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Background: Möbius (Moebius) and Poland's syndromes are two rare congenital syndromes characterized by non-progressive bilateral (and often asymmetric) dysfunction of the 6 th and 7 th cranial nerves and hypoplasia of the pectoral muscles associated with chest wall and upper limb anomalies respectively. Manifest simultaneously as Poland-Möbius (Poland-Moebius) syndrome, debate continues as to whether this is a distinct nosological entity or represents phenotypic variation as part of a spectrum of disorders of rhomboencephalic development. Etiological hypotheses implicate both genetic and environmental factors. The PLXND1 gene codes for a protein expressed in the fetal central nervous system and vascular endothelium and is thus involved in embryonic neurogenesis and vasculogenesis. It is located at chromosome region 3q21-q22, a locus of interest for Möbius syndrome., Case Presentation: We present the first report of a patient with Poland-Möbius syndrome and a mutation in the PLXND1 gene. A child with Poland-Möbius syndrome and a maternally inherited missense variant (NM_015103.2:ex14:c.2890G > Ap.V964M) in the PLXND1 gene is described. In order to contextualize these findings, the literature was examined to identify other confirmed cases of Poland-Möbius syndrome for which genetic data were available. Fourteen additional cases of Poland-Möbius syndrome with genetic studies are described in the literature. None implicated the PLXND1 gene which has previously been implicated in isolated Möbius syndrome., Conclusions: This report provides further evidence in support of a role for PLXND1 mutations in Möbius syndrome and reasserts the nosological link between Möbius and Poland's syndromes., Level of Evidence: Level V, Descriptive Study., (© 2022. The Author(s).)

Background: New Zealand has a multiethnic population and a national cardiac inherited disease registry (Cardiac Inherited Disease Registry New Zealand [CIDRNZ]). Ancestry is reflected in the spectrum and prevalence of genetic variants in long QT syndrome (LQTS)., Objective: The purpose of this study was to study the genetic testing yield and mutation spectrum of CIDRNZ LQTS probands stratified by self-identified ethnicity., Methods: A 15-year retrospective review of clinical CIDRNZ LQTS probands with a Schwartz score of ≥2 who had undergone genetic testing was performed., Results: Of the 264 included LQTS probands, 160 (61%) reported as European, 79 (30%) NZ Māori and Pacific peoples (Polynesian), and 25 (9%) Other ethnicities, with comparable clinical characteristics across ethnic groups (cardiac events in 72%; age at presentation 28±19 years; corrected QT interval 512±55 ms). Despite comparable testing (5.3±1.4 LQTS genes), a class III-V LQTS variant was identified in 35% of Polynesian probands as compared with 63% of European and 72% of Other probands (P<.0001). Among variant-positive CIDRNZ LQTS probands (n=148), Polynesians were more likely to have non-missense variants (57% vs 39% and 25% in probands of European and Other ethnicity, respectively; P=.005) as well as long QT syndrome type 1-3 variants not reported elsewhere (71% vs European 22% and Other 28%; P<.0001). Variants found in multiple probands were more likely to be shared within the same ethnic group; P<.01)., Conclusion: Genetic testing of Polynesian LQTS probands has a lower diagnostic yield, despite comparable testing and clinical disease severity. Rare LQTS variants are more common in Polynesian LQTS probands. These data emphasize the importance of increasing the knowledge of genetic variation in the Polynesian population., (Copyright © 2020 Heart Rhythm Society. Published by Elsevier Inc. All rights reserved.)

