Saponins are structurally diverse bioactive metabolites found in more than 100 plant families and are synthesized by plants as protection against insects, fungi, and other organisms. The mode of action is related to their interference with membranes, and particularly the membrane sterols. As membrane sterol composition varies across kingdoms and species, species-specific differences in toxicity may therefore be expected. The aim of this study was to elucidate the structure-activity relationships of different saponins across four different organisms (Daphnia magna, Enchytraeus crypticus, Saccharomyces cerevisiae, Raphidocelis subcapitata) representing three eukaryotic kingdoms of life using either immobility tests or growth inhibition assays. We hypothesized that monodesmosidic saponins are more bioactive due to higher amphiphilicity/polarity and that species susceptibility depends on sterol composition, with organisms containing plant sterols being less susceptible than those with animal or fungal sterols. The hypothesis was supported for monodesmosidic saponins, as α-hederin and hederacolchiside A1 exhibited significant cytotoxicity (EC50 values ranging from 8.7 to 36.9 and 2.0- 68.9mg/L, respectively, for the different organisms), whereas bidesmosidic saponins such as hederacoside C and ginsenoside-Ro were inactive at concentrations up to 100mg/L. The aglycone backbone and sugar moiety composition, however, also play critical roles, with simpler, linear saccharide chains leading to increased toxicity. C-23 hydroxylation has been shown to enhance mortality against insects; however, its absence did not affect the ability of hederacolchiside A1 to exhibit toxic properties. Additionally, species-specific sensitivities varied, with the crustacean D. magna being the most sensitive species, followed by the anelid worm E. crypticus, yeast S. cerevisiae, and the least-sensitive was, as hypothesized, the algae R. subcapitata. These insights contribute to a deeper understanding of saponin structure-activity relationships and open new avenues for the targeted development of saponin-based applications in agriculture, medicine, and biotechnology.

Leccinum is an ecologically significant and taxonomically complex genus of ectomycorrhizal fungi widely distributed across boreal, temperate, Mediterranean, and selected tropical regions. Despite its ecological, nutritional, and applied importance, no comprehensive review has previously synthesized global knowledge on this genus. This work provides the first integrative assessment of Leccinum research, combining a bibliometric analysis of 293 peer-reviewed publications with an in-depth qualitative synthesis of ecological, biochemical, and environmental findings. Bibliometric results show increasing scientific attention since the mid-20th century, with major contributions from Europe, Asia, and North America, and dominant research themes spanning taxonomy, ecology, chemistry, and environmental sciences. The literature review highlights substantial advances in phylogenetic understanding, species diversity, and host specificity. Leccinum forms ectomycorrhizal associations with over 60 woody host genera, underscoring its functional importance in forest ecosystems. Nutritionally, Leccinum species are rich in proteins, carbohydrates, minerals, bioactive polysaccharides, phenolic compounds, and umami-related peptides, with demonstrated antioxidant, immunomodulatory, and antitumor activities. At the same time, the genus exhibits notable bioaccumulation capacity for heavy metals (particularly Hg, Cd, and Pb) and radionuclides, making it both a valuable food source and a sensitive environmental bioindicator. Applications in biotechnology, environmental remediation, forest restoration, and functional food development are emerging but remain insufficiently explored. Identified research gaps include the need for global-scale phylogenomic frameworks, expanded geographic sampling, standardized biochemical analyses, and deeper investigation into physiological mechanisms and applied uses. This review provides the first holistic synthesis of Leccinum, offering an integrated perspective on its taxonomy, ecology, nutritional composition, environmental significance, and practical applications. The findings serve as a foundation for future mycological, ecological, and biotechnological research on this diverse and understudied fungal genus.

Brown seaweeds are marine bioresources rich in bioactive compounds such as carbohydrates, proteins, pigments, fatty acids, polyphenols, vitamins, and minerals. Among these substances, brown algae-derived polysaccharides (alginate, fucoidan, and laminarin) have promising industrial prospects owing to their distinctive structural features and diverse biological activities. Consequently, processing technologies have advanced substantially to address industrial requirements for biopolymer quality, cost-effectiveness, and sustainability. Over the years, significant progress has been made in developing various advanced methods for the sake of extracting, purifying, and structurally characterizing polysaccharides. Aside from that, numerous studies reported their broad spectrum of biological activities, such as antioxidant, anti-inflammatory, anticoagulant, and antimicrobial properties. Furthermore, these substances have various industrial, pharmaceutical, bioenergy, food, and other biotechnology applications. The present review systematically outlines the brown algae-derived polysaccharides treatment process, covering the entire value chain from seaweed harvesting to advanced extraction methods, while highlighting their biological activities and industrial potential as well.

