Single Variant Analysis Research Articles

OP20 Expanding the genetic spectrum of inflammatory bowel disease through large-scale exome sequencing

Abstract Background Inflammatory bowel disease (IBD), including Crohn's disease (CD) and ulcerative colitis (UC), is highly heritable, with more than 50% of the variation in disease risk explained by genetics 1. Genome-wide association studies (GWASs) have identified over 300 IBD-associated loci 2, primarily underpinned by non-coding variants. Initial sequencing efforts have identified rare coding variants associated with CD 3, offering novel insights into disease mechanisms. However, the impact of low-frequency and rare coding variants in IBD remains underexplored. Methods We conducted single variant and meta-analysis across six different whole-exome/genome sequencing datasets. After quality control, 167,172 non-synonymous variants with minor allele frequency (MAF) between 0.0001 and 0.1 in gnomAD v4.1 (Non-Finnish European, NFE) were tested for association across 86,213 IBD cases (44,131 CD, 32,748 UC, and 9,334 IBD-uncertain) and 478,363 controls. Variants with P < 3×10-7 were considered significant. Additionally, very rare (MAF < 0.001) non-synonymous (NSYN) and protein-truncating variants (PTVs) were analysed for gene-level burden differences in each dataset and meta-analysed, with P < 2.5×10⁻⁶ as the threshold for significance. Results In the single variant analysis, we found 55 CD, 17 UC, and 20 IBD associated variants. After checking their linkage disequilibrium (LD) with known signals, 27 (CD), 14 (UC), and 19 (IBD) novel associations were defined (Figure 1). For example, a frameshift variant in TNFRSF6B (E45fs, MAF = 0.00055), missense variants in FAM120B (S431P, MAF = 0.0018) and IFNL2 (H32R, MAF = 0.0045) were significantly associated with CD, UC and IBD, respectively. Our results expand the list of TNF receptors involved in IBD susceptibility, and provide additional support to the role of IFNλ signalling on IBD pathogenesis. The burden test identified 18 genes (Figure 2) with a significantly different burden of very rare NSYNs or PTVs in CD (N = 9), UC (N = 4), or IBD (N = 12) cases versus controls. Among them, NOD2, ATG4C, ADCY7, and ITGAV had orthogonal support from GWASs, either from independent common coding variants or non-coding variants driving gene-expression differences, suggesting the joint contribution of common and rare-coding variants in these genes to IBD susceptibility. In addition, IBD patients showed an increased burden of very rare variants in IL10RA, a known monogenic IBD gene, expanding the phenotypic spectrum to include a more complex form of IBD. Conclusion In conclusion, we report novel rare variants and genes associated with IBD based on the meta-analysis of 86,213 IBD cases and 478,363 controls. This study provides novel insights into the biology of IBD and may prompt further investigation into new therapeutics.

Exome sequencing identifies genes for socioeconomic status in 350,770 individuals

Socioeconomic status (SES) is a critical factor in determining health outcomes and is influenced by genetic and environmental factors. However, our understanding of the genetic structure of SES remains incomplete. Here, we conducted a large-scale exome study of SES markers (household income, occupational status, educational attainment, and social deprivation) in 350,770 individuals. For rare coding variants, we identified 56 significant associations by gene-based collapsing tests, unveiling 7 additional SES-associated genes (NRN1, CCDC36, RHOB, EP400, NCAM1, TPTEP2-CSNK1E, and LINC02881). Exome-wide single common variant analysis revealed nine lead single-nucleotide polymorphisms (SNPs) associated with household income and 34 lead SNPs associated with EduYears, replicating previous GWAS findings. The gene-environment correlations had a substantial impact on the genetic associations with SES, as indicated by the significantly increased P values in several associations after controlling for geographic regions. Furthermore, we observed the pleiotropic effects of SES-associated genetic factors on a wide range of health outcomes, such as cognitive function, psychosocial status, and diabetes. This study highlights the contribution of coding variants to SES and their associations with health phenotypes.

