Search Results (2,059)

In Lolium perenne, a novel growth habit mutant, named VIROIZ, was recovered following colchicine treatment, and it was confirmed to maintain the diploid chromosome number (2n = 2x = 14). The mutation affected the stem morphology by inducing prolific axillary shoot formation at nodal zones, resulting in a spreading growth habit that can extend to ~70 cm in width. Inheritance analysis based on single-plant evaluations in crosses with wild-type plants (F₁, n = 285; F₂, n = 380) and in selfed progeny (S₁, n = 255) consistently showed ~40% expression of the spreading phenotype, deviating from classical Mendelian ratios and indicating complex genetic control. Phenotypic selection further distinguished divergent classes: positively selected lines (C1⁺) averaged 3.90 axillary tillers per stem, whereas negatively selected lines (C1⁻) averaged only 0.22. Partial sequencing of 11 candidate genes implicated in shoot architecture, covering 40–90% of full-length DNA, did not provide a conclusive explanation for the altered stem growth. Notably, a single point mutation was observed in CRT3 (an endoplasmic reticulum chaperone that interacts with brassinosteroid signaling) highlighting it as a primary target for future studies. Cytological analysis of meiosis in F₁ hybrids between VIROIZ and wild-type plants revealed irregular chromosome pairing with persistent univalents (2–4 per cell), supporting the presence of structural chromosomal rearrangements that may disrupt gene organization and function in VIROIZ. The non-Mendelian segregation of the spreading phenotype, together with the observed meiotic irregularities, suggests that the mutation affects regulatory genes responsive to hormonal signals controlling axillary meristem initiation. The mutant represents a valuable resource for turf-type L. perenne breeding and for studying hormonal regulation of shoot morphogenesis in Poaceae. Full article

Environmental and ancient DNA are mostly present in degraded forms in nature. Plant forensics is necessary for plants like Taxus (Taxaceae), which is a medicinal, as well as poisonous, endangered plant. We designed a study to develop high-efficiency PCR mini-barcoding primers for the identification of Taxus. We collected environmental materials, fresh and old Taxus specimens from natural habitats, herbaria, and ex situ propagation sites. Taxon-specific mini-barcoding primers were prepared through primer3. All the primers were amplified onto Taxus specimens and environmental samples having Taxus DNA, while no amplification on fresh and herbarium specimens other than Taxus was noted. DNA sequencing of amplified regions of matK, ITS, and rbcL yielded lengths of 117, 175, and 200 bp. Blast taxonomy showed 100% identification power at the genus level, while 75–93% at the species level, and identified a total of 30 taxa within the genus Taxus, comprising 16 species, 5 varieties, 2 hybrids, and 7 variants. ITS was the most specific for genus identification, followed by matK and rbcL. Environmental, trade, socio-economic, and toxicological crimes were also identified. Our high-efficiency PCR mini-barcoding method can be useful in the prevention of Taxus illegal trade and habitat degradation to mitigate climate change in the Himalayan region of Pakistan. Full article

