers.

The engineering strategy exploited one of the most thoroughly characterized signaling systems in eukaryotic biology: the yeast mating pheromone pathway. Haploid Saccharomyces cerevisiae cells exist as one of two mating types, a or α, and each secretes a small peptide pheromone that binds to a specific receptor on cells of the opposite type. Receptor activation triggers a signaling cascade that induces morphological changes, cell-cycle arrest, and the expression of mating-specific genes. Among these, the FIG1 gene is switched on roughly 100-fold within just 20 minutes of receptor activation — a speed and dynamic range that makes it an ideal handle for synthetic biologists.

To turn B. subtilis into a yeast-language speaker, the team built on a signal peptide toolbox they had developed previously. The codon-optimized α-factor gene was inserted into an evaluation vector, preceded by the secretion signal peptide of the bacterial protein YfkD to route the peptide through B. subtilis’s native Sec secretion system. Expression was placed under the control of P_liaI, a promoter inducible by the antibiotic bacitracin. The final construct was integrated into the amyE locus of the protease-deficient strain WB800N, yielding the producer strain TMB6428. When induced with bacitracin, this strain secreted α-pheromone at concentrations of up to 8 nanomolar — modest by industrial standards, but consistent and robust, and sufficient for the task at hand.

On the receiving end, the researchers used a haploid yeast strain carrying deletions in BAR1 and FAR1. BAR1 encodes an aspartyl protease that degrades and inactivates α-factor, so its removal prevents the secreted signal from being destroyed before it can act. FAR1 encodes a cyclin-dependent kinase inhibitor responsible for the G1 growth arrest that normally follows pheromone exposure; deleting it keeps the sensor cells alive and dividing rather than frozen in mating mode. Into this background, the team integrated a single genomic copy of the fluorescent protein mNeongreen under control of the α-inducible FIG1 promoter. Genomic integration was necessary because B. subtilis cannot grow properly in yeast minimal medium, which would have made plasmid selection by auxotrophy impractical. The choice of mNeongreen over standard GFP was deliberate: originally discovered in the lancelet Branchiostoma lanceolatum, mNeongreen is the brightest green fluorescent protein known, resists photobleaching better than GFP, and matures faster — all advantages when the expected signal is weak.

Getting the two organisms to grow together posed its own challenges. B. subtilis doubles roughly every 30 minutes, while haploid yeast needs about 90 minutes in rich medium, so the team had to balance starting densities carefully. They settled on a ratio of approximately 1:6, with yeast at an OD600 of 0.1 and bacteria at 0.6 — a bias toward the bacterial producer that compensates for the fact that the engineered strain secretes only a single copy of α-factor, whereas yeast’s native MFα gene encodes four tandem copies processed into four separate peptides. The cultivation medium also required compromise: standard LB broth supplemented with 2% glucose satisfied the nutritional needs of both organisms at a shared incubation temperature of 31.5 °C.

The proof of principle came when mid-exponential co-cultures were induced with bacitracin. Within two hours, Western blot analysis detected mNeongreen protein in co-cultures containing the α-factor-producing bacteria, and the signal intensified at four and twenty-two hours as pheromone accumulated. Flow cytometry of 20,000 cells per sample confirmed statistically significant fluorescence in induced co-cultures, while negative controls — uninduced cultures and co-cultures with wild-type B. subtilis W168, which cannot produce α-factor — showed no detectable signal. The positive control, yeast treated with 250 nanomolar synthetic α-factor, produced brighter fluorescence than the co-culture, consistent with the bacterial secretion rate being lower than the bolus concentration used artificially. Under physiological conditions, 100 nanomolar synthetic α-factor is sufficient to induce mating in BAR1-deleted yeast, so the bacterial output, though modest, falls within a biologically meaningful range.

Perhaps the most visually striking evidence came from epifluorescence microscopy. Beyond the green glow of mNeongreen, the microscope revealed yeast cells undergoing shmoo formation — the characteristic pear-shaped morphological change that haploid cells adopt in response to authentic mating pheromone. Shmoo formation appeared about four hours after induction and increased over the following eighteen hours, demonstrating that the α-factor secreted by B. subtilis was not merely recognized at the receptor level but was fully functional, capable of driving the complete physiological differentiation program of yeast mating. The bacterial cells, meanwhile, showed no morphological changes, confirming that the signal flowed in one direction only.

The study also revealed a manageable imperfection: the P_liaI promoter exhibits slight leakiness, producing faint fluorescence in uninduced co-cultures after four and twenty-two hours. The researchers attribute this to known intrinsic transition-state induction of the LiaRS system, and note that the yeast-only positive control did not show this time-dependent increase because synthetic α-factor was added only once at the start rather than continuously. Looking ahead, the team envisions a synthetic sensor-actuator circuit in which engineered B. subtilis cells detect environmental signals — antibiotics, for example — and respond by secreting yeast pheromones that trigger the production of degradation enzymes in yeast. Since engineered S. cerevisiae capable of degrading β-lactam antibiotics in response to α-pheromone has already been described, the two pieces of such a self-regulating loop already exist. With accumulation of degradation end products potentially serving as a built-in negative feedback to shut the circuit down, the prospect of autonomous, self-contained microbial consortia — communicating across kingdoms to clean up pollutants or produce chemicals sustainably — has moved measurably closer to reality.

