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18 pages, 7479 KiB  
Article
Development and Validation of a Custom-Built System for Real-Time Monitoring of In Vitro Rumen Gas Fermentation
by Zhen-Shu Liu, Bo-Yuan Chen, Jacky Peng-Wen Chan and Po-Wen Chen
Animals 2025, 15(15), 2308; https://doi.org/10.3390/ani15152308 - 6 Aug 2025
Abstract
While the Ankom RF system facilitates efficient high-throughput in vitro fermentation studies, its high cost and limited flexibility constrain its broader applicability. To address these limitations, we developed and validated a low-cost, modular gas monitoring system (FerME), assembled from commercially available components. To [...] Read more.
While the Ankom RF system facilitates efficient high-throughput in vitro fermentation studies, its high cost and limited flexibility constrain its broader applicability. To address these limitations, we developed and validated a low-cost, modular gas monitoring system (FerME), assembled from commercially available components. To evaluate its performance and reproducibility relative to the Ankom RF system (Ankom Technology, Macedon, NY, USA), in vitro rumen fermentation experiments were conducted under strictly controlled and identical conditions. Whole rumen contents were collected approximately 2 h post-feeding from individual mid- or late-lactation dairy cows and immediately transported to the laboratory. Each fermenter received 50 mL of processed rumen fluid, 100 mL of anaerobically prepared artificial saliva buffer, and 1.2 g of the donor cow’s diet. Bottles were sealed with the respective system’s pressure sensors, flushed with CO2, and incubated in a 50 L water bath maintained at 39 °C. FerME (New Taipei City, Taiwan) and Ankom RF fermenters were placed side-by-side to ensure uniform thermal conditions. To assess the effect of filter bag use, an additional trial employed Ankom F57 filter bags (Ankom Technology, Macedon, NY, USA; 25 μm pore size). Trial 1 revealed no significant differences in cumulative gas production, volatile fatty acids (VFAs), NH3-N, or pH between systems (p > 0.05). However, the use of filter bags reduced gas output and increased propionate concentrations (p < 0.05). Trial 2, which employed filter bags in both systems, confirmed comparable results, with the FerME system demonstrating improved precision (CV: 4.8% vs. 13.2%). Gas composition (CH4 + CO2: 76–82%) and fermentation parameters remained consistent across systems (p > 0.05). Importantly, with 12 pressure sensors, the total cost of FerME was about half that of the Ankom RF system. Collectively, these findings demonstrate that FerME is a reliable, low-cost alternative for real-time rumen fermentation monitoring and could be suitable for studies in animal nutrition, methane mitigation, and related applications. Full article
(This article belongs to the Section Animal System and Management)
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16 pages, 2650 KiB  
Article
Inhibition of Tyrosinase and Melanogenesis by a White Mulberry Fruit Extract
by Nuttawadee Prasawang, Nareerat Sutjarit, Athisri Sitthipunya, Prasit Suwannalert, Wutarak Monsuwan and Nisamanee Charoenchon
Int. J. Mol. Sci. 2025, 26(15), 7589; https://doi.org/10.3390/ijms26157589 - 6 Aug 2025
Abstract
Ultraviolet B (UVB) radiation is a key factor in the overproduction of melanin in the skin. Melanocytes produce melanin through melanogenesis to protect the skin from UVB radiation-induced damage. However, excessive melanogenesis can lead to hyperpigmentation and increase the risk of malignant melanoma. [...] Read more.
