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Contains fulltext : 52397.pdf (Publisher’s version ) (Closed access) BACKGROUND: Usher syndrome, a combination of retinitis pigmentosa (RP) and sensorineural hearing loss with or without vestibular dysfunction, displays a high degree of clinical and genetic heterogeneity. Three clinical subtypes can be distinguished, based on the age of onset and severity of the hearing impairment, and the presence or absence of vestibular abnormalities. Thus far, eight genes have been implicated in the syndrome, together comprising 347 protein-coding exons. METHODS: To improve DNA diagnostics for patients with Usher syndrome, we developed a genotyping microarray based on the arrayed primer extension (APEX) method. Allele-specific oligonucleotides corresponding to all 298 Usher syndrome-associated sequence variants known to date, 76 of which are novel, were arrayed. RESULTS: Approximately half of these variants were validated using original patient DNAs, which yielded an accuracy of >98%. The efficiency of the Usher genotyping microarray was tested using DNAs from 370 unrelated European and American patients with Usher syndrome. Sequence variants were identified in 64/140 (46%) patients with Usher syndrome type I, 45/189 (24%) patients with Usher syndrome type II, 6/21 (29%) patients with Usher syndrome type III and 6/20 (30%) patients with atypical Usher syndrome. The chip also identified two novel sequence variants, c.400C>T (p.R134X) in PCDH15 and c.1606T>C (p.C536S) in USH2A. CONCLUSION: The Usher genotyping microarray is a versatile and affordable screening tool for Usher syndrome. Its efficiency will improve with the addition of novel sequence variants with minimal extra costs, making it a very useful first-pass screening tool.

Editor—Mutations in GJB2 , the connexin 26 (Cx26) gene, are thought to account for over 50% of autosomal recessive, non-syndromic, congenital deafness, the most common form of genetic deafness1 2 and 10-30% of sporadic cases.2 3 Over 50 recessive mutations in the GJB2 gene have been reported since it was originally described4 (the connexin 26 (GJB2) deafness homepage athttp://www.iro.es/cx26deaf.html). The most common mutation is a deletion of a guanosine nucleotide at position 30-35 (35delG), accounting for approximately 30-63% of mutations in white populations with a carrier frequency of 1:31 in Mediterranean populations.1 2 However, the 35delG mutation is present at a lower prevalence in different ethnic groups,5-8 with other mutations occurring at a higher prevalence, such as 167delT in the Jewish population.9-12 Both the high carrier frequency of GJB2 mutations and the prevalence of non-35delG mutations in non-white populations implies that mutations other than 35delG may be more common in non-white ethnic minorities who have settled in the UK, particularly those in which consanguinity is prevalent. In order to examine strategies suitable for sensitive, medium throughput mutation detection in GJB2 , we used denaturing high performance liquid chromatography (DHPLC) to screen for mutations in a cohort of 51 multi-ethnic patients with non-syndromic deafness who presented at our centre for genetic counselling. We found that DHPLC detected all the control mutations in the sample and that no mutations were identified by sequencing that were not detected by DHPLC. Three mutations, W24X, W77X, and Q124X, found in Indian, Pakistani, and Bangladeshi families in this study, have been previously observed in families from the Indian subcontinent, suggesting that they may be common mutations in these ethnic groups. Fifty one subjects with non-syndromic hearing loss were ascertained through genetic counselling and audiological medicine clinics at Great …

Mutations in the human gap junction β-2 gene (GJB2) that encodes connexin-26 have been shown to cause non-syndromic sensorineural hearing loss (NSSNHL) at the DFNB1 locus on 13q11. Functional and genetic data regarding the disease causing potential of one particular GJB2 sequence variant, 101 T→C (M34T), have proven contradictory. In this study, we found the prevalence of the M34T allele in a cohort of white sib pairs and sporadic cases with NSSNHL from the United Kingdom and Ireland to be 3.179% of chromosomes screened. Significantly, we identified the first M34T/M34T genotype cosegregating in a single family with mid to high frequency NSSNHL. Screening a control population of 630 subjects we identified 25 M34T heterozygotes; however, no M34T homozygotes were detected. Surprisingly, the majority of M34T alleles (88%) were in cis with a 10 bp deletion in the 5' non-coding sequence. This non-coding deletion was also homozygous in the homozygous M34T subjects. Microsatellite analysis of flanking loci in M34T heterozygotes and controls does not define an extensive ancestral haplotype but preliminary data suggest two common alleles in subjects with the M34T allele. In summary, we provide data that support M34T acting as a recessive GJB2 allele associated with mild-moderate prelingual hearing impairment. Keywords: GJB2; connexin-26; M34T; hearing loss

