Molecular diagnosis of MCAD in the Macedonian neonates with elevated medium-chain acylcarnitines identified through MS/MS-based newborn screening

Abstract

Introduction: Medium Cchain acyl-CoA dehydrogenase (MCAD) deficiency is an autosomal recessive disorder of fatty acid oxidation, with potential fatal outcome. It is diagnosed by acylcarnitine analysis on newborn screening blood spot cards by tandem mass spectrometry. Early diagnosis of MCAD and presymptomatic treatment can potentially reduce morbidity and mortality. Materials and Methods: A total of 38,578 newborns were screened for inborn errors of metabolism during May 2014 - Jan 2022, using the LC/MS/MS method. Eight newborns showed elevations of medium-chain acylcarnitines with predominance of octanoylcarnitine, and C8/C10 ratio as well. Molecular ACADM gene analysis was performed by whole exome sequencing using NovaSeq 6000 (Illumina), and confirmed by Sanger sequencing. Sequencing data were analysed using the bcbio_nextgen bioinformatics pipeline (version 1.2.7; GRCh37 genome). Results: Molecular analysis of the ACADM gene was performed in eight patients with positive metabolic screening for MCAD deficiency. Two different ACADM mutations were obtained in a total of 15/16 (93.75%) alleles of the patients. The common c.958A>G mutation (p.Lys329Glu) was detected in 12/16 (75%) alleles, while 3/16 (18.75%) alleles had c.244dupT pathogenic variant (p.Trp81fs). Five of the patients were homozygous for c.985A>G variant, and one was homozygous for c.244dupT. One of the patients was compound heterozygote (c.985A>G/c.244dupT) while another was a heterozygote for c.958A>G without second mutant allele detection. Conclusions: The sensitivity of medium-chain acylcarnitines as screening markers for early detection of MCAD was confirmed through the molecular analysis of the ACADM gene. Early detection and treatment have successfully prevented adverse health outcomes in patients with MCAD.