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The finding of a genotype-negative hypertrophic cardiomyopathy (HCM) pedigree with several affected members indicating a familial origin of the disease has driven this study to discover causative gene variants. Genetic testing of the proband and subsequent family screening revealed the presence of a rare variant in the MYBPC3 gene, c.3331-26T>G in intron 30, with evidence supporting cosegregation with the disease in the family. An analysis of potential splice-altering activity using several splicing algorithms consistently yielded low scores. Minigene expression analysis at the mRNA and protein levels revealed that c.3331-26T>G is a spliceogenic variant with major splice-altering activity leading to undetectable levels of properly spliced transcripts or the corresponding protein. Minigene and patient mRNA analyses indicated that this variant induces complete and partial retention of intron 30, which was expected to lead to haploinsufficiency in carrier patients. As most spliceogenic MYBPC3 variants, c.3331-26T>G appears to be non-recurrent, since it was identified in only two additional unrelated probands in our large HCM cohort. In fact, the frequency analysis of 46 known splice-altering MYBPC3 intronic nucleotide substitutions in our HCM cohort revealed 9 recurrent and 16 non-recurrent variants present in a few probands (≤ 4), while 21 were not detected. The identification of non-recurrent elusive MYBPC3 spliceogenic variants that escape detection by in silico algorithms represents a challenge for genetic diagnosis of HCM and contributes to solving a fraction of genotype-negative HCM cases., (© 2022. The Author(s).)

Missense and frameshift pathogenic variants and microdeletions involving TBL1XR1 gene have been described in patients with intellectual disability, autism, Rett-like features and schizophrenia, some of them with the clinical diagnosis of Pierpont syndrome, a rare pattern of multiple congenital anomalies, but others without dysmorphic findings or with non-specific ones, and also patients with only some of the features associated with Pierpont syndrome. We here present a case with a de novo novel missense variant in TBL1XR1 gene with overlapping features with Pierpont syndrome and autism, a neurobehavioral manifestation not previously reported in Pierpont syndrome. This patient expands the phenotypic spectrum of TBL1XR1 gene pathogenic variants., (© 2021 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Usher syndrome is a rare disorder causing retinitis pigmentosa, together with sensorineural hearing loss. Due to the phenotypic and genetic heterogeneity of this disease, the best method to screen the causative mutations is by high-throughput sequencing. In this study, we tested a semiconductor chip based sequencing approach with 77 unrelated patients, as a molecular diagnosis routine. In addition, Multiplex Ligation-dependent Probe Amplification and microarray-based Comparative Genomic Hybridization techniques were applied to detect large rearrangements, and minigene assays were performed to confirm the mRNA processing aberrations caused by splice-site mutations. The designed panel included all the USH causative genes (MYO7A, USH1C, CDH23, PCDH15, USH1G, CIB2, USH2A, ADGRV1, WHRN and CLRN1) as well as four uncertainly associated genes (HARS, PDZD7, CEP250 and C2orf71). The outcome showed an overall mutation detection ratio of 82.8% and allowed the identification of 42 novel putatively pathogenic mutations. Furthermore, we detected two novel nonsense mutations in CEP250 in a patient with a disease mimicking Usher syndrome that associates visual impairment due to cone-rod dystrophy and progressive hearing loss. Therefore, this approach proved reliable results for the molecular diagnosis of the disease and also allowed the consolidation of the CEP250 gene as disease causative for an Usher-like phenotype.

Partial duplications involving the long arm of the X chromosome are associated with mental retardation, short stature, microcephaly, hypopituitarism and a wide range of physical findings. We identified an inherited Xq26.2-Xq26.3 duplication in two brothers with severe mental retardation, hypotonia, growth delay, craniofacial disproportion and dental malocclusion. Chromosome analysis was normal and multiplex ligation-dependent probe amplification analysis detected duplication on Xq26. Further characterization by array comparative genomic hybridization and quantitative PCR helped to determine proximal and distal duplication breakpoints giving a size of approximately 2.8 Mb. The duplication encompasses 24 known genes, including the X-linked mental retardation genes ARHGEF6, PHF6, HPRT1 and SLC9A6. Clinical and molecular characterization of Xq duplications will shed more light into the phenotypic implication of functional disomy of X-chromosome genes.

Spinal muscular atrophy (SMA) is caused by mutations in the SMN1 gene. We have studied the molecular pathology of SMA in 745 unrelated Spanish patients using PCR-RFLP, SMN gene dosage analysis, linkage studies, long-range PCR and direct sequencing. Our systematic approach allowed us to complete genetic testing and risk assessment in 736 SMA patients (98.8%). Females were more frequently affected by the acute form of the disease (type I), whereas chronic forms (type II-III) predominated in males (p<0.008). Absence of the SMN1 gene was detected in 671 patients (90%), and hybrid SMN1-SMN2 genes were observed in 37 cases (5%). Furthermore, we detected 13 small mutations in 28 patients (3.8%), four of which were previously identified in other populations (c.91dupT; c.770&#95;780dup11; p.Tyr272Cys and p.Thr274Ile), while five mutations were found to date only in Spanish patients (c.399&#95;402delAGAG, p.Ile116Phe, p.Gln136Glu, c.740dupC and c.834+2T>G). The c.399&#95;402delAGAG mutation accounted for 1.9% of all Spanish SMA patients. Finally, we discovered four novel mutations: c.312dupA, c.411delT, p.Trp190X and p.Met263Thr. Our results confirm that most SMA cases are due to large genetic rearrangements in the repetitive region of the SMA locus, resulting in absence-dysfunction of the SMN1 gene. By contrast, ancestrally inherited small mutations are responsible for only a small number of cases. Four prevalent changes in exons 3 and 6 (c.399&#95;402delAGAG; c.770&#95;780dup11; p.Tyr272Cys; p.Thr274Ile) accounted for almost 70% of our patients with these subtle mutations. An SMN-SMN dimer model featuring tight hydrophobic-aromatic interactions is proposed to explain the impact of mutations at the C-terminal end of the protein.