Background and Objectives: Thrombophilia is a prothrombotic disorder that increases the risk of blood clotting and can pose serious health problems. It is considered a condition of gene–gene or gene–environment interactions. Variation in the prevalence of thrombophilia mutations and their interaction among populations necessitates localized genetic assessments. However, population-based genetic data remains limited for developing effective preventive strategies. Materials and Methods: This cross-sectional observational study was conducted over five years (2020–2024) at a tertiary university hospital in Northern Greece. A total of 2961 individuals aged 18–85 years (mean: 50.5) were registered based on family or medical history of venous thromboembolism (VTE) or clinical symptoms of VTE. The final analysis included 2078 participants comprising 1143 males (55%) and 935 females (45%), who met all the inclusion criteria. Inclusion criteria were absence of acute illness or malignancy, informed consent, and an adequate DNA quantity for genotyping, whereas excluded criteria included incomplete laboratory data, active inflammatory or malignant disease, and cognitive or psychiatric conditions. Peripheral blood samples were collected in 2 mL K₃-EDTA tubes, and genomic DNA was analyzed using real-time polymerase chain reaction (PCR) with melting curve analysis and hybridization probes (LightMix^® in vitro diagnostics, TIB MolBiol, Berlin, Germany). Five thrombophilia-related polymorphisms, Factor V Leiden (F5 G1691A), prothrombin (F2 G20210A), methylenetetrahydrofolate reductase (MTHFR C677T and MTHFR A1298C), and Plasminogen Activator Inhibitor-1 (PAI-1) 4G/5G, were examined for allele and genotype frequencies, Hardy–Weinberg equilibrium testing, pairwise linkage disequilibrium (D′ and r²), and power analysis. For subjects tested for Factor V Leiden (n = 1476), the activated protein C resistance (APC) ratio was additionally evaluated using the ACL TOP 750 analyzer. Results: Allele frequencies were 7.3% for FV Leiden and 3.7% for FII. The PAI-1 allele was distributed at 44%, while the MTHFR (C677T and A1298C) alleles were each present at 33%. Significant linkage disequilibrium was identified between MTHFR (C677T and A1298C) and between MTHFR A1298C and PAI-1. No evolutionary pressure or demographic bias was found in the Hardy–Weinberg equilibrium. The APC ratio demonstrated a high sensitivity (99.2%) and specificity (96.6%), indicating that it may serve as a reliable screening method. Conclusions: Our findings highlight informative patterns in the genetic predisposition to thrombophilia, which may help develop rule-based strategies for implementing thromboprophylaxis guidelines and personalized medical interventions. Full article

Hepatitis C represents a critical global health crisis, causing approximately 1.4 million deaths annually. Although 98% of cases are treatable, only about 20% of infected individuals know their hepatitis C virus (HCV) status, highlighting the urgent need for rapid and more efficient diagnostic management. Viral genetic material can be detected in serum or plasma within just one week of exposure, making it the most reliable marker and the gold standard for active HCV infection diagnosis. In this study, a biosensor was developed to detect conserved nucleotide sequences of HCV using a 3D surface electrode composed of poly-L-lysine (PLL) and carbon nanotubes (CNTs). PLL is a positively charged biocompatible polymer rich in amine groups, attractive for the immobilization of proteins, DNA, and other biomolecules. PLL was employed to construct a 3D surface with vertically aligned CNTs, achieving a high electron transfer rate. Cyclic voltammetry technique and scanning electron microscopy (SEM) were used to characterize the sensor platform, and analytical responses were measured by differential pulse voltammetry. This HCV biosensor detected the hybridization event by a significant reduction in DPV peaks in the presence of the ferri/ferrocyanide redox probe, without any intercalator agents. DNA responses were observed in phosphate-buffered saline (PBS) and cDNA-spiked serum samples, demonstrating its analytical specificity. These findings represent advances in analytical tools that can effectively address the challenges of timely diagnosis for asymptomatic HCV carriers. Full article

Deoxyribonucleic acid (DNA)-based biosensors are rapid, cost-effective, and portable devices for monitoring crop pathogens. However, their on-field operations rely on a laboratory-bound heating block, which controls temperature during sample preparation. This study aimed to develop a field-deployable heating block to assist in the DNA hybridization protocol of DNA-based biosensors. It should maintain $95 °\mathrm{C}$, $55 °\mathrm{C}$, and $20 °\mathrm{C}$ for 5, 10, and 5 min, respectively. It had aluminum bars, positive thermal coefficient ceramic heaters, a Peltier thermoelectric module, and DS18B20 thermistors, serving twelve 0.2 mL polymerase chain reaction (PCR) tubes. An Arduino microcontroller employing a proportional–integral–derivative (PID) algorithm with a solid-state relay was utilized. Machine performance for distilled water-filled PCR tubes showed a maximum $10 °\mathrm{C}$ thermal variation. The machine maintained $(96.00\pm 0.97) °\mathrm{C}$, $(55.15\pm 2.17) °\mathrm{C}$, and $(17.75\pm 0.71) °\mathrm{C}$ with root mean square errors (RMSEs) of $1.40 °\mathrm{C}$, $2.18 °\mathrm{C}$, and $2.36 °\mathrm{C}$, respectively. The average thermal rates were $(0.16\pm 0.11) °\mathrm{C}/\mathrm{s}$, $(0.29\pm 0.11) °\mathrm{C}/\mathrm{s}$, and $(0.14\pm 0.07) °\mathrm{C}/\mathrm{s}$ from ambient to $95 °\mathrm{C}$, $95 °\mathrm{C}$ to $55 °\mathrm{C}$, and $55 °\mathrm{C}$ to $20 °\mathrm{C}$, respectively. Overall, the low standard deviations and RMSEs demonstrate thermostable results and robust temperature control. Full article

