Characterising the spectrum of autosomal recessive hereditary hearing loss in Iran.
ABSTRACT: Countries with culturally accepted consanguinity provide a unique resource for the study of rare recessively inherited genetic diseases. Although hereditary hearing loss (HHL) is not uncommon, it is genetically heterogeneous, with over 85 genes causally implicated in non-syndromic hearing loss (NSHL). This heterogeneity makes many gene-specific types of NSHL exceedingly rare. We sought to define the spectrum of autosomal recessive HHL in Iran by investigating both common and rarely diagnosed deafness-causing genes.Using a custom targeted genomic enrichment (TGE) panel, we simultaneously interrogated all known genetic causes of NSHL in a cohort of 302 GJB2-negative Iranian families.We established a genetic diagnosis for 67% of probands and their families, with over half of all diagnoses attributable to variants in five genes: SLC26A4, MYO15A, MYO7A, CDH23 and PCDH15. As a reflection of the power of consanguinity mapping, 26 genes were identified as causative for NSHL in the Iranian population for the first time. In total, 179 deafness-causing variants were identified in 40 genes in 201 probands, including 110 novel single nucleotide or small insertion-deletion variants and three novel CNV. Several variants represent founder mutations.This study attests to the power of TGE and massively parallel sequencing as a diagnostic tool for the evaluation of hearing loss in Iran, and expands on our understanding of the genetics of HHL in this country. Families negative for variants in the genes represented on this panel represent an excellent cohort for novel gene discovery.
Project description:BACKGROUND:Non-syndromic hearing loss (NSHL) is the most common sensory impairment in humans. Until recently its extreme genetic heterogeneity precluded comprehensive genetic testing. Using a platform that couples targeted genomic enrichment (TGE) and massively parallel sequencing (MPS) to sequence all exons of all genes implicated in NSHL, we tested 100 persons with presumed genetic NSHL and in so doing established sequencing requirements for maximum sensitivity and defined MPS quality score metrics that obviate Sanger validation of variants. METHODS:We examined DNA from 100 sequentially collected probands with presumed genetic NSHL without exclusions due to inheritance, previous genetic testing, or type of hearing loss. We performed TGE using post-capture multiplexing in variable pool sizes followed by Illumina sequencing. We developed a local Galaxy installation on a high performance computing cluster for bioinformatics analysis. RESULTS:To obtain maximum variant sensitivity with this platform 3.2-6.3 million total mapped sequencing reads per sample were required. Quality score analysis showed that Sanger validation was not required for 95% of variants. Our overall diagnostic rate was 42%, but this varied by clinical features from 0% for persons with asymmetric hearing loss to 56% for persons with bilateral autosomal recessive NSHL. CONCLUSIONS:These findings will direct the use of TGE and MPS strategies for genetic diagnosis for NSHL. Our diagnostic rate highlights the need for further research on genetic deafness focused on novel gene identification and an improved understanding of the role of non-exonic mutations. The unsolved families we have identified provide a valuable resource to address these areas.
Project description:A significant contribution to the causes of hereditary hearing impairment comes from genetic factors. More than 120 genes and 160 loci have been identified to be involved in hearing impairment. Given that consanguine populations are more vulnerable to most inherited diseases, such as hereditary hearing loss (HHL), the genetic picture of HHL among the Iranian population, which consists of at least eight ethnic subgroups with a high rate of intermarriage, is expected to be highly heterogeneous. Using an electronic literature review through various databases such as PubMed, MEDLINE, and Scopus, we review the current picture of HHL in Iran. In this review, we present more than 39 deafness genes reported to cause non-syndromic HHL in Iran, of which the most prevalent causative genes include GJB2, SLC26A4, MYO15A, and MYO7A. In addition, we highlight some of the more common genetic causes of syndromic HHL in Iran. These results are of importance for further investigation and elucidation of the molecular basis of HHL in Iran and also for developing a national diagnostic tool tailored to the Iranian context enabling early and efficient diagnosis of hereditary hearing impairment.
