"Working Together": Perspectives of Healthcare Professionals in Providing Virtual Care to Youth with Chronic Pain during the COVID-19 Pandemic.

BACKGROUND: The onset of the coronavirus disease in 2019 necessitated a rapid transition to virtual care for chronic pain treatment. METHODS: A mixed methods design was implemented using qualitative interviews and quantitative satisfaction surveys. Interviews were conducted in February 2021 with a sample of healthcare professionals (HCPs; n = 6) who had provided multidisciplinary treatment (MDT) through an outpatient hospital pediatric chronic pain program. Satisfaction surveys were distributed to all MDT professionals employed by the clinic in April 2021 (n = 13 of 20 eligible; 65% response rate). Participants represented medicine, rehabilitation, and mental health professionals. RESULTS: Analysis of interviews generated five themes: (1) adaptation to virtual care, (2) benefits of virtual care, (3) limitations of virtual care, (4) shifting stance on virtual care over time, and (5) considerations for implementing virtual care. The satisfaction survey data revealed that respondents were able to effectively provide appropriate diagnoses, recommendations, and/or care plans for pediatric chronic pain via virtual care (n = 12, 92.3%). Detailed survey responses are presented by discipline. CONCLUSIONS: This study provides a rich exploration of HCPs' experiences in providing MDT for pediatric chronic pain within a virtual care model. The current results may contribute to the future development of guidelines for virtual care delivery with pediatric chronic pain populations.

Virtual multidisciplinary pain treatment: Experiences and feedback from children with chronic pain and their caregivers.

BACKGROUND: The onset of the coronavirus disease 2019 (COVID-19) necessitated a rapid transition to virtual care for chronic pain treatment. OBJECTIVE: This study examined experiences of patients and caregivers who received virtual multidisciplinary pain treatment (MDT) for pediatric chronic pain between March 2020 and August 2021. METHODS: A mixed methods design was implemented using qualitative interviews and quantitative satisfaction surveys. Satisfaction surveys were administered to a convenience sample of patients (aged 8 to 18; N = 20) and their caregivers (N = 20) who received MDT through an outpatient hospital pediatric chronic pain program. Interviews were conducted with a subset of these patients and their caregivers (n = 6). RESULTS: Analysis of interviews revealed four themes: 1) benefits of virtual care; 2) challenges of virtual care; 3) recommendations and evaluation of virtual care; and 4) patient preferences. Analysis of the satisfaction survey data revealed that while patients and caregivers were satisfied with many aspects of virtual care, 65% (n = 13) of patients reported a preference for in-person appointments, with caregivers showing equal preference for in-person and virtual appointments, though this was a non-significant difference (p = .37). Overall, both patients and caregivers stated a stronger preference for in-person physiotherapy sessions but were willing to have psychology sessions provided virtually. Finally, the most reported preference was for a hybrid model of care incorporating at least some in-person contact with providers. CONCLUSION: This study provides a rich exploration of virtual care for multidisciplinary pediatric chronic pain treatment. The current results may inform the future development of guidelines for virtual care delivery with pediatric chronic pain populations.

Resolution of post-traumatic chronic testicular pain in a pediatric patient after microsurgical denervation of the spermatic cord: a case report.

PURPOSE: Chronic scrotal content pain, chronic orchialgia, or testicular pain can present after trauma, vasectomy, and hernia repair, among other triggers. Microsurgical denervation of the spermatic cord is an option for definitive pain control. While this practice is established in adult urology, access to diagnostic intervention and definitive denervation surgery is limited in the pediatric population. CLINICAL FEATURES: We report a case of definitive resolution of testicular pain with microsurgical denervation of the spermatic cord in a pediatric patient with post-traumatic chronic orchialgia that significantly reduced his daily activities and worsened his anxiety prior to this treatment. The patient underwent attempts at conservative medication-based management, followed by diagnostic spermatic cord nerve block before definitive denervation surgery. CONCLUSIONS: The incidence of chronic pain in pediatrics is substantial and is estimated to be around 20%. Orchialgia remains difficult and problematic to treat. Mental health diagnoses such as anxiety and depression are also significantly associated with chronic pain. Following consideration and implementation of steps for all these concerns, a diagnostic block and microsurgical denervation led to successful resolution of chronic testicular pain in a pediatric patient.