Two bacterial strains, designated LXY-1^T and LXY-4^T, were isolated from soil samples collected at a heavy metal smelting plant located in Guangxi, China. Phylogenetic analysis indicated that these strains formed two distinct lineages within the genus Neobacillus. Both strains were characterized as facultative anaerobic, Gram-positive staining, endospore-forming, non-motile, short-rod bacteria. The major cellular fatty acids identified in these strains included C_16:0, iso-C_15:0, antéiso-C_15:0, and antéiso-C_17:0. The predominant polar lipids comprised diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), and phosphatidylglycerol (PG). The average nucleotide identity (ANI) values between the newly isolated strains and their closest phylogenetic relatives, the type strains of the genus Neobacillus, were found to be below 95%, with corresponding digital DNA–DNA hybridization (dDDH) values remaining below 70%. Based on a comprehensive polyphasic taxonomic analysis incorporating chemotaxonomic, phenotypic, phylogenetic, and genomic data, we proposed that strains LXY-1^T and LXY-4^T represent two novel species of the genus Neobacillus, for which the names Neobacillus terrisolis sp. nov. and Neobacillus solisequens sp. nov. are designated. The type strains are LXY-1^T (= CGMCC 30313^T = JCM 37671^T) and LXY-4^T (= CGMCC 1.62901^T = JCM 37672^T), respectively. Full article

Periodontitis is a biofilm-driven inflammatory disease in which conventional indices (probing depth, clinical attachment level, and radiographs) quantify tissue destruction without capturing the biology of infection. In this review, we synthesized microbiological diagnostics, from chairside tools to omics. We outline sampling strategies and emphasize the quantitative monitoring of bacterial load. Enzymatic assays (e.g., N-benzoyl-DL-arginine-2-naphthylamide hydrolysis assay test) measure functional activity at the point of care. Immunological methods include rapid immunochromatography for Porphyromonas gingivalis and enzyme-linked immunosorbent assay for the high-throughput measurement of bacterial antigens. Molecular platforms encompass quantitative polymerase chain reaction (qPCR) (TaqMan, SYBR, multiplex panels; propidium monoazide quantitative-qPCR for viable cells), checkerboard DNA–DNA hybridization for semi-quantitative community profiling, loop-mediated isothermal amplification (LAMP)/molecular beacon-LAMP for portable isothermal detection, and microarrays. Complementary modalities such as fluorescent in situ hybridization, next-generation sequencing, and Fourier transform infrared spectroscopy provide spatial, ecological, and biochemical resolutions. We discuss the limitations of current approaches, including sampling bias, presence–activity discordance, semi-quantitation, method biases, limited strain/function resolution, low-biomass artifacts, and lack of validated cutoffs. To address these challenges, we propose a pragmatic hybrid strategy: site-specific quantitative panels combined with activity and host-response markers interpreted alongside clinical metrics under standardized quality assurance/quality control. Priorities include outcome-linked thresholds, strain-aware/functional panels, robust point-of-care chemistry, and harmonized protocols to enable personalized periodontal care. Full article