Project description:Recent advances in targeted genomic enrichment with massively parallel sequencing (TGE+MPS) have made comprehensive genetic testing for non-syndromic hearing loss (NSHL) possible. After excluding NSHL subjects with causative mutations in GJB2 and the MT-RNR1 (1555A>G) variant by Sanger sequencing, we completed TGE+MPS on 194 probands with presumed NSHL identified across Japan. We used both publicly available minor allele frequency (MAF) datasets and ethnic-specific MAF filtering against an in-house database of 200 normal-hearing Japanese controls. Ethnic-specific MAF filtering allowed us to re-categorize as common 203 variants otherwise annotated as rare or novel in non-Japanese ethnicities. This step minimizes false-positive results and improves the annotation of identified variants. Causative variants were identified in 27% of probands with solve rates of 35%, 35% and 19% for dominant, recessive and sporadic NSHL, respectively. Mutations in MYO15A and CDH23 follow GJB2 as the frequent causes of recessive NSHL; copy number variations in STRC are a major cause of mild-to-moderate NSHL. Ethnic-specific filtering by allele frequency is essential to optimize the interpretation of genetic data.
Project description:BACKGROUND:Nonsyndromic hearing loss (NSHL) is the most common sensorineural disorder and one of the most common human defects. Autosomal recessive inheritance accounts for a huge percentage of familial cases. Next-generation sequencing (NGS) is a powerful molecular diagnostic strategy for NSHL. The combination of a microarray gene chip and NGS can better delineate the etiology and genetic cause of deafness in many cases. METHODS:One hundred and thirty-one unrelated students with NSHL who attend a special education school in Yunnan Province were recruited. Firstly, four common deafness-related genes (GJB2, GJB3, SLC26A4, and mtDNA 12S rRNA) were evaluated for mutations using a microarray kit. Furthermore, 227 known human deafness genes were sequenced to identify the responsible genetic variant of the proband in three Chinese families with autosomal recessive hearing loss. The mutational status of family members of the probands was validated by Sanger sequencing. RESULTS:Five novel variants were found in three families using NGS. In family 1, we identified compound heterozygosity at the MYO15A (OMIM, #600316), including an duplication variant c.3866dupC, p.His1290Alafs*25 and a 3-bp deletion (c.10251_10253del, p.Phe3420del), resulting in protein length changes and premature protein truncation, respectively. In family 2, two affected siblings from a consanguineous Chinese Dai family harbored an c.1274G>C, p.Arg425Pro missense variant in the OTOF (OMIM, #601071). In family 3, we identified compound heterozygosity for c.129_130del, p.His43Glnfs*28 and c.76_79del, p.Lys26* in the RDX gene (OMIM, #611022). CONCLUSION:Five novel variants were found in three families with NSHL. Our findings extend the mutational spectrum in deafness-related genes and will help physicians in better understanding the etiology of hearing loss.
Project description:<h4>Background</h4>Hearing loss (HL) is one of the most common heterogeneous congenital disabilities worldwide. Gap junction protein ?-3 (<i>GJB3</i>) gene encodes Connexin31 protein (Cx31). The hereditary type of hearing impairment in this gene are known to cause both autosomal recessive and autosomal dominant form. In addition, <i>GJB3</i> mutations have been involved in sensorineural deafness, erythrokeratodermia variabilis (EKV), and neuropathy diseases. We aimed to investigate <i>GJB3</i> mutations in people suffering from HL among three different ethnicities of Iranian population (Baloch, Kurd, and Turkmen).<h4>Methods</h4>In this descriptive study, 50 <i>GJB2</i>-negative non-syndromic hearing loss (NSHL) Iranian individuals from 3 ethnic groups of Baloch (n=17), Kurd (n =15) and Turkmen (n=18) were enrolled. DNA extractions, PCR, and mutation detection was carried out for the two large deletions of the <i>GJB6</i>, del (GJB6 -D13S1830,) and del (<i>GJB6</i> -D13S1854) followed by direct DNA sequencing method for the <i>GJB3</i>.<h4>Results</h4>DNA sequencing of <i>GJB3</i> was shown a missense heterozygous mutation rs199689484 (NM_024009.3) GJB3: c.340G>A (p.Ala114Thr) in a Baloch patient, and a polymorphism rs35983826 (NM_024009.3) <i>GJB3</i>: c.798C>T (p.Asn266=) in a Turkman patient, in coding region of the <i>GJB3</i>. We did not detect del (GJB6 -D13S1830) and del (GJB6 -D13S1854) among these three ethnicities in Iran.<h4>Conclusion</h4>Deafness is a heterogeneous disorder. Specific genes and mutations contribute to hearing loss that varies from locus to locus as well as from population to population.
