Introduction and rationale
Hearing loss is the most common invisible sensory deficit. It represents a common birth defect that ranges from 1 to 3 per 1000 live births (NCHAM, 2006). In developing countries, it is estimated to be twice as that in the developed countries (Prasansuk, 2000). Permanent childhood hearing loss has been described as ‘neurologic emergency’ as auditory deprivation has a significant impact on the overall brain development and sensory integration of the child (Sheth and McHugh, 2007).
Genetics plays a fundamental role in the pathogenesis of hearing loss. Genetics accounts for approximately half of the cases of hearing impairment worldwide (Sheth and McHugh, 2007). Alterations in one or more of 400 genes involved in determination of the structure and function of the organ of hearing have been implicated in its pathogenesis (Marazita et al., 1993). The autosomal recessive pattern of inheritance is considered to be the most common form.
Recognition of the etiology of childhood hearing impairment is important in terms of prognosis and counseling. Genetic workup could facilitate establishing a cause of hearing loss in a significant number of deaf children. Moreover, the importance of early detection of hearing loss is well recognized and the use of genetic tests in a newborn screening program might be beneficial in identifying this major cause of hearing loss (Milunsky et al., 2000). It might provide insights into the efficacy of early intervention aiming to improve the communication skills acquired by hearing-impaired children.
Our understanding of genetics has increased markedly in the last century, especially in the last quarter of the 20th century. Mapping and identification of genes involved in hereditary hearing loss began only in 1990 (Sheth and McHugh, 2007). Molecular genetic techniques have led to significant advances in understanding of the genes involved in hearing loss. The Human Genome Project (2000) has led to better diagnostic and treatment options. More than 100 genes involved in nonsyndromic and syndromic hearing loss have been mapped and over 40 genes have been cloned (Burton et al., 2006).
Mutations in the gap junction β2 subunit (GJB2) gene that encodes the β2 polypeptide chain of connexin 26 (Cx26) protein have been described as a leading cause of deafness in almost all the populations studied. It was considered to be responsible for half of the cases of recessive hearing impairment in Caucasians (Zelante et al., 1997). Three mutations in GJB2 (35delG, 167delT, and 235delC) are particularly common in specific populations (Caucasoid, Jewish Ashkenazi, and Oriental, respectively), with a carrier frequency between one in 30 and one in 75 (Rabionet et al., 2000).
During the last decade, mutations in the GJB2 gene were studied in the Egyptian population. The main objective of these studies was to explore the prevalence of GJB2 gene mutations and their characteristics, if any, across different geographical locations in Egypt. This article reviews the different studies carried out in Egypt on genetics of hearing loss focusing on GJB2 gene mutations.
Review of the literature
Genetic hearing loss in Egypt
Interest in the genetics of hearing loss in Egypt began in the 1990s. Samy et al. (1990) reported that genetic hearing loss constituted 27.7% of the causes of hearing loss among Egyptian children. El Sayed et al. (1992) carried out a retrospective study to determine the profile and characteristics of hereditary hearing loss among Egyptian children. They collected data from the files of children up to 15 years of age with hereditary hearing loss from 1985 to 1990. Analysis of data showed that hereditary hearing loss was present in 9.5% of children. Hearing loss had occurred in the first 4 years of life in 85% of children. Hereditary sensorineural type was the most common and constituted 20% of all sensorineural hearing loss. Most of the children had severe and profound degrees of hearing loss, with a high-frequency slope configuration in 53%. However, 0.8% of children had conductive hearing loss. Systemic disorders in association with hereditary hearing loss were noted in 10.4%, which led to a delay in the diagnosis of hearing loss. The authors reported that consanguinity constituted 88% of the studied Egyptian families and recommended genetic counseling to prevent hearing loss in Egyptian children.
Galal et al. (1993) carried out a study to develop an etiological classification of hearing loss among Egyptian children. The study was carried out on 500 hearing-impaired children who were subjected to pediatric evaluation, audiological testing, laboratory investigations, and radiological examination. The results showed that genetic hearing loss was present in 43.6%, whereas acquired hearing loss was present in 50% of children. Idiopathic hearing loss was noted in 6.4% of children. The authors reported a highly significant increase in consanguinity up to 85%. First-degree cousins constituted 74%. The study showed that 17% of children had syndromic hearing loss with an age of onset below 5 years. Analysis of syndromic hearing loss showed that the most common syndromes, in order of frequency, were as follows: hearing loss with integumentary system disorders (2.8%), hearing loss with craniofacial anomalies (2.4%), and hearing loss with metabolic disorders (2.4%). The authors recommended adding genetic testing to the protocol of pediatric hearing evaluation.
