RNA-KG is a recently developed biomedical knowledge graph that integrates the interactions involving coding and non-coding RNA molecules extracted from public data sources. It can be used to support the classification of new molecules, identify new interactions through the use of link prediction methods, and reveal hidden patterns among the represented entities. In this paper, we propose RNA-KG v2.0, a new release of RNA-KG that integrates around 100M manually curated interactions sourced from 91 linked open data repositories and ontologies. Relationships are characterized by standardized properties that capture the specific context (e.g. cell line, tissue, pathological state) in which they have been identified. In addition, the nodes are enriched with detailed attributes, such as descriptions, synonyms, and molecular sequences sourced from platforms such as OBO ontologies, NCBI repositories, RNAcentral, and Ensembl. The enhanced repository enables the expression of advanced queries that take into account the context in which the experiments were conducted. It also supports downstream applications in RNA research, including ‘context-aware’ link prediction techniques that combine both topological and semantic information. Finally, the recent integration of RNA-KG relationships into the RNAcentral portal provides a powerful resource for linking RNA-centric relationships with non-coding gene expression in human tissues, RNA secondary structures, and their functional roles in biological pathways, which can accelerate the discovery of novel therapeutic targets.

Early-life microbial exposures shape immune development and allergy risk. Food allergen sensitization, reflected by the presence of food allergen-specific immunoglobulin E (IgE), is an early indication of impaired immune tolerance. Here we show that early-life transmission of aromatic lactate-producing bifidobacteria strains in 147 children followed from birth to 5 years of age, facilitated by vaginal delivery, exposure to older siblings and exclusive breastfeeding for the first 2 months, led to increased levels of aromatic lactates in the infant gut. This microbiota–metabolite signature was inversely associated with the development of food allergen-specific IgE until 5 years and atopic dermatitis at 2 years. The observed effect was mediated by 4-hydroxy-phenyllactate, which inhibited IgE, but not IgG, production in ex vivo human immune cell cultures. Together, these findings define an early-life microbiota–metabolite–immune axis linking microbial transmission and feeding practices with reduced allergic sensitization. Vaginal birth, exclusive breastfeeding and early contact with siblings promote colonization of the infant gut with bifidobacteria capable of producing aromatic lactates, a microbial and metabolite signal that is inversely related to the risk of allergen-specific sensitization and dermatitis later in life.

Rhizosphere microbes benefit plant growth and health. How plant-microbe interactions regulate fruit quality remains poorly understood. Here, we elucidate the multi-level modulation of vitamin accumulation in tomato by flavonoid-mediated crosstalk between host plants and rhizosphere microbes. SlMYB12-overexpressing plants with up-regulated flavonoid biosynthesis accumulate higher levels of vitamins C and B6 in fruits compared to wild-type plants grown in natural soil. Flavonoid-mediated improvement of fruit quality depends on the presence of soil microbiomes and relates to rhizosphere enrichment of key taxa (e.g. Lysobacter). Multi-omics analyses reveal that flavonoids attract Lysobacter soli by stimulating its twitching motility and spermidine biosynthesis, which in turn boosts vitamin accumulation in fruits across tomato cultivars and soil types. RpoN acts as a dual regulator in L. soli that is responsive to flavonoids, controlling bacterial motility and spermidine production. Our study provides insight into flavonoid-mediated rhizosphere signalling and underscores plant-microbiome orchestration for improved tomato fruit quality. How fruit quality is regulated by plant microbiome remains poorly understood. Here, the authors reveal that flavonoids secreted by tomato roots can recruit specific soil microbes to the rhizosphere and stimulate spermidine biosynthesis, which can induce vitamin accumulation in tomato fruits.

In vitro antibiotic testing is important for guiding therapy and drug development. Current methods are focused on growth inhibition in bulk bacterial populations but often fail to accurately predict treatment responses. Here we introduce Antimicrobial Single-Cell Testing (ASCT), a large-scale live-cell imaging approach that quantifies bacterial killing in real time at single-cell resolution. By tracking over 140 million mycobacteria and analysing ~20,000 time–kill curves, we identify key determinants of antibiotic killing and its clinical relevance. For Mycobacterium tuberculosis, we found that drug-specific killing dynamics in starved bacteria, rather than growth inhibition or killing of growing cells, predict regimen efficacy in mice and humans. Extending this approach to Mycobacterium abscessus and comparing 405 bacterial strains, we show that antibiotic killing is also a genetically encoded bacterial trait (drug tolerance). We demonstrate that tolerance patterns cluster by antibiotic targets, identify a phage protein that modulates antibiotic killing, and show that strain-specific killing dynamics are associated with individual patient outcomes independent of drug resistance. Together, these findings establish a framework that reveals how drug properties and bacterial diversity shape treatment responses, offering a path to more effective and personalized therapies. Via high-throughput imaging and tracking over 140 million single mycobacteria, the authors show that drug- and strain-specific killing predict treatment outcomes, with potential to improve drug development and personalized therapy.

