Motivation Unique molecular identifiers (UMIs) enable efficient error correction in amplicon sequencing but UMI-aware analysis workflows for long-read sequencing and particularly for nanopore data are still sparse. Existing approaches lack portability, real-time sequencing support, GPU acceleration, and efficient use of resources.Results We present umi-pipeline-nf, a portable, fully containerized, modular and scalable workflow to create single-molecule consensus sequences from UMI-tagged long-read nanopore amplicon data. Umi-pipeline-nf supports flexible UMI-designs and is built in Nextflow DSL2 for seamless deployment across computing platforms and a high degree of parallelization, allowing analysis of several targets at once. It scales linearly from single samples to large cohorts, outperforming existing tools in efficiency and flexibility. Additionally, we integrated real-time read processing, robust UMI clustering, and GPU-accelerated consensus polishing. Umi-pipeline-nf supports two different polishing strategies [reference sequence-based and partial order alignment (POA)-based]. Implementation of GPU-accelerated, reference sequence-based polishing results in up to 100-fold runtime improvements and reduced usage of computational resources, compared to other UMI analysis pipelines and POA-based polishing.Availability and implementation The umi-pipeline-nf analysis pipeline and test data are available at https://github.com/genepi/umi-pipeline-nf, and a frozen snapshot is available at DOI: 10.5281/zenodo.18607956. Scripts and configuration files for the analyses in the present manuscript can be found at https://github.com/AmstlerStephan/umi-pipeline-nf_Paper.

Apolipoprotein A-IV (apoA-IV) plays key roles in lipid metabolism, reverse cholesterol transport, and kidney function, yet its genetic determinants remain poorly defined. We conduct a genome-wide association study (GWAS) meta-analysis of apoA-IV concentrations measured by ELISA in 25,181 individuals and combine these with proteomic data from 33,995 UK Biobank participants (Olink platform), yielding a total sample of 59,176. We perform genetic correlations and colocalization analyses to explore links with lipid, renal, and other complex traits. The GWAS identifies several novel loci to be associated with apoA-IV concentrations (TDRD5, DPP4, MYL3, MORC1, MCUB, GATA4, ZPR1, UMOD, GLP2R, SLC38A10 and APOE) besides two previously identified loci (APOA5-A4-C3-A1 cluster and KLKB1). Cross-platform comparison shows strong concordance of effect directions and magnitudes, underscoring the robustness of findings across measurement techniques. Global genetic correlation reveals significant shared genetic architecture between apoA-IV, kidney function and HDL-cholesterol, while colocalization supports shared causal variants with lipid, renal, and hematological phenotypes, suggesting biologically relevant pathways. These results provide a comprehensive overview of the genetic architecture of apoA-IV and suggest mechanistic links to lipid metabolism, kidney function and blood-related traits. Our findings refine the role of apoA-IV as a potential biomarker and inform future epidemiological research.

In the study at hand, 2-sample Mendelian randomization supported a causal effect of interleukin-6 receptor (IL-6R) signaling on Lp(a) concentrations, revealing a direct interplay between 2 causal risk factors for cardiovascular disease (CVD). Importantly, epidemiological analyses in UK Biobank showed that neglecting the IL-6R functional state can confound the association between IL-6 levels and Lp(a). However, the effect of circulating IL-6 on Lp(a) concentrations was modest even when the functional state of the receptor was considered. In line, factorial Mendelian randomization indicated that down-regulation of IL-6R signaling and Lp(a) independently reduce CVD risk, highlighting complementary opportunities for CVD risk mitigation.

India is the most populous country globally, yet genetic studies involving Indian individuals remain limited. The Indian population is composed of many founder groups and has a mixed genetic ancestry, including an ancestral component not observed anywhere outside of India. This presents a unique opportunity to uncover novel disease variants and develop tailored medical interventions. To facilitate genetic research in India, a crucial first step is to create a foundational resource that serves as a benchmark for future population studies and methods development. Thus, we constructed the largest and most nationally representative linkage disequilibrium (LD) and genotype imputation reference panels in India to date, using high-coverage whole-genome sequencing data of 2,680 participants from the Longitudinal Aging Study in India-Harmonized Diagnostic Assessment of Dementia (LASI-DAD). As an LD reference panel, LASI-DAD includes 69.5 million variants, representing 170% and 213% increases relative to the 1000 Genomes Project and TOP-LD South Asian panels, respectively. Besides serving as an LD lookup panel, LASI-DAD facilitates various statistical analyses relying on precise LD estimates. In polygenic risk score (PRS) analyses, LASI-DAD improved the PRS predictive performance by 2.1%-35.1% across traits and studies. As an imputation reference panel, LASI-DAD enhanced imputation accuracy, measured by the Pearson correlation between imputed and true genotypes, by 3%-101% (mean 38%) compared with the TOPMed panel and by 3%-73% (mean 27%) compared with the Genome Asia Pilot panel across different allele frequencies. The LASI-DAD reference panel is publicly available to benefit future studies.