RéSUMé: OBJECTIF: La douleur chronique au niveau du contenu scrotal, l'orchialgie chronique ou la douleur testiculaire peuvent apparaître à la suite d'un traumatisme, d'une vasectomie, et d'une réparation herniaire, entre autres déclencheurs. La dénervation microchirurgicale du cordon spermatique est une option pour le contrôle définitif de la douleur. Bien que cette pratique soit établie en urologie de l'adulte, l'accès à l'intervention diagnostique et à une chirurgie de dénervation définitive est limité pour la population pédiatrique. CARACTéRISTIQUES CLINIQUES : Nous rapportons un cas de résolution définitive d'une douleur testiculaire par dénervation microchirurgicale du cordon spermatique chez un patient pédiatrique souffrant d'orchialgie chronique post-traumatique, laquelle réduisait de manière significative ses activités quotidiennes et empirait son anxiété, avant ce traitement. Le patient a subi plusieurs tentatives de prises en charge conservatrices à l'aide de traitements médicamenteux, suivies d'un bloc nerveux diagnostique du cordon spermatique avant chirurgie de dénervation définitive. CONCLUSION: L'incidence de douleur chronique en pédiatrie est importante et est estimée à environ 20%. L'orchialgie reste difficile et problématique à traiter. Des diagnostics de santé mentale tels que l'anxiété et la dépression sont également associés de manière significative à la douleur chronique. Après examen et mise en œuvre d'étapes pour tenir compte de toutes ces préoccupations, un bloc diagnostique et une dénervation microchirurgicale ont conduit à la résolution réussie de la douleur testiculaire chronique chez un patient pédiatrique.

Genetic Diagnosis and the Severity of Cardiovascular Phenotype in Patients With Elastin Arteriopathy.

BACKGROUND: Elastin insufficiency causes recurrent vascular stenoses. Hemizygous deletion of the elastin gene (ELN) causes Williams-Beuren syndrome (WBS), while single nucleotide variants in ELN cause nonsyndromic supravalvar aortic stenosis (SVAS). Our objective was to compare cardiovascular disease outcomes in patients with WBS and nonsyndromic SVAS. METHODS: Patients (81 WBS, 42 nonsyndromic SVAS) with cardiovascular disease were included in this retrospective single center study. Freedom from surgical and catheter interventions and reinterventions was compared. Vascular tissue from 8 patients and 6 controls was analyzed for arterial wall architecture. RESULTS: Patients with nonsyndromic SVAS presented at a younger age (median 0.3 [0.4-0.7] years) compared with patients with WBS (1.3 [0.2-3.0] years) and had lower freedom from surgical/catheter interventions compared with patients with WBS, with median event-free survival 1.1 (0.3-5.9) versus 4.7 (2.4-13.3) years, respectively (hazard ratio, 1.62 [95% CI, 1.02-2.56]; P=0.04). Patients with nonsyndromic SVAS also had a lower freedom from reinterventions (P=0.054 by log-rank test). This was related in part to a higher frequency of primary and reinterventions for concomitant valvar aortic stenosis. Histology revealed abnormal intimal and medial thickening, disorganized and fragmented elastic fibers, reduced smooth muscle calponin expression, and increased macrophage marker, CD68, expression in the arterial walls in patients with WBS and nonsyndromic SVAS compared with controls. CONCLUSIONS: Patients with nonsyndromic SVAS require early and more frequent vascular and valvular interventions and reinterventions, in particular for concomitant valvar aortic stenosis compared with patients with WBS. This provides important prognostic information to guide counseling of affected families with cardiovascular disease and may guide primary intervention strategies based on predicted risk of restenosis.

Rapid mobilization of a virtual pediatric chronic pain clinic in Canada during the COVID-19 pandemic.

BACKGROUND: Studies have been conducted describing the potential for using virtual care software during disasters and public health emergencies. However, limited data exist on ways in which the Canadian health care system utilizes virtual care during disasters or public health emergencies. AIMS: Due to the need for social distancing and reduction of nonessential ambulatory services during the COVID-19 pandemic, the SickKids Chronic Pain Clinic sought to transition care delivery from in person to virtual. The virtual clinic aimed to reduce risks associated with physical contact and environmental exposure without reducing access to care itself. METHODS: Harnessing of various digital tools including Ontario Telemedicine Network Guestlink, Zoom, and Microsoft Teams. The Chronic Pain Clinic Team worked together to communicate with patients and families, schedule virtual visits, establish remote access to clinical data collection tools, digitize the after-visit summary, and add resources on pain self-management to the clinic's website. RESULTS: The Chronic Pain Clinic successfully transitioned all clinic appointments (multidisciplinary and individual; 77 appointments) over a 2-week period to virtual care. Virtual clinics did not surpass the usual time taken pre-COVID-19, suggesting that the clinic workflow was readily adaptable to virtual care. CONCLUSIONS: Access to quality virtual care is essential to prevent chronic pain from taking a toll on the lives of patients and families. Rapid establishment of a virtual clinic without gaps in service delivery to patients is possible given institutional support and a team culture centered around collaboration and flexibility.