Malonyl-CoA serves as a central precursor for the biosynthesis of diverse high-value compounds, including lipids, organic acids, and polyketides, in engineered microbial fermentation systems. However, insufficient malonyl-CoA supply often limits the production of these products. Intracellular malonyl-CoA levels are tightly regulated by the activities of acetyl-CoA carboxylase (ACC) and fatty acid synthesis pathway enzymes. Although strategies have been developed to redirect malonyl-CoA flux from fatty acid biosynthesis toward desired products, native ACC-mediated synthesis remains constrained by slow kinetics, complex regulation, and ATP consumption. To overcome these limitations, two alternative malonyl-CoA biosynthetic pathways have recently been developed. The malonate assimilation pathway enables direct uptake and CoA ligation of exogenous malonate, providing precise control over malonyl-CoA metabolism. The non-carboxylative malonyl-CoA (NCM) pathway converts pyruvate to malonyl-CoA through a novel intermediate, eliminating both ATP and CO2 loss while simultaneously regenerating NADPH. This review highlights recent advances in these two alternative malonyl-CoA biosynthetic pathways and their applications across diverse microbial hosts, underscoring their potential to enhance the sustainable production of valuable biochemicals.

The presence of intracellular microbiota in epithelial cells of gastrointestinal tracts (GITs) of dairy cows, as well as their associations with rumen development, remains unclear.  Using a single-cell analysis of host-microbiome interactions (SAHMI) within a single-cell atlas derived from ten GITs tissue types collected from new-born (NB) and adult (AD) cows, we found that 20.5% of the single-cell RNA sequencing reads aligned to reference microbial genomes after filtering low-quality single cells and doublets. Comparative analysis revealed that abomasum tissue exhibited the highest proportion of cells detected microbial signals, with Paneth cells possessing the most genes classified as both marker genes and those related to microbial signals. In the NB rumen, Basal cells demonstrated the greatest overlap between differentially expressed genes in AD vs. NB comparison and the microbial signal-related genes. Notably, these microbiota-associated genes, which are mainly linked to Aliiroseovarius crassostreae, Enterobacter sp. T2, and Enzebya pacifica, are implicated in nucleotide excision repair mechanisms, including DNA replication and the cell cycle. Furthermore, bacterial fluorescence in situ hybridization (FISH) analysis indicated that these three microbial species were partially localized within the cytoplasm and nucleus of rumen epithelial cells in NB cattle.  These findings provide substantial evidence supporting the existence of an intracellular microbiome within the GITs of dairy cattle and highlight their potential relationships with rumen development. This research enhances our understanding of the crosstalk between hosts and microbiome during the maturation of ruminants.

Prime editing (PE) enables precise genome modifications without donor DNA or double-strand breaks, but its application in dicot plants has faced challenges due to low efficiency, locus dependence, and poor heritability. Here, we develop an ultra-efficient prime editing (UtPE) system for dicots by integrating evolved PE6 variants (PE6c and PE6ec), an altered pegRNA (aepegRNA), an RNA chaperone, and a geminiviral replicon. UtPE significantly improves editing performance in tomatoes, with UtPEv1 excelling in simple edits (unstructured RTTs) and UtPEv3 effective for complex targets (structured RTTs or multiple nucleotide changes). Compared to a PE2max-based tool, UtPE increases desired editing efficiency by 3.39 to 8.89-fold, enables editing at previous inaccessible sites, achieves an average of 16.0% desired editing efficiency in calli, and produces high-frequency desired edits in up to 87.5% of T0 plants. Multiplexed editing at up to three loci and stable T1 inheritance are also achieved, resulting in traits such as jointless pedicels and glyphosate resistance, while minimizing off-target effects. Low efficiency, locus-dependence, and poor heritability affect the application of prime editing to dicot plants. Here, the authors solve these problems by developing an ultra-efficient prime editing (UtPE) system for dicots by integrating evolved PE6 variants, an altered pegRNA, an RNA chaperone, and a geminiviral replicon.