Search Results (804)

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

Chromosomal microarray analysis (CMA) was performed for 40 patients with B-ALL undergoing treatment according to the ALL-2016 protocol to investigate the copy number alterations (CNAs) and copy neutral loss of heterozygosity (cnLOH) associated with minimal residual disease (MRD)-positive remission. Aberrations involving over 20,000 genes were identified, and a random forest approach was applied to isolate a subset of genes whose CNAs and cnLOH are significantly associated with poor therapeutic response. We have assembled the triple matched healthy population data and used that data as a reference, but not as a matched control. We identified a recurrent cluster of cnLOH in the 19q13.2–19q13.31 region, significantly enriched in MRD-positive patients (70% vs. 47% in the reference group vs. 16% in MRD-negative patients). This region includes the pregnancy-specific glycoprotein (PSG) gene family and the oncogene ERF, suggesting a potential role in leukemic persistence and treatment resistance. Additionally, we observed significant deletions involving 7p22.3 and 16q13, often as part of large-scale losses affecting almost the entire chromosomes 7 and 16, indicative of global chromosomal instability. These findings highlight specific genomic regions potentially involved in therapy resistance and may contribute to improved risk stratification in B-ALL. Our findings emphasize the value of high-resolution CMA in diagnostics and risk stratification and suggest that PSG genes and other candidate genes could serve as biomarkers for predicting treatment outcomes. Full article

Irradiation with ultraviolet light-emitting diodes (UV-LEDs) represents a promising method for viral inactivation, but a detailed understanding of the wavelength-dependent action spectra remains limited, particularly across different viral components. In this study, we established standardized UV action spectra for infectivity reduction in pathogenic viruses using a system equipped with interchangeable LEDs at 13 different peak wavelengths (250–365 nm). The reduction in viral infectivity induced by UV-LED exposure was strongly related to viral genome damage, whereas no significant degradation of viral structural proteins was detected. Peak virucidal efficiency was observed at 267–270 nm across all tested viruses, representing a slight shift from the traditionally expected 260 nm nucleic acid absorption peak. Enveloped RNA viruses, including influenza A virus, respiratory syncytial virus, and coronavirus, exhibited greater UV sensitivity than nonenveloped viruses such as feline calicivirus and adenovirus. These observations indicate that structural characteristics, such as the presence of an envelope and genome organization, influence UV susceptibility. The wavelength-specific action spectra established in this study provide critical data for optimizing UV-LED disinfection systems to achieve efficient viral inactivation while minimizing energy consumption in healthcare, food safety, and environmental sanitation. Full article

Background: Cell-free DNA (cfDNA) has become a cornerstone of liquid biopsy applications, offering promise for early disease detection and monitoring. However, its widespread clinical adoption is limited by variability in pre-analytical processing, especially during isolation. Current extraction methods face challenges in yield, purity, and reproducibility. Methods: We developed and optimized SafeCAP 2.0, a novel magnetic bead-based cfDNA extraction kit, focusing on efficient recovery, minimal genomic DNA contamination, and PCR compatibility. Optimization involved systematic evaluation of magnetic bead chemistry, buffer composition, and reagent volumes. Performance was benchmarked against a commercial reference kit (Apostle MiniMax) using spiked oligonucleotides and plasma from patients with stage IV NSCLC. Results: The optimized protocol demonstrated superior recovery with a limit of detection (LoD) as low as 0.3 pg/µL and a limit of quantification (LoQ) of 1 pg/μL with no detectable PCR inhibition. In comparative studies, SafeCAP 2.0 showed equivalent or improved performance over the commercial kit. Clinical validation using 47 patient plasma samples confirmed robust cfDNA recovery and fragment integrity. Conclusions: SafeCAP 2.0 offers a cost-effective, high-performance solution for cfDNA extraction in both research and clinical workflows. Its design and validation address key pre-analytical barriers, supporting integration into routine diagnostics and precision medicine platforms. Full article