Whole-genome sequencing identifies novel loci for keratoconus and facilitates risk stratification in a Han Chinese population.

Keratoconus (KC) is a prevalent corneal condition with a modest genetic basis. Recent studies have reported significant genetic associations in multi-ethnic cohorts. However, the situation in the Chinese population remains unknown. This study was conducted to identify novel genetic variants linked to KC and to evaluate the potential applicability of a polygenic risk model in the Han Chinese population. A total of 830 individuals diagnosed with KC and 779 controls from a Chinese cohort were enrolled and genotyped by whole-genome sequencing(WGS). Common and rare variants were respectively subjected to single variant association analysis and gene-based burden analysis. Polygenic risk score (PRS) models were developed using top single-nucleotide polymorphisms (SNPs) identified from a multi-ethnic meta-analysis and then evaluated in the Chinese cohort. The characterization of germline variants entailed correction for population stratification and validation of the East Asian ancestry of the included samples via principal component analysis. For rare protein-truncating variants (PTVs) with minor allele frequency (MAF) < 5%, ZC3H11B emerged as the top prioritized gene, albeit failing to reach the significance threshold. We detected three common variants reaching genome-wide significance (P ≤ 5 × 10-8), all of which are novel to KC. Our study validated three well known predisposition loci, COL5A1, EIF3A and FNDC3B. Additionally, a significant correlation of allelic effects was observed for suggestive SNPs between the largest multi-ethnic meta-genome-wide association study (GWAS) and our study. The PRS model, generated using top SNPs from the meta-GWAS, stratified individuals in the upper quartile, revealing up to a 2.16-fold increased risk for KC. Our comprehensive WGS-based GWAS in a large Chinese cohort enhances the efficiency of array-based genetic studies, revealing novel genetic associations for KC and highlighting the potential for refining clinical decision-making and early prevention strategies.

Tracking cryptic SARS-CoV-2 hospital outbreak through quasispecies analysis

BackgroundSince the beginning of the pandemic, contact tracing has been one of the most relevant issues to understand SARS-CoV-2 transmission dynamics and, in this context, the analysis of quasispecies may turn out to be a useful tool for outbreak investigations. Analysis of the intra-host single nucleotide variants (iSNVs) found in the nsp2, ORF3, and ORF7 genes of SARS-CoV-2 was conducted in order to correctly identify virus transmission chain among patients hospitalized in Brescia Civic Hospital.MethodsDuring the period between August and October 2023, 13 nasopharyngeal specimens, collected from patients admitted to Brescia Civic Hospital, were tested for SARS-CoV-2 positivity and molecularly characterized. Firstly, a phylogenetic analysis was performed to evaluate if they were epidemiologically linked and, then, the Beta-binomial method was used to estimate the transmission bottleneck size (Nb) and quantify the number of viral particles transmitted from one individual (donor) to another (recipient).ResultsAccording to the molecular characterization of specimens, we identified two transmission clusters in the cardiology unit: the first cluster concerned patients tested positive for the HV.1/EG.5.1.6 lineage, while the second cluster concerned patients tested positive for the FL.10.1 lineage. Moreover, evaluating the bottleneck size, we were able to solve SARS-CoV-2 transmission chain among infected patients.ConclusionOur method shows that it is possible to conduct a tracing study using a genomic approach based on iSNVs analysis.

Swedish Genome-Wide Haplotype Association Analysis Suggests Breast Cancer Loci with Varying Risk-Modifying Effects.

Background: To find support for risk-modifying genes in breast cancer, a haplotype GWAS in sporadic breast cancer cases was undertaken. The results were compared with the results from previous analyses in familial cases and all cases from the same Swedish cohort. Methods: In total, 2550 women with sporadic invasive breast cancer and 5021 healthy controls were included in a sliding-window haplotype GWAS using PLINK 1.07. Results: The analysis of sporadic cases confirmed the loci on chromosomes 10q26.13, 11q13.3, and 16q12.1 and suggested one novel locus on chromosome 12p11.21 (OR = 1.42 p = 4.55 × 10-8). A comparison between these loci and the same loci in the analyses of familial cases and all breast cancer cases was undertaken. Conclusions: Haplotype GWAS in sporadic cases of Swedish breast cancer cases supported known risk loci and suggested another risk locus. The loci identified in the analysis of sporadic and all breast cancer cases were suggested to act as modifiers of the risk of breast cancer. Haplotype analysis identified other loci with higher odds ratios than single-variant analysis. Further studies are needed to find out how to best include the findings in breast cancer prevention.