Subject of Research: Engineered interkingdom pheromone signaling between Bacillus subtilis and Saccharomyces cerevisiae

Article Title: Interkingdom communication 2.0: pheromone-based control between Bacillus subtilis and Saccharomyces cerevisiae

Article References: Döring, J., Vološen, T., Ostermann, K., & Wolf, D. (2026). Interkingdom communication 2.0: pheromone-based control between Bacillus subtilis and Saccharomyces cerevisiae. Applied Microbiology and Biotechnology, 110(1), Article 310. https://doi.org/10.1007/s00253-026-14069-5

Image Credits: AI Generated

DOI: 10.1007/s00253-026-14069-5

Keywords: interkingdom communication, Bacillus subtilis, Saccharomyces cerevisiae, pheromone signaling, synthetic biology, quorum sensing, mNeongreen, co-culture, FIG1 promoter, microbial consortia, biotechnology, signal transduction

The research team, led by Sylvie Skalickova and Pavel Horky, set out to test whether two commercially available and fast-growing bacteria could unlock the nutritional potential locked inside milling waste. They chose Lactiplantibacillus plantarum, a homofermentative lactic acid bacterium equipped with enzymes capable of degrading cellulose-rich polysaccharides, and Bacillus subtilis, a sporulating species whose enzymatic repertoire includes proteases, amylases and lipases. Both organisms can also produce secondary metabolites such as organic acids, phenolic compounds and flavonoids, which could enrich the fermented material beyond simple nutrient release. Prior work on fermentation-based biofertilizers had focused mostly on rice bran, fruit pomace, olive waste and other substrates rich in readily digestible nutrients, leaving heterogeneous grain milling waste largely unexplored.

In the first stage of the experiment, the researchers optimized the fermentation process in vitro. Substrates of wheat bran, feed-grade wheat flour and mill dust were mixed in three ratios, submerged in water, pre-treated with xylanase at 45 degrees Celsius for 24 hours, and then inoculated with either bacterial species at two different doses. Cultivation proceeded aerobically at 37 degrees Celsius under mild agitation for 48 hours. The team then measured markers of microbial activity, including glucose, lactic acid, total polyphenols and total protein, alongside the bioavailability of phosphorus and micronutrients in both the liquid supernatant and the solid pellet.

The results were striking. Fermented milling waste showed a two-fold increase in total protein concentration and a 6.5-fold increase in lactic acid concentration compared with the unfermented control. Free phosphorus availability doubled, and soluble mineral ions rose significantly across the board: calcium increased by 3 to 9 milligrams per liter, iron by 4.5 to 9.9 milligrams per liter, magnesium by 7.5 to 13 milligrams per liter, manganese by 13 to 15 milligrams per liter and zinc by 8.5 to 9.3 milligrams per liter. Importantly, these effects were not strictly dose-dependent, and the outcome depended on both the bacterial strain and the substrate composition. Most microelements accumulated in the liquid fraction, while calcium concentrated in the pellet, a pattern the authors attribute in part to microbial acidification of the medium, which promotes the release of mineral ions from polysaccharide complexes.

The substrate ratios mattered too. Bacillus subtilis fermentation of the bran-heavy 6:3:1 mixture produced a sharp rise in lactic acid alongside a steep drop in free glucose, whereas L. plantarum continued to consume glucose without being inhibited by rising acidity. The authors note that the elevated acid readings in the Bacillus treatments may reflect other organic acids interfering with the lactic acid assay rather than lactic acid itself. Fermentation also significantly increased total polyphenol content in all treatments, consistent with microbial hydrolysis of cell-wall components releasing bound phenolic compounds. Based on the combined in vitro data, the team selected L. plantarum and the 6:2:2 substrate ratio for the next stage, because this combination maximized the inclusion of hard-to-use mill dust while delivering strong mineral solubilization and high lactic acid production, which can also help protect plants against pathogens.

With the fermented material, or FMW, in hand, the researchers moved to a greenhouse pot experiment with maize (Zea mays). FMW was applied in three ways: incorporated directly into the soil, sprayed onto leaves as a foliar treatment, or poured onto the soil as a drench. Each route was benchmarked against mineral NPK fertilizer, compost, vermicompost and an unfertilized control, with fertilizer doses converted from an intensive field rate of 250 kilograms of nitrogen per hectare. Over three 14-day sampling intervals, the team tracked aboveground biomass, leaf chlorophyll as a proxy for nitrogen status, soil mineral nitrogen, soil pH and the concentrations of macro- and micronutrients in both soil and plant tissue.

The growth results revealed a clear hierarchy. NPK fertilizer produced the highest and statistically most significant biomass at every sampling date, as expected from its instantly soluble nutrients. Direct soil incorporation of FMW, by contrast, yielded the lowest biomass, between 9 and 25 percent below the unfertilized control, likely because the carbon-rich substrate stimulated rapid microbial proliferation that temporarily immobilized nitrogen needed by young plants. Yet the liquid application routes told a different story. Foliar FMW produced the relatively highest dry weight gains in the first two sampling terms, at 12 and 9 percent above control, while soil drenching with FMW delivered a 21 percent biomass advantage by the final harvest. The authors suggest that foliar and drench applications bypass soil microbial competition altogether, delivering amino acids and bioactive compounds directly through the leaf cuticle or the root zone.

