Custom Next-Generation Sequencing Identifies Novel Mutations Expanding the Molecular and clinical spectrum of isolated Hearing Impairment or along with defects of the retina, the thyroid, and the kidneys.

BACKGROUND: In the Tunisian population, the molecular analysis of hearing impairment remains based on conventional approaches, which makes the task laborious and enormously expensive. Exploration of the etiology of Hearing Impairment and the early diagnosis of causal mutations by next-generation sequencing help significantly alleviate social and economic problems. METHODS: We elaborated a custom SureSelectQXT panel for next-generation sequencing of the coding sequences of 42 genes involved in isolated hearing impairment or along with defects of the retina, the thyroid, and the kidneys. RESULTS: We report eight pathogenic variants, four of which are novel in patients with isolated hearing impairment, hearing impairment, and renal tubular acidosis, Usher syndrome and Pendred syndrome. Functional studies using molecular modeling showed the severe impact of the novel missense mutations on the concerned proteins. Basically, we identified mutations in nuclear as well as mitochondrial genes in a Tunisian family with isolated hearing impairment, which explains definitely the phenotype detected since 2006. CONCLUSION: Our results expanded the mutation spectrum and genotype-phenotype correlation of isolated and syndromic hearing loss and also emphasized the importance of combining both targeted next-generation sequencing and detailed clinical evaluation to elaborate a more accurate diagnosis for hearing impairment and related phenotypes especially in North African populations.

Novel pathogenic mutations and further evidence for clinical relevance of genes and variants causing hearing impairment in Tunisian population.

Introduction: Hearing impairment (HI) is characterized by complex genetic heterogeneity. The evolution of next generation sequencing, including targeted enrichment panels, has revolutionized HI diagnosis. Objectives: In this study, we investigated genetic causes in 22 individuals with non-GJB2 HI. Methods: We customized a HaloplexHS kit to include 30 genes known to be associated with autosomal recessive nonsyndromic HI (ARNSHI) and Usher syndrome in North Africa. Results: In accordance with the ACMG/AMP guidelines, we report 11 pathogenic variants; as follows; five novel variants including three missense (ESRRB-Tyr295Cys, MYO15A-Phe2089Leu and MYO7A-Tyr560Cys) and two nonsense (USH1C-Gln122Ter and CIB2-Arg104Ter) mutations; two previously reported mutations (OTOF-Glu57Ter and PNPT1-Glu475Gly), but first time identified among Tunisian families; and four other identified mutations namely WHRN-Gly808AspfsX11, SLC22A4-Cys113Tyr and two MYO7A compound heterozygous splice site variants that were previously described in Tunisia. Pathogenic variants in WHRN and CIB2 genes, in patients with convincing phenotype ruling out retinitis pigmentosa, provide strong evidence supporting their association with ARNSHI. Moreover, we shed lights on the pathogenic implication of mutations in PNPT1 gene in auditory function providing new evidence for its association with ARNSHI. Lack of segregation of a previously identified causal mutation OTOA-Val603Phe further supports its classification as variant of unknown significance. Our study reports absence of otoacoustic emission in subjects using bilateral hearing aids for several years indicating the importance of screening genetic alteration in OTOF gene for proper management of those patients. Conclusion: In conclusion, our findings do not only expand the spectrum of HI mutations in Tunisian patients, but also improve our knowledge about clinical relevance of HI causing genes and variants.

High levels of autoantibodies against catalase and superoxide dismutase in nasopharyngeal carcinoma.

INTRODUCTION: Autoantibodies against the antioxidant enzymes have been described in Epstein-Barr virus-associated diseases. Here, we hypothesize that Epstein-Barr virus (EBV), which is associated with nasopharyngeal carcinoma (NPC), induces anticatalase and/or antisuperoxide dismutase autoantibodies that inhibit catalase and/or superoxide dismutase activities and thereby contribute to the oxidative stress status described in this pathology. METHODS: Using a standard enzyme-linked immunosorbent assay (ELISA), the levels of immunoglobulin G (IgG), and M (IgM) directed against catalase and superoxide dismutase in the sera of 30 NPC patients and 30 healthy control individuals were evaluated. The antioxidative profile was tested among the same patients by measuring serum catalase and superoxide dismutase activities. To investigate the implication of EBV in the establishment of autoantibody production in NPC patients, a correlation study between serological testing for EBV viral capsid antigen immunoglobulin G (VCA IgG) and autoantibodies against both enzymes was undertaken. FINDINGS: The levels of IgG against superoxide dismutase and catalase were found to be increased in sera patients compared to controls (P<0.001). NPC patients exhibited decreased catalase (P<0.001) and superoxide dismutase activities (P<0.001) in their sera. However, a positive correlation between superoxide dismutase IgM antibody and IgG antibody titers to VCA (P<0.05, r=0.483, n=21) was found. A positive correlation between catalase (IgM) antibodies and IgG antibody titers to VCA (P<0.05, r=0.546, n=30) was also found. CONCLUSION: High levels of anticatalase and antisuperoxide dismutase antibodies in the sera of NPC patients were found.

Identification of a novel frameshift mutation in the DFNB31/WHRN gene in a Tunisian consanguineous family with hereditary non-syndromic recessive hearing loss.

Approximately 80% of hereditary hearing loss is non-syndromic. Non-syndromic deafness is the most genetically heterogeneous trait. The most common and severe form of hereditary hearing impairment is autosomal recessive non-syndromic hearing loss (ARNSHL), accounting for approximately 80% of cases of genetic deafness. To date, 22 genes implicated in ARNSHL have been identified. Recently a gene, DFNB31/WHRN, which encodes a putative PDZ scaffold protein called whirlin, was found to be responsible for the ARNSHL DFNB31. We found evidence for linkage to the DFNB31locus in a consanguineous Tunisian family segregating congenital profound ARNSHL. Mutation screening of DFNB31/WHRNrevealed four nonpathogenic sequence variants and a novel frameshift mutation [c.2423delG] + [c.2423delG] that changed the reading frame and induced a novel stop codon at amino acid 818 ([p.Gly808AspfsX11] + [p.Gly808AspfsX11]). To determine the contribution of the DFNB31locus in the childhood deafness, we performed linkage analysis in 62 unrelated informative families affected with ARNSHL. No linkage was found to this locus. From this study, we concluded that DFNB31/WHRN is most likely to be a rare cause of ARNSHL in the Tunisian population.