De novo inherited Xq25 deletion: hints from preimplantation genetic testing in alobar holoprosencephaly

This study investigates the association between alobar holoprosencephaly (HPE) and de novo germline microdeletions in the Xq25 region. To develop a Preimplantation Genetic Testing for Monogenic Disorders (PGT-M) based workflow enabling high-resolution preimplantation detection of sub-Mb microdeletions, overcoming the >1 Mb resolution limit of conventional whole genome amplification(WGA) copy number variation(CNV) sequencing to identify causative Xq25 variants and prevent pathogenic microdeletion transmission.

This study presents a clinical case involving a couple with an adverse obstetric history accompanied by two occurrences of HPE. Genetic analysis and counseling were performed, followed by haplotype analysis and PGT-M as interventions. The Mutated Allele Revealed by Sequencing with Aneuploidy and Linkage Analyses (MARSALA) technique was utilized as a simple, efficient, and reliable approach for PGT-M.

The primary outcomes were the detection of chromosome microdeletions and the successful selection of embryos lacking Xq25 fragment microdeletions. Pregnancy outcomes, embryo development, and genetic counseling results served as secondary outcomes. We analyzed conception products using chromosome microarray analysis and performed single-nucleotide polymorphism haplotype linkage analysis on biopsy tissues amplified by WGA prior to sequencing. Chromosome abnormalities with microdeletions were identified using the PGT-M-based workflow. Embryo euploidy status was assessed using high-depth copy number variation sequencing.

Among five tested blastocysts, three were euploid (two carrying a maternal mutation and one non-mutant.). Following implantation of the mutation-free euploid blastocyst, a successful pregnancy was achieved. Chromosomal karyotype and chromosome microarray analysis (CMA) analysis of mid-pregnancy amniotic fluid showed normal chromosomes and genetic content, leading to the birth of a healthy, full-term infant.

This study underscores the necessity of maintaining detailed obstetric records, providing genetic counseling, and preserving nonviable fetal genetic material. By integrating the MARSALA technique with selected genetic marker-based SNP linkage analysis under the framework of PGT-M, this study established a method enabling single-cell-level preimplantation diagnosis of various chromosomal microabnormalities.