Abstract: Extracellular vesicles (EVs) can disseminate replication-competent viral genomes complexed with selected host proteins, enabling stealth cell-to-cell transfer within lipid membrane-enclosed bubbles. In addition to complementing free-virion spread, EV-associated genomes can be protected from neutralizing antibodies and persist under conditions in which classical virion production decreases. Here, we propose a route-resolved framework in which interconnected cellular secretory pathways, including endoplasmic reticulum (ER) remodeling, multivesicular body (MVB) biogenesis, secretory autophagy, and plasma-membrane budding, jointly generate EV heterogeneity and create discrete opportunities for the capture, protection, and export of infectious cargo. We highlight reticulon-3 (RTN3), an ER-shaping protein, as an upstream regulator that can couple infection-induced ER microdomains to endosomal docking and autophagy-linked trafficking decisions that bias intermediates toward secretion rather than degradation. Supporting this view, transmission electron microscopy of dengue virus-infected cells reveals extensive vesicular remodeling, including irregular MVBs adjacent to the plasma membrane and autophagosome-like double-membrane structures, consistent with altered vesicular routing following RTN3 perturbation. Collectively, these route-resolved, spatially organized spatio-organelle changes support a pathomechanistic model in which RTN3-mediated ER remodeling reshapes ER-endosome-autophagy trafficking interfaces, creating regulated decision points that can be leveraged to stratify infectious EV subsets (with infectivity-linked single-vesicle and quantitative proteomics approaches) and to inform host-directed strategies that curb non-lytic viral dissemination.

Abstract: Developmental processes are usually described through dynamical systems and gradient-driven cellular rearrangements, yet their topological constraints are not well characterized. We introduce a mathematical approach linking morphogenesis with the Gömböc, a convex body whose equilibrium structure is minimal under topological constraints. We model developmental dynamics as gradient flows defined on a configuration space of tissue states where a morphogenetic potential integrates mechanical, chemical and adhesive cellular interactions. To explore how varying landscape parameters affect the stability of critical configurations and developmental trajectories, we simulated morphogenetic systems governed by gradient flows with Morse-type potentials. We found that systems approaching minimal critical-point structures display large basins of attraction and convergent trajectories despite diverse initial states. Developmental systems may operate near Gömböc-like dynamical regimes in which the topological properties of the configuration space constrain the number of accessible states, while attractors and gradient dynamics may induce a causal order. Our framework generates testable predictions. Developmental trajectories should concentrate into a small number of preferred channels, with transverse dispersion showing an exponential decay over time. In exponential morphogen gradients, migration time is expected to scale approximately linearly with the initial distance from the source. Saddle-like transitional configurations should appear as intermediate states in morphogenetic landscapes, detectable as brief phases of reduced migration speed and increased directional fluctuations. Overall, a quantitative framework is provided for analyzing developmental robustness, identifying transition bottlenecks in morphogenetic landscapes and predicting how physical or biochemical parameters could reshape developmental trajectories in synthetic and regenerative contexts.

Abstract: TBEV is a major cause of viral central nervous system infections in Europe, with heterogeneous geographical distribution and substantial underdiagnosis in low-incidence regions. This study aimed to evaluate the validity of regional TBE risk classification in Poland by combining surveillance-based incidence data with serological markers of TBEV exposure. Plasma samples from 5,541 blood donors residing in nine regions were tested by anti-TBEV IgG ELISA, followed by confirmatory VNT, IFA and anti-NS1 IgG ELISA to differentiate infection-induced from vaccine-induced antibodies. Regions were classified based on average TBE incidence from 2015–2019. Overall, anti-TBEV IgG screening reactivity was detected in 4.9% of donors, with significant regional variation (p < 0.001). The highest seroprevalence was observed in highly affected regions; however, unexpectedly elevated seroprevalence was also detected in regions classified as low affected. Markers consistent with TBEV infection (anti-NS1 IgG) were identified in only 2.6% of donors, whereas vaccine-induced immunity accounted for the majority of seropositive results. Male sex was independently associated with higher odds of seropositivity. Our findings suggest that passive surveillance data alone may insufficiently capture population-level exposure to TBEV, particularly in regions considered non-endemic. Integrating sero-epidemiological data with surveillance systems may improve risk assessment and inform targeted prevention strategies.