Ataxia with isolated vitamin E deficiency is a rare autosomal recessive neurodegenerative disease due to mutations in the alpha-tocopherol transfer protein gene. In ataxia with isolated vitamin E deficiency, the biochemical hallmark is the low plasmatic levels of vitamin E and, in most of the patients, vitamin E supplementation allows a stabilization of the neurologic conditions. We have investigated the genetic cause of ataxia and reduced levels of vitamin E, and apolipoproteins A1 and B in a 16-y-old patient. Results revealed that our propositus is a compound heterozygote for the c.227&#95;229delinsATT/c.744delA mutations in the alpha-tocopherol transfer protein gene, each inherited from one of the two parents. His sister is also a compound heterozygote for both mutations, and she presents a biochemical pattern similar to that of his brother. After receiving the vitamin E supplementation, plasmatic levels of vitamin E and apolipoprotein A1 have been normalized in the propositus. The detected mutations would justify the undetectable levels of vitamin E, but would not explain the also decreased levels of the apolipoproteins, as neither that after treatment with vitamin E, the levels of apolipoprotein B do not become normal. These findings suggest that other genes may play a role in producing this atypical biochemical profile.

Purpose: Routine protocols for the study of mental retardation include karyotype, analysis for fragile X syndrome, and subtelomeric rearrangements. Nevertheless, detection of cryptic rearrangements requires more sensitive techniques. Mutation screening in all known genes responsible for X-linked mental retardation is not feasible, and linkage analysis is sometimes limited. Multiplex ligation probe amplification is a recently developed technique based on the amplification of specific probes that allows relative quantification of 40 to 46 different target DNA sequences in a single reaction., Methods: In the present study, we assessed multiplex ligation probe amplification for the detection of microduplications/microdeletions in 80 male patients with suspicion of X-linked mental retardation., Results: We detected four copy number aberrations (5%): three duplications (GDI1, RPS6KA3, and ARHGEF6) and one deletion (OPHN1). All these changes were confirmed by other molecular techniques, and patients were clinically re-evaluated., Conclusions: We strongly recommend the use of multiplex ligation probe amplification as a first screening method for the detection of copy number aberrations in patients with mental retardation because of its cost-effectiveness.

Adult dominant polycystic kidney disease is an hereditary condition responsible for 6% of end-stage renal failure in Spain. Two genes were located in chromosomes 16 and 4 as related to this age-dependent disease in the 90s (PKD1 and PKD2). The diagnosis can be easily achieved by sonographic study, but molecular analysis by means of linkage analysis has the advantage of an early diagnosis in asymptomatic genetic carriers, with a view to the preventive follow-up of these subjects and genetic counselling. In this paper we present the results of molecular analysis of 30 families with Adult Dominant Polycystic Kidney Disease (from the province of Las Palmas Spain), carried out linkage analysis with two series of microsatellite markers located within or in the vicinity ofPKD1 (D16S521, KG8, AC2.5, CW2, SM7) and PKD2 (D4S1538, D4S1534, D4S423,D4S414) genes. The objectives of the study were: first, to verify the informativeness, and therefore, the usefulness of these markers for family studies in our population; and second,to assess the sensitivity and specificity of the genetic analysis in our population. Most of the markers showed a high heterozygosity, comparable to data in other studies. Considering the alleles of the different markers together in a chromosome as an haplotype increased the informativeness of the markers, and allowed the unequivocal identification of genetic data in 97.7% of patients and 88.7% of healthy subjects. The sensitivity and specificity of the genetic analysis were 90.7% (CI 95%: 85.7-95.7) and 86.8% (CI 95%: 80.6-93.0), respectively.

Mutations in the Connexin-26 gene are responsible for up to 60% of nonsyndromic, neurosensory autosomal recessive deafness (NSRD). Amongst all the mutations described to date, 35delG (a deletion of a G in a tract of five Gs at positions 30-35) is the most common and has been found in virtually all of the populations studied. Because its frequency varies in different populations, a rapid and simple method of detection of this mutation would be very helpful in population studies. A wide variety of methods for this detection have been described, but we herein present a very simple method using a PCR with primers designed to provide an amplicon of 94 or 93 nucleotides for the normal or mutant alleles, respectively, that can be easily distinguished in an 8% polyacrylamide gel. The entire protocol can be completed in a morning, thus supporting multiple runs. This assay will be useful in screening the large sample sizes required for population studies.

Case Reports: We report the cases of four males from four different families, who presented paroxysmal episodes from the 1st 2nd year. These episodes were characterised by asymmetrical bilateral dystonia of the upper limbs, predominantly in both hands, and were associated with orofacial dyskinesias, stereotipies (jumping, arm flapping, etc.), facial tics and, occasionally, phonic tics. Consciousness is not affected in any of the cases. These movements are triggered in situations where the patient is relaxed or excited. They occur daily and last from a few seconds to 30 minutes. Between the bouts, they remain asymptomatic. Family cases suggest it is inherited by autosomal dominant transmission, perhaps linked to the X chromosome; in addition, two cases are sporadic. In the only adult, the movements progress to a series of rhythmic bilateral dystonic myoclonias and facial tics dyskinesias. All the studies carried out, EEG, hemogram, biochemical analysis, neuroimaging, copper and ceruloplasmin levels, were normal., Conclusions: 1. We report a non epileptic paroxysmal disorder originating in the extrapyramidal tracts with its own characteristics, with onset during early childhood, which is associated with stereotipies, tics and dystonia; 2. It occurs predominantly in males; 3. It is inherited by autosomal dominant transmission, or perhaps sex linked autosomal dominant inheritance, and there are also sporadic cases; 4. The range of clinical features is very wide and includes cases in which there are few symptoms to others where the extent and gravity of the disorder is very significant.

Introduction and Clinical Cases: The Pitt Rogers Danks syndrome is characterized by prenatal and postnatal retardation of growth, mental retardation, microcephaly, convulsions and a peculiar facies. It is believed to represent a clinical variant of the Wolf Hirschhorn syndrome, since there is a deletion in the 4p16.3 region in both syndromes. We report two cases in the same family caused by maternal mal segregation of a 4:8 balanced translocation. We describe the clinical characteristics, investigations done and a review of the literature.