Genome-wide association study analysis of single nucleotide variants in L. infantum associated with IL-6 inflammatory response in visceral leishmaniasis.

Elevated levels of IL-6 in plasma are associated with the severity of visceral leishmaniasis (VL). The clinical manifestations of VL vary among patients, influenced by host factors and the virulence of the Leishmania infantum parasite. Considering that severe VL may result from an exaggerated inflammatory response, this study investigated whether IL-6 could serve as a biomarker to identify pro-inflammatory virulence factors. We conducted a genome-wide association study (GWAS) analysis on L. infantum isolates from patients with VL, whose IL-6 concentrations were measured. The analysis revealed that the relationship between IL-6 levels and clinical outcomes (survival vs mortality) had an area under the curve (AUC) of 0.67 (95% CI 0.52–0.81). A cut-off of 391.7 pg mL−1 for IL-6 was established to conduct a logistic regression analysis. We identified 10 029 single nucleotide variants (SNVs) across 62 genomes, resulting in 6,948 SNVs after filtering, of which 6,341 are located in protein-coding regions. The association analysis with PLINK identified 722 variants, of which 35 showed significant associations, with odds ratios ≥3.3, primarily in coding regions. These findings demonstrate that IL-6 levels tended to be associated with the fatal outcome of VL and highlight 35 novel genetic variants that could serve as potential biomarkers for prognosis. Further research into the biological role of these variants may lead to new therapeutic targets and improve the clinical management of VL, especially in identifying high-risk patients.

Integrated eQTL mapping approach reveals genomic regions regulating candidate genes of the E8-r3 locus in soybean.

Deciphering the gene regulatory networks of critical quantitative trait loci associated with early maturity provides information for breeders to unlock soybean's (Glycine max (L.) Merr.) northern potential and expand its cultivation range. The E8-r3 locus is a genomic region regulating the number of days to maturity under constant short-day photoperiodic conditions in two early-maturing soybean populations (QS15524F2:F3 and QS15544RIL) belonging to maturity groups MG00 and MG000. In this study, we developed a combinatorial expression quantitative trait loci mapping approach using three algorithms (ICIM, IM, and GCIM) to identify the regions that regulate three candidate genes of the E8-r3 locus (Glyma.04G167900/GmLHCA4a, Glyma.04G166300/GmPRR1a, and Glyma.04G159300/GmMDE04). Using this approach, a total of 2,218 trans (2,061 genes)/7 cis (7 genes) and 4,073 trans (2,842 genes)/3,083 cis (2,418 genes) interactions were mapped in the QS15524F2:F3 and QS15544RIL populations, respectively. From these interactions, we successfully identified two hotspots (F2_GM15:49,385,092-49,442,237 and F2_GM18:1,434,182-1,935,386) and three minor regions (RIL_GM04:17,227,512-20,251,662, RIL_GM04:31,408,946-31,525,671 and RIL_GM13:37,289,785-38,620,690) regulating the candidate genes of E8-r3 and several of their homologs. Based on co-expression network and single nucleotide variant analyses, we identified ALTERED PHLOEM DEVELOPMENT (Glyma.15G263700) and DOMAIN-CONTAINING PROTEIN 21 (Glyma.18G025600) as the best candidates for the F2_GM15:49,385,092-49,442,237 and F2_GM18:1,434,182-1,935,386 hotspots. These findings demonstrate that a few key regions are involved in the regulation of the E8-r3 candidates GmLHCA4a, GmPRR1a, and GmMDE04.