Ultraviolet B (UVB) radiation is a key factor in the overproduction of melanin in the skin. Melanocytes produce melanin through melanogenesis to protect the skin from UVB radiation-induced damage. However, excessive melanogenesis can lead to hyperpigmentation and increase the risk of malignant melanoma. Tyrosinase is the rate-limiting enzyme in melanogenesis; it catalyzes the oxidation of tyrosine to 3,4-dihydroxy-L-phenylalanine and subsequently to dopaquinone. Thus, inhibiting tyrosinase is a promising strategy for preventing melanogenesis and skin hyperpigmentation. White mulberry (Morus alba L.) is rich in antioxidants, and mulberry fruit extracts have been used as cosmetic skin-lightening agents. However, data on the capacity of mulberry fruit extracts to inhibit tyrosinase under UVB radiation-induced melanogenic conditions remain scarce, especially in an in vivo model. In this study, we evaluated the effects of a mulberry crude extract (MCE) on UVB radiation-induced melanogenesis in B16F10 melanoma cells and zebrafish embryos. The MCE significantly reduced tyrosinase activity and melanogenesis in a dose-dependent manner without inducing cytotoxicity. These effects are likely attributable to the antioxidant constituents of the extract. Our findings highlight the potential of this MCE as an effective tyrosinase inhibitor for the prevention of UVB radiation-induced skin hyperpigmentation. Full article
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17 pages, 1396 KiB  
Article
Dose-Dependent Effect of the Polyamine Spermine on Wheat Seed Germination, Mycelium Growth of Fusarium Seed-Borne Pathogens, and In Vivo Fusarium Root and Crown Rot Development
by Tsvetina Nikolova, Dessislava Todorova, Tzenko Vatchev, Zornitsa Stoyanova, Valya Lyubenova, Yordanka Taseva, Ivo Yanashkov and Iskren Sergiev
Agriculture 2025, 15(15), 1695; https://doi.org/10.3390/agriculture15151695 - 6 Aug 2025
Abstract
Wheat (Triticum aestivum L.) is a crucial global food crop. The intensive crop farming, monoculture cultivation, and impact of climate change affect the susceptibility of wheat cultivars to biotic stresses, mainly caused by soil fungal pathogens, especially those belonging to the genus [...] Read more.
Wheat (Triticum aestivum L.) is a crucial global food crop. The intensive crop farming, monoculture cultivation, and impact of climate change affect the susceptibility of wheat cultivars to biotic stresses, mainly caused by soil fungal pathogens, especially those belonging to the genus Fusarium. This situation threatens yield and grain quality through root and crown rot. While conventional chemical fungicides face resistance issues and environmental concerns, biological alternatives like seed priming with natural metabolites are gaining attention. Polyamines, including putrescine, spermidine, and spermine, are attractive priming agents influencing plant development and abiotic stress responses. Spermine in particular shows potential for in vitro antifungal activity against Fusarium. Optimising spermine concentration for seed priming is crucial to maximising protection against Fusarium infection while ensuring robust plant growth. In this research, we explored the potential of the polyamine spermine as a seed treatment to enhance wheat resilience, aiming to identify a sustainable alternative to synthetic fungicides. Our findings revealed that a six-hour seed soak in spermine solutions ranging from 0.5 to 5 mM did not delay germination or seedling growth. In fact, the 5 mM concentration significantly stimulated root weight and length. In complementary in vitro assays, we evaluated the antifungal activity of spermine (0.5–5 mM) against three Fusarium species. The results demonstrated complete inhibition of Fusarium culmorum growth at 5 mM spermine. A less significant effect on Fusarium graminearum and little to no impact on Fusarium oxysporum were found. The performed analysis revealed that the spermine had a fungistatic effect against the pathogen, retarding the mycelium growth of F. culmorum inoculated on the seed surface. A pot experiment with Bulgarian soft wheat cv. Sadovo-1 was carried out to estimate the effect of seed priming with spermine against infection with isolates of pathogenic fungus F. culmorum on plant growth and disease severity. Our results demonstrated that spermine resulted in a reduced distribution of F. culmorum and improved plant performance, as evidenced by the higher fresh weight and height of plants pre-treated with spermine. This research describes the efficacy of spermine seed priming as a novel strategy for managing Fusarium root and crown rot in wheat. Full article
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22 pages, 2517 KiB  
Article
Characterization and Engineering of Two Novel Strand-Displacing B Family DNA Polymerases from Bacillus Phage SRT01hs and BeachBum
by Yaping Sun, Kang Fu, Wu Lin, Jie Gao, Xianhui Zhao, Yun He and Hui Tian
Biomolecules 2025, 15(8), 1126; https://doi.org/10.3390/biom15081126 - 5 Aug 2025
Viewed by 73
Abstract
Polymerase-coupled nanopore sequencing requires DNA polymerases with strong strand displacement activity and high processivity to sustain continuous signal generation. In this study, we characterized two novel B family DNA polymerases, SRHS and BBum, isolated from Bacillus phages SRT01hs and BeachBum, respectively. Both enzymes [...] Read more.
