Genotyping for Cx26 and Cx30 mutations in cases with congenital hearing loss

Hearing loss is the most frequent sensory defect in human being. The 13q11-q12 region contains the GJB2 and GJB6 genes, which code connexin 26 (CX26) and connexin 30 (CX30) proteins, respectively. The 35delG, 167delT, and 235delC mutations in the Cx26 gene are the main cause for sporadic nonsyndromic hearing loss (NSHL) in many populations. The 342-kb deletion [del(GJB6-D13S1830)] of the Cx30 gene is the second most common connexin mutation after the 35delG mutation in some NSHL populations. In our study 47 hearing-impaired students were included. The Cx26 gene and the Cx30 gene were analyzed for presence of the 35delG, 167delT, and 342-kb deletion [del(GJB6-D13S1830)]. Genotyping were performed for detecting 35delG, 167delT, and del(GJB6-D13S1830) mutations using the PCR-ELISA techniques. According to the results obtained from 47 cases, the 35delG mutation was detected in 7 cases ( approximately 14.9%). Four of these mutations were determined as homozygote mutant ( approximately 8.5%), and three were determined as heterozygote mutant ( approximately 6.4%). However, 167delT and del(GJB6-D13S1830) mutations were not detected in the study group. These results support the overwhelming majority of 35delG in our study group from deafness school in our study. In conclusion, the 35delG mutation was determined as the most frequently shown mutation that leads to congenital hearing loss as in previous studies from Turkey.     

Differential rates of frameshift alterations in four repeat sequences of hereditary nonpolyposis colorectal cancer tumors

In some Palestinian communities, the prevalence of inherited prelingual deafness is among the highest in the world. As an initial step towards understanding the genetic causes of hearing loss in the Palestinian population, 48 independently ascertained probands with non-syndromic hearing loss were evaluated for mutations in the connexin 26 gene. Of the 48 deaf probands, 11 (23%) were homozygous or compound heterozygous for mutations in GJB2. Five different mutations were identified: ivs1(+1) G-->A, 35delG, 167delT, T229C, 235delC. Nine deaf probands were homozygous and only two compound heterozygous. Among 400 hearing Palestinian controls, one carrier was observed (for 167delT). We show that GJB2 ivs1(+1) G-->A disrupts splicing, yielding no detectable message. Linkage disequilibrium analysis suggests, in the Palestinian and Israeli populations, a common origin of the 35delG mutation, which is worldwide, and of 167delT, which appears specific to Israeli Ashkenazi and Palestinian populations. A high prevalence of deafness, high frequency of homozygosity rather than compound heterozygosity among deaf, and low mutation carrier frequency together reflect the high levels of consanguinity of many extended Palestinian families. Some of the 25 families with multiple cases of inherited prelingual deafness and wildtype GJB2 sequences may represent as-yet-unknown genes for inherited hearing loss.     

GJB2 and mitochondrial A1555G gene mutations in nonsyndromic profound hearing loss and carrier frequencies in healthy individuals

This study aimed to assess mutations in GJB2 gene (connexin 26), as well as A1555G mitochondrial mutation in both the patients with profound genetic nonsyndromic hearing loss and healthy controls. Ninety-five patients with profound hearing loss (>90 dB) and 67 healthy controls were included. All patients had genetic nonsyndromic hearing loss. Molecular analyses were performed for connexin 26 (35delG, M34T, L90P, R184P, delE120, 167delT, 235delC and IVS1+1 A-->G) mutations, and for mitochondrial A1555G mutation. Twenty-two connexin 26 mutations were found in 14.7% of the patients, which were 35delG, R184P, del120E and IVS1+1 A-->G. Mitochondrial A1555G mutation was not encountered. The most common GJB2 gene mutation was 35delG, which was followed by del120E, IVS1+1 A-->G and R184P, and 14.3% of the patients segregated with DFNB1. In consanguineous marriages, the most common mutation was 35delG. The carrier frequency for 35delG mutation was 1.4% in the controls. 35delG and del120E populations, seems the most common connexin 26 mutations that cause genetic nonsyndromic hearing loss in this country. Nonsyndromic hearing loss mostly shows DFNB1 form of segregation.     

