BACKGROUND Whole-exome sequencing is a diagnostic approach for the identification of molecular defects in patients with suspected genetic disorders. METHODS We developed technical, bioinformatic, interpretive, and validation pipelines for whole-exome sequencing in a certified clinical laboratory to identify sequence variants underlying disease phenotypes in patients. RESULTS We present data on the first 250 probands for whom referring physicians ordered whole-exome sequencing. Patients presented with a range of phenotypes suggesting potential genetic causes. Approximately 80% were children with neurologic pheno-types. Insurance coverage was similar to that for established genetic tests. We identified 86 mutated alleles that were highly likely to be causative in 62 of the 250 patients, achieving a 25% molecular diagnostic rate (95% confidence interval, 20 to 31). Among the 62 patients, 33 had autosomal dominant disease, 16 had auto-somal recessive disease, and 9 had X-linked disease. A total of 4 probands received two nonoverlapping molecular diagnoses, which potentially challenged the clinical diagnosis that had been made on the basis of history and physical examination. A total of 83% of the autosomal dominant mutant alleles and 40% of the X-linked mutant alleles occurred de novo. Recurrent clinical phenotypes occurred in patients with mutations that were highly likely to be causative in the same genes and in different genes responsible for genetically heterogeneous disorders. CONCLUSIONS Whole-exome sequencing identified the underlying genetic defect in 25% of consecutive patients referred for evaluation of a possible genetic condition. (Funded by the National Human Genome Research Institute.)

IMPORTANCE Clinical whole-exome sequencing is increasingly used for diagnostic evaluation of patients with suspected genetic disorders. OBJECTIVE To perform clinical whole-exome sequencing and report (1) the rate of molecular diagnosis among phenotypic groups, (2) the spectrum of genetic alterations contributing to disease, and (3) the prevalence of medically actionable incidental findings such as FBN1 mutations causing Marfan syndrome. DESIGN, SETTING, AND PATIENTS Observational study of 2000 consecutive patients with clinical whole-exome sequencing analyzed between June 2012 and August 2014. Whole-exome sequencing tests were performed at a clinical genetics laboratory in the United States. Results were reported by clinical molecular geneticists certified by the American Board of Medical Genetics and Genomics. Tests were ordered by the patient’s physician. The patients were primarily pediatric (1756 [88%]; mean age, 6 years; 888 females [44%], 1101 males [55%], and 11 fetuses [1% gender unknown]), demonstrating diverse clinical manifestations most often including nervous system dysfunction such as developmental delay. MAIN OUTCOMES AND MEASURES Whole-exome sequencing diagnosis rate overall and by phenotypic category, mode of inheritance, spectrum of genetic events, and reporting of incidental findings. RESULTS A molecular diagnosis was reported for 504 patients (25.2%) with 58% of the diagnostic mutations not previously reported. Molecular diagnosis rates for each phenotypic category were 143/526 (27.2%; 95% CI, 23.5%–31.2%) for the neurological group, 282/1147 (24.6%; 95% CI, 22.1%–27.2%) for the neurological plus other organ systems group, 30/83 (36.1%; 95% CI, 26.1%–47.5%) for the specific neurological group, and 49/244 (20.1%; 95% CI, 15.6%–25.8%) for the nonneurological group. The Mendelian disease patterns of the 527 molecular diagnoses included 280 (53.1%) autosomal dominant, 181 (34.3%) autosomal recessive (including 5 with uniparental disomy), 65 (12.3%) X-linked, and 1 (0.2%) mitochondrial. Of 504 patients with a molecular diagnosis, 23 (4.6%) had blended phenotypes resulting from 2 single gene defects. About 30% of the positive cases harbored mutations in disease genes reported since 2011. There were 95 medically actionable incidental findings in genes unrelated to the phenotype but with immediate implications for management in 92 patients (4.6%), including 59 patients (3%) with mutations in genes recommended for reporting by the American College of Medical Genetics and Genomics. CONCLUSIONS AND RELEVANCE Whole-exome sequencing provided a potential molecular diagnosis for 25% of a large cohort of patients referred for evaluation of suspected genetic conditions, including detection of rare genetic events and new mutations contributing to disease. The yield of whole-exome sequencing may offer advantages over traditional molecular diagnostic approaches in certain patients.

