Augmenting Diagnostic Yield From Genomic Sequencing.

NHS genetics centres in Scotland sought to investigate the Genomics England 100,000 Genomes Project diagnostic utility to evaluate genome sequencing for in rare, inherited conditions. Four regional services recruited 999 individuals from 394 families in 200 rare phenotype categories, with negative historic genetic testing. Genome sequencing was performed at Edinburgh Genomics, and phenotype and sequence data were transferred to Genomics England for variant calling, gene-based filtering and variant prioritisation. NHS Scotland genetics laboratories performed interpretation, validation and reporting. New diagnoses were made in 23% cases – 19% in genes implicated in disease at the time of variant prioritisation, and 4% from later review of additional genes. Diagnostic yield varied considerably between phenotype categories and was minimal in cases with prior exome testing. Genome sequencing with gene panel filtering and reporting achieved improved diagnostic yield over previous historic testing but not over now routine trio-exome sequence tests. Re-interpretation of genomic data with updated gene panels modestly improved diagnostic yield at minimal cost. However, to justify the additional costs of genome vs exome sequencing, efficient methods for analysis of structural variation will be required and / or cost of genome analysis and storage will need to decrease.

Systematic evaluation of genome sequencing for the diagnostic assessment of autism spectrum disorder and fetal structural anomalies

The aim of this study is to evaluate the diagnostic value of genome sequencing in children with epilepsy, and to provide genome sequencing-based insights into the molecular genetic mechanisms of epilepsy to help establish accurate diagnoses, design appropriate treatments and assist in genetic counselling.We performed genome sequencing on 320 Chinese children with epilepsy, and interpreted single-nucleotide variants and copy number variants of all samples. The complete pedigree and clinical data of the probands were established and followed up. The clinical phenotypes, treatments, prognoses and genotypes of the patients were analysed.Age at seizure onset ranged from 1 day to 17 years, with a median of 4.3 years. Pathogenic/likely pathogenic variants were found in 117 of the 320 children (36.6%), of whom 93 (29.1%) had single-nucleotide variants, 22 (6.9%) had copy number variants and two had both single-nucleotide variants and copy number variants. Single-nucleotide variants were most frequently found in SCN1A (10/95, 10.5%), which is associated with Dravet syndrome, followed by PRRT2 (8/95, 8.4%), which is associated with benign familial infantile epilepsy, and TSC2 (7/95, 7.4%), which is associated with tuberous sclerosis. Among the copy number variants, there were three with a length <25 kilobases. The most common recurrent copy number variants were 17p13.3 deletions (5/24, 20.8%), 16p11.2 deletions (4/24, 16.7%), and 7q11.23 duplications (2/24, 8.3%), which are associated with epilepsy, developmental retardation and congenital abnormalities. Four particular 16p11.2 deletions and two 15q11.2 deletions were considered to be susceptibility factors contributing to neurodevelopmental disorders associated with epilepsy.The diagnostic yield was 75.0% in patients with seizure onset during the first postnatal month, and gradually decreased in patients with seizure onset at a later age. Forty-two patients (13.1%) were found to be specifically treatable for the underlying genetic cause identified by genome sequencing. Three of them received corresponding targeted therapies and demonstrated favourable prognoses.Genome sequencing provides complete genetic diagnosis, thus enabling individualized treatment and genetic counselling for the parents of the patients. Genome sequencing is expected to become the first choice of methods for genetic testing of patients with epilepsy.

Genome sequencing increases diagnostic yield in clinically diagnosed Alagille syndrome patients with previously negative test results

The content of the 13th Mutation Detection meeting (Leiden, April 2015) is summarized in this report. Topics discussed at the meeting included current challenges of clinical NGS, advances in bioinformatics, data quality control, single cell analysis and RNA sequencing, among others. Social, ethical and regulatory challenges of genomic data handling and data sharing were the focus of an expert panel debate. The 14th International Symposium on Variants in the Genome will take place in Santiago de Compostela, June 5-8, 2017. http://isv.variome.org.

