Global warming driven by carbon emissions makes transitioning to a circular carbon economy increasingly urgent. Methanol is a promising energy carrier and a favorable substrate for biotechnology applications due to its high energy conversion efficiency and easy integration with existing infrastructure. Eubacterium limosum has the native ability to convert methanol into valuable chemicals, including acetate, butyrate, and hexanoate. However, oxidized cosubstrates such as CO2 or formate are generally required due to limitations in energy and redox balance. In this study, we demonstrate a novel approach using extracellular electron transfer (EET) with ferric iron (ferrihydrite) as an electron acceptor to enable methanol fermentation without carbon cosubstrates. This ferrihydrite-assisted methanol fermentation yielded a product spectrum like methanol-formate cofermentation, producing more reduced chemicals such as hexanoate. Physiological evidence indicated that cysteine (Cys-SH), an amino acid containing a thiol group (R-SH) and typically included in anaerobic media, was essential in enabling the indirect EET process between E. limosum and ferrihydrite. Thiol regeneration from cystine (Cys–S–S–Cys) confirmed that E. limosum catalyzed the cysteine/cystine recycling. Proteomic analysis revealed the metabolic pathways of this cysteine-mediated EET process. This EET-driven process advances our understanding of microbial carbon cycling and provides a sustainable biotechnology for chemical production.

Virus-induced genome editing (VIGE) using compact RNA-guided endonucleases is a transformational new approach in plant biotechnology, enabling tissue-culture-independent and transgene-free genome editing. We recently established a transgene-free VIGE approach for heritable editing at single loci in Arabidopsis by delivering ISYmu1 TnpB (Ymu1) and its guide RNA (gRNA) via Tobacco Rattle Virus (TRV). Here, we greatly improved this approach by devising a multiple gRNA expression system and by utilizing an engineered high-activity Ymu1 variant (Ymu1-WFR) to develop an efficient multiplexed genome editing approach.

The emergence of antibiotic-resistant bacteria has rendered conventional antibiotic treatments ineffective, necessitating the development of antibacterial agents with unique mechanisms of action. To address this challenge, researchers have increasingly resorted to synthetic and bioengineered nano-materials to augment the antibacterial activity of nonantibiotic antibacterials (nonantibiotic antibacterial agents), including antimicrobial peptides (AMPs), metallic nanoparticles (MNPs), bacteriophages (phages), and phage derivatives such as endolysins, which are under extensive investigation. In this review, we discuss how modifications and syntheses of these agents, leveraging advancements in nanoscience and nanotechnology, have and can significantly enhance their antibacterial properties and overcome limitations such as cytotoxicity, instability, and poor bioavailability for in vivo or clinical use. Furthermore, we highlight supramolecular strategies for improved delivery, including phage-based, AMP-based, and endolysin-based systems and their demonstrated efficacy against persistent bacterial infections. Additionally, we highlight how the integration of artificial intelligence and machine learning ultimately promises to revolutionize the design, optimization, and clinical translation of these precision antimicrobials, paving the way for targeted and highly effective treatments.

Adenine deaminase (ADE) plays a critical role in food purine degradation and in preventing hyperuricemia. Herein, the ADE genes from four different sources were expressed using pMA5, pMA0911, and pP43NMK as the vectors and Bacillus subtilis WB800 as the host. B. subtilis WB800/p43-AK-ADE exhibited the highest ADE activity of 45.41 U/mL. The optimum pH and temperature for AK-ADE were 7.5 and 45 °C, respectively. The enzyme remained stable within pH 6.0–8.5 and temperature 35–45 °C, respectively. The Km and Kcat values were 0.30 mM and 0.28 s–1, respectively. Furthermore, molecular docking and active pocket prediction revealed that AK-ADE binds to adenine with the lowest binding free energy, indicating the strongest affinity. In contrast, ADE from other sources exhibited markedly reduced binding and suboptimal residue positioning. Finally, when AK-ADE was applied to beer wort, it resulted in the removal of 71% of the adenine content, establishing its potential for industrial application.