Polymerase-coupled nanopore sequencing requires DNA polymerases with strong strand displacement activity and high processivity to sustain continuous signal generation. In this study, we characterized two novel B family DNA polymerases, SRHS and BBum, isolated from Bacillus phages SRT01hs and BeachBum, respectively. Both enzymes exhibited robust strand displacement, 3′→5′ exonuclease activity, and maintained processivity under diverse reaction conditions, including across a broad temperature range (10–45 °C) and in the presence of multiple divalent metal cofactors (Mg2+, Mn2+, Fe2+), comparable to the well-characterized Phi29 polymerase. Through biochemical analysis of mutants designed using AlphaFold3-predicted structural models, we identified key residues (G96, M97, D486 in SRHS; S97, M98, A493 in BBum) that modulated exonuclease activity, substrate specificity and metal ion utilization. Engineered variants SRHS_F and BBum_Pro_L efficiently incorporated unnatural nucleotides in the presence of Mg2+—a function not observed in Phi29 and other wild-type strand-displacing B family polymerases. These combined biochemical features highlight SRHS and BBum as promising enzymatic scaffolds for nanopore-based long-read sequencing platforms. Full article
(This article belongs to the Section Enzymology)
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21 pages, 10626 KiB  
Article
Comparative Metabolomic Analysis Reveals Tissue- and Species-Specific Differences in the Abundance of Dammarane-Type Ginsenosides in Three Panax Species
by Shu He, Ying Gong, Shuangfei Deng, Yaquan Dou, Junmin Wang, Hoang Van Sam, Xingliang Chen, Xiahong He and Rui Shi
Horticulturae 2025, 11(8), 916; https://doi.org/10.3390/horticulturae11080916 (registering DOI) - 5 Aug 2025
Viewed by 160
Abstract
The genus Panax contains traditional herbs that have been widely used in traditional medicine. The active constituents, collectively known as ginsenosides, are well characterized in the most representative species, P. notoginseng. However, the major bioactive chemical constituents of P. stipuleanatus together with [...] Read more.
The genus Panax contains traditional herbs that have been widely used in traditional medicine. The active constituents, collectively known as ginsenosides, are well characterized in the most representative species, P. notoginseng. However, the major bioactive chemical constituents of P. stipuleanatus together with P. vietnamensis are relatively less studied. In this study, an untargeted metabolomic analysis was performed in P. notoginseng, P. stipuleanatus, and P. vietnamensis using root and leaf organs. Further metabolomic differences in P. stipuleanatus were compared with those of the two most prevalent species. The analysis results revealed tissue-specific qualitative and quantitative metabolic differences in each species. Several differentially accumulated metabolites were enriched in the biosynthesis of secondary metabolites, including the biosynthesis of ginsenosides I. The ginsenosides Rb1, Rf, Rg1, Rh1, Rh8, and notoginsenosides E, M, and N had a higher abundance level in the roots of both P. notoginseng and P. vietnamensis. In P. stipuleanatus, the accumulation of potentially important ginsenosides is mainly found in the leaf. In particular, the dammarane-type ginsenosides Rb3, Rb1, Mx, and F2 as well as the notoginsenosides A, Fe, Fa, Fd, L, and N were identified to have a higher accumulation in the leaf. The strong positive correlation network of different ginsenosides probably enhanced secondary metabolism in each species. The comparative analysis revealed a significant differential accumulation of metabolites in the leaves of both species. The various compounds of dammarane-type ginsenoside, such as Rb1, Rg1, Rg6, Rh8, Rh10, Rh14, and majoroside F2, had a significantly higher concentration level in the leaves of P. stipuleanatus. In addition, several notoginsenoside compounds such as A, R1, Fe, Fd, and Ft1 showed a higher abundance in the leaf. These results show that the abundance level of major ginsenosides is significant in P. stipuleanatus and provides an important platform to improve the ginsenoside quality of Panax species. Full article
(This article belongs to the Section Medicinals, Herbs, and Specialty Crops)
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12 pages, 472 KiB  
Communication
LAMPOX: A Portable and Rapid Molecular Diagnostic Assay for the Epidemic Clade IIb Mpox Virus Detection
by Anna Rosa Garbuglia, Mallory Draye, Silvia Pauciullo, Daniele Lapa, Eliana Specchiarello, Florence Nazé and Pascal Mertens
Diagnostics 2025, 15(15), 1959; https://doi.org/10.3390/diagnostics15151959 - 4 Aug 2025
Viewed by 178
Abstract
The global spread of Mpox virus (MPXV) underscores the urgent need for rapid, field-deployable diagnostic tools, especially in low-resource settings. We evaluated a loop-mediated isothermal amplification (LAMP) assay, termed LAMPOX, developed by Coris BioConcept. The assay was tested in three formats—two liquid versions [...] Read more.
