Food Research International

Edible seaweed has the potential to become a valuable marine resource for food applications due to its potential health benefits and ecological sustainability. The brown seaweed Alaria esculenta is rich in essential minerals, vitamins, and dietary fibers, making it a nutritious food source. Fermentation, as a traditional preservation method, can enhance seaweed shelf-life and be useful for the development of new foods/ beverages. In this study, the effects of fermentation of A. esculenta, by the lactic acid bacterium (LAB) Lactiplantibacillus plantarum, on the nutritional profile, and the content of potentially toxic elements, was investigated. L. plantarum was successfully cultivated on A. esculenta using two modes of operation, submerged (SmF) and solid-state fermentation (SSF), resulting in production of cells and lactic acid, and reduction of the pH to below 4.3 within 3 days, which was not achieved in parallel spontaneous fermentations using indigenous seaweed microbiota. A. esculenta s macro-nutritional profile was altered, reducing mannitol but increasing fucose and glucose content (after acid hydrolysis) while also concentrating the protein content. LAB fermentation significantly increased the concentration of antioxidant phenolic compounds, such as phloroglucinol, syringic acid, and epicatechin, compared to untreated samples. However, lipophilic compounds like carotenoids decreased after both spontaneous and LAB-fermentation. A reduction in total mineral content was observed after LAB fermentation and water soaking, and SmF with L. plantarum effectively reduced arsenic and iodine levels. Overall, fermentation using L. plantarum showed potential as a bio-preservation method for the edible brown seaweed, A. esculenta, improving its nutritional profile and enhancing food safety.

To evaluate the international interoperability of food composition databases, we assessed the compatibility of seven national food composition tables with USDA FoodData Central (FDC) using the LLM-based matching method reported previously (Nakagawa and Yamamoto, 2026). Databases from four English-speaking countries (Canada, United Kingdom, Australia, and New Zealand), South Korea, and Japan were compared with 8,158 USDA FDC entries (SR Legacy and Foundation Foods, excluding Survey/FNDDS). Match rates varied by country (62.0-89.7%) and food category. After excluding six USDA categories unsuitable for cross-national comparison, 45.2% of the remaining 6,290 entries were not matched by any country. Canada showed the highest concordance, reflecting shared North American food supply. Japan and South Korea showed similar low coverage for vegetables and spices. These findings suggest that while USDA FDC represents a practical foundation for a globally comprehensive food composition database given its breadth, systematic incorporation of country-specific foods and classification schemes will be necessary to achieve true international interoperability.

Seaweed cultivation has recently gained increased attention in North-West Europe as a sustainable source of biomass for biobased products. However, yields need to increase to make the seaweed sector economically viable. To achieve this, higher yielding varieties can be bred but this requires variation for yield and yield-related traits among genotypes. To reliably select high-yielding genotypes, an understanding is required of how both within-farm and between-farm environmental differences affect phenotypes and how to identify simple and reliable proxies for yield. In this study we evaluated growth of nine Saccharina latissima genotypes on two farms, 12 km apart, within the same season. We observed a threefold difference in yield among genotypes, demonstrating the potential for improvement through selection and breeding. Blade thickness and blade size-related traits were strongly correlated with yield, highlighting their potential to serve as rapid and non-destructive proxies for yield, thereby accelerating selection. Furthermore, we demonstrated the importance of adequate replication in farm trials to improve genotype performance estimation by correcting for within-farm spatial variation. Moreover, phenotypic variation was most explained by the genotype and environment, highlighting the importance of both genotype and site selection. Although genotype by environment interactions (GxE) were significant, its contributions were small, indicating stable genotype ranking across farms. Overall, these results are promising for breeding improved S. latissima as it indicates that genotype performance is consistent across close by locations and that local S. latissima populations harbour substantial phenotypic variation that can be used to breed for increased yield. Highlights- Local genetic resources harbour substantial variation in yield and morphology for breeding. - Minor GxE allows for breeding across farms. - Blade thickness and blade size related traits are good predictors of yield. - Correction for on-farm spatial variation improves genotype performance estimation.