Molecular screening programs in mentally retarded individuals have been performed in several populations worldwide. One finding has been an excess of FMR1 intermediate alleles in a population with learning difficulties. However, other published reports with similar characteristics did not corroborate those previous results. In order to contribute additional data from our population, we studied 563 patients affected with nonspecific mental retardation (MRX) that did not present a CGG expansion in the FMR1 gene and 208 individuals as a control population. Forty MRX patients presented alleles within the intermediate range. Among them, one case showed a pattern of expression of the FMR1 protein (FMRP) concordant with a fragile X syndrome case with an intermediate allele/full mutation mosaicism, although it was not detected by Southern blot analysis. Statistical analysis was performed again showing no statistically significant difference regarding the intermediate allele frequency in the MRX and control populations. This finding is in agreement with the hypothesis that the incidence of intermediate FMR1 alleles in MRX populations does not seem to be higher than in control populations, and it emphasizes the importance of FMRP detection as a diagnostic tool for fragile X syndrome.

Background: TBL1XR1 encodes a F-box-like/WD40 repeat-containing protein that plays a role in transcription mediated by nuclear receptors and is a known genetic cause of neurodevelopmental disease of childhood (OMIM# 608628). Yet the developmental trajectory and progression of neurologic symptoms over time remains poorly understood. Methods: We developed and distributed a survey to two closed Facebook groups devoted to families of patients with TBL1XR1-related disorder. The survey consisted of 14 subsections focused upon the developmental trajectories of cognitive, behavioral, motor, and other neurological abnormalities. Data were collected and managed using REDCap electronic data capture tools. Results: Caregivers of 41 patients with a TBL1XR1-related disorder completed the cross-sectional survey. All reported variants affecting a single amino acid, including missense mutations and in-frame deletions, were found in the WD40 repeat regions of Tbl1xr1. These are domains considered important for protein–protein interactions that may plausibly underlie disease pathology. The majority of patients were diagnosed with a neurologic condition before they received their genetic diagnosis. Language appeared most significantly affected with only a minority of the cohort achieving more advanced milestones in this domain. Conclusion: TBL1XR1-related disorder encompasses a spectrum of clinical presentations, marked by early developmental delay ranging in severity, with a subset of patients experiencing developmental regression in later childhood. [ABSTRACT FROM AUTHOR]

Focuses on the R608del mutation in the acid sphingomyelinase gene (SMPDI) which is most prevalent among patients from Gran Canaria Island with Niemann-Pick disease type B. Causes of Niemann-Pick type A and B diseases; Description of the R608del mutation; Clinical, biochemical and genetical findings in people living in Canaria island.

Background: Spinal muscular atrophy (SMA) is an autosomal recessive disease caused by mutations and deletions in SMN1 at exon 7. The carrier frequency for SMN1 mutations ranges from 2 to 4% in the general population. Methods: We examined allelic, genotypic relatedness and copy number (CN) variations and frequencies of SMN1 and SMN2, in 13,426 samples from Qatar biobank (QBB) to provide a precise estimation of SMA carrier frequency in Qatar in comparison to other populations. Results: The SMA carrier frequency was found to be (2.8%) and the rs143838139 was found in 491/13426 (3.66%) of individuals. The SNP rs121909192, which is a pathogenic risk factor, was found in 321/13500 (2.38%). In Addition 242/11379 (2.13%) had two copies of SMN1 and the rs143838139, which may explain the (2 + 0) silent carrier. Additionally, two participants were found to be SMA type 4 with 0 and 4 copy numbers in SMN1 and SMN2, respectively. Conclusion: The SMA carrier frequency in Qatar was found to be comparable to Saudi Arabia and Caucasians. The likely pathogenic variant, rs121909192, was found to be significantly higher when compering with other in our study. The rs143838139 variant, which has a strong association with the silent carrier genotype, has been found. Consequently, testing for this SNP may enhance the precision of evaluating the likelihood of a patient having an affected child. We conclude that the frequency of SMA carriers varies within the Qatar population and other ethnic groups. [ABSTRACT FROM AUTHOR]

Purpose. Therapeutic advances in the treatment of spinal muscular atrophy (SMA) prompt the need for robust and efficient molecular diagnosis of this disease. Approximately five percent of SMA cases are attributable to one copy of SMN1 with a hypomorphic or inactivating variant in trans with a deleted or converted allele. These intragenic variants are challenging to definitively localize to SMN1 due to its sequence homology with the SMN2 gene. To enhance the clinical sensitivity of SMA diagnostic testing, we present an optimized gene-specific sequencing assay to localize variants to either SMN1 or SMN2. Methods. SMN1 and SMN2 genes are independently amplified by long-range allele-specific PCR. Long-range products are used in subsequent nested PCR reactions to amplify the coding exons of SMN1 and SMN2. The resulting products are sequenced using standard Sanger-based methodologies and analyzed for disease-associated alterations. Results. 83 probands suspicious for a clinical diagnosis of SMA with a nondiagnostic SMN dosage result were sequenced for intragenic variants in the SMN1 gene. Gene-specific sequencing revealed likely disease-associated variants in SMN1 in 42 cases (50.6%). Of the 42 variants, 27 are unique including 16 loss-of-function variants, 9 missense variants, 1 in-frame deletion variant, and 1 splice site variant. Conclusions. Herein, we describe an optimized assay for clinical sequencing of the full coding region of SMN1 and SMN2. This assay uses standard techniques and equipment readily available to most molecular diagnostic laboratories. [ABSTRACT FROM AUTHOR]

Autism spectrum disorder (ASD) is among the most widespread neurodevelopmental diseases, with an approximate prevalence rate of 1 in 59. From a genetic point of view, this disorder is highly heterogeneous. This disorder is associated with both inheritable and de novo mutations in several genes. In addition to genetic loci that are identified through early karyotype analyses, recent advent of high throughput sequencing methods has facilitated identification of several genetic loci that confer risk of ASD. The current review provides an overview of different types of identified mutations including missense and nonsense mutations and copy number variations in various genes in individuals affected with ASD. [ABSTRACT FROM AUTHOR]