Background/Objectives: Mutations in NACHT, LRR and PYD domain-containing protein 2 (NLRP2) and NLRP7 genes, members of the NOD-like receptor (NLR) family of innate immune sensors, result in recurrent miscarriages and reproductive wastage in women. These genes have been identified to be maternal effect genes in humans and mice regulating early embryo development. Previous research in vitro suggests that NLRP2 and NLRP7 regulate DNA methylation and/or immune signaling through inflammasome formation. However, the exact mechanisms underlying NLRP2 and NLRP7 function are not well defined. Methods: To determine the interacting proteins required for NLRP2/NLRP7-mediated regulation of DNA methylation, yeast 2-hybrid screens, coimmunoprecipitation, and FRET studies were performed and verified the ability of novel protein interactions to affect global DNA methylation by 5-methylcytosine-specific ELISA. Results: Various methodologies employed in this research demonstrate a novel protein interaction between human ErbB3-binding protein 1 (EBP1, also known as proliferation-associated protein 2G4 (PA2G4) and NLRP2 or NLRP7. In addition, NLRP2 and NLRP7 regulate EBP1 gene expression. Functionally, global DNA methylation levels appeared to decrease further when NLRP2 and NLRP7 were co-expressed with EBP1, although additional studies may need to confirm the significance of this effect. Conclusions: Since EBP1 is implicated in apoptosis, cell proliferation, DNA methylation, and differentiation, our discovery significantly advances our understanding of how mutations in NLRP2 or NLRP7 may contribute to reproductive wastage in women through EBP1. Full article

Background: Currently, there are limited therapeutic or preventative strategies for neurodegenerative disorders due to the challenges in alleviating the progressive neuronal loss and neuroinflammation which are the primary characteristics of these diseases, ultimately leading to cell death and functional impairment. Cocoa-derived flavanols (Theobroma cacao) have been studied as potential bioactive compounds to modify and reverse various inflammation-associated diseases because of their remarkable antioxidant properties and capacity to modulate metabolic imbalance and reactive inflammatory responses. The faba bean (Vicia faba) extract obtained through nondenaturing biotechnological processes is a potent dopamine (DA) enhancer that has shown promising results as a neuroprotective agent against degeneration. Objective: This study will examine the synergistic effects of Neurofabine-C, a hybrid compound derived from cocoa and faba bean extracts, on various brain biomarkers in mice related to inflammatory, metabolic, and neurodegenerative processes. Methods: A triple-transgenic mouse model of neurodegeneration was treated with Neurofabine-C, and biomolecular data were obtained by performing biochemical and immunohistochemical analysis. Results: Neurofabine-C prevented neuronal degeneration (NeuN), mitigated the neuro-inflammatory processes triggered (decreased expression of reactive astrocytes (GFAP)), and induced an increase in neurogenesis in the treated cortical mice brain (PAX6). Epigenetic analysis revealed significant chromatin remodeling in the hippocampus. Neuroprotective genes, including FOXO3, ATM, and TRP73, were upregulated, whereas the expression of HIF1α and APOE decreased. In parallel, DNMT3A expression increased 20-fold, HDAC3 decreased by 60%, and global 5-methylcytosine levels increased four-fold. These coordinated changes suggest that Neurofabine-C promotes neuroprotective programs through enhanced DNA methylation and reduced histone deacetylation. Conclusions: The findings indicate that Neurofabine-C exhibits multiple neuroprotective mechanisms, making it a potent bioproduct for mitigating neuroinflammatory processes associated with neurodegenerative disorders. Full article