Introduction: Intellectual Disability (ID) is the most common cause of referral to pediatric genetic centers, as it affects around 1–3% of the general population and is characterized by a wide genetic heterogeneity. The Genome Sequencing (GS) approach is expected to achieve a higher diagnostic yield than exome sequencing given its wider and more homogenous coverage, and, since theoretically, it can more accurately detect variations in regions traditionally not well captured and identify structural variants, or intergenic/deep intronic putatively pathological events. The decreasing cost of sequencing, the progress in data-management and bioinformatics, prompted us to assess GS efficiency as the first line procedure to identify the molecular diagnosis in patients without obvious ID etiology. This work is being carried out in the framework of the national French initiative for genomic medicine (Plan France Médecine Génomique 2025). Methods and Analysis: This multidisciplinary, prospective diagnostic study will compare the diagnostic yield of GS trio analysis (index case, father, mother) with the French core minimal reference strategy (Fragile-X testing, chromosomal microarray analysis and Gene Panel Strategy of 44 selected ID genes). Both strategies are applied in a blinded fashion, in parallel, in the same population of 1275 ID index cases with no obvious diagnosis (50% not previously investigated). Among them, a subgroup of 196 patients are randomized to undergo GS proband analysis in addition to GS trio analysis plus the French core minimal reference strategy, in order to compare their efficiency. The study also aims to identify the most appropriate strategy according to the clinical presentation of the patients, to evaluate the impact of deployment of GS on the families’ diagnostic odyssey and the modification of their care, and to identify the advantages/difficulties for the patients and their families. Ethics Statement: The protocol was approved by the Ethics Committee Sud Méditerranée I and the French data privacy commission (CNIL, authorization 919361). Trial Registration: ClinicalTrials.gov identifier NCT04154891 (07/11/2019).

Genetic testing strategies used to determine the etiology of congenital heart disease/defects (CHD/CHDs) vary between and within institutions, leading to potentially missed diagnostic opportunities. There has been little investigation comparing the diagnostic utility of gene panels among more comprehensive strategies used in the genetic evaluation of patients with CHD. In this descriptive study, we investigated the diagnostic yields of different genetic testing strategies in a real-world cohort of 263 patients with CHDs with genetic diagnoses. We counterfactually determined the diagnostic yield of a virtual gene panel designed for this study. We compared the diagnostic yield of the gene panel to other testing strategies, including chromosomal microarray (CMA), CMA + the gene panel, and genome sequencing. We assessed diagnostic yield differences according to clinical presentations to determine if phenotypes can inform optimal testing strategies. The virtual gene panel would have identified 51.3% of genetic disorders in this cohort, and 25.9% of genetic disorders would have remained undetected; another 22.8% may have needed additional testing to fully characterize the diagnoses. A combined approach of the virtual gene panel and CMA increased the diagnostic yield compared with panel-only testing or CMA alone (87.8% vs. 51.3% and 63.1%, respectively). The gene panel plus CMA would have increased the diagnostic yield by 24%-35% compared with CMA or panel testing alone in patients with extracardiac anomalies, 19%-41% in syndromic patients, and 0%-70% across CHD classifications. This combined approach also eliminated the potential need for follow-up testing; however, genome sequencing had a higher diagnostic yield across all clinical presentations (99.6%). CHD gene panels and CMA used individually or in combination are suboptimal first-line testing strategies, missing up to 36.5% of genetic disorders in our sample. Given the wide spectrum of phenotypes and genetic etiologies, our results support consideration of standardized genome sequencing for patients with CHDs.

