Clinical utility of exome sequencing in hearing loss: a retrospective cohort study
Background: Hearing loss (HL) is a prevalent sensorineural disorder with a highly heterogeneous etiology. Next-generation sequencing (NGS) has revolutionized the genetic testing landscape for diseases characterized by high genetic and allelic heterogeneity, enabling the simultaneous screening of hundreds of genes.
Methods: One hundred and seventy-one unrelated patients with non-syndromic or syndromic HL were enrolled in this study. Exome sequencing (ES) was applied to explore molecular etiology in the cohort, and clinical reports were provided by geneticists and genetic counselors. Multidisciplinary team forums were conducted to ensure accurate diagnoses and improved patient management.
Results: The molecular cause of HL was determined in 78 of 171 probands (45.6%): 54 with an autosomal recessive (AR) inheritance pattern, 23 with an autosomal dominant (AD) pattern, and 1 with both AR/AD inheritance patterns. Candidate variants in 33 genes were identified in the study cohort: 14 with an AR inheritance pattern, 18 with an AD pattern, and 1 with both AR/AD inheritance patterns. Twenty-eight of the variants identified in the study were novel.
Conclusion: Exome sequencing facilitates genetic diagnosis and improves the management of patients with HL in clinical practice. Identifying the etiology of HL may improve patient care, refine genetic counseling, and facilitate the estimation of recurrence risk.
1 Introduction
According to estimates from the World Health Organization, approximately 466 million individuals globally are affected by disabling hearing loss (HL), including 34 million children under the age of fifteen years (Wilson et al., 2017). HL is a prevalent sensorineural disorder with implications that extend far beyond sensory impairment (World Health Organization, 2018). Insufficient auditory stimulation or inadequate language exposure during early childhood may alter brain connectivity and processing; thus, children who have not acquired adequate auditory stimulation may encounter significant challenges in their subsequent linguistic acquisition, cognitive development, and psychosocial functioning (World Health Organization, 2018; Kral and O’Donoghue, 2010; Lieu et al., 2020). HL can manifest at any stage of life, impairing communication, affecting social interactions, and creating difficulties in professional settings (World Health Organization, 2018).
The complex etiology of HL, coupled with the highly variable and often overlapping presentations of different types of HL, poses challenges for traditional clinical diagnosis (Alford et al., 2014). It is estimated that up to 60% of educationally significant congenital and early-onset HL is associated with genetic factors (Alford et al., 2014; Schimmenti et al., 2004). Although the role of genetic factors in adult-onset HL remains less clear, an increasing number of susceptibility loci have been identified, and a substantial proportion of cases have been attributed to genetic causes (Alford et al., 2014; Michels et al., 2019). Approximately 120 non-syndromic loci have been identified in humans (hereditaryhearingloss.org), and over 400 genetic syndromes include HL as a characteristic feature (Alford et al., 2014; García-García et al., 2020). Recent recommendations advocate genetic testing as the initial diagnostic step for children with bilateral sensorineural hearing loss (SNHL), following cytomegalovirus (CMV) testing. Identifying the genetic etiology of HL can provide several potential benefits for patients and their families, including improved clinical management, informed planning for future medical and educational needs, and more accurate estimation of recurrence risk (Alford et al., 2014). Furthermore, given the rapid advancement of SNHL gene therapy research, understanding the genetic etiology of HL is crucial for assessing the eligibility for imminent clinical trials.
Traditional molecular diagnostic tests for HL primarily involved genotyping or DNA sequencing to identify specific HL variants or screen a limited number of genes associated with it. In the last decade, next-generation sequencing (NGS) has revolutionized the genetic testing landscape for diseases characterized by high genetic and allelic heterogeneity, such as HL, enabling the simultaneous screening of hundreds of genes (García-García et al., 2020). In the present study, exome sequencing (ES) was utilized to facilitate genetic diagnosis and improve the management of patients with HL in clinical practice.
2 Materials and methods
2.1 Ethics statement
The study was approved by the Medical Ethics Committee of Guangdong Women and Children Hospital. Written informed consent was obtained from all participants and their parents or legal guardians (in the case of children under 18). The authors had access to identifiable patient information, which was anonymized prior to submission. All the procedures conducted in the study adhered to the Declaration of Helsinki, as previously described (Liu et al., 2022).
2.2 Patients and samples
A total of 171 unrelated patients diagnosed with non-syndromic or syndromic HL were enrolled from January 2017 to September 2024 under an institutional review board-approved protocol of informed consent. A comprehensive history and physical examination were obtained for each participant, including clinical history, severity of HL, age and cause of onset, infection history, exposure to aminoglycoside antibiotics, and genetic factors associated with hearing impairment (Yin et al., 2014). The recruited patients exhibited sensorineural or mixed HL. The severity of HL was established as mild (hearing thresholds 26 dB–40 dB), moderate (hearing thresholds 41 dB–60 dB), severe (hearing thresholds 61 dB–80 dB), or profound (hearing thresholds more than 80 dB). A stepwise approach was provided as an alternative (Yin et al., 2014), and four patients in the study cohort opted for this approach and were pre-screened for the GJB2, SLC26A4, and MT-RNR1 gene variants prior to ES.
Peripheral blood samples were collected from the patients and their first-degree relatives if available. Genomic DNA was extracted using the SolPure Blood DNA Kit (Magen, Shanghai, China), following the manufacturer’s instructions, which included optional RNase treatment. The DNA was then fragmented using a Q800R Sonicator (Qsonica, CT, United States). The paired-end libraries were prepared following the Illumina library preparation protocol.
2.3 Exome sequencing
All cases underwent genetic evaluations using either whole-exome sequencing (WES) or clinical exome sequencing (CES). Custom-designed NimbleGen SeqCap probes (Roche NimbleGen, Madison, WI) were used for in-solution hybridization to enrich target sequences for WES or, in the case of clinical ES, the target sequences included ∼5,000 genes that were potentially associated with known Mendelian genetic diseases (AmCare Genomic Laboratory, Guangzhou, China). Low-quality reads (Phred score < Q20) were removed before demultiplexing. Sequences were aligned to the hg19 reference genome using NextGENe software (SoftGenetics, State College, PA) with standard parameters for single-nucleotide variant (SNV) and insertion/deletion (indel) detection.