USH1G with unique retinal findings caused by a novel truncating mutation identified by genome-wide linkage analysis.

PURPOSE: Usher syndrome (USH) is an autosomal recessive disorder divided into three distinct clinical subtypes based on the severity of the hearing loss, manifestation of vestibular dysfunction, and the age of onset of retinitis pigmentosa and visual symptoms. To date, mutations in seven different genes have been reported to cause USH type 1 (USH1), the most severe form. Patients diagnosed with USH1 are known to be ideal candidates to benefit from cochlear implantation. METHODS: Genome-wide linkage analysis using Affymetrix GeneChip Human Mapping 10K arrays were performed in three cochlear implanted Saudi siblings born from a consanguineous marriage, clinically diagnosed with USH1 by comprehensive clinical, audiological, and ophthalmological examinations. From the linkage results, the USH1G gene was screened for mutations by direct sequencing of the coding exons. RESULTS: We report the identification of a novel p.S243X truncating mutation in USH1G that segregated with the disease phenotype and was not present in 300 ethnically matched normal controls. We also report on the novel retinal findings and the outcome of cochlear implantation in the affected individuals. CONCLUSIONS: In addition to reporting a novel truncating mutation, this report expands the retinal phenotype in USH1G and presents the first report of successful cochlear implants in this disease.

A comprehensive introduction to the genetic basis of non-syndromic hearing loss in the Saudi Arabian population.

BACKGROUND: Hearing loss is a clinically and genetically heterogeneous disorder. Mutations in the DFNB1 locus have been reported to be the most common cause of autosomal recessive non-syndromic hearing loss worldwide. Apart from DFNB1, many other loci and their underlying genes have also been identified and the basis of our study was to provide a comprehensive introduction to the delineation of the molecular basis of non-syndromic hearing loss in the Saudi Arabian population. This was performed by screening DFNB1 and to initiate prioritized linkage analysis or homozygosity mapping for a pilot number of families in which DFNB1 has been excluded. METHODS: Individuals from 130 families of Saudi Arabian tribal origin diagnosed with an autosomal recessive non-syndromic sensorineural hearing loss were screened for mutations at the DFNB1 locus by direct sequencing. If negative, genome wide linkage analysis or homozygosity mapping were performed using Affymetrix GeneChip® Human Mapping 250K/6.0 Arrays to identify regions containing any known-deafness causing genes that were subsequently sequenced. RESULTS: Our results strongly indicate that DFNB1 only accounts for 3% of non-syndromic hearing loss in the Saudi Arabian population of ethnic ancestry. Prioritized linkage analysis or homozygosity mapping in five separate families established that their hearing loss was caused by five different known-deafness causing genes thus confirming the genetic heterogeneity of this disorder in the kingdom. CONCLUSION: The overall results of this study are highly suggestive that underlying molecular basis of autosomal recessive non-syndromic deafness in Saudi Arabia is very genetically heterogeneous. In addition, we report that the preliminary results indicate that there does not seem to be any common or more prevalent loci, genes or mutations in patients with autosomal recessive non-syndromic hearing loss in patients of Saudi Arabian tribal origin.

The first three years of screening for medium chain acyl-CoA dehydrogenase deficiency (MCADD) by newborn screening ontario.