Duckweed has attracted considerable attention for its high protein content, rapid growth, and broad potential in biotechnological applications. Understanding key phenotypic traits is crucial for unlocking and maximizing this potential. While most studies on duckweed growth have been conducted under natural or non-sterile conditions, here we minimize environmental influences and focus on the genetic component of growth by assessing growth performance under axenic culture. In this study, we measured relative growth rate (RGR) in four duckweed species, Landoltia punctata (G. Mey.) Les & D. J. Crawford, Lemna aequinoctialis Welw., Spirodela polyrhiza (L.) Schleid., and Wolffia globosa (Roxb.) Hartog & Plas. collected from various natural locations across Thailand. A total of six to seven strains were tested for each species. The relative growth rates of studied species ranged from 0.012 day−1 in S. polyrhiza to 0.162 day−1 in W. globosa. Significant intraspecific variation was observed in L. punctata, S. polyrhiza, and W. globosa, with the coefficients of variation between 9.6 to 109.9 percent. Each strain showed distinct growth characteristics: Most displayed a steady growth pattern, whereas W. globosa showed exponential growth at Day 35 after the start of experiment. The results provide the first systematic comparisons of baseline growth rate data for duckweed species in Thailand. These findings advance the understanding of strain-specific growth traits in duckweed and establish a standardized protocol for evaluating growth traits under axenic conditions, providing a basis for future research and applications.

Digital twins (DTs) have emerged in recent years as a very promising technology for individualized health management and recovery. Each DT is built upon a computational model that captures some aspect of human biology and is typically intended to be used with individuals to either restore their health or maintain it. In many cases, DT projects reach "from bench to bedside"and often involve commercialization to bring the final product to the end-user. The purpose of this perspective is to highlight DT technology as an opportunity for the mathematical modeling community and look at DT development from both the academic and commercial viewpoints.Since DT technology is applied widely in biology, beyond health care, there are several references at the end to applications in ecology, biotechnology, agriculture, and synthetic biology.

Early regulatory networks driving somatic embryogenesis in Saccharum spp. L. revealed by time-resolved proteomics.

Natural products continue to play a vital role as sources of bioactive compounds with promising applications in medicine, agriculture, and biotechnology. Fraxinus floribunda Wallich, a member of the Oleaceae family, is an important medicinal plant native to the Sikkim Himalayan region. The species produces a wide range of secondary metabolites, including coumarins, flavonoids, triterpenoids, and phenolic compounds, many of which exhibit strong antioxidant, anti-inflammatory, hepatoprotective, and antidiabetic properties. Structurally, these metabolites are characterized by aromatic frameworks bearing hydroxyl, methoxy, and glycosidic groups that enhance their biological reactivity. The coumarin (benzopyrone) and flavonoid cores contribute significantly to free-radical scavenging and facilitate interactions with biological enzymes and receptors, thereby supporting multiple pharmacological effects. In addition to its medicinal value, the species holds considerable economic importance as a source of bioactive coumarins, flavonoids, and triterpenoids utilized in pharmaceutical, cosmetic, and nutraceutical industries. Traditionally, F. floribunda has been employed to relieve joint pain, arthritis, inflammation, gout, fractures, and diabetes. Experimental investigations have confirmed its anti-nociceptive, hepatoprotective, and antioxidant potential through both extract-based and compound-specific studies. Spectroscopic analyses such as UV-Vis, FTIR, NMR, and MS have been utilized for the identification and structural confirmation of its bioactive constituents. Beyond its therapeutic importance, the plant also holds economic value due to its timber and potential for producing value-added phytochemical products. This review consolidates available data on the ethnobotanical relevance, phytochemistry, and biological activities of F. floribunda, underscoring the necessity for further pharmacokinetic, mechanistic, and spectroscopic studies to support its rational development as a natural source of therapeutic agents.