BACKGROUND & AIMS:Molecular phenotyping of Crohn's disease (CD) trajectories after ileocolonic resection (ICR) enables the identification of evolving disease mechanisms and related biomarker discovery. Here, we aimed at identifying a pathogenic node with predictive value for endoscopic disease recurrence after ICR in CD. METHODS:We performed unbiased transcriptomics of the postoperative ileum, comparing patients with an endoscopic Rutgeerts score i0 vs those with ≥i2b (N = 36). A potential biomarker from this analysis was then validated at the protein level by quantitative confocal microscopy on tissue slides, and its value for predicting endoscopic recurrence at resection was assessed in 3 independent ICR cohorts (total N = 241). Functional implications and therapeutic potential were investigated in mouse enteritis models. RESULTS:Patients with endoscopic CD recurrence exhibited a transcriptional node characterized by reduced glutathione peroxidase 4 (GPX4) expression, which was confined to the intestinal epithelium. Reduced intestinal epithelial GPX4 expression was already observed at the time of ICR and predicted endoscopic recurrence in addition to established clinical risk factors in a patient-level meta-analysis. Intestinal epithelial GPX4 expression was a surrogate of its enzymatic activity, inversely correlated with a mucosal endoplasmic reticulum stress signature, but was unrelated to a GPX4 genetic variant associated with CD susceptibility, histologic severity of inflammation, or clinical risk factors for recurrence. In mice, reduced intestinal epithelial Gpx4 expression fueled endoplasmic reticulum stress and enabled severe enteritis, which was ameliorated by restoration of epithelial GPX4 expression with selenium supplementation. CONCLUSIONS:Collectively, our study identifies a druggable biomarker that improves the prediction of endoscopic CD recurrence, potentially guiding patient management and therapy in the future. One of the studied ICR cohorts was part of the REMIND (Groupe de Recherche sur les Maladies inflammatoires digestives) study, registered at ClinicalTrials.gov (NCT03458195).

Introduction:The rate of decline in estimated glomerular filtration rate (GFR, eGFR) is increasingly recognized as a quantitative marker of chronic kidney disease (CKD) progression. However, data on eGFR slopes have mainly been reported in cohorts enriched for fast progression and the heterogeneity of eGFR slopes across the spectrum of CKD remains poorly defined. Methods:In 5214 participants of the German CKD (GCKD) study, we modeled eGFR slopes using per-protocol and clinical measurements. We used linear-mixed effects models, with eGFR slope as the outcome and baseline demographics as independent variables to (i) describe eGFR slope heterogeneity; (ii) assess differences by CKD etiology, eGFR and urinary albumin-to-creatinine ratio (UACR) categories, sex, and age; and (iii) determine associations of slopes with estimated eGFR decline (30%, 40%, and 57%) and observed end points (kidney failure with replacement therapy, mortality). Results:On average, 9 eGFR values per participant (interquartile range: 7-12) over 6.5 years were used for slope calculation. The adjusted mean annual eGFR slope was -1.43 ml/min per 1.73 m2. Slopes were similar across eGFR categories, but steeper with higher UACR. Faster eGFR decline was observed in participants of younger age and in those with polycystic kidney disease or diabetic kidney disease (DKD). Although eGFR slopes did not consistently differ by sex, women with diabetes as the leading cause of CKD had lower slopes than their male counterparts. A rapid annual decline (> 5 ml/min per 1.73 m2) occurred in 4.3%, with variation in frequency by CKD cause and UACR. Conclusion:In conclusion, though the average eGFR slope was low, it varied considerably, depending on CKD etiology and UACR. This data may help to put slope estimates in individual patients and defined subpopulations into perspective.