Fungal contamination of food with yeast and moulds is associated with major economic losses due to spoilage and also poses health risks in the form of mycotoxin production. The strain Pantoea agglomerans APC 4211 isolated from leaves of Ilex aquifolium (holly tree) has broad spectrum antifungal activity against a variety of food spoilage fungi. Genomic analysis of the strain confirmed the presence of biosynthetic gene clusters potentially encoding for the enzymatic machinery required for the production of the antifungal lipopeptide herbicolin A. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis of the cell-free supernatant (CFS) confirmed the presence of molecular masses corresponding to herbicolin A (1300.8 Da), and herbicolin B (1138 Da). Purified herbicolin A has desirable properties for biotechnological applications, including potent antifungal activity against a range of spoilage fungi, thermal stability and resistance to proteases. Herbicolin A has low cytotoxicity against epithelial cell lines and has minimum inhibitory concentrations (MICs) lower than those of some commercial antifungal drugs (0.2 - 2.5 {micro}g/ml). In a model dairy system (10% skim milk), herbicolin A demonstrated excellent solubility and stability, effectively eliminating Aspergillus niger and Penicillium notatum at a concentration of 5 {micro}g/mL. In conclusion, herbicolin A is a potent, naturally occurring antifungal agent with the potential to be applied as a biopreservative in food systems, providing a safe, clean-label, and efficient compound for synthetic preservatives replacement. HighlightsO_LIHerbicolin A has a strong potential as a natural preservative for food protection C_LIO_LIHerbicolin A shows lower MICs than several antifungal agents C_LIO_LIHerbicolin A is stable under heat and resistant to proteolytic degradation C_LIO_LIHerbicolin A has strong solubility and stability in a model dairy system C_LIO_LIHerbicolin A indicates low cytotoxicity against epithelial cell lines C_LI Data summaryThe authors confirm all supporting data, code and protocols have been provided within the article or through supplementary data files.

Adipogenesis is the process of adipose-derived stem cells (ADSCs) responding to extracellular signals from the stem cell niche to differentiate into adipocytes (fat cells) and may be studied in vitro using a cocktail of chemicals that promote adipogenic differentiation to produce differentiated ADSCs (dADSCs). The global membrane N- and O-glycosylation changes of this process have been previously analysed and compared to native adipocytes as a benchmark for a true adipocyte profile, and revealed that bisecting GlcNAc type N-glycans are characteristic of adipogenesis. As stem cell differentiation has been widely reported to result in cellular protein changes, the same cells (ADSCs, dADSCs and mature adipocytes) were characterised for their membrane proteome here using label-free quantitative shotgun proteomics analysis. The membrane proteome displayed more differences in protein numbers between the cell types compared to the previously reported N-glycome which had shown high identical glycomes between stem cells and in vitro dADSCs, suggesting that the proteome is more dynamic during in vitro adipogenesis. Following the global shotgun proteomics analysis, a more targeted approach of carrying out proteomic analysis of de-N-glycosylated peptides of gel-separated proteins unearthed new glycoproteins not detected in the shotgun proteomic analysis. This approach identified the adipogenic marker, CD36, to be under-represented in the shotgun proteome analysis, but as the dominant (glyco)protein in the adipocyte membrane proteome that was also up-regulated at the mRNA transcript level in both the in vitro differentiated ADSCs (7.1-fold increase) and mature adipocytes (102.9-fold increase). A comparison of CD36 sequence coverage in the global shotgun analysis with the de-N-glycosylated CD36 revealed a 41% increase when N-glycans were removed prior to trypsin digestion, explaining its observed increased abundance and highlights the crucial need for de-N-glycosylation of proteins in proteomics experiments for increased identification of glycoproteins. The systems glycobiology approach by the integration of previously reported glycomics data and the proteomics and transcriptomics analyses in this work extended the investigation of membrane protein glycosylation changes in adipose-derived stem cell differentiation. The work provides a framework for future glycoproteomics-based investigations into the differentiation of stem cells into adipocytes, and will allow their related pathologies and potential therapeutic applications to be discovered. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=121 SRC="FIGDIR/small/722121v1_ufig1.gif" ALT="Figure 1"> View larger version (44K): org.highwire.dtl.DTLVardef@189a786org.highwire.dtl.DTLVardef@5563b8org.highwire.dtl.DTLVardef@5cb5borg.highwire.dtl.DTLVardef@69e11f_HPS_FORMAT_FIGEXP M_FIG C_FIG