Molecular genetic studies
There has been increasing interest in the GJB2 gene as one of a family of genes that encodes polypeptide components of intercellular gap junctions. It encodes the β2 polypeptide chain for the Cx26 protein, which is expressed in discrete regions of the cochlea (Goodenough et al., 1996). The expression of the GJB2 gene in the cochlea is essential for audition and other connexins in the cochlea cannot compensate for its loss in the auditory epithelial cells. The presence of Cx26 protein in the inner ear-supporting cells is responsible for the rapid removal of potassium ions away from the region of the sensory cells during transduction to maintain hair cell homeostasis and sensitivity (Kelsell et al., 1997).
A preliminary study on GJB2 gene mutations was carried out in the Delta area of Egypt by Kolkila et al. (2004). The data were collected from four families with a positive history of consanguineous marriage. The percentage of mutations in the GJB2 gene was 70.4%. Four types of mutations were found. They were 35delG (deletion/frame shift mutation), R143W mutation (missense mutation with nucleotide change 426C→T at codon 143R→W), V27I mutation (polymorphic mutation with nucleotide change 79G→A at codon 27V→I), and 235delC (deletion/frame shift mutation). The 35delG mutation was the most frequent type of mutation as it was detected in 78.9% of cases. Moreover, 70% of cases with this mutation were homozygous, whereas 30% were heterozygous. The percentage of compound mutations was 33.3%, which reflected the high level of consanguinity in the families studied.
A larger scale study on mutations of the GJB2 gene in a hearing-impaired Egyptian population was carried out by a team from the Audiology Unit of Ain Shams University in collaboration with the Department of Medical Genetics, University of Antwerp, Belgium (Snoeckx et al., 2005). Patients were recruited from the great Cairo area. The study was carried out on 170 patients, representing 120 families with nonsyndromic bilateral sensorineural hearing loss (SNHL) of variable onset and course. Seventy normal-hearing adult participants were chosen as a control group for the genetic workup. The participants and families were recruited from Cairo and Delta region.
Molecular diagnostic tests were used for the study and control groups. Mutation analysis of the complete coding region of the GJB2 gene was screened and the presence of the 342-kb deletion comprising the GJB6 gene was also investigated. In addition, complete audiological evaluation was carried out on patients in the study group to define the characteristic hearing profile. The performance of children with the GJB2 gene mutation while using hearing aids was compared with children with nongenetic hearing loss. This was done to observe the impact of such a mutation on the prognosis and communication strategies.
According to the results of the mutation analysis, patients were categorized as positive for GJB2 gene mutations and negative for GJB2 gene mutations. The results showed that GJB2 gene mutations were detected in only 15% of the Egyptian children. Ten different variants in the GJB2 gene were found, of whom six were described previously (Table 1). The 35delG mutation was the most frequent pathogenic mutations in the Egyptian population.
- Analysis of evolutionary conservation of the amino acid variants was carried out by ConSeq (http://conseq.bioinfo.tau.ac.il), a web server for the identification of structurally and functionally important residues in protein sequences. Conservation scores calculated by ConSeq are a relative measure of evolutionary conservation at each sequence site of the query sequence. These scores were converted into a conservation scale ranging from 1 (variable) to 9 (highly conserved).
- IC1, IC2: intracytoplasmic domain 1 and 2, respectively; TM1, TM3: transmembrane domain 1, 2, respectively.