Rationale & Objective: Afamin is a vitamin Ebinding glycoprotein primarily expressed in the liver and kidney. This study investigated whether serum afamin concentrations are associated with kidney function and incident kidney failure. Study Design: Prospective cohort study with 6.5 years follow-up. Setting & Participants: 5,041 White patients enrolled in the German Chronic Kidney Disease (GCKD) study with measured afamin concentrations and either an estimated glomerular filtration rate (eGFR) of 30-60 mL/min/1.73 m2 or an eGFR > 60 mL/min/1.73 m2 with a urinary albumin-creatinine ratio (UACR) of >= 300 mg/g at study entry. Exposure: Serum afamin concentrations (mg/L). Outcome: Incident kidney failure (initiation of kidney replacement therapy or kidney-related death). Analytical Approach: Generalized linear regression and quantile regression models fit to investigate the association of afamin concentrations with eGFR and UACR. Adjusted Cox regression analysis to examine the association of afamin concentrations with incident kidney failure. Results: The mean +/- SD afamin concentration at study entry was 73.2 +/- 17.6 mg/L. Higher afamin concentrations were associated with better kidney function with a 2.60 mL/min/1.73 m2 higher eGFR (95% CI, 2.30-2.8 9) and a 5.97 mg/g lower UACR (95% CI, 3.04-8.9 0) for each 10 mg/L higher level of afamin concentration in adjusted analysis. During the follow-up period, each 10 mg/L higher level of afamin concentration was associated with a 14% lower risk of kidney failure (HR, 0.86 [95%CI, 0.810.92], P < 0.001). Limitations: Residual confounding, and potential limited generalizability to non-White populations and people with mild stages of chronic kidney disease (CKD) or no CKD. Conclusions: Higher serum afamin concentrations appear to be associated with a higher eGFR, less albuminuria, and a lower risk for future kidney failure in patients with CKD.

Polygenic scores (PGSs) for body mass index (BMI) may guide early prevention and targeted treatment of obesity. Using genetic data from up to 5.1 million people (4.6% African ancestry, 14.4% American ancestry, 8.4% East Asian ancestry, 71.1% European ancestry and 1.5% South Asian ancestry) from the GIANT consortium and 23andMe, Inc., we developed ancestry-specific and multi-ancestry PGSs. The multi-ancestry score explained 17.6% of BMI variation among UK Biobank participants of European ancestry. For other populations, this ranged from 16% in East Asian-Americans to 2.2% in rural Ugandans. In the ALSPAC study, children with higher PGSs showed accelerated BMI gain from age 2.5 years to adolescence, with earlier adiposity rebound. Adding the PGS to predictors available at birth nearly doubled explained variance for BMI from age 5 onward (for example, from 11% to 21% at age 8). Up to age 5, adding the PGS to early-life BMI improved prediction of BMI at age 18 (for example, from 22% to 35% at age 5). Higher PGSs were associated with greater adult weight gain. In intensive lifestyle intervention trials, individuals with higher PGSs lost modestly more weight in the first year (0.55 kg per s.d.) but were more likely to regain it. Overall, these data show that PGSs have the potential to improve obesity prediction, particularly when implemented early in life.

Introduction The rate of decline in eGFR is increasingly recognized as a quantitative marker of chronic kidney disease (CKD) progression. However, data on eGFR slopes have mainly been reported in cohorts enriched for fast progression and the heterogeneity of eGFR slopes across the spectrum of CKD remains poorly defined. Methods In 5,214 participants of the German CKD (GCKD) study, we modeled eGFR slopes using per-protocol and clinical measurements. We used linear-mixed effects models, with eGFR slope as the outcome and baseline demographics as independent variables, to (i) describe eGFR slope heterogeneity, (ii) assess differences by CKD etiology, eGFR and UACR categories, sex and age, and (iii) determine associations of slopes with estimated eGFR decline (30%, 40%, and 57%) and observed endpoints (kidney failure with replacement therapy, mortality). Results On average 9 eGFR values/participant (IQR 7-12) over 6.5 years were used for slope calculation. The adjusted mean annual eGFR slope was -1.43mL/min/1.73m2. Slopes were similar across eGFR categories, but steeper with higher UACR. Faster eGFR decline was observed in participants of younger age and in those with polycystic or diabetic kidney disease. While eGFR slopes did not consistently differ by sex, women with diabetes as the leading cause of CKD had lower slopes than their male counterparts. A rapid annual decline (> 5 mL/min/1.73m2) occurred in 4.3%, with variation in frequency by CKD cause and UACR. Conclusion In conclusion, while the average eGFR slope was low, it varied considerably, depending on CKD etiology and UACR. This data may help to put slope estimates in individual patients and defined subpopulations into perspective.