Comprehensive analysis of Mycobacterium tuberculosis genomes reveals genetic variations in bacterial virulence

Tuberculosis, a disease caused by Mycobacterium tuberculosis (Mtb), is a significant health problem worldwide. Here, we developed a method to detect large insertions and deletions (indels), which have been generally understudied. Leveraging this framework, we performed a comprehensive analysis of single nucleotide variants and small and large indels across 1,960 Mtb clinical isolates. Comparing the distribution of variants demonstrated that gene disruptive variants are underrepresented in genes essential for bacterial survival. An evolutionary analysis revealed that Mtb genomes are enriched in partially deleterious mutations. Genome-wide association studies identified small and large deletions in eccB2 significantly associated with patient prognosis. Additionally, we unveil significant associations with antibiotic resistance in 23 non-canonical genes. Among these, large indels are primarily found in genetic regions of Rv1216c, Rv1217c, fadD11, and ctpD. This study provides a comprehensive catalog of genetic variations and highlights their potential impact for the future treatment and risk prediction of tuberculosis.

Genome-wide association analysis of treatment resistant schizophrenia for variant discovery and polygenic assessment

BackgroundTreatment resistant schizophrenia (TRS) is broadly defined as inadequate response to adequate treatment and is associated with a substantial increase in disease burden. Clozapine is the only approved treatment for TRS, showing superior clinical effect on overall symptomatology compared to other drugs, and is the prototype of atypical antipsychotics. Risperidone, another atypical antipsychotic with a more distinctive dopamine 2 antagonism, is commonly used in treatment of schizophrenia. Here, we conducted a genome-wide association study on patients treated with clozapine (TRS) vs. risperidone (non-TRS) and investigated whether single variants and/or polygenic risk score for schizophrenia are associated with TRS status. We hypothesized that patients who are treated with clozapine and risperidone might exhibit distinct neurobiological phenotypes that match pharmacological profiles of these drugs and can be explained by genetic differences. The study population (n = 1286) was recruited from a routine therapeutic drug monitoring (TDM) service between 2005 and 2022. History of a detectable serum concentration of clozapine and risperidone (without TDM history of clozapine) defined the TRS (n = 478) and non-TRS (n = 808) group, respectively.ResultsWe identified a suggestive association between TRS and a common variant within the LINC00523 gene with a significance just below the genome-wide threshold (rs79229764 C > T, OR = 4.89; p = 1.8 × 10−7). Polygenic risk score for schizophrenia was significantly associated with TRS (OR = 1.4, p = 2.1 × 10−6). In a large post-mortem brain sample from schizophrenia donors (n = 214; CommonMind Consortium), gene expression analysis indicated that the rs79229764 variant allele might be involved in the regulation of GPR88 and PUDP, which plays a role in striatal neurotransmission and intellectual disability, respectively.ConclusionsWe report a suggestive genetic association at the rs79229764 locus with TRS and show that genetic liability for schizophrenia is positively associated with TRS. These results suggest a candidate locus for future follow-up studies to elucidate the molecular underpinnings of TRS. Our findings further demonstrate the value of both single variant and polygenic association analyses for TRS prediction.

Yield of genetic association signals from genomes, exomes and imputation in the UK Biobank

Whole-genome sequencing (WGS), whole-exome sequencing (WES) and array genotyping with imputation (IMP) are common strategies for assessing genetic variation and its association with medically relevant phenotypes. To date, there has been no systematic empirical assessment of the yield of these approaches when applied to hundreds of thousands of samples to enable the discovery of complex trait genetic signals. Using data for 100 complex traits from 149,195 individuals in the UK Biobank, we systematically compare the relative yield of these strategies in genetic association studies. We find that WGS and WES combined with arrays and imputation (WES + IMP) have the largest association yield. Although WGS results in an approximately fivefold increase in the total number of assayed variants over WES + IMP, the number of detected signals differed by only 1% for both single-variant and gene-based association analyses. Given that WES + IMP typically results in savings of lab and computational time and resources expended per sample, we evaluate the potential benefits of applying WES + IMP to larger samples. When we extend our WES + IMP analyses to 468,169 UK Biobank individuals, we observe an approximately fourfold increase in association signals with the threefold increase in sample size. We conclude that prioritizing WES + IMP and large sample sizes rather than contemporary short-read WGS alternatives will maximize the number of discoveries in genetic association studies.

