Jan 2026· Human Mutation· Vol 2026· 0 citations· 46 references
Medicine
TL;DR
In a challenging, ES‐negative ARID cohort, GS provided an additional ~5.3% diagnostic yield by uncovering a noncoding splice alteration and a large intragenic deletion and highlight significant interpretation challenges in noncoding regions.
Abstract
Intellectual disability (ID) affects approximately 1%–3% of the population and spans diverse clinical presentations with marked genetic heterogeneity, especially in consanguineous populations where autosomal‐recessive ID is common. Despite advances in diagnostic methods, ~50% of individuals with ID remain without a molecular diagnosis. Genome sequencing (GS) can detect variant classes poorly captured by other methodologies, including deep intronic splice changes and structural variants. We performed short‐read GS on 38 Iranian autosomal recessive intellectual disability (ARID) families that remained unsolved after exome sequencing (ES) and two phases of reanalysis. Sequencing and variant calling were performed on DRAGEN Bio‐IT Platform with GRCh38 and variants were annotated in Golden helix Varseq software and the AnnotSV tool. GS yielded diagnoses in 2 of 38 families (5.3%), identifying a homozygous deep‐intronic ATP8A2 variant (c.1473 + 519C > T) that creates a cryptic donor site and a 57‐bp pseudoexon, and a homozygous ~103‐kb CNTNAP2 deletion removing the in‐frame Exon 2. Additionally, GS uncovered a homozygous NCOR1 missense variant that represents a novel candidate gene for ARID, but further studies are required to confirm the gene–disease association. The ATP8A2 and CNTNAP2 variants were uniquely detectable by GS. In conclusion, in a challenging, ES‐negative ARID cohort, GS provided an additional ~5.3% diagnostic yield by uncovering a noncoding splice alteration and a large intragenic deletion. These results underscore GS as a valuable, though still modest, tool over ES and highlight significant interpretation challenges in noncoding regions. Continued advances in functional assays and complementary long‐read technologies will be essential to further reduce the diagnostic gap in unresolved ID.
Background Muscular dystrophies (MDs) are a genetically heterogeneous group of disorders, posing significant diagnostic challenges, especially in populations with high consanguinity. Despite advances in genetic testing, a substantial proportion of patients remain undiagnosed. Whole‐exome sequencing (WES) has emerged as a powerful tool for identifying causal variants in such unresolved cases. To identify the genetic basis of nondystrophinopathic MDs in Iranian families with inconclusive prior genetic testing and to evaluate the diagnostic yield and mutational spectrum in this population. Methods We performed WES on one affected individual from each of 10 unrelated Iranian families with clinically diagnosed MD. Candidate variants were prioritized based on in silico prediction tools (SIFT, PolyPhen‐2, CADD, SpliceAI), population frequency databases (gnomAD, 1000 Genomes, EVS), and ACMG/AMP guidelines. Findings were validated by Sanger sequencing, MLPA, and STR haplotype analysis. Segregation analysis was performed in available family members. Results WES led to a diagnostic yield of 69.2% (9/13 variants in 10 families) after segregation analysis and ACMG‐based reclassification. We identified 13 candidate variants in 10 known MD‐associated genes, including DYSF, SGCA, TK2, MAP3K20, LMNA, COL6A1, COL6A2, ITGA7, MICU1, and SGCB. Among these, seven variants (54%) were novel. The majority of cases (84.6%) followed an autosomal recessive pattern, consistent with high parental consanguinity (70%). Notably, a de novo splice‐site variant in COL6A2 (c.1053+1G>T) was identified in a sporadic case, confirming an autosomal dominant inheritance. Challenges in variant interpretation were observed in families with variants in tightly linked genes (COL6A1 and COL6A2), highlighting the role of linkage disequilibrium in founder populations. Conclusion WES is a highly effective diagnostic strategy for genetically heterogeneous MDs, particularly in consanguineous populations. Our study expands the mutational spectrum of MDs in Iran and provides critical data for genetic counseling, prenatal diagnosis, and future therapeutic development. The high rate of novel variants underscores the importance of population‐specific genomic studies.
Nasibeh Soltani, Zahra Shahbazi, M. Fallah et al.· Human Mutation· 0 citations
Background Inherited ocular disorders are a leading cause of early-onset visual impairment, particularly in populations with high consanguinity such as Iran, where a substantial proportion of affected individuals remain without a molecular diagnosis after conventional evaluation. We aimed to determine the diagnostic yield and variant spectrum of whole-exome sequencing (WES) in Iranian probands with congenital or early-onset ocular disorders that were genetically unresolved by prior testing. Methods Thirty unrelated probands were recruited consecutively (July–September 2024). Genomic DNA underwent exome capture (Agilent SureSelect V7) and paired-end sequencing (Illumina NovaSeq 6000). Reads were cleaned with SOAPnuke, aligned with BWA-MEM, and variants were called with GATK HaplotypeCaller and DeepVariant, filtered by GATK VQSR, and annotated against public and Iranian-specific databases (GEMIRAN, IRANOM). Variants were classified per ACMG/AMP criteria. Diagnostic yield was defined as the proportion of probands with a causative or candidate variant concordant with phenotype. Results A causative or strong candidate variant concordant with the phenotype was identified in 21 of 30 probands (diagnostic yield 70.0%; 95% CI 50.6–85.3%). Pathogenic or likely-pathogenic variants were found in established genes including ABCA4, USH2A, RP1, CRB1, CEP290, GUCY2D, CYP1B1 and TYR. Among the identified genotypes, 54% were homozygous and 12% hemizygous (X-linked), consistent with consanguinity in 11/30 (36.7%) families; 35% were single heterozygous findings in autosomal-recessive genes, interpreted as incomplete genotypes pending detection of a second allele. Onset was infantile in 73% of probands. Conclusions WES is an effective first-tier test for congenital and early-onset ocular disorders in the Iranian population, resolving roughly 70% of previously undiagnosed probands. Single-allele findings in recessive genes indicate that complementary copy-number and structural-variant analysis, deep-intronic assessment, periodic reanalysis, and reflex whole-genome sequencing are needed to maximise yield and support accurate genetic counselling.
