A combined genomic and functional approach to One-Carbon Metabolism disorders in a population-based newborn screening

Genomic first-tier approaches are increasingly explored as a strategy for newborn screening, raising major challenges for variant interpretation in asymptomatic populations. Within the BabyDetect project, we evaluated the clinical relevance of variants detected in three key one-carbon metabolism genes – MTHFR , CBS , and MAT1A – by comparing next-generation sequencing with biochemical data. Variants of uncertain significance (VUS) in those three genes were observed in approximately 4% of newborns. All likely pathogenic or pathogenic variants were present in the heterozygous state. Methionine levels, measured prospectively as part of routine newborn screening, and total homocysteine levels, assessed retrospectively, remained within neonatal reference ranges for both VUS and likely pathogenic/pathogenic variant carriers, with no significant differences between variant groups. These findings indicate that neither VUS nor heterozygous likely pathogenic/pathogenic variants in these genes exert a measurable biochemical impact in the neonatal period. Our results demonstrate the critical value of integrating biochemical data to contextualize genomic findings. This combined approach supports cautious interpretation of genomic results in population-based newborn screening and argues against the necessity to consider VUS in the absence of biochemical or clinical abnormalities.