Contexte: Des études ont été menées pour décrire le potentiel d'utilisation des logiciels de soins virtuels lors de catastrophes et d'urgences de santé publique. Toutefois, il existe peu de données sur les moyens par lesquels le systéme de soins de santé canadien utilise les soins virtuels lors de catastrophes ou d'urgences de santé publique.But: En raison de la nécessité d'une distanciation sociale et de la réduction des services ambulatoires non essentiels pendant la pandémie de COVID-19, la clinique de douleur chronique de SickKids a cherché à assurer une transition de la prestation des soins en personne vers des soins virtuels. La clinique virtuelle visait à réduire les risques liés au contact physique et à l'exposition environnementale sans réduire l'accès aux soins en tant que tels.Méthodes: Exploitation de divers outils numériques, dont le Réseau de télémédecine de l'Ontario Guestlink, Zoom et Microsoft Teams. L'équipe de la Clinique de la douleur chronique a travaillé ensemble pour communiquer avec les patients et les familles, programmer des visites virtuelles, établir un accès à distance aux outils de collecte de données cliniques, numériser le résumé après la visite et ajouter des ressources sur l'auto-prise en charge de la douleur sur le site web de la clinique.Résultats: La Clinique de la douleur chronique a réussi à faire passer tous les rendez-vous de la clinique (multidisciplinaires et individuels; 77 rendez-vous) sur une période de deux semaines à des soins virtuels. Les cliniques virtuelles n'ont pas dépassé le temps habituel avant la COVID-19, ce qui indique que le flux de travail de la clinique était facilement adaptable aux soins virtuels.Conclusions: L'accès à des soins virtuels de qualité est essentiel pour éviter que la douleur chronique ne fasse des ravages dans la vie des patients et des familles. L'établissement rapide déune clinique virtuelle sans interruption dans la prestation des services aux patients est possible grâce à un soutien institutionnel et à une culture d'èquipe centrèe sur la collaboration et la souplesse.

Genetic architecture of laterality defects revealed by whole exome sequencing.

Aberrant left-right patterning in the developing human embryo can lead to a broad spectrum of congenital malformations. The causes of most laterality defects are not known, with variants in established genes accounting for <20% of cases. We sought to characterize the genetic spectrum of these conditions by performing whole-exome sequencing of 323 unrelated laterality cases. We investigated the role of rare, predicted-damaging variation in 1726 putative laterality candidate genes derived from model organisms, pathway analyses, and human phenotypes. We also evaluated the contribution of homo/hemizygous exon deletions and gene-based burden of rare variation. A total of 28 candidate variants (26 rare predicted-damaging variants and 2 hemizygous deletions) were identified, including variants in genes known to cause heterotaxy and primary ciliary dyskinesia (ACVR2B, NODAL, ZIC3, DNAI1, DNAH5, HYDIN, MMP21), and genes without a human phenotype association, but with prior evidence for a role in embryonic laterality or cardiac development. Sanger validation of the latter variants in probands and their parents revealed no de novo variants, but apparent transmitted heterozygous (ROCK2, ISL1, SMAD2), and hemizygous (RAI2, RIPPLY1) variant patterns. Collectively, these variants account for 7.1% of our study subjects. We also observe evidence for an excess burden of rare, predicted loss-of-function variation in PXDNL and BMS1- two genes relevant to the broader laterality phenotype. These findings highlight potential new genes in the development of laterality defects, and suggest extensive locus heterogeneity and complex genetic models in this class of birth defects.

Changes to Neural Activation Patterns (c-fos Labeling) in Chinchilla Auditory Midbrain following Neonatal Exposure to an Enhanced Sound Environment.

