Adadey, S.M.2023-03-102023-03-102020-07Adadey, S.M (2020) The Genetics Of Congenital Non-Syndromic Hearing Impairment In Ghana ,University of Ghana, Legon, http://ugspace.ug.edu.gh:8080/handle/123456789/38755http://ugspace.ug.edu.gh:8080/handle/123456789/38755PhD. BiochemistryAbstract Background: The partial or total inability of an individual to hear sound is known as hearing impairment (HI). Globally, over 466 million people are living with HI, with the majority of cases from developing countries. Although over 123 genes have been associated with HI, only one gene (GJB2) has been studied in Ghana. This thesis therefore sought to investigate variants in GJB2, GJB4, GJB6, and GJC3 genes that are associated with HI in Ghana as well as to investigate other non-genetic causes of HI among Ghanaian children. Method: Hearing-impaired students from 11 schools for the deaf and Adamorobe, a village in Ghana, were enrolled and categorized as familial or non-familial. Control participants who did not have any known family history of HI were also enrolled. DNA was obtained from the blood samples collected from all the participants in 81 families from the schools for the deaf, 8 families from Adamorobe, and 166 non-familial cases. From the DNA samples, the regions that code for GJB2, GJC3, and GJB4 proteins were polymerase chain reaction (PCR) amplified using specific primers, Sanger sequenced and analyzed. The large genomic deletion of the GJB6 gene (GJB6-D3S1830) was investigated using multiplex PCR and confirmed with Sanger sequencing. A rapid diagnostic test was designed for GJB2-p.Arg143Trp (rs80338948) using restriction fragment length polymorphism (RFLP) and NciI restriction enzyme. The test was optimized and validated using Sanger sequencing. Bioinformatic tools and online databases were employed in predicting the clinical significance/pathogenicity of the identified variants in the connexin genes. In silico protein modeling techniques were used to model the protein structure for a likely pathogenic GJB4-p.Asn119Thr (rs190460237) mutant and wild-type proteins. The ligand-binding properties of the modeled proteins were studied. Results: The hearing-impaired participants enrolled on the study had severe to profound HI with the majority (68.3%) of them having prelingual HI. Nearly all the prelingual HI cases seemed congenital based on their parent’s reports, however, 54% of the hearing-impaired students received the first comprehensive hearing test when they had grown past the age of language development, thus between the ages of 6-11years. Cerebrospinal meningitis (CSM) was found to be the most frequent environmental cause of HI. The genetic analyses revealed that GJB2-Arg143Trp accounted for HI in 25.9% of the families studied, and 7.9% of isolated cases. A carrier frequency of 1.4% was estimated from randomly selected hearing controls in Ghana. Seven out of eight families from Adamorobe tested positive for the GJB2-Arg143Trp founder mutation. We were the first to report the presence of a GJB2-Trp44Ter variant in a hearing-impaired family in Ghana. To facilitate rapid screening of the variant within the population, a rapid GJB2- p.Arg143Trp-Nci-RFLP test was developed and found to be 100% sensitive, with no false positive or false negative observed. The test is highly specific for any variant within the recognition site of the restriction enzymes; however, it cannot differentiate between these variants. When screening other genes associated with HI, we identified one GJC3 variant that may not associate with HI. Also identified were seven GJB4 variants, of which 5 were predicted to be as either benign or synonymous, and the remaining 2 were predicted likely pathogenetic. One of the two variants may not be associated with HI because the variant’s homozygous form was observed in both patients and controls. We modelled the protein structure and function of the other likely pathogenic GJB4 variant (p.Asn119Thr) and found subtle but important alterations in the structure and binding characteristics of the mutated protein compared to the wildtype. Conclusion: We have obtained many important results through these studies. We have confirmed meningitis as the major cause of environmental HI in Ghana. Variations within the GJB2 gene account for the most HI cases of genetic origin in Ghana and hence we have identified the need to include GJB2 gene investigations in the national newborn hearing screening (NHS) program. The GJB2- p.Arg143Trp-Nci-RFLP test will therefore be instrumental in this capacity. Finally, based on data presented in this thesis, GJB4, GJB6, and GJC3 gene variants were not likely associated with HI in the Ghanaian population.enGhanaGeneticspolymorphismThe hearing-impairedThesis