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Abstract: A diagnosis of choroideraemia (CHM) can be made clinically, based on the fundus examination and a family history consistent with X-linked inheritance. Molecular genetic testing offers a means of confirming the clinical diagnosis, establishing carrier status and allows presymptomatic diagnosis for families who wish to pursue these options. The aim of this study was to examine the uptake and assess the results from a diagnostic molecular genetics service for CHM. We have carried out a comprehensive audit of all molecular genetic results of UK NHS patients and families referred to the North West Regional Molecular Genetics Laboratory in Manchester, UK over a 55 month period. 110 people were referred to this service for testing including diagnostic, carrier and predictive requests. Putative pathogenic mutations were identified in 65/83 (78%) of male index cases. The identification of a familial pathogenic change enabled carrier testing in 16 asymptomatic females and predictive testing in 3 males. Case examples illustrate the range of cases referred for testing and also reflect the need for genetic counselling that results from offering a molecular diagnostic service such as this. Clinical molecular testing for CHM is available clinically and can be used to support the clinical diagnosis and management of patients with choroideraemia as well as their families. Case studies demonstrate the need to provide genetic testing to families and the potential clinical utility of testing. [Copyright &y& Elsevier]

Background: Prader-Willi syndrome (PWS) and Angelman syndrome (AS) are clinically distinct neurodevelopmental genetic disorders that map to 15q11-q13. The primary phenotypes are attributable to loss of expression of imprinted genes within this region which can arise by means of a number of mechanisms. The most sensitive single approach to diagnosing both PWS and AS is to study methylation patterns within 15q11-q13; however many techniques exist for this purpose. Given the diversity of techniques available, there is a need for consensus testing and reporting guidelines. Methods: Testing and reporting guidelines have been drawn up and agreed in accordance with the procedures of the UK Clinical Molecular Genetics Society and the European Molecular Genetics Quality Network. Results: A practical set of molecular genetic testing and reporting guidelines has been developed for these two disorders. In addition, advice is given on appropriate reporting policies, including advice on test sensitivity and recurrence risks. In considering test sensitivity, the possibility of differential diagnoses is discussed. Conclusion: An agreed set of practice guidelines has been developed for the diagnostic molecular genetic testing of PWS and AS. [ABSTRACT FROM AUTHOR]

A letter to the editor is presented in response to the article on external quality assessment schemes for real-time polymerase chain reaction (PCR).

The article discusses the guidelines regarding the clinical utility of genetic testing for the disease choroideremia or tapetochoroidal dystrophy. Topics discussed include the mutational spectrum caused by mutations in the CHM gene, life-time risk of developing the disease if the test is positive, and a molecular diagnosis in an affected individual concerning the disease's genetic situation.

The article offers information on the genetic mutations in the BEST1-related dystrophies or Bestrophinopathies. It informs that only single families have been identified as having BEST1 mutations. It reports that the p.Thr6Pro, mutation associated with the Best disease, has a particularly high frequency in the Netherlands.

Pseudoachondroplasia (PSACH) and autosomal dominant multiple epiphyseal dysplasia (MED) are chondrodysplasias resulting in short-limbed dwarfism, joint pain and stiffness and early onset osteoarthritis. All PSACH, and the largest proportion of MED, result from mutations in cartilage oligomeric matrix protein (COMP). The first mutations in COMP were identified in 1995 in patients with both PSACH and MED and subsequently there has been over 30 publications describing COMP mutations in at least 250 PSACH-MED patients. However, despite these discoveries, a methodical analysis of the relationship between COMP mutations and phenotypes has not been undertaken. In particular, there has, to date, been little correlation between the type and location of a COMP mutation and the resulting phenotype of PSACH or MED. To determine if genotype to phenotype correlations could be derived for COMP, we collated 300 COMP mutations, including 25 recently identified novel mutations. The results of this analysis demonstrate that mutations in specific residues and/or regions of the type III repeats of COMP are significantly associated with either PSACH or MED. This newly derived genotype to phenotype correlation may aid in determining the prognosis of PSACH and MED, including the prediction of disease severity, and in the long term guide genetic counselling and contribute to the clinical management of patients with these diseases. [ABSTRACT FROM AUTHOR]