BACKGROUND: Medium chain acyl-CoA dehydrogenase deficiency (MCADD) is a disorder of mitochondrial fatty acid oxidation and is one of the most common inborn errors of metabolism. Identification of MCADD via newborn screening permits the introduction of interventions that can significantly reduce associated morbidity and mortality. This study reports on the first three years of newborn screening for MCADD in Ontario, Canada. METHODS: Newborn Screening Ontario began screening for MCADD in April 2006, by quantification of acylcarnitines (primarily octanoylcarnitine, C8) in dried blood spots using tandem mass spectrometry. Babies with positive screening results were referred to physicians at one of five regional Newborn Screening Treatment Centres, who were responsible for diagnostic evaluation and follow-up care. RESULTS: From April 2006 through March 2009, approximately 439 000 infants were screened for MCADD in Ontario. Seventy-four infants screened positive, with a median C8 level of 0.68 uM (range 0.33-30.41 uM). Thirty-one of the screen positive infants have been confirmed to have MCADD, while 36 have been confirmed to be unaffected. Screening C8 levels were higher among infants with MCADD (median 8.93 uM) compared to those with false positive results (median 0.47 uM). Molecular testing was available for 29 confirmed cases of MCADD, 15 of whom were homozygous for the common c.985A > G mutation. Infants homozygous for the common mutation tended to have higher C8 levels (median 12.13 uM) relative to compound heterozygotes for c.985A > G and a second detectable mutation (median 2.01 uM). Eight confirmed mutation carriers were identified among infants in the false positive group. The positive predictive value of a screen positive for MCADD was 46%. The estimated birth prevalence of MCADD in Ontario is approximately 1 in 14 000. CONCLUSIONS: The birth prevalence of MCADD and positive predictive value of the screening test were similar to those identified by other newborn screening programs internationally. We observed some evidence of correlation between genotype and biochemical phenotype (C8 levels), and between C8 screening levels and eventual diagnosis. Current research priorities include further examining the relationships among genotype, biochemical phenotype, and clinical phenotype, with the ultimate goal of improving clinical risk prediction in order to provide tailored disease management advice and genetic counselling to families.

Renal failure associated with APECED and terminal 4q deletion: evidence of autoimmune nephropathy.

Autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) is a rare autosomal recessive disorder caused by mutations in the autoimmune regulator gene (AIRE). Terminal 4q deletion is also a rare cytogenetic abnormality that causes a variable syndrome of dysmorphic features, mental retardation, growth retardation, and heart and limb defects. We report a 12-year-old Saudi boy with mucocutaneous candidiasis, hypoparathyroidism, and adrenocortical failure consistent with APECED. In addition, he has dysmorphic facial features, growth retardation, and severe global developmental delay. Patient had late development of chronic renal failure. The blastogenesis revealed depressed lymphocytes' response to Candida albicans at 38% when compared to control. Chromosome analysis of the patient revealed 46,XY,del(4)(q33). FISH using a 4p/4q subtelomere DNA probe assay confirmed the deletion of qter subtelomere on chromosome 4. Parental chromosomes were normal. The deleted array was further defined using array CGH. AIRE full gene sequencing revealed a homozygous mutation namely 845_846insC. Renal biopsy revealed chronic interstitial nephritis with advanced fibrosis. In addition, there was mesangial deposition of C3, C1q, and IgM. This is, to the best of our knowledge, the first paper showing evidence of autoimmune nephropathy by renal immunofluorescence in a patient with APECED and terminal 4q deletion.

Cornelia de Lange syndrome: parental preferences regarding the provision of medical information.

We surveyed 57 caregivers of children with Cornelia de Lange syndrome (CdLS) using a self-report questionnaire designed to elicit their experiences with receiving information regarding health complications associated with this multisystem disorder. Their most frequent sources of information at the time of diagnosis were a health care provider (86%) and the CdLS Foundation (72%). Although most caregivers (82%) indicated a desire to receive information about all possible CdLS related health complications, on average they recalled receiving information on only 64% of their child's current or future health complications. For two of the top three complications (as ranked by caregivers), information was available to fewer than half of the caregivers at diagnosis. Only 40% of caregivers were satisfied with the amount of information received at diagnosis, while only 45% felt that the information they received was useful. Caregivers indicated a preference for receiving information in written format (59%) or verbally (39%). Our results indicate that caregivers of children with CdLS report a high need for information at diagnosis, regarding their child's actual or potential health complications. Dissatisfaction may result both from a deficiency in information provided, as well as a mismatch between issues mentioned and those that caregivers deem most important. Caregivers may benefit maximally from receiving information in person at the time of diagnosis as well as having an additional written source of information. These findings highlight the importance of complementary sources of information for caregivers and are likely to be relevant for other multisystem disorders.