involves karyotyping, whereas in the Netherlands, patients who undergo amniocentesis have a more limited assessment only for trisomies 13, 18, and 21 and the sex chromosomes; thus, the tradeoff is a bit different.In addition, cfDNA is provided as a screening test for trisomies 13, 18, and 21, not just for Down syndrome. However, the performance characteristics of cfDNA for trisomies 13 and 18 are not as favorable as for T21, with a higher rate of falsenegative and false-positive results. In addition, a percentage of patients-somewhere between 1.5% and 8%-fail to obtain a result, usually because of insufficient fetal DNA. Such "low fetal fraction" is associated with obesity, which is a significant problem affecting a high percentage of reproductive-aged women in the United States. It is estimated that 20% to 50% of cfDNA tests fail to provide adequate fetal DNA in obese women. In addition, low fetal fraction is also associated with aneuploidy; therefore, women with cfDNA test failure should be considered high risk and offered follow-up with diagnostic testing (as well as a second attempt at cfDNA). When these potential outcomes are all considered, the performance characteristics of cfDNA versus traditional screening are not as clearly superior.Like the authors of this abstracted paper, several other authors and experts (Prenat Diagn 2013;33 (7):636-642) have suggested a contingent approach, using multiple marker screening as an initial screening tool, and then offering NIPT to intermediate-risk patients and either cfDNA or invasive testing to the highest-risk patients. Before completely changing the current standard of care, we need to understand this tradeoff. These authors consider comparative costs, but do not really provide incremental cost-effectiveness ratios, which are the best way to compare these strategies. While cfDNA is a better test if we are looking at a very precise test for a single disorder, only in patients in whom the test is successful at providing a result current screening may be preferable if we are looking to screen the entire population for a broad range of birth defects. Cost utility analyses, conducted by independent investigators and considering all important outcomes, are clearly needed before our approach completely changes. -MEN)

Importance: While congenital malformations and genetic diseases are a leading cause of early infant death, the contribution of single-gene disorders in this group is undetermined. Objective: To determine the diagnostic yield and utility of clinical exome sequencing in critically ill infants. Design, setting, participants: Clinical exome sequencing was performed on 278 unrelated infants within the first 100 days of life, admitted to Texas Children’s Hospital in Houston, over a period of five years, between December 2011 and January 2017. Exome sequencing types included proband exome, trio exome, and critical trio exome, a rapid genomic assay for seriously-ill infants. Main outcomes and measures: Indications for testing, diagnostic yield of clinical exome sequencing, turnaround time, molecular findings, patient age at diagnosis, and impact on medical management in a group of critically ill infants suspected to have genetic disorders. Results: Clinical indications for exome sequencing included a wide range of medical concerns. Overall, molecular diagnosis was achieved in 102/278 infants by clinical exome sequencing with a diagnostic yield of 36.7%. The diagnosis affected medical management in 53/102 (52.0%) of infants, with substantial impact on informed redirection of care, initiation of new subspecialist care, medication/dietary modifications, and furthering life-saving procedures in select patients. Critical trio exome revealed a molecular diagnosis in 32/63 infants (50.8%) at 33.1±5.6 days of life with turnaround time (TAT) of 13.0 ± 0.4 days. Clinical care was altered by the diagnosis in 23/32 (71.9%) patients. The diagnostic yield, patient age at diagnosis, and medical impact in the group that underwent critical trio exome is significantly different comparing to regular exome testing. For deceased infants (n=81), genetic disorders were molecular diagnosed in 39 (48.1%) by exome sequencing with implications for recurrence risk counseling. Conclusions and relevance: Exome sequencing is a powerful tool for the diagnostic evaluation of critically ill infants with suspected monogenic disorders in the neonatal and pediatric ICUs, leading to notable impact on clinical decision-making.