GJB2 gene mutations in the Egyptian population were found in the majority of participants by congenital, profound, high frequency, stationary, and symmetrical SNHL mainly with a positive family history. Moreover, they belonged to the group of children with delayed language development and communication through total mode of communication. Nevertheless, the results highlighted the involvement of GJB2 gene mutations in some patients with postlingual progressive and with a mild degree of SNHL. Both the GJB2-positive and GJB2-negative categories were similar in their audiological profile, but had different communicative skills. Moreover, children with severe to profound SNHL related to GJB2 gene mutations had the worst response to hearing aid amplification in comparison with matched children with hereditary or nonhereditary etiology of hearing loss. The authors attributed the difference in the performance among the subgroups to the extensive destruction and degeneration of the whole organ of corti structures in GJB2-positive group, with a possible neuronal involvement. It was concluded that identification of GJB2 variants in infants with hearing loss has many clinical implications. These included ruling out the risk of effects on other systems in case of syndromic complications and prediction of severe to profound hearing loss requiring aggressive intervention that potentially indicated cochlear implant. It also enables genetic counseling for parents in terms of recurrence rates. Young family members should be monitored and followed up for the possibility of postlingual hearing loss.
The results of this study showed that it would not be reasonable or cost-effective to test for GJB2 gene mutations in every individual with hearing loss or in neonatal hearing screening programs as implemented now in many developed countries. Given the low prevalence of GJB2-related hearing loss in Egyptian population, a molecular diagnosis for GJB2 gene mutations in Egypt should only be proposed in neonates or infants with profound hearing loss. In such cases, GJB2 testing would not be for the diagnosis of hearing loss, but it would aim to tailor intervention rehabilitation programs.
Moreover, this study showed that other genes for hereditary hearing loss in Egypt are involved that deserve further investigations. This confirms that nonsyndromic deafness is genetically highly heterogeneous, with large differences between populations. Therefore, it is important to investigate the differences in mutation frequencies between populations. Population-specific data are a prerequisite for efficient and useful DNA diagnostics and genetic counseling for hereditary deafness.
Another study by Abou Elew et al. (2006) investigated the contribution of GJB2 (Cx26), 35delG mutation, toward congenital nonsyndromic autosomal recessive hearing loss in thirty eight patients and 40 normal-hearing Egyptian participants. The 35delG mutation was found in four (10.5%) patients; one was homozygous and three were heterozygous for this mutation. Bilateral profound SNHL was detected in the four patients. None of the controls carried the 35delG mutation. The authors concluded that the 35delG mutation is a relatively important contributor toward nonsyndromic hearing loss among the Egyptian population. The 35delG mutation has a major impact on the degree of hearing impairment.
Mutations in the GJB2 gene and other mutations in southern Egyptians were investigated by Mohamed et al. (2010). They studied 97 families with nonsyndromic hereditary hearing loss (155 individuals). The results showed the presence of 35delG mutations in 6.2% of the families, with a carrier frequency of 1.6%. The allelic frequency for 35delG was 8.7%, with 11 homozygous patients and five heterozygous patients. All homozygous patients showed a phenotype of bilateral severe to profound SNHL, whereas heterozygous patients were normal carriers, except for one patient, who had bilateral moderate SNHL. The authors compared the allelic frequency of 35delG in a south Egypt group with some other ethnic groups. They reported that the Egyptians had one of the lowest frequencies (1.6%) in comparison with other countries (Greece, 3.5%; Italy, 4%; Lebanon, 2.3%; Cyprus, 2.4%). In contrast to the rather frequent 35delG mutation, sample collection did not show any other known mutation, except in one family, where a novel amino acid substitution (p.I71N) was found. The authors concluded that the GJB2 gene mutation was not the gene of highest impact on the hereditary hearing loss in a southern Egyptian population. They recommended further studies on other genes that were detected by linkage study such as MYO15A.
Farid et al. (2012) investigated the carrier frequency of the 35delG mutation in patients with nonsyndromic hearing loss and its relation to the degree of hearing impairment. The study was carried out on 37 patients with congenital SNHL and 70 control participants with normal hearing. Patients had variable degrees of bilateral SNHL ranging in severity from mild to profound. Results indicated the presence of the 35delG mutation in seven (18.9%) patients, five homozygous and two heterozygous for this mutation. The frequency of the mutant allele among patients was 16.21%. Genotype–phenotype relation of the 35delG mutation in the patients indicated bilateral profound SNHL in five patients, three homozygous and two heterozygous for the mutation. The allelic frequency of the 35delG mutation in the control group was 0.71%. It was concluded that the 35delG mutation of the GJB2 gene is an important contributor to autosomal recessive non-syndromic hearing loss (ARNSHL) in the Egyptian population. It has a relevant impact on the degree of hearing loss. They recommended further studies on a larger scale to identify other mutations in GJB2 gene as well as other genes implicated in the development of ARNSHL.