Conventional semen cryopreservation involves equilibration at 4{degrees}C and optimum freezing rates. We hypothesized that a cholesterol-based semen extender obviates the need for equilibration, minimizing total processing time for semen cryopreservation. Experiments were conducted to determine the effects of semen extender (egg yolk- or cholesterol-based) and freezing method (routine or fast) on post-thaw sperm characteristics and fertility of beef and bison semen. In Experiment 1, beef semen diluted in tris-egg yolk-glycerol (TEYG) or cholesterol-cyclodextrin tris-glycerol (CCTG) extender underwent routine or fast freezing method. Cholesterol from animal and plant origins were compared. The routine method included 90-min equilibration at 4{degrees}C and routine freezing (RE-RF, total time 97 min) whereas the fast method included no equilibration and fast freezing (NE-FF, total time 14 min). Post-thaw sperm quality was assessed by CASA, and in vitro fertilization. Post-thaw sperm motility was not affected by the origin of cholesterol (animal or plant), but was lowest in the TEYG NE-FF group (24% vs 43-51%, P < 0.05). In vitro cleavage and blastocyst development rates did not differ between RE-RF and NE-FF groups. In Experiment 2, bison semen was diluted in TEYG or plant-CCTG extender and frozen as in Experiment 1. Post-thaw sperm motility was lowest in the TEYG NE-FF group (10% vs 39-51%, P < 0.05). In Experiment 3, beef semen diluted in TEYG or plant-CCTG extender underwent either a routine (RE-RF) or modified freezing (NE-RF, total time 25 min) method. Post-thaw sperm characteristics did not differ between extenders but were greater using routine freezing (RE-RF) compared to the modified method of freezing (NE-RF). Pregnancy rates were similar between extenders (TEYG vs plant-CCTG) using the modified freezing method without equilibration and insemination at 72 h after progesterone device removal. In conclusion, beef and bison semen diluted in cholesterol-based extender may be cryopreserved without equilibration.

Purpose: Modified Barium Swallow (MBS) studies utilize videofluoroscopy, a dynamic X-ray technique for evaluating swallowing anatomy and physiology. Each MBS exam typically includes multiple bolus trials, often involving different bolus consistencies. Accurate classification of bolus types is essential, as swallowing dynamics, aspiration risks, and residue levels vary with bolus consistency. In this preliminary study, we propose a deep learning-based approach for automated bolus type classification in MBS, aiming to provide a standardized and efficient framework for automated processing of swallowing assessments. Methods: A total of 206 patients (Mean +/- SD age: 60.24 +/- 9.02 years; 89.32% men) underwent MBS examinations, comprising 277 individual MBS studies. The dataset included 2,752 bolus-level video segments, categorized by bolus type as follows: 1,711 liquid (IDDSI 0-3, 62.17%), 521 pudding (IDDSI 4, 18.93%), and 520 solid boluses (IDDSI 7, cookie or cracker, 18.89%). To standardize variable video lengths for the data pipeline, each MBS video was temporally segmented into a fixed-length frame sequence, with shorter videos padded using static frames and longer videos randomly cropped to the target length. We employed an Inflated 3D convolutional neural network to develop the deep learning model. Results: Each video segment contained an average of 273.03 +/- 195.81 frames. On the independent test set, the deep learning model achieved an overall accuracy of 96.13%, and the macro F1-score was 95.05% in classifying food bolus types within MBS videos. Conclusions: The developed AI-based system demonstrated effective automated classification of food bolus types in MBS videos, representing an important step toward fully automated MBS analysis for swallowing efficiency assessment. The AI model reduces the reliance on manual labels, thereby promising to streamline clinical and research workflows.

Fermented foods are increasingly recognized for their health-boosting potential, yet the mechanisms involved are not fully resolved. Here, we tested whether kombucha reshapes the gastrointestinal microbiome and whether these changes are associated with stress-related behaviors under contrasting dietary backgrounds. Male C57BL/6 mice were fed either a total Western diet (TWD) or a control diet (CTRL) supplemented with kombucha or water three times weekly for seven weeks. Depressive-like and anxiety-related behaviors were evaluated using the forced swimming (FST) and marble burying tests (MBT). Ileum, cecum, and colon microbiomes were profiled via 16S rRNA, ITS2, and shotgun metagenomics, while feces and whole brains were profiled by LC-MS metabolomics. Serum cytokines were measured by ELISA. Results highlight diet-dependent effects of Kombucha on behavioral, microbial and metabolic outcomes. Kombucha reduced immobility in the FST under both diets, whereas fewer marbles buried were observed only under TWD. Kombucha intake enriched Bifidobacterium pseudolongum in the ileum under CTRL and TWD diets, while cecal microbial functions related to amino acid metabolism were stimulated mainly under CTRL. Only CTRL mice receiving kombucha showed higher fecal acetate and butyrate together with lower fecal levels of neurochemically relevant amino acids, including glutamine, phenylalanine, tryptophan, and tyrosine. Under TWD, kombucha was associated with lower spleen weight and altered brain tryptophan/kynurenine profiles. These findings identify kombucha as a food intervention that can remodel gastrointestinal microbial and neuroactive metabolism in a diet depending manner. Associations with reduced depressive and anxiety-related behaviors are promising but warrant further exploration. Key HighlightsO_LIKombucha supplementation reshaped the mice gastrointestinal microbiome and its neuroactive potential C_LIO_LIKombucha intake was associated reduced depressive and anxious like behaviors C_LIO_LIThe potential of kombucha to modulate microbial, metabolic and behavioral outcomes may be dependent on subject dietary background C_LI

