Antibiotic resistance is a growing global health threat. Although antibiotic activity is well studied in homogeneous liquid cultures, many infections are caused by spatially structured multicellular populations where consumption of scarce nutrients establishes strong spatial variations in their abundance. These nutrient variations have long been hypothesized to help bacterial populations tolerate antibiotics, since liquid culture studies link antibiotic tolerance to metabolic activity, and thus, local nutrient availability. Here, we test this hypothesis by visualizing cell death in structured E. coli populations exposed to select nutrients and antibiotics. We find that nutrient availability acts as a bottleneck to antibiotic killing, causing death to propagate through the population as a traveling front. By integrating our measurements with biophysical theory and simulations, we establish quantitative principles that explain how collective nutrient consumption can limit the progression of this “death front,” protecting a population from a nominally deadly antibiotic dose. While increasing nutrient supply can overcome this bottleneck, in some cases, excess nutrient unexpectedly promotes the regrowth of resistant cells. Altogether, this work provides a key step toward predicting and controlling antibiotic treatment of spatially structured bacterial populations, yielding biophysical insights into collective behavior and guiding strategies for effective antibiotic stewardship. In this work, authors provide evidence that bacteria in spatially structured populations protect each other from antibiotics through collective nutrient consumption, creating ‘death fronts’ that sweep through the colony– explaining why infections often survive treatments that work in lab tests.

With antimicrobial resistance emerging as a top global health concern, new approaches to prevent and treat infections are urgently needed. Antimicrobial peptides are recognized as a promising alternative to conventional antibiotics. Considering the costly, labor-intensive, and time-consuming nature of wet lab screening, in recent years, proteomics research has increasingly relied on in silico methods to predict physicochemical, structural, and biological properties of candidate peptides. Here, we present the AMPSeek protocol to enable scientists to screen peptides and predict their potential antimicrobial activity, toxicity, and associated three-dimensional structure with a single command. The AMPSeek framework integrates AMPlify, tAMPer, and LocalColabFold, scales efficiently with increasing sample size, and produces an interactive HyperText Markup Language report to facilitate result interpretation. We provide step-by-step instructions for installing and running AMPSeek on a ten-peptide test set, along with two additional datasets (n = 20 and n = 100; mean peptide length ≈121.3) to demonstrate AMPSeek's near-linear scalability.

Intestinal motility is a function of the enteric nervous system involving secretion of the excitatory neurotransmitter acetylcholine (ACh). Although gut commensal bacteria are key regulators of intestinal physiology, the molecular mechanisms underlying microbial influence on intestinal peristalsis and constipation remain unclear. Here we report a link between microbial nitrogen metabolism and intestinal motility regulation via ammonia production. We observed compensatory elevation of intestinal ammonia levels and urease activity in mouse models of intestinal dysmotility, induced by ACh deficiency, and in patients with constipation. Ammonia supplementation or intervention in mice with the urease-positive Lysinibacillus fusiformis isolated from patient stool, or engineered urease-expressing bacteria, effectively restored colonic ACh levels. In vitro, ammonia upregulated the expression of voltage-gated calcium channels on enteric neurons, driving Ca2+ influx to potentiate ACh secretion. Our study reveals a microbial compensatory mechanism that responds to fluctuating ACh levels in the intestine and provides microbial targets for intestinal motility disorders. Intestinal motility defects caused by lower acetylcholine levels in the gut are compensated by bacteria that generate ammonia from urea which facilitates the release of acetylcholine via upregulation of voltage-gated channels on enteric neurons.

Genome editing has revolutionized plant biology research, yet the efficient delivery of editing reagents remains a challenge. Current methods are labour intensive, involving lengthy tissue culture and complex transformation and regeneration steps. Viral delivery can mitigate these issues but CRISPR–Cas nucleases exceed viral cargo limits, restricting guide RNA (gRNA) delivery into Cas9-expressing transgenic plants. This requires generating an initial Cas9 transgenic line. Furthermore, gRNAs delivered by plant viral vectors can induce somatic edits, although only a few produce heritable edits. Some engineered plant negative-strand rhabdoviruses can deliver both Cas9 and gRNA, but they face other challenges, including the need for tissue regeneration or pruning infected plants, and some rhabdoviruses can be delivered only through vector transmission. Recently, smaller editors such as TnpBs were discovered, but they are significantly less active than Cas9. Here we optimized a tobacco rattle virus-based system to deliver recently engineered, highly active ISDra2 TnpB variants. The eTnpBc variant enables effective somatic editing in systemic leaves and achieves up to 90% editing efficiency at target loci. In addition, up to 89% of offspring exhibit a mutant phenotype, with editing efficiencies reaching 100%. The design principles outlined here could promote wider use of eTnpBc for efficient, transformation- and transgene-free plant genome editing. Although genome editing has transformed plant research, delivering editing reagents remains challenging. This study reports the use of viral vectors to deliver engineered eTnpBc, achieving high levels of somatic and heritable editing in plants.