Genomic Landscape of Osteosarcoma of Bone in an Older-Aged Patient Population and Analysis of Possible Etiologies Based on Molecular Signature.

Background: Osteosarcoma (OS), the most common primary malignant bone tumor, occurs mostly in the pediatric and adolescent (P/A) population where it has been subject to intense study whereas OS arising in the older-aged adult population has undergone less scrutiny. Materials and Methods: In this study, we assess the molecular aberrations detected in eight older adult patients (>59 years of age) with OS of bone by whole-exome sequencing (WES) on formalin-fixed, paraffin-embedded tissue and quantified the contributions of endogenous and exogenous mutational processes to tumor mutational burden and to tumorigenesis through computational analysis. Results: We identified 86 clinically significant somatic mutations. TP53 mutations occurred in OSs of three patients and one patient harbored a pathogenic germline mutation of TP53. Loss-of-heterozygosity of DNA-damage repair genes occurred in all six tumors evaluated. Computational analysis of single nucleotide variants within each tumor detected eight distinct mutagenic processes of which age-associated mutational processes, thiopurine chemotherapy, and defective homologous DNA recombination repair contributed the most to both tumor mutation burden and tumor pathogenesis. Conclusion: The genomic landscape of our older OS patients deciphered by WES is extremely diverse with only 15% of mutated somatic genes uncovered in our study previously described in P/A-enriched OS studies. Endogenous age-related mutagenic processes, defective DNA homologous recombination repair, and exogenous effects of chemotherapy are mainly responsible for pathogenic mutations in OS occurring in our cohort.

Innovative all-in-one exome sequencing strategy for diagnostic genetic testing in male infertility: Validation and 10-month experience.

Current guidelines indicate that patients with extreme oligozoospermia or azoospermia should be tested for chromosomal imbalances, azoospermia factor (AZF) deletions and/or CFTR variants. For other sperm abnormalities, no genetic diagnostics are recommended. To determine whether exome sequencing (ES) with combined copy number variant (CNV) and single nucleotide variant (SNV) analysis is a reliable first-tier method to replace current methods (validation study), and to evaluate the diagnostic yield after 10 months of implementation (evaluation study). In the validation study, ES was performed on DNA of patients already diagnosed with AZF deletions (n=17), (non-)mosaic sex chromosomal aneuploidies or structural chromosomal anomalies (n=37), CFTR variants (n=26), or variants in known infertility genes (n=4), and 90 controls. The data were analyzed using our standard diagnostic pipeline, with a bioinformatic filter for 130 male infertility genes. In the evaluation study, results of 292 clinical exomes were included. All previously reported variants in the validation cohort, including clinically relevant Y-chromosomal microdeletions, were correctly identified and reliably detected. In the evaluation study, we identified one or more clinically relevant genetic anomalies in 67 of 292 of all cases (22.9%): these included aberrations that could have been detected with current methods in 30 of 67 patients (10.2% of total), (possible) (mono)genetic causes in the male infertility gene panel in 28 of 67 patients (9.6%), and carriership of cystic fibrosis in nine of 67 patients (3.1%). ES is a reliable first-tier method to detect the most common genetic causes of male infertility and, as additional genetic causes can be detected, in our evaluation cohort the diagnostic yield almost doubled (10.2%-19.8%, excluding CF carriers). A genetic diagnosis provides answers on the cause of infertility and helps the professionals in the counseling for treatment, possible co-morbidities and risk for offspring and/or family members. Karyotyping will still remain necessary for detecting balanced translocations or low-grade chromosomal mosaicism.

Shared germline genomic variants in two patients with double primary gastrointestinal stromal tumours (GISTs)

BackgroundGastrointestinal stromal tumours (GISTs) are prevalent mesenchymal tumours of the gastrointestinal tract, commonly exhibiting structural variations in KIT and PDGFRA genes. While the mutational profiling of somatic tumours is well...