The chemical constituents and cytoprotective potential of Cyathea podophylla, a Vietnamese fern, remain poorly investigated. This study aimed to isolate its compounds and evaluate their in vitro cytoprotective activity against 6-hydroxydopamine (6-OHDA)-induced toxicity in F11 cells. Compounds were chromatographically isolated and structurally characterized using NMR and HR-ESI-MS. Seven compounds were identified: five phenolics (trans-cinnamic acid, (E)-4-(3,4-dihydroxyphenyl)but-3-en-2-one, p-coumaric acid, 3,4-dihydroxybenzoic acid, 4-O-acetyl-caffeic acid), 5-hydroxymethylfurfural, and butyl-{beta}-D-fructofuranoside. Six of these are newly reported for the Cyathea genus. In MTT assays, butyl-{beta}-D-fructofuranoside exhibited the strongest cytoprotective effect (69.6% cell protection at 10 {micro}M, p < 0.001), followed by (E)-4-(3,4-dihydroxyphenyl)but-3-en-2-one (39.2% at 10 {micro}M). The remaining compounds lacked significant activity. These findings expand the phytochemical profile of Cyathea podophylla and provide preliminary evidence of its cytoprotective properties against 6-OHDA-induced injury, warranting further mechanistic and in vivo validation.

Chitosan is an oligosaccharide derived from chitin that is protonated at acidic pH to form a polycation. Its positive charge promotes the interaction with negatively charged components of the yeast cell surface, which has been associated with increased cell permeability and growth inhibition. In this study, we investigated the interaction of chitosan with the cell surface and its permeabilizing capacity in three yeast species displaying distinct susceptibility profiles, Saccharomyces cerevisiae, Candida albicans and Debaryomyces hansenii. We evaluated the correlation between differential susceptibility and chitosan association at the cell surface, as well as cell permeabilization, by integrating growth analyses with surface-binding assays, including FITC-conjugated chitosan to monitor surface association and cellular integration over time, and ultrastructural examination by transmission electron microscopy (TEM). Our results showed that chitosan exhibited varying effects on the growth and permeability of each yeast strain, with D. hansenii being the most susceptible. Furthermore, we observed the incorporation of chitosan onto the cell surface and confirmed its role as a permeabilizing agent. Finally, we used chitosan-induced permeabilization as a method to measure the activity of selected enzymes in situ, demonstrating its potential for studying metabolic functions in permeabilized yeast cells. Overall, our findings establish chitosan as a strain-dependent antifungal agent and a useful tool for functional biochemical analyses in yeast.