Tellurite (TeO₃²⁻) is a highly soluble and toxic oxyanion that inhibits the growth of Escherichia coli at concentrations as low as ~1 µg/mL. This toxicity has been primarily attributed to the generation of reactive oxygen species (ROS) during its intracellular reduction by thiol-containing molecules and NAD(P)H-dependent enzymes. However, under anaerobic conditions, E. coli exhibits significantly increased tellurite tolerance—up to 100-fold in minimal media—suggesting the involvement of additional, ROS-independent mechanisms. In this study, we combined chemical-genomic screening, untargeted metabolomics, and targeted biochemical assays to investigate the effects of tellurite under both aerobic and anaerobic conditions. Our findings reveal that tellurite perturbs amino acid and nucleotide metabolism, leading to intracellular imbalances that impair protein synthesis. Additionally, tellurite induces notable changes in membrane lipid composition, particularly in phosphatidylethanolamine derivatives, which may influence biophysical properties of the membrane, such as fluidity or curvature. This membrane remodeling could contribute to the increased resistance observed under anaerobic conditions, although direct evidence of altered membrane fluidity remains to be established. Overall, these results demonstrate that tellurite toxicity extends beyond oxidative stress, impacting central metabolic pathways and membrane-associated functions regardless of oxygen availability. Full article

The CRISPR/Cas9 system is a powerful genome-editing tool that is applied in baculovirus engineering. In this study, we present the first report of the AcMNPV genome deletions for bioproduction purposes, using a dual single-guide RNA (sgRNA) CRISPR/Cas9 approach. We used this method to remove nonessential genes for the budded virus and boost recombinant protein yields when applied as BEVS. We show that the co-delivery of two distinct ribonucleoprotein (RNP) complexes, each assembled with a sgRNA and Cas9, into Sf9 insect cells efficiently generated deletions of fragments containing tandem genes in the genome. To evaluate the potential of this method, we assessed the expression of two model proteins, eGFP and HRPc, in insect cells and larvae. The gene deletions had diverse effects on protein expression: some significantly enhanced it while others reduced production. These results indicate that, although the targeted genes are nonessential, their removal can differentially affect recombinant protein yields depending on the host. Notably, HRPC expression increased up to 3.1-fold in Spodoptera frugiperda larvae. These findings validate an effective strategy for developing minimized baculovirus genomes and demonstrate that dual-guide CRISPR/Cas9 editing is a rapid and precise tool for baculovirus genome engineering. Full article

Prolonged expression of gene-editing components in CRISPR-modified plants can interfere with phenotypic analysis of target traits, increase the risk of off-target mutations, and lead to unnecessary metabolic burden. To mitigate these issues in peanut (Arachis hypogaea L.), callus-specific promoters were screened to restrict Cas9 expression to the callus stage, minimizing its activity in regenerated plants. In this study, six callus-specific genes in peanut were identified by mining RNA sequencing datasets and validating their expression profiles using quantitative reverse transcriptase PCR. The promoters of Arahy.H0FE8D, Arahy.WT3AEF, Arahy.I20Q6X, Arahy.ELJ55T, and Arahy.N9CMH4 were cloned and assessed for their expression activity. Beta-glucuronidase (GUS) histochemical staining confirmed that all five promoters were functional in peanut callus. Further investigation revealed their ability to drive cytosine base editing via a deaminase-nCas9 fusion protein, with all promoters successfully inducing precise base substitutions in peanut. Notably, P_Ah-H0FE8D, P_Ah-WT3AEF, P_Ah-ELJ55T, and P_Ah-N9CMH4 exhibited comparable or higher editing efficiencies than the commonly used cauliflower mosaic virus 35S promoter. These findings provide valuable tools for improving the biosafety of CRISPR-based genome editing in peanut breeding programs. Full article

