Search Results (381)

Accurate differentiation between Coilia brachygnathus and Coilia nasus is imperative for the effective management of fisheries, the conservation of aquatic ecosystems, and the mitigation of commercial fraud. Current morphological identification remains challenging due to their high morphological similarity—particularly for processed samples—while conventional molecular methods often lack the speed or specificity required for field applications or high-throughput screening. In this study, a novel integrated approach was developed and validated, combining TaqMan quantitative real-time PCR (qPCR). for precise genotyping of C. brachygnathus and C. nasus with Recombinase Polymerase Amplification coupled with Lateral Flow Dipstick (RPA-LFD) for rapid on-site screening. First, species-specific RPA-LFD assays were designed to target the mitochondrial COI gene sequence. This enabled visual detection within 10 min at 37 °C, with a sensitivity of 10² copies/μL, and required no complex equipment. A dual TaqMan MGB qPCR assay was further developed by validating stable differentiating SNPs (chr21:3798155, C/T) between C. brachygnathus and C. nasus, using FAM/VIC dual-labeled MGB probes. Results showed that this assay could distinguish the two species in a single tube: for C. brachygnathus, Ct values in the FAM channel were significantly earlier than those in the VIC channel (ΔCt ≥ 1), with a FAM detection limit of 125 copies/reaction; for C. nasus, only VIC channel amplification was observed, with a detection limit as low as 12.5 copies/reaction. Validation with 171 known tissue samples demonstrated 100% concordance with expected species identities. This integrated approach effectively combines the high accuracy and quantitative capacity of TaqMan qPCR for confirmatory laboratory genotyping with the speed, simplicity, and portability of RPA-LFD for initial field or point-of-need screening. This reliable, efficient, and user-friendly technique provides a powerful tool for resource management, biodiversity monitoring, and ensuring the authenticity of high-quality C. brachygnathus and C. nasus. Full article

Forest ecosystems undergo seasonal shifts in bacterial and fungal communities, but little is known about the specific microbiota associated with Quercus robur–Boletus edulis systems. This study represents the first examination of seasonal changes in soil microbiota in pedunculate oak habitats in Galicia (NW Spain) and their relationship with Boletus edulis and Boletus reticulatus mycelium prevalence and concentration. Soil microbiota richness, diversity, and composition, as well as seasonal variation in Boletus mycelium, were assessed using DNA metabarcoding and qPCR, respectively. Sampling was conducted in autumn at two 30–40-year-old Q. robur stands. Bacterial communities were dominated by Acidobacteria (34%) and Proteobacteria (33%), with Acidobacterium (12%), Paludibaculum (9%), and Edaphobacter (7%) identified as most abundant. Fungal communities were primarily Basidiomycota (93%), led by Russula (46%). For both bacteria and fungi, the highest OTU richness was observed in September, followed by a significant decrease in October and a partial recovery in November. Boletus species were found to exhibit positive correlations with specific bacteria (e.g., Massilia, Rhizobium) and fungi (e.g., Amanita, Clavaria, Inocybe, Scleroderma, Suillus and Mortierella), suggesting a potential influence of these microbes on mycelium development. This study provides novel insights into the seasonal dynamics of soil microbiota and their potential role in Boletus ecology, thereby advancing understanding of host–microbe interactions in temperate forests. Full article