Objectives: Molecular diagnostic laboratories screen for mutations in disease-causing genes in order to confirm a clinical diagnosis. The classification of DNA variants as 'pathogenic' or 'likely pathogenic' mutations creates a workflow bottleneck, which becomes increasingly challenging as greater number of genes are screened. The classification challenge is also acute if there are conflicting reports regarding pathogenicity and differing classification criteria between laboratories. This study aimed to compare two procedures for the classification of variants in the breast cancer ( BRCA ) 1 gene., Methods: This bioinformatic study was conducted at LabPLUS, Auckland, New Zealand, from February to June 2017. DNA was extracted from peripheral blood samples of 30 patients and gene library construction was carried out using a commercially available targeted panel for the BRCA1 and BRCA2 genes. The genes were subsequently sequenced and the sequence data analysed. The guidelines published by the American College of Medical Genetics and Genomics and the Association for Molecular Pathology (ACMG/AMP) provides a comprehensive framework for the interpretation of variants in genes that are associated with Mendelian disorders. The use of these guidelines were compared to the variant classifications that were achieved by reference to those reported in the BRCA Exchange database., Results: The results showed concordance between the two classification protocols for a panel of 30 BRCA1 gene variants, although the transparency in following the ACMG/AMP guidelines provides a diagnostic laboratory with a generalisable approach that allows laboratory-directed revisions to be undertaken in light of new information., Conclusion: The ACMG/AMP-based guidelines were applied to a cohort of patients with BRCA1 gene variants. The use of these guidelines provides a system which creates consistency in variant interpretation and supports subsequent clinical management., Competing Interests: CONFLICT OF INTEREST The authors declare no conflicts of interest., (© Copyright 2019, Sultan Qaboos University Medical Journal, All Rights Reserved.)

Camurati-Engelmann disease (OMIM 31300) is a rare cranio-tubular bone dysplasia characterized by osteosclerosis of the long bones and skull caused by dominantly-inherited mutations in the transforming growth factor beta 1 (TGFB1) gene. A wide variation in phenotype has been recognized, even within families carrying the same mutation. In addition, aspects of the natural history of the disorder, in particular whether it is always progressive or can remit spontaneously, remain uncertain. In a large kindred carrying a TGFB1 gene mutation (c.653G > A; p.R218H) we have attempted to clarify the extent of phenotypic variability and the natural history of the disease through detailed individual histories of symptoms, and skeletal imaging by both radiography and scintigraphy. Only one subject had the classical childhood onset with bone pain in the legs and gait disturbance. Eight subjects reported the onset of leg pain in their teenage years that, by their early 20s, had either resolved or persisted at a low level. Two of these eight later developed cranial nerve palsies. There was a wide variation in the radiographic appearance in adults, but disease extent and activity in long bones, as assessed by scintigraphy, was inversely correlated with age (p < 0.025). In younger subjects the radiographic and scintigraphic appearances were concordant, but in older subjects the scintigram could be quiescent despite florid radiographic changes. Sequential scintigrams in two subjects showed reduced activity in the later scan. One subject had suffered meningoencephalitis in early childhood that resulted in paresis of one arm. The affected arm showed markedly less disease involvement, implicating mechanical or growth factors in its etiology. Our data suggest that the natural history of Camurati-Engelmann disease can be benign, and that disease activity commonly attenuates in adulthood. Severe cases of childhood onset and/or with cranial nerve involvement, may occur only in a minority of mutation carriers. © 2019 American Society for Bone and Mineral Research., (© 2019 American Society for Bone and Mineral Research.)

Sudden cardiac death (SCD) in people before the age of 35 years is a devastating event for any family. The causes of SCD in the young can be broadly divided into two groups: heritable cardiac disorders that affect the heart structure (cardiomyopathies) and primary electrical disorders (cardiac ion channelopathies). Genetic testing is vital as those suffering from cardiac ion channelopathies have structurally normal hearts, and those with cardiomyopathies may only show subtle abnormalities in the heart and these signs may not be detected during an autopsy. Post-mortem genetic testing of SCD victims is important to identify the underlying genetic cause. This is important as family cascade screening may be undertaken to identify those who may be at risk and provide vital information about risk stratification and clinical management. The development of massively parallel sequencing (MPS) has made it possible for the simultaneous screening of multiple patients for hundreds of genes. In light of this, we opted to develop an MPS approach for SCD analysis that would allow us to screen for mutations in genes implicated in cardiomyopathies and cardiac ion channelopathies. The rationale behind this panel was to limit it to genes carrying the greatest mutation load. If no likely pathogenic gene variant were found then testing could cascade to whole exome/genome sequencing as a gene-discovery exercise. The overarching aim was to design and validate a custom-cardiac panel that satisfies the diagnostic requirements of LabPLUS (Auckland City Hospital, Auckland, NZ) and the guidelines provided by the Royal College of Pathologists of Australasia and the Association for Clinical Genetic Science., Competing Interests: The authors declare no conflict of interest.