Abstract: Background/Objectives: Nuclear receptor corepressors NCOR1 and NCOR2 are key regulators of transcriptional repression, chromatin remodelling, and immunometabolic signalling. While NCOR1 has already been linked to vascular biology, its relevance in abdominal aortic aneurysm (AAA) remains unclear, particularly for NCOR2. This study aimed to investigate the expression, cellular localisation, and molecular interactions of NCOR1/2 in human AAA tissue. Methods: Human AAA samples (elective and ruptured) (n=45) and non-aneurysmal control aortas (n=18) were obtained from our Swiss Vascular Biobank. Transcriptomic profiling was performed using ribosomal RNA-depleted RNA sequencing. Differential expression and correlation analyses were performed using DESeq2/EdgeR and Spearman rank correlation with Benjamini–Hochberg correction. Cellular localisation was assessed through immunohistochemistry (IHC). Results: Bulk transcriptomic analyses showed no significant differences in NCOR1 or NCOR2 expression between AAA and controls. IHC revealed that NCOR1 was found in endothelial cells (ECs), smooth muscle cells (SMCs), and inflammatory infiltrates, while NCOR2 was primarily associated with macrophages. Correlation analyses suggest NCOR1 linking with various cellular markers, proteolytic enzymes, inflammatory mediators, and epigenetic regulators, including lncRNA MALAT1. NCOR2 showed distinct associations with remodelling enzymes, TGFB1 signalling, selective epigenetic modifiers, and lncRNA H19. Conclusions: The lack of transcriptional differences in NCOR1 and NCOR2 between AAA and controls does not exclude cell-type-specific regulation or functional relevance. The specific cellular distributions and molecular associations in human AAA imply that NCOR1 and NCOR2 play non-redundant roles in vascular remodelling, inflammation, and epigenetic regulation. Our findings highlight NCOR pathways as potential modulators of AAA pathophysiology and promising targets for future therapies.

Abstract: Haploid embryos constitute a valuable model for genetic and epigenetic studies; however, their developmental competence is reduced compared with diploid counterparts. This study evaluated whether supplementation of the culture medium with specific small molecules could improve developmental competence and outgrowth establishment of parthenogenetic haploid embryos. The effects of TGF-β inhibition (A83-01), WNT pathway modulation (CHIR99021 and IWR-1), and activin A (AA) supplementation were assessed from the morula stage onward under serum-free conditions. A83-01 treatment did not improve blastocyst formation or morphology and was associated with reduced total cell numbers relative to IVF controls. CHIR99021 supplementation increased the number of SOX2-positive cells compared with IWR-1 and vehicle-treated embryos, suggesting partial support of pluripotency; however, overall developmental progression remained inferior to diploid controls. In contrast, activin A significantly increased the proportion of haploid morulae developing into blastocyst and improved hatching rates. Nevertheless, AA supplementation did not restore CDX2-positive cell numbers or total cell counts to diploid levels. Furthermore, neither CHIR99021 nor AA affect DNA fragmentation levels, although a tendency toward increased TUNEL-positive cells was observed. Activin A treatment also failed to improve embryonic outgrowth formation. Collectively, these findings demonstrate that although activin A enhances blastocyst yield and hatching in bovine haploid embryos, modulation of TGF-β or WNT signaling alone is insufficient to restore diploid-like proliferative developmental competence.

Abstract: The question of how consciousness arises from physical systems remains one of the most profound challenges in neuroscience and philosophy. This essay examines two leading models that attempt to explain the emergence of consciousness from both biological and synthetic neural networks: Integrated Information Theory (IIT) and Global Workspace Theory (GWT). Each offers a distinct approach—one grounded in intrinsic informational structure, the other in functional accessibility and cognitive architecture. By comparing their principles, empirical support, and criticisms, this essay aims to clarify how these models contribute to our understanding of consciousness and its potential replication in artificial systems. Recent adversarial testing reveals that both theories face substantial empirical challenges, suggesting the field may need to resolve fundamental conceptual questions before definitive adjudication between theories becomes possible.