Whole exome sequencing analyses identified novel genes for Alzheimer's disease and related dementia.

The heritability of Alzheimer's disease (AD) is estimated to be 58%-79%. However, known genes can only partially explain the heritability. Here, we conducted gene-based exome-wide association study (ExWAS) of rare variants and single-variant ExWAS of common variants, utilizing data of 54,569 clinically diagnosed/proxy AD and related dementia (ADRD) and 295,421 controls from the UK Biobank. Gene-based ExWAS identified 11 genes predicting a higher ADRD risk, including five novel ones, namely FRMD8, DDX1, DNMT3L, MORC1, and TGM2, along with six previously reported ones, SORL1, GRN, PSEN1, ABCA7, GBA, and ADAM10. Single-variant ExWAS identified two ADRD-associated novel genes, SLCO1C1 and NDNF. The identified genes were predominantly enriched in amyloid-β process pathways, microglia, and brain regions like hippocampus. The druggability evidence suggests that DDX1, DNMT3L, TGM2, SLCO1C1, and NDNF could be effective drug targets. Our study contributes to the current body of evidence on the genetic etiology of ADRD. Gene-based analyses of rare variants identified five novel genes for Alzheimer's disease and related dementia (ADRD), including FRMD8, DDX1, DNMT3L, MORC1, and TGM2. Single-variant analyses of common variants identified two novel genes for ADRD, including SLCO1C1 and NDNF. The identified genes were predominantly enriched in amyloid-β process pathways, microglia, and brain regions like hippocampus. DDX1, DNMT3L, TGM2, SLCO1C1, and NDNF could be effective drug targets.

Whole genome sequencing based analysis of inflammation biomarkers in the Trans-Omics for Precision Medicine (TOPMed) consortium.

Inflammation biomarkers can provide valuable insight into the role of inflammatory processes in many diseases and conditions. Sequencing based analyses of such biomarkers can also serve as an exemplar of the genetic architecture of quantitative traits. To evaluate the biological insight, which can be provided by a multi-ancestry, whole-genome based association study, we performed a comprehensive analysis of 21 inflammation biomarkers from up to 38 465 individuals with whole-genome sequencing from the Trans-Omics for Precision Medicine (TOPMed) program (with varying sample size by trait, where the minimum sample size was n = 737 for MMP-1). We identified 22 distinct single-variant associations across 6 traits-E-selectin, intercellular adhesion molecule 1, interleukin-6, lipoprotein-associated phospholipase A2 activity and mass, and P-selectin-that remained significant after conditioning on previously identified associations for these inflammatory biomarkers. We further expanded upon known biomarker associations by pairing the single-variant analysis with a rare variant set-based analysis that further identified 19 significant rare variant set-based associations with 5 traits. These signals were distinct from both significant single variant association signals within TOPMed and genetic signals observed in prior studies, demonstrating the complementary value of performing both single and rare variant analyses when analyzing quantitative traits. We also confirm several previously reported signals from semi-quantitative proteomics platforms. Many of these signals demonstrate the extensive allelic heterogeneity and ancestry-differentiated variant-trait associations common for inflammation biomarkers, a characteristic we hypothesize will be increasingly observed with well-powered, large-scale analyses of complex traits.

Whole exome sequencing analysis identifies genes for alcohol consumption

Alcohol consumption is a heritable behavior seriously endangers human health. However, genetic studies on alcohol consumption primarily focuses on common variants, while insights from rare coding variants are lacking. Here we leverage whole exome sequencing data across 304,119 white British individuals from UK Biobank to identify protein-coding variants associated with alcohol consumption. Twenty-five variants are associated with alcohol consumption through single variant analysis and thirteen genes through gene-based analysis, ten of which have not been reported previously. Notably, the two unreported alcohol consumption-related genes GIGYF1 and ANKRD12 show enrichment in brain function-related pathways including glial cell differentiation and are strongly expressed in the cerebellum. Phenome-wide association analyses reveal that alcohol consumption-related genes are associated with brain white matter integrity and risk of digestive and neuropsychiatric diseases. In summary, this study enhances the comprehension of the genetic architecture of alcohol consumption and implies biological mechanisms underlying alcohol-related adverse outcomes.