Background: Approximately 75% of clinically definite long QT syndrome (LQTS) cases are caused by mutations in the KCNQ1 , KCNH2 and SCN5A genes. Of these mutations, a small proportion (3.2-9.2%) are predicted to affect splicing. These mutations present a particular challenge in ascribing pathogenicity., Methods: Here we report an analysis of the transcriptional consequences of two mutations, one in the KCNQ1 gene (c.781&#95;782delinsTC) and one in the SCN5A gene (c.2437-5C>A), which are predicted to affect splicing. We isolated RNA from lymphocytes and used a directed PCR amplification strategy of cDNA to show mis-spliced transcripts in mutation-positive patients., Results: The loss of an exon in each mis-spliced transcript had no deduced effect on the translational reading frame. The clinical phenotype corresponded closely with genotypic status in family members carrying the KCNQ1 splice variant, but not in family members with the SCN5A splice variant. These results are put in the context of a literature review, where only 20% of all splice variants reported in the KCNQ1 , KCNH2 and SCN5A gene entries in the HGMDPro 2015.4 database have been evaluated using transcriptional assays., Conclusions: Prediction programmes play a strong role in most diagnostic laboratories in classifying variants located at splice sites; however, transcriptional analysis should be considered critical to confirm mis-splicing. Critically, this study shows that genuine mis- splicing may not always imply clinical significance, and genotype/phenotype cosegregation remains important even when mis-splicing is confirmed., Competing Interests: The authors have no conflicts of interest to declare, and alone are responsible for the content and writing of the paper.

Two male siblings from a consanguineous union presented in early infancy with marked truncal hypotonia, a general paucity of movement, extrapyramidal signs and cognitive delay. By mid-childhood they had made little developmental progress and remained severely hypotonic and bradykinetic. They developed epilepsy and had problems with autonomic dysfunction and oculogyric crises. They had a number of orthopaedic problems secondary to their hypotonia. Cerebrospinal fluid (CSF) neurotransmitters were initially normal, apart from mildly elevated 5-hydroxyindolacetic acid, and the children did not respond favourably to a trial of levodopa-carbidopa. The youngest died from respiratory complications at 10 years of age. Repeat CSF neurotransmitters in the older sibling at eight years of age showed slightly low homovanillic acid and 5-hydroxyindoleacetic acid levels. Whole-exome sequencing revealed a novel mutation homozygous in both children in the monoamine transporter gene SLC18A2 (p.Pro237His), resulting in brain dopamine-serotonin vesicular transport disease. This is the second family to be described with a mutation in this gene. Treatment with the dopamine agonist pramipexole in the surviving child resulted in mild improvements in alertness, communication, and eye movements. This case supports the identification of the causal mutation in the original case, expands the clinical phenotype of brain dopamine-serotonin vesicular transport disease and confirms that pramipexole treatment may lead to symptomatic improvement in affected individuals.

The increasing diagnostic use of gene sequencing has led to an expanding dataset of novel variants that lie within consensus splice junctions. The challenge for diagnostic laboratories is the evaluation of these variants in order to determine if they affect splicing or are merely benign. A common evaluation strategy is to use in silico analysis, and it is here that a number of programmes are available online; however, currently, there are no consensus guidelines on the selection of programmes or protocols to interpret the prediction results. Using a collection of 222 pathogenic mutations and 50 benign polymorphisms, we evaluated the sensitivity and specificity of four in silico programmes in predicting the effect of each variant on splicing. The programmes comprised Human Splice Finder (HSF), Max Entropy Scan (MES), NNSplice, and ASSP. The MES and ASSP programmes gave the highest performance based on Receiver Operator Curve analysis, with an optimal cut-off of score reduction of 10%. The study also showed that the sensitivity of prediction is affected by the level of conservation of individual positions, with in silico predictions for variants at positions -4 and +7 within consensus splice sites being largely uninformative.

