Late-Onset Autosomal Dominant Macular Degeneration Caused by Deletion of the CRX Gene.

PURPOSE: To characterize the phenotype observed in a case series with macular disease and determine the cause. DESIGN: Multicenter case series. PARTICIPANTS: Six families (7 patients) with sporadic or multiplex macular disease with onset at 20 to 78 years, and 1 patient with age-related macular degeneration. METHODS: Patients underwent ophthalmic examination; exome, genome, or targeted sequencing; and/or polymerase chain reaction (PCR) amplification of the breakpoint, followed by cloning and Sanger sequencing or direct Sanger sequencing. MAIN OUTCOME MEASURES: Clinical phenotypes, genomic findings, and a hypothesis explaining the mechanism underlying disease in these patients. RESULTS: All 8 cases carried the same deletion encompassing the genes TPRX1, CRX, and SULT2A1, which was absent from 382 control individuals screened by breakpoint PCR and 13 096 Clinical Genetics patients with a range of other inherited conditions screened by array comparative genomic hybridization. Microsatellite genotypes showed that these 7 families are not closely related, but genotypes immediately adjacent to the deletion breakpoints suggest they may share a distant common ancestor. CONCLUSIONS: Previous studies had found that carriers for a single defective CRX allele that was predicted to produce no functional CRX protein had a normal ocular phenotype. Here, we show that CRX whole-gene deletion in fact does cause a dominant late-onset macular disease.

Novel homozygous mutations in the transcription factor NRL cause non-syndromic retinitis pigmentosa.

Purpose: To describe the clinical phenotype and genetic basis of non-syndromic retinitis pigmentosa (RP) in one family and two sporadic cases with biallelic mutations in the transcription factor neural retina leucine zipper (NRL). Methods: Exome sequencing was performed in one affected family member. Microsatellite genotyping was used for haplotype analysis. PCR and Sanger sequencing were used to confirm mutations in and screen other family members where they were available. The SMART tool for domain prediction helped us build the protein schematic diagram. Results: For family MM1 of Pakistani origin, whole-exome sequencing and microsatellite genotyping revealed homozygosity on chromosome 14 and identified a homozygous stop-loss mutation in NRL, NM_006177.5: c.713G>T, p.*238Lext57, which is predicted to add an extra 57 amino acids to the normal protein chain. The variant segregated with disease symptoms in the family. For case RP-3051 of Spanish ancestry, clinical exome sequencing focusing on the morbid genome highlighted a homozygous nonsense mutation in NRL, c.238C>T, p.Gln80*, as the most likely disease candidate. For case RP-1553 of Romanian ethnicity, targeted-exome sequencing of 73 RP/LCA genes identified a homozygous nonsense mutation in NRL, c.544C>T, p.Gln182*. The variants were either rare or absent in the gnomAD database. Conclusions: NRL mutations predominantly cause dominant retinal disease, but there have been five published reports of mutations causing recessive disease. Here, we present three further examples of recessive RP due to NRL mutations. The phenotypes observed are consistent with those in the previous reports, and the observed mutation types and distribution further confirm distinct patterns for variants in NRL causing recessive and dominant diseases.

New missense variants in RELT causing hypomineralised amelogenesis imperfecta.

Amelogenesis imperfecta (AI) is a heterogeneous group of genetic diseases characterised by dental enamel malformation. Pathogenic variants in at least 33 genes cause syndromic or non-syndromic AI. Recently variants in RELT, encoding an orphan receptor in the tumour necrosis factor (TNF) superfamily, were found to cause recessive AI, as part of a syndrome encompassing small stature and severe childhood infections. Here we describe four additional families with autosomal recessive hypomineralised AI due to previously unreported homozygous mutations in RELT. Three families carried a homozygous missense variant in the fourth exon (c.164C>T, p.(T55I)) and a fourth family carried a homozygous missense variant in the 11th exon (c.1264C>T, p.(R422W)). We found no evidence of additional syndromic symptoms in affected individuals. Analyses of tooth microstructure with computerised tomography and scanning electron microscopy suggest a role for RELT in ameloblasts' coordination and interaction with the enamel matrix. Microsatellite genotyping in families segregating the T55I variant reveals a shared founder haplotype. These findings extend the RELT pathogenic variant spectrum, reveal a founder mutation in the UK Pakistani population and provide detailed analysis of human teeth affected by this hypomineralised phenotype, but do not support a possible syndromic presentation in all those with RELT-variant associated AI.