Sensory brain regions show neuroplastic changes following deficits or experimental augmentation of peripheral input during a neonatal period. We have previously shown reorganization of cortical tonotopic maps after neonatal cochlear lesions or exposure to an enhanced acoustic environment. Such experiments probe the cortex and show reorganization, but it is unclear if such changes are intrinsically cortical or reflect projections from modified subcortical regions. Here, we ask whether an enhanced neonatal acoustic environment can induce midbrain (inferior colliculus (IC)) changes. Neonatal chinchillas were chronically exposed to a 70 dB SPL narrowband (2 ± 0.25 kHz) sound stimulus for 4 weeks. In line with previous studies, we hypothesized that such exposure would induce widening of the 2 kHz tonotopic map region in IC. To probe c-fos expression in IC (central nucleus), sound-exposed and nonexposed animals were stimulated with a 2 kHz stimulus for 90 minutes. In sound-exposed subjects, we find no change in the width of the 2 kHz tonotopic region; thus, our hypothesis is not supported. However, we observed a significant increase in the number of c-fos-labeled neurons over a broad region of best frequencies. These data suggest that neonatal sound exposure can modify midbrain regions and thus change the way neurons in IC respond to sound stimulation.

Whole exome sequencing in 342 congenital cardiac left sided lesion cases reveals extensive genetic heterogeneity and complex inheritance patterns.

BACKGROUND: Left-sided lesions (LSLs) account for an important fraction of severe congenital cardiovascular malformations (CVMs). The genetic contributions to LSLs are complex, and the mutations that cause these malformations span several diverse biological signaling pathways: TGFB, NOTCH, SHH, and more. Here, we use whole exome sequence data generated in 342 LSL cases to identify likely damaging variants in putative candidate CVM genes. METHODS: Using a series of bioinformatics filters, we focused on genes harboring population-rare, putative loss-of-function (LOF), and predicted damaging variants in 1760 CVM candidate genes constructed a priori from the literature and model organism databases. Gene variants that were not observed in a comparably sequenced control dataset of 5492 samples without severe CVM were then subjected to targeted validation in cases and parents. Whole exome sequencing data from 4593 individuals referred for clinical sequencing were used to bolster evidence for the role of candidate genes in CVMs and LSLs. RESULTS: Our analyses revealed 28 candidate variants in 27 genes, including 17 genes not previously associated with a human CVM disorder, and revealed diverse patterns of inheritance among LOF carriers, including 9 confirmed de novo variants in both novel and newly described human CVM candidate genes (ACVR1, JARID2, NR2F2, PLRG1, SMURF1) as well as established syndromic CVM genes (KMT2D, NF1, TBX20, ZEB2). We also identified two genes (DNAH5, OFD1) with evidence of recessive and hemizygous inheritance patterns, respectively. Within our clinical cohort, we also observed heterozygous LOF variants in JARID2 and SMAD1 in individuals with cardiac phenotypes, and collectively, carriers of LOF variants in our candidate genes had a four times higher odds of having CVM (odds ratio = 4.0, 95% confidence interval 2.5-6.5). CONCLUSIONS: Our analytical strategy highlights the utility of bioinformatic resources, including human disease records and model organism phenotyping, in novel gene discovery for rare human disease. The results underscore the extensive genetic heterogeneity underlying non-syndromic LSLs, and posit potential novel candidate genes and complex modes of inheritance in this important group of birth defects.

Assessment of large copy number variants in patients with apparently isolated congenital left-sided cardiac lesions reveals clinically relevant genomic events.

Congenital left-sided cardiac lesions (LSLs) are a significant contributor to the mortality and morbidity of congenital heart disease (CHD). Structural copy number variants (CNVs) have been implicated in LSL without extra-cardiac features; however, non-penetrance and variable expressivity have created uncertainty over the use of CNV analyses in such patients. High-density SNP microarray genotyping data were used to infer large, likely-pathogenic, autosomal CNVs in a cohort of 1,139 probands with LSL and their families. CNVs were molecularly confirmed and the medical records of individual carriers reviewed. The gene content of novel CNVs was then compared with public CNV data from CHD patients. Large CNVs (>1 MB) were observed in 33 probands (∼3%). Six of these were de novo and 14 were not observed in the only available parent sample. Associated cardiac phenotypes spanned a broad spectrum without clear predilection. Candidate CNVs were largely non-recurrent, associated with heterozygous loss of copy number, and overlapped known CHD genomic regions. Novel CNV regions were enriched for cardiac development genes, including seven that have not been previously associated with human CHD. CNV analysis can be a clinically useful and molecularly informative tool in LSLs without obvious extra-cardiac defects, and may identify a clinically relevant genomic disorder in a small but important proportion of these individuals.