Acid sphingomyelinase deficiency (ASMD) or Niemann–Pick disease type A (NPA), type B (NPB) and type A/B (NPA/B), is a rare lysosomal storage disease characterized by progressive accumulation of sphingomyelin (SM) in the liver, lungs, bone marrow and, in severe cases, neurons. A disease model was established by generating liver organoids from a NPB patient carrying the p.Arg610del variant in the SMPD1 gene. Liver organoids were characterized by transcriptomic and lipidomic analysis. We observed altered lipid homeostasis in the patient-derived organoids showing the predictable increase in sphingomyelin (SM), together with cholesterol esters (CE) and triacylglycerides (TAG), and a reduction in phosphatidylcholine (PC) and cardiolipins (CL). Analysis of lysosomal gene expression pointed to 24 downregulated genes, including SMPD1, and 26 upregulated genes that reflect the lysosomal stress typical of the disease. Altered genes revealed reduced expression of enzymes that could be involved in the accumulation in the hepatocytes of sphyngoglycolipids and glycoproteins, as well as upregulated genes coding for different glycosidases and cathepsins. Lipidic and transcriptome changes support the use of hepatic organoids as ideal models for ASMD investigation. [ABSTRACT FROM AUTHOR]

For urinalysis teaching and routine clinical imaging of urine, urine microscopic test needs slides that can be clearly examined for long term, which is still under focus. This study hypothesized, that poly(vinyl chloride) (PVC) and polystyrene (PS) could be able to reserve the urine components for a long time. Therefore, we explored, for the first time upon our knowledge, the potential of PVC and PS polymers to adhere the cover slide. The physical properties, viscosity, average molecular weight, the tensile strength and intrinsic viscosity of the polymers were determined. A normal slide (8 × 3 cm × 1 mm) was used to test a urine sample in the presence of either 3% PVC in tetra hydro‐furan(THF) or 3% PS. The results of imaging the PVC and PS slide indicate a clear and transparent vision. The PVC urine slide maintains normal components of urine up to a year, which did not observe in the PS slide. These results suggest that PVC is an effective polymer to protect urine compositions from hydrolyses or decomposition, which makes this novel urine slide a successful candidate for long‐term teaching of urinalyses. [ABSTRACT FROM AUTHOR]

Hypertrophic cardiomyopathy (HCM) is the most prevalent genetically inherited cardiomyopathy that follows an autosomal dominant inheritance pattern. The majority of HCM cases can be attributed to mutation of the MYBPC3 gene, which encodes cMyBP-C, a crucial structural protein of the cardiac muscle. The manifestation of HCM's morphological, histological, and clinical symptoms is subject to the complex interplay of various determinants, including genetic mutation and environmental factors. Approximately half of MYBPC3 mutations give rise to truncated protein products, while the remaining mutations cause insertion/deletion, frameshift, or missense mutations of single amino acids. In addition, the onset of HCM may be attributed to disturbances in the protein and transcript quality control systems, namely, the ubiquitin–proteasome system and nonsense-mediated RNA dysfunctions. The aforementioned genetic modifications, which appear to be associated with unfavorable lifelong outcomes and are largely influenced by the type of mutation, exhibit a unique array of clinical manifestations ranging from asymptomatic to arrhythmic syncope and even sudden cardiac death. Although the current understanding of the MYBPC3 mutation does not comprehensively explain the varied phenotypic manifestations witnessed in patients with HCM, patients with pathogenic MYBPC3 mutations can exhibit an array of clinical manifestations ranging from asymptomatic to advanced heart failure and sudden cardiac death, leading to a higher rate of adverse clinical outcomes. This review focuses on MYBPC3 mutation and its characteristics as a prognostic determinant for disease onset and related clinical consequences in HCM. [ABSTRACT FROM AUTHOR]

TBL1XR1, which encodes transducing β‐like 1 X‐linked receptor 1, is implicated in both Pierpont syndrome and intellectual developmental disorder, autosomal dominant‐41 (MRD‐41, OMIM #616944). While both conditions are autosomal dominant, variants associated with Pierpont syndrome are believed to behave in a dominant negative fashion, whereas those causing MRD‐41 result in haploinsufficiency. Here, we present a patient with a de novo novel variant in TBL1XR1 (c.977G > A,p.S326N) identified by trio exome sequencing. Though a different variant at this same residue has previously been associated with MRD‐41, our patient's presentation is suggestive of Pierpont syndrome. The patient's clinical phenotype, which includes short stature, developmental delay, dysmorphic craniofacial features, and plantar fat pads, more closely resembles that of known patients with Pierpont syndrome than MRD‐41. Furthermore, this missense variant is directly adjacent to one previously associated with Pierpont syndrome and exists in the same region as all variants associated with Pierpont, on the inner surface of a WD40 ring. We propose this variant is a newly identified cause of Pierpont syndrome. [ABSTRACT FROM AUTHOR]

Background: Acid Sphingomyelinase Deficiency (ASMD) is a rare autosomal recessive disorder caused by mutations in the SMPD1 gene. This rarity contributes to misdiagnosis, delayed diagnosis and barriers to good care. There are no published national or international consensus guidelines for the diagnosis and management of patients with ASMD. For these reasons, we have developed clinical guidelines that defines standard of care for ASMD patients. Methods: The information contained in these guidelines was obtained through a systematic literature review and the experiences of the authors in their care of patients with ASMD. We adopted the Appraisal of Guidelines for Research and Evaluation (AGREE II) system as method of choice for the guideline development process. Results: The clinical spectrum of ASMD, although a continuum, varies substantially with subtypes ranging from a fatal infantile neurovisceral disorder to an adult-onset chronic visceral disease. We produced 39 conclusive statements and scored them according to level of evidence, strengths of recommendations and expert opinions. In addition, these guidelines have identified knowledge gaps that must be filled by future research. Conclusion: These guidelines can inform care providers, care funders, patients and their carers about best clinical practice and leads to a step change in the quality of care for patients with ASMD with or without enzyme replacement therapy (ERT). [ABSTRACT FROM AUTHOR]