Bacterial cellulose (BC) hydrogels produced by Gluconacetobacter species hold considerable promise for a wide range of applications owing to their exceptional mechanical properties, biocompatibility, and biodegradability. Achieving precise control over their structural and mechanical characteristics is crucial for the engineering of BC-based materials. In this study, we investigated the formation dynamics and structural features of BC hydrogels, emphasizing the complex interplay between cellulose nanofibril secretion and bacterial motility. Comprehensive tracking of bacterial movement during hydrogel formation has validated mechanisms underlying the development of branching and merging junctions, which are key elements that define the network's physical properties. Additionally, we observed the emergence of vortex-lattice and chiral-nematic structures during hydrogel development, depending on bacterial and cellulose densities. These insights contribute to a fundamental understanding of bottom-up 3D fabrication of BC hydrogels that harness the collective behavior of cellulose-producing bacteria.

Transposon insertion sequencing (TIS) encompasses methods such as Transposon Directed Insertion sequencing (TraDIS) and Transposon-Sequencing (Tn-Seq); these methods are widely uses for genome scale screens of essential and conditionally essential genes. Early limitations centred on mutant generation, today the biggest factor is sequencing and data generation and the costs associated. Here, we present and evaluate four methods TIS sequencing library generation protocols compatible with Illumina whole genome sequencing (WGS) workflows to maximise sequence efficiency and minimise turnaround times and costs. Using E.coli BW25113 transposon mutagenesis libraries generated with Tn5 and mariner (Himar1) transposons, we compare the methods in terms of reagent cost, workflow complexity, and target enrichments for the recovery of unique insertion sites and essential genes identified. All methods generated TIS data suitable for gene essentiality analysis. Illumina Flex based protocols were 4-6 times cheaper than the traditional Illumina Nextera based approach with similar or superior Transposon-Chromosome (Tn-Chr) junction enrichment. Across all methods, sequencing depth and mutant library density were the dominant factor in useful biological insight. Subsampling demonstrated that for a good quality mutant library, five million reads were sufficient to identify essential genes in E.coli. Whereas deeper sequencing reduced the statistical power and included contaminating background noise, seen primarily with Tn5. We conclude that an Illumina Flex based approach, especially when integrating with routine WGS, provides an excellent balance of speed, cost and data quality. Assuming five million reads and a robust Illumina Flex approach, a TIS library can be sequenced for around &pound40.

To counter challenges from bacteriophages (phages), bacteria use defense mechanisms that can reside on mobile genetic elements or within chromosomes. These immune systems are easily gained and lost, allowing adaptation to threats. However, the mechanism of mobilization of chromosomally encoded defense genes remains poorly understood. Here, we show that phage- and phage-inducible chromosomal island (PICI)–mediated lateral transduction (LT), a highly efficient horizontal gene transfer HGT mechanism, facilitates the transfer of these defense genes between bacteria. Using several bacterial models, we demonstrate that defense systems are often positioned near phage or PICI attachment sites, allowing them to exploit LT for their mobility. In addition, LT diversifies defense genes carried by prophages and PICIs, driving immune system evolution and turnover. These processes provide phage resistance to new bacterial hosts and profoundly affect population genomics. Our findings reveal LT as a crucial mechanism shaping bacterial evolution and influencing the trajectory of pathogenic clones in nature.

In natural and engineered ecosystems, diverse species interact in complex ways to form highly efficient microecologies. One key orchestrator of these interactions is autoinducer-2 (AI-2), a signaling molecule that plays a crucial role in microbial community assembly, metabolic flux, and resilience to environmental disturbances. This review provides the systematic synthesis of AI-2’s dual structural dynamics (S-THMF-borate/R-THMF interconversion) and its context-dependent roles in mediating bacterial crosstalk. It also reveals the receptor diversity (such as LuxP and LsrB) of AI-2 in bacterial kingdom and the signal transduction mechanism. Systematically elaborated on AI-2’s regulation of cellular metabolic flux and its ability to autonomously exhibit a series of coordinated behaviors in response to environmental changes. The review explores the ramifications of AI-2 on bacterial community interactions in synthetic biology and natural ecosystems. The wide application of AI-2-mediated interspecific communication in various fields including host health, agriculture, industry and environmental ecology has also been widely discussed. Factors influencing AI-2 production are thoroughly examined, including internal factors such as strain specificity, cell density, growth form and the phenotypic heterogeneity. Additionally, external biological factors (such as nutritional status and environmental stress) and abiotic factors (aggregation, diffusion, and flow) are discussed in detail. By examining knowledge gaps in AI-2-mediated spatial heterogeneity and multi-QS system coordination, this work charts a roadmap for harnessing microbial communication in chemical engineering and environmental sustainability.