Current therapeutic techniques for cancer often lack specificity. They also cause systemic toxicity and lack genetic control. Thus, cancer ranks among the most complex and crucial global health issues. The novel concept of smart nano-capsules is discussed in this Perspective. These oral medications modify genes using CRISPR technology and integrate biosensing and laser-guided activation to enable more personalized cancer therapies. The creation of these versatile nanocapsules is driven by three objectives. First, they aim to enable controlled gene editing in the gastrointestinal tract. Second, they deliver treatments to specific target areas. Third, they detect tumors in real time. Nanocapsules equipped with biosensing components provide microenvironmental input. An external laser can trigger the release of light-absorbing agents. Moreover, these features reduce off-target effects and allow spatiotemporal precision, the enteric-coated architecture ensures oral stability. Surface functionalization enhances selective tumor accumulation. AI-guided control algorithms can manage diagnostic interpretation and activation. The CRISPR-based cancer medicines offer the potential for improved safety, specificity, and translational use in the future. Combining advanced nanotechnology, gene editing, and AI-guided control could create innovative solutions.

Phages are a valuable resource for the genetic engineering of Streptomyces antibiotic-producing bacteria. Indeed, a few integrative vectors based on phage integrase are available to insert transgenes at specific genomic loci. Chromosome conformation captures previously demonstrated that the Streptomyces linear chromosome is organized in two spatial compartments: the central compartment encompassing the most conserved and highly expressed genes in exponential phase, and the terminal compartments enriched in poorly conserved sequences including specialized metabolite biosynthetic gene clusters. This study introduces a new integrative tool based on a recently described phage, Samy, which specifically targets the terminal compartment of its native host chromosome. Samy is related to PhiC31 phage and, like the latter, encodes a serine integrase. Whereas PhiC31 targets a site generally located near the origin of replication, the Samy integration site is one of the farthest known attB sites from it. We demonstrated that the Samy integrase efficiently mediates the specific integration of a non-replicating plasmid in six Streptomyces strains from distinct clades. Bioinformatic analyses revealed that the Samy-attB site is rather conserved and located in the terminal compartment of most Streptomyces chromosomes. Finally, heterologous expression of the albonoursin biosynthetic gene cluster from the Samy-, PhiC31-, and R4-attB sites yields quantitatively equivalent levels of production, though qualitative differences were observed. Altogether, these results demonstrate that the att-int Samy system expands Streptomyces genetic engineering tools by enabling targeted integration in the terminal chromosomal compartment.

Intracellular bacterial pathogens can cause high levels of morbidity and mortality in humans. Host immune responses that protect against these infections include pyroptosis, a form of lytic cell death caused by the insertion of large gasdermin (GSDM) pores into the host plasma membrane. Here we review recent advances in our understanding of how the five GSDM proteins, GSDMA–E, are activated by distinct signalling pathways. Pyroptosis can both eliminate intracellular niches and release cytosolic interleukin-1 family cytokines that further prime host immune responses against the invading pathogen. Because pyroptosis targets microbes, host-adapted intracellular pathogens have evolved strategies to efficiently subvert it. However, environmental pathogens fail to evade, making pyroptosis a potent barrier against infection. We summarize recent findings for the host-adapted bacterial pathogens Shigella flexneri, Salmonella enterica serovar Typhimurium and Mycobacterium tuberculosis, contrasted with the environmental bacteria Burkholderia thailandensis and Chromobacterium violaceum. This Review discusses recent advances in our understanding of how gasdermins are activated by distinct signalling pathways and summarizes recent findings from host-adapted bacterial pathogens contrasted with environmental bacteria.

RNA design aims to find a sequence that can fold into a target secondary structure. It can create artificial RNA molecules for specific functions, with wide applications in medicine. It is computationally challenging due to two levels of combinatorial explosion: the exponentially large design space and the exponentially many competing structures per design. Popular methods such as local search cannot keep up with these combinatorial explosions. We instead employ two techniques from machine learning, continuous optimization and Monte-Carlo sampling. We start from a distribution over all valid sequences, and use gradient descent to improve the expectation of an arbitrary objective function. We define novel coupled-variable distributions to model the correlation between nucleotides. We then use sampling to approximate the objective, estimate the gradient, and select the final candidate. Our work consistently outperforms state-of-the-art methods in key metrics including Boltzmann probability and ensemble defect, especially on long and hard-to-design structures. RNA design aims to find a sequence that folds into a target structure. Here, the authors formulate it as a continuous optimization over a coupled-variable distribution and leverage sampling to optimize arbitrary objectives. On Eterna100, it outperforms state-of-the-art methods across key metrics.