The goal of this study is to develop an efficient machine learning framework for designing high-hardness multi-metal nitride coatings, overcoming the limitations of traditional trial-and-error methods. The development of multicomponent metal nitride hard coatings via multi-arc ion plating remains a significant challenge due to the vast compositional search space. Although theoretical studies in macroscopic, mesoscopic, and microscopic domains exist, these often focus on idealized models and lack effective coupling across scales, leading to time-consuming and labor-intensive traditional methods. With advancements in materials genomics and data mining, machine learning has become a powerful tool in material discovery. In this work, we construct a compositional search space for multicomponent nitrides based on electronic configuration, valence electron count, electronegativity, and oxidation states of metal elements in unary nitrides. The search space is further constrained by FCC crystal structure and hardness theory. By incorporating a feature library with micro-, meso-, and macro-structural characteristics and using clustering analysis with theoretical intermediate variables, the model enriches dataset information and enhances predictive accuracy by reducing experimental errors. This model is successfully applied to design multicomponent metal nitride coatings using a literature-derived database of 233 entries. Experimental validation confirms the model’s predictions, and clustering is used to minimize experimental and data errors, yielding a strong agreement between predicted optimal molar ratios of metal elements and nitrogen and measured hardness performance. Of the 100 Vickers hardness (HV) predictions made by the model using input features like molar ratios of metal elements (e.g., Ti, Al, Cr, Zr) and atomic size mismatch, 82 exceeded the dataset’s maximum hardness, with the best sample achieving a prediction accuracy of 91.6% validated against experimental measurements. This approach offers a robust strategy for designing high-performance coatings with optimized hardness. Full article

CRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats-associated protein 9)-mediated genome editing has emerged as a transformative tool in medicine, offering significant potential for cancer therapy because of its capacity to precisely target and alter the genetic modifications associated with the disease. However, a major challenge for its clinical translation is the safe and efficient in vivo delivery of CRISPR/Cas9 components to target cells. Nanotechnology is a promising solution to this problem. Nanocarriers, owing to their tunable physicochemical properties, can encapsulate and protect CRISPR/Cas9 components, enabling targeted delivery and enhanced cellular uptake. This review provides a comprehensive examination of the synergistic potential of CRISPR/Cas9 and nanotechnology in cancer therapy and explores their integrated therapeutic applications in gene editing and immunotherapy. A critical aspect of in vivo CRISPR/Cas9 application is to achieve effective localization at the tumor site while minimizing off-target effects. Nanocarriers can be engineered to overcome biological barriers, thereby augmenting tumor-specific delivery and facilitating intracellular uptake. Furthermore, their design allows for controlled release of the therapeutic payload, ensuring sustained efficacy and reduced systemic toxicity. The optimization of nanocarrier attributes, including size, shape, surface charge, and composition, is crucial for improving the cellular internalization, endosomal escape, and nuclear localization of CRISPR/Cas9. Moreover, surface functionalization with targeting ligands can enhance the specificity of cancer cells, leading to improved gene-editing accuracy. This review thoroughly discusses the challenges associated with in vivo CRISPR/Cas9 delivery and the innovative nanotechnological strategies employed to overcome them, highlighting their combined potential for advancing cancer treatment for clinical application. Full article

Thyroid cancer (TC) is the most common endocrine malignancy, with a rising global incidence. In Ecuador, TC rates are among the highest worldwide. Generally, fine-needle aspiration (FNA) remains the standard diagnostic tool; however, due to its limitations, alternative or complementary approaches are required. In this context, liquid biopsy, particularly circulating tumor DNA (ctDNA), offers a promising, minimally invasive option for tumor genotyping. Objective: This study evaluated the concordance between genetic variants identified in ctDNA and tumor tissue. Thirty-six women with papillary thyroid cancer were included. Tumor tissue and blood samples were collected, and DNA was extracted. Next-Generation Sequencing (NGS) using the TruSight Tumor 15 panel identified genetic variants in both ctDNA and tumor DNA. Variant pathogenicity was assessed following ACMG guidelines. Genetic ancestry was determined using Ancestry Informative Markers (AIMs). A total of 71 cancer-associated variants were detected, with 81.69% concordance between tumor DNA and ctDNA. TP53 was the most frequently mutated gene. While most pathogenic variants were found in tumor tissue, some variants appeared exclusively in ctDNA samples on specific patients, suggesting tumor heterogeneity. Ancestry analysis revealed a predominant Native American component (62.4%). Liquid biopsy demonstrates high concordance with tumor tissue analysis and holds potential as a complementary diagnostic tool for thyroid cancer. However, challenges such as low ctDNA yield and underrepresentation in genetic databases highlight the need for improved protocols and increased inclusion of admixed populations in genomic studies. Full article