Abstract:

Toxoplasma gondii, the causative agent of toxoplamosis, a disease widely distributed, is an intracellular parasite that invades host cells of different tissues using specialized endocytic activity. Recent studies suggest that tunneling nanotubes (TNTs), thin cell surface projections, may participate in the parasite-host cell interaction process. We report results on the involvement of host cells TNTs in the adhesion and internalization of T. gondii tachyzoites to epithelial LLC-MK² cells. Microscopy analysis showed that incubating cells in 0.45 M sucrose induces reversible assembly of TNTs without affecting cell viability. The presence of extended TNTs correlated with increase on parasite adhesion and reduction of parasite entry, suggesting a structural or signaling role in mediating adhesion. TNTs assembled following sucrose incubation contain both actin and tubulin components. These results highlight the functional relevance of TNTs in T. gondii host cell interaction, especially in parasite adhesion, opening new perspectives for understanding T. gondii-host cell interaction.

Abstract: The origin of the transmembrane potential (TMP) in living cells is one of the foundational questions of cellular physiology. The dominant explanatory framework---the ion-pump model---attributes TMP to the active, \ATP-dependent displacement of ions (principally \Naplus\ and \Kplus) across the plasma membrane by dedicated protein complexes. This view, consolidated through the seminal work of Hodgkin and Huxley and the structural characterisation of the Na,K-ATPase, has shaped decades of research in neuroscience, cardiology, and cell biology. A competing framework, the \textit{murburn concept} developed by Manoj and colleagues, proposes a fundamentally different mechanism. According to this view, TMP is not the product of mechanical ion pumping but emerges spontaneously from asymmetric redox chemistry at the membrane interface. Diffusible reactive species (DRS), generated continuously during aerobic respiration, accumulate differentially on either side of the membrane, producing effective charge separation analogous to that observed in electrochemical cells. This perspective examines both frameworks critically, identifies the core points of disagreement, and evaluates the explanatory scope and empirical challenges of each. We argue that the murburn framework raises legitimate and underexplored questions about the thermodynamic sufficiency of the ion-pump model, and that a productive synthesis may lie in recognising redox chemistry as a primary contributor to membrane polarisation---rather than a secondary consequence of it.

Abstract: The successful application of 3D and 4D food printing is fundamentally governed by the rheology and microstructure of edible inks. These factors control every step, from extrusion and nozzle deposition to the final product functionality. This review systematically examines how formulation variables, including starch/protein composition, water content, and hydrocolloids, determine the network architecture and critical rheological properties, such as yield stress and viscoelasticity. These properties determine printing outcomes such as filament formation, stacking accuracy, and the stability of sensitive components. This review explores 4D printing as a "3D + 1D function," where printed structures provide additional features over time, such as a controlled color change or bioactive release, while post-printing treatment often activates these features. Through case studies of novel inks, we show how interfacial chemistry and process parameters influence texture and stability. Finally, we discuss the application of rheological metrics for predicting printability and outline the critical need for developing multi-parameter, process-relevant printability indices to advance the field of digital food manufacturing.