Background: Acid sphingomyelinase deficiency (ASMD) disorder, also known as Niemann–Pick disease (NPD) is a rare genetic disease caused by mutations in SMPD1 gene, which encodes sphingomyelin phosphodiesterase (ASM). Except for liver and spleen enlargement and lung disease, two subtypes (Type A and B) of NDP have different onset times, survival times, ASM activities, and neurological abnormalities. To comprehensively explore NPD's genotype-phenotype association and pathophysiological characteristics, we collected 144 NPD cases with strict quality control through literature mining. Results: The difference in ASM activity can differentiate NPD type A from other subtypes, with the ratio of ASM activity to the reference values being lower in type A (threshold 0.045 (4.45%)). Severe variations, such as deletion and insertion, can cause complete loss of ASM function, leading to type A, whereas relatively mild missense mutations generally result in type B. Among reported mutations, the p.Arg3AlafsX76 mutation is highly prevalent in the Chinese population, and the p.R608del mutation is common in Mediterranean countries. The expression profiles of SMPD1 from GTEx and single-cell RNA sequencing data of multiple fetal tissues showed that high expressions of SMPD1 can be observed in the liver, spleen, and brain tissues of adults and hepatoblasts, hematopoietic stem cells, STC2_TLX1-positive cells, mesothelial cells of the spleen, vascular endothelial cells of the cerebellum and the cerebrum of fetuses, indicating that SMPD1 dysfunction is highly likely to have a significant effect on the function of those cell types during development and the clinicians need pay attention to these organs or tissues as well during diagnosis. In addition, we also predicted 21 new pathogenic mutations in the SMPD1 gene that potentially cause the NPD, signifying that more rare cases will be detected with those mutations in SMPD1. Finally, we also analysed the function of the NPD type A cells following the extracellular milieu. Conclusions: Our study is the first to elucidate the effects of SMPD1 mutation on cell types and at the tissue level, which provides new insights into the genotype-phenotype association and can help in the precise diagnosis of NPD. [ABSTRACT FROM AUTHOR]

Background: Obsessive–Compulsive Disorder (OCD) is a common and chronic psychiatric disorder with significant morbidity characterized by intrusive, uncontrollable and reoccurring thoughts (i.e., obsessions) and/or ritualistic behaviours (i.e., compulsions). Conradi-Hünerman-Happle Syndrome (CHHS) is a rare inherited X-linked dominant variant of chondrodysplasia punctata, a heterogeneous group of rare bone dysplasias characterized by punctate epiphyseal calcifications of complex etiology and pathophysiology that remain to be defined. Available literature reveals a lacuna in regards to the coexistence of the entities with no clinical reports described. Case presentation: A 12 year old female patient with diagnosis of CHHS, presents to psychiatric consultation due to aggravation of her OCD clinical picture, with aggravation of hand-washing frequency during the Covid-19 pandemic with significant functional impact. Psychopharmacological treatment aimed at OCD with Selective Serotonin Reuptake Inhibitor (SSRI) and antipsychotic was instituted with favourable, albeit partial response. Conclusions: The authors aim to describe a clinical case in which the patient presents with Conradi-Hünerman-Happle Syndrome and Obsessive–Compulsive Disorder. Clinical descriptions of CHHS and OCD are not available in the literature. Through this case description the authors aim to present a rare case as well as discuss an eventual association between etiology and/or pathophysiology of the two disorders. [ABSTRACT FROM AUTHOR]

Increasing evidence suggests that both coding and non-coding regions of sarcomeric protein genes can contribute to hypertrophic cardiomyopathy (HCM). Here, we introduce an experimental workflow (tested on four patients) for complete sequencing of the most common HCM genes (MYBPC3, MYH7, TPM1, TNNT2, and TNNI3) via long-range PCR, Oxford Nanopore Technology (ONT) sequencing, and bioinformatic analysis. We applied Illumina and Sanger sequencing to validate the results, FastQC, Qualimap, and MultiQC for quality evaluations, MiniMap2 to align data, Clair3 to call and phase variants, and Annovar's tools and CADD to assess pathogenicity of variants. We could not amplify the region encompassing exons 6–12 of MYBPC3. A higher sequencing error rate was observed with ONT (6.86–6.92%) than with Illumina technology (1.14–1.35%), mostly for small indels. Pathogenic variant p.Gln1233Ter and benign polymorphism p.Arg326Gln in MYBPC3 in a heterozygous state were found in one patient. We demonstrated the ability of ONT to phase single-nucleotide variants, enabling direct haplotype determination for genes TNNT2 and TPM1. These findings highlight the importance of long-range PCR efficiency, as well as lower accuracy of variant calling by ONT than by Illumina technology; these differences should be clarified prior to clinical application of the ONT method. [ABSTRACT FROM AUTHOR]

The purpose of this study was to expand the mutation spectrum by searching the causative mutations in nine Lebanese families with Usher syndrome (USH) using whole-exome sequencing. The pathogenicity of candidate mutations was first evaluated according to their frequency, conservation, and in silico prediction tools. Then, it was confirmed via Sanger sequencing, followed by segregation analysis. Finally, a meta-analysis was conducted to calculate the prevalence of USH genes in the Lebanese population. Three missense mutations, two splice site mutations, and one insertion/deletion were detected in eight of the families. Four of these variants were novel: c.5535C > A; p.(Asn1845Lys) in exon 41 of CDH23 , c.7130G > A; p.(Arg2377Gln) in exon 32 of ADGRV1 , c.11390-1G > A in USH2A , and c.3999–6A > G in PCDH15. All the identified mutations were shown to be likely disease-causing through our bioinformatics analysis and co-segregated with the USH phenotype. The mutations were classified according to the ACMG standards. Finally, our meta-analysis showed that the mutations in ADGRV1 , USH2A , and CLRN1 are the most prevalent and responsible for approximately 75% of USH cases in Lebanon. Of note, the frequency USH type 3 showed a relatively high incidence (23%) compared to the worldwide prevalence, which is around 2–4%. In conclusion, our study has broadened the mutational spectrum of USH and showed a high heterogeneity of this disease in the Lebanese population. [ABSTRACT FROM AUTHOR]

Spinal muscular atrophy (SMA) is caused by bi‐allelic loss or pathogenic variants in the SMN1 gene. SMN2, the highly homologous copy of SMN1, is considered the major phenotypic modifier of the disease. Determination of SMN2 copy number is essential to establish robust genotype–phenotype correlations and predict disease evolution, to stratify patients for clinical trials, as well as to define those eligible for treatment. Discordant genotype–phenotype correlations are not uncommon in SMA, some of which are due to intragenic SMN2 variants that may influence the amount of complete SMN transcripts and, therefore, of full‐length SMN protein. Detection of these variants is crucial to predict SMA phenotypes in the present scenario of therapeutic advances and with the perspective of SMA neonatal screening and early diagnosis to start treatments. Here, we present a novel, affordable, and versatile method for complete sequencing of the SMN2 gene based on long‐range polymerase chain reaction and next‐generation sequencing. The method was validated by analyzing samples from 53 SMA patients who lack SMN1, allowing to characterize paralogous, rare variants, and single‐nucleotide polymorphisms of SMN2 as well as SMN2–SMN1 hybrid genes. The method identifies partial deletions and can be adapted to determine rare pathogenic variants in patients with at least one SMN1 copy. [ABSTRACT FROM AUTHOR]

