First-Tier Versus Last-Tier Trio Whole-Genome Sequencing for the Diagnosis of Pediatric-Onset Rare Diseases

Abstract

Despite advances in diagnostics, children with rare genetic disorders still face extended diagnostic odysseys, delaying appropriate clinical management, and placing burdens on families and healthcare resources. Whole-genome sequencing (WGS) offers a more comprehensive interrogation of the genome than other genetic tests, but its use in clinical practice remains limited. This study compared diagnostic rates, turnaround times, and clinical utility of first-tier versus last-tier trio-WGS for patients with suspected genetic pediatric-onset conditions, including 97 critical and 104 non-critical patients. Eighty-five patients (42.3%), including 57 (58.8%) critical and 28 (26.9%) non-critical patients, received a molecular diagnosis. The diagnostic rate was higher for first-tier (57%) than for last-tier (32.8%) trio-WGS. Of 121 causative variants identified, 19.8% would have been missed by whole-exome sequencing. Laboratory processing time was 4 days for all patients. The clinical setting had the greatest impact on time to reporting, averaging 5 days for critical patients versus 74 days for outpatients. WGS results impacted clinical decision-making for 34% of all critical and 14.3% of WGS-positive non-critical patients. This is the first Italian clinical study to demonstrate the diagnostic and clinical utility of a genome-first approach for both critical and non-critical patients with suspected genetic pediatric-onset disorders and feasibility in a public healthcare system.

Keywords: critical illness; delayed diagnosis; neonatal intensive care units; newborn infant diseases; outpatients; pediatric intensive care units; public health practice; rare diseases; whole genome sequencing.

FIGURE 1

Study flow chart. Abbreviations: NICU/PICU, neonatal intensive care unit/pediatric intensive care unit; pt., patients; WGS, whole‐genome sequencing.

FIGURE 2

Distribution of cohort by age groups and practice setting. Abbreviations: NICU/PICU, neonatal intensive care unit/pediatric intensive care unit.

FIGURE 3

Distribution of clinical indications for WGS among critical and non‐critical subcohorts. For NICU/PICU patients, “other” includes cancer (3.1%), connective tissue disease, and genodermatosis (2% each), and skeletal dysplasia (1%). For outpatients, “other” includes connective tissue disease, cancer, and skeletal dysplasia (2% each), and growth disorder and genodermatosis (1% each). Abbreviations: NICU/PICU, neonatal intensive care unit/pediatric intensive care unit; WGS, whole‐genome sequencing.

FIGURE 4

Process flow from patient's clinical indication to the final report. Red text, tubes, and arrows represent critical samples; blue represents non‐critical samples. A–C refer to critical samples; A1–C1 to non‐critical samples. A and A1 correspond to elapsed time between onset of symptoms and blood sampling for WGS. T₀ corresponds to time of sample arrival in the laboratory. B and B1 correspond to elapsed time between sample arrival in the laboratory and accumulation of the 7 trios to be processed simultaneously. T₁ corresponds to the start time of sample processing and T₂ corresponds to time of communication of WGS results to the referring clinician. T₁–T₂ represents the processing time from DNA isolation up to the initial communication of results. T₀–T₂ represents laboratory TAT. C and C1 correspond to time for reverse phenotyping. T₀–T₃ corresponds to elapsed time from sample arrival in the laboratory to final report. Abbreviations used: NICU/PICU, neonatal intensive care unit/pediatric intensive care unit; TAT, turnaround time.