Cost comparison of genetic and clinical screening in families with hereditary hemorrhagic telangiectasia.

Endoglin (ENG) and ALK-1 mutations cause hereditary hemorrhagic telangiecstasia (HHT), an autosomal dominant disorder leading to vascular dysplasia in the form of mucocutaneous telangiectasia and visceral arteriovenous malformations (AVMs). We proposed to compare two alternative strategies for management of HHT: screening HHT families with molecular diagnostic tests followed by targeted clinical screening versus conventional clinical screening. A decision analytic model was constructed to compare screening strategies for a hypothetical HHT family. The family consists of 1 index case and 13 relatives. The clinical screening protocol in use at the Canadian HHT Center in Toronto was assumed to be the standard of care. Unit costs for clinical screening (in Canadian dollars) were obtained from the 2003 Ontario Health Insurance Schedule of Benefits. Genetic screening costs were estimated for quantitative multiplex PCR and sequencing of Endoglin (ENG) and ALK-1 genes, as performed at HHT Solutions, Toronto. The genetic screening strategy resulted in a net cost of $4,060 per individual versus $5,975 for the clinical screening strategy. The genetic screening strategy would save $1,915 per family member or $26,810 saved per family. Sensitivity analyses revealed that the genetic screening strategy was cost saving over all plausible ranges of input variables for all hypothetical families tested. We concluded that a genetic screening strategy with targeted clinical screening is more economically attractive than conventional clinical screening and results in a reduction in the number of clinical tests for family members who do not have HHT.

Newly recognized autosomal recessive acrofacial dysostosis syndrome resembling Nager syndrome.

We report on two patients with a unique constellation of anomalies resembling the Nager acrofacial dysostosis syndrome. Clinical manifestations which differentiate their condition from Nager syndrome include: microcephaly, cleft lip and palate, a peculiar beaked nose, blepharophimosis, microtia, symmetrical involvement of the thumbs, and great toes and developmental delay. We postulate that the inheritance is autosomal recessive on the basis of similarly affected male and female sibs.

A novel missense mutation in the galactosyltransferase-I (B4GALT7) gene in a family exhibiting facioskeletal anomalies and Ehlers-Danlos syndrome resembling the progeroid type.

The Ehlers-Danlos syndrome (EDS) is a heterogeneous group of heritable connective tissue disorders characterized by skin hyperextensibility, joint hypermobility, and tissue fragility. Several genes have been implicated to result in EDS phenotypes. The progeroid type of EDS is characterized by wrinkled, loose skin on the face, curly fine hair, scanty eyebrows and eyelashes, in addition to the classical features of EDS. Here we describe two similarly affected individuals in two sibships of a large consanguineous family from Qatar. DNA samples from affected and unaffected members of the family were analyzed for homozygosity of polymorphic markers associated with genes that have been implicated in EDS. Among 28 markers analyzed, homozygosity was only observed for D5S469 and D5S2111, which were markers for galactosyltransferase-I (B4GALT7) located on chromosome 5q35.2, where the previously reported progeroid-like variant of EDS has been mapped. Exons harboring the coding regions and exon-intron junctions of B4GALT7 were amplified by PCR and examined for mutations. A homozygous misssense C to T substitution at nucleotide 808 in the coding region was discovered in both affected individuals. The carrier parents were heterozygous for this mutation, which was not found among 76 DNA samples from control individuals of the same ethnicity. Segregation of this novel mutation in the family further confirmed the allelic variant and its recessive mode of inheritance in this type of EDS. The C to T substitution results in an arginine to cysteine change at amino acid residue 270 that is located in the catalytically active extracellular C-terminal domain. This change could result in abnormal protein folding and/or aberrant interactions of mutated galactosyltransferase-I with other extracellular matrix proteins leading to the development of a progeroid-like phenotype in affected individuals.

Clinical and genetic aspects of trigonocephaly: a study of 25 cases.

