Back to 2026 Abstracts
Digital Karyogram Over Karyotype in DSD and Severe Hypospadias Evaluation
Elizabeth Lynch, MD, Ugur Hodoglugil, MD, PhD, Mikaela Kuan, BS, Sawona Biswas, MS, CGC, Fion Ma, MS, CGC, Neil Risch, PhD, MS, Laurence S. Baskin, MD.
University of California San Francisco, San Francisco, CA, USA.
BackgroundKaryotype analysis is a cornerstone of the initial evaluation for disorders of sex development (DSD) in patients presenting with atypical genitalia. The sex chromosome constitution directs all subsequent differential diagnoses and management decisions. Standard G-banded karyotyping carries a turnaround time of 5-10 days and resolution limit of 5-10 Mb. Adjunctive modalities such as FISH, fluorescence PCR, and chromosomal microarray (CMA) offer faster results but carry limitations including inability to detect balanced rearrangements or low-level mosaicism. Whole genome sequencing (WGS) is increasingly accessible with improving turnaround times and declining costs. WGS data can be analyzed to generate a digital karyogram providing high-resolution chromosomal copy number assessment. We hypothesize WGS-derived karyograms can serve as a reliable replacement test in the DSD workup.
MethodsFollowing IRB approval (IRB# 24-41714), 124 pediatric patients were prospectively enrolled. Eligible diagnoses included hypospadias, atypical genitalia, congenital adrenal hyperplasia (CAH), and DSD. Patients with known syndromes or congenital anomalies (e.g., imperforate anus, cardiac defects) were excluded. Genomic DNA was obtained via buccal swab/saliva or peripheral blood and submitted for WGS on a NovaSeq X platform, with alignment and variant calling performed using DRAGEN software. Tertiary analysis was performed using Emedgene software utilizing the Genome View tab and digital karyogram feature. WGS-derived karyograms were compared to a standard G-banded karyotype or SNP microarray as the gold standard; available NIPT results were reviewed secondarily.
ResultsOf 45 patients with available reference cytogenetic studies — including 36 postnatal peripheral blood karyotypes, 4 prenatal amniocentesis karyotypes, 3 postnatal SNP microarrays, 1 cord blood SNP microarray, and 1 postnatal aCGH — 42 had an accessible WGS digital karyogram; 3 were excluded due to inaccessible WGS data. Karyotypes included 31 with 46,XX or 46,XY, 2 with 47,XXY, 1 non-mosaic isodicentric Y 45,X, and 8 mosaic cases: 4 mosaic isodicentric Y 45,X (10%, 52%, 52%, 86% abnormal cells), 3 mosaic 45,X0/46,XY (30%, 43%, 54%), and 1 mosaic 46,XY/46,XX (55%/45%). WGS demonstrated 100% concordance across all 42 patients. Of these, 18 had NIPT data, of which 15 included sex chromosome data. Among 46,XX patients (n=7), NIPT was concordant in 3, unavailable in 4. Among 46,XY patients (n=23), 8 had NIPT, all concordant. Of 8 mosaic patients, 5 had NIPT; 3 had sex chromosome data: 2 mosaic isodicentric Y 45,X positive for 45,X0/46,XY mosaicism (isodicentric Y not specified), and 1 mosaic 45,X0/46,XY with possible X chromosome mosaicism, unconfirmed. (Table 1)
| Karyotype | | N | | Karyogram Concordance | | NIPT (n) | | NIPT Sex Chr. Data |
| 46,XX | | 7 | | 7/7 (100%) | | 3 | | 3/3 concordant |
| 46,XY | | 23 | | 23/23 (100%) | | 8 | | 8/8 concordant |
| 47,XXY | | 2 | | 2/2 (100%) | | — | | — |
| Non-mosaic idic Y, 45,X | | 1 | | 1/1 (100%) | | — | | — |
| Mosaic idic Y, 45,X | | 4 | | 4/4 (100%) | | 2 | | 2/2 positive for 45,X0/46,XY‡ |
| Mosaic 45,X0/46,XY | | 3 | | 3/3 (100%) | | 1 | | Possible X mosaicism, unconfirmed |
| Mosaic 46,XY/46,XX | | 1 | | 1/1 (100%) | | — | | — |
| Total | | 42 | | 42/42 (100%) | | 18 | | 15 with sex chr. data |
Table 1. WGS Digital Karyogram and NIPT Results (N=42). ‡Isodicentric Y not specified by NIPT.ConclusionsWGS-derived digital karyograms were fully concordant with standard cytogenetic results for 46,XX, 46,XY, and complex sex chromosome mosaicism, while NIPT demonstrated variable performance in detecting mosaic cases. These findings support WGS as a potential first-line screening tool for DSD; however, limitations remain in detection and characterization of translocations, isodicentric Y mosaicism, and low-level mosaicism. This analysis was not blinded, and prospective blinded validation in a larger cohort is needed.
Back to 2026 Abstracts