IntroductionInborn errors of immunity (IEIs) constitute a diverse group of more than 500 disorders resulting from pathogenic variants in over 500 causative genes, with most being monogenic diseases. The use of exome sequencing based on next-generation sequencing technologies has significantly advanced the discovery of causative variants underlying IEIs and has achieved diagnostic yields of up to 40%. Despite these advances, a substantial proportion of patients still remain genetically undiagnosed due to limitations in detecting deep intronic or structural variants. Accordingly, we applied whole-genome sequencing to a cohort of patients suspected of IEIs in order to evaluate its diagnostic yield and capacity to identify novel structural genomic alterations.MethodsWe analyzed data from 25 probands presenting with suspected IEIs based on clinical features, who were enrolled through the National Bio-Big Data Program's whole genome sequencing (WGS) project at Samsung Medical Center, spanning July 2020 to February 2022. The study utilized a stepwise analytical protocol involving initial candidate gene panel analysis for detecting small variants, subsequent investigation of structural variants, and then a genotype-driven approach utilizing in-house bioinformatics pipelines. All identified variants were assessed for pathogenicity in accordance with the 2015 ACMG/AMP guidelines for the interpretation of sequence variants.ResultsCausative variants were detected in 10 (40%) probands using candidate gene panel analysis, which included BTK, CYBB, DKC1, DNAH11, DNAH5, IL2RG, NFKB2, PIK3CD and SH2D1A. Genotype-driven analysis identified pathogenic variants in two (8%) probands involving NF1 and PTPN11, while an additional five (20%) probands were found to have structural variants, including BTK, LRBA and SH2D1A. In total, genetic analysis revealed causative variants in 60% of patients. Variants of uncertain significance were identified in four cases among three probands (12%).ConclusionWGS facilitated the robust identification of causative genetic variants, including complex structural changes. These results suggest that employing WGS in patients suspected of IEIs could provide additional diagnostic yield. Supplementary InformationThe online version contains supplementary material available at 10.1007/s10875-025-01947-2.

Achieving a definitive genetic diagnosis of unexplained multiple congenital anomalies (MCAs) in neonatal intensive care units (NICUs) infants is challenging because of the limited diagnostic capabilities of conventional genetic tests. Although the implementation of whole genome sequencing (WGS) has commenced for diagnosing MCAs, due to constraints in resources and faculty, many NICUs continue to utilize chromosomal microarray (CMA) and/or karyotyping as the initial diagnostic approach. We aimed to evaluate the diagnostic efficacy of WGS in infants with MCAs who have received negative results from karyotyping and/or CMA. In this prospective study, we enrolled 80 infants with MCAs who were admitted to a NICU at a single center and had received negative results from CMA and/or karyotyping. The phenotypic characteristics were classified according to the International Classification of Diseases and the Human Phenotype Ontology. We assessed the diagnostic yield of trio-WGS in infants with normal chromosomal result and explored the process of diagnosing by analyzing both phenotype and genotype. Also, we compared the phenotype and clinical outcomes between the groups diagnosed with WGS and the undiagnosed group. The diagnostic yield of WGS was 26% (21/80), of which 76% were novel variants. There was a higher diagnostic yield in cases of craniofacial abnormalities, including those of the eye and ear, and a lower diagnostic yield in cases of gastrointestinal and genitourinary abnormalities. In addition, higher rates of rehabilitation therapy and gastrostomy were observed in WGS-diagnosed infants than in undiagnosed infants. This prospective cohort study assessed the usefulness of trio-WGS following chromosomal analysis for diagnosing MCAs in the NICU and revealed improvements in the diagnostic yield and clinical utility of WGS.

