Diagnostic Utility of Next-Generation Sequencing for Unconjugated Hyperbilirubinemia in Children

Unconjugated hyperbilirubinemia can cause severe neurologic complications. Next-generation sequencing (NGS) is widely used for molecular diagnosis in pediatric disorders. This study evaluated the clinical features, diagnostic utility of NGS, and prognosis of children with unconjugated hyperbilirubinemia.

Medical records of patients aged <18 years with unconjugated hyperbilirubinemia persisting >3 months from January 2015 to November 2024 were retrospectively reviewed. Hyperbilirubinemia was defined as total bilirubin >1.5 mg/dL. DNA was extracted from blood, and panel-based NGS targeting 96 genes was performed.

Among 98 patients, NGS was performed in 30 (30.6%), identifying pathogenic variants in 12 (40.0%). The most frequent diagnosis was Gilbert syndrome (20%), followed by hereditary spherocytosis (16.7%); one patient (3.3%) had Rotor syndrome. Among patients with hereditary spherocytosis, only one showed elevated reticulocyte counts, and none had anemia. Resolution of hyperbilirubinemia occurred in 34 patients (34.7%) with a median duration of 6.6 months. Persistent hyperbilirubinemia >1 year occurred in 35 patients (35.7%), although levels >5 mg/dL were not observed during the maximum follow-up period. Multivariate analysis showed peak bilirubin level (odds ratio, 1.65; 95% confidence interval, 0.087-2.895) was significantly associated with abnormal NGS results.

NGS provides effective diagnosis for unconjugated hyperbilirubinemia, with a 40% yield. Hemolytic disease should be considered as an underlying cause, even without anemia.