Abstract: Behcet disease (BD) is an inflammatory disorder with manifestation in mucosal tissues. Unlike autoimmune diseases that generate autoantibodies, BD is believed to be an autoinflammatory disease triggered by innate immune cells rather than adaptive cells. Hyperactivation of neutrophils causes vasculitis and thrombosis, and they migrate into cutaneous and ocular lesions. Dominance of M1 macrophages promotes the differentiation of Th1 cells. Moreover, the cross-reaction of bacterial heat shock proteins induces production of cytokines such as IL-4 and IFN-γ, in γδT cells, which alters the balance between Th1 and Th2 phenotypes. Nevertheless, natural killer (NK) cells play more critical roles in BD pathogenesis than other innate immune cells because not only their activity is precisely controlled by the interaction between ligands and receptors but NK1 shift also elicits Th1 dominance. The genetic factors associated with BD are HLA-B51 and major histocompatibility complex class I-related chain A (MICA), which stimulate NK receptors as ligands. Improperly processed peptides dysregulate their interaction with NK receptors, triggering the inflammatory response. NK1 and NK2 subsets represent cytokine production in relapse and remission periods; however, the cytotoxicity of NK cells in relapse is lower than that in remission periods. It still remains unclear how NK cells are activated recurrently and expand cytokine production. This review highlights the regulation of gene expression encoding NK receptors, tissue-resident NK cells, and adaptive NK cells to discuss their potential for relapsing. Splicing variants and readthrough genes encoding NK receptors easily alter cytokine production. Moreover, tissue-resident NK cells in mucosal tissues and adaptive NK cells that memorize the virus infection have the potential to trigger hyperactivation in relapse.

Abstract: Fusarium spp. are active producers of mycotoxins that enter the food chain and pose risks to human health. Identifying pathogenic agents is a key step in developing disease management strategies. For the first time in Bulgaria, we identified Fusarium species in wheat, harvest 2024÷2025, through the application of DNA barcoding. For genetic marker and construction of phylogenetic tree, the protein-coding gene β-tub was chosen. Among 26 identified isolates, F. sporotrichioides (42.3%) dominated, followed by F. proliferatum 23.1%), F. avenaceum (7,7%), F. armeniacum (7.7%), аnd F. poae (7,7%). F. tricinctum (3.8%), F. oxisporum (3.8%), and F. equiaseti (3.9%) were weakly expressed. Phylogenetic analysis classified the isolates into 5 species complexes: FSAMSC, FFSC, FTSC, FIESC, and FOSC and highlighted the genetic distances between them.Molecular genetic analysis showed that 84.6% of the wheat samples contained only one species of Fusarium, and in 15.4% the co-presence of two species was established. The largest share was in samples with low infestation 2÷4%, which represented 35% (n=32) of all positives. No statistically significant difference was found between the varieties and the level of contamination, as well as between the origin of the selected varieties and the level of contamination.

Abstract: Traumatic exposure does not uniformly lead to persistent posttraumatic symptoms, suggesting that vulnerability depends on more than event intensity alone. The Symbolic Objectification Hypothesis (SOH) proposes that an important determinant of traumatic outcome is the representational form in which threat is processed during activation. Specifically, risk increases when threat cannot be maintained as a bounded, identifiable, and cognitively manipulable object while executive continuity remains intact. Under these conditions, threat is more likely to become immersive, increasing the probability of defensive capture. SOH integrates findings showing that threat activation is graded and can coexist with organized cognition, and it proposes a mechanism linking representational failure to persistent re-experiencing. When symbolic objectification breaks down during encoding or reactivation, attentional narrowing increases, sequential processing becomes less stable, and temporal and contextual integration are weakened. Subsequent retrieval may therefore be more likely to evoke a present-oriented reliving state rather than an autobiographically situated memory. The hypothesis is operationalized through repeated indicators of symbolic objectification maintenance and restoration during activation, together with performance-based markers of discontinuity and recovery that index defensive capture. SOH generates falsifiable predictions regarding trauma vulnerability, symptom persistence, and treatment response. Clinically, the model suggests that trauma-focused interventions may be strengthened by pairing exposure-based engagement with explicit training in symbolic objectification, with the goal of reducing immersive threat experience while preserving executive continuity.

Abstract: This review explores the modulation of the host cellular flexibility “kinome" (protein kinases) and "phosphatome" (protein phosphatases) by dietary nutrients and gut microbiota metabolites, proposing a potential paradigm in the strategies for healthy aging and metabolic disease prevention. While mainstream nutrition approaches focus on population-wide guidelines, precision nutrition exploits the innovations in personal molecular networks and systems medicine, integrating genomics and metabolomics to address "metabolic rigidity"—the cell inability to switch between fuel sources. The review examines how master molecular regulators like AMPK and mTOR, and "metabolic brakes" like PTP1B and PTEN, are affected by single nucleotide polymorphisms (SNPs) and microbial signals (SCFAs, secondary bile acids, indoles). Specifically, the "microbial kinomic interference" hypothesis is discussed, where gut metabolites act as remote ligands for host signaling enzymes. Finally, the potential role of a personalized phosphoproteomics strategy is highlighted as an effective functional readout to guide nutritional interventions, aiming to restore metabolic plasticity through a gut microbiota/multi-omics approach.