Microbial colonization is a major cause of deterioration in paintings, leading to discoloration, pigment degradation, and loss of structural integrity. While biodeterioration of artworks has been studied in temperate climates, tropical environments remain underexplored despite their high humidity and temperature, which promote microbial growth. This study assessed the microbiological deterioration of two eighteenth-century oil paintings, La Muerte de San Jose and Virgen de Guadalupe, located in Orosis Colonial Church and Religious Art Museum, Costa Rica. Microorganisms were isolated and identified using VITEK(R) 2, microscopy, and MALDI-ToF analysis, and their biofilm-forming capacity was evaluated. Additionally, the antimicrobial activity of six essential oil components was tested using direct and indirect contact assays. Twenty-three bacterial species and fifteen fungal genera were identified, with Bacillus, Staphylococcus, Cladosporium, and Aspergillus among the most common. Notably, La Virgen de Guadalupe displayed the highest microbial diversity, reflected in a high Shannon index, indicative of a more complex microbial community. Several isolates displayed strong biofilm formation, particularly Bacillus subtilis/amyloliquefaciens/vallismortis and Staphylococcus saprophyticus. Linalool exhibited the strongest inhibitory activity, achieving complete bacterial growth inhibition in non-contact assays. Environmental monitoring revealed persistently elevated relative humidity and CO2 levels during the study period. Together, these results reveal the complex microbial ecology of tropical heritage paintings and demonstrate that volatile essential oil components can serve as candidates for low-impact antimicrobial strategies in preventive conservation. ImportanceUnderstanding the microbiological deterioration of cultural heritage in tropical environments is crucial for designing sustainable conservation strategies. While microbial colonization of artworks has been widely studied in temperate regions, data from tropical climates remain limited despite inherently favorable conditions for microbial proliferation. This study integrates microbiological, environmental, and physicochemical analyses to characterize microbial communities colonizing eighteenth-century oil paintings in Orosi, Costa Rica. By combining microbial identification, biofilm quantification, and essential oil biocide testing, it bridges applied microbiology and cultural heritage conservation. The finding that volatile components such as linalool inhibit biofilm-forming bacteria without direct contact highlights their potential as eco-friendly, noninvasive antimicrobial alternatives to conventional biocides. These results expand the understanding of biodeterioration dynamics under tropical conditions and offer a practical framework for developing sustainable, evidence-based conservation protocols that protect both heritage materials and the environment. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=171 SRC="FIGDIR/small/723565v1_ufig1.gif" ALT="Figure 1"> View larger version (98K): org.highwire.dtl.DTLVardef@16cd608org.highwire.dtl.DTLVardef@57aa00org.highwire.dtl.DTLVardef@159fcbeorg.highwire.dtl.DTLVardef@e0363b_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 0.C_FLOATNO Artistic visualization of the geographical context of the studied artworks and the multidisciplinary analytical approaches applied, highlighting the diversity of microorganisms identified (illustration by Keylin Urena-Alvarado). C_FIG

This work aimed to establish a translationally viable, xeno-free, serum-free platform and protocol for the isolation and expansion of human salivary stem/progenitor cells (hS/PCs) suitable for regulatory qualification and future FDA-approved first-in-human autologous regenerative therapy trials for the treatment of hyposalivation disorders. Parotid gland specimens from non-cancerous regions/tissues were collected from consented surgical patients. Primary hS/PCs were isolated from tissue specimens, cultured in animal-component-free conditions, expanded to produce millions of cells, then enriched for CD44+ stem/progenitor cells by magnetic cell sorting. Normal epithelial purity was assessed using cytokeratins 5/14. Anti-CD133/PROM1 (cancer marker) and anti- fibroblast (clone TE-7) antibodies were used to demonstrate a lack of contaminating cells. Phenotype validation was performed by flow cytometry and immunocytochemistry on both CD44+ sorted and unsorted populations. Senescence-associated beta-galactosidase (SA-{beta}-gal) assays were performed across serial passages (P1-P6). Pluripotency was demonstrated by culture under conditions supporting lineage-specific differentiation. Primary hS/PCs demonstrated consistent expansion and epithelial morphology under serum-free conditions. CD44 expression remained high (>95%) throughout expansion, with negligible detection of CD133 or fibroblast markers, confirming epithelial purity and absence of tumorigenic or stromal contamination. Immunocytochemistry corroborated these expression profiles. SA-{beta}-gal staining revealed only a minor, passage-dependent increase (5-16%) in senescent cells from multiple donors, indicating retention of proliferative potential. Our defined, animal-free culture system supports stable expansion of pure low passage hS/PCs under conditions compatible with good manufacturing practice (GMP).

Monoclonal antibody (mAb) glycosylation is a critical quality attribute that is difficult to rationally engineer and rapidly assess during cell line development. Here, we investigate whether cell-surface glycosylation can serve as a predictive indicator of mAb product glycosylation following targeted glycogene engineering in CHO cells. Five key glycogenes (COSMC, FUT8, B4GALT1, ST3GAL4, ST6GAL1) were investigated in two mAb-producing CHO cell lines. Product glycan analysis revealed consistent, gene-specific effects across hosts, including loss of core fucosylation, and tuneable galactosylation and sialylation. Lectin-based surface profiling reliably reflected product outcomes for COSMC and FUT8 modifications but showed limited predictive power for galactosylation and 2,3-sialylation, highlighting glycosylation pathway redundancy and context dependence. This study provides the first systematic, cross-cell line evaluation of lectin-based cell-surface glycan profiling as a predictor of mAb product glycosylation, establishing its practical utility and inherent limitations for CHO glycoengineering workflows. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=118 SRC="FIGDIR/small/724788v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@6d5cfborg.highwire.dtl.DTLVardef@1f38e0aorg.highwire.dtl.DTLVardef@f25fa2org.highwire.dtl.DTLVardef@64a0dc_HPS_FORMAT_FIGEXP M_FIG C_FIG