Background: Meniere’s disease (MD) is a rare inner ear disorder characterized by endolymphatic hydrops and symptoms such as vertigo and hearing loss, with no curative treatment currently available. XuanYunNing tablets (XYN) have been clinically used to treat MD, but their molecular mechanisms remain unclear. Objective: This study aimed to systematically evaluate the pharmacological effects of XYN in a guinea pig model of MD and to elucidate the underlying molecular mechanisms of both MD pathogenesis and XYN intervention through integrated multi-omics analyses, including transcriptomics, proteomics, and bioinformatics. Methods: A guinea pig model of endolymphatic hydrops was induced by intraperitoneal injection of desmopressin acetate (dDAVP). Pharmacodynamic efficacy was evaluated via behavioral scoring and histopathological analysis. The differentially expressed genes (DEGs) and differentially expressed proteins (DEPs) modulated by XYN treatment were identified using high-throughput transcriptomic and proteomic sequencing. These data were integrated through multi-omics bioinformatic analysis. Key molecular targets and signaling pathways were further validated using RT-qPCR and Western blotting. Results: Pharmacological evaluations showed that guinea pigs in the model group exhibited a 26% increase in endolymphatic hydrops area, while high-dose XYN treatment reduced this area by 19% and significantly improved functional parameters, including overall physiological condition (e.g., weight and general appearance), auricular reflexes to low-, medium-, and high-frequency sound stimuli, nystagmus, and the righting reflex. High-throughput sequencing combined with integrative omics analysis identified 513 potential molecular targets of XYN. Subsequent network and module analyses pinpointed the JAK-STAT signaling pathway as the central axis. Mendelian randomization (MR) analysis further supported a causal relationship between MD and metabolic, immune, and inflammatory traits, reinforcing the central role of JAK-STAT signaling in both MD progression and XYN-mediated intervention. Mechanistic studies confirmed that XYN downregulated IFNG, IFNGR1, JAK1, p-STAT3/STAT3, and AOX at both mRNA and protein levels, thereby inhibiting aberrant JAK-STAT pathway activation in MD model animals. In addition, a total of 125 chemical constituents were identified in XYN by UHPLC-MS analysis. ZBTB20 and other molecules were identified as potential blood-based biomarkers for MD. Conclusions: This study reveals that XYN alleviates MD symptoms by disrupting a pathological cycle driven by JAK-STAT signaling, inflammation, and metabolic dysfunction. These findings support the clinical potential of XYN in the treatment of Meniere’s disease and may inform the development of novel therapeutic strategies. Full article

Citrinin (CTN), a mycotoxin commonly found in contaminated food and animal feed, impairs intestinal barrier integrity through oxidative stress and cytotoxicity. However, its link to ferroptosis, an iron-dependent form of regulated cell death, remains unclear. This study investigated whether CTN induces ferroptosis in intestinal epithelial cells and evaluated the protective role of Euphorbia hypericifolia (EH) against CTN-induced oxidative damage and tight junction (TJ) disruption. Using IPEC-J2 cells exposed to CTN, intracellular ferrous ion (Fe²⁺) levels, reactive oxygen species (ROS) accumulation, and TJ integrity were assessed using FerroOrange and DCFH-DA staining, RT-qPCR, immunofluorescence, and WST-1 assays. Additionally, a high-throughput screen of 459 natural products identified EH extract as a top candidate in mitigating CTN toxicity. The CTN treatment significantly elevated intracellular Fe²⁺ and ROS levels, downregulated antioxidant genes (notably CAT), and disrupted ZO-1 expression and TJ morphology in IPEC-J2 cells, all hallmarks of ferroptosis-like cell death. Co-treatment with EH extract effectively reversed these effects, restoring antioxidant gene expression, reducing Fe²⁺ and ROS accumulation, and preserving TJ structure. Phytochemical profiling of EH extract revealed several bioactive compounds potentially responsible for its protective effects. These findings suggest that CTN induces ferroptosis-related cytotoxicity in IPEC-J2 cells, but EH alleviates this toxicity by modulating oxidative stress and iron homeostasis, supporting its potential use as a natural feed additive for intestinal protection Full article