Microbial alginate lyases are essential biocatalysts for analyzing alginate structure and sustainably producing bioactive alginate oligosaccharides (AOS). In this study, we characterized Aly94, a novel alginate lyase from the polysaccharide lyase family 6 (PL6) family, identified from a marine sediment metagenomic library. Biochemical analyses showed Aly94 exhibits optimal activity at 40℃ in 50mM NaH₂PO₄-Na₂HPO₄ buffer (pH 7.0). Adding 20mM NaCl significantly increases its catalytic efficiency. The enzyme exhibits a strong preference for polyguluronate (polyG) over polymannuronate (polyM), with specific activities of 4.19U/mg (polyG), 0.25U/mg (polyM), and 2.45U/mg (alginate). When degrading substrates-particularly polyG-Aly94 primarily generates trisaccharides. Although Aly94 acts as an endolytic alginate lyase, it also could digest the monosaccharides from small oligosaccharide chains (∆G3, ∆G4). These catalytic properties, combined with its polyG-specific depolymerization, made Aly94 a promising candidate for biotechnological applications requiring controlled alginate saccharification and high-value AOS production.

Brain vasculature is a complex and heterogeneous structure that serves specialized roles in maintaining brain health and homeostasis. There is substantial interest in developing representative human models of the brain vasculature for drug screening and disease modeling applications. Many contemporary strategies have focused on culturing neurovascular cell types in hydrogels and microdevices, but it remains challenging to achieve anatomically relevant vascular structures that have similar function to their in vivo counterparts. Here, we present a strategy for isolating microvessels from cryopreserved human cortical tissue and culturing these vessels in a biomimetic gelatin-based hydrogel contained in a microfluidic device. We provide histological evidence of arteriole and capillary architectures within hydrogels, as well as anastomosis to the hydrogel edges allowing lumen perfusion. In capillaries, we demonstrate restricted diffusion of a 10kDa dextran, indicating intact passive blood-brain barrier function. We anticipate this bona fide human brain vasculature-on-a-chip will be useful for various biotechnology applications.

Delivering plasmid DNA (pDNA) into cells is essential for numerous biotechnological and biomedical applications. Among available nanocarriers, nonviral lipid-based vesicles are particularly promising for transfecting mammalian cells. Nevertheless, further development is required to create delivery systems that are both broadly effective across cell types and scalable for clinical use. Here, we explore stable nanovesicles composed of the sterol derivative cholesteryl N-(2-dimethylaminoethyl)carbamate (DC-CHOL) and myristalkonium chloride (MKC) as a platform for pDNA delivery. These nanovesicles, previously shown to efficiently deliver small RNAs to neuroblastoma cells, exhibit favorable physicochemical properties, such as high morphological uniformity and long-term colloidal stability, positioning them as strong candidates for DNA transfection. Using suspension-adapted human embryonic kidney 293 (HEK293) cells, which are widely employed for producing viral vectors and complex biotherapeutics, we evaluated the delivery performance of DC-CHOL/MKC nanovesicles with a reporter plasmid encoding enhanced green fluorescent protein. A Design of Experiments (DoE) approach was applied to identify and optimize critical transfection parameters, namely, the DNA concentration, DNA-to-vesicle ratio, and NaCl concentration in the complexing medium. This study demonstrates the capability of these nonviral vectors to deliver double-stranded plasmid DNA and emphasizes the critical role of the physicochemical characteristics of the pDNA/lipid complex in achieving efficient transfection.

Somatic embryogenesis (SE) enables somatic cells to develop directly into embryos. SE is a major approach of regeneration, but recalcitrance to SE has become one of the main obstacles to biotechnology-aided breeding, especially for perennial woody plants. Citrus is one of the most important fruit crops in the world, and glycerol has long been used to induce SE from the embryogenic callus (EC) of citrus. Recently, we reported that CsIAA4-mediated repression of auxin signaling plays a critical role in glycerol-induced citrus SE, but the downstream signaling cascade remains to be elucidated. In this study, the HD-Zip transcription factor CsHAT14 was identified as a key downstream regulator of auxin signaling in citrus SE. CsARF5 directly promoted CsHAT14 expression, which repressed SE through suppression of critical regeneration-related genes (CsDOF3.4 and CsWOX13) and the auxin efflux gene CsPILS5. CsIAA4 interacted with CsARF5, and this interaction attenuated CsARF5-mediated transcriptional activation of CsHAT14, thereby de-repressed CsHAT14- directly suppressed genes including CsDOF3.4, and thus promoted SE. Knockdown of CsDOF3.4 resulted in downregulation of cell cycle-related genes and impaired SE. Our findings established the CsIAA4-CsARF5 and CsHAT14-CsDOF3.4 modules-mediated auxin signaling cascade that coordinates citrus SE, which advanced our understanding of the mechanisms underlying SE and supported improvement of regeneration efficiency in citrus biotechnology applications.