Moderate hearing loss and pseudodominant inheritance due to L90P/35delG mutations in the GJB2 (connexin 26) gene

Mutations in the GJB2 (connexin 26-Cx26) gene are responsible for 20-50% of cases with prelingual non-syndromic deafness in a large part of the world including Turkey. Although most of the cases with Cx26 deafness have a recessive mode of inheritance, a small group of families demonstrated dominant or pseudodominant inheritance. In this report we present a Turkish family in which the proband had congenital profound deafness and was found to be homozygous for the 35delG mutation, whereas the father and a paternal uncle who had milder, late-onset sensorineural hearing loss had compound heterozygous 35delG and L90P mutations. This family and previous reports with the L90P mutation demonstrate that the hearing loss associated with the L90P/35delG genotype is consistently milder than that of 35delG homozygotes. GJB2 gene screening should be considered in families with seemingly dominant inheritance and late-onset moderate hearing loss.     

Connexin 26 (GJB2) mutations in the Turkish population: implications for the origin and high frequency of the 35delG mutation in Caucasians

Mutations in the Connexin 26 (GJB2/Cx26) gene are responsible for more than half of all cases of prelingual non-syndromic recessive deafness in many Caucasian populations. To determine the importance of Cx26 mutations as a cause of deafness in Turks we screened 11 families with prelingual non-syndromic deafness, seven (64%) of which were found to carry the 35delG mutation. We subsequently screened 674 Turkish subjects with no known hearing loss and found twelve 35delG heterozygotes (1.78%; 95% confidence interval: 0.9%-3%) but no examples of the 167delT mutation. To search for possible founder effects, we typed chromosomes carrying the 35delG mutation for closely linked polymorphic markers in samples from Turkey and United States and compared the allele frequencies with those of hearing subjects. The data showed a modest degree of disequilibrium in both populations. Analyses of two pedigrees from Turkey demonstrated both conserved and different haplotypes, suggesting possible founder effects and multiple origins of the 35delG mutation.     

Connexin-26 gene analysis in hearing-impaired newborns

The efficacy and utility of the Connexin-26 (Cx-26) gene (also called GJB2) analysis from DNA isolated from Guthrie newborn screening cards is demonstrated. This analysis precisely defined a major cause of prelingual nonsyndromic deafness in those children requiring amplification in our study. Guthrie cards were obtained from 49 deaf children requiring amplification identified over the last 5 years by the Rhode Island Newborn Screening Program. Children with syndromes or other recognizable causes of hearing loss were excluded. DNA was extracted from the Guthrie cards and analyzed sequentially for the Cx-26 35delG mutation and then for the 167delT mutation followed by gene sequencing on remaining heterozygotes. Three of 42 children were 35delG homozygotes; 2/42 children were 35delG/167delT compound heterozygotes. One child was identified as being a 35delG heterozygote with no other mutation found by sequencing. Nine Guthrie cards yielded no amplification or uninterpretable results. Cx-26 mutations were identified as causing 11.9% of the deafness in the children studied. In conclusion, Cx-26 analysis is an important test that identifies a major cause of prelingual nonsyndromic deafness. Molecular analysis of hearing-impaired newborns will be important for genetic counseling in these families. Failures with Guthrie cards may make use of other collection methods preferable.     

High frequency of GJB2 mutation W24X among Slovak Romany (Gypsy) patients with non-syndromic hearing loss (NSHL)

Mutations in the GJB2 gene (connexin 26) represent a major cause of autosomal recessive non-syndromic hearing loss (NSHL) worldwide. In most Caucasian populations, the 35delG mutation in this gene was found to account for up to 50% of cases of the genetic non-syndromic childhood deafness. In populations of non-European ethnic background, other GJB2 gene mutations are occasionally common, e.g. 167delT in Ashkenazi Jews, R143W in Africaans and 235delC in Koreans. In this work, DNA samples from 54 unrelated NSHL patients from endogamous and inbred population of Slovak Roms (Gypsies) from Eastern Slovakia were screened for GJB2 mutations. The coding region of the GJB2 gene of patients was sequenced and mutations W24X, R127H, V153I, L90P and V37I were found. In Slovak Romany population, mutation W24X accounts for 23.2%, R127H for 19.4%, 35delG for 8.3%, V153I for 3.7%, L90P for 3.7% and V37I for 0.9% of screened chromosomes. As the W24X mutation was previously found in India and Pakistan, were from the European Romanies originate, it was brought by the European Romnanies from their Indian homeland. The carrier frequency of 35delG was estimated for Slovak non-Romany population to be 3.3%, and for Slovak Romany population to 0.88%. The carrier frequency of W24X varied in different Slovak Romany subpopulations from 0.0% up to 26.1%.     