In the Division of Human Genetics and Genome Research of the National Research Centre, El Bagoury et al. (2014) studied the frequency of mutations in the GJB2 gene and the two most common deletions in the GJB6 gene [del (GJB6-D13S1830) and del (GJB6-D13S1854)] in 36 Egyptian patients from 36 unrelated Egyptian families. The patients were from different areas of upper and lower Egypt and they presented with mild, moderate, or severe prelingual hearing loss. Results of the study were in agreement with the previous studies in terms of frequency of the GJB2 gene mutations. It was found that the homozygous form of 35delG in the GJB2 gene was detected in 13.8% of patients. No deletions were detected in the GJB6 gene as reported by Snoeckx et al. (2005).
Molecular genetic studies in Egypt versus the Middle East and Mediterranean countries
GJB2 gene mutations were studied in other countries in the Middle East. The results showed that the DFNB1 locus accounts for 16% of the families tested in Jordan (Medlej-Hashim et al., 2002) and 10% in Iranians (Najmabadi et al., 2002). These findings are similar to those from Egyptian studies. However, results were higher (23%) in some Palestinian communities (Shahin et al., 2002) and in Lebanon (33%) (Mustapha et al., 2001). A study in an Omani population showed that it played no role in nonsyndromic recessive deafness (Simsek et al., 2001).
Lucotte (2007) carried out a comparable study on the percentage of carriers in seven populations of the Mediterranean area. The study was carried out on the genomic DNAs out of a total of 886 healthy individuals from Sevilla (Spain), Genoa, Sicily (Italy), Alexandria (Egypt), Libya, Algier (Algeria), and Tangier (Morocco), genotyped by the Taqman assay. The approximate proportions of the 35delG mutation are less than 1/149 in Sevilla, 1/129 in Genoa, 1/34 in Sicily, 1/54 in Alexandria, 1/41 in Libya, 1/141 in Algier, and 1/123 in Tangier. When compared with other Mediterranean populations, the mean prevalence of the mutation was 1/49. One of the most increased values in the prevalence of 35delG corresponds to Greece (1/28); the pattern of various prevalences of 35delG was interpreted in the present meta-analysis as being a result of Ancient Greek colonization of the ‘Magna Grecian’ in historical times.
Abidi et al. (2007) studied the prevalence and spectrum of GJB2 mutations, including the GJB6-D13S1830 deletion, in Moroccan patients and estimated the carrier frequency of the 35delG mutation in the general population. GJB2 mutations were found in 43.20% of the deaf patients. Among these patients, 35.80% were 35delG/35delG homozygous, 2.47% were 35delG/wt heterozygous, 3.70% were V37I/wt heterozygous, and one patient was E47X/35delG compound heterozygous. The carrier frequency of GJB2-35delG in the normal Moroccan population was 2.65%. These findings indicate that the GJB2-35delG mutation is the major cause of autosomal recessive nonsyndromic hearing loss in a Moroccan population, which is different from the findings in Egyptian population.
Al Achkar et al. (2011) studied the frequency of six GJB2 mutations in 50 Syrian families with congenital deafness and 180 controls. The data showed a high prevalence of the 35delG mutation among deaf families. Homozygous 35delG was detected in fifteen of the Syrian families (30%). A compound heterozygous genotype was observed in two families: one with the 35delG/167delT mutation (2%) and one with the 35delG/M34T mutation (2%). Nine families were heterozygous, with no second identified mutation in Cx26. It was concluded that these results have important implications for the diagnosis and counseling of families with Cx26 deafness.
This comparative review indicates wide variability in the prevalence of the GJB2 mutation in the Middle East area and Mediterranean countries because of ethnic differences. However, all studies indicate that 35delG was the most common mutation in these areas.
Conclusion and recommendations
Genetic hearing loss is found in almost 50% of the causes of pediatric hearing loss in Egypt because of the high prevalence of consanguineous marriages in the Egyptian population. GJB2 gene mutations are not common mutations as they were present in almost 15% of participants in the studies carried out in different locations in Egypt. The 35delG mutation of the GJB2 gene is an important contributor to ARNSHL in the Egyptian population. Bilateral symmetrical severe to profound SNHL is the characteristic profile in GJB2, especially in homozygous patients.