Mutations in the polyglutamylase gene TTLL5, expressed in photoreceptor cells and spermatozoa, are associated with cone-rod degeneration and reduced male fertility.

Hereditary retinal degenerations encompass a group of genetic diseases characterized by extreme clinical variability. Following next-generation sequencing and autozygome-based screening of patients presenting with a peculiar, recessive form of cone-dominated retinopathy, we identified five homozygous variants [p.(Asp594fs), p.(Gln117*), p.(Met712fs), p.(Ile756Phe), and p.(Glu543Lys)] in the polyglutamylase-encoding gene TTLL5, in eight patients from six families. The two male patients carrying truncating TTLL5 variants also displayed a substantial reduction in sperm motility and infertility, whereas those carrying missense changes were fertile. Defects in this polyglutamylase in humans have recently been associated with cone photoreceptor dystrophy, while mouse models carrying truncating mutations in the same gene also display reduced fertility in male animals. We examined the expression levels of TTLL5 in various human tissues and determined that this gene has multiple viable isoforms, being highly expressed in testis and retina. In addition, antibodies against TTLL5 stained the basal body of photoreceptor cells in rat and the centrosome of the spermatozoon flagellum in humans, suggesting a common mechanism of action in these two cell types. Taken together, our data indicate that mutations in TTLL5 delineate a novel, allele-specific syndrome causing defects in two as yet pathogenically unrelated functions, reproduction and vision.

Biallelic mutations in the autophagy regulator DRAM2 cause retinal dystrophy with early macular involvement.

Retinal dystrophies are an overlapping group of genetically heterogeneous conditions resulting from mutations in more than 250 genes. Here we describe five families affected by an adult-onset retinal dystrophy with early macular involvement and associated central visual loss in the third or fourth decade of life. Affected individuals were found to harbor disease-causing variants in DRAM2 (DNA-damage regulated autophagy modulator protein 2). Homozygosity mapping and exome sequencing in a large, consanguineous British family of Pakistani origin revealed a homozygous frameshift variant (c.140delG [p.Gly47Valfs(∗)3]) in nine affected family members. Sanger sequencing of DRAM2 in 322 unrelated probands with retinal dystrophy revealed one European subject with compound heterozygous DRAM2 changes (c.494G>A [p.Trp165(∗)] and c.131G>A [p.Ser44Asn]). Inspection of previously generated exome sequencing data in unsolved retinal dystrophy cases identified a homozygous variant in an individual of Indian origin (c.64_66del [p.Ala22del]). Independently, a gene-based case-control association study was conducted via an exome sequencing dataset of 18 phenotypically similar case subjects and 1,917 control subjects. Using a recessive model and a binomial test for rare, presumed biallelic, variants, we found DRAM2 to be the most statistically enriched gene; one subject was a homozygote (c.362A>T [p.His121Leu]) and another a compound heterozygote (c.79T>C [p.Tyr27His] and c.217_225del [p.Val73_Tyr75del]). DRAM2 encodes a transmembrane lysosomal protein thought to play a role in the initiation of autophagy. Immunohistochemical analysis showed DRAM2 localization to photoreceptor inner segments and to the apical surface of retinal pigment epithelial cells where it might be involved in the process of photoreceptor renewal and recycling to preserve visual function.

Homozygous single base deletion in TUSC3 causes intellectual disability with developmental delay in an Omani family.