CRISPR-dependent base editing (BE) enables the modeling and correction of genetic mutations at single-base resolution. Base editing screens, where point mutations are queried en masse, are powerful tools to systematically draw genotype–phenotype associations and characterise the function of genes and other genomic elements. However, the lack of user-friendly web-based tools for designing base editing screens can hinder broad technology adoption. Here, we introduce CRISPR-BEasy (https://crispr-beasy.cerc-genomic-medicine.ca), a free, automated web-based server that streamlines the creation of single guide (sg)RNA tiling libraries for base editing screens. Researchers can provide their genes or genomic features of interest, their base editors of choice, and target sequences to act as positive and negative controls. The server designs and annotates sgRNA libraries by integrating custom code with publicly available tools such as crisprVerse and Ensembl’s Variant Effect Predictor. CRISPR-BEasy provides downloadable results, including sgRNA on/off-target scores, predicted mutational outcomes per base editor, and intuitive interactive visualizations for data quality assessment. CRISPR-BEasy also provides a separate tool that assembles sgRNA libraries into oligonucleotides for cloning following the detailed protocol documented in the searchable web server manual. Together, CRISPR-BEasy ensures the seamless design of cloning-ready sgRNA libraries, seeking to democratise access to base editing screening technologies.

BACKGROUND & AIMS:Pathogenic variants in the cellular iron exporter ferroportin (SLC40A1) cause hepatic and splenic iron overload. Low to normal transferrin saturation (TSAT) and iron accumulation in Kupffer cells with high splenic iron distinguish ferroportin disease (FD) from SLC40A1-related hemochromatosis (SLC40A1-HC), which are both caused by variants in SLC40A1. The aim of our study was to describe pathogenic mutations in SLC40A1, phenotypic variability in affected patients and compare outcomes with HFE-related hemochromatosis (HFE-HC). METHODS:The international EASL non-HFE hemochromatosis patient registry prospectively collected clinical, radiological, biochemical, and genetic data for 95 patients with SLC40A1 variants from six centers. Additionally, 363 patients were identified by a systematic literature review. As a comparator, 603 patients diagnosed with HFE-HC were included. RESULTS:The FD phenotype presented in 65.5% of affected individuals. Patients with FD were younger at diagnosis and more often female than those with SLC40A1-HC. SLC40A1 variants were associated with higher hepatic and splenic iron concentrations compared to the HFE-HC group. Variability in phenotypic presentation was high among patients with SLC40A1 variants, and a genotype-to-phenotype correlation could only explain a small proportion of this variation. Variants that directly affect the metal binding site in ferroportin more likely presented with high TSAT. Patients with the SLC40A1-HC phenotype (TSAT >45%) had a higher risk of fibrosis. Life expectancy was similar between patients with SLC40A1 variants and matched patients with HFE-HC. Most individuals with SLC40A1 variants (73.2%) received regular phlebotomies, which were not associated with differences in life expectancy. CONCLUSIONS:Mutations in SLC40A1 cause a highly variable disease spectrum with hepatic and splenic iron overload. Fibrosis risk is higher in patients with elevated TSAT. IMPACT AND IMPLICATIONS:Clinical management of individuals with SLC40A1 variants has largely been extrapolated from HFE-related hemochromatosis despite fundamental pathophysiological differences. Our study provides detailed phenotypic characterization that supports diagnosis and distinction of these rare iron overload disorders. Long-term follow-up shows preserved life expectancy, unaffected by phlebotomy, underscoring the need to critically assess phlebotomy on an individualized basis. Patients with SLC40A1-related hemochromatosis (transferrin saturation >45%) had a higher prevalence of chronic liver disease than those with ferroportin disease, suggesting that elevated transferrin saturation and hepatic iron drive disease progression, which can guide risk stratification and clinical decision making. CLINICAL TRIAL NUMBER:Not applicable.