Freshwater salinization, an escalating global environmental stressor, poses a significant threat to freshwater biodiversity, including fish communities. This study investigates the grass carp (Ctenopharyngodon idellus), a species with the highest aquaculture output in China, to elucidate the molecular underpinnings of its physiological adaptations to fluctuating salinity gradients. We used high-throughput mRNA sequencing and differential gene expression profiling to analyze transcriptional dynamics in intestinal and kidney tissues of grass carp exposed to heterogeneous salinity stressors. Concurrent serum biochemical analyses showed salinity stress significantly increased Na⁺, Cl⁻, and osmolarity, while decreasing lactate and glucose. Salinity stress exerted a profound impact on the global transcriptomic landscape of grass carp. A substantial number of co-regulated differentially expressed genes (DEGs) in kidney and intestinal tissues were enriched in immune and metabolic pathways. Specifically, genes associated with antigen processing and presentation (e.g., cd4-1, calr3b) and apoptosis (e.g., caspase17, pik3ca) exhibited upregulated expression, whereas genes involved in gluconeogenesis/glycolysis (e.g., hk2, pck2) were downregulated. KEGG pathway enrichment analyses revealed that metabolic and cellular structural pathways were predominantly enriched in intestinal tissues, while kidney tissues showed preferential enrichment of immune and apoptotic pathways. Rigorous validation of RNA-seq data via qPCR confirmed the robustness and cross-platform consistency of the findings. This study investigated the core transcriptional and physiological mechanisms regulating grass carp’s response to salinity stress, providing a theoretical foundation for research into grass carp’s resistance to salinity stress and the development of salt-tolerant varieties. Full article

Browning significantly impacts the commercial value of luffa (luffa cylindrica) and is primarily driven by the metabolic processes of phenolic acids. Investigating changes in phenolic acids during browning aids in understanding the physiological mechanisms underlying this process and provides a basis for improving storage, processing, variety breeding, and utilization of germplasm resources. This study compared browning-resistant (‘30’) and browning-sensitive (‘256’) luffa varieties using high-throughput sequencing and metabolomics techniques. The results revealed 55 genes involved in the phenylpropanoid biosynthesis pathway, including 8 phenylalanine ammonia-lyase (PAL) genes, 20 peroxidase (POD) genes, 2 polyphenol oxidase (PPO) genes associated with tyrosine metabolism, and 37 peroxisome-related genes. Real-time quantitative (qPCR) was employed to validate 15 browning-related genes, revealing that the expression levels of LcPOD21 and LcPOD6 were 12.5-fold and 25-fold higher in ‘30’ compared to ‘256’, while LcPAL5 and LcPAL4 were upregulated in ‘30’. Enzyme analysis showed that catalase (CAT) and phenylalanine ammonia-lyase (PAL) activities were higher in ‘30’ than in ‘256’. Conversely, superoxide dismutase (SOD) and polyphenol oxidase (PPO) activities were reduced in ‘30’, whereas CAT activity was upregulated. The concentrations of cinnamic acid, p-coumaric acid, trans-5-O-(4-coumaroyl)mangiferic acid, and caffealdehyde were lower in browning-resistant luffa ‘30’ than in browning-sensitive luffa ‘256’, suggesting that their levels influence browning in luffa. These findings elucidate the mechanisms underlying browning and inform strategies for the storage, processing, and genetic improvement of luffa. Full article

L-carnitine and Mildronate are substances that can significantly rearrange the energy metabolism of cells. This can potentially cause changes in the bacterial composition of the gut microbiome and affect testis functionality and male sexual health. Mice of the C57Bl/6 line were used. Sexual behavior was assessed using physiological tests, and gene expression patterns were assessed by qPCR. High-throughput sequencing of mouse fecal microbiota was performed. We showed that long-term administration of Mildronate has no significant effect on the intestinal microbiome, and there was a compensatory increase in the expression of genes involved in fatty acid and leptin metabolism. No impairment of sexual motivation in male mice was observed. Prolonged L-carnitine supplementation caused a decrease in alpha diversity of bacteria and a decrease in some groups of microorganisms that are components of a healthy gut microflora. A correlation was observed between the level of bacteria from Firmicutes phylum, indicators of sexual motivation of mice, and the dynamics of body weight gain. Our results may indicate that metabolic modulators can have a significant impact on the structure of the bacterial community of the gut microbiome, which may influence male sexual health through the gut–semen axis. Full article