Exome sequencing identifies novel genetic variants associated with varicose veins.

Varicose veins (VV) are one of the common human diseases, but the role of genetics in its development is not fully understood. We conducted an exome-wide association study of VV using whole-exome sequencing data from the UK Biobank, and focused on common and rare variants using single-variant association analysis and gene-level collapsing analysis. A total of 13,823,269 autosomal genetic variants were obtained after quality control. We identified 36 VV-related independent common variants mapping to 34 genes by single-variant analysis and three rare variant genes (PIEZO1, ECE1, FBLN7) by collapsing analysis, and most associations between genes and VV were replicated in FinnGen. PIEZO1 was the closest gene associated with VV (P = 5.05 × 10-31), and it was found to reach exome-wide significance in both single-variant and collapsing analyses. Two novel rare variant genes (ECE1 and METTL21A) associated with VV were identified, of which METTL21A was associated only with females. The pleiotropic effects of VV-related genes suggested that body size, inflammation, and pulmonary function are strongly associated with the development of VV. Our findings highlight the importance of causal genes for VV and provide new directions for treatment.

Whole-genome sequencing identifies variants in ANK1, LRRN1, HAS1, and other genes and regulatory regions for stroke in type 1 diabetes

Individuals with type 1 diabetes (T1D) carry a markedly increased risk of stroke, with distinct clinical and neuroimaging characteristics as compared to those without diabetes. Using whole-exome or whole-genome sequencing of 1,051 individuals with T1D, we aimed to find rare and low-frequency genomic variants associated with stroke in T1D. We analysed the genome comprehensively with single-variant analyses, gene aggregate analyses, and aggregate analyses on genomic windows, enhancers and promoters. In addition, we attempted replication in T1D using a genome-wide association study (N = 3,945) and direct genotyping (N = 3,263), and in the general population from the large-scale population-wide FinnGen project and UK Biobank summary statistics. We identified a rare missense variant on SREBF1 exome-wide significantly associated with stroke (rs114001633, p.Pro227Leu, p-value = 7.30 × 10–8), which replicated for hemorrhagic stroke in T1D. Using gene aggregate analysis, we identified exome-wide significant genes: ANK1 and LRRN1 displayed replication evidence in T1D, and LRRN1, HAS1 and UACA in the general population (UK Biobank). Furthermore, we performed sliding-window analyses and identified 14 genome-wide significant windows for stroke on 4q33-34.1, of which two replicated in T1D, and a suggestive genomic window on LINC01500, which replicated in T1D. Finally, we identified a suggestively stroke-associated TRPM2-AS promoter (p-value = 5.78 × 10–6) with borderline significant replication in T1D, which we validated with an in vitro cell-based assay. Due to the rarity of the identified genetic variants, future replication of the genomic regions represented here is required with sequencing of individuals with T1D. Nevertheless, we here report the first genome-wide analysis on stroke in individuals with diabetes.

OA08 GWAS meta-analysis of ankylosing spondylitis and Behçet's disease identifies novel MHC-I-opathy genes