Ali Asadi, Seyed Ataollah Sadat Shandiz, Amirhossein Ebrahimi et al.· Molecular Genetics and Metab...· 0 citations
Whole-genome sequencing (WGS) projects for rare disease diagnosis typically yield a diagnostic rate of 25-41%, depending on the methods for patient selection and the extent of prior genetic testing. The Scottish Genomes Partnership (SGP) is a collaborative programme using genome sequencing to diagnose rare disease patients with presumed monogenic aetiology in the Scottish NHS. Within SGP, short-read sequencing (SRS) had previously achieved a diagnostic rate of 23% in affected families. To increase diagnostic yield, we applied Oxford Nanopore Technologies (ONT) long-read sequencing (LRS) to a cohort of 24 SGP families (74 individuals) that remained undiagnosed after SRS-based SNV/indel analysis. We also retrospectively reviewed SRS-derived structural variant (SV) calls to assess whether LRS results could have been detected in SRS data. After quality, rarity, panel-based and inheritance-based filtering, 392 candidate SVs were retained across de novo, homozygous recessive, compound heterozygous and X-linked inheritance models, of which 8 overlapped PanelApp diagnostic-grade "green" genes. Pathogenic or likely pathogenic de novo SVs were identified in 3 of 24 families: an AUTS2 inversion, a DLX5/6 locus inversion, and an FN1 deletion. All three SVs were independently confirmed by retrospective SRS re-analysis using SVRare. These findings demonstrate that LRS-based SV analysis, supported by orthogonal SRS re-analysis, can resolve clinically significant SVs in families who remain unsolved after standard rare disease testing.
P. Dutta, A. Pagnamenta, Christelle Robert et al.· European Journal of Human Ge...· 0 citations
As the first genetic study of hypertrophic cardiomyopathy in an Egyptian population, this work expands the global mutational spectrum, demonstrates the utility of genetic testing for risk stratification and personalised management, and underscores the need to diversify genomic datasets for equitable precision medicine.
Rana E Negm, R. Gabre, A. A. El‐Sherif et al.· Cardiology in the Young· 0 citations
Short-read sequencing (SRS)-based disease-targeted NGS gene panels have revolutionized rare disease diagnostics but often leave autosomal recessive cases unsolved when only one pathogenic allele is detected. Missing variants may reside in deep intronic regions or involve structural variants (SVs) undetectable by SRS. To improve diagnostic yield, we implemented a cost-effective target capture-based long-read sequencing (LRS) assay covering 56 genes and retrospectively analyzed 78 patients suspected of autosomal recessive disorders who remained undiagnosed after SRS. Functional validation using reverse transcription PCR (RT-PCR) and minigene assays was performed to further determine pathogenicity. Target capture-based LRS solved 25.6% (20/78) of cases by identifying 10 SVs, 3 deep intronic variants experimentally confirmed to cause aberrant splicing, and 7 cases in which haplotype phasing confirmed that variants were in trans with the known pathogenic variant, leading to reclassification of the VUS as likely pathogenic. This study demonstrates that target capture-based LRS effectively detects diverse types of variants missed by SRS. Integrating this assay into stepwise diagnostic workflows offers a practical and cost-effective strategy to enhance diagnostic yield in autosomal recessive diseases. However, because this cohort was retrospectively defined based on a prior single-allele detection by SRS, this 25.6% (20/78) diagnostic yield reflects performance within a highly enriched population and should not be directly extrapolated to unselected rare disease cohorts.
Jee-Soo Lee, K. Ryu, Hyesu Lee et al.· European Journal of Human Ge...· 0 citations
Advances in DNA sequencing technology are increasing the rate of molecular diagnosis for patients and families with inherited Mendelian diseases. However, some patients remain unsolved following standard-of-care testing (typically either exome or genome sequencing), with structural variants (SVs) likely to account for a significant proportion of these missed diagnoses. We re-analysed ten research cases with the visual disorder foveal hypoplasia (FH) that were previously unsolved through whole exome sequencing (WES). We used SVRare, a tool that integrates outputs from Canvas and Manta and combines these with allele frequencies and information regarding their genomic context to highlight plausible pathogenic SVs. This analysis solved 2/10 cases. Expanding this strategy, we then used SVRare in a reverse genetics approach to analyse an FH-relevant gene panel in the 100,000 Genomes Project (100KGP) rare disease cohort. This identified potentially pathogenic SVs as the cause of disease in a further 11 previously unsolved cases with phenotypes overlapping FH. In total, 11 SVs in five genes were identified as the likely cause of FH in 13 patients: SLC38A8 (1 patient), PAX6 (3), OCA2 (3), GPR143 (5) and CACNA1F (1). This analysis also identified multiple carriers of an apparently novel deletion in OCA2. However, further analysis suggested that this is, in fact, the relatively common founder variant responsible for oculocutaneous albinism (OCA) in African populations, first identified over 30 years ago. These findings show that SVRare effectively identifies deleterious SVs and illustrates the challenges in reporting SVs, including the importance of accurate and consistent reporting of such variants.
Mohammed A M Derar, Jing Yu, C. Watson et al.· European Journal of Human Ge...· 0 citations