Objectives: Screening for mutations in large genes is challenging in a molecular diagnostic environment. Sanger-based DNA sequencing methods are largely used; however, massively parallel sequencing (MPS) can accommodate increasing test demands and financial constraints. This study aimed to establish a simple workflow to amplify and screen all coding regions of the BRCA1 and BRCA2 (BRCA1/2) genes by Sanger-based sequencing as well as to assess a MPS approach encompassing multiplex polymerase chain reaction (PCR) and pyrosequencing., Methods: This study was conducted between July 2011 and April 2013. A total of 20 patients were included in the study who had been referred to Genetic Health Services New Zealand (Northern Hub) for BRCA1/2 mutation screening. Patients were randomly divided into a MPS evaluation and validation cohort (n = 10 patients each). Primers were designed to amplify all coding exons of BRCA1/2 (28 and 42 primer pairs, respectively). Primers overlying known variants were avoided to circumvent allelic drop-out. The MPS approach necessitated utilisation of a complementary fragment analysis assay to eliminate apparent false-positives at homopolymeric regions. Variants were filtered on the basis of their frequency and sequence depth., Results: Sanger-based sequencing of PCR-amplified coding regions was successfully achieved. Sensitivity and specificity of the combined MPS/homopolymer protocol was determined to be 100% and 99.5%, respectively., Conclusion: In comparison to traditional Sanger-based sequencing, the MPS workflow led to a reduction in both cost and analysis time for BRCA1/2 screening. MPS analysis achieved high analytical sensitivity and specificity, but required complementary fragment analysis combined with Sanger-based sequencing confirmation in some instances.

Background: Catecholaminergic polymorphic ventricular tachycardia (CPVT) is a heritable cardiac disorder characterized by life-threatening ventricular tachycardia caused by exercise or acute emotional stress. The standard diagnostic screening involves Sanger-based sequencing of 45 of the 105 translated exons of the RYR2 gene, and copy number changes of a limited number of exons that are detected using multiplex ligation-dependent probe amplification (MLPA)., Methods: In the current study, a previously validated bespoke array comparative genomic hybridization (aCGH) technique was used to detect copy number changes in the RYR2 gene in a 43-year-old woman clinically diagnosed with CPVT., Results: The CGH array detected a 1.1 kb deletion encompassing exon 3 of the RYR2 gene. This is the first report using the aCGH technique to screen for mutations causing CPVT., Conclusions: The aCGH method offers significant advantages over MLPA in genetic screening for heritable cardiac disorders.

Background: Large gene rearrangements, not detectable by standard molecular genetic sequencing techniques, are present in a minority of patients with long QT syndrome. We aimed to screen for large rearrangements in genes responsible for long QT syndrome as part of the molecular autopsy of a 36-year-old woman who died suddenly and had a negative autopsy. A retrospective analysis of an ECG identified a long QT interval, but sequencing of known LQT genes was uninformative., Methods and Results: Array comparative genomic hybridization was used to screen for deletions and duplications in 101 genes implicated in cardiac disorders and sudden death using a postmortem blood sample. A 542 kb deletion encompassing the entire KCNJ2 gene was identified in the decedent. The mother had electrocardiographic U-wave changes consistent with Andersen-Tawil syndrome and exaggerated by exercise but none of the characteristic noncardiac features. Fluorescence in situ hybridization confirmed the deletion in the decedent and established its presence in the mother., Conclusions: A novel application of array comparative genomic hybridization and fluorescence in situ hybridization has identified that long QT syndrome and sudden cardiac death may occur as a result of a deletion of an entire gene. The case also supports recent research suggesting that noncardiac features of Andersen-Tawil syndrome occur only with missense or minor gene rearrangements in the KCNJ2 gene, resulting in a dominant negative effect on Kir2.x channels.

Genetic testing at a distance is commonplace where members of a family with a segregating germline mutation are geographically separated. For the most part, this challenge is addressed through the intervention of health professionals in taking and/or processing blood samples for subsequent couriering of DNA to a referral laboratory. In some circumstances, however, the collecting of pivotal clinical material may involve direct patient involvement. We describe such a situation where noninvasive saliva samples were provided by members of a family manifesting Von Hippel-Lindau (VHL) disease. The analysis identified a novel mutation in the VHL gene that was used to exclude other family members as being at risk of VHL disease.

The mitochondrial myopathies consist of a heterogeneous group of disorders caused by structural and functional abnormalities in mitochondria leading to involvement of the nervous system and muscles as well as other organ systems. The peculiar genetic characteristics of mitochondrial DNA impart distinctive properties to these disorders. The pathophysiology is presented. The methods employed in making the correct diagnosis, the preoperative patient assessment and correction of metabolic dysfunctions and anaesthetic techniques used, are highlighted. The conditions are briefly reviewed and suggestions are made for the safe anaesthetic management of affected patients.