The soil chemistry data added further nuance. Among the organic treatments, FMW consistently provided the highest soil mineral nitrogen, reaching 22.2 milligrams per kilogram in the final sampling compared with 12.95 for the control, 13.37 for compost and 13.23 for vermicompost. FMW also produced the highest ammonium nitrogen fractions among the organic treatments in every term, a form of nitrogen that persists longer in soil and serves as a reserve against leaching. Correlations between soil mineral nitrogen and leaf chlorophyll were strong to very strong, with Pearson coefficients ranging from 0.79 to 0.94. Soil drenching with FMW also produced the highest soil pH values at every sampling, a buffering effect the authors link to basic cations such as calcium and magnesium displacing exchangeable hydrogen and aluminum ions, in welcome contrast to the acidifying tendency of mineral fertilizers.

Plant tissue analysis underscored the biostimulant potential of the fermented waste. FMW applied to soil or as a drench produced the second-highest concentrations of phosphorus, calcium, iron and copper after NPK, and delivered statistically the highest tissue levels of magnesium and zinc of any treatment. Relative to the unfertilized control, FMW soil application or drenching raised plant content of phosphorus by 23 percent, potassium by 50 percent, calcium by 31 percent, sulfur by 36 percent, iron by 40 percent and manganese by a remarkable 86 percent. The authors attribute this micronutrient mobilization to chelation by organic acids and humified compounds released during fermentation, which keep metals soluble and plant-available even in near-neutral soils.

The study’s conclusion is deliberately measured: fermented milling waste is not a replacement for mineral fertilizer, which remains unmatched for short-term yield, but rather a complementary organic amendment with a genuine biostimulant dimension whose value depends heavily on how it is applied. Compared with compost and vermicompost, which take weeks to months to produce, controlled fermentation offers a far faster route from waste to usable product, fitting neatly into circular bioeconomy goals. The authors caution that pot experiments cannot be directly extrapolated to the field, and that environmental and safety questions, including salt accumulation, phytotoxic compounds, pathogen contamination and heavy-metal loading, remain to be assessed. Multi-year field trials will be needed to quantify cumulative effects on soil organic carbon, microbial activity and yield stability. Still, the prospect of turning one of agriculture’s most stubborn waste streams into a liquid amendment that feeds soils, nourishes crops and buffers acidity marks a compelling step toward more sustainable cereal processing.

Subject of Research: Microbial fermentation of grain milling waste as an organic soil amendment and biostimulant for crop nutrition

Article Title: Utilization of grain milling waste for plant growth and nutrient availability in soil

Article References: Skalickova, S., Antosovsky, J., Ridoskova, A., Ryant, P., & Horky, P. (2026). Utilization of grain milling waste for plant growth and nutrient availability in soil. Frontiers in Sustainability, 7, Article 1958097. https://doi.org/10.3389/frsus.2026.1958097

Image Credits: AI Generated

DOI: 10.3389/frsus.2026.1958097

Keywords: grain milling waste, fermentation, Lactiplantibacillus plantarum, Bacillus subtilis, biofertilizer, soil health, plant nutrition, biostimulant, circular economy, wheat bran, organic amendment, maize

The motivation is stark. Nigeria’s Niger Delta, home to the vast majority of the country’s petroleum production, endures an average of roughly 240,000 barrels of spilled crude oil every year, according to figures cited in the study. Decades of chronic spills, oily waste disposal, and industrial activity have degraded farmland, groundwater, and aquatic ecosystems, undermining soil fertility and agricultural productivity. Petroleum hydrocarbons are chemically stable, toxic, and persistent, allowing them to accumulate in soils and sediments and disrupt microbial balance and ecosystem function. Effective, affordable, and sustainable remediation has therefore become an urgent environmental priority for the region.

Nature already possesses the raw machinery for cleanup. Indigenous microorganisms can use hydrocarbons as carbon and energy sources, gradually detoxifying polluted ground. But natural attenuation is routinely throttled by nutrient limitation, particularly shortages of nitrogen and phosphorus, by the low bioavailability of hydrophobic hydrocarbons, and by the sheer structural complexity of crude oil, which blends aliphatic, aromatic, resin, and asphaltene fractions. Incomplete degradation can even leave behind intermediate metabolites more toxic than the parent compounds. Bioremediation strategies attempt to overcome these constraints: biostimulation feeds the native microbial community with added nutrients, while bioaugmentation introduces specialized degraders. The new study combined both approaches in a single experimental design.

The team, led by Senyene Umana of Akwa Ibom State University with colleagues from the University of Uyo, first characterized the biostimulant itself. The brewer’s spent grain proved remarkably nutritious for microbes: 76.0 percent organic carbon, 27.5 percent protein, 2.33 percent nitrogen, and a calorific value of about 399 kilocalories. Its protein-derived nitrogen addresses the classic bottleneck in petroleum-contaminated soils, where microbes starve for nitrogen even while drowning in carbon. The grain’s carbohydrate fraction can serve as a co-substrate for cometabolic degradation, and its fibrous matrix may improve soil structure, moisture retention, aeration, and microbial attachment. Notably, the spent grain contained only trace hydrocarbons of its own, meaning its principal contribution was nutrition rather than added contamination.