The frequency of GJB2 mutations was symmetrical in different locations of Egypt. The results in Delta area showed higher frequency as the study included four consanguineous families, whereas in other studies large series of one patient per family were studied.
It is recommended to raise public awareness of the consequences of genetic hearing loss because of high rate of consanguineous marriages in the Egyptian population. Molecular genetic studies for a large number of families with nonsyndromic recessive hearing loss in different geographical areas in Egypt should be considered to pinpoint the common genes involved in deafness in the Egyptian population. It is also recommended to apply the new strategy of exome sequencing for all genes implicated in hearing loss, for at least one patient in each family, to identify the spectrum of gene mutations in the Egyptian population.
Conflicts of interest
There are no conflicts of interest.
Abidi O, Boulouiz R, Nahili H, Ridal M, Alami MN, Tlili A, et al..GJB2 (connexin 26) gene mutations in Moroccan patients with autosomal recessive non-syndromic hearing loss
and carrier frequency of the common GJB2-35delG mutation.Int J Pediatr Otorhinolaryngol2007;71:1239–1245.
Abou Elew H, Abou Elew M, Abdel Rahman M, El Abd S.Contribution of the GJB2 (connexin 26) 35delG mutation to nonsyndromic autosomal recessive hearing loss in Egyptian patients.Egypt J Lab Med2006;18:25–28.
Al Achkar W, Moassass F, Al Halabi B, Al Ablog A.Mutations of the connexin 26 gene in families with non-syndromic hearing loss
.Mol Med Reports2011;4:331–335.
Burton S, Withrow K, Arnos K, Kalfoglou A, Pandya A.A focus group study of consumer attitudes toward genetic testing and newborn screening for deafness.Genetics in medicine2006;8:779–783.
Denoyelle F, Weil D, Maw MA, Wilcox SA, Lench NJ, Allen Powell DR, et al..Prelingual deafness: high prevalence of a 30delG mutation in the connexin 26 gene.Hum Mol Genet1997;6:2173–2177.
El Bagoury N, Soliman H, Mohammed O, Ghorab E, El Ruby M, El Bassyouni HT, Essawi M.Mutation analysis of the GJB2 and GJB6 genes in Egyptian patients with autosomal recessive sensorineural nonsyndromic hearing loss.Middle East J Med Genet2014;3:11–15.
El Sayed A, Soliman S, Kamal N1992Profile of hereditary hearing loss in Egyptian children: a retrospective study [master thesis]. Cairo: Ain Shams University.
Farid S, Abou Elew M, Shabana M, Elfouly H, Shaker O, Abou-Elew H, El-Akad M.Carrier frequency of the 35delG Mutations in non syndromic hearing loss
patients and its relation to the degree of hearing impairment.Egypt J Lab Med2012;24:25–30.
Galal A, Khattab A, Mohamed R, Osman Z, ElSawy M, Kamal N1993Genetic study of childhood deafness with associated anomalies among Egyptians [unpublished MD thesis]. Cairo: Genetics, Ain Shams University.
Goodenough DA, Goliger JA, Paul DL.Connexins, connexons and intercellular communication.Annu Rev Biochem1996;65:475–502.
Green GE, Scott DA, McDonald JM, Woodworth GG, Sheffield VC, Smith RJH.Carrier rates in the midwestern United States for GJB2 mutations causing inherited deafness.J Am Med Assoc1999;281:2211–2216.
Kelley PM, Harris DJ, Comer BC, Askew JW, Fowler T, Smith SD, Kimberling WJ.Novel mutations in the connexin 26 gene (GJB2) that cause autosomal recessive (DFNB1) hearing loss.Am J Hum Genet1998;62:792–799.
Kelsell DP, Dunlop J, Stevens HP, Lench NJ, Liang JN, Parry G, et al..Connexin 26 mutations in hereditary non-syndromic sensorineural deafness.Nature1997;387:80–83.
Kenna M, Wu B, Cotanche D, Korf B, Rehm H.Connexin 26 studies in patients with sensorineural hearing loss.Arch Otolaryngol Head Neck Surg2001;127:1037–42.