The global spread of Mpox virus (MPXV) underscores the urgent need for rapid, field-deployable diagnostic tools, especially in low-resource settings. We evaluated a loop-mediated isothermal amplification (LAMP) assay, termed LAMPOX, developed by Coris BioConcept. The assay was tested in three formats—two liquid versions and a dried, ready-to-use version—targeting only the ORF F3L (Liquid V1) or both the ORF F3L and N4R (Liquid V2 and dried) genomic regions. Analytical sensitivity and specificity were assessed using 60 clinical samples from confirmed MPXV-positive patients. Sensitivity on clinical samples was 81.7% for Liquid V1 and 88.3% for Liquid V2. The dried LAMPOX assay demonstrated a sensitivity of 88.3% and a specificity of 100% in a panel of 112 negative controls, with most positive samples detected in under 7 min. Additionally, a simplified sample lysis protocol was developed to facilitate point-of-care use. While this method showed slightly reduced sensitivity compared to standard DNA extraction, it proved effective for samples with higher viral loads. The dried format offers key advantages, including ambient-temperature stability and minimal equipment needs, making it suitable for point-of-care testing. These findings support LAMPOX as a promising tool for rapid MPXV detection during outbreaks, especially in resource-limited settings where traditional PCR is impractical. Full article
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20 pages, 3354 KiB  
Article
An Assessment of the Population Structure and Stock Dynamics of Megalobrama skolkovii During the Early Phase of the Fishing Ban in the Poyang Lake Basin
by Xinwen Huang, Qun Xu, Bao Zhang, Chiping Kong, Lei Fang, Xiaoping Gao, Leyi Sun, Lekang Li and Xiaoling Gong
Fishes 2025, 10(8), 378; https://doi.org/10.3390/fishes10080378 - 4 Aug 2025
Viewed by 90
Abstract
The ten-year fishing ban on the Yangtze River aims to restore aquatic biodiversity and rebuild fishery resources. Megalobrama skolkovii, a key species in the basin, was investigated using 2024 data to provide a preliminary assessment of its population structure, stock dynamics, and [...] Read more.
The ten-year fishing ban on the Yangtze River aims to restore aquatic biodiversity and rebuild fishery resources. Megalobrama skolkovii, a key species in the basin, was investigated using 2024 data to provide a preliminary assessment of its population structure, stock dynamics, and early recovery. Age analysis (n = 243) showed that 1–6-year-olds were dominated by fish aged 3 (35%), with few older than 4, indicating moderate structural truncation. Growth parameters modeled by the von Bertalanffy Growth Function yielded L = 61.89 cm and k = 0.25 year1, with a weight–growth inflection age of 4.4 years. Natural mortality (M = 0.48 year−1) was estimated using Pauly’s empirical formula, and total mortality (Z = 0.55 year−1) was estimated from the catch curve analysis. While fishing mortality (F) was statistically indistinguishable from zero, a plausible low-intensity fishing scenario was explored to assess potential impacts of residual activities. Length-based indicators (LBIs) showed Pmat = 46.05%, Popt = 9.51%, and Pmega = 6.88%, suggesting reproductive recovery but incomplete structural restoration. These preliminary findings reveal an asymmetrical recovery trajectory, whereby physiological improvements and enhanced recruitment have occurred, yet full structural restoration remains incomplete. This underscores the need for continued, long-term conservation and monitoring to support population resilience. Full article
(This article belongs to the Section Biology and Ecology)
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15 pages, 24657 KiB  
Article
Identification and Genetic Analysis of Downy Mildew Resistance in Intraspecific Hybrids of Vitis vinifera L.
by Xing Han, Yihan Li, Zhilei Wang, Zebin Li, Nanyang Li, Hua Li and Xinyao Duan
Plants 2025, 14(15), 2415; https://doi.org/10.3390/plants14152415 - 4 Aug 2025
Viewed by 137
Abstract
Downy mildew caused by Plasmopara viticola is an important disease in grape production, particularly in the highly susceptible, widely cultivated Vitis vinifera L. Breeding for disease resistance is an effective solution, and V. vinifera intraspecific crosses can yield progeny with both disease resistance [...] Read more.