For the degrader, the researchers turned to Eniong River sediment in Akwa Ibom State. After enrichment cultures in mineral salt medium with crude oil as the sole carbon source, one isolate stood out for its vigorous growth on hydrocarbons. Molecular identification by 16S rRNA gene sequencing, deposited in GenBank under accession number PZ556056, revealed a 99.38 percent match to Bacillus subtilis. Bacillus species are prized in bioremediation for producing extracellular enzymes and biosurfactants that break down and solubilize hydrophobic compounds, and the isolate showed no lag phase before consuming oil, suggesting prior adaptation or constitutive expression of degradation enzymes.

The experimental setup was elegantly simple. Six-kilogram portions of sandy loam topsoil were placed in perforated wooden boxes and spiked with Bonny Light crude oil to reach a measured initial total petroleum hydrocarbon concentration of 2784.01 milligrams per kilogram, a low-to-moderate pollution scenario by Niger Delta field standards. Treatments received the standardized Bacillus suspension at approximately 1.5 times ten to the eighth cells per milliliter, plus brewer’s spent grain at 1, 10, or 20 percent by weight. Untreated contaminated soil and soil inoculated with bacteria alone served as controls. Over 80 days at ambient tropical temperature, the team tracked microbial populations every ten days and measured residual hydrocarbons by gas chromatography at the end.

The headline result was a dose-response surprise. Bacillus alone degraded 49.93 percent of the hydrocarbons, a respectable demonstration of intrinsic capability. But the combined treatments shattered that benchmark: 88.58 percent removal with 1 percent spent grain, 76.85 percent with 10 percent, and 68.61 percent with 20 percent. Counterintuitively, the smallest nutrient dose delivered the biggest cleanup. The 10 percent treatment did produce the highest total heterotrophic bacterial abundance, peaking at 8.82 log10 colony-forming units per gram, roughly a 1.5-log increase over the unamended control. Yet abundance of the specialized oil-degrading bacteria told a different story, climbing most sustainably in the 1 percent treatment, where they rose about 2.5 logs, while collapsing to undetectable levels by day 60 in the 10 percent soil.

The explanation lies in microbial economics. Heavy organic loading supplied readily biodegradable substrates that let non-hydrocarbon-utilizing heterotrophs proliferate, shifting metabolism away from petroleum and toward the easy carbon in the amendment. Intensified competition for nutrients, oxygen, and ecological niches then squeezed out the specialized degraders, and rapid decomposition of the amendment may have raised biological oxygen demand, creating microenvironments hostile to aerobic hydrocarbon oxidation. Nitrifying bacteria felt the strain too: Nitrosomonas and Nitrobacter populations declined sharply in the controls and were undetected in the higher spent grain treatments, though the 1 percent amendment preserved comparatively better nitrifier abundance, suggesting moderate supplementation supports broader microbial functionality. The study’s authors are candid about limitations, noting that without an abiotic control, the reported efficiencies represent apparent hydrocarbon removal rather than proven biological degradation alone, since volatilization, adsorption, and photooxidation could contribute.

The chromatographic fingerprints added technical depth. Degradation preferentially targeted low- and mid-molecular-weight n-alkanes from C10 to C22, which are more bioavailable and structurally simpler than their heavier counterparts, while high-molecular-weight fractions from C23 to C35 also declined progressively in amended soils. Critically, pristane and phytane, isoprenoid compounds widely used as biodegradation indicators because of their recalcitrance, were substantially reduced, consistent with genuine hydrocarbon alteration during remediation. Some individual fractions showed transient increases or newly detectable peaks in treated soils, which the authors attribute cautiously to analytical variability, changes in extractability, or transformation-related processes rather than new petroleum inputs. Meanwhile, the amendment moderated soil chemistry toward remediation-friendly territory: pH shifted from acidic 5.05 in the contaminated baseline to a near-neutral 7.26 to 8.21 range, redox potential climbed from around 20 to as high as 90 millivolts, and moisture content rose from about 1.4 percent to nearly 16 percent, all within ranges favorable for hydrocarbon-degrading microbes.

The broader implications are compelling. Brewer’s spent grain is generated in enormous quantities by the brewing industry worldwide, and in beer-producing regions like Nigeria it is abundant, cheap, and locally available, sidestepping the transport and cost barriers of commercial fertilizers. The study demonstrates that synergy between nutrient-driven stimulation, functional degraders, and optimized amendment rates can dramatically outperform either strategy alone, while also warning that more is not better: excessive organic loading can backfire by feeding the wrong microbes. The authors acknowledge that field-scale application will need to grapple with seasonal availability, compositional variation among breweries, and handling requirements to prevent secondary microbial growth or odor. They also note that ecological recovery cannot be inferred without plant-based toxicity assays. Still, as a proof of concept under tropical conditions, the work offers a persuasive vision: turning one industry’s waste into another industry’s cleanup tool, and letting biology do the heavy lifting where shovels and chemicals have too often failed.

