PROJECT SUMMARY/ABSTRACT Hearing impairment (HI) is the most common sensory deficit in the world. Congenital HI occurs in 1-2 per 1000 newborns. Understanding the mechanism of hearing will greatly aid in the development of treatment strategies for HI. Additionally, identification of new HI genes and pathogenic variants is highly beneficial for genetic screening so that HI can be diagnosed early, and intervention can occur to maximize the child’s cognitive, social-emotional, speech and language development. The first step is to identify genes involved in the etiology of nonsyndromic (NS) HI. Although 73 genes have been identified for autosomal recessive (AR) NSHI, the vast majority of NSHI genes have yet to be uncovered. The extreme genetic heterogeneity of NSHI is due to the complex inner ear architecture. Identification of genes involved in HI is the first step in improving knowledge of the auditory process, which in turn will aid in the development of diagnostic modalities and therapeutic interventions. Annually we will ascertain ~110 ARNSHI families with at least two members affected with severe to profound bilateral sensorineural ARNSHI from Hungary (Magyar and Romani), Nigeria, and Pakistan. DNA samples from hearing-impaired family members will be screened for genes/variants that are common causes of ARNSHI within their respective population. There will be ~92 families to identify new ARNSHI genes through next-generation sequencing and data analysis that uses information on variant frequency, deleterious status, and conservation. Information clinical etiology, animal models, and inner ear expression will also be used. Candidate variants will be validated and tested for segregation with ARNSHI status within the pedigrees. Once a putative causal variant is identified we will screen our in-house collection of ARNSHI exome data, use GeneMatcher, and contact our long-term collaborators to find additional ARNSHI families that segregate potentially causal variants within the same gene. For ~20 newly identified ARNSHI genes, expression and functional studies will also be performed using zebrafish and mice models to ensure gene pathogenicity as well as to elucidate a better understanding of the role the genes play in the etiology of HI. Due to the trans-Atlantic slave trade the majority of African Americans have west African ancestry; therefore, the study of Nigerian families will impact our understanding of the etiology of HI and improve diagnostics and treatment in this minority population. Families from Pakistan and Hungary (Magyar and Romani) should yield new ARNSHI gene that also have public health importance in other populations.