Fermentation of C1 gases is an emerging technology where waste gases are bio converted into value-added products. This study navigates the gas fermentation potential of Gordonia rubripertincta to produce carotenoids. The crucial carbon monoxide dehydrogenase (CODH) enzyme, necessary for gas uptake by the microbe, was found to be present in G. rubripertincta through blastp on NCBI website. The organism was then used for gas fermentation experiments in a continuous stirred tank reactor (CSTR) in different modes of reactor operation resulting in the production of about 500 mg pigment/g WCW (wet cell weight). Two important reactor parameters, molybdenum content and pH, were optimized for enhanced carotenoid production. Overall, G. rubripertincta was observed to be an efficient candidate organism for C1 gas fermentation. KEY HIGHLIGHTSO_LIGordonia rubripertincta synthesises aerobic carbon monoxide dehydrogenase enzyme. C_LIO_LIIt is a potential gas fermenting microbe that gives carotenoids as product. C_LIO_LIThe gas uptake efficiency of the microbe is more in fed-batch discontinued mode. C_LIO_LIIn FB-D, the resultant carotenoids are 500+9 mg/g wet cell weight (WCW). C_LIO_LIMo/pH of 20 mg/7.0 resulted in highest carotenoids, i.e., 134+41 mg/g WCW. C_LI GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=87 SRC="FIGDIR/small/722808v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@8b1185org.highwire.dtl.DTLVardef@2b6f90org.highwire.dtl.DTLVardef@1a9697dorg.highwire.dtl.DTLVardef@14c9dc8_HPS_FORMAT_FIGEXP M_FIG C_FIG

Fresh produce encounters pathogens at various stages of production and supply, with the harvesting process serving as one of these stages. To evaluate contamination associated with harvesting, we systematically swabbed zone 1 harvester surfaces and quantified Bacteroidales as a fecal biomarker using quantitative polymerase chain reaction (qPCR). Baseline contamination was dominated by non-detects, with occasional low-level detections (<25 copies/cm2) near the assay limit of detection (LoD). Detection occurred more frequently post-harvest (overall [~]4% pre-harvest and 10% post-harvest), while microbial loads remained low, indicating that harvesting primarily affected the likelihood of low-level contamination rather than increasing contamination abundance. Additionally, we developed and field-deployed a portable loop- mediated isothermal amplification (LAMP) assay for rapid harvester hygiene assessment and benchmarked its field performance against qPCR. Together, these results support a practical molecular tool for monitoring fecal contamination and informing cleaning and sanitization decisions.

Though currently a minor crop, faba bean is a promising source of plant-based protein as global diets shift towards more plant-based nutrition. To realise this potential, advances in breeding and cultivation are crucial. To exploit heterosis, faba bean breeding frequently utilises synthetic cultivars, which involves open pollination of inbred lines to produce a mixture of F1 hybrid seeds and self-pollinated offspring. Pure F1 hybrid cultivars are currently unavailable due to unstable cytoplasmic male sterility (CMS) systems. An ability to distinguish F1 seeds from their parental inbreds via characteristics associated with xenia effects could change this. The xenia effect refers to the influence of paternal pollen on seed traits, for example seed weight and cotyledon cells in faba bean. In this study, we exploited the xenia effect captured in hyperspectral imaging data to develop machine learning scenarios for discriminating between parental and F1 seeds of open pollinated synthetic combinations (Syn-1). The hyperspectral data were pre-processed using Savitzky-Golay filtering to reduce noise and smooth the spectra. Various machine learning algorithms were applied, incorporating Bayesian hyperparameter optimisation. The scenarios achieved up to 98.9 % accuracy in separating parental components of Syn-1. When including all seeds, the model achieved 40.7 %, indicating moderate detection and classification performance. As the harmonic mean of precision and recall, the F1 score accounts for both the correctness of F1 seed detections and the completeness with which F1 seeds were detected. While this approach does not yet enable the development of full hybrid cultivars, it paves the way for hybrid-enriched cultivars. These could help to streamline breeding for synthetic cultivars and potentially increase yields, for example by increasing the proportion of F1 hybrid seeds in synthetic cultivars. This study extends knowledge of the xenia effect in faba bean and provides a basis for further research aimed at enhancing breeding methods and productivity.