Subject of Research: Combined biostimulation with brewer's spent grain and Bacillus bioaugmentation for crude oil biodegradation in contaminated soil

Article Title: Combined effects of brewer’s spent grain biostimulation and Bacillus bioaugmentation on crude oil biodegradation in contaminated soil

Article References: Umana, S., Nkanang, A., Udosen, C., & Eduok, S. (2026). Combined effects of brewer’s spent grain biostimulation and Bacillus bioaugmentation on crude oil biodegradation in contaminated soil. BMC Environmental Science, 3(1), Article 25. https://doi.org/10.1186/s44329-026-00066-6

Image Credits: AI Generated

DOI: 10.1186/s44329-026-00066-6

Keywords: bioremediation, brewer's spent grain, biostimulation, bioaugmentation, Bacillus subtilis, petroleum hydrocarbons, soil remediation, Niger Delta, hydrocarbon degradation, total petroleum hydrocarbons, oil-degrading bacteria, agro-industrial waste

The enzyme in question is pyrimidine-nucleoside phosphorylase, or PyNP, from the bacterium Bacillus subtilis. PyNP participates in the production of nucleic-acid building blocks by cleaving and reassembling nucleosides, the sugar-and-base units that form the letters of DNA and RNA. Because the same reaction chemistry can be redirected to process synthetic nucleoside analogues, the enzyme is a workhorse in the design of anticancer, antiviral, and antibacterial drugs, and it is also used to manufacture artificial nucleosides, fluorescently labeled nucleosides, and stable-isotope-labeled nucleic acids for research and diagnostics. Despite decades of practical use, the molecular basis of how PyNP’s changing shape governs molecular recognition has remained unclear, leaving a gap between what chemists observe at the bench and what is actually happening at atomic resolution inside the active site.

Addressing that gap, a research team led by Professor Akihiko Hatano, together with researcher Kousei Takeshima of the Course of Chemistry and Biotechnology, Department of Materials Science and Chemistry, College of Engineering at Shibaura Institute of Technology, set out to test how different conformations of the enzyme influence ligand retention and sugar preference. Their strategy was to treat the enzyme not as one structure but as a family of structures. Using all-atom molecular dynamics simulations, the team sampled four distinct conformational states of PyNP, probed the active site with 13 structural probes, and accumulated molecular dynamics trajectories totaling 15.6 microseconds of simulation time. The findings were published online in the journal ACS Omega on September 12, 2026.

The motivation, Professor Hatano explains, grew out of years of bench work. His group has long used this enzyme to synthesize artificial nucleosides, and a recurring question kept surfacing: why does changing the sugar component of a substrate alter the reaction? Experiments could document the outcomes, but they could not directly expose the atomic-scale motions responsible. “We wanted to understand this question from the perspective of atomic-scale motions that experiments alone cannot reveal,” said Prof. Hatano. “This led us to examine how the enzyme and its bound molecules behave across different structural states.” In other words, the team wanted to watch the enzyme and its cargo in real time at the level of individual atoms, rather than infer behavior from static crystal structures.

The first result concerns the geometry of the active-site pocket itself. As the protein transitions between its most closed and most open conformational states, the pocket that cradles the ligand expands by approximately 1.4-fold. That is a substantial change for a recognition site whose job is to grip molecules precisely. A pocket that breathes to this degree cannot be characterized by a single volume or a single set of contact distances; the microenvironment experienced by a bound nucleoside depends on which conformational state the enzyme happens to occupy at any moment. This observation alone suggests that docking studies and structure-based drug design built on one static structure may systematically misjudge how ligands behave in the real, dynamic enzyme.

The simulations then connected this breathing motion directly to function. Across the trajectories, 75.6 percent of those in the closed group retained their ligands in a bound state, compared with only 55.1 percent in the open group. The difference is striking: simply shifting the enzyme’s conformation changed the odds that a bound molecule would stay in place by roughly twenty percentage points. Ligand retention, in this view, is not a fixed property of the enzyme-ligand pair but a joint product of molecular identity and enzyme shape. A compound that appears loosely bound in an open structure might be firmly anchored in a closed one, and vice versa, which has immediate implications for how inhibitors targeting PyNP should be evaluated.

The study also resolved the sugar-preference puzzle. Ribose- and 2′-deoxyribose-containing compounds, it turns out, do not show a universal preference for one sugar over the other. Instead, sugar preference depended on both the individual compound and the enzyme’s conformation, meaning there is no single rule of thumb that predicts which sugar PyNP will handle better. The simulations did, however, reveal a consistent geometric signature: ribose occupied the North, or C3′-endo, ring conformation for 64.3 percent of the sampled time, whereas 2′-deoxyribose occupied the South, or C2′-endo, conformation for 57.9 percent. This conformational divergence was strongest for the unsubstituted compound and became progressively smaller as the substituent at the 6-position of the base grew larger, indicating that bulky base modifications can partially override the intrinsic tendencies of the two sugars.

A single amino acid residue emerged as a key player in this dynamic picture. Tyrosine 165, or Tyr165, behaves as a moving lid over the active site, an effect that was particularly pronounced in the closed reference structure. To test whether this lid genuinely contributes to ligand capture, the researchers ran additional simulations in which the residue was removed. Ligand retention dropped in those simulations, supporting the proposed lid mechanism at the computational level. The authors are careful to note a caveat: the strongest Tyr165 signal came mainly from a closed structure belonging to a related species, so confirmation using a closed-state structure from B. subtilis itself is still needed before the lid mechanism can be considered fully established for this organism.

The broader implications reach well beyond one bacterial enzyme. Because PyNP is already used to synthesize fluorescently labeled nucleosides and stable-isotope-labeled nucleic acids, understanding how enzyme shape affects molecular capture could help researchers design more efficient enzymatic synthesis routes and select reaction conditions that favor the desired conformational behavior. The same understanding could guide the design of inhibitors that exploit the closed state or the lid residue, a strategy relevant to anticancer and antiviral drug development. There is also a sustainability angle: enzymatic synthesis performed in water at mild temperatures is an inherently greener manufacturing route than many conventional chemical syntheses, and improving its efficiency through conformation-aware enzyme engineering could amplify that advantage.