The pupal stage in necrophagous flies represents the longest and least morphologically distinct phase of development, posing a persistent challenge for accurately estimating postmortem intervals (PMI) in forensic investigations. Here, we present a novel molecular approach to pupal age estimation in Sarcophaga peregrina, a forensically important species, by profiling microRNA (miRNA) expression dynamics. High-throughput sequencing across early, mid, and late pupal stages identified 191 known miRNAs, of which nine exhibited distinct monotonic temporal trends. Six miRNAs (miR-210-3p, miR-285, miR-927-5p, miR-956-3p, miR-92b, and miR-275-5p) were validated by qRT-PCR and demonstrated consistent time-dependent expression patterns. Polynomial regression models revealed a strong correlation between miRNA abundance and developmental age (R² = 0.88–0.99). Functional enrichment analyses of predicted miRNA targets highlighted their roles in key regulatory pathways, including ecdysteroid signaling, hypoxia response, autophagy, and energy metabolism. This study establishes, for the first time, a robust miRNA-based framework for estimating pupal age in forensic entomology, underscoring the potential of miRNAs as temporally precise biomarkers for PMI estimation. Full article

Hippeastrum, a perennial herbaceous plant belonging to the Amaryllidaceae family, is widely cultivated for its large, vibrant flowers with diverse petal colors, which have significant ornamental and economic value. However, the mechanisms underlying anthocyanin accumulation in Hippeastrum petals remain poorly understood. To fully explore the involved regulation mechanism was significant for the breeding of Hippeastrum and other Amaryllidaceae family plants. In this study, we selected six Hippeastrum cultivars with distinctly different petal colors. We used metabolomic profiling and high-throughput transcriptomic sequencing to assess varied anthocyanin profiles and associated expression of genes in their biosynthetic pathways. Four key anthocyanins were identified: cyanidin, cyanidin-3-O-rutinoside, delphinidin-3-glucoside, and delphinidin-3-rutinoside. Weighted gene co-expression network analysis (WGCNA) correlated the abundance of these four anthocyanins with transcriptomic data, to suggest three regulatory modules. Nine transcription factors families in these modules were identified and some of them were validated using qRT-PCR. Y2H assay isolated some transcription factors interacted with TTG1 (WD40 protein), including MYB3/39/44/306 and bHLH13/34/110, illustrating the possibility of forming MBW complexes. Our study provides a comprehensive characterization of anthocyanin composition. These findings laid a theoretical foundation for future research on the regulatory mechanisms of pigment accumulation and the breeding of Hippeastrum cultivars with novel petal colors. Full article

The land-use conversion of Masson pine forests to tea fields is extensively practiced across subtropical China, primarily driven by its economic benefit. However, the effects of this conversion on soil fungal communities and functional guilds are poorly understood. Herein, a field experiment was conducted in a Masson pine forest (F), a 5-year-old tea plantation without (FT-CK) fertilization or with (FT-N), and a 30-year-old tea plantation (FT-O) to assess the impact of Masson pine forest-to-tea conversion on soil fungal abundance, community structure, and functional guilds by using qPCR and high-throughput sequencing. Compared to F, fungal abundance significantly decreased by 95%, 68%, and 79% in FT-CK, FT-N, and FT-O, respectively, probably caused by the decreased total nitrogen content and habitat disruption. Fungal alpha diversity significantly increased in FT-N and FT-O compared to FT-CK. FT-O presented the highest percentages of Mortierella among treatments, which favours soil organic carbon accumulation. FUNGuild-based predictions showed that FT-CK and FT-N had higher relative abundances of plant pathogens than F and FT-O. FT-O presented the highest percentages of litter and soil saprotrophs but exhibited the lowest percentages of ectomycorrhizal fungi among treatments, likely driven by increased soil organic carbon, total nitrogen, and total phosphorus content. Our findings demonstrate that Masson pine forest-to-tea conversion significantly degrades soil fungal community and function, highlighting the urgent need for soil management strategies (e.g., organic amendments) to enhance soil health in tea agroecosystems. Full article