We reviewed 25 patients ascertained through the finding of trigonocephaly/metopic synostosis as a prominent manifestation. In 16 patients, trigonocephaly/metopic synostosis was the only significant finding (64%); 2 patients had metopic/sagittal synostosis (8%) and in 7 patients the trigonocephaly was part of a syndrome (28%). Among the nonsyndromic cases, 12 were males and 6 were females and the sex ratio was 2 M:1 F. Only one patient with isolated trigonocephaly had an affected parent (5.6%). All nonsyndromic patients had normal psychomotor development. In 2 patients with isolated metopic/sagittal synostosis, FGFR2 and FGFR3 mutations were studied and none were detected. Among the syndromic cases, two had Jacobsen syndrome associated with deletion of chromosome 11q 23 (28.5%). Of the remaining five syndromic cases, different conditions were found including Say-Meyer syndrome, multiple congenital anomalies and bilateral retinoblastoma with no detectable deletion in chromosome 13q14.2 by G-banding chromosomal analysis and FISH, I-cell disease, a new acrocraniofacial dysostosis syndrome, and Opitz C trigonocephaly syndrome. The last two patients were studied for cryptic chromosomal rearrangements, with SKY and subtelomeric FISH probes. Also FGFR2 and FGFR3 mutations were studied in two syndromic cases, but none were found. This study demonstrates that the majority of cases with nonsyndromic trigonocephaly are sporadic and benign, apart from the associated cosmetic implications. Syndromic trigonocephaly cases are causally heterogeneous and associated with chromosomal as well as single gene disorders. An investigation to delineate the underlying cause of trigonocephaly is indicated because of its important implications on medical management for the patient and the reproductive plans for the family.

Genetic analysis of patients with the Saethre-Chotzen phenotype.

Saethre-Chotzen syndrome is a common craniosynostosis syndrome characterized by craniofacial and limb anomalies. Intragenic mutations of the TWIST gene within 7p21 have been identified as a cause of this disorder. There is phenotypic overlap with other craniosynostosis syndromes, and intragenic mutations in FGFR2 (fibroblast growth factor receptor 2) and FGFR3 (fibroblast growth factor receptor 3) have been demonstrated in the other conditions. Furthermore, complete gene deletions of TWIST have also been found in a significant proportion of patients with Saethre-Chotzen syndrome. We investigated 11 patients clinically identified as having the Saethre-Chotzen phenotype and 4 patients with craniosynostosis but without a clear diagnosis. Of the patients with the Saethre-Chotzen phenotype, four were found to carry the FGFR3 P250R mutation, three were found to be heterozygous for three different novel mutations in the coding region of TWIST, and two were found to have a deletion of one copy of the entire TWIST gene. Developmental delay was a distinguishing feature of the patients with deletions, compared to patients with intragenic mutations of TWIST, in agreement with the results of Johnson et al. [1998: Am J Hum Genet 63:1282-1293]. No mutations were found for the four patients with craniosynostosis without a clear diagnosis. Therefore, 9 of our 11 patients (82%) with the Saethre-Chotzen phenotype had detectable genetic changes in FGFR3 or TWIST. We propose that initial screening for the FGFR3 P250R mutation, followed by sequencing of TWIST and then fluorescence in situ hybridization (FISH) for deletion detection of TWIST, is sufficient to detect mutations in > 80% of patients with the Saethre-Chotzen phenotype.

Charcot-Marie-Tooth disease and related neuropathies: mutation distribution and genotype-phenotype correlation.