High frequency of 35delG GJB2 mutation and absence of del(GJB6-D13S1830) in Greek Cypriot patients with nonsyndromic hearing loss

Mutations in the GJB2 (Connexin 26) gene are responsible for more than half of all cases of prelingual, recessive, inherited, nonsyndromic deafness in Europe. This paper presents a mutation analysis of the GJB2 and GJB6 (Connexin 30) genes in 30 Greek Cypriot patients with sensorineural nonsyndromic hearing loss compatible with recessive inheritance. Ten of the patients (33.3%) had the 35delG mutation in the GJB2 gene. Moreover, 9 of these were homozygous for the 35delG mutation, whereas 1 patient was in the compound heterozygous state with the disease causing E47X nonsense mutation. Another patient with severe sensorineural hearing loss was heterozygous for the V153I missense mutation. Finally, no GJB6 mutations or the known del(GJB6-D13S1830) were identified in any of the investigated Greek Cypriot nonsyndromic hearing loss patients. This work confirms that the GJB2 35delG mutation is an important pathogenic mutation for hearing loss in the Greek Cypriot population. This finding will be used toward the effective diagnosis of nonsyndromic hearing loss, improve genetic counseling, and serve as a potential therapeutic platform in the future for the affected patients in Cyprus.     

Changes in the connexin 26 gene (<Emphasis Type="Italic">GJB2</Emphasis>) in Russian patients with hearing loss: Results of long-term molecular diagnostics of hereditary nonsyndromic hearing loss

Molecular testing for mutations in the connexin 26 gene (GJB2) is a routine diagnostic analysis for subjects with hereditary hearing loss worldwide. However, till now there is no assessment of the diagnostic significance of this analysis for Russian patients, and there are difficulties in interpretation of the results of DNA diagnostics. In the present study, a sample of 705 patients with nonsyndromic autosomal recessive hearing loss from different regions of Russian Federation was investigated. A portion of DFNB1 hearing loss caused by mutations in the GJB2 gene among the sample was 46%. The frequency of DFNB1 hearing loss was 1:1000, that is, the frequency of isolated autosomal recessive hearing loss 1:500 in the population. It was found that each sixteenth individual in Russia is a heterozygous carrier of the mutation in the GJB2 gene. Totally, 20 pathological GJB2 alleles were detected; among them, a c.35delG mutation with the allelic frequency 81% prevails. Six most frequent mutations (c.35delG, c.313_326del14, c.23+1G>A (IVS1+1G>A), c.235delC, c.167delT, and p.Glu120del), which account for 95% of pathological GJB2 alleles, were detected. Mutations previously not described in the GJB2 gene (c.129delG, p.Gly200Arg, and c[Arg127His, Gly160Ser]) were found. An optimal algorithm of molecular testing of Russian patients which detects up to 100% of mutations in the GJB2 gene was suggested. Data concerning a clinical significance of p.Met34Thr and p.Val37Ile mutations are confirmed in the study. Eight polymorphic substitutions in the GJB2 gene which do not have clinical significance (p.Val27Ile, c.*3C>A, p.Val153Ile, p.Gly160Ser, c.Arg127His, p.Glu114Gly (c.341A>G), c.-45C>A, and p.Ala149Thr) were also detected.     

GJB2 and GJB6 mutations in 165 Danish patients showing non-syndromic hearing impairment

Thirty-two genes causing non-syndromic hearing impairment (NSHI) have been cloned, including GJB2 and GJB6 encoding the gap junction subunits connexin 26 and connexin 30, respectively. One mutation in GJB2, 35delG, accounts for a large percentage of GJB2 hearing impairment in Southern Europe whereas a considerably lower frequency has been reported from Northern European populations. Recently, a 342-kb deletion implicating GJB6 was found in 22 out of 44 NSHI patients of Spanish origin with only one mutated allele of GJB2. We report the first study of GJB2 and GJB6 mutations in Danish patients with NSHI. We tested 165 individuals and found GJB2 mutations in 16 individuals. The deletion implicating GJB6 was found in two individuals out of 9 heterozygous for GJB2 mutation. Furthermore, we screened 509 unselected samples from the Danish newborn population for the 35delG mutation in GJB2. We found 9 samples heterozygous for 35delG and 11 samples heterozygous for mutations leading to amino acid variants in GJB2 protein. In conclusion, our data are in accordance with results from other Northern European populations. Furthermore, our data on the GJB6 deletion suggest that routine screening for this deletion could help to explain hearing impairment in some Northern European NSHI patients heterozygous for a mutation in GJB2.     