Intellectual disability (ID) is the term used to describe a diverse group of neurological conditions with congenital or juvenile onset, characterized by an IQ score of less than 70 and difficulties associated with limitations in cognitive function and adaptive behavior. The condition can be inherited or caused by environmental factors. The genetic forms are heterogeneous, with mutations in over 500 known genes shown to cause the disorder. We report a consanguineous Omani family in which multiple individuals have ID and developmental delay together with some variably present features including short stature, microcephaly, moderate facial dysmorphism, and congenital malformations of the toes or hands. Homozygosity mapping combined with whole exome next generation sequencing identified a novel homozygous single base pair deletion in TUSC3, c.222delA, p.R74 fs. The mutation segregates with the disease phenotype in a recessive manner and is absent in 60,706 unrelated individuals from various disease-specific and population genetic studies. TUSC3 mutations have been previously identified as causing either syndromic or non-syndromic ID in patients from France, Italy, Iran and Pakistan. This paper supports the previous clinical descriptions of the condition caused by TUSC3 mutations and describes the seventh family with mutations in this gene, thus contributing to the genetic spectrum of mutations. This is the first report of a family from the Arabian peninsula with this form of ID. © 2016 Wiley Periodicals, Inc.

Novel C8orf37 mutations cause retinitis pigmentosa in consanguineous families of Pakistani origin.

PURPOSE: To investigate the molecular basis of retinitis pigmentosa in two consanguineous families of Pakistani origin with multiple affected members. METHODS: Homozygosity mapping and Sanger sequencing of candidate genes were performed in one family while the other was analyzed with whole exome next-generation sequencing. A minigene splicing assay was used to confirm the splicing defects. RESULTS: In family MA48, a novel homozygous nucleotide substitution in C8orf37, c.244-2A>C, that disrupted the consensus splice acceptor site of exon 3 was found. The minigene splicing assay revealed that this mutation activated a cryptic splice site within exon 3, causing a 22 bp deletion in the transcript that is predicted to lead to a frameshift followed by premature protein truncation. In family MA13, a novel homozygous null mutation in C8orf37, c.555G>A, p.W185*, was identified. Both mutations segregated with the disease phenotype as expected in a recessive manner and were absent in 8,244 unrelated individuals of South Asian origin. CONCLUSIONS: In this report, we describe C8orf37 mutations that cause retinal dystrophy in two families of Pakistani origin, contributing further data on the phenotype and the spectrum of mutations in this form of retinitis pigmentosa.

Faecal Microbiota Transplantation [FMT] in the Treatment of Chronic Refractory Pouchitis: A Systematic Review and Meta-analysis.

BACKGROUND: The aim of this systematic review and meta-analysis is to assess the efficacy and safety of faecal microbiota transplantation [FMT] in the treatment of chronic pouchitis. METHODS: A PRISMA-compliant systematic review and meta-analysis was conducted using the following databases and clinical trial registers: Medline, Embase, Scopus, Cochrane Database of Systematic Reviews [CENTRAL], clinical trials.gov, ScienceDirect, and VHL [virtual health library]. The primary outcome was clinical response/remission in patients treated with FMT. Secondary outcomes included safety profile, quality of life, and changes in the gut microbiome. RESULTS: Seven observational cohort studies/case series and two randomised, controlled trials with a total of 103 patients were included. The route, preparation, and quantity of FMT administered varied among the included studies. Clinical response rate of 42.6% with a remission rate of 29.8% was estimated in our cohort following FMT therapy. Minor, self-limiting, adverse events were reported, and the treatment was well tolerated with good short- and long-term safety profiles. Successful FMT engraftment in recipients varied and, on average, microbial richness and diversity was lower in patients with pouchitis. In some instances, shifts with specific changes towards abundance of species, suggestive of a 'healthier' pouch microbiota, were observed following treatment with FMT. CONCLUSION: The evidence for FMT in the treatment of chronic pouchitis is sparse, which limits any recommendations being made for its use in clinical practice. Current evidence from low-quality studies suggests a variable clinical response and remission rate, but the treatment is well tolerated, with a good safety profile. This review emphasises the need for rationally designed, well-powered, randomised, placebo-controlled trials to understand the efficacy of FMT for the treatment of pouchitis.