Downy mildew caused by Plasmopara viticola is an important disease in grape production, particularly in the highly susceptible, widely cultivated Vitis vinifera L. Breeding for disease resistance is an effective solution, and V. vinifera intraspecific crosses can yield progeny with both disease resistance and high quality. To assess the potential of intraspecific recurrent selection in V. vinifera (IRSV) in improving grapevine resistance to downy mildew and to analyze the pattern of disease resistance inheritance, the disease-resistant variety Ecolly was selected as one of the parents and crossed with Cabernet Sauvignon, Marselan, and Dunkelfelder, respectively, creating three reciprocal combinations, resulting in 1657 hybrid F1 progenies. The primary results are as follows: (1) significant differences in disease resistance among grape varieties and, significant differences in disease resistance between different vintages of the same variety were found; (2) the leaf downy mildew resistance levels of F1 progeny of different hybrid combinations conformed to a skewed normal distribution and showed some maternal dominance; (3) the degree of leaf bulbous elevation was negatively correlated with the level of leaf downy mildew resistance, and the correlation coefficient with the level of field resistance was higher; (4) five progenies with higher levels of both field and in vitro disease resistance were obtained. Intraspecific hybridization can improve the disease resistance of offspring through super-parent genetic effects, and Ecolly can be used as breeding material for recurrent hybridization to obtain highly resistant varieties. Full article
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16 pages, 1313 KiB  
Article
Mycorrhizas Promote Total Flavonoid Levels in Trifoliate Orange by Accelerating the Flavonoid Biosynthetic Pathway to Reduce Oxidative Damage Under Drought
by Lei Liu and Hong-Na Mu
Horticulturae 2025, 11(8), 910; https://doi.org/10.3390/horticulturae11080910 (registering DOI) - 4 Aug 2025
Viewed by 137
Abstract
Flavonoids serve as crucial plant antioxidants in drought tolerance, yet their antioxidant regulatory mechanisms within mycorrhizal plants remain unclear. In this study, using a two-factor design, trifoliate orange (Poncirus trifoliata (L.) Raf.) seedlings in the four-to-five-leaf stage were either inoculated with Funneliformis [...] Read more.
Flavonoids serve as crucial plant antioxidants in drought tolerance, yet their antioxidant regulatory mechanisms within mycorrhizal plants remain unclear. In this study, using a two-factor design, trifoliate orange (Poncirus trifoliata (L.) Raf.) seedlings in the four-to-five-leaf stage were either inoculated with Funneliformis mosseae or not, and subjected to well-watered (70–75% of field maximum water-holding capacity) or drought stress (50–55% field maximum water-holding capacity) conditions for 10 weeks. Plant growth performance, photosynthetic physiology, leaf flavonoid content and their antioxidant capacity, reactive oxygen species levels, and activities and gene expression of key flavonoid biosynthesis enzymes were analyzed. Although drought stress significantly reduced root colonization and soil hyphal length, inoculation with F. mosseae consistently enhanced the biomass of leaves, stems, and roots, as well as root surface area and diameter, irrespective of soil moisture. Despite drought suppressing photosynthesis in mycorrhizal plants, F. mosseae substantially improved photosynthetic capacity (measured via gas exchange) and optimized photochemical efficiency (assessed by chlorophyll fluorescence) while reducing non-photochemical quenching (heat dissipation). Inoculation with F. mosseae elevated the total flavonoid content in leaves by 46.67% (well-watered) and 14.04% (drought), accompanied by significantly enhanced activities of key synthases such as phenylalanine ammonia-lyase (PAL), chalcone synthase (CHS), chalcone isomerase (CHI), 4-coumarate:coA ligase (4CL), and cinnamate 4-hydroxylase (C4H), with increases ranging from 16.90 to 117.42% under drought. Quantitative real-time PCR revealed that both mycorrhization and drought upregulated the expression of PtPAL1, PtCHI, and Pt4CL genes, with soil moisture critically modulating mycorrhizal regulatory effects. In vitro assays showed that flavonoid extracts scavenged radicals at rates of 30.07–41.60% in hydroxyl radical (•OH), 71.89–78.06% in superoxide radical anion (O2•−), and 49.97–74.75% in 2,2-diphenyl-1-picrylhydrazyl (DPPH). Mycorrhizal symbiosis enhanced the antioxidant capacity of flavonoids, resulting in higher scavenging rates of •OH (19.07%), O2•− (5.00%), and DPPH (31.81%) under drought. Inoculated plants displayed reduced hydrogen peroxide (19.77%), O2•− (23.90%), and malondialdehyde (17.36%) levels. This study concludes that mycorrhizae promote the level of total flavonoids in trifoliate orange by accelerating the flavonoid biosynthesis pathway, hence reducing oxidative damage under drought. Full article
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26 pages, 486 KiB  
Article
Towards Characterizing the Download Cost of Cache-Aided Private Updating
by Bryttany Stark, Ahmed Arafa and Karim Banawan
Entropy 2025, 27(8), 828; https://doi.org/10.3390/e27080828 - 4 Aug 2025
Viewed by 192
Abstract
We consider the problem of privately updating a message out of K messages from N replicated and non-colluding databases where a user has an outdated version of the message W^θ of length L bits that differ from the current version [...] Read more.