Abstract Background/Aims Spondyloarthropathies (SpA) and Behçet's disease (BD) have clinical and HLA locus overlap and have been grouped under MHC-I-opathy [1]. Genetic overlap can help elucidate MHC-I-opathy pathogenesis. Ankylosing spondylitis (AS) is a primary SpA. Here, a cluster-based approach helps to identify overlapping loci between AS and BD of even modest significance. Methods Association clustering methods such as OASIS [2] reduce the multiple-testing burden and are therefore more powerful than single variant analysis for identifying modest genetic effects. Here, two large genome-wide association studies (GWAS) of AS (921 cases, 907 controls) and BD (1215 cases and 1278 healthy controls) from Turkiye, were subjected to OASIS meta-analyses to identify common non-HLA loci [3, 4]. Statistics used to identify significant loci included the novel OASIS locus index (OLI), defined as the product of maximum -logP at a locus with the ratio of actual to predicted number of significant SNPs. Results GWAS for both diseases had the highest significance at the HLA-locus. OASIS avoids the proportional signals problem, inherent in GWAS, by separately analysing highly significant loci (P &amp;lt; 10-8). Hence, of the 234 independent modestly significant non-HLA loci, there were 16 loci common to both AS and BD (Figure 1). These included known MHC-I-opathy loci, 1p31.3 for IL23R (P = 5.37 E-06, OLI = 52.7) and 13q14.11 for LACC1 (P = 7.41 E-06, OLI= 65.3) [1, 5]. Novel loci included 3p21.31 containing CCR9 (P = 2.46 E-05, OLI=25.8), 3q22.3 containing CLDN18 (P = 5.37 E-05, OLI=31.4) and 4q27-4q28.1 containing FGF2 (P = 1.1 E-04, OLI=29.5). Conclusion Five loci containing pathologically relevant genes for MHC-I-opathy were identified using a cluster-based approach in AS and BD GWAS datasets. These large meta-analyses will help identify pathogenic pathways for MHC-I-opathy, an entity that has so far largely remained elusive [6]. Disclosure M. Saeed: None.

Whole-genome sequencing in prenatally detected congenital malformations: prospective cohort study in clinical setting.

To investigate the diagnostic yield of trio whole-genome sequencing (WGS) in fetuses with various congenital malformations referred to a tertiary center for prenatal diagnosis. In this prospective study, 50 pregnancies with different congenital malformations, negative for trisomies and causative copy-number variants, were analyzed further with fetal-parental trio WGS analysis. Parents were eligible for inclusion if they accepted further investigation following the detection of isolated or multiple malformations on prenatal ultrasound. Cases with isolated increased nuchal translucency, gamete donation or multiple pregnancy were excluded. WGS with the Illumina Inc. 30× polymerase-chain-reaction-free short-read sequencing included analysis of single-nucleotide variants, insertions and deletions, structural variants, short tandem repeats and copy-number identification of SMN1 and SMN2 genes. A molecular diagnosis was achieved in 13/50 (26%) cases. Causative sequence variants were identified in 12 genes: FGFR3 (n = 2), ACTA1 (n = 1), CDH2 (n = 1), COL1A2 (n = 1), DHCR7 (n = 1), EYA1 (n = 1), FBXO11 (n = 1), FRAS1 (n = 1), L1CAM (n = 1), OFD1 (n = 1), PDHA1 (n = 1) and SOX9 (n = 1). The phenotypes of the cases were divided into different groups, with the following diagnostic yields: skeletal malformation (4/9 (44%)), multisystem malformation (3/7 (43%)), central nervous system malformation (5/15 (33%)) and thoracic malformation (1/10 (10%)). Additionally, two cases carried variants that were considered potentially clinically relevant, even though they were assessed as variants of uncertain significance, according to the guidelines provided by the American College of Medical Genetics and Genomics. Overall, we identified a causative or potentially clinically relevant variant in 15/50 (30%) cases. We demonstrate a diagnostic yield of 26% with clinical WGS in prenatally detected congenital malformations. This study emphasizes the benefits that WGS can bring to the diagnosis of fetal structural anomalies. It is important to note that causative chromosomal aberrations were excluded from our cohort before WGS. As chromosomal aberrations are a well-known cause of prenatally detected congenital malformations, future studies using WGS as a primary diagnostic test, including assessment of chromosomal aberrations, may show that the detection rate exceeds the diagnostic yield of this study. WGS can add clinically relevant information, explaining the underlying cause of the fetal anomaly, which will provide information concerning the specific prognosis of the condition, as well as estimate the risk of recurrence. A genetic diagnosis can also provide more reproductive choice for future pregnancies. © 2024 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of International Society of Ultrasound in Obstetrics and Gynecology.