The analysis of gut microbiota in patients with bile acid diarrhoea treated with colesevelam.

Introduction: Bile acid diarrhoea (BAD) is a common disorder that results from an increased loss of primary bile acids and can result in a change in microbiome. The aims of this study were to characterise the microbiome in different cohorts of patients with BAD and to determine if treatment with a bile acid sequestrant, colesevelam, can alter the microbiome and improve microbial diversity. Materials and methods: Patients with symptoms of diarrhoea underwent 75-selenium homocholic acid (75SeHCAT) testing and were categorised into four cohorts: idiopathic BAD, post-cholecystectomy BAD, post-operative Crohn's disease BAD and 75SeHCAT negative control group. Patients with a positive 75SeHCAT (<15%) were given a trial of treatment with colesevelam. Stool samples were collected pre-treatment, 4-weeks, 8-weeks and 6-12 months post-treatment. Faecal 16S ribosomal RNA gene analysis was undertaken. Results: A total of 257 samples were analysed from 134 patients. α-diversity was significantly reduced in patients with BAD and more specifically, in the idiopathic BAD cohort and in patients with severe disease (SeHCAT <5%); p < 0.05. Colesevelam did not alter bacterial α/ß-diversity but patients who clinically responded to treatment had a significantly greater abundance of Fusobacteria and Ruminococcus, both of which aid in the conversion of primary to secondary bile acids. Conclusion: This is the first study to examine treatment effects on the microbiome in BAD, which demonstrated a possible association with colesevelam on the microbiome through bile acid modulation in clinical responders. Larger studies are now needed to establish a causal relationship with colesevelam and the inter-crosstalk between bile acids and the microbiome.

Long-Read Nanopore Sequencing of RPGR ORF15 is Enhanced Following DNase I Treatment of MinION Flow Cells.

INTRODUCTION: RPGR ORF15 is an exon present almost exclusively in the retinal transcript of RPGR. It is purine-rich, repetitive and notoriously hard to sequence, but is a hotspot for mutations causing X-linked retinitis pigmentosa. METHODS: Long-read nanopore sequencing on MinION and Flongle flow cells was used to sequence RPGR ORF15 in genomic DNA from patients with inherited retinal dystrophy. A flow cell wash kit was used on a MinION flow cell to increase yield. Findings were confirmed by PacBio SMRT long-read sequencing. RESULTS: We showed that long-read nanopore sequencing successfully reads through a 2 kb PCR-amplified fragment containing ORF15. We generated reads of sufficient quality and cumulative read-depth to detect pathogenic RP-causing variants. However, we observed that this G-rich, repetitive DNA segment rapidly blocks the available pores, resulting in sequence yields less than 5% of the expected output. This limited the extent to which samples could be pooled, increasing cost. We tested the utility of a MinION wash kit containing DNase I to digest DNA fragments remaining on the flow cell, regenerating the pores. Use of the DNase I treatment allowed repeated re-loading, increasing the sequence reads obtained. Our customised workflow was used to screen pooled amplification products from previously unsolved inherited retinal disease (IRD) in patients, identifying two new cases with pathogenic ORF15 variants. DISCUSSION: We report the novel finding that long-read nanopore sequencing can read through RPGR-ORF15, a DNA sequence not captured by short-read next-generation sequencing (NGS), but with a more reduced yield. Use of a flow cell wash kit containing DNase I unblocks the pores, allowing reloading of further library aliquots over a 72-h period, increasing yield. The workflow we describe provides a novel solution to the need for a rapid, robust, scalable, cost-effective ORF15 screening protocol.

Impaired protein hydroxylase activity causes replication stress and developmental abnormalities in humans.