We consider the problem of privately updating a message out of K messages from N replicated and non-colluding databases where a user has an outdated version of the message W^θ of length L bits that differ from the current version Wθ in at most f bits. The user also has a cache containing coded combinations of the K messages (with a pre-specified structure), which are unknown to the N databases (unknown prefetching). The cache Z contains linear combinations from all K messages in the databases with r=lL being the caching ratio. The user needs to retrieve Wθ correctly using a private information retrieval (PIR) scheme without leaking information about the message index θ to any individual database. Our objective is to jointly design the prefetching (i.e., the structure of said linear combinations) and the PIR strategies to achieve the least download cost. We propose a novel achievable scheme based on syndrome decoding where the cached linear combinations in Z are designed to be bits pertaining to the syndrome of Wθ according to a specific linear block code. We derive a general lower bound on the optimal download cost for 0r1, in addition to achievable upper bounds. The upper and lower bounds match for the cases when r is exceptionally low or high, or when K=3 messages for arbitrary r. Such bounds are derived by developing novel cache-aided arbitrary message length PIR schemes. Our results show a significant reduction in the download cost if f<L2 when compared with downloading Wθ directly using typical cached-aided PIR approaches. Full article
(This article belongs to the Special Issue Information-Theoretic Security and Privacy)
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25 pages, 4241 KiB  
Article
Deep Learning for Comprehensive Analysis of Retinal Fundus Images: Detection of Systemic and Ocular Conditions
by Mohammad Mahdi Aghabeigi Alooghareh, Mohammad Mohsen Sheikhey, Ali Sahafi, Habibollah Pirnejad and Amin Naemi
Bioengineering 2025, 12(8), 840; https://doi.org/10.3390/bioengineering12080840 - 3 Aug 2025
Viewed by 327
Abstract
The retina offers a unique window into both ocular and systemic health, motivating the development of AI-based tools for disease screening and risk assessment. In this study, we present a comprehensive evaluation of six state-of-the-art deep neural networks, including convolutional neural networks and [...] Read more.
The retina offers a unique window into both ocular and systemic health, motivating the development of AI-based tools for disease screening and risk assessment. In this study, we present a comprehensive evaluation of six state-of-the-art deep neural networks, including convolutional neural networks and vision transformer architectures, on the Brazilian Multilabel Ophthalmological Dataset (BRSET), comprising 16,266 fundus images annotated for multiple clinical and demographic labels. We explored seven classification tasks: Diabetes, Diabetic Retinopathy (2-class), Diabetic Retinopathy (3-class), Hypertension, Hypertensive Retinopathy, Drusen, and Sex classification. Models were evaluated using precision, recall, F1-score, accuracy, and AUC. Among all models, the Swin-L generally delivered the best performance across scenarios for Diabetes (AUC = 0.88, weighted F1-score = 0.86), Diabetic Retinopathy (2-class) (AUC = 0.98, weighted F1-score = 0.95), Diabetic Retinopathy (3-class) (macro AUC = 0.98, weighted F1-score = 0.95), Hypertension (AUC = 0.85, weighted F1-score = 0.79), Hypertensive Retinopathy (AUC = 0.81, weighted F1-score = 0.97), Drusen detection (AUC = 0.93, weighted F1-score = 0.90), and Sex classification (AUC = 0.87, weighted F1-score = 0.80). These results reflect excellent to outstanding diagnostic performance. We also employed gradient-based saliency maps to enhance explainability and visualize decision-relevant retinal features. Our findings underscore the potential of deep learning, particularly vision transformer models, to deliver accurate, interpretable, and clinically meaningful screening tools for retinal and systemic disease detection. Full article
(This article belongs to the Special Issue Machine Learning in Chronic Diseases)
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18 pages, 2432 KiB  
Article
Alkali Lignin-Based Biopolymer Formulations for Electro-Assisted Drug Delivery of Natural Antioxidants in Breast Cancer Cells—A Preliminary Study
by Severina Semkova, Radina Deneva, Georgi Antov, Donika Ivanova and Biliana Nikolova
Int. J. Mol. Sci. 2025, 26(15), 7481; https://doi.org/10.3390/ijms26157481 - 2 Aug 2025
Viewed by 271
Abstract
Recently, a number of natural biologically active substances have been proven to be attractive alternatives to conventional anticancer medicine or as adjuvants in contemporary combination therapies. Although lignin-based materials were previously accepted as waste materials with limited usefulness, recent studies increasingly report the [...] Read more.