Administration of ciliary neurotrophic factor (CNTF) reduces food intake and body weight in both humans and experimental animals, where it also ameliorates hyperglycemia, hyperinsulinemia, and dyslipidemia. To exert its anti-obesogenic and anti-diabetogenic effects, CNTF targets brain feeding centers as well as multiple peripheral organs inducing the phosphorylation of the transcription factor signal transducer and activator of transcription 3 (p-STAT3). However, data showing which peripheral cytotypes are specifically targeted by exogenous CNTF in vivo in metabolically relevant organs are currently lacking. Here, we first evaluated the gene expression levels of the subunits of the tripartite CNTF receptor (Cntfr) complex, i.e., the Cntfr, the leukemia inhibitory factor receptor {beta} (Lifr{beta}) and the glycoprotein 130 (gp130), by quantitative real-time PCR in metabolically relevant organs of adult male mice: gastrointestinal (GI) tract, pancreas, liver, visceral and subcutaneous white (WAT) and interscapular brown adipose tissue (iBAT), skeletal muscle and the sciatic nerve. We then quantified p-STAT3 by Western blotting in these organs after intraperitoneal administration of CNTF (0.3 mg/kg) or saline. Finally, we mapped CNTF-responsive cells by immunohistochemistry, followed by morphometric quantification and confocal microscopy in both CNTF- and saline-treated mice. Lifr{beta} and gp130 were ubiquitously detected across all the investigated organs; the Cntfr showed the highest expression levels in the skeletal muscle, sciatic nerve, and iBAT, whereas it was found to be expressed to a lesser extent in the other sites. Administration of CNTF led to a significant increase of p-STAT3/STAT3 protein ratio in all organs examined, except the duodenum, and induced a distinctive pattern of cell nuclear p-STAT3 immunoreactivity. Notably, along the analyzed GI tract CNTF induced nuclear STAT3 phosphorylation in neurons of the submucosal and myenteric plexuses of the enteric nervous system and in contractile cells of the muscularis externa, where the response peaked in the mesenteric gut and colon. In the pancreas, CNTF triggered a higher activation within the endocrine component compared to the exocrine parenchyma. In the liver, CNTF induced STAT3 phosphorylation not only in parenchymal cells but also in sinusoids and resident macrophages. The cytokine activated p-STAT3 in subcutaneous and visceral white adipocytes, but also in brown adipocytes, with a prominent response observed in the beige subcutaneous adipocytes; adipose resident macrophages and endothelial cells of numerous blood vessels were also CNTF-responsive. Lastly, in skeletal muscle, a major site for glucose/lipid utilization, CNTF induced widespread nuclear p-STAT3 immunoreactivity in muscle fibers and in connective and Schwann cells of the peripheral nerves, including the sciatic nerve, supplying the gastrocnemius. In conclusion, our data indicate that CNTF acts across diverse cytotypes within metabolically relevant organs and tissues, likely fostering its peripheral metabolic effects through this cellular heterogeneity.

O_LIPlants and fungi are major sources of natural products beneficial to society, making the study of distinct species essential for discovering new drugs and bioactive compounds. The medicinal mushroom "Lingzhi" or "Reishi" (Ganoderma lingzhi) is widely used in traditional medicine and extensively studied for its bioactive triterpenoids, yet it is commonly identified as Ganoderma lucidum, the type species of the genus, which lacks a type specimen. C_LIO_LIWe sequenced a G. lucidum specimen preserved in the Kew fungarium, which agreed with the original description and was collected from wood of Corylus avellana in southern England. Using this reference specimen, we compiled genomic and ITS barcoding datasets to explore the genetic and geographic variation within this species. C_LIO_LIWe showed that G. lingzhi and G. lucidum diverged more than 12 million years ago and that all seven "G. lucidum" genomes deposited in public databases belong to other species. More than 1000 barcoding sequences showed that the widely used homology-based ITS barcoding is not working in this group, which can be mitigated by a phylogenetic placement approach. The 149 sequences assigned to G. lucidum with high confidence showed a Eurasian distribution and introductions to North and South America and Africa. C_LIO_LIOur study underscores the importance of accurate species identification and provides guidance for a group of pharmaceutical and socially significant species. To further support future studies and the wider public in differentiating between G. lingzhi and G. lucidum, we propose using "False Lingzhi" as the English name for G. lucidum. C_LI Societal Impact StatementTraditional Chinese Medicine has expanded far beyond Asia, with growing markets in North America and Europe for supplements and functional foods. Lingzhi or Reishi (Ganoderma lingzhi), a well-known medicinal mushroom, is valued for its anti-inflammatory and anticancer properties. However, it is often misidentified with species that may not provide the same health benefits. This confusion poses risks to consumer safety, product regulation, and research. Here, we establish a reference using morphological and molecular tools for the most commonly misidentified species (Ganoderma lucidum) and propose the name "False Lingzhi" to distinguish it, supporting accurate identification, safer product development, and reliable research.