“Enzymes should not be viewed as single, static structures when we think about molecular recognition,” said Prof. Hatano. “Our results show that different conformations can change how compounds are retained and how sugar-related preferences emerge.” The study points toward future calculations and experiments aimed at connecting these dynamic binding behaviors to the actual chemical steps of the phosphorylase reaction, a link that remains to be established. What the work already demonstrates, however, is methodological as much as mechanistic: examining multiple enzyme conformations with molecular simulations can reveal binding behavior that any single static structure may miss. By showing that ligand retention and sugar preference depend jointly on molecular identity and enzyme shape, the research provides a framework for dynamic substrate recognition and opens concrete paths toward future enzyme engineering, rational inhibitor design, and sustainable biocatalytic synthesis of the nucleoside analogues on which modern medicine and biological research increasingly depend.

Subject of Research: Conformation-dependent ligand retention and sugar recognition in pyrimidine-nucleoside phosphorylase from Bacillus subtilis studied by molecular dynamics simulations

Article Title: Molecular simulations reveal how an enzyme’s shape guides molecular recognition

Article References: Molecular simulations reveal how an enzyme’s shape guides molecular recognition. (n.d.). Original publication

Image Credits: AI Generated

DOI: Not provided

Keywords: pyrimidine-nucleoside phosphorylase, molecular dynamics simulations, Bacillus subtilis, molecular recognition, ligand retention, ribose, 2'-deoxyribose, active site, conformational states, enzyme engineering, nucleoside synthesis, ACS Omega

The team began where microbiology has always begun: with morphology and biochemistry. All three representative isolates proved to be Gram-positive, catalase-positive and Voges-Proskauer-positive, a classic Bacillus signature that reflects a thick peptidoglycan cell wall, the ability to decompose hydrogen peroxide, and the capacity to ferment sugars into acetoin and related neutral products. Yet the tests were not merely confirmatory. The Methyl Red test, which detects stable mixed-acid fermentation, separated AMU-K-1 from AMU-UP-1 and AMU-MY-1, revealing genuine metabolic heterogeneity among bacteria that looked superficially alike. This kind of phenotypic split matters for anyone hoping to deploy soil bacilli in agriculture, because fermentation end products such as acetoin and 2,3-butanediol are directly relevant to industrial bioprocesses and to plant-microbe signaling in the rhizosphere.

Antimicrobial susceptibility testing added a second layer of differentiation. In agar well diffusion assays, the isolates produced clear inhibition zones against tetracycline, amoxicillin and azithromycin, indicating susceptibility to three clinically important antibiotic classes. But when the researchers moved to broth microdilution, a quantitative method that determines minimum inhibitory concentrations rather than relying on visible zones, they found substantial variability in resistance profiles between isolates. The discrepancy is a useful caution for the field: diffusion-based assays can be influenced by diffusion rates, growth conditions and inoculum density, and the authors’ decision to pair them with microdilution follows current best practice in antimicrobial susceptibility testing. For soil-derived bacteria intended as plant growth promoters or biocontrol agents, antibiotic resistance profiles are not an academic detail, since resistance genes carried by beneficial strains can potentially spread to pathogens in the environment.

The molecular core of the study rests on the 16S rRNA gene, the workhorse of bacterial taxonomy. The researchers amplified roughly 1,500-base-pair fragments of the gene and obtained high-quality Sanger sequencing reads of 1,048 bases for AMU-K-1 and 1,102 bases for AMU-UP-1. BLAST searches against public databases identified AMU-K-1 as closely related to Bacillus safensis, a species originally isolated from spacecraft cleanrooms but now recognized in soils worldwide. AMU-UP-1, however, presented a familiar taxonomic headache: its sequence showed 100 percent identity to multiple closely related taxa within the Bacillus subtilis and Bacillus velezensis species complex. The 16S rRNA gene, for all its utility, simply evolves too slowly in this clade to separate species that diverged recently, and the authors were candid that unambiguous species-level resolution was not possible from this gene alone.

To place the isolates in an evolutionary framework, the team reconstructed phylogenetic trees using the Tamura 3-parameter model with invariant sites, a substitution model designed for sequences with strong transition-transversion bias and skewed guanine-cytosine content. The resulting trees confirmed that both isolates fall within well-supported Bacillus lineages. Nucleotide substitution analysis across more than 1,300 aligned positions revealed a pronounced excess of transitions over transversions, meaning that purine-to-purine and pyrimidine-to-pyrimidine changes dominate the mutational landscape of these ribosomal sequences. That pattern is expected in conserved RNA-coding genes, where structural constraints penalize the more disruptive transversions, but quantifying it provides a baseline for future comparative work on Indian soil bacilli and reinforces the choice of the T92+I model for this dataset.