A reassessment of the risk-benefit balance of the two lipid nanoparticle (LNP)-based vaccines, Pfizer’s Comirnaty and Moderna’s Spikevax, is currently underway. While the FDA has approved updated products, their administration is recommended only for individuals aged 65 years or older and for those aged 6 months or older who have at least one underlying medical condition associated with an increased risk of severe COVID-19. Among other factors, this change in guidelines reflect an expanded spectrum and increased incidence of adverse events (AEs) and complications relative to other vaccines. Although severe AEs are relatively rare (occurring in <0.5%) in vaccinated individuals, the sheer scale of global vaccination has resulted in millions of vaccine injuries, rendering post-vaccination syndrome (PVS) both clinically significant and scientifically intriguing. Nevertheless, the cellular and molecular mechanisms of these AEs are poorly understood. To better understand the phenomenon and to identify research needs, this review aims to highlight some theoretically plausible connections between the manifestations of PVS and some unique structural properties of mRNA-LNPs. The latter include (i) ribosomal synthesis of the antigenic spike protein (SP) without natural control over mRNA translation, diversifying antigen processing and presentation; (ii) stabilization of the mRNA by multiple chemical modification, abnormally increasing translation efficiency and frameshift mutation risk; (iii) encoding for SP, a protein with multiple toxic effects; (iv) promotion of innate immune activation and mRNA transfection in off-target tissues by the LNP, leading to systemic inflammation with autoimmune phenomena; (v) short post-reconstitution stability of vaccine nanoparticles contributing to whole-body distribution and mRNA transfection; (vi) immune reactivity and immunogenicity of PEG on the LNP surface increasing the risk of complement activation with LNP disintegration and anaphylaxis; (vii) GC enrichment and double proline modifications stabilize SP mRNA and prefusion SP, respectively; and (viii) contaminations with plasmid DNA and other organic and inorganic elements entailing toxicity with cancer risk. The collateral immune anomalies considered are innate immune activation, T-cell- and antibody-mediated cytotoxicities, dissemination of pseudo virus-like hybrid exosomes, somatic hypermutation, insertion mutagenesis, frameshift mutation, and reverse transcription. Lessons from mRNA-LNP vaccine-associated AEs may guide strategies for the prediction, prevention, and treatment of AEs, while informing the design of safer next-generation mRNA vaccines and therapeutics. Full article

Background: Food allergies represent a growing global health concern, yet the current diagnostic methods often fail to distinguish between true allergies and food sensitivities, leading to misdiagnoses and inadequate treatment. Epigenetic alterations, such as DNA methylation (DNAm), may offer novel biomarkers for precise diagnosis. Methods: This study employed a computational machine learning framework integrated with DNAm data to identify potential biomarkers and enhance diagnostic accuracy. Differential methylation analysis was performed using the limma package to identify informative CpG features, which were then analyzed with advanced algorithms, including SVM (polynomial and RBF kernels), k-NN, Random Forest, and artificial neural networks (ANN). Deep learning via a stacked autoencoder (SAE) further enriched the analysis by uncovering epigenetic patterns and reducing feature dimensionality. To ensure robustness, the identified biomarkers were independently validated using the external dataset GSE114135. Results: The hybrid machine learning models revealed LDHC and SLC35G2 methylation as promising biomarkers for food allergy prediction. Notably, the methylation pattern of the LDHC gene showed significant potential in distinguishing individuals with food allergies from those with food sensitivity. Additionally, the integration of machine learning and deep learning provided a robust platform for analyzing complex epigenetic data. Importantly, validation on GSE114135 confirmed the reproducibility and reliability of these findings across independent cohorts. Conclusions: This study demonstrates the potential of combining machine learning with DNAm data to advance precision medicine in food allergy diagnosis. The results highlight LDHC and SLC35G2 as robust epigenetic biomarkers, validated across two independent datasets (GSE114134 and GSE114135). These findings underscore the importance of developing clinical tests that incorporate these biomarkers to reduce misdiagnosis and lay the groundwork for exploring epigenetic regulation in allergic diseases. Full article

The mitochondrial DNA of trypanosomatid parasites consists of thousands of catenated minicircles and dozens of maxicircles that form a complex network structure, the kinetoplast (kDNA). Although kDNA replication and segregation during mitotic division are well studied, its inheritance during genetic exchange events remains unclear. In Trypanosoma brucei, hybrids inherit minicircles biparentally but retain maxicircles from a single parent. Although biparental inheritance of minicircles has been described in natural Trypanosoma cruzi hybrids, this process has not been explored in laboratory-generated hybrids of this parasite. In the present study, we analyzed kDNA inheritance in T. cruzi experimental hybrids using a comprehensive minicircle hypervariable region (mHVR) database and genome sequencing data. Our findings revealed biparental inheritance of minicircles, with hybrid lines retaining mHVRs from both parents for over 800 generations. In contrast, maxicircles were exclusively inherited from one parent. Unexpectedly, we observed an increase in kDNA content in hybrids, affecting both minicircles and maxicircles, and exhibiting instability over time. To explain these findings, we propose a Replicative Mixing (REMIX) model, where the hybrid inherits one kinetoplast from each parent and they are replicated allowing minicircle mixing. Instead maxicircle networks remain physically separated, leading to uniparental fixation after segregation in the first cell division of the hybrid. This model challenges previous assumptions regarding kDNA inheritance and provides a new framework for understanding kinetoplast dynamics in hybrid trypanosomes. Full article