Although protein hydroxylation is a relatively poorly characterized posttranslational modification, it has received significant recent attention following seminal work uncovering its role in oxygen sensing and hypoxia biology. Although the fundamental importance of protein hydroxylases in biology is becoming clear, the biochemical targets and cellular functions often remain enigmatic. JMJD5 is a "JmjC-only" protein hydroxylase that is essential for murine embryonic development and viability. However, no germline variants in JmjC-only hydroxylases, including JMJD5, have yet been described that are associated with any human pathology. Here we demonstrate that biallelic germline JMJD5 pathogenic variants are deleterious to JMJD5 mRNA splicing, protein stability, and hydroxylase activity, resulting in a human developmental disorder characterized by severe failure to thrive, intellectual disability, and facial dysmorphism. We show that the underlying cellular phenotype is associated with increased DNA replication stress and that this is critically dependent on the protein hydroxylase activity of JMJD5. This work contributes to our growing understanding of the role and importance of protein hydroxylases in human development and disease.

Meta-analysis of the demographic and prognostic significance of gastrointestinal symptoms in COVID-19 patients.

Background and Aim: To evaluate the demographic and prognostic significance of gastrointestinal (GI) symptoms in patients with coronavirus disease 2019 (COVID-19). Methods: A systematic search of electronic information sources was conducted. Combined overall effect sizes were calculated using random-effects models for baseline demographic factors and outcomes including mortality, intensive care unit (ICU) admission, and length of hospital stay. Results: Twenty-four comparative observational studies reporting a total of 51 522 COVID-19 patients with (n = 6544) or without (n = 44 978) GI symptoms were identified. The patients with GI symptoms were of comparable age (mean difference [MD]: 0.25, 95% confidence interval [CI] -2.42 to 2.92, P = 0.86), rate of pre-existing hypertension (odds ratio [OR]: 1.11, 95% CI 0.86-1.42, P = 0.42), diabetes mellitus (OR: 1.14, 95% CI 0.91-1.44, P = 0.26), and coronary artery disease (OR: 1.00, 95% CI 0.86-1.16, P = 0.98) compared with those without GI symptoms. However, there were significantly more male patients in the GI symptoms group (OR: 0.85, 95% CI 0.75-0.95, P = 0.005). The presence of GI symptoms was associated with similar risk of mortality (OR: 0.73; 95% CI 0.47-1.13, P = 0.16), ICU admission (OR: 1.15; 95% CI 0.67-1.96, P = 0.62), and length of hospital stay (MD: 0.43; 95% CI -0.73 to 1.60, P = 0.47) when compared with their absence. Conclusion: Meta-analysis of the best possible available evidence demonstrated that GI symptoms in COVID-19 patients do not seem to affect patients with any specific demographic patterns and may not have any important prognostic significance. Although no randomized studies can be conducted on this topic, future high-quality studies can provide stronger evidence to further understand the impact of GI symptoms on outcomes of COVID-19 patients.

Patient Derived Organoids Confirm That PI3K/AKT Signalling Is an Escape Pathway for Radioresistance and a Target for Therapy in Rectal Cancer.

Objectives: Partial or total resistance to preoperative chemoradiotherapy occurs in more than half of locally advanced rectal cancer patients. Several novel or repurposed drugs have been trialled to improve cancer cell sensitivity to radiotherapy, with limited success. We aimed to understand the mechanisms of resistance to chemoradiotherapy in rectal cancer using patient derived organoid models. Design: To understand the mechanisms underlying this resistance, we compared the pre-treatment transcriptomes of patient-derived organoids (PDO) with measured radiotherapy sensitivity to identify biological pathways involved in radiation resistance coupled with single cell sequencing, genome wide CRISPR-Cas9 and targeted drug screens. Results: RNA sequencing enrichment analysis revealed upregulation of PI3K/AKT/mTOR and epithelial mesenchymal transition pathway genes in radioresistant PDOs. Single-cell sequencing of pre & post-irradiation PDOs showed mTORC1 and PI3K/AKT upregulation, which was confirmed by a genome-wide CRSIPR-Cas9 knockout screen using irradiated colorectal cancer (CRC) cell lines. We then tested the efficiency of dual PI3K/mTOR inhibitors in improving cancer cell sensitivity to radiotherapy. After irradiation, significant AKT phosphorylation was detected (p=0.027) which was abrogated with dual PI3K/mTOR inhibitors and lead to significant radiosensitisation of the HCT116 cell line and radiation resistant PDO lines. Conclusions: The PI3K/AKT/mTOR pathway upregulation contributes to radioresistance and its targeted pharmacological inhibition leads to significant radiosensitisation in CRC organoids, making it a potential target for clinical trials.