Background/Objectives: The implementation of polygenic scores (PGSs) and multifactorial risk assessments (MFRAs) has the potential to enhance breast cancer risk stratification, particularly in carriers of moderate-penetrance pathogenic variants (PVs), whose risk profiles often remain unclear if testing is limited to monogenic risk factors. Methods: To enhance breast cancer risk stratification, we included the BCAC313 polygenic score, together with MFRA, for carriers of moderate-penetrance pathogenic variants (PVs) during routine diagnostics and assessed its effect on the classification of patients’ risk categories in a real-world cohort at our center in its first year of implementation. Seventeen carriers with PVs in moderate-risk breast cancer genes were included in this study. Thirteen of them qualified for analysis for a full MFRA, including PGS, according to ancestry estimation and clinical criteria. The MFRA was performed using the CanRisk tool, which incorporates clinical, lifestyle, familial, and genetic data, including the BCAC313 score. Results: PGS z-scores were significantly higher in breast cancer patients compared to the unaffected control cohort (p = 0.016). The MFRA, including PGS, increased risk estimates for contralateral breast cancer in seven of eight patients with breast cancer and for primary breast cancer in three of five healthy carriers, compared to the risk conferred by the MFRA and moderate-penetrance pathogenic variant alone. Risk estimates varied widely, demonstrating the value of MFRA in personalized care. In five cases, one with a CHEK2-PV and four with an ATM-PV, the modified risk assessment contributed to the surgical decision for a prophylactic mastectomy. Conclusions: The MFRA, including PGS, provides the clinically meaningful refinement of breast cancer risk estimates in individuals with moderate-risk PVs. Personalized risk predictions can inform clinical management and support decision-making, which highlights the utility of this approach in clinical practice.

A hyperinflammatory state with highly elevated concentrations of inflammatory biomarkers such as C-reactive protein (CRP) is a characteristic feature of severe coronavirus disease 2019 (COVID-19). To examine a potential role of common genetic factors that may influence COVID-19 outcomes, we investigated whether individuals with a polygenic predisposition for a pro-inflammatory response (in the form of Polygenic Scores) are more likely to develop severe COVID-19. The innovative approach of polygenic scores to investigate genetic factors in COVID-19 severity should provide a comprehensive approach beyond single-gene studies. In our cohort of 156 patients of European ancestry, two overlapping Polygenic Scores (PGS) predicting a genetic predisposition to basal CRP concentrations were significantly different between non-severe and severe COVID-19 cases and were associated with less severe COVID-19 outcomes. Furthermore, specific single nucleotide polymorphisms (SNPs) that contribute to either of the two Polygenic Scores predicting basal CRP levels are associated with different traits that represent risk factors for COVID-19 disease initiation (ACE2 receptor, viral replication) and progression (CRP). We suggest that genetically determined enforced CRP formation may contribute to strengthening of innate immune responses and better initial pathogen control thereby reducing the risk of subsequent hyperinflammation and adverse course of COVID-19.

In early March 2020, a SARS-CoV-2 outbreak occurred in the ski resort of Ischgl, in Austria. After an initial seroprevalence study in April 2020, a follow-up study in November 2020 showed persistence of high levels of seropositivity. The impacts of SARS-CoV-2 infections and non-pharmaceutical interventions required to reduce transmission were wide-ranging, including worsened mental and physical health and economic damage. We analysed data from the Ischgl follow-up study. Of the 1,259 adults that participated in the Ischgl-1 study (Ischgl-1), 801 were followed-up. Seropositivity was defined using presence of binding and neutralizing antibodies at Ischgl-1. At follow-up, 7 months later (Ischgl-2), participants reported changes to self-rated mental and physical health, physical activity, alcohol consumption, smoking and economic status. Changes were compared by serological status, using multivariable logistic and multinomial regression models, where appropriate, and adjusting for factors including age, sex, and morbidity. 1 in 2 participants reported experiencing a moderate or severe impact of the pandemic. One fifth of participants reported a worsening in their mental health from November 2019 to November 2020; women and participants aged ≥35 to <70 years were disproportionately affected. Seropositivity was associated with a decline in physical health but no decline in mental health or behaviour changes. Very few participants reported any changes in behaviours. The overriding impact the population of Ischgl was economic-50% of participants reported a worsening of their professional and/or financial situation. Declines in self-reported mental health were associated with the overall experience of the pandemic and economic factors. The population of Ischgl demonstrated a high level of resilience to the pandemic as measured by health. However, certain segments of the population were disproportionately affected, particularly with regard to mental health and economic wellbeing. Future pandemic preparedness must consider how pandemic mitigation strategies can be responsive to context and the wider impacts on mental health and social and economic wellbeing while minimising mortality and safeguarding health systems.