Background: Usher syndrome (USH) is a recessive inherited disease characterized by sensorineural hearing loss, retinitis pigmentosa, and sometimes, vestibular dysfunction. Although the molecular epidemiology of Usher syndrome has been well studied in Europe and United States, there is a lack of studies in other regions like Africa or Central and South America. Methods: We designed a NGS panel that included the 10 USH causative genes (MYO7A , USH1C , CDH23 , PCDH15 , USH1G , CIB2 , USH2A , ADGRV1 , WHRN , and CLRN1), four USH associated genes (HARS , PDZD7 , CEP250 , and C2orf71), and the region comprising the deep-intronic c.7595-2144A>G mutation in USH2A. Results: NGS sequencing was performed in 11 USH patients from Cuba. All the cases were solved. We found the responsible mutations in the USH2A , ADGRV1 , CDH23 , PCDH15 , and CLRN1 genes. Four mutations have not been previously reported. Two mutations are recurrent in this study: c.619C>T (p.Arg207∗) in CLRN1 , previously reported in two unrelated Spanish families of Basque origin, and c.4488G>C (p.Gln1496His) in CDH23 , first described in a large Cuban family. Additionally, c.4488G>C has been reported two more times in the literature in two unrelated families of Spanish origin. Conclusion: Although the sample size is very small, it is tempting to speculate that the gene frequencies in Cuba are distinct from other populations mainly due to an "island effect" and genetic drift. The two recurrent mutations appear to be of Spanish origin. Further studies with a larger cohort are needed to elucidate the real genetic landscape of Usher syndrome in the Cuban population. [ABSTRACT FROM AUTHOR]

Spinal muscular atrophy (SMA) is a neuromuscular disorder caused by the mutations in survival motor neuron 1 gene (SMN1). The molecular pathology of missense mutations in SMN1 is not thoroughly investigated so far. Therefore, we collected all missense mutations in the SMN1 protein, using all possible search terms, from three databases (PubMed, PMC and Google Scholar). All missense mutations were subjected to in silico pathogenicity, conservation, and stability analysis tools. We used statistical analysis as a QC measure for validating the specificity and accuracy of these tools. PolyPhen-2 demonstrated the highest specificity and accuracy. While PolyPhen-1 showed the highest sensitivity; overall, PolyPhen2 showed better measures in comparison to other in silico tools. Three mutations (D44V, Y272C, and Y277C) were identified as the most pathogenic and destabilizing. Further, we compared the physiochemical properties of the native and the mutant amino acids and observed loss of H-bonds and aromatic stacking upon the cysteine to tyrosine substitution, which led to the loss of aromatic rings and may reduce protein stability. The three mutations were further subjected to Molecular Dynamics Simulation (MDS) analysis using GROMACS to understand the structural changes. The Y272C and Y277C mutants exhibited maximum deviation pattern from the native protein as compared to D44V mutant. Further MDS analysis predicted changes in the stability that may have been contributed due to the loss of hydrogen bonds as observed in intramolecular hydrogen bond analysis and physiochemical analysis. A loss of function/structural impact was found to be severe in the case of Y272C and Y277C mutants in comparison to D44V mutation. Correlating the results from in silico predictions, physiochemical analysis, and MDS, we were able to observe a loss of stability in all the three mutants. This combinatorial approach could serve as a platform for variant interpretation and drug design for spinal muscular dystrophy resulting from missense mutations. [ABSTRACT FROM AUTHOR]

Xq25q26 duplication syndrome has been reported in individuals with clinical features such as short stature, intellectual disability, syndromic facial appearance, small hands and feet, and genital abnormalities. The symptoms are related to critical chromosome regions including Xq26.1-26.3. In this particular syndrome, no patient with congenital heart disease was previously reported. Here, we report a 6-year-old boy with typical symptoms of Xq25q26 duplication syndrome and double outlet right ventricle (DORV) with pulmonary atresia (PA). He had the common duplicated region of Xq25q26 duplication syndrome extending to the distal region including the MOSPD1 locus. MOSPD1 regulates transforming growth factor beta (TGFβ) 2,3 and may be responsible for cardiac development including DORV. In the patient's lymphocytes, mRNA expression of TGFβ2 was lower than control, and might cause DORV as it does in TGFβ2-deficient mice. Therefore, MOSPD1 is a possible candidate gene for DORV, probably in combination with GPC3. Further studies of the combined functions of MOSPD1 and GPC3 are needed, and identification of additional patients with MOSPD1 and GPC3 duplication should be pursued. [ABSTRACT FROM AUTHOR]

Conradi-Hünermann-Happle syndrome, or X-linked dominant chondrodysplasia punctata type 2 (CDPX2), is a genodermatosis caused by mutations in EBP. While typically lethal in males, females with CDPX2 generally manifest by infancy or childhood with variable features including congenital ichthyosiform erythroderma, chondrodysplasia punctata, asymmetric shortening of the long bones, and cataracts. We present a 36-year-old female with short stature, rhizomelic and asymmetric limb shortening, severe scoliosis, a sectorial cataract, and no family history of CDPX2. Whole exome sequencing (WES) revealed a p.Arg63del mutation in EBP, and biochemical studies confirmed a diagnosis of CDPX2. Short stature in combination with ichthyosis or alopecia, cataracts, and limb shortening in an adult should prompt consideration of a diagnosis of CDPX2. As in many genetic syndromes, the hallmark features of CDPX2 in pediatric patients are not readily identifiable in adults. This demonstrates the utility of WES as a diagnostic tool in the evaluation of adults with genetic disorders. © 2015 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Background: Mutations in the acid alpha-glucosidase (GAA) gene usually lead to reduced GAA activity. In this study, we analyzed the mutations of GAA and GAA enzyme activity from one sibling suspected Pompe disease and their first-degree relatives. Materials and Methods: In this cross-sectional study, GAA enzyme activity assay was assessed using tandem mass spectrometry. Polymerase chain reaction and Sanger sequencing were performed for GAA analysis. Results: GAA enzyme activity was significantly decreased in patients compared to the normal range (P = 0.02). Two individuals showed ten alterations in the GAA sequence, in which one of them (c. 1650del G) has not been previously described in the literature. A single Guanine deletion (del-G) was detected at codon 551 in exon 12. Conclusion: According to the literature, the detected change is a novel mutation. We hypothesized that the discovered deletion in the GAA might lead to a reduced activity of the gene product. [ABSTRACT FROM AUTHOR]