Midbrain Frequency Representation following Moderately Intense Neonatal Sound Exposure in a Precocious Animal Model (Chinchilla laniger).

Auditory brain areas undergo reorganization resulting from abnormal sensory input during early postnatal development. This is evident from studies at the cortical level but it remains unclear whether there is reorganization in the auditory midbrain in a species similar to the human, that is, with early hearing onset. We have explored midbrain plasticity in the chinchilla, a precocious species that matches the human in terms of hearing development. Neonatal chinchillas were chronically exposed to a 2 kHz narrowband sound at 70 dB SPL for 4 weeks. Tonotopic maps in inferior colliculus (central nucleus) were defined based on single neuron characteristic frequency. We hypothesized an overrepresentation of the 2 kHz region of the maps. However, we observed a significant decrease in the proportion of neurons dedicated to the 2 kHz octave band and also away from the exposure frequency at 8 kHz. In addition, we report a significant increase in low frequency representation (<1 kHz), again a change to tonotopic mapping distant to the 2 kHz region. Thus in a precocious species, tonotopic maps in auditory midbrain are altered following abnormal stimulation during development. However, these changes are more complex than the overrepresentation of exposure related frequency regions that are often reported.

Bi-allelic Mutations in PKD1L1 Are Associated with Laterality Defects in Humans.

Disruption of the establishment of left-right (L-R) asymmetry leads to situs anomalies ranging from situs inversus totalis (SIT) to situs ambiguus (heterotaxy). The genetic causes of laterality defects in humans are highly heterogeneous. Via whole-exome sequencing (WES), we identified homozygous mutations in PKD1L1 from three affected individuals in two unrelated families. PKD1L1 encodes a polycystin-1-like protein and its loss of function is known to cause laterality defects in mouse and medaka fish models. Family 1 had one fetus and one deceased child with heterotaxy and complex congenital heart malformations. WES identified a homozygous splicing mutation, c.6473+2_6473+3delTG, which disrupts the invariant splice donor site in intron 42, in both affected individuals. In the second family, a homozygous c.5072G>C (p.Cys1691Ser) missense mutation was detected in an individual with SIT and congenital heart disease. The p.Cys1691Ser substitution affects a highly conserved cysteine residue and is predicted by molecular modeling to disrupt a disulfide bridge essential for the proper folding of the G protein-coupled receptor proteolytic site (GPS) motif. Damaging effects associated with substitutions of this conserved cysteine residue in the GPS motif have also been reported in other genes, namely GPR56, BAI3, and PKD1 in human and lat-1 in C. elegans, further supporting the likely pathogenicity of p.Cys1691Ser in PKD1L1. The identification of bi-allelic PKD1L1 mutations recapitulates previous findings regarding phenotypic consequences of loss of function of the orthologous genes in mice and medaka fish and further expands our understanding of genetic contributions to laterality defects in humans.

Utility of genetic evaluation in infants with congenital heart defects admitted to the cardiac intensive care unit.

Congenital heart defects (CHDs) are heterogeneous and present with a spectrum of severity, with roughly 25% of patients requiring intervention before age 1. The etiology of disease is unknown in many individuals; however, there is a rapidly expanding understanding of genetic risk factors that may contribute to pathogenesis. Through this work, we sought to evaluate the diagnostic yield of a clinical genetics evaluation and associated genetic testing among infants with critical CHDs. Furthermore, we aimed to both determine the utility of microarray and establish a strong baseline that can be used in future studies of the impact of exome sequencing in this population. We completed a retrospective chart review of 364 infants with CHDs admitted to the Cardiac Intensive Care Unit who underwent a clinical genetics evaluation. A genetic diagnosis was established in 25% of patients: 9% of infants were diagnosed prenatally, while 16% were diagnosed postnatally. Cardiac lesion subtype greatly influenced the diagnostic yield. On physical exam, the presence of dysmorphic features, as assessed by a clinical geneticist, was associated with a sevenfold increased likelihood of reaching a diagnosis. Directed by clinical acumen, diagnostic rates varied by testing modality with rates of 23% for karyotype, 12% for fluorescent in situ hybridization or multiplex-dependent ligation probe analysis, 9% for genome wide microarray, and 17% for targeted gene sequencing. Careful consideration of lesion subtype and physical exam findings clarify populations of infants with CHD that benefit from a genetics evaluation and inform an efficient testing paradigm. © 2016 Wiley Periodicals, Inc.