PDZD8 Disruption Causes Cognitive Impairment in Humans, Mice, and Fruit Flies.

BACKGROUND: The discovery of coding variants in genes that confer risk of intellectual disability (ID) is an important step toward understanding the pathophysiology of this common developmental disability. METHODS: Homozygosity mapping, whole-exome sequencing, and cosegregation analyses were used to identify gene variants responsible for syndromic ID with autistic features in two independent consanguineous families from the Arabian Peninsula. For in vivo functional studies of the implicated gene's function in cognition, Drosophila melanogaster and mice with targeted interference of the orthologous gene were used. Behavioral, electrophysiological, and structural magnetic resonance imaging analyses were conducted for phenotypic testing. RESULTS: Homozygous premature termination codons in PDZD8, encoding an endoplasmic reticulum-anchored lipid transfer protein, showed cosegregation with syndromic ID in both families. Drosophila melanogaster with knockdown of the PDZD8 ortholog exhibited impaired long-term courtship-based memory. Mice homozygous for a premature termination codon in Pdzd8 exhibited brain structural, hippocampal spatial memory, and synaptic plasticity deficits. CONCLUSIONS: These data demonstrate the involvement of homozygous loss-of-function mutations in PDZD8 in a neurodevelopmental cognitive disorder. Model organisms with manipulation of the orthologous gene replicate aspects of the human phenotype and suggest plausible pathophysiological mechanisms centered on disrupted brain development and synaptic function. These findings are thus consistent with accruing evidence that synaptic defects are a common denominator of ID and other neurodevelopmental conditions.

A Recessively Inherited Risk Locus on Chromosome 13q22-31 Conferring Susceptibility to Schizophrenia.

We report a consanguineous family in which schizophrenia segregates in a manner consistent with recessive inheritance of a rare, partial-penetrance susceptibility allele. From 4 marriages between 2 sets of siblings who are half first cousins, 6 offspring have diagnoses of psychotic disorder. Homozygosity mapping revealed a 6.1-Mb homozygous region on chromosome 13q22.2-31.1 shared by all affected individuals, containing 13 protein-coding genes. Microsatellite analysis confirmed homozygosity for the affected haplotype in 12 further apparently unaffected members of the family. Psychiatric reports suggested an endophenotype of milder psychiatric illness in 4 of these individuals. Exome and genome sequencing revealed no potentially pathogenic coding or structural variants within the risk haplotype. Filtering for noncoding variants with a minor allele frequency of <0.05 identified 17 variants predicted to have significant effects, the 2 most significant being within or adjacent to the SCEL gene. RNA sequencing of blood from an affected homozygote showed the upregulation of transcription from NDFIP2 and SCEL. NDFIP2 is highly expressed in brain, unlike SCEL, and is involved in determining T helper (Th) cell type 1 and Th2 phenotypes, which have previously been implicated with schizophrenia.

The ciliary Frizzled-like receptor Tmem67 regulates canonical Wnt/ß-catenin signalling in the developing cerebellum via Hoxb5.