Most families with heritable neuromuscular disorders do not receive a molecular diagnosis. Here we evaluate diagnostic utility of exome, genome, RNA sequencing, and protein studies and provide evidence-based recommendations for their integration into practice. In total, 247 families with suspected monogenic neuromuscular disorders who remained without a genetic diagnosis after standard diagnostic investigations underwent research-led massively parallel sequencing: neuromuscular disorder gene panel, exome, genome, and/or RNA sequencing to identify causal variants. Protein and RNA studies were also deployed when required. Integration of exome sequencing and auxiliary genome, RNA and/or protein studies identified causal or likely causal variants in 62% (152 out of 247) of families. Exome sequencing alone informed 55% (83 out of 152) of diagnoses, with remaining diagnoses (45%; 69 out of 152) requiring genome sequencing, RNA and/or protein studies to identify variants and/or support pathogenicity. Arrestingly, novel disease genes accounted for <4% (6 out of 152) of diagnoses while 36.2% of solved families (55 out of 152) harbored at least one splice-altering or structural variant in a known neuromuscular disorder gene. We posit that contemporary neuromuscular disorder gene-panel sequencing could likely provide 66% (100 out of 152) of our diagnoses today. Our results emphasize thorough clinical phenotyping to enable deep scrutiny of all rare genetic variation in phenotypically consistent genes. Post-exome auxiliary investigations extended our diagnostic yield by 81% overall (34-62%). We present a diagnostic algorithm that details deployment of genomic and auxiliary investigations to obtain these diagnoses today most effectively. We hope this provides a practical guide for clinicians as they gain greater access to clinical genome and transcriptome sequencing.

PurposeIndividuals with intellectual disability (ID) and/or neurodevelopment disorders (NDDs) are currently investigated with several different approaches in clinical genetic diagnostics. MethodsWe compared the results from 3 diagnostic pipelines in patients with ID/NDD: genome sequencing (GS) first (N = 100), GS as a secondary test (N = 129), or chromosomal microarray (CMA) with or without FMR1 analysis (N = 421). ResultsThe diagnostic yield was 35% (GS-first), 26% (GS as a secondary test), and 11% (CMA/FMR1). Notably, the age of diagnosis was delayed by 1 year when GS was performed as a secondary test and the cost per diagnosed individual was 36% lower with GS first than with CMA/FMR1. Furthermore, 91% of those with a negative result after CMA/FMR1 analysis (338 individuals) have not yet been referred for additional genetic testing and remain undiagnosed. ConclusionOur findings strongly suggest that genome analysis outperforms other testing strategies and should replace traditional CMA and FMR1 analysis as a first-line genetic test in individuals with ID/NDD. GS is a sensitive, time- and cost-effective method that results in a confirmed molecular diagnosis in 35% of all referred patients.

Importance:Epilepsy is the most common neurological disorder of childhood. Identifying genetic diagnoses underlying epilepsy is critical to developing effective therapies and improving outcomes. Most children with non-acquired (unexplained) epilepsy remain genetically unsolved, and the utility of genome sequencing after nondiagnostic exome sequencing is unknown.Objective:To determine the diagnostic (primary) and clinical (secondary) utility of genome sequencing after nondiagnostic exome sequencing in individuals with unexplained pediatric epilepsy.Design:This cohort study performed genome sequencing and comprehensive analyses for 125 participants and available biological parents enrolled from August 2018 to May 2023, with data analysis through April 2024 and clinical return of diagnostic and likely diagnostic genetic findings. Clinical utility was evaluated.Setting:Pediatric referral centerParticipants:Participants with unexplained pediatric epilepsy and previous nondiagnostic exome sequencing; biological parents when availableExposure(s):Short-read genome sequencing and analysisMain Outcome(s) and Measure(s):Primary outcome measures were the diagnostic yield of genome sequencing, defined as the percentage of participants receiving a diagnostic or likely diagnostic genetic finding, and the unique diagnostic yield of genome sequencing, defined as the percentage of participants receiving a diagnostic or likely diagnostic genetic finding that required genome sequencing. The secondary outcome measure was clinical utility of genome sequencing, defined as impact on evaluation, treatment, or prognosis for the participant or their family.Results:125 participants (58 [46%] female) were enrolled with median age at seizure onset 3 [IQR 1.25, 8] years, including 44 (35%) with developmental and epileptic encephalopathies. The diagnostic yield of genome sequencing was 7.2% (9/125), with diagnostic genetic findings in five cases and likely diagnostic genetic findings in four cases. Among the solved cases, 7/9 (78%) required genome sequencing for variant detection (small copy number variant, three noncoding variants, and three difficult to sequence small coding variants), for a unique diagnostic yield of genome sequencing of 5.6% (7/125). Clinical utility was documented for 4/9 solved cases (44%).Conclusions and Relevance:These findings suggest that genome sequencing can have diagnostic and clinical utility after nondiagnostic exome sequencing and should be considered for patients with unexplained pediatric epilepsy.