The most distinctive element of the study is its thermodynamic treatment of the 16S rRNA molecule itself. Using RNA secondary structure prediction tools, the researchers computed minimum free energy structures for the sequenced regions. AMU-K-1 yielded a minimum free energy of minus 530.80 kilocalories per mole, while AMU-UP-1 yielded minus 523.80 kilocalories per mole, both indicating highly stable ribosomal conformations. More intriguing was the ensemble diversity, a measure of how many alternative folding states a sequence can adopt at equilibrium: AMU-K-1 scored 357.82 against 151.53 for AMU-UP-1, suggesting that the ribosomal RNA of the B. safensis-like isolate explores a much broader conformational space in silico. The authors are appropriately careful here, noting that whether such computational differences translate into physiological adaptability requires direct experimental verification. Still, the approach hints at a future in which the folding energetics of housekeeping genes become part of the standard characterization toolkit.

Why does this matter beyond taxonomy? Bacillus species are among the most commercially important bacteria in agriculture. Bacillus velezensis in particular has emerged as a model plant-associated beneficial bacterium, producing lipopeptides that suppress fungal pathogens and colonizing roots in ways that enhance nutrient uptake. Bacillus safensis and related strains have been associated with drought tolerance in crops such as sugarcane, and Bacillus-based inoculants are central to the push for sustainable crop protection. Accurate identification is the precondition for all of these applications, because a mislabeled strain can carry unwanted traits, including antibiotic resistance determinants or pathogenicity factors, into field deployments. The Indian isolates, deposited in NCBI GenBank under accession numbers PZ619164 and PZ619172, now form a verified reference resource for such efforts.

The study also sits within a broader methodological debate. Full-length 16S rRNA sequencing, including nanopore-based approaches, has been shown to achieve species-level resolution in some bacterial groups, and whole-genome comparisons are increasingly regarded as the gold standard for distinguishing members of the B. subtilis complex. By combining phenotypic, biochemical, phylogenetic and thermodynamic evidence, the authors demonstrate a pragmatic middle path for laboratories without routine access to genome sequencing: extract the maximum information from a single locus, quantify its evolutionary dynamics, and treat computational predictions as hypotheses to be tested rather than conclusions. The explicit caveat that in silico ensemble diversity does not equal physiological flexibility is a model of how such integrative papers should be written.

For India specifically, the work contributes to a growing catalog of regionally characterized soil bacilli, joining earlier studies of Bacillus subtilis and Priestia megaterium from eastern Indian farmlands and Bacillus licheniformis and Bacillus cereus from sorghum rhizospheres. Soil biodiversity is under pressure from land-use change and a warming climate, and documenting native microbial diversity before it erodes has become a quiet urgency in environmental microbiology. Each verified isolate with published sequences, biochemical profiles and susceptibility data is a permanent, reusable asset for bioprospecting, whether the target is a biocontrol agent, a bioremediation workhorse or an industrial enzyme producer.

The eight isolates characterized here are unlikely to be the last word on Indian soil Bacillus, and the authors themselves frame the study as a foundation for future exploitation in plant growth promotion, biological control, environmental bioremediation and microbial biotechnology. What the paper ultimately delivers is a template: a demonstration that old-fashioned petri dish biochemistry, when fused with careful sequencing, model-based phylogenetics and even RNA folding thermodynamics, can still generate new knowledge about the organisms beneath our feet. As the search for sustainable alternatives to chemical fertilizers and pesticides accelerates, the humble soil isolate, rigorously identified and honestly annotated, remains one of biotechnology’s most valuable raw materials.

Subject of Research: Integrated phenotypic, molecular, phylogenetic and thermodynamic characterization of soil Bacillus isolates from India

Article Title: Integrated morpho-biochemical, molecular, phylogenetic, and thermodynamic characterization of Bacillus isolates from ecologically distinct Indian soils

Article References: Mohanty, A., Sahoo, J. P., Mohapatra, A., Samal, P. P., Sahoo, L., Mahapatra, S. S., & Hossain, S. N. (2026). Integrated morpho-biochemical, molecular, phylogenetic, and thermodynamic characterization of Bacillus isolates from ecologically distinct Indian soils. Molecular Biology Reports, 53(1), Article 1692. https://doi.org/10.1007/s11033-026-12911-7

Image Credits: AI Generated

DOI: 10.1007/s11033-026-12911-7

Keywords: Bacillus, soil microbiology, 16S rRNA, phylogenetics, Bacillus subtilis, Bacillus velezensis, Bacillus safensis, antimicrobial susceptibility, RNA secondary structure, plant growth promotion, India, Sanger sequencing

The central obstacle the team set out to address is a familiar one in cereal science. When bran is milled into flour, its sharp, fibrous particles physically interrupt the continuous protein films that gluten proteins normally form during mixing and resting. The result is a weakened network with fewer cohesive junctions, less elasticity, and poorer gas retention. In products like Chinese steamed buns, or mantou, where a light, springy, uniformly porous crumb is the benchmark of quality, that weakness translates directly into lower specific volume, higher hardness, and a shorter shelf appeal. Whole-wheat versions of these staples routinely underperform their refined-flour counterparts on nearly every textural metric.

Fermentation has been used for millennia to modify dough properties, but the choice of microorganism matters enormously. The Beijing group, led by Maosi Fan and corresponding authors Yingli Liu and Jing Wang, applied four distinct strains of Bacillus subtilis to whole-wheat dough and then subjected both the dough and the finished steamed buns to an unusually comprehensive battery of measurements. Rheological testing, intermolecular force analysis, microscopy, pasting behavior assays, texture profiling, and volatile metabolomics were all brought to bear on the question of what, precisely, the bacteria were doing to the flour matrix during fermentation.