Charcot-Marie-Tooth disease (CMT) is a genetically heterogeneous disorder that has been associated with alterations of several proteins: peripheral myelin protein 22, myelin protein zero, connexin 32, early growth response factor 2, periaxin, myotubularin related protein 2, N-myc downstream regulated gene 1 product, neurofilament light chain, and kinesin 1B. To determine the frequency of mutations in these genes among patients with CMT or a related peripheral neuropathy, we identified 153 unrelated patients who enrolled prior to the availability of clinical testing, 79 had a 17p12 duplication (CMT1A duplication), 11 a connexin 32 mutation, 5 a myelin protein zero mutation, 5 a peripheral myelin protein 22 mutation, 1 an early growth response factor 2 mutation, 1 a periaxin mutation, 0 a myotubularin related protein 2 mutation, 1 a neurofilament light chain mutation, and 50 had no identifiable mutation; the N-myc downstream regulated gene 1 and the kinesin 1B gene were not screened for mutations. In the process of screening the above cohort of patients as well as other patients for CMT-causative mutations, we identified several previously unreported mutant alleles: two for connexin 32, three for myelin protein zero, and two for peripheral myelin protein 22. The peripheral myelin protein 22 mutation W28R was associated with CMT1 and profound deafness. One patient with a CMT2 clinical phenotype had three myelin protein zero mutations (I89N+V92M+I162M). Because one-third of the mutations we report arose de novo and thereby caused chronic sporadic neuropathy, we conclude that molecular diagnosis is a necessary adjunct for clinical diagnosis and management of inherited and sporadic neuropathy.

Detecting rearrangements in children using subtelomeric FISH and SKY.

The etiology of mental retardation (MR), often presenting as developmental delay in childhood, is unknown in approximately one-half of cases. G-banding is the standard method for investigating those suspected of having a chromosomal etiology; however, detection of structural abnormalities is limited by the size and pattern of the G-bands involved. Rearrangements involving subtelomeric regions have been shown to cause MR and this has generated interest in investigating the prevalence of these rearrangements using telomere-specific probes. In addition, because cryptic interchromosomal rearrangements may not be small or confined to chromosomal ends, spectral karyotyping (SKY) using chromosome-specific painting probes may be of value. We report here a study using these two FISH-based techniques in 50 children with idiopathic MR or developmental delay and normal GTG-banded karyotypes. Our objective was to assess the prevalence of cryptic rearrangements in this population using subtelomeric FISH and SKY. Three rearrangements were detected by subtelomeric FISH: a derivative 5 from a maternal t(5;21); a recombinant 11 from a paternal pericentric inversion; and a 2q deletion that was also present in the mother. Only the derivative 5 was detected by SKY. SKY did not detect any interstitial interchromosomal rearrangement. The prevalence of clinically significant cryptic rearrangements by subtelomeric FISH and SKY was thus 4% (95% confidence interval 0.5-13.7) and 2% (95% CI 0.05-10.7), respectively. This study supports the view that G-banding does not detect all clinically significant chromosomal abnormalities and that subtelomeric FISH and SKY can detect some of these abnormalities.

Severe and mild phenotypes in Pfeiffer syndrome with splice acceptor mutations in exon IIIc of FGFR2.

Pfeiffer syndrome is clinically and genetically heterogeneous. Three clinical subtypes have been delineated based on the severity of acrocephalysyndactyly and associated manifestations. Severe cases are usually sporadic and caused by a number of different mutations in exons IIIa and IIIc of the fibroblast growth factor receptor 2 (FGFR2) gene. Mild cases are either sporadic or familial and are caused by mutations in FGFR2 or FGFR1, respectively. We report on two individuals with different novel de novo mutations in FGFR2. The first is a 17-year-old male who has a severe phenotype, within the spectrum of subtype 1 including severe ocular proptosis, elbow ankylosis, visceral anomalies, and normal intelligence. This patient was found to have a novel complex mutation at the 3' acceptor site of exon IIIc of FGFR2, denoted as C952-3 del10insACC. The other patient, a 2-year-old female, has a mild phenotype, typical of the classic subtype 1 including brachycephaly with coronal synostosis and hypertelorism. She was also found to have a mutation at the 3' acceptor site (the same splice site) of exon IIIc of FGFR2, a point mutation designated as 952-1G-->A. Speculation on the molecular mechanisms that cause severe and mild phenotypes is presented in relation to these two cases.