Abstract: In this study, we compared the effects of microwave-assisted extraction (MAE) and ultrasound-assisted extraction (UAE) on the total phenolic content, antioxidant activity, morphological characteristics, and identification of the bioactive compounds in pomegranate seeds. We conducted a phytochemical characterization of the extracts by determining the total phenolic content and total flavonoids. Antioxidant activity was evaluated using ferric reducing antioxidant power (FRAP) and free radical inhibition methods (DPPH and ABTS). Morphological characteristics were analyzed via scanning electron microscopy, UV-Vis and FTIR of the extracts were recorded. Additionally, the main bioactive compounds were identified using HPLC-MS. Our results demonstrated that MAE was the most efficient technique, yielding a higher content of total phenols (35.47 mg GAE/g), total flavonoids (14.44 mg CAE/g) and antioxidant activity (0.19 and 0.41 mmol TEAC/g, as determined by FRAP and ABTS, respectively). In terms of morphological characteristics, UAE induced more changes in the structure of the plant material compared to MAE. According to HPLC-MS analysis, the extract obtained using MAE notably contained coumaric acid, cyanidin, and quercetin, whereas the UAE extract included coumaric acid, cyanidin, kaempferol, and epicatechin. In conclusion, this study demonstrated that MAE is a more efficient method than UAE for extracting bioactive compounds. Pomegranate seeds may represent a potential source of these compounds for application in various industrial areas.

Abstract: What if cotton could grow already colored — eliminating the need for dyes altogether? Today nearly all cotton is harvested white and later dyed using chemical processes that account for roughly 17–20% of global industrial water pollution. Billions of liters of water and large quantities of synthetic chemicals are used each year simply to give fabrics their color. This work explores a transformative alternative: cotton that produces its own colors while growing. We present a unified biological design framework for cotton fibers capable of naturally producing six shades — the existing brown and green, along with engineered pink, blue, and, for the first time, black cotton. Instead of dyeing fabric after harvest, the plant itself is programmed to create pigments directly inside the fiber. A key innovation is a dual-pigment strategy that enables the production of black cotton by combining two natural pigment systems commonly found in plants and biological materials. By carefully activating these pathways only in the developing fiber, the plant can generate stable coloration without affecting normal growth. Beyond proposing the concept, this study provides a practical roadmap for turning naturally colored cotton into a real agricultural technology. The framework outlines the full journey from laboratory design to field deployment, including gene construction, plant transformation, greenhouse testing, field trials, regulatory approval, and large-scale seed production. Methods for combining color traits with existing pest-resistant cotton varieties are also discussed to ensure compatibility with modern farming. If successfully implemented, naturally colored cotton could dramatically reduce the environmental footprint of the textile industry by eliminating large portions of the dyeing process. In the long term, this approach points toward a future where the colors of clothing are not manufactured in factories but grown directly in the field.