Project description:Through gene mutation analysis of patients with non-syndromic hearing loss (NSHL) correct genetic counseling for patients with NSHL and their family members were provided. A total of 116 patients suffering from NSHL were selected, and Sanger sequencing was applied to analyze 31 mutation sites in four deafness genes [gap junction β-2 (GJB2), solute carrier family 26, member 4 (SLC26A4), GJB3 and mitochondria 12S ribosomal ribonucleic acid (12SrRNA)]. Based on detection results, for the families with reproductive needs, amniotic fluid was extracted from pregnant women during proper gestational weeks to identify fetal genotypes and predict hearing state. Among 116 patients with NSHL, 51 patients carrying definite pathogenic mutation were found, including 35 patients with GJB2 mutations, 14 patients with SLC26A4 gene mutations and 2 patients with mitochondrial deoxyribonucleic acid 12SrRNA (mtDNA 12SrRNA) mutations. No GJB3 gene mutation site was detected. In addition, prenatal diagnosis to 17 pregnant women who had given birth to babies with deafness was performed, and results suggested that genotypes of 6 fetuses were consistent with those of probands, genotypes of 8 fetuses were consistent with those of their parents, and no mutation was found in the other 3 fetuses. Gene mutation analysis of patients with NSHL can identify the etiology and provide appropriate genetic counseling and birth guiding for patients with NSHL and their family members. In addition, prenatal diagnosis to the families who plan to give birth again can avoid the natality of fetuses with hearing loss.
Project description:BACKGROUND:Hereditary hearing loss (HL) is heterogeneous in terms of their phenotypic features, modes of inheritance, and causative gene mutations. The contribution of genetic variants to sporadic HL remains largely expanding. Either recessive or de novo dominant variants could result in an apparently sporadic occurrence of HL. In an attempt to find such variants we recruited 128 Chinese patients with sporadic nonsyndromic sensorineural HL (NSHL) and performed targeted deafness multigene sequencing in these unrelated trios-families to elucidate the molecular basis. METHODS:We analyzed a total of 384 available members (probands and their two parents) from 128 unrelated Chinese families presenting with bilateral sensorineural HL, in which previous screening had found no mutations with the GJB2, SLC26A4, and MT-RNR1 genes. We used a targeted genomic enrichment platform to simultaneously capture exons, splicing sites, and immediate flanking intron sequences of 127 known deafness genes. Sanger sequencing was used to identify probands and their two parents segregating causative variants in the candidate gene. RESULTS:We observed that two heterozygous de novo WFS1 mutations in exon 8: c.2051C>T (p.A684V) and c.2590G>A (p.E864K) in five families. The two de novo WFS1 mutations were found in 3.9% (5/128) of sporadic HL patients. We found that four of the five patients had the same de novo p.A684V mutation, and their audiograms showed symmetrical bilateral and profound sensorineural hearing impairments at all frequencies, but only the proband with de novo p.E864K mutation demonstrated significantly bilateral moderate low-mid frequency sensorineural HL. Our data suggest that this WFS1 p.A684V is likely to be a de novo mutational hot spot. CONCLUSIONS:We found 3.9% (5/128) of sporadic NSHL is caused by de novo WFS1 mutations. Our data provide that the de novo p.E864K mutation is first identified and de novo p.A684V mutation is likely to be a mutational hot spot in WFS1. It is the first study to highlight that WFS1 gene with the two de novo mutations has been indicated to classify the distinct hearing impairment phenotypes. Furthermore, de novo p.A684V serves as a WFS1 mutational hot spot that was found in the Chinese population with sporadic childhood NSHL, and our study also provides pointers toward the necessity for sequencing of asymptomatic parents of a sporadic case with an apparent dominant pathogenic variant.