High temperatures are one of the most important abiotic stressors affecting the survival and growth of American shad (Alosa sapidissima). Building on previous omics sequencing studies of A. sapidissima liver and gills under high temperature stress, this study focused on investigating the regulatory role of miR-1236-3p and its target gene hsp90b1. The results indicate that the full-length cDNA of the hsp90b1 gene is 2023 bp and comprises a 5’ end of 58 bp, a 3’ end of 84 bp, and a coding region of 1881 bp, encoding 626 amino acids. Sequence alignment and phylogenetic tree analysis reveal that the hsp90b1 sequence is highly conserved across species. In situ hybridization showed that hsp90b1 is mainly localized in the cytoplasm. Software prediction identified a potential binding site between miR-1236-3p and hsp90b1. Through the construction of wild-type and mutant 3’UTR hsp90b1 dual luciferase reporter plasmids, the targeted relationship between the two was confirmed. In addition, the spatiotemporal expression levels of the hsp90b1 was found to be highest in the multicellular stage and liver tissue at a cultivation temperature of 27 °C; miR-1236-3P was highly expressed in the hatching stage and heart tissue at 30 °C. These findings provide a theoretical foundation for further investigating the regulatory role of non-coding RNA in A. sapidissima heat stress and offer data for subsequent molecular breeding studies. Full article

Microbial forensics involves analyzing biological evidence to evaluate weaponized microorganisms or their toxins. This study aimed to detect and type Yersinia pestis from four simulated forensic samples—human plasma diluted in phosphate-buffered saline (#24-2), tomato juice (#24-5), grape juice (#24-8), and a surgical mask (#24-10). Notably, samples #24-10 may have contained live bacteria other than Y. pestis. A real-time polymerase chain reaction confirmed the presence of Y. pestis in all samples; however, whole-genome sequencing (WGS) coverage of the Y. pestis chromosome ranged from 0.46% to 97.1%, largely due to host DNA interference and low abundance. To address these limitations and enable strain-level identification, we designed a hybridization-based target enrichment approach focused on multilocus variable number tandem repeat analysis (MLVA). Next-generation sequencing (NGS) using whole-genome amplification revealed that the accuracy of the 25 MLVA profiles of Y. pestis for samples #24-2, #24-5, #24-8, and #24-10 was 4%, 100%, 52%, and 0%, respectively. However, all samples showed 100% accuracy with target-enriched NGS, confirming they all belong to the same strain. These findings demonstrate that a targeted enrichment strategy for MLVA loci can overcome common obstacles in microbial forensics, particularly when working with trace or degraded samples where conventional WGS proves challenging. Full article

A novel species of the genus Methylocystis is proposed based on polyphasic evidence from strain SC2^T, isolated from the heavily polluted Saale River near Wichmar, Germany. Digital DNA–DNA hybridization and phylogenomic analyses demonstrate that strain SC2^T represents a distinct species within the genus, clearly separated from its closest relatives, namely Methylocystis suflitae NLS-7^T, Methylocystis rosea SV97^T, Methylocystis silviterrae FS^T, and Methylocystis hirsuta CSC1^T. As is typical of the family Methylocystaceae, cells possess intracytoplasmic membranes arranged parallel to the cytoplasmic membrane, and the dominant fatty acids are C18:1ω8c and C18:1ω7c. The strain grows aerobically on methane as the primary carbon and energy source and expresses both low- and high-affinity particulate methane monooxygenase (pMMO), but lacks the soluble form. The species epithet reflects the strain’s ability to utilize hydrogen as an alternative energy source. A further feature is its use of asparagine as an osmoprotectant, enhancing salt tolerance. Genomic analysis reveals complete pathways for nitrogen fixation, denitrification, and hydrogen oxidation. These genetic and physiological characteristics support the designation of a novel species, for which the name Methylocystis hydrogenophila sp. nov. is proposed. The type strain is SC2^T (=DSM 114506 = NCIMB 15437). Full article