BACKGROUND The landscape and clinical utility of comprehensive genomic investigations for a wide range of pediatric gastrointestinal (GI) disorders have not been fully characterized in the Middle East. AIM To characterize the diagnostic yield and clinical utility of genomic investigations in a Middle Eastern pediatric cohort of GI disorders, and to dissect the pathogenic landscape of those disorders in this region. METHODS Sixty-nine pediatric patients of diverse Arab and Asian origins, were clinically and genetically assessed for a spectrum of GI diseases, including liver disease, inflammatory bowel disease, chronic diarrhea, and pancreatitis. Clinical genomic investigations included mainly (87%) next generation sequencing-based gene panels and whole exome or genome sequencing. Clinical information, including demographics, symptoms, management and clinical outcomes, was extracted from medical records. RESULTS The overall positive yield was 55%, whereas multiple molecular diagnoses were made in 3 patients (4%) including 2 with triple genetic findings, highlighting the utility of genetic investigations in delineating the phenotypic complexity in this cohort. A secondary medically actionable finding (MYBPC3 -associated cardiomyopathy) was identified in one out of 12 patients (8%) who received exome or genome sequencing. Among all disease groups, the diagnostic yield was highest in patients with chronic diarrhea (73.3%) followed by those with cholestasis (62.5%). Copy number variants contributed substantially (18%) to the pathogenic variation spectrum. Consistent with consanguinity rates in this region, autosomal recessive conditions accounted for 66% of all diagnosed patients. Importantly, genetic findings guided clinical management plans and interventions in most cases (97%). Finally, we highlight a putative candidate gene, NR1I3 , possibly associated with cholestasis identified in an undiagnosed Yemeni family with episodic transient disease. CONCLUSION Our study provides new insights into the pathogenic variation landscape in pediatric GI disorders in the Middle East and emphasizes the clinical utility of genomic investigations in managing those patients.

267 Multiple fetal anomalies are associated with actionable postnatal genome sequencing results

Genomic sequencing is a powerful diagnostic tool in critically ill infants, but performing exome or genome sequencing (ES/GS) in the context of a research study is different from implementing these tests clinically. We investigated the integration of rapid ES into routine clinical care after a pilot research study in a Level IV Neonatal Intensive Care Unit (NICU). We performed a retrospective cohort analysis of infants admitted with suspected genetic disorders to the NICU from December 1, 2018 to March 31, 2021 and compared results to those obtained from a previous research study cohort (March 1, 2017 to November 30, 2018). Clinical rapid ES was performed in 80/230 infants (35%) with a suspected genetic disorder and identified a genetic diagnosis in 22/80 infants (28%). The majority of diagnoses acutely impacted clinical management (14/22 (64%)). Compared to the previous research study, clinically integrated rapid ES had a significantly lower diagnostic yield and increased time from NICU admission and genetics consult to ES report, but identified four genetic diagnoses that may have been missed by the research study selection criteria. Compared to other genetic tests, rapid ES had similar or higher diagnostic yield and similar or decreased time to result. Overall, rapid ES was utilized in the NICU after the pilot research study, often as the first-tier sequencing test, and could identify the majority of disease-causing variants, shorten the diagnostic odyssey, and impact clinical care. Based on our experience, we have identified strategies to optimize the clinical implementation of rapid ES in the NICU.