Evaluation of optical genome mapping in clinical genetic testing of facioscapulohumeral muscular dystrophy

Background Facioscapulohumeral muscular dystrophy (FSHD) is the third most common hereditary muscular dystrophy, caused by the contraction of the D4Z4 repeats on the permissive 4qA haplotype on chromosome 4, resulting in the faulty expression of the DUX4 gene. Traditional diagnostics is based on Southern blot, a time- and effort-intensive method that can be affected by single nucleotide (SNV), and copy number variants (CNV), as well as by the similarity of the D4Z4 repeats located on chromosome 10. We aimed to evaluate optical genome mapping (OGM) as an alternative molecular diagnostic method for the detection of FSHD. Methods We first performed optical genome mapping with EnFocus™ FSHD analysis using DLE-1 labeling and the Saphyr instrument, for our patients with inconclusive diagnostic Southern blot results, negative FSHD2 results, and clinically evident FSHD. Secondly, we performed OGM in parallel with the classical Southern blot analysis for our prospectively collected new FSHD cases. Finally, we used panel exome sequencing to confirm FSHD2. Results By using OGM we could resolve two cases with diagnostically inconclusive Southern blot results as having shortened D4Z4 repeats on the permissive 4qA allele, consistent with their clinical presentation. Results of the prospectively collected patients tested in parallel using Southern blot and OGM showed full concordance, showing OGM to be a useful alternative to the classical Southern blot method for detecting FSHD1. For a patient showing clinical FSHD but no shortened D4Z4 repeat on the 4qA allele using OGM or Southern blot, a likely pathogenic variant in SMCHD1 was detected using exome sequencing, confirming FSHD2. OGM and panel exome sequencing can be used consecutively to detect FSHD2.