Primary cilia defects result in a group of related pleiotropic malformation syndromes known as ciliopathies, often characterised by cerebellar developmental and foliation defects. Here, we describe the cerebellar anatomical and signalling defects in the Tmem67tm1(Dgen)/H knockout mouse. At mid-gestation, Tmem67 mutant cerebella were hypoplastic and had aberrantly high canonical Wnt/ß-catenin signalling, proliferation and apoptosis. Later in development, mutant cerebellar hemispheres had severe foliation defects and inferior lobe malformation, characterized by immature Purkinje cells (PCs). Early postnatal Tmem67 mutant cerebellum had disrupted ciliogenesis and reduced responsiveness to Shh signalling. Transcriptome profiling of Tmem67 mutant cerebella identified ectopic increased expression of homeobox-type transcription factors (Hoxa5, Hoxa4, Hoxb5 and Hoxd3), normally required for early rostral hindbrain patterning. HOXB5 protein levels were increased in the inferior lobe, and increased canonical Wnt signalling, following loss of TMEM67, was dependent on HOXB5. HOXB5 occupancy at the ß-catenin promoter was significantly increased by activation of canonical Wnt signalling in Tmem67-/- mutant cerebellar neurones, suggesting that increased canonical Wnt signalling following mutation or loss of TMEM67 was directly dependent on HOXB5. Our results link dysregulated expression of Hox group genes with ciliary Wnt signalling defects in the developing cerebellum, providing new mechanistic insights into ciliopathy cerebellar hypoplasia phenotypes.

LHFPL5 mutation: A rare cause of non-syndromic autosomal recessive hearing loss.

Hearing loss is a debilitating disorder that impairs language acquisition, resulting in disability in children and potential isolation in adulthood. Its onset can have a genetic basis, though environmental factors, which are often preventable, can also cause the condition. The genetic forms are highly heterogeneous, and early detection is necessary to arrange appropriate patient support. Here we report the molecular basis of hereditary hearing loss in a consanguineous family with multiple affected members from Oman. Combining homozygosity mapping with whole exome sequencing identified a novel homozygous nucleotide substitution c.575T > C in the lipoma HMGIC fusion partner-like 5 gene (LHFPL5), that converted the 192nd amino acid residue in the protein from a leucine to a proline, p.(Leu192Pro). Sanger sequencing confirmed segregation with the disease phenotype as expected for a recessive condition and the variant was absent in 123,490 subjects from various disease-specific and population genetic studies as well as 150 unrelated individuals and 35 deaf patients of Omani ethnicity. This study, which describes a novel LHFPL5 mutation in a family of Omani origin with hereditary hearing loss, supports previous clinical descriptions of the condition and contributes to the genetic spectrum of mutations in this form of deafness.

Identification of Inherited Retinal Disease-Associated Genetic Variants in 11 Candidate Genes.

Inherited retinal diseases (IRDs) display an enormous genetic heterogeneity. Whole exome sequencing (WES) recently identified genes that were mutated in a small proportion of IRD cases. Consequently, finding a second case or family carrying pathogenic variants in the same candidate gene often is challenging. In this study, we searched for novel candidate IRD gene-associated variants in isolated IRD families, assessed their causality, and searched for novel genotype-phenotype correlations. Whole exome sequencing was performed in 11 probands affected with IRDs. Homozygosity mapping data was available for five cases. Variants with minor allele frequencies ≤ 0.5% in public databases were selected as candidate disease-causing variants. These variants were ranked based on their: (a) presence in a gene that was previously implicated in IRD; (b) minor allele frequency in the Exome Aggregation Consortium database (ExAC); (c) in silico pathogenicity assessment using the combined annotation dependent depletion (CADD) score; and (d) interaction of the corresponding protein with known IRD-associated proteins. Twelve unique variants were found in 11 different genes in 11 IRD probands. Novel autosomal recessive and dominant inheritance patterns were found for variants in Small Nuclear Ribonucleoprotein U5 Subunit 200 (SNRNP200) and Zinc Finger Protein 513 (ZNF513), respectively. Using our pathogenicity assessment, a variant in DEAH-Box Helicase 32 (DHX32) was the top ranked novel candidate gene to be associated with IRDs, followed by eight medium and lower ranked candidate genes. The identification of candidate disease-associated sequence variants in 11 single families underscores the notion that the previously identified IRD-associated genes collectively carry > 90% of the defects implicated in IRDs. To identify multiple patients or families with variants in the same gene and thereby provide extra proof for pathogenicity, worldwide data sharing is needed.