Kolkila I, Emara A, El Sharnooby J.Connexin 26 gene mutations in patients with non-syndromic hearing loss
and the use of multi-frequency puretone audiometry.Sci J Al-Azhar Med Fac (Girls)2004;25:939–955.
Lucotte G.High prevalences of carriers of the 35delG mutation of connexin 26 in the Mediterranean area.Int J Pediatr Otorhinolaryngol2007;71:741–746.
Marazita ML, Ploughman LM, Rawlings B, Remington E, Arnos KS, Nance WE.Genetic epidemiological studies of early-onset deafness in the U.S. school-age population.Am J Med Genet1993;46:486–491.
Marlin S, Garabédian ÉN, Roger G, Moatti L, Matha N, Lewin P, et al..Connexin 26 gene mutations in congenitally deaf children: pitfalls for genetic counseling.Arch Otolaryngol Head Neck Surg2001;127:927–933.
Medlej-Hashim M, Mustapha M, Chouery E, Weil D, Parronaud J, Salem N, et al..Non syndromic recessive deafness in Jordan: mapping of a new locus to chromosome 9q343 and prevalence of DFNB1 mutations.Eur J Hum Genet2002;10:391–394.
Milunsky JM, Maher TA, Yosunkaya E, Vohr BR.Connexin-26 gene analysis in hearing-impaired newborns.Genet Test2000;4:345–349.
Mohamed MR, Alesutan I, Föller M, Sopjani M, Bress A, Baur M, et al..Functional analysis of a novel I71N mutation in the GJB2 gene among southern Egyptians causing autosomal recessive hearing loss.Cell Physiol Biochem2010;26:959–966.
Mustapha M, Salem N, Delague V, Chouery E, Ghassibeh M, Rai M, et al..Autosomal recessive non-syndromic hearing loss
in the Lebanese population: prevalence of the 30delG mutation and report of two novel mutations in the connexin 26 (GJB2) gene.J Med Genet2001;38:E36.
Najmabadi H, Cucci RA, Sahebjam S, Kouchakian N, Farhadi M, Kahrizi K, et al..GJB2 mutations in Iranians with autosomal recessive non-syndromic sensorineural hearing loss.Hum Mutat2002;19:572.
Roeser R, Valente M, Hosford- Dunn H.Genetics of hearing loss.In Pediatric Audiology Diagnosis, Technology & Management2006Thieme Medical pub;100–121.
Prasansuk S.Incidence/prevalence of sensorineural hearing impairment in Thailand and Southeast Asia.Audiology2000;39:207–211.
Rabionet R, Gasparini P, Estivill X.Molecular genetics of hearing impairment due to mutations in gap junction genes encoding beta connexins.Hum Mutat2000;16:190–202.
Samy H, Soliman S, Tawfik S1990Hearing loss among Egyptian children: etiology and psychological aspect [unpublished Master thesis]. Cairo: Audiology Unit, Ain Shams University.
Shahin H, Walsh T, Sobe T, Lynch E, King MC, Avraham KB, et al..Genetics of congenital deafness in the Palestinian population: multiple connexin 26 alleles with shared origins in the Middle East.Hum Genet2002;110:284–289.
Sheth S, Mchugh KRoeser R, Valente M, Hosford- Dunn H.Genetics of hearing loss.Pediatric Audiology Diagnosis, technology & Management, Chapter 62007Thieme Medical publisher;100–121.
Simsek M, Al Wardy N, Al Khayat A, Shanmugakonar M, Al Bulushi T, Al Khabory M, et al..Absence of deafness-associated connexin-26 (GJB2) gene mutations in the Omani population.Hum Mutat2001;18:545–546.
Snoeckx RL, Hassan DM, Kamal NM, Van Den Bogaert K, Van Camp G.Mutation analysis of the GJB2 (connexin 26) gene in Egypt.Hum Mutat2005;26:60–61.
Zelante L, Gasparini P, Estivill X, Melchionda S, D’Agruma L, Govea N, et al..Connexin 26 mutations associated with the most common form of non-syndromic neurosensory autosomal recessive deafness (DFNB1) in Mediterraneans.Hum Mol Genet1997;6:1605–1609.