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.

Over the past decade, there has been tremendous progress in understanding brain somatic mosaicism in epilepsy in the research setting. Access to resected brain tissue samples from patients with medically refractory epilepsy undergoing epilepsy surgery has been key to making these discoveries. In this review, we discuss the gap between making discoveries in the research setting and bringing results back to the clinical setting. Current clinical genetic testing mainly uses clinically accessible tissue samples, like blood and saliva, and can detect inherited and de novo germline variants and potentially non-brain-limited mosaic variants that have resulted from post-zygotic mutation (also called "somatic mutations"). Methods developed in the research setting to detect brain-limited mosaic variants using brain tissue samples need to be further translated and validated in the clinical setting, which will allow post-resection brain tissue genetic diagnoses. However, obtaining a genetic diagnosis after surgery for refractory focal epilepsy, when brain tissue samples are available, is arguably "too late" to guide precision management. Emerging methods using cerebrospinal fluid (CSF) and stereoelectroencephalography (SEEG) electrodes hold promise for establishing genetic diagnoses pre-resection without the need for actual brain tissue. In parallel, development of curation rules for interpreting the pathogenicity of mosaic variants, which have unique considerations compared to germline variants, will assist clinically accredited laboratories and epilepsy geneticists in making genetic diagnoses. Returning results of brain-limited mosaic variants to patients and their families will end their diagnostic odyssey and advance epilepsy precision management.

Autism spectrum disorder (ASD) is a genetically heterogenous neurodevelopmental disorder. In the early years of next-generation sequencing, de novo germline variants were shown to contribute to ASD risk. These germline mutations are present in all of the cells of an affected individual and can be detected in any tissue, including clinically accessible DNA sources such as blood or saliva. In recent years, studies have also implicated de novo somatic variants in ASD risk. These somatic mutations arise postzygotically and are present in only a subset of the cells of an affected individual. Depending on the developmental time and progenitor cell in which a somatic mutation occurs, it may be detectable in some tissues and not in others. Somatic mutations detectable at relatively low sequencing coverage in clinically accessible tissues are suggested to contribute to 3–5% of simplex ASD diagnoses, and “brain limited” somatic mutations have been identified in postmortem ASD brain tissue. Somatic mutations likely represent the genetic diagnosis in a proportion of otherwise unexplained individuals with ASD, and brain limited somatic mutations can be used as markers to discover risk genes, cell types, brain regions, and cellular pathways important for ASD pathogenesis and to potentially target for therapeutics.

Although somatic mutations have well-established roles in cancer and certain focal epilepsies, the extent to which mutational mosaicism shapes the developing human brain is poorly understood. Here we characterize the landscape of somatic mutations in the human brain using ultra-deep (~250×) whole-genome sequencing of brains from 59 autism spectrum disorder (ASD) cases and 15 controls. We observe a mean of 26 (±10, range 10-60) somatic single nucleotide variants (sSNVs) per brain present in ≥4% of cells, with enrichment of mutations in coding and putative regulatory regions. Our analysis reveals that the first cell division after fertilization produces ~3.4 mutations, followed by 2-3 mutations in subsequent generations. This rate suggests that a typical individual possesses ~80 sSNVs present in ≥2% of cells—comparable to the number of de novo germline mutations per generation—with about half of individuals having at least one potentially function-altering somatic mutation somewhere in the cortex. Although limited by sample size, ASD brains show an excess of somatic mutations in neural enhancer sequences compared to controls, suggesting that mosaic enhancer mutations may contribute to ASD risk.