Fungi synthesize a wide variety of secondary metabolites (SMs). The genes of the biosynthetic pathways of many of these compounds are encoded by biosynthetic gene clusters (BGCs), which typically consist of a core biosynthetic enzyme, tailoring enzymes, transporters, and pathway-specific regulators. One of the well-studied fungal SMs is the polyketide terrein, which is produced by Aspergillus terreus and exhibits a wide range of biological activities, such as cytotoxic, phytotoxic, and antibacterial effects. The structure and function of the terrein BGC, the functions of the encoded proteins, and the processes controlling the transcriptional regulation of the BGC are summarized in this mini review. Both pathway-specific and global regulators and epigenetic regulation are presented. Furthermore, similar BGCs identified in other fungal taxa are introduced in short. Despite significant advances, key aspects of terrein biosynthesis, such as some protein functions, details of the BGC regulation, and SM ecological functions remain unresolved. Filling in these gaps will help us better understand the biology of fungal SMs and could pave the way for biotechnological applications.

Recombinant protein expression can be a limiting step in the production of protein reagents for drug discovery and other biotechnology applications. We introduce RP3Net (Recombinant Protein Production Prediction Network), an AI model of small-scale heterologous soluble protein expression in Escherichia coli. RP3Net utilizes the most recent protein and genomic foundational models. A curated dataset of internal experimental results from AstraZeneca (AZ) and publicly available data from the Structural Genomics Consortium (SGC) was used for training, validation and testing of RP3Net. RP3Net achieves an increase in Area Under Receiver Operator Curve (AUROC) of 0.15, compared to a baseline model. When experimentally validated on an independent, prospective, manually selected set of 97 constructs, RP3Net outperformed currently available models, with an AUROC of 0.83, delivering accurate predictions in 77% of the cases, and correctly identifying successfully expressing constructs in 92% of cases. The model, along with installation and running instructions, is available under an MIT license at https://github.com/RP3Net/RP3Net, DOI 10.5281/zenodo.17243498. Supplementary data are available at Bioinformatics online.

In the present study, a new glucose specific lectin isolated from roots of an endemic plant Globularia alypum (designated as GaGBL1) is tested for its immunomodulatory, antihyperglycemic, antibacterial, anti-acetylcholinesterase and antioxidant activities. The obtained lectin was exposed to agglutinate human erythrocytes of ABO blood group system as well as the rabbit erythrocytes. Furthermore, the carbohydrate binding property of lectin was confirmed with the inhibition of HA. The fractions showed hemagglutination activity (HA). GaGBL1 was found to be glucose specific lectin with a purification yield of 557.83 μg/mL. It was isolated from a protein capable of agglutinating only human erythrocytes type O. Moreover, its relative agglutinating activity of 128 UH/mg protein was inhibited only by D-glucose with MIC value of 200mM. Meanwhile, D-galactose, D-lactose, D-fructose, D-mannitol, and D-sucrose did not exert this effect. The pure protein GaGBL1 possessed an immunosuppressor action. Meanwhile, this lectin showed no inhibitory effects neither against bacterial strains nor for acetylcholinesterase enzyme. In addition, the results indicated a negative antioxidant activity. Interestingly, it was seen that regular oral administration of GaGBL1 injected by intraperitoneal route to normoglycemic mice showed highly potent antihyperglycemic effect in comparison with the synthetic drug glibenclamide. These data reveal many interesting biological activities of GaGBL1 and suggest that complementary investigations to explore its immunosuppressive and antidiabetic potentials. Hence, further characterization of this novel lectin and understanding of its mechanism of action will help to broaden its scope of biotechnological, pharmaceutical and agricultural applications.

Novel insights into hákarl: A deep dive into the microbiological and physico-chemical features of Iceland's traditional fermented shark.