Recommendations for the integration of genomics into clinical practice.

The introduction of diagnostic clinical genome and exome sequencing (CGES) is changing the scope of practice for clinical geneticists. Many large institutions are making a significant investment in infrastructure and technology, allowing clinicians to access CGES, especially as health-care coverage begins to extend to clinically indicated genomic sequencing-based tests. Translating and realizing the comprehensive clinical benefits of genomic medicine remain a key challenge for the current and future care of patients. With the increasing application of CGES, it is necessary for geneticists and other health-care providers to understand its benefits and limitations in order to interpret the clinical relevance of genomic variants identified in the context of health and disease. New, collaborative working relationships with specialists across diverse disciplines (e.g., clinicians, laboratorians, bioinformaticians) will undoubtedly be key attributes of the future practice of clinical genetics and may serve as an example for other specialties in medicine. These new skills and relationships will also inform the development of the future model of clinical genetics training curricula. To address the evolving role of the clinical geneticist in the rapidly changing climate of genomic medicine, two Clinical Genetics Think Tank meetings were held that brought together physicians, laboratorians, scientists, genetic counselors, trainees, and patients with experience in clinical genetics, genetic diagnostics, and genetics education. This article provides recommendations that will guide the integration of genomics into clinical practice.Genet Med 18 11, 1075-1084.

A genome-wide association study of congenital cardiovascular left-sided lesions shows association with a locus on chromosome 20.

Congenital heart defects involving left-sided lesions (LSLs) are relatively common birth defects with substantial morbidity and mortality. Previous studies have suggested a high heritability with a complex genetic architecture, such that only a few LSL loci have been identified. We performed a genome-wide case-control association study to address the role of common variants using a discovery cohort of 778 cases and 2756 controls. We identified a genome-wide significant association mapping to a 200 kb region on chromosome 20q11 [P= 1.72 × 10-8 for rs3746446; imputed Single Nucleotide Polymorphism (SNP) rs6088703 P= 3.01 × 10-9, odds ratio (OR)= 1.6 for both]. This result was supported by transmission disequilibrium analyses using a subset of 541 case families (lowest P in region= 4.51 × 10-5, OR= 1.5). Replication in a cohort of 367 LSL cases and 5159 controls showed nominal association (P= 0.03 for rs3746446) resulting in P= 9.49 × 10-9 for rs3746446 upon meta-analysis of the combined cohorts. In addition, a group of seven SNPs on chromosome 1q21.3 met threshold for suggestive association (lowest P= 9.35 × 10-7 for rs12045807). Both regions include genes involved in cardiac development-MYH7B/miR499A on chromosome 20 and CTSK, CTSS and ARNT on chromosome 1. Genome-wide heritability analysis using case-control genotyped SNPs suggested that the mean heritability of LSLs attributable to common variants is moderately high ([Formula: see text] range= 0.26-0.34) and consistent with previous assertions. These results provide evidence for the role of common variation in LSLs, proffer new genes as potential biological candidates, and give further insight to the complex genetic architecture of congenital heart disease.

Exome sequencing identifies rare variants in multiple genes in atrioventricular septal defect.

PURPOSE: The genetic etiology of atrioventricular septal defect (AVSD) is unknown in 40% cases. Conventional sequencing and arrays have identified the etiology in only a minority of nonsyndromic individuals with AVSD. METHODS: Whole-exome sequencing was performed in 81 unrelated probands with AVSD to identify potentially causal variants in a comprehensive set of 112 genes with strong biological relevance to AVSD. RESULTS: A significant enrichment of rare and rare damaging variants was identified in the gene set, compared with controls (odds ratio (OR): 1.52; 95% confidence interval (CI): 1.35-1.71; P = 4.8 × 10(-11)). The enrichment was specific to AVSD probands, compared with a cohort without AVSD with tetralogy of Fallot (OR: 2.25; 95% CI: 1.84-2.76; P = 2.2 × 10(-16)). Six genes (NIPBL, CHD7, CEP152, BMPR1a, ZFPM2, and MDM4) were enriched for rare variants in AVSD compared with controls, including three syndrome-associated genes (NIPBL, CHD7, and CEP152). The findings were confirmed in a replication cohort of 81 AVSD probands. CONCLUSION: Mutations in genes with strong biological relevance to AVSD, including syndrome-associated genes, can contribute to AVSD, even in those with isolated heart disease. The identification of a gene set associated with AVSD will facilitate targeted genetic screening in this cohort.

