Evaluating use of changing technologies for rapid next-generation sequencing in pediatrics.

BACKGROUND: Rapid next-generation sequencing (NGS) offers the potential to shorten the diagnostic process and improve the care of acutely ill children. The goal of this study was to report our findings, including benefits and limitations, of a targeted NGS panel and rapid genome sequencing (rGS) in neonatal and pediatric acute clinical care settings. METHODS: Retrospective analysis of patient characteristics, diagnostic yields, turnaround time, and changes in management for infants and children receiving either RapSeq, a targeted NGS panel for 4500+ genes, or rGS, at the University of Utah Hospital and Primary Children's Hospital, from 2015 to 2020. RESULTS: Over a 5-year period, 142 probands underwent rapid NGS: 66 received RapSeq and 76 rGS. Overall diagnostic yield was 39%. In the majority of diagnostic cases, there were one or more changes in clinical care management. Of note, 7% of diagnoses identified by rGS would not have been identified by RapSeq. CONCLUSIONS: Our results indicate that rapid NGS impacts acute pediatric care in real-life clinical settings. Although affected by patient selection criteria, diagnostic yields were similar to those from clinical trial settings. Future studies are needed to determine relative advantages, including cost, turnaround time, and benefits for patients, of each approach in specific clinical circumstances. IMPACT: The use of comprehensive Mendelian gene panels and genome sequencing in the clinical setting allows for early diagnosis of patients in neonatal, pediatric, and cardiac intensive care units and impactful change in management. Diagnoses led to significant changes in management for several patients in lower acuity inpatient units supporting further exploration of the utility of rapid sequencing in these settings. This study reviews the limitations of comparing sequencing platforms in the clinical setting and the variables that should be considered in evaluating diagnostic rates across studies.

Rapid genome sequencing identifies a novel de novo SNAP25 variant for neonatal congenital myasthenic syndrome.

Congenital myasthenic syndrome (CMS) is a group of 32 disorders involving genetic dysfunction at the neuromuscular junction resulting in skeletal muscle weakness that worsens with physical activity. Precise diagnosis and molecular subtype identification are critical for treatment as medication for one subtype may exacerbate disease in another (Engel et al., Lancet Neurol 14: 420 [2015]; Finsterer, Orphanet J Rare Dis 14: 57 [2019]; Prior and Ghosh, J Child Neurol 36: 610 [2021]). The SNAP25-related CMS subtype (congenital myasthenic syndrome 18, CMS18; MIM #616330) is a rare disorder characterized by muscle fatigability, delayed psychomotor development, and ataxia. Herein, we performed rapid whole-genome sequencing (rWGS) on a critically ill newborn leading to the discovery of an unreported pathogenic de novo SNAP25 c.529C > T; p.Gln177Ter variant. In this report, we present a novel case of CMS18 with complex neonatal consequence. This discovery offers unique insight into the extent of phenotypic severity in CMS18, expands the reported SNAP25 variant phenotype, and paves a foundation for personalized management for CMS18.

Comprehensive variant calling from whole-genome sequencing identifies a complex inversion that disrupts ZFPM2 in familial congenital diaphragmatic hernia.

BACKGROUND: Genetic disorders contribute to significant morbidity and mortality in critically ill newborns. Despite advances in genome sequencing technologies, a majority of neonatal cases remain unsolved. Complex structural variants (SVs) often elude conventional genome sequencing variant calling pipelines and will explain a portion of these unsolved cases. METHODS: As part of the Utah NeoSeq project, we used a research-based, rapid whole-genome sequencing (WGS) protocol to investigate the genomic etiology for a newborn with a left-sided congenital diaphragmatic hernia (CDH) and cardiac malformations, whose mother also had a history of CDH and atrial septal defect. RESULTS: Using both a novel, alignment-free and traditional alignment-based variant callers, we identified a maternally inherited complex SV on chromosome 8, consisting of an inversion flanked by deletions. This complex inversion, further confirmed using orthogonal molecular techniques, disrupts the ZFPM2 gene, which is associated with both CDH and various congenital heart defects. CONCLUSIONS: Our results demonstrate that complex structural events, which often are unidentifiable or not reported by clinically validated testing procedures, can be discovered and accurately characterized with conventional, short-read sequencing and underscore the utility of WGS as a first-line diagnostic tool.