The novel goose parvovirus (NGPV) and the novel duck reovirus (NDRV) are pathogens that can substantially affect the growth and development of ducklings, causing considerable economic losses to duck farms. Therefore, a timely, rapid, accurate, and high-throughput diagnosis and identification of viral infections are critical for preventing the spread of epidemics. In this study, a TaqMan probe-based duplex one-step RT-qPCR was established for the simultaneous detection and qualitative and quantitative identification of the two viruses. It demonstrated greater sensitivity than conventional PCR, detecting as low as 2.42 copies/μL of NGPV genome and 70.1 copies/μL of NDRV genome. Additionally, it exhibited remarkable specificity, responding exclusively to the nucleic acids of target pathogens. It also demonstrated excellent reproducibility and availability, particularly in clinical settings, with a coinfection detection rate of 13.3%, contributing to the development of NGPV- and NDRV-testing technologies. Full article

Background: Oral squamous cell carcinoma (OSCC), which accounts for over 90% of all oral malignancies, remains a major global health challenge due to its aggressive clinical course and poor prognosis. Periodontitis, a widespread chronic inflammatory condition affecting the supporting structures of the teeth, has increasingly been implicated as a potential risk factor for the development of various cancers. Emerging evidence suggests that microbial dysbiosis within the oral cavity may contribute to the creation of a pro-tumorigenic microenvironment, thereby promoting tumor initiation and progression. Nevertheless, the precise mechanisms linking periodontitis to OSCC, particularly through alterations in the oral microbiota, remain insufficiently understood. This article seeks to comprehensively analyze the association between periodontitis and OSCC and to elucidate the potential role of oral microbiota dysbiosis in mediating this relationship. Methods: In this study, a ligature-induced periodontitis model was established in C57BL/6J mice, and after two weeks, an OSCC model was introduced by the subcutaneous injection of SCC-7 cells to investigate the impact of periodontitis on OSCC progression. The effects of periodontitis on OSCC cell proliferation and invasion were assessed using scratch wound healing assays and CCK-8 proliferation assays. 16S rDNA high-throughput sequencing was conducted to profile the microbial communities present in the oral cavity and OSCC tissues, with particular emphasis on α-diversity indices (including Pielou’s evenness and Chao1 richness) and taxonomic composition at both the phylum and class levels. Furthermore, qPCR was utilized to assess the expression levels of cytokines in both periodontal and OSCC tissues, thereby elucidating the inflammatory milieu, potentially linking periodontitis to OSCC progression. Results: Our findings demonstrated that periodontitis significantly promoted OSCC growth and enhanced the invasive potential of OSCC cells. Microbial profiling revealed marked alterations in both the oral and OSCC microbiota, characterized by significant shifts in community composition and increased microbial diversity. Notably, these microbial changes exhibited consistent patterns between the oral cavity and the OSCC microenvironment, suggesting a potential mechanistic link between periodontitis-associated dysbiosis and OSCC progression. Consistently, qPCR analysis revealed elevated expression levels of IL-1β, IL-10, and IL-18 in both periodontal and OSCC tissues, providing evidence that the microbial alterations were accompanied by intensified inflammatory responses, which may contribute to OSCC progression. Conclusions: This study underscores the intricate interplay between periodontitis-induced microbial dysbiosis and the development of oral squamous cell carcinoma (OSCC). The findings suggest that periodontal inflammation, together with associated shifts in the oral microbiota, acts synergistically to drive OSCC progression. The elevated expression of cytokines further supports the role of a pro-inflammatory tumor microenvironment in mediating this interaction. These results offer important insights into the microbial and inflammatory mechanisms underlying the connection between periodontitis and OSCC, highlighting the critical role of maintaining periodontal health in the prevention and management of OSCC. Full article

Macrophages participate in cutaneous wound healing by adopting M1 pro-inflammatory and M2 immunoregulatory/pro-repair phenotypes. Chronic wounds associated with a deficient macrophage response could benefit from treatments that restore an acute inflammatory response and promote healing. Calcium-alginate dressings release calcium ions, which are potent bioactivators of macrophage function in wounds. Here, the effects of two calcium-alginate dressings, Algosteril^® (ALG, pure Ca²⁺ alginate) and Biatain^® Alginate (BIA, Ca²⁺ alginate with carboxymethyl cellulose), and a 3 mM CaCl₂ solution were compared in human macrophages polarized to M1 or M2. ALG and CaCl₂ preserved monocyte viability, and BIA reduced it. Both alginates and CaCl₂ reinforced the M1 pro-inflammatory transcriptional profile and phenotype, with significant increases in IL-6 and TNF-α secretion by ALG only. In M2 macrophages, all conditions increased the M1-specific gene expression and reduced M2 markers, suggesting an orientation toward an inflammatory profile. Only ALG significantly increased the secretion of CCL18 and VEGF, suggesting pro-repair activity. All conditions increased M2 phagocytic activity. This work demonstrates the interest in calcium alginates for stimulating macrophage subtypes, which could help restore wound healing, especially in patients with compromised innate immunity. It highlights the differences among the calcium-alginate dressings. The pure alginate shows higher stimulation of macrophage pro-inflammatory and pro-repair functions. Full article