Specific Alleles of CLN7/MFSD8, a Protein That Localizes to Photoreceptor Synaptic Terminals, Cause a Spectrum of Nonsyndromic Retinal Dystrophy.

Purpose: Recessive mutations in CLN7/MFSD8 usually cause variant late-infantile onset neuronal ceroid lipofuscinosis (vLINCL), a poorly understood neurodegenerative condition, though mutations may also cause nonsyndromic maculopathy. A series of 12 patients with nonsyndromic retinopathy due to novel CLN7/MFSD8 mutation combinations were investigated in this study. Methods: Affected patients and their family members were recruited in ophthalmic clinics at each center where they were examined by retinal imaging and detailed electrophysiology. Whole exome or genome next generation sequencing was performed on genomic DNA from at least one affected family member. Immunofluorescence confocal microscopy of murine retina cross-sections were used to localize the protein. Results: Compound heterozygous alleles were identified in six cases, one of which was always p.Glu336Gln. Such combinations resulted in isolated macular disease. Six further cases were homozygous for the variant p.Met454Thr, identified as a founder mutation of South Asian origin. Those patients had widespread generalized retinal disease, characterized by electroretinography as a rod-cone dystrophy with severe macular involvement. In addition, the photopic single flash electroretinograms demonstrated a reduced b- to a-wave amplitude ratio, suggesting dysfunction occurring after phototransduction. Immunohistology identified MFSD8 in the outer plexiform layer of the retina, a site rich in photoreceptor synapses. Conclusions: This study highlights a hierarchy of MFSD8 variant severity, predicting three consequences of mutation: (1) nonsyndromic localized maculopathy, (2) nonsyndromic widespread retinopathy, or (3) syndromic neurological disease. The data also shed light on the underlying pathogenesis by implicating the photoreceptor synaptic terminals as the major site of retinal disease.

Disease Expression in Autosomal Recessive Retinal Dystrophy Associated With Mutations in the DRAM2 Gene.

PURPOSE: To determine the disease course of retinal dystrophy caused by recessive variants in the DRAM2 (damage-regulated autophagy modulator 2) gene. METHODS: Sixteen individuals with DRAM2-retinopathy were examined (six families; age range, 19-56 years, includes one pre-symptomatic case). The change in visual acuity over time was studied, and electrophysiology (n = 6), retina-tracking perimetry (n = 1), fundus autofluorescence (FAF) imaging (n = 6), and optical coherence tomography (OCT; n = 12) were performed. RESULTS: All symptomatic patients presented with central visual loss (15/15) unaccompanied either by nyctalopia or light-hypersensitivity; most (11/15) developed symptoms in the third decade of life. A granular macular appearance, often with associated white/yellow dots, was an early fundoscopic feature. There was an ill-defined ring of hyperautofluorescence on FAF. Optical coherence tomography revealed loss of the ellipsoid zone perifoveally in a 19-year-old pre-symptomatic individual. The central atrophic area enlarged over time and fundoscopy showed peripheral degeneration in seven of the nine individuals that were examined ≥ 10 years after becoming symptomatic; some of these subjects developed nyctalopia and light hypersensitivity. Electrophysiology revealed generalized retinal dysfunction in three of the five individuals that were tested ≥ 10 years after becoming symptomatic. CONCLUSIONS: Patients with DRAM2-retinopathy are typically asymptomatic in the first two decades of life and present with central visual loss and a maculopathy. A faint hyperautofluorescent ring on FAF can be a suggestive feature. The retinal periphery is frequently affected later in the disease process. Photoreceptor degeneration is likely to be the primary event and future studies on DRAM2-retinopathy are expected to provide important insights into retinal autophagy.