Spinal muscular atrophy (SMA) is an autosomal recessive, lower motor neuron disease. Clinical heterogeneity is pervasive: three infantile (type I-III) and one adult-onset (type IV) forms are recognized. Type I SMA is the most common genetic cause of death in infancy and accounts for about 50% of all patients with SMA. Most forms of SMA are caused by mutations of the survival motor neuron ( SMN1) gene. A second gene that is 99% identical to SMN1 ( SMN2) is located in the same region. The only functionally relevant difference between the two genes identified to date is a C → T transition in exon 7 of SMN2, which determines an alternative spliced isoform that predominantly excludes exon 7. Thus, SMN2 genes do not produce sufficient full length SMN protein to prevent the onset of the disease. Since the identification of the causative mutation, biomedical research of SMA has progressed by leaps and bounds: from clues on the function of SMN protein, to the development of different models of the disease, to the identification of potential treatments, some of which are currently in human trials. The aim of this review is to elucidate the current state of knowledge, emphasizing how close we are to the solution of the puzzle that is SMA, and, more importantly, to highlight the missing pieces of this puzzle. Filling in these gaps in our knowledge will likely accelerate the development and delivery of efficient treatments for SMA patients and be a prerequisite towards achieving our final goal, the cure of SMA. © 2013 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Niemann-Pick disease ( NPD) types A and B are autosomal, recessively inherited, lysosomal storage disorders caused by deficient activity of acid sphingomyelinase (E.C. 3.1.4.12) because of mutations in the sphingomyelin phosphodiesterase-1 ( SMPD1) gene. Here, we present the molecular analysis and clinical characteristics of 15 NPD type A and B patients. Sequencing the SMDP1 gene revealed eight previously described mutations and seven novel mutations including four missense [c.682T>C (p. Cys228Arg), c.1159T>C (p. Cys387Arg), c.1474G>A (p. Gly492Ser), and c. 1795C>T (p. Leu599Phe)], one frameshift [c. 169delG (p. Ala57Leufs*20)] and two splicing (c.316+1G>T and c. 1341delG). The most frequent mutations were p.Arg610del (21%) and p. Gly247Ser (12%). Two patients homozygous for p.Arg610del and initially classified as phenotype B showed different clinical manifestations. Patients homozygous for p. Leu599Phe had phenotype B, and those homozygous for c. 1341delG or c.316+1G>T presented phenotype A. The present results provide new insight into genotype/phenotype correlations in NPD and emphasize the difficulty of classifying patients into types A and B, supporting the idea of a continuum between these two classic phenotypes. [ABSTRACT FROM AUTHOR]

Chondrodysplasia punctata (CDP) is an etiologically heterogeneous disorder characterized by the radiographic finding of stippled epiphyses (punctate calcifications). It is often accompanied by a characteristic facial appearance, known as the Binder phenotype, which is attributed to hypoplasia of the nasal cartilages; abnormal distal phalanges (brachytelephalangy) are a common component manifestation as well. We report eight patients with a Binder phenotype with or without CDP who all shared a known or suspected maternal deficiency of vitamin K. We suspect that this phenotype is probably under recognized, and we hope to increase awareness about the maternal risk factors, especially hyperemesis gravidarum, which lead to nutritional deficiency. © 2013 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Type I spinal muscular atrophy (SMA) is an autosomal recessive disorder caused by loss or mutations of the survival motor neuron 1 ( SMN1) gene. The reduction in SMN protein levels in SMA leads to degeneration and death of motor neurons. In this study, we have analyzed the nuclear reorganization of Cajal bodies, PML bodies and nucleoli in type I SMA motor neurons with homozygous deletion of exons 7 and 8 of the SMN1 gene. Western blot analysis revealed a marked reduction of SMN levels compared to the control sample. Using a neuronal dissociation procedure to perform a careful immunocytochemical and quantitative analysis of nuclear bodies, we demonstrated a severe decrease in the mean number of Cajal bodies per neuron and in the proportion of motor neurons containing these structures in type I SMA. Moreover, most Cajal bodies fail to recruit SMN and spliceosomal snRNPs, but contain the proteasome activator PA28γ, a molecular marker associated with the cellular stress response. Neuronal stress in SMA motor neurons also increases PML body number. The existence of chromatolysis and eccentric nuclei in SMA motor neurons correlates with Cajal body disruption and nucleolar relocalization of coilin, a Cajal body marker. Our results indicate that the Cajal body is a pathophysiological target in type I SMA motor neurons. They also suggest the Cajal body-dependent dysfunction of snRNP biogenesis and, therefore, pre-mRNA splicing in these neurons seems to be an essential component for SMA pathogenesis. [ABSTRACT FROM AUTHOR]

Usher syndrome is an autosomal recessive disorder characterized by congenital hearing loss combined with retinitis pigmentosa, and in some cases, vestibular areflexia. Three clinical subtypes are distinguished, and MYO7A and USH2A represent the two major causal genes involved in Usher type I, the most severe form, and type II, the most frequent form, respectively. Massively parallel sequencing was performed on a cohort of patients in the context of a molecular diagnosis to confirm clinical suspicion of Usher syndrome. We report here 231 pathogenic MYO7A and USH2A genotypes identified in 73 Usher type I and 158 Usher type II patients. Furthermore, we present the ACMG classification of the variants, which comprise all types. Among them, 68 have not been previously reported in the literature, including 12 missense and 16 splice variants. We also report a new deep intronic variant in USH2A. Despite the important number of molecular studies published on these two genes, we show that during the course of routine genetic diagnosis, undescribed variants continue to be identified at a high rate. This is particularly pertinent in the current era, where therapeutic strategies based on DNA or RNA technologies are being developed. [ABSTRACT FROM AUTHOR]

