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Congenital diaphragmatic hernia (CDH) is a life-threatening birth defect that presents as either an isolated diaphragm defect or as part of a complex disorder with a wide array of anomalies (complex CDH). Some patients with complex CDH display distinct craniofacial anomalies such as craniofrontonasal dysplasia or craniosynostosis, defined by the premature closure of cranial sutures. Using clinical whole exome sequencing (WES), we found a BCL11B missense variant in a patient with a left-sided congenital diaphragmatic hernia as well as sagittal suture craniosynostosis. We applied targeted sequencing of BCL11B in patients with craniosynostosis or with a combination of craniosynostosis and CDH. This resulted in three additional BCL11B missense mutations in patients with craniosynostosis. The phenotype of the patient with both CDH as well as craniosynostosis was similar to the phenotype of previously reported patients with BCL11B missense mutations. Although these findings imply that both craniosynostosis as well as CDH may be associated with BCL11B mutations, further studies are required to establish whether BCL11B variants are causative mutations for both conditions or if our finding was coincidental.

Spatial normalization is an important step for group image processing and evaluation of mean brain perfusion in anatomical regions using arterial spin labeling (ASL) MRI and is typically performed via high-resolution structural brain scans. However, structural segmentation and/or spatial normalization to standard space is complicated when gray-white matter contrast in structural images is low due to ongoing myelination in newborns and infants. This problem is of particularly clinical relevance for imaging infants with inborn or acquired disorders that impair normal brain development. We investigated whether the ASL MRI perfusion contrast is a viable alternative for spatial normalization, using a pseudo-continuous ASL acquired using a 1.5 T MRI unit (GE Healthcare). Four approaches have been compared: (1) using the structural image contrast, or perfusion contrast with (2) rigid, (3) affine, and (4) nonlinear transformations – in 16 healthy controls [median age 0.83 years, inter-quartile range (IQR) ± 0.56] and 36 trigonocephaly patients (median age 0.50 years, IQR ± 0.30) – a non-syndromic type of craniosynostosis. Performance was compared quantitatively using the real-valued Tanimoto coefficient (TC), visually by three blinded readers, and eventually by the impact on regional cerebral blood flow (CBF) values. For both patients and controls, nonlinear registration using perfusion contrast showed the highest TC, at 17.51 (CI 6.66–49.38) times more likely to have a higher rating and 17.45–18.88 ml/100 g/min higher CBF compared with the standard normalization. Using perfusion-based contrast improved spatial normalization compared with the use of structural images, significantly affected the regional CBF, and may open up new possibilities for future large pediatric ASL brain studies.

Background: Genetic tests for primary immunodeficiency disorders (PIDs) are expensive, time-consuming, and not easily accessible in developing countries. Therefore, we studied the feasibility of a customized single nucleotide variant (SNV) microarray that we developed to detect disease-causing variants and copy number variation (CNV) in patients with PIDs for only 40 Euros.Methods: Probes were custom-designed to genotype 9,415 variants of 277 PID-related genes, and were added to the genome-wide Illumina Global Screening Array (GSA). Data analysis of GSA was performed using Illumina GenomeStudio 2.0, Biodiscovery Nexus 10.0, and R-3.4.4 software. Validation of genotype calling was performed by comparing the GSA with whole-genome sequencing (WGS) data of 56 non-PID controls. DNA samples of 95 clinically diagnosed PID patients, of which 60 patients (63%) had a genetically established diagnosis (by Next-Generation Sequencing (NGS) PID panels or Sanger sequencing), were analyzed to test the performance of the GSA. The additional SNVs detected by GSA were validated by Sanger sequencing.Results: Genotype calling of the customized array had an accuracy rate of 99.7%. The sensitivity for detecting rare PID variants was high (87%). The single sample replication in two runs was high (94.9%). The customized GSA was able to generate a genetic diagnosis in 37 out of 95 patients (39%). These 37 patients included 29 patients in whom the genetic variants were confirmed by conventional methods (26 patients by SNV and 3 by CNV analysis), while in 8 patients a new genetic diagnosis was established (6 patients by SNV and 2 patients suspected for leukemia by CNV analysis). Twenty-eight patients could not be detected due to the limited coverage of the custom probes. However, the diagnostic yield can potentially be increased when newly updated variants are added.Conclusion: Our robust customized GSA seems to be a promising first-line rapid screening tool for PIDs at an affordable price, which opens opportunities for low-cost genetic testing in developing countries. The technique is scalable, allows numerous new genetic variants to be added, and offers the potential for genetic testing not only in PIDs, but also in many other genetic diseases.