Abstract: Background: Aging is shaped by interdependent molecular processes captured by the hallmarks framework, in which epigenetic alterations stand out as a potentially modifiable regulatory layer. DNA methylation (DNAm) patterns change with age and can be summarized by epigenetic clocks that estimate biological age, pace of aging, and risk-related phenotypes. Yet, the extent to which interventions reproducibly modulate DNAm-based biomarkers across tissues and species remains uncertain. Methods: A systematized review of longitudinal intervention studies (2010–2025; English/Spanish) was conducted in PubMed, Scopus, and Cochrane CENTRAL, with selection documented using PRISMA. Human eligibility included randomized controlled trials (RCTs), non-randomized controlled studies, and pre–post designs (n≥10; adults ≥18 years). Preclinical eligibility included longitudinal mammalian studies (n≥5 per group). Outcomes were changes in DNAm-based epigenetic age (years) and/or pace of aging (e.g., DunedinPACE). Data were extracted into a standardized matrix (clock, tissue, effect direction/magnitude, safety, RoB_overall) and synthesized narratively; meta-analysis was not performed due to heterogeneity. Results: Thirty-five longitudinal studies were included (29 human, 6 preclinical). Lifestyle interventions in humans generally showed modest effects, with more consistent signals when exposure was sustained and accompanied by plausible physiological changes (e.g., prolonged calorie restriction affecting DunedinPACE, with effect sizes up to d=−0.43 at 12 months and d=−0.40 at 24 months in higher-adherence participants). Exogenous compounds showed higher heterogeneity and mixed evidence, including robust null epigenetic findings in some trials (e.g., metformin adjusted ITT differences ranging from −0.91 to +0.82 years across clocks, all p≥0.18) alongside favorable signals in smaller analytic subsets or open-label settings (e.g., bezisterim sub-study with reductions of −3.68 years in SkinBloodAge, −5.00 in Hannum, and −4.77 in InflammAge). Blood/circulation-derived interventions produced some of the largest reported effect sizes but also raised interpretation challenges: therapeutic plasma exchange with a sham arm reported epigenetic age decreases of ~1.3–2.6 years depending on the clock and regimen, with pronounced shifts in immune/inflammation-sensitive clocks; the apparent benefits waned after treatment cessation. Unexpectedly, repeated plasmapheresis in donors was associated with increases in several clocks and DunedinPACE per procedure (~+0.16–0.26 years per session across GrimAge-family clocks and ~0.003±0.001 DunedinPACE units per session). In rodents, plasma fractions/exosome-rich preparations and heterochronic parabiosis reported large percentage reductions across tissues, with strong dependence on exposure duration and concerns about translational uncertainty (up to ~77.6% in liver and ~68.2% in blood in one plasma-fraction study). Evidence for partial reprogramming (OSKM) was limited to a single rat study with small, near-significant trends in hippocampus-based clocks (two-sided p=0.064–0.088 across three clocks). Conclusions: DNAm-based epigenetic biomarkers are modifiable by interventions in mammals, but effects are heterogeneous and depend on the intervention, clock construct (age vs pace/risk signatures), biological matrix, tissue, follow-up duration, and study design. A single notion of “epigenetic rejuvenation” is not supported; instead, intervention effects appear domain-specific and must be interpreted in relation to what each clock measures.

Abstract: The integration of three-dimensional (3D) printing and virtual surgical planning (VSP) has introduced a new standard in spinal surgery, enabling highly individualized preoperative preparation and intraoperative execution. By virtually reconstructing patient anatomy, surgeons can identify critical vascular corridors, analyze endplate morphology, determine optimal interbody cage geometry for best anatomical fit, and predefine screw trajectories and entry points with exceptional accuracy. This level of planning reduces intraoperative uncertainty and minimizes the risk of implant malposition, particularly in anatomically complex scenarios, such as the current case report. Quantitative assessment of alignment correction can also be performed preoperatively, allowing the surgeon to model and predict postoperative spinal balance with greater precision. This technology is particularly advantageous in congenital spinal anomalies, such as hemivertebrae, where orientation can be challenging and anatomical landmarks are often distorted or partially obscured. We report the application of 3D printed patient-specific implants (PSIs) in conjunction with VSP to perform a single-level, stand-alone anterior lumbar interbody fusion (ALIF) for the treatment of congenital scoliosis due to a left hemivertebra, associated with progressive left L5/S1 foraminal stenosis. The patient presented with severe left L5 radicular pain and motor–sensory radiculopathy. The use of a PSI facilitated precise reconstruction of the spinal column, optimized implant fit and enhanced surgical accuracy in this technically complex case.