Adeno-associated virus (AAV) vectors have emerged as powerful tools for in vivo gene therapy, enabling long-term transgene expression in targeted tissues with minimal pathogenicity. This review examines the AAV serotypes used in clinical gene therapy trials for neurodegenerative (central nervous system, CNS) diseases, highlighting their tropisms, engineering advances, and translational progress. We discuss how capsid modifications, cell-specific promoters, and novel delivery routes are enhancing AAV tropism and reducing immunogenicity to overcome current limitations. Key clinical trials in neurodegenerative disorders (such as Parkinson’s, Alzheimer’s, and Huntington’s disease) are summarized, including delivery methods (intravenous, intracoronary, intrathecal, etc.) and outcomes. We further outline the regulatory landscape with recent approvals of AAV-based therapies and ongoing efforts to address safety challenges like immune responses and vector dose toxicity. A more translational, forward-looking perspective is adopted to consider combination therapies (e.g., AAV with immune modulation or genome editing) and strategic directions to improve the next generation of AAV vectors. Overall, continued innovation in AAV vector design and delivery, alongside careful clinical evaluation, is accelerating the translation of gene therapies for neurodegenerative diseases. Full article

The Luhua chicken is an outstanding local breed in China that has been placed under conservation due to the impact of specialized breeding and the widespread adoption of commercial varieties. As such, this study analyzed reproductive traits across three consecutive generations and utilized whole-genome resequencing data from 60 Luhua chickens to assess conservation efficacy through genetic diversity, run of homozygosity (ROH) distribution, kinship, and population structure so as to better conserve the breed. The results show that, across generations, the body weight at first egg increased, the age at first egg was delayed, and the egg weight at first laying increased. No significant variations were found in the body weight at 300 d or the total egg number. The key genetic parameters of the polymorphism information content (PIC), expected heterozygosity (HE), observed heterozygosity (HO), and mean identical-by-state (IBS) distance were 0.234, 0.351, 0.277, and 0.782, respectively. The majority of ROHs ranged from 0.5 to 1 Mb, and the inbreeding coefficient based on ROHs was calculated at 0.021. The findings reveal that these traits remained unchanged across the three generations. Our research suggests that optimizing the mating plan of Luhua chickens is essential to minimize inbreeding risk. Furthermore, the methodology applied in this study provides a valuable reference for the conservation monitoring of other indigenous chicken breeds. Full article

Mitochondria are currently of great interest to scientists. The role of mitochondrial DNA (mtDNA) mutations has been proven in the genesis of more than 200 pathologies, which are called mitochondrial disorders. Therefore, the study of mitochondria and mitochondrial DNA is of great interest not only for understanding cell biology but also for the treatment and prevention of many mitochondria-related pathologies. There are two main trends of mitochondrial therapy: mitochondrial replacement therapy (MRT) and mitochondrial transplantation therapy (MTT). Also, there are two main categories of MRT based on the source of mitochondria. The heterologous approach includes the following methods: pronuclear transfer technique (PNT), maternal spindle transfer (MST), Polar body genome transfer (PBT) and germinal vesicle transfer (GVT). An alternative approach is the autologous method. One promising autologous technique was the autologous germline mitochondrial energy transfer (AUGMENT), which involved isolating oogonial precursor cells from the patient, extracting their mitochondria, and then injecting them during ICSI. Transmission of defective mtDNA to the next generation can also be prevented by using these approaches. The development of a healthy child, free from genetic disorders, and the prevention of the occurrence of lethal mitochondrial disorders are the main tasks of this method. However, a number of moral, social, and cultural objections have restricted its exploration, since humanity first encountered the appearance of a three-parent baby. Therefore, this review summarizes the causes of mitochondrial diseases, the various methods involved in MRT and the results of their application. In addition, a new technology, mitochondrial transplantation therapy (MTT), is currently being actively studied. MTT is an innovative approach that involves the introduction of healthy mitochondria into damaged tissues, leading to the replacement of defective mitochondria and the restoration of their function. This technology is being actively studied in animals, but there are also reports of its use in humans. A bibliographic review in PubMed and Web of Science databases and a search for relevant clinical trials and news articles were performed. A total of 81 publications were selected for analysis. Methods of MRT procedures were reviewed, their risks described, and the results of their use presented. Results of animal studies of the MTT procedure and attempts to apply this therapy in humans were reviewed. MRT is an effective way to minimize the risk of transmission of mtDNA-related diseases, but it does not eliminate it completely. There is a need for global legal regulation of MRT. MTT is a new and promising method of treating damaged tissues by injecting the body’s own mitochondria. The considered methods are extremely good in theory, but their clinical application in humans and the success of such therapy remain a question for further study. Full article