Aims To describe the UK NEQAS for Molecular Genetics scheme for gastrointestinal stromal tumour (GIST) and to report and interpret the findings of four rounds of circulation of this quality assurance programme for KIT/PDGFRA mutation analyses. Methods Samples of GISTs from formalin-fixed paraffinembedded tissue blocks were circulated to registered participants of the UK NEQAS for Molecular Genetics scheme for GIST. Three samples were provided per annual circulation from 2008 to 2011 inclusive. The participants were required to analyse the samples for KIT and/or PDGFRA mutations using their routine protocols, and the anonymised participants' reports were assessed and an annual scheme report issued. Results The genotyping error rates for the 2008, 2009, 2010 and 2011 circulations were 13%, 33%, 19% and 4%, respectively. These errors were either missed or incorrectly described mutations. There was an overall false negative rate of 2% and false positive rate of 0%. The main recommendations that arose from these circulations were: (1) inclusion of reference accession numbers in reports; (2) avoidance of the term 'heterozygous' when analysing DNA from tumour tissue unless there was certainty that only neoplastic DNA was studied; and (3) the need to screen KIT exons 9, 11, 13 and 17 and PDGFRA exons 12, 14 and 18 before classifying a GIST as 'wild type'. Conclusions The UK NEQAS for Molecular Genetics scheme emphasises the potential complexities of KIT/ PDGFRA mutation analyses for GISTs and provides recommendations to help optimise such genotyping and reporting. The scheme has also demonstrated its educational value among participating laboratories. [ABSTRACT FROM AUTHOR]

Purpose To assess the clinical usefulness of genetic testing in a pediatric population with inherited retinal disease (IRD). Design Single-center retrospective case series. Participants Eighty-five unrelated children with a diagnosis of isolated or syndromic IRD who were referred for clinical genetic testing between January 2014 and July 2016. Methods Participants underwent a detailed ophthalmic examination, accompanied by electrodiagnostic testing (EDT) and dysmorphologic assessment where appropriate. Ocular and extraocular features were recorded using Human Phenotype Ontology terms. Subsequently, multigene panel testing (105 or 177 IRD-associated genes) was performed in an accredited diagnostic laboratory, followed by clinical variant interpretation. Main Outcome Measures Diagnostic yield and clinical usefulness of genetic testing. Results Overall, 78.8% of patients (n = 67) received a probable molecular diagnosis; 7.5% (n = 5) of these had autosomal dominant disease, 25.4% (n = 17) had X-linked disease, and 67.2% (n = 45) had autosomal recessive disease. In a further 5.9% of patients (n = 5), a single heterozygous ABCA4 variant was identified; all these participants had a spectrum of clinical features consistent with ABCA4 retinopathy. Most participants (84.7%; n = 72) had undergone EDT and 81.9% (n = 59) of these patients received a probable molecular diagnosis. The genes most frequently mutated in the present cohort were CACNA1F and ABCA4 , accounting for 14.9% (n = 10) and 11.9% (n = 8) of diagnoses respectively. Notably, in many cases, genetic testing helped to distinguish stationary from progressive IRD subtypes and to establish a precise diagnosis in a timely fashion. Conclusions Multigene panel testing pointed to a molecular diagnosis in 84.7% of children with IRD. The diagnostic yield in the study population was significantly higher compared with that in previously reported unselected IRD cohorts. Approaches similar to the one described herein are expected to become a standard component of care in pediatric ophthalmology. We propose the introduction of genetic testing early in the diagnostic pathway in children with clinical and/or electrophysiologic findings, suggestive of IRD. [ABSTRACT FROM AUTHOR]

Background: Although the majority of small in-frame insertions/deletions (indels) has no/little affect on protein function, a small subset of these changes has been causally associated with genetic disorders. Notably, the molecular mechanisms and frequency by which they give rise to disease phenotypes remain largely unknown. The aim of this study is to provide insights into the role of in-frame indels (≤21 nucleotides) in two genetically heterogeneous eye disorders.Results: One hundred eighty-one probands with childhood cataracts and 486 probands with retinal dystrophy underwent multigene panel testing in a clinical diagnostic laboratory. In-frame indels were collected and evaluated both clinically and in silico. Variants that could be modeled in the context of protein structure were identified and analysed using integrative structural modeling. Overall, 55 small in-frame indels were detected in 112 of 667 probands (16.8 %); 17 of these changes were novel to this study and 18 variants were reported clinically. A reliable model of the corresponding protein sequence could be generated for 8 variants. Structural modeling indicated a diverse range of molecular mechanisms of disease including disruption of secondary and tertiary protein structure and alteration of protein-DNA binding sites.Conclusions: In childhood cataract and retinal dystrophy subjects, one small in-frame indel is clinically reported in every ~37 individuals tested. The clinical utility of computational tools evaluating these changes increases when the full complexity of the involved molecular mechanisms is embraced. [ABSTRACT FROM AUTHOR]