The rheological results were among the clearest signals. Fermented dough showed reduced apparent viscosity alongside increased creep recovery, a combination that describes a dough which is easier to deform under stress but better able to spring back afterward. In practical terms, that is exactly the behavior a baker or steamed-bun producer wants: a mass that handles well during shaping, resists collapse, and recovers its structure as gas cells expand. The finding suggests that the bacterial fermentation did not simply soften the dough by degrading it, but rather reorganized the underlying network into something more pliable and more resilient at the same time.

To understand how that reorganization happened, the researchers examined the forces holding the protein network together. Protein fractions associated with hydrogen bonding and disulfide bonds increased after fermentation, while fractions held together by ionic and hydrophobic interactions decreased. This shift matters because hydrogen bonds and disulfide bridges are the connective tissue of a well-ordered gluten network. Disulfide bonds covalently link glutenin polymers into the long chains that give dough its elasticity, while dense hydrogen bonding between proteins and between proteins and starch granules provides the cohesive, continuous matrix that traps fermentation gases. A network rebuilt around these stronger, more cooperative interactions is structurally better suited to holding the open, even crumb that defines a good steamed bun.

Spectroscopic and microscopic evidence supported that interpretation. Fourier-transform infrared analysis indicated changes consistent with re-established hydrogen bonding and a higher degree of short-range order in the starch fraction, while microscopy revealed a more continuous protein phase threading through the dough. In an unfermented whole-wheat system, bran particles tend to sit as discontinuities in the protein matrix, creating weak points where cracks can initiate. After B. subtilis fermentation, the protein phase appeared to reconnect around and between those particles, effectively healing the damage that bran incorporation inflicts. The starch order findings are notable as well, since greater short-range molecular order in starch is generally associated with slower retrogradation and a crumb that stays softer for longer.

The textural consequences showed up clearly in the finished steamed buns. Fermented buns exhibited higher specific volume and higher porosity, meaning they rose taller and trapped more, and more evenly distributed, gas. At the same time, their hardness was reduced and the crumb structure was softer. Those are not trivial gains. Specific volume and crumb softness are the two attributes that most reliably separate a premium steamed bun from a disappointing one, and both are notoriously difficult to preserve when refined flour is replaced with whole wheat. The fact that a single biological intervention improved both at once is the study’s most commercially significant result.

Perhaps the most surprising dimension of the work is its effect on aroma. Whole-wheat products often carry off-flavors traced to lipid oxidation products and to compounds derived from lignin in the bran itself, and these odors are a major reason consumers cite for avoiding whole-grain foods. Using volatile metabolomics and relative odor activity values, the researchers found that fermented buns contained lower levels of these lipid-oxidation- and lignin-derived off-odorants, alongside elevated levels of aromatic alcohols, compounds generally associated with pleasant, yeasty, fermented fragrance profiles. In other words, the bacteria did not merely mask the undesirable smells; they appear to have shifted the underlying metabolic balance of the dough toward a more favorable volatile signature.

Multivariate analysis across all four strains identified one, designated SH, as the most effective performer, suggesting that strain selection is a critical variable rather than an afterthought. Different B. subtilis isolates produce different suites of enzymes, including proteases, amylases, and lipases, and the precise enzymatic cocktail a strain secretes will determine how aggressively it remodels proteins, starch, and lipids during fermentation. The SH strain’s dominance implies that the optimal outcome, a restructured protein network, ordered starch, softened crumb, and improved aroma, reflects a particular balance of enzymatic activity rather than fermentation intensity alone. That opens a practical path for producers: rather than accepting whatever ambient microbes happen to be present, manufacturers could inoculate dough with a characterized, food-grade strain chosen for these specific effects.

The study, supported by the National Natural Science Foundation of China, stops short of claiming that B. subtilis fermentation solves every challenge of whole-wheat processing, and the published version reports instrumentally measured properties rather than large-scale consumer trials. But the implications reach well beyond steamed buns. Whole-grain consumption is consistently linked to better dietary fiber intake and health outcomes, yet the textural penalties of bran have limited how much of it food manufacturers can incorporate without losing customers. A controlled bacterial fermentation that simultaneously restores gluten continuity, softens the crumb, and cleans up the aroma profile offers a low-cost, non-chemical route to closing that gap. If the strain-specific findings hold up at industrial scale, the humble soil bacterium Bacillus subtilis, already a workhorse of fermented foods across Asia, may become one of the most useful tools in the effort to make whole grains taste as good as they are for you.

Subject of Research: Bacillus subtilis fermentation to improve the structure, texture, and volatile flavor profile of whole-wheat dough and steamed buns

Article Title: Bacillus subtilis fermentation improves the structural, textural and volatile properties of whole-wheat dough and steamed buns

Article References: Fan, M., Lin, S., He, X., Gu, Y., Xu, L., Liu, K., Wei, X., Jiao, Y., Tang, W., Liu, Y., & Wang, J. (2026). Bacillus subtilis fermentation improves the structural, textural and volatile properties of whole-wheat dough and steamed buns. npj Science of Food. https://doi.org/10.1038/s41538-026-01172-0

Image Credits: AI Generated

DOI: 10.1038/s41538-026-01172-0

Keywords: Bacillus subtilis, whole-wheat dough, fermentation, steamed buns, gluten network, food rheology, volatile metabolomics, food texture, bran, starch order, food microbiology, cereal science