Purpose:To prospectively evaluate the diagnostic and clinical utility of singleton whole-exome sequencing (WES) as a first-tier test in infants with suspected monogenic disease. Methods:Singleton WES was performed as a first-tier sequencing test in infants recruited from a single pediatric tertiary center. This occurred in parallel with standard investigations, including single-or multigene panel sequencing when clinically indicated. The diagnosis rate, clinical utility, and impact on management of singleton WES were evaluated.Results: Of 80 enrolled infants, 46 received a molecular genetic diagnosis through singleton WES (57.5%) compared with 11 (13.75%) who underwent standard investigations in the same patient group. Clinical management changed following exome diagnosis in 15 of 46 diagnosed participants (32.6%). Twelve relatives received a genetic diagnosis following cascade testing, and 28 couples were identified as being at high risk of recurrence in future pregnancies. Conclusions:This prospective study provides strong evidence for increased diagnostic and clinical utility of singleton WES as a firsttier sequencing test for infants with a suspected monogenic disorder. Singleton WES outperformed standard care in terms of diagnosis rate and the benefits of a diagnosis, namely, impact on management of the child and clarification of reproductive risks for the extended family in a timely manner.

Background-The chromosome 17q21.31 microdeletion syndrome is a novel genomic disorder that has originally been identified using high resolution genome analyses in patients with unexplained mental retardation.

IMPORTANCE Optimal use of whole-exome sequencing (WES) in the pediatric setting requires an understanding of who should be considered for testing and when it should be performed to maximize clinical utility and cost-effectiveness.OBJECTIVES To investigate the impact of WES in sequencing-naive children suspected of having a monogenic disorder and evaluate its cost-effectiveness if WES had been available at different time points in their diagnostic trajectory. DESIGN, SETTING, AND PARTICIPANTS This prospective study was part of the MelbourneGenomics Health Alliance demonstration project. At the ambulatory outpatient clinics of the Victorian Clinical Genetics Services at the Royal Children's Hospital, Melbourne, Australia, children older than 2 years suspected of having a monogenic disorder were prospectively recruited from May 1 through November 30, 2015, by clinical geneticists after referral from general and subspecialist pediatricians. All children had nondiagnostic microarrays and no prior single-gene or panel sequencing.EXPOSURES All children underwent singleton WES with targeted phenotype-driven analysis. MAIN OUTCOMES AND MEASURESThe study examined the clinical utility of a molecular diagnosis and the cost-effectiveness of alternative diagnostic trajectories, depending on timing of WES. RESULTSOf 61 children originally assessed, 44 (21 [48%] male and 23 [52%] female) aged 2 to 18 years (mean age at initial presentation, 28 months; range, 0-121 months) were recruited, and a diagnosis was achieved in 23 (52%) by singleton WES. The diagnoses were unexpected in 8 of 23 (35%), and clinical management was altered in 6 of 23 (26%). The mean duration of the diagnostic odyssey was 6 years, with each child having a mean of 19 tests and 4 clinical genetics and 4 nongenetics specialist consultations, and 26 (59%) underwent a procedure while under general anesthetic for diagnostic purposes. Economic analyses of the diagnostic trajectory identified that WES performed at initial tertiary presentation resulted in an

Disorders of sex development (DSD), ranging in severity from mild genital abnormalities to complete sex reversal, represent a major concern for patients and their families. DSD are often due to disruption of the genetic programs that regulate gonad development. Although some genes have been identified in these developmental pathways, the causative mutations have not been identified in more than 50% 46,XY DSD cases. We used the Affymetrix Genome-Wide Human SNP Array 6.0 to analyse copy number variation in 23 individuals with unexplained 46,XY DSD due to gonadal dysgenesis (GD). Here we describe three discrete changes in copy number that are the likely cause of the GD. Firstly, we identified a large duplication on the X chromosome that included DAX1 (NR0B1). Secondly, we identified a rearrangement that appears to affect a novel gonad-specific regulatory region in a known testis gene, SOX9. Surprisingly this patient lacked any signs of campomelic dysplasia, suggesting that the deletion affected expression of SOX9 only in the gonad. Functional analysis of potential SRY binding sites within this deleted region identified five putative enhancers, suggesting that sequences additional to the known SRY-binding TES enhancer influence human testis-specific SOX9 expression. Thirdly, we identified a small deletion immediately downstream of GATA4, supporting a role for GATA4 in gonad development in humans. These CNV analyses give new insights into the pathways involved in human gonad development and dysfunction, and suggest that rearrangements of non-coding sequences disturbing gene regulation may account for significant proportion of DSD cases.