Purpose To compare the efficacy of whole genome sequencing (WGS) with targeted next-generation sequencing (NGS) in the diagnosis of inherited retinal disease (IRD). Design Case series. Participants A total of 562 patients diagnosed with IRD. Methods We performed a direct comparative analysis of current molecular diagnostics with WGS. We retrospectively reviewed the findings from a diagnostic NGS DNA test for 562 patients with IRD. A subset of 46 of 562 patients (encompassing potential clinical outcomes of diagnostic analysis) also underwent WGS, and we compared mutation detection rates and molecular diagnostic yields. In addition, we compared the sensitivity and specificity of the 2 techniques to identify known single nucleotide variants (SNVs) using 6 control samples with publically available genotype data. Main Outcome Measures Diagnostic yield of genomic testing. Results Across known disease-causing genes, targeted NGS and WGS achieved similar levels of sensitivity and specificity for SNV detection. However, WGS also identified 14 clinically relevant genetic variants through WGS that had not been identified by NGS diagnostic testing for the 46 individuals with IRD. These variants included large deletions and variants in noncoding regions of the genome. Identification of these variants confirmed a molecular diagnosis of IRD for 11 of the 33 individuals referred for WGS who had not obtained a molecular diagnosis through targeted NGS testing. Weighted estimates, accounting for population structure, suggest that WGS methods could result in an overall 29% (95% confidence interval, 15–45) uplift in diagnostic yield. Conclusions We show that WGS methods can detect disease-causing genetic variants missed by current NGS diagnostic methodologies for IRD and thereby demonstrate the clinical utility and additional value of WGS. [ABSTRACT FROM AUTHOR]

Prader Willi and Angelman syndromes are clinically distinct genetic disorders both mapping to chromosome region 15q11-q13, which are caused by a loss of function of paternally or maternally inherited genes in the region, respectively. With clinical diagnosis often being difficult, particularly in infancy, confirmatory genetic diagnosis is essential to enable clinical intervention. However, the latter is challenged by the complex genetics behind both disorders and the unmet need for characterised reference materials to aid accurate molecular diagnosis. With this in mind, a panel of six genotyping reference materials for Prader Willi and Angelman syndromes was developed, which should be stable for many years and available to all diagnostic laboratories. The panel comprises three Prader Willi syndrome materials (two with different paternal deletions, and one with maternal uniparental disomy (UPD)) and three Angelman syndrome materials (one with a maternal deletion, one with paternal UPD or an epigenetic imprinting centre defect, and one with a UBE3A point mutation). Genomic DNA was bulk-extracted from Epstein-Barr virus-transformed lymphoblastoid cell lines established from consenting patients, and freeze-dried as aliquots in glass ampoules. In total, 37 laboratories from 26 countries participated in a collaborative study to assess the suitability of the panel. Participants evaluated the blinded, triplicate materials using their routine diagnostic methods against in-house controls or externally sourced uncertified reference materials. The panel was established by the Expert Committee on Biological Standardization of the World Health Organization as the first International Genetic Reference Panel for Prader Willi and Angelman syndromes. [ABSTRACT FROM AUTHOR]

Fragile X syndrome is the most common inherited form of mental retardation. It is caused by expansion of a trinucleotide (CGG)n repeat sequence in the 5′ untranslated region of the FMR1 gene, resulting in promoter hypermethylation and suppression of FMR1 transcription. Additionally, pre-mutation alleles in carrier males and females may result in Fragile X tremor ataxia syndrome and primary ovarian insufficiency, respectively. Fragile X is one of the most commonly requested molecular genetic tests worldwide. Quality assessment schemes have identified a wide disparity in allele sizing between laboratories. It is therefore important that clinical laboratories have access to characterized reference materials (RMs) to aid accurate allele sizing and diagnosis. With this in mind, a panel of genotyping RMs for Fragile X syndrome has been developed, which should be stable over many years and available to all diagnostic laboratories. Immortalized cell lines were produced by Epstein-Barr virus transformation of lymphocytes from consenting patients. Genomic DNA was extracted in bulk and RM aliquots were freeze-dried in glass ampoules. Twenty-one laboratories from seventeen countries participated in a collaborative study to assess their suitability. Participants evaluated the samples (blinded, in triplicate) in their routine methods alongside in-house and commercial controls. The panel of five genomic DNA samples was endorsed by the European Society of Human Genetics and approved as an International Standard by the Expert Committee on Biological Standardization at the World Health Organization. [ABSTRACT FROM AUTHOR]