(1) BackgroundSaffron crocus (Crocus sativus) is the source of saffron, the most expensive spice in the world. It evolved about 3000 years ago as a sterile triploid clone in Greece. Since then, saffron has spread across the globe, where regionally distinct practices of saffron cultivation have developed. Despite differences in morpho-physiological traits, genetic variability is low, if present at all. Here, we aim to resolve chromosomal and sequence-associated variability across saffron crocus cultivars from the crops main cultivation areas in Africa, Asia and Europe. (2) MethodsWe used genome-wide DNA polymorphisms obtained through genotyping-by-sequencing (GBS) of 33 saffron and 14 closely related Crocus accessions, which we place into a phylogenetic context. For karyotyping, we compare nine saffron accessions by multi-color fluorescent in situ hybridisation (FISH) with repetitive DNA probes. (3) Key resultsPhylogenetic analyses confirmed the single origin and clonal nature of all saffron accessions. We detected slight DNA differences among saffron crocus genotypes, which were minor compared with those in wild C. cartwrightianus populations. Still, the Iranian saffron accessions form a genetically very narrow group that differs from the other proveniences in population genetic analyses. However, chromosomes of some saffron accessions display variable FISH signals, likely resulting from gains and losses of tandemly repeated DNA. (4) Main conclusionsBased on the high genetic identity and small karyotypic differences, we confirm the clonal origin of the saffron accessions. Nevertheless, as we detected small and regional chromosomal variability, we conclude that at least four somaclonal saffron lineages emerged after saffrons origin. Societal Impact StatementFor millennia, many cultures developed cultivation practices and regional crop varieties. A notable case is saffron, the worlds most expensive spice that is harvested from stigmas of saffron crocus. This flower crop arose 3000 years ago in a singular genome triplication event and since then spread clonally across the globe. By identifying genetic and chromosomal variability in clonal saffron accessions, we highlight regional diversity, support the preservation of traditional knowledge, and underscore the risk of relying on only one clonal lineage. This informs strategies for saffron cultivation, linking cultural heritage with modern genomics to address biodiversity, evolution, and food security.

BackgroundThe use of autologous or allogeneic cell therapies has now entered to the clinical practice in several fields of medicine, especially in oncology and hematology. From this regard, 2D-cell manufacturing is complex and costly and bioreactors have attracted major interest for efficient and cost-effective mass production of cells. Bioreactors have several advantages such as homogeneous repartition of nutrients and gas, control of all culture parameters and increased yield. However, the important shear stress generated by those bioreactors is an important disadvantage as it can affect cell survival or cell quality. This important shear stress is the result of the mixing method using either blades (used in stirred-tanked bioreactors) or gas bubbles (used in airlift bioreactors). Another downside of the use of bioreactors is the difficulty to scale-up. As the volume increases, the shear stress generated by blades radically increases leading to cell death and a decrease of cell quality. DescriptionIn this study, we describe a bioreactor developed using a different mixing method effectively reducing the shear stress and facilitating scale-up. This bladeless method uses an inclination of the bioreactor as well as rotation to mix fluids in a container. Here we described different steps that led to the adaptation of this bioreactor, initially developed for fragile microalgae culture, for mammalian cell culture amplification. The bioreactor was tested to amplify a natural killer (NK) cell line NK92 which is an IL-2 dependent cell line used in clinical trials for cancer therapy. We have tested the influence of 1-The number of cells seeded; 2-The influence of the rotation speed on cell growth and viability; 3-The influence of the bioreactor angle on the above parameters; 4-The duration of the culture. ResultsCells were initially seeded at 2.5.105 / ml in a volume of 380 ml. According to the rotation speed of 15, 30, 45 and 60 rpm, we have observed an increase of cell numbers at day 3 (3-fold), day 5 (7-fold) and day 7 (10-fold) compared to seeding, the best expansion being obtained at day 7 with a rotation speed of 45 rpm. The optimal angle of rotation was found to be 3 degree, with an optimal amplification at day 7 versus day 3 (p < 0.01). The viability was also found to be optimal in the latter condition. ConclusionsThese preliminary results demonstrate that NK92 cells could be amplified using this bioreactor. In the best tested condition, neither cell viability nor cell growth was impacted. These results strongly suggest the potential use of this device in future clinically applicable conditions.