Progressive hearing loss,and recurrent sudden sensorineural hearing loss associated with GJB2 mutations--phenotypic spectrum and frequencies of GJB2 mutations in Austria

Mutations of GJB2 (encoding connexin 26) are the most common cause of hearing loss (HL) in different populations, and a broad spectrum of GJB2 mutations has been identified. We screened 204 consecutive patients with non-syndromic sensorineural hearing loss for GJB2 mutations. Causative GJB2mutations were identified in 31 (15.2%) patients, and two common mutations, c.35delG and L90P (c.269T>C), accounted for 72.1% and 9.8% of GJB2 disease alleles. In four additional patients (2.0%) only one recessive GJB2 mutation was identified, making genetic counselling difficult. No genotype-phenotype correlation was established. We found, however, that homozygotes for truncating mutations were more likely to have a more severe degree of HL compared with other genotypes. Moreover, we showed by co-segregation studies that L90P is a GJB2 disease allele, and that compound heterozygotes for L90P and any recessive mutation share a mild to moderate phenotype. GJB2-associated HL was linked with progressive HL or with recurrent sudden sensorineural hearing loss (SSNHL) in three of 15 cases being analysed retrospectively. We extended the phenotypic spectrum of GJB2-related disease and recommend GJB2 mutation screening also in cases of progressive HL, and recurrent SSNHL. In addition, a carrier frequency of 1/110 (0.9%) for the most common Caucasian mutation in this gene, c.35delG, was determined in 1,212 blood donors from West-Austria, supporting the prevailing hypothesis of a Mediterranean founder mutation. Based on population and patient data, an overall GJB2 mutation carrier frequency of 1.3% was estimated for West-Austria.     

Detection of the 35delG/GJB2 and del(GJB6-D13S1830) mutations in Venezuelan patients with autosomal recessive nonsyndromic hearing loss

Severe to profound hearing impairment affects 1 of every 1000 newborn children each year. Inheritance accounts for 60% of these cases, of which 70% are nonsyndromic. The most common cause of autosomal recessive nonsyndromic hearing loss (ARNSHL) is mutation in GJB2, a gene on chromosome 13, which encodes a gap junction protein named Connexin 26. Mutations in GJB2 are responsible for 40% of genetic childhood deafness. The most common mutation, 35delG, predominates in many ethnic groups. Some families with linkage to the DFNB1 locus have none or only one mutated allele in GJB2, however, some subjects can exhibit a large deletion in another connexin gene, GJB6, resulting in a monogenic or digenic pattern of inheritance in this complex DFNB1 locus that contains both genes (GJB2 and GJB6). The aim of the study was to determine (1) the frequency for the 35delG (27.5%), del(GJB6-D13S1830) (2.5%) and del(GJB6-D13S1854) (0.0%) mutations in a cohort of 40 Venezuelan patients with ARNSHL and (2) the carrier frequency 35delG (4%), del(GJB6-D13S1830) (0%) and del(GJB6-D13S1854) (0%) in the Venezuelan population with no familial history of hearing impairment. One patient (2.5%) was detected as double heterozygote for the deletion del(GJB6-D13S1830) and 35delG mutation. This result has direct clinical implications because we include the molecular detection of the deletion del(GJB6-D13S1830) during the evaluation of the diagnosis of deafness in the Venezuelan population.     

Contribution of GJB2 Mutations to Hearing Loss in the Hazara Division of Pakistan

Mutations of GJB2, which encodes connexin 26, are the most common cause of hereditary hearing loss in many human populations. This study was initiated to determine the prevalence of GJB2 mutations in individuals with hearing loss from the Hazara Division in Pakistan. We recruited 70 participants with nonsyndromic deafness segregating as an apparently recessive trait and directly sequenced the GJB2 coding region from their DNA. The homozygous mutations c.71 G→A (p.W24X), c.104 T→G (p.I35S), and c.35delG (p.G12VfsX1) were identified as the cause of hearing loss in three participants (4.28%); in populations from other areas of Pakistan, frequencies of 6–7% have been observed. The mutations c.104 T→G and c.35delG were identified in Pakistan for the first time. These results confirm the low prevalence of GJB2 mutations in Hazara and suggest that mutations in other genes may play a significant role in the etiology of deafness in this population.     