Circulating cell-free DNA (cfDNA) is an important biomarker for various cancer types, enabling a non-invasive testing approach. However, pre-analytical variables, including sample collection, tube type, processing conditions, and extraction methods, can significantly impact the yield, integrity, and overall quality of cfDNA. This study presents a comprehensive analytical validation of a magnetic bead-based, high-throughput cfDNA extraction system, with a focus on assessing its efficiency, reproducibility, and compatibility with downstream molecular applications. The validation was performed using a range of sample types: synthetic cfDNA spiked into DNA-free plasma, multi-analyte ctDNA plasma controls, Seraseq ctDNA reference material in a plasma-like matrix, extraction specificity controls, residual clinical specimen from patients, and samples from healthy individuals stored at room temperature or 4 °C for up to 48 h to assess stability. Extracted cfDNA was analyzed for concentration, percentage, and fragment size, using the Agilent TapeStation. Variant detection was evaluated using a next-generation sequencing (NGS) assay on the Seraseq ctDNA reference material. The results demonstrated high cfDNA recovery rates, consistent fragment size distribution (predominantly mononucleosomal and dinucleosomal), minimal genomic DNA (gDNA) contamination, and strong concordance between detected and expected variants in reference materials. The workflow also showed robust performance under different study parameters, variable sample conditions, including sample stability and integrity. Together, these findings confirm the efficiency and reliability of the evaluated cfDNA extraction system and underscore the importance of standardized pre-analytical workflows for the successful implementation of liquid biopsy for early cancer detection, therapeutic monitoring, and improved patient outcomes. Full article

Background and Objectives: Blood-borne inflammatory ratios have been proposed as inexpensive prognostic tools across a range of diseases, but their role in pulmonary tuberculosis (TB) remains uncertain. In this retrospective case–control analysis, we explored whether composite indices derived from routine haematology—namely the neutrophil-to-lymphocyte ratio (NLR), the platelet-to-lymphocyte ratio (PLR), the systemic immune–inflammation index (SII) and a novel CRP–Fibrinogen Index (CFI)—could enhance risk stratification beyond established cytokine measurements among Romanian adults with culture-confirmed pulmonary T. Materials and Methods: Data were drawn from 80 consecutive TB in-patients and 50 community controls. Full blood counts, C-reactive protein, fibrinogen, and four multiplex cytokines were extracted from electronic records, and composite indices were calculated according to standard formulas. The primary outcomes were in-hospital mortality within 90 days and length of stay (LOS). Results: Among TB patients, the median NLR was 3.70 (IQR 2.54–6.14), PLR was 200 (140–277) and SII was 1.36 × 10⁶ µL⁻¹ (0.74–2.34 × 10⁶), compared with 1.8 (1.4–2.3), 117 (95–140) and 0.46 × 10⁶ µL⁻¹ (0.30–0.60 × 10⁶) in controls. Those with SII above the cohort median exhibited more pronounced acute-phase responses (median CRP 96 vs. 12 mg L⁻¹; fibrinogen 578 vs. 458 mg dL⁻¹), yet median LOS remained virtually identical (29 vs. 28 days) and early mortality was low in both groups (8% vs. 2%). The CFI showed no clear gradient in hospital stay across its quartiles, and composite ratios—while tightly inter-correlated—demonstrated only minimal association with cytokine levels and LOS. Conclusions: Composite cell-count indices were markedly elevated but did not predict early death or prolonged admission. In low-event European cohorts, their chief value may lie in serving as cost-free gatekeepers, flagging those who should proceed to more advanced cytokine or genomic testing. Although routine reporting of NLR and SII may support low-cost surveillance, validation in larger, multicentre cohorts with serial sampling is needed before these indices can be integrated into clinical decision-making. Full article