Usher syndrome (USH) is an autosomal recessive syndromic ciliopathy characterized by sensorineural hearing loss, retinitis pigmentosa and, sometimes, vestibular dysfunction. There are three clinical types depending on the severity and age of onset of the symptoms; in addition, ten genes are reported to be causative of USH, and six more related to the disease. These genes encode proteins of a diverse nature, which interact and form a dynamic protein network called the "Usher interactome". In the organ of Corti, the USH proteins are essential for the correct development and maintenance of the structure and cohesion of the stereocilia. In the retina, the USH protein network is principally located in the periciliary region of the photoreceptors, and plays an important role in the maintenance of the periciliary structure and the trafficking of molecules between the inner and the outer segments of photoreceptors. Even though some genes are clearly involved in the syndrome, others are controversial. Moreover, expression of some USH genes has been detected in other tissues, which could explain their involvement in additional mild comorbidities. In this paper, we review the genetics of Usher syndrome and the spectrum of mutations in USH genes. The aim is to identify possible mutation associations with the disease and provide an updated genotype–phenotype correlation. [ABSTRACT FROM AUTHOR]

The centrosome linker serves to hold the duplicated centrosomes together until they separate in late G2/early mitosis. Precisely how the linker is assembled remains an open question. In this study, we identify Cep44 as a novel component of the linker in human cells. Cep44 localizes to the proximal end of centrioles, including mother and daughter centrioles, and its ablation leads to loss of centrosome cohesion. Cep44 does not impinge on the stability of C-Nap1 (also known as CEP250), LRRC45 or Cep215 (also known as CDK5RAP2), and vice versa, and these proteins are independently recruited to the centrosome. Rather, Cep44 associates with rootletin and regulates its stability and localization to the centrosome. Our findings reveal a role of the previously uncharacterized protein Cep44 for centrosome cohesion and linker assembly. [ABSTRACT FROM AUTHOR]

Spinal muscular atrophy (SMA) is an autosomal recessive neurodegenerative disease and the most common genetic cause of infant mortality, affecting ∼1 in 10,000 live births. The disease is characterized by progressive symmetrical muscle weakness resulting from the degeneration and loss of anterior horn cells in the spinal cord and brain stem nuclei. The disease is classified on the basis of age of onset and clinical course. SMA is caused by mutations in the telomeric copy of the survival motor neuron 1 (SMN1) gene, but all patients retain a centromeric copy of the gene, SMN2. The homozygous absence of the SMN1 exon 7 has been observed in the majority of patients and is being utilized as a reliable and sensitive SMA diagnostic test. In the majority of cases, the disease severity correlates inversely with an increased SMN2 gene copy number. Carrier detection, in the deletion cases, relies on the accurate determination of the SMN1 gene copies. Since SMA is one of the most common lethal genetic disorders, with a carrier frequency of 1 in 40 to 1 in 60, direct carrier dosage testing has been beneficial to many families. This unit attempts to highlight the molecular genetics of SMA with a focus on the advantages and limitations of the current molecular technologies., (Copyright © 2016 John Wiley & Sons, Inc.)

Background: This study aims to provide genetic diagnoses for 30 cases of fetal skeletal dysplasia, and a molecular basis for the future prenatal diagnosis of fetal skeletal dysplasia. Methods: A total of 30 cases of fetal skeletal dysplasia detected with ultrasound between January 2014 and June 2017 were analyzed. Among these fetuses, 15 fetuses had local skeletal malformations, while 15 fetuses had short limb malformations. Samples of fetal umbilical cord blood, amniotic fluid, and/or aborted tissue were collected from all cases. Karyotyping, whole genome sequencing, and targeted next-generation sequencing of skeletal disease-related pathogenic genes were performed, as needed. Blood samples were taken from the parents for verification using Sanger sequencing. Results: Among the 30 cases of fetal skeletal dysplasia, two cases were diagnosed with trisomy 18. However, none of these cases were identified with any microdeletions or microreplications associated with skeletal dysplasia. Among the 28 chromosomally normal cases with fetal skeletal dysplasia, 21 cases were detected with mutations in genes related to skeletal diseases. Furthermore, collagen gene mutations were detected in six fetuses with short limb malformations, while heterozygous disease-causing mutations in the fibroblast growth factor receptor 3 (FGFR3) gene were detected in seven fetuses. The remaining fetuses carried mutations in other various genes, including tumor protein p63 (TP63), cholestenol delta-isomerase (EBP), cholinergic receptor nicotinic gamma subunit (CHRNG), filamin B (FLNB), and SRY-box 9 (SOX9). Three compound heterozygous mutations in CHRNG, COL11A2 and SOX9 were carried by phenotypically healthy parents. Conclusion: Targeted next-generation sequencing can significantly improve the prenatal diagnoses of fetal skeletal dysplasia, providing parents with more precision medicine, and improved genetic counseling. [ABSTRACT FROM AUTHOR]

Background: High resolution genome-wide copy number analysis, routinely used in clinical diagnosis for several years, retrieves new and extremely rare copy number variations (CNVs) that provide novel candidate genes contributing to disease etiology. The aim of this work was to identify novel genetic causes of neurodevelopmental disease, inferred from CNVs detected by array comparative hybridization (aCGH), in a cohort of 325 Portuguese patients with intellectual disability (ID).Results: We have detected CNVs in 30.1% of the patients, of which 5.2% corresponded to novel likely pathogenic CNVs. For these 11 rare CNVs (which encompass novel ID candidate genes), we identified those most likely to be relevant, and established genotype-phenotype correlations based on detailed clinical assessment. In the case of duplications, we performed expression analysis to assess the impact of the rearrangement. Interestingly, these novel candidate genes belong to known ID-related pathways. Within the 8% of patients with CNVs in known pathogenic loci, the majority had a clinical presentation fitting the phenotype(s) described in the literature, with a few interesting exceptions that are discussed.Conclusions: Identification of such rare CNVs (some of which reported for the first time in ID patients/families) contributes to our understanding of the etiology of ID and for the ever-improving diagnosis of this group of patients. [ABSTRACT FROM AUTHOR]