Editor—The branchio-oto-renal (BOR) syndrome was first reported in the last century by Heusinger, but not clinically defined until 1976 by Melnick et al .1 The major clinical features associated with the BOR syndrome are hearing loss, branchial defects, ear pits, and renal anomalies.1 Hearing loss can be conductive, sensorineural, or mixed, ranging from mild to profound deafness.2 Renal abnormalities are also variable, including renal aplasia, hypoplasia, and dysplasia, as well as anomalies of the collecting system.3 Branchial defects including fistulas or cysts and ear pits are often observed, and minor features such as external ear abnormalities, ear tags, and lacrimal duct aplasia are sometimes present. The BOR syndrome is inherited in an autosomal dominant manner and penetrance is high, although expressivity can be extremely variable.3-5 The BOR syndrome gene was localised to 8q following the description of a person with an inherited rearrangement, dir ins(8)(q24.11:q13.3:q21.13) presenting with features of both the branchio-oto (BO) syndrome and trichorhinophalangeal syndrome.6 Linkage analysis confirmed that the BOR syndrome mapped to 8q13.3.7 8 Further fine mapping defined the BOR region to be a 450-650 kb interval.9-12In 1997, the gene was cloned by sequencing P1/PAC clones from a contig spanning the region, yielding sequences homologous to the Drosophila developmental gene eyes absent ( eya). Point mutations and deletions were subsequently detected in the human homologue, EYA1 , in families with the BOR syndrome.13 14 EYA1 consists of 16 coding exons and encodes a 559 amino acid protein. There are two additional EYA1 isoforms and all show significant homology to the Drosophila eya gene, as well as to the murine homologue, constituting a novel gene family.14 15 The Drosophila eya gene is essential in the formation of the fly compound eye and the pathway has …

Background Usher syndrome (USH) is an autosomal recessive disorder comprising retinitis pigmentosa, hearing loss and, in some cases, vestibular dysfunction. It is clinically and genetically heterogeneous with three distinctive clinical types (I–III) and nine Usher genes identified. This study is a comprehensive clinical and genetic analysis of 172 Usher patients and evaluates the contribution of digenic inheritance. Methods The genes MYO7A, USH1C, CDH23, PCDH15, USH1G, USH2A, GPR98, WHRN, CLRN1 and the candidate gene SLC4A7 were sequenced in 172 UK Usher patients, regardless of clinical type. Results No subject had definite mutations (nonsense, frameshift or consensus splice site mutations) in two different USH genes. Novel missense variants were classified UV1-4 (unclassified variant): UV4 is ‘probably pathogenic’, based on control frequency A being the most common USH1 mutation in the cohort). USH2A was responsible for 79.3% of USH2 families and GPR98 for only 6.6%. No mutations were found in USH1G, WHRN or SLC4A7. Conclusions One or two pathogenic/likely pathogenic variants were identified in 86% of cases. No convincing cases of digenic inheritance were found. It is concluded that digenic inheritance does not make a significant contribution to Usher syndrome; the observation of multiple variants in different genes is likely to reflect polymorphic variation, rather than digenic effects.

Genetic factors are the major causes of childhood hearing impairment. Whereas autosomal recessive mutations account for the majority of prelingual non-syndromic sensorineural hearing impairment (NSSHI), the relative contribution of mitochondrial DNA (mtDNA) mutations to childhood onset NSSHI has not been established. We screened 202 subjects with congenital/childhood onset NSSHI, consisting of 110 sporadic cases, 75 sib pairs, and 17 families with affected subjects in more than one generation, in order to determine the prevalence of mtDNA mutations associated with NSSHI. mtDNA mutations were found in three of 10 families (30%) in whom the affected members were related through the maternal lineage. One sporadic case (0.9%) was also found to have a known mtDNA mutation but none was found in the sib pairs. Although the prevalence of mtDNA mutations was low in the group as a whole (2%), we suggest that screening should be considered in cases of childhood hearing impairment when it is progressive and particularly in families where transmission is compatible with maternal inheritance. Keywords: mitochondrial DNA; point mutation; hearing impairment