GJB2 mutations and degree of hearing loss: a multicenter study

Hearing impairment (HI) affects 1 in 650 newborns, which makes it the most common congenital sensory impairment. Despite extraordinary genetic heterogeneity, mutations in one gene, GJB2, which encodes the connexin 26 protein and is involved in inner ear homeostasis, are found in up to 50% of patients with autosomal recessive nonsyndromic hearing loss. Because of the high frequency of GJB2 mutations, mutation analysis of this gene is widely available as a diagnostic test. In this study, we assessed the association between genotype and degree of hearing loss in persons with HI and biallelic GJB2 mutations. We performed cross-sectional analyses of GJB2 genotype and audiometric data from 1,531 persons, from 16 different countries, with autosomal recessive, mild-to-profound nonsyndromic HI. The median age of all participants was 8 years; 90% of persons were within the age range of 0-26 years. Of the 83 different mutations identified, 47 were classified as nontruncating, and 36 as truncating. A total of 153 different genotypes were found, of which 56 were homozygous truncating (T/T), 30 were homozygous nontruncating (NT/NT), and 67 were compound heterozygous truncating/nontruncating (T/NT). The degree of HI associated with biallelic truncating mutations was significantly more severe than the HI associated with biallelic nontruncating mutations (P<.0001). The HI of 48 different genotypes was less severe than that of 35delG homozygotes. Several common mutations (M34T, V37I, and L90P) were associated with mild-to-moderate HI (median 25-40 dB). Two genotypes--35delG/R143W (median 105 dB) and 35delG/dela(GJB6-D13S1830) (median 108 dB)--had significantly more-severe HI than that of 35delG homozygotes.     

Frequency of the 35delG allele causing nonsyndromic recessive deafness in the Algerian patients

Deafness is a heterogeneous disorder showing different patterns of inheritance and involving a multitude of different genes. Mutations in the GJB2 gene encoding connexin 26 (Cx26) protein are a major cause for non-syndromic autosomal recessive and sporadic deafness. Among these mutations, the c.35delG deletion is the most common mutation for sensorineural deafness. One hundred sixteen persons from fifty-eight families were tested by the method based on the principle of PCR-mediated-site-directed mutagenesis (PSDM), followed by a Bsl1 digestion. Mutation c.35delG was diagnosed in sixteen families (11 homozygotes and 5 heterozygotes). The low allelic frequency (17.24%) and low ratio of individuals homozygous (13.8%) and heterozygous (6.9%) for the c.35delG mutation suggest that there are other mutations in the GJB2 gene or other genes responsible for deafness in the Algerian population. This study reports a significant association (P=0.003) between first cousin consanguinity and non-syndromic prelingual deafness.     

Prevalence of GJB2 (CX26) gene mutations in south Iranian patients with autosomal recessive nonsyndromic sensorineural hearing loss

Hereditary hearing loss is a genetically heterogeneous disorder. Mutations in connexin 26 (CX26), are a major cause in many countries and are largely dependent on ethnic groups. The purpose of our study was to evaluate the prevalence of GJB2 mutations among affected individuals from south of Iran. Fifty patients presenting with autosomal recessive non-syndromic hearing loss from Fars, province in south of Iran, were studied for mutations in GJB2 gene and screened by direct sequencing. Mutations were detected in 15 out of 50 patients (30?%). Eight different mutations were identified; six of them were previously identified (35delG, V27I M34V, V153I, A149T, V198M). The remaining two alleles, L28I and N169T, were novel variants. The most common mutations were 35delG followed by V153I with an allele frequency of 7 and 6?%, respectively. In this study, 30?% of our subjects were found to have the causative variants or polymorphisms in GJB2 and the c.35delG mutation was the most common cause in our patients. However, more study with larger sample size as well as in vitro functional study for these new variants in Xenopus oocytes is required.     

Spectrum of the GJB2 mutations in Belarussian patients with hearing loss. Findings of pilot genetic screening of hearing impairment in newborns

A total of 111 unrelated probands and their 8 sibs from Grodno oblast (Belarus) with bilateral isolated sensorineural hearing impairment were studied for the presence of mutations in the connexin 26 (GJB2) gene. Mutations were detected in 51 probands (46% of the sample). A significantly higher frequency of the GJB2 gene mutations was observed in familial cases of the disease with the autosomal recessive mode of inheritance (in 78% of families). Detected characteristics of the GJB2 gene mutation spectrum demonstrated that the using the algorithm, which was designed for Russian patients, is optimal for the molecular study of patients from Belarus. In the sample of patients with hearing loss, the highest (among other similar samples studied in the world) allele frequency of c.313_326del14 mutation (7% of all pathological GJB2 alleles) was registered; Polish origin of this deletion was suggested. It was demonstrated that detection of the GJB2 gene mutation on one patient’s chromosome only is insufficient to confirm a molecular genetic diagnosis of hearing loss of the DFNB1 genetic type (autosomal recessive hearing loss caused by the GJB2 gene mutations). Pilot screening for the GJB2 gene mutations in newborns from Grodno oblast was performed. The material from 235 children was studied during the screening; nine heterozygous carriers of the mutation were found. The c.35delG mutation was detected in a homozygous state in a single newborn (hearing loss of moderate severity was subsequently audiologically confirmed in this child).     