Recently, a number of natural biologically active substances have been proven to be attractive alternatives to conventional anticancer medicine or as adjuvants in contemporary combination therapies. Although lignin-based materials were previously accepted as waste materials with limited usefulness, recent studies increasingly report the possibility of their use for novel applications in various industrial branches, including biomedicine. In this regard, the safety, efficiency, advantages and limitations of lignin compounds for in vitro/in vivo applications remain poorly studied and described. This study was carried out to investigate the possibility of using newly synthesized, alkali lignin-based micro-/nano-biopolymer formulations (Lignin@Formulations/L@F) as carriers for substances with antioxidant and/or anticancer effectiveness. Moreover, we tried to assess the opportunity for using an electro-assisted approach for achieving improved intracellular internalization. An investigation was conducted on an in vitro panel of breast cell lines, namely two breast cancer lines with different metastatic potentials and one non-tumorigenic line as a control. The characterization of all tested formulations was performed via DLS (dynamic light scattering) analysis. We developed an improved separation procedure via size/charge unification for all types of Lignin@Formulations. Moreover, in vitro applications were investigated. The results demonstrate that compared to healthy breast cells, both tested cancer lines exhibited slight sensitivity after treatment with different formulations (empty or loaded with antioxidant substances). This effect was also enhanced after applying electric pulses. L@F loaded with Quercetin was also explored only on the highly metastatic cancer cell line as a model for the breast cancer type most aggressive and non-responsive to traditional treatments. All obtained data suggest that the tested formulations have potential as carriers for the electro-assisted delivery of natural antioxidants such as Quercetin. Full article
(This article belongs to the Special Issue Natural Products in Cancer Prevention and Treatment)
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20 pages, 462 KiB  
Article
Chromatographic and Chemometric Characterization of the Two Wild Edible Mushrooms Fistulina hepatica and Clitocybe nuda: Insights into Nutritional, Phenolic, and Antioxidant Profiles
by Ana Saldanha, Mikel Añibarro-Ortega, Adriana K. Molina, José Pinela, Maria Inês Dias and Carla Pereira
Separations 2025, 12(8), 204; https://doi.org/10.3390/separations12080204 - 2 Aug 2025
Viewed by 221
Abstract
Fistulina hepatica (Schaeff.) With. and Clitocybe nuda (Bull.) H.E. Bigelow & A.H. Sm. are wild edible mushrooms with nutritional and functional potential that remain insufficiently characterized. This study provides the first comparative assessment of their nutritional profiles, phenolic composition, and antioxidant activity, using [...] Read more.
Fistulina hepatica (Schaeff.) With. and Clitocybe nuda (Bull.) H.E. Bigelow & A.H. Sm. are wild edible mushrooms with nutritional and functional potential that remain insufficiently characterized. This study provides the first comparative assessment of their nutritional profiles, phenolic composition, and antioxidant activity, using specimens collected from Montesinho Natural Park (Portugal). Proximate composition, organic and phenolic acids, free sugars, and fatty acids were analyzed by chromatographic methods, and antioxidant capacity was assessed through OxHLIA and TBARS assays. F. hepatica showed higher carbohydrates (9.3 ± 0.2 g/100 g fw) and estimated energy values (43 ± 1 kcal/100 g fw), increased phenolic acids content (2.7 ± 0.1 mg/g extract), and the exclusive presence of p-coumaric and cinnamic acids, along with OxHLIA activity (IC50 = 126 ± 5 µg/mL at Δt = 60 min). C. nuda displayed higher protein (2.5 ± 0.1 g/100 g dw) and quinic acid contents (4.13 ± 0.02 mg/g extract), a PUFA-rich profile, and greater TBARS inhibition (EC50 = 303 ± 17 µg/mL). These findings highlight distinct and complementary bioactive traits, supporting their valorization as natural functional ingredients. Their compositional features offer promising applications in sustainable food systems and nutraceutical development, encouraging further investigations into safety, bioaccessibility, and formulation strategies. Notably, F. hepatica is best consumed at a young developmental stage, as its sensory properties tend to decline with maturity. Full article
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23 pages, 2268 KiB  
Article
Potential for Drought Stress Alleviation in Lettuce (Lactuca sativa L.) with Humic Substance-Based Biostimulant Applications
by Santiago Atero-Calvo, Francesco Magro, Giacomo Masetti, Eloy Navarro-León, Begoña Blasco and Juan Manuel Ruiz
Plants 2025, 14(15), 2386; https://doi.org/10.3390/plants14152386 - 2 Aug 2025
Viewed by 282
Abstract
In the present study, we evaluated the potential use of a humic substance (HS)-based biostimulant in mitigating drought stress in lettuce (Lactuca sativa L.) by comparing both root and foliar modes of application. To achieve this, lettuce plants were grown in a [...] Read more.