Abstract: An extensive survey of early blight of tomato was conducted in Peshawar and Hazara divisions of Khyber Pakhtunkhwa province, during the fruit bearing period of 2014 of the crop. Comparatively more disease incidence and severity was observed in Peshawar than Hazara Division. Data also revealed that more disease was prevalent in district Haripur than Abbottabad and Mansehra. The isolates of Alternaria solani collected during the survey were different in terms of their cultural characteristics and aggressiveness. Isolates from Peshawar division showed rapid growth on Potato Dextrose Agar medium and produced higher number of spores ml−1 as compared to isolates collected from Hazara Division. Moreover these also produced the largest size lesion (20.6mm) when compared with those collected from Hazara Division. A positive linear trend was observed when lesion size was regressed over colony diameter and spores concentration indicating that isolates showing aggressiveness also showed more radial growth and produced more spores mlˉ1. The studies also confirmed the existence of cultivar specific aggressiveness amongst the isolates of A. solani in screen house experiment. Isolates adapted on respective cultivars caused high disease severity, number of lesions per plant and lesion size with concurrent reduction in yield. Isolate AsRJ previously adapted on variety Red Jambo when inoculated on the same variety produced high disease severity (64.02%) and lesion size (8.2mm), with the lowest yield (436.71g). A similar trend was observed for other isolate and cultivar combinations which could have serious implications for cultivation of a particular variety on vast acreages over time.

Abstract: Inferring demographic history from whole-genome data is a fundamental objective in evolutionary and conservation genomics. However, the Pairwise Sequentially Markovian Coalescent (PSMC) framework, the most widely used demographic inference method for whole-genome sequence data, is highly sensitive to sequencing coverage, with low coverage producing systematic underestimation of heterozygosity and biased effective population size trajectories. Here, we present PSMC-FAC, an automated method designed to optimize false-negative rate correction in low-coverage genomes by minimizing geometric distances between corrected and high-coverage demographic trajectories. Whole-genome datasets from humans, gray wolves, and cattle were downsampled across multiple coverage levels and processed through standard demographic inference pipelines. Corrected trajectories were compared using Hausdorff and discrete Fréchet distance metrics projected onto a common temporal grid, and optimal correction factors were modeled as a function of sequencing depth using polynomial regression. Across species and demographic contexts, PSMC-FAC markedly improved concordance between low- and high-coverage trajectories and revealed highly predictable coverage-dependent correction patterns. Overall, PSMC-FAC provides a reproducible and mathematically grounded alternative to subjective correction approaches, enabling reliable demographic inference from moderate-coverage genomes and facilitating broader population-scale genomic analyses.

Abstract: Background: Vitamin D (25(OH)D) deficiency affects over one billion people globally and is associated with type 2 diabetes (T2D) and cardiometabolic diseases. However, causal relationships remain unclear, as vitamin D supplementation have shown limited benefit in reducing T2D risk. Genetic studies have identified variants influencing circulating 25(OH)D levels, but whether genetically determined vitamin D status predicts cardiometabolic outcomes is still uncertain. We therefore used bidirectional Mendelian randomization with genome-wide polygenic scores to evaluate the causal relationship and directionality between vitamin D status and T2D. Methods and Results: We analyzed multi-ethnic populations from the UK Biobank (N = 471,861), and 3,486 participants from the Asian Indian Diabetic Heart Study/Sikh Diabetes Study with serum 25(OH)D measures and genome-wide genotype data. A global polygenic score of vitamin D–raising alleles did not significantly reduce the risk of T2D, coronary artery disease, stroke, or other cardiometabolic risk factors. In contrast, a higher T2D polygenic risk score (PRS) was strongly associated with increased risk for 25(OH)D deficiency (<50 nmol/L). Genetically instrumented analyses showed per SD increase in T2D PRS significantly reduced circulating 25(OH)D levels (β = −8.9 nmol/L; 95% CI: −9.3 to −8.5; p = 3.6 × 10⁻²⁸). Conclusions: Our findings suggest low circulating vitamin D levels are unlikely to causally predict T2D risk but may serve as a marker for secondary prevention in endocrine and cardiovascular health. Instead, genetic susceptibility to T2D appears to contribute to reduced vitamin D levels. Further studies are needed to clarify the mechanisms underlying vitamin D deficiency in diabetes.