Background: The 2C protein of foot-and-mouth disease virus (FMDV), a member of helicase superfamily 3 (SF3), drives viral genome replication and serves as a critical target for antiviral drug development. Methods: A fluorescence resonance energy transfer (FRET)-based high-throughput screening (HTS) platform was developed to identify 2C helicase inhibitors. Primary screening evaluated 4424 compounds for helicase inhibition. Molecular docking analyzed inhibitor interactions with the N207 residue within the catalytic core and helicase inhibition assays classified the inhibitor type (mixed, competitive, noncompetitive). Differential scanning fluorimetry (nanoDSF) quantified 2C thermal destabilization. Antiviral activity was assessed via indirect immunofluorescence, RT-qPCR, and plaque reduction assays. Results: Six compounds inhibited 2C helicase activity at >620 μM. Molecular docking revealed hydrogen bonding, hydrophobic interactions, and π-cation stabilization at the catalytic core. 2-MPO and MPPI were classified as mixed-type inhibitors, 5-TzS⁻ and 2-PyOH as competitive, and DCMQ/Spiro-BD-CHD-dione as noncompetitive. NanoDSF showed a ΔT_m ≥ 1.5 °C (2.5 mM compounds), with reduced destabilization in N207A mutants. Antiviral assays identified 2-MPO and MPPI as optimal inhibitors. MPPI achieved effective FMDV suppression at 160 μM, exhibiting two orders of magnitude higher potency than 2-MPO (400 μM). Conclusions: The established FRET-based HTS platform targeting 2C helicase facilitates anti-FMDV lead discovery, while 2C inhibitors may serve as an effective therapeutic strategy against other picornaviruses. Full article

Background: Birnaviruses infect a wide range of aquatic and terrestrial hosts, including several economically important fish species. This study aimed to isolate and characterize a novel birnavirus strain from mandarin fish (Siniperca chuatsi), a high-value freshwater species in Chinese aquaculture. Methods: A novel strain, designated mandarin fish birnavirus (MFBV), was isolated from diseased fish and propagated in SCK cells. The complete genome was determined using high-throughput sequencing and RACE. Viral replication kinetics, tissue distribution, and pathogenicity were assessed through in vitro infection, RT-qPCR, histopathology, and experimental challenges. In addition, disinfectant sensitivity and environmental stability were evaluated. Results: The MFBV genome comprises two segments (A: 3539 bp; B: 2719 bp), and phylogenetic analysis revealed close relatedness to largemouth bass birnavirus (LBBV) and Lates calcarifer birnavirus (LCBV). MFBV displayed rapid replication in SCK cells, completing a replication cycle in 8–10 h. In juvenile and fry fish, an experimental infection caused acute disease with cumulative mortality ranging from 41.8% to 83.6%, with fry showing higher susceptibility. Viral RNA was detected in multiple tissues (7.9 × 10⁶–7.9 × 10⁷ copies/μg RNA), and histopathological lesions were observed in the intestine, spleen, and kidney. MFBV was highly sensitive to glutaraldehyde (20 ppm), while other disinfectants showed reduced efficacy. Viral half-life ranged from 36.5 to 144.5 h at room temperature. Conclusions: These findings demonstrate that MFBV can induce acute systemic infection in mandarin fish. The results offer new insights into the genomic and biological features of birnaviruses, contributing to improved disease management and viral taxonomy. Full article