High throughput exome coverage of clinically relevant cardiac genes.

BACKGROUND: Given the growing use of whole-exome sequencing (WES) for clinical diagnostics of complex human disorders, we evaluated coverage of clinically relevant cardiac genes on WES and factors influencing uniformity and depth of coverage of exonic regions. METHODS: Two hundred and thirteen human DNA samples were exome sequenced via Illumina HiSeq using different versions of the Agilent SureSelect capture kit. 50 cardiac genes were further analyzed including 31 genes from the American College of Medical Genetics (ACMG) list for reporting of incidental findings and 19 genes associated with congenital heart disease for which clinical testing is available. Gene coordinates were obtained from two databases, CCDS and Known Gene and compared. Read depth for each region was extracted from the exomes and used to assess capture variability between kits for individual genes, and for overall coverage. GC content, gene size, and inter-sample variability were also tested as potential contributors to variability in gene coverage. RESULTS: All versions of capture kits (designed based on Consensus coding sequence) included only 55% of known genomic regions for the cardiac genes. Although newer versions of each Agilent kit showed improvement in capture of CCDS regions to 99%, only 64% of Known Gene regions were captured even with newer capture kits. There was considerable variability in coverage of the cardiac genes. 10 of the 50 genes including 6 on the ACMG list had less than the optimal coverage of 30X. Within each gene, only 32 of the 50 genes had the majority of their bases covered at an interquartile range ≥30X. Heterogeneity in gene coverage was modestly associated with gene size and significantly associated with GC content. CONCLUSIONS: Despite improvement in overall coverage across the exome with newer capture kit versions and higher sequencing depths, only 50% of known genomic regions of clinical cardiac genes are targeted and individual gene coverage is non-uniform. This may contribute to a bias with greater attribution of disease causation to mutations in well-represented and well-covered genes. Improvements in WES technology are needed before widespread clinical application.

Excitatory and inhibitory tonotopic bands in chinchilla inferior colliculus revealed by c-fos immuno-labeling.

We describe in detail a reliable experimental protocol for c-fos immuno-labeling of patterns of neural activation in the chinchilla (chinchilla laniger). We report on resting-level neural activity in inferior colliculus (IC) of auditory midbrain, and on tonotopic bands present following 90 min of pure-tone sound stimulation. Neurons activated by 6-kHz sound stimulation lay ventro-medial to those activated at 2 kHz. This is consistent with the known tonotopic organization of IC, and verified in the present report by multi-unit neuron response recordings in central nucleus of IC. Of particular interest, we observe a significant reduction in cell labeling adjacent to the tonotopic bands, and suggest that such decreases represent inhibitory regions. C-fos-labeled bands and lateral regions of reduced labeling resemble excitatory and lateral-inhibitory response areas of IC neurons.

Rare variants in NR2F2 cause congenital heart defects in humans.

Congenital heart defects (CHDs) are the most common birth defect worldwide and are a leading cause of neonatal mortality. Nonsyndromic atrioventricular septal defects (AVSDs) are an important subtype of CHDs for which the genetic architecture is poorly understood. We performed exome sequencing in 13 parent-offspring trios and 112 unrelated individuals with nonsyndromic AVSDs and identified five rare missense variants (two of which arose de novo) in the highly conserved gene NR2F2, a very significant enrichment (p = 7.7 × 10(-7)) compared to 5,194 control subjects. We identified three additional CHD-affected families with other variants in NR2F2 including a de novo balanced chromosomal translocation, a de novo substitution disrupting a splice donor site, and a 3 bp duplication that cosegregated in a multiplex family. NR2F2 encodes a pleiotropic developmental transcription factor, and decreased dosage of NR2F2 in mice has been shown to result in abnormal development of atrioventricular septa. Via luciferase assays, we showed that all six coding sequence variants observed in individuals significantly alter the activity of NR2F2 on target promoters.