Project description:The genetics of both syndromic (SHL) and non-syndromic hearing loss (NSHL) is characterized by a high degree of genetic heterogeneity. We analyzed whole exome sequencing data of 102 unrelated probands with apparently NSHL without a causative variant in known NSHL genes. We detected five causative variants in different SHL genes (SOX10, MITF, PTPN11, CHD7, and KMT2D) in five (4.9%) probands. Clinical re-evaluation of these probands shows that some of them have subtle syndromic findings, while none of them meets clinical criteria for the diagnosis of the associated syndrome (Waardenburg (SOX10 and MITF), Kallmann (CHD7 and SOX10), Noonan/LEOPARD (PTPN11), CHARGE (CHD7), or Kabuki (KMT2D). This study demonstrates that individuals who are evaluated for NSHL can have pathogenic variants in SHL genes that are not usually considered for etiologic studies.
Project description:: Hearing loss (HL) is a common sensory disorder affecting over 5% of the global population. The etiology underlying HL includes congenital and acquired causes; genetic factors are the main cause in over 50% of congenital cases. Pathogenic variants in the GJB2 gene are a major cause of congenital non-syndromic hearing loss (NSHL), while their distribution is highly heterogeneous in different populations. To the best of our knowledge, there is no data regarding the genetic etiologies of HL in Peru. In this study, we screened 133 Peruvian families with NSHL living in Lima. We sequenced both exons of the GJB2 gene for all probands. Seven probands with familial NSHL that remained negative for GJB2 variants underwent whole genome sequencing (WGS). We identified biallelic pathogenic variants in GJB2 in 43 probands; seven were heterozygous for only one allele. The c.427C>T variant was the most common pathogenic variant followed by the c.35delG variant. WGS revealed three novel variants in MYO15A in two probands, one of them was predicted to affect splicing and the others produce a premature stop codon. The Peruvian population showed a complex profile for genetic variants in the GJB2 gene, this particular profile might be a consequence of the admixture history in Peru.
Project description:Hereditary hearing loss (HHL) is a common disorder characterized by a huge genetic heterogeneity. The definition of a correct molecular diagnosis is essential for proper genetic counseling, recurrence risk estimation, and therapeutic options. From 20 to 40% of patients carry mutations in GJB2 gene, thus, in more than half of cases it is necessary to look for causative variants in the other genes so far identified (~100). In this light, the use of next-generation sequencing technologies has proved to be the best solution for mutational screening, even though it is not always conclusive. Here we describe a combined approach, based on targeted re-sequencing (TRS) of 96 HHL genes followed by high-density SNP arrays, aimed at the identification of the molecular causes of non-syndromic HHL (NSHL). This strategy has been applied to study 103 Italian unrelated cases, negative for mutations in GJB2, and led to the characterization of 31% of them (i.e., 37% of familial and 26.3% of sporadic cases). In particular, TRS revealed TECTA and ACTG1 genes as major players in the Italian population. Furthermore, two de novo missense variants in ACTG1 have been identified and investigated through protein modeling and molecular dynamics simulations, confirming their likely pathogenic effect. Among the selected patients analyzed by SNP arrays (negative to TRS, or with a single variant in a recessive gene) a molecular diagnosis was reached in ~36% of cases, highlighting the importance to look for large insertions/deletions. Moreover, copy number variants analysis led to the identification of the first case of uniparental disomy involving LOXHD1 gene. Overall, taking into account the contribution of GJB2, plus the results from TRS and SNP arrays, it was possible to reach a molecular diagnosis in ~51% of NSHL cases. These data proved the usefulness of a combined approach for the analysis of NSHL and for the definition of the epidemiological picture of HHL in the Italian population.