Engineering synthetic intrinsically disordered proteins (synIDPs) enables regulation of biomolecular condensation and protein solubility. However, limited understanding of how sequence-dependent interaction cooperativity relates to the fitness impacts of synIDPs on endogenous cellular processes constrains our design capability. Here, to circumvent this design challenge, we present a systematic directed evolution method for the evolution of synIDPs capable of mediating diverse phase behaviors in living cells. The selection methods allow us to evolve a toolbox of synIDPs with distinct phase behaviors and thermoresponsive features in living cells, leading to the evolution of synthetic condensates. The reverse-selection method further allows us to select synIDPs as solubility tags. We demonstrate the applications of the evolved synIDPs in protein circuits to (1) regulate intracellular protein activity and (2) reverse antibiotic resistance. Our systematic evolution and selection strategies provide a versatile platform for developing synIDPs for broad applications in synthetic biology and biotechnology.

The pathogenic bacterium Vibrio parahaemolyticus represents a substantial economic and public health concern; however, elucidating its virulence mechanisms has been significantly impeded by its inherent resistant to genetic manipulation, primarily attributed to sophisticated immune defense systems including restriction-modification (R-M) modules, CRISPR-Cas systems, standalone DNases, and DdmDE systems. Paradoxically, while genetic modification is essential for overcoming these barriers, the very barriers themselves obstruct DNA introduction. Our investigation focused on the V. parahaemolyticus X1 strain, where initial plasmid transformation attempts proved unsuccessful. However, low-efficiency conjugation allowed knockout of defense genes, thereby silencing the host's defense mechanisms. Our findings revealed a standalone DNase, Vpn, as the predominant obstacle to foreign DNA entry in the X1 strain, while a DdmDE system executes elimination of invaded plasmids. Leveraging these insights, we created the V. parahaemolyticus X2 strain via sequential depletion of the Vpn nuclease and the DdmDE system. Capitalizing on the bacterium's exceptional growth rate, characterized by a generation time of approximately 10.5 min, we established a highly efficient molecular cloning platform capable of creating a new plasmid construct within a single day. This work not only presents a strategic framework for genetic manipulation of previously recalcitrant bacterial species but also underscores the potential of fast-growing marine bacteria as promising candidates for next-generation biotechnological applications.

Here, we present the first complete, fully phased diploid genome of type strain Yarrowia lipolytica YB-423 (=ATCC 18942™), constructed using a combination of Oxford Nanopore long-read and Illumina short-read sequencing. Y. lipolytica is an industrially relevant yeast species known for its metabolic versatility, particularly its ability to degrade hydrophobic compounds and express useful products such as fatty acids. Despite its growing use in biotechnology, a high-quality genome assembly of the species' diploid type-strain has been lacking. The assembly and annotations presented here span six chromosomes of paired "haplotigs" and a mitochondrial genome, capturing large-scale structural variations and prominent levels of genome-wide heterozygosity. Variant analysis revealed 13,908 heterozygous alleles, of which 3,201 alleles were distributed among 1,237 protein-coding genes. Gene set enrichment analysis showed that these variants are enriched among genes involved in transmembrane transport, suggesting a role in environmental adaptability. Comparative analysis of matched haplotigs for the same chromosome uncovered multiple inversions and transpositions, as well as allele-specific insertions of retrotransposons, providing new insights into the structural complexity and evolutionary dynamics of the genome. The fully phased, finished diploid genome of Y. lipolytica YB-423 represents a crucial step toward unlocking the full genetic potential of Y. lipolytica. Our work will provide a valuable foundation for future comparative and functional genomics and strain engineering studies, particularly for industrial microbiology and biotechnology applications.

Coastal marine bacteria with hydrocarbon-degrading capacity: Isolation, screening, and genomic insights.

Engineering DNA polymerases to efficiently synthesize artificial or noncognate nucleic acids remains an essential challenge in synthetic biology. Here we describe an evolutionary campaign designed to convert a family of highly selective DNA polymerases into an unnatural homolog with strong RNA synthesis activity. Starting from a homologous recombination library, a short evolutionary path was achieved using a single-cell droplet-based microfluidic selection strategy to produce C28, a newly engineered polymerase that can synthesize RNA with a rate of ~3 nt s-1 and of >99% fidelity. C28 is capable of long-range RNA synthesis, reverse transcription and chimeric DNA-RNA amplification using the PCR. Despite strong discrimination against other genetic systems, C28 readily accepts several 2'F and base-modified RNA analogs. Together, these findings highlight the power of directed evolution as an approach for reprogramming DNA polymerases with activities that could help drive future applications in biotechnology and medicine.