Low incidence of GJB2, GJB6 and mitochondrial DNA mutations in North Indian patients with non-syndromic hearing impairment

Mutations at the DFNB1 locus which encode connexin 26 (CX26) and connexin 30 (CX30) proteins, respectively, are main cause for sporadic and familial non-syndromic hearing impairment (NSHI) in many populations. 342-kb deletion [del (GJB6-D13S1830)] of Cx30 gene is second most common connexin mutation. Specific mitochondrial DNA (mtDNA) mutations have been found to be associated with NSHI. In this study, we screened 210 NSHI patients for GJB2 mutations, ΔGJB6-D13S1830 deletion and three point mutations in mtDNA (A1555G, A3243G, A7445G) using PCR, DHPLC and sequencing in North Indian cohort. 35delG was found to be the most common mutation (10.9%), followed by W24X (3.8%) and W77X (1.9%) mutations. We did not observe GJB6-D13S1830 deletion and three mitochondrial point mutations in our cohort. Most of patients (50/58) carried monoallelic variations. Our results reveal different spectrum of GJB2 mutations specific to North Indian cohort, with 35delG being most prevalent. These results suggest that different types of GJB2 mutations affect autosomal recessive NSHI according to ethnic background.     

Prevalence and evolutionary origins of the del(GJB6-D13S1830) mutation in the DFNB1 locus in hearing-impaired subjects: a multicenter study

Mutations in GJB2, the gene encoding connexin-26 at the DFNB1 locus on 13q12, are found in as many as 50% of subjects with autosomal recessive, nonsyndromic prelingual hearing impairment. However, genetic diagnosis is complicated by the fact that 10%-50% of affected subjects with GJB2 mutations carry only one mutant allele. Recently, a deletion truncating the GJB6 gene (encoding connexin-30), near GJB2 on 13q12, was shown to be the accompanying mutation in approximately 50% of these deaf GJB2 heterozygotes in a cohort of Spanish patients, thus becoming second only to 35delG at GJB2 as the most frequent mutation causing prelingual hearing impairment in Spain. Here, we present data from a multicenter study in nine countries that shows that the deletion is present in most of the screened populations, with higher frequencies in France, Spain, and Israel, where the percentages of unexplained GJB2 heterozygotes fell to 16.0%-20.9% after screening for the del(GJB6-D13S1830) mutation. Our results also suggest that additional mutations remain to be identified, either in DFNB1 or in other unlinked genes involved in epistatic interactions with GJB2. Analysis of haplotypes associated with the deletion revealed a founder effect in Ashkenazi Jews and also suggested a common founder for countries in Western Europe. These results have important implications for the diagnosis and counseling of families with DFNB1 deafness.     

High frequency hearing loss correlated with mutations in the GJB2 gene

Genetic hearing impairment affects approximately 1/2000 live births. Mutations in one gene, GJB2, coding for connexin 26 cause 10%-20% of all genetic sensorineural hearing loss. Mutation analysis in the GJB2 gene and audiology were performed on 106 families presenting with at least one child with congenital hearing loss. The families were recruited from a hospital-based multidisciplinary clinic, which functions to investigate the aetiology of sensorineural hearing loss in children and which serves an ethnically diverse population. In 74 families (80 children), the aetiology was consistent with non-syndromic recessive hearing loss. Six different connexin 26 mutations, including one novel mutation, were identified. We show that GJB2 mutations cause a range of phenotypes from mild to profound hearing impairment and that loss of hearing in the high frequency range (4000-8000 Hz) is a characteristic feature in children with molecularly diagnosed connexin 26 hearing impairment. We also demonstrate that this type of audiology and high frequency hearing loss is found in a similar-sized group of deaf children in whom a mutation could only be found in one of the connexin 26 alleles, suggesting connexin 26 involvement in the aetiology of hearing loss in these cases. In our study of the M34T mutation, only compound heterozygotes exhibited hearing loss, suggesting autosomal recessive inheritance.