In the present study, we evaluated the potential use of a humic substance (HS)-based biostimulant in mitigating drought stress in lettuce (Lactuca sativa L.) by comparing both root and foliar modes of application. To achieve this, lettuce plants were grown in a growth chamber on a solid substrate composed of vermiculite and perlite (3:1). Plants were exposed to drought conditions (50% of Field Capacity, FC) and 50% FC + HS applied as radicular (‘R’) and foliar (‘F’) at concentrations: R-HS 0.40 and 0.60 mL/L, respectively, and 7.50 and 10.00 mL/L, respectively, along with a control (100% FC). HSs were applied three times at 10-day intervals. Plant growth, nutrient concentration, lipid peroxidation, reactive oxygen species (ROS), and enzymatic and non-enzymatic antioxidants were estimated. Various photosynthetic and chlorophyll fluorescence parameters were also analyzed. The results showed that HS applications alleviated drought stress, increased plant growth, and reduced lipid peroxidation and ROS accumulation. HSs also improved the net photosynthetic rate, carboxylation efficiency, electron transport flux, and water use efficiency. Although foliar HSs showed a greater tendency to enhance shoot growth and photosynthetic capacity, the differences between the application methods were not significant. Hence, in this preliminary work, the HS-based product evaluated in this study demonstrated potential for alleviating drought stress in lettuce plants at the applied doses, regardless of the mode of application. This study highlights HS-based biostimulants as an effective and sustainable tool to improve crop resilience and support sustainable agriculture under climate change. However, further studies under controlled growth chamber conditions are needed to confirm these results before field trials. Full article
(This article belongs to the Special Issue Biostimulation for Abiotic Stress Tolerance in Plants)
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19 pages, 993 KiB  
Article
Antibacterial Properties of Submerged Cultivated Fomitopsis pinicola, Targeting Gram-Negative Pathogens, Including Borrelia burgdorferi
by Olga Bragina, Maria Kuhtinskaja, Vladimir Elisashvili, Mikheil Asatiani and Maria Kulp
Sci 2025, 7(3), 104; https://doi.org/10.3390/sci7030104 - 2 Aug 2025
Viewed by 161
Abstract
The rise in multidrug-resistant bacterial strains and persistent infections such as Lyme disease caused by Borrelia burgdorferi highlights the need for novel antimicrobial agents. The present study explores the antioxidant, antibacterial, and cytotoxic properties of extracts from submerged mycelial biomass of Fomitopsis pinicola [...] Read more.
The rise in multidrug-resistant bacterial strains and persistent infections such as Lyme disease caused by Borrelia burgdorferi highlights the need for novel antimicrobial agents. The present study explores the antioxidant, antibacterial, and cytotoxic properties of extracts from submerged mycelial biomass of Fomitopsis pinicola, cultivated in synthetic and lignocellulosic media. Four extracts were obtained using hot water and 80% ethanol. The provided analysis of extracts confirmed the presence of various bioactive compounds, including flavonoids, alkaloids, and polyphenols. All extracts showed dose-dependent antioxidant activity (IC50: 1.9–6.7 mg/mL). Antibacterial tests revealed that Klebsiella pneumoniae was most sensitive, with the L2 extract producing the largest inhibition zone (15.33 ± 0.47 mm), while the strongest bactericidal effect was observed against Acinetobacter baumannii (MBC as low as 0.5 mg/mL for L1). Notably, all extracts significantly reduced the viability of stationary-phase B. burgdorferi cells, with L2 reducing viability to 42 ± 2% at 5 mg/mL, and decreased biofilm mass, especially with S2. Cytotoxicity assays showed minimal effects on NIH 3T3 cells, with slight toxicity in HEK 293 cells for S2 and L1. These results suggest that F. pinicola extracts, particularly ethanolic L2 and S2, may offer promising natural antimicrobial and antioxidant agents for managing resistant infections. Full article
(This article belongs to the Section Biology Research and Life Sciences)
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