Background: FCI is an R code for analyzing data from real-time PCR experiments. This algorithm estimates standard curve features as well as nucleic acid concentrations and confidence intervals according to Fieller's theorem. Results: In order to describe the features of FCI four situations were selected from real data collected during an international external quality assessment program for quantitative assays based on real-time PCR. The code generates a diagnostic figure suitable for assessing the quality of the quantification process. Conclusion: We have provided a freeware programme using this algorithm specifically designed to increase the information content of the real-time PCR assay. [ABSTRACT FROM AUTHOR]

Individuals who have ocular features of albinism and skin pigmentation in keeping with their familial background present a considerable diagnostic challenge. Timely diagnosis through genomic testing can help avert diagnostic odysseys and facilitates accurate genetic counselling and tailored specialist management. Here, we report the clinical and gene panel testing findings in 12 children with presumed ocular albinism. A definitive molecular diagnosis was made in 8/12 probands (67%) and a possible molecular diagnosis was identified in a further 3/12 probands (25%). TYR was the most commonly mutated gene in this cohort (75% of patients, 9/12). A disease-causing TYR haplotype comprised of two common, functional polymorphisms, TYR c.[575 C > A;1205 G > A] p.[(Ser192Tyr);(Arg402Gln)], was found to be particularly prevalent. One participant had GPR143-associated X-linked ocular albinism and another proband had biallelic variants in SLC38A8, a glutamine transporter gene associated with foveal hypoplasia and optic nerve misrouting without pigmentation defects. Intriguingly, 2/12 individuals had a single, rare, likely pathogenic variant in each of TYR and OCA2 – a significant enrichment compared to a control cohort of 4046 individuals from the 100,000 genomes project pilot dataset. Overall, our findings highlight that panel-based genetic testing is a clinically useful test with a high diagnostic yield in children with partial/ocular albinism. [ABSTRACT FROM AUTHOR]

The article presents a case study of a male infant in 2002 who was diagnosed with non-syndromic infantile-onset retinal dystrophy and whose younger sister in 2006, was also diagnosed the same condition. Whole exome sequencing was used to test the boy's DNA in 2013 and identified a homozygous two-nucleotide delection in the IQCB1 gene. Further investigation of the boy at age twelve proved he has renal impairment and anaemia.

Purpose To assess the utility of integrating genomic data from next-generation sequencing and phenotypic data to enhance the diagnosis of bilateral congenital cataract (CC). Design Evaluation of diagnostic technology. Participants Thirty-six individuals diagnosed with nonsyndromic or syndromic bilateral congenital cataract were selected for investigation through a single ophthalmic genetics clinic. Methods Participants underwent a detailed ophthalmic examination, accompanied by dysmorphology assessment where appropriate. Lenticular, ocular, and systemic phenotypes were recorded. Mutations were detected using a custom-designed target enrichment that permitted parallel analysis of 115 genes associated with CC by high-throughput, next-generation DNA sequencing (NGS). Thirty-six patients and a known positive control were tested. Suspected pathogenic variants were confirmed by bidirectional Sanger sequencing in relevant probands and other affected family members. Main Outcome Measures Molecular genetic results and details of clinical phenotypes were identified. Results Next-generation DNA sequencing technologies are able to determine the precise genetic cause of CC in 75% of individuals, and 85% patients with nonsyndromic CC were found to have likely pathogenic mutations, all of which occurred in highly conserved domains known to be vital for normal protein function. The pick-up rate in patients with syndromic CC also was high, with 63% having potential disease-causing mutations. Conclusions This analysis demonstrates the clinical utility of this test, providing examples where it altered clinical management, directed care pathways, and enabled more accurate genetic counseling. This comprehensive screen will extend access to genetic testing and lead to improved diagnostic and management outcomes through a stratified medicine approach. Establishing more robust genotype–phenotype correlations will advance knowledge of cataract-forming mechanisms. [ABSTRACT FROM AUTHOR]

A correction to the article "Preparation and validation of the first WHO international genetic reference panel for Fragile X syndrome" that was published in the August 25, 2010 issue is presented.

The article presents a study to analyze data on all test requests for fragile X syndrome in children received over a five-year period in the North West of Great Britain using a standard performa. It mentions that Great Britain guidance requires fragile X testing as a first